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Treatment-Resistant Anxiety Disorders: Neurotrophic PerspectivesOctober 31st 2006
Anxiety disorders are the most prevalent psychiatric disorders in the United States. Although effective treatments are available, such as the SSRIs and cognitive-behavioral therapy (CBT), it is estimated that in about 40% of patients, anxiety disorders are partially or completely resistant to first-line treatment.
Enlisting Family Members to Address Treatment Refusal in Substance AbusersOctober 31st 2006
Anyone who is close to someone who abuses alcohol or drugs knows all too well that substance abusers do not typically seek treatment until they have experienced years of substance-related problems. During the first year after onset of a diagnosable substance use disorder, only 1 of 5 alcohol-dependent persons and 1 of 4 drug-dependent persons receive treatment.
Neurotransmitters, Pharmacologic Synergy, and Clinical StrategiesOctober 31st 2006
Although studies now suggest that some psychotropic medication regimens have a somewhat higher success rate than the one-third rule would have predicted, psychiatrists are still left with the problem of why it is that only one third to one half of patients who are treated get better, and why fewer still sustain that improvement over time
Pathology and Management of Treatment Resistance in Bipolar DisorderOctober 30th 2006
The problem of treatment resistance in bipolar disorder begins with its definition. Characterizing the phases of bipolar disorder as manic, mixed, hypomanic, or depressed does not do justice to the reality for many persons with this disorder.
Treatment Resistance in Schizophrenia: The Role of Alternative TherapiesOctober 30th 2006
In 1931, Gananath Sen and Kartick Chandra Bose reported on the use of an alkaloid extract from the Rauwolfia serpentina plant in the treatment of hypertension and "insanity with violent maniacal symptoms." They noted that dosages "of 20 to 30 grains of the powder twice daily produce not only a hypnotic effect but also a reduction of blood pressure and violent symptoms
Better Tools Needed to Measure Treatment OutcomeOctober 1st 2006
The need for better tools, as well as better use of existing tools, to measure treatment response in clinical trials was a principle focus of the 46th annual NIMH-sponsored NCDEU (New Clinical Drug Evaluation Unit) meeting, held June 12-15 in Boca Raton, Fla. Improved clinical research techniques are needed to better separate treatment effect from placebo response, to distinguish between active comparators, and to facilitate development of novel treatments, according to several presenters at the conference.
NIDA Confirms Benefit of Drug Treatment Programs for Criminal OffendersOctober 1st 2006
Effective treatment of drug abuse and addiction in the criminal justice system saves the community money and reduces crime, the NIH's National Institute on Drug Abuse (NIDA) reported in a recently released booklet, Principles of Drug Abuse Treatment for Criminal Justice Populations.
IOM Report Takes on Medication Errors . . . AgainOctober 1st 2006
In its latest report on medication errors, a committee assembled by the Institute of Medicine (IOM) included some sidebars on psychiatric drugs. The report, issued in July, said that there is too little data on misadministration of psychiatric drugs and that clinical trials with psychiatric drugs have been small and incapable of providing pragmatic, comparative information.
Book Review: Principles and Practice of Psychopharmacotherapy, 4th EditionOctober 1st 2006
A new edition of an established text is an opportunity for its authors and editors to review new information and prune what is no longer useful. The 4th edition of Principles and Practice of Psychopharmacotherapy is more successful at the former than it is at the latter.
BasicNeeds: Successful Mental Health Programs With Scarce ResourcesOctober 1st 2006
Ahmed Abdullai often wandered aimlessly and slept in a graveyard in Tamale, northern Ghana. Now, with the help of BasicNeeds--an international development charity--Abdullai takes medications to stabilize his mental illness, learns gardening skills, lives at home, contributes to the family farm, and feels like a "human being once again."
Youth Aggression: Economic Impact, Causes, Prevention, and TreatmentOctober 1st 2006
The increase in youth violence and aggression in the past 50 years has been called an "epidemic." This epidemic has had a tremendous impact on society. From an economic and public health perspective, primary prevention of youth violence is obviously desirable.
Treating Delirium: When the Brain Goes Off TrackOctober 1st 2006
Typically, delirium worsens at night ("sundowning"), with lucid intervals often present in the morning. It is important to realize that delirium may appear before any abnormal laboratory values are detected and may persist after the resolution of these abnormalities.
Cortisol and Seasonal Changes in Mood and BehaviorOctober 1st 2006
The degree to which season changes affect mood, energy, sleep, appetite, food preference, or desire to socialize with others has been called "seasonality." Identification of a seasonal pattern can only be made if both the patient and physician actively look for it.
Children's and Parents' Attitudes About Participation in Treatment ResearchOctober 1st 2006
Since children are a vulnerable population, ethical issues in the conduct of medication studies involving them are extremely important. We recently reported the results of a study that examined youths' and parents' attitudes about, and experiences with, participation in psychopharmacology treatment research.
Addressing Psychiatric Comorbidities in Patients With EpilepsyOctober 1st 2006
In a presentation given at the midyear meeting of the American Epilepsy Society, Andres Kanner, MD cited studies from the literature showing that the rates of depression, anxiety, psychosis, and attention-deficit/hyperactivity disorder (ADHD) are significantly higher among persons with epilepsy than among the general population
Understanding and Using the Placebo EffectOctober 1st 2006
Most physicians make at least some use of the placebo effect to enhance treatments, whether they realize it or not. This article examines the extent of the placebo effect in patients with psychiatric illness, and reviews what is known about how placebos work.
Treating Adolescents With Major Depression and an Alcohol Use DisorderOctober 1st 2006
Alcohol is the drug of choice for adolescents, with cigarettes and marijuana being second and third. Contrary to widespread belief, alcohol dependence is most common in 18- to 20-year-olds, with progressively decreasing rates of alcohol dependence in older age groups.
Depression Management in Cancer PatientsOctober 1st 2006
Depressive disorders and symptoms are common in cancer patients (up to 58% have depressive symptoms and up to 38% have major depression), worsen over the course of cancer treatment, persist long after cancer therapy, recur with the recurrence of cancer, and significantly impact quality of life.
Bipolar Disorder: Defining Remission and Selecting TreatmentOctober 1st 2006
The longitudinal course of bipolar disorder (BD) is characterized by a low rate of recovery, a high rate of recurrence, and poor interepisodic functioning. There is a need to invoke a chronic disease management model (CDMM) when treating individuals with BD.
Should Emergency Medicine Physicians Screen for Psychiatric Disorders?October 1st 2006
Emergency department (ED) visits have increased from 89 million in 1992 to more than 110 million in 2002, while the number of EDs decreased by about 15% during the same period. One suspected consequence of ED overcrowding is an increased tendency to disregard a psychiatric problem, especially if it is not the chief complaint.
The Camelford Hysteria: A Lesson for ECT?October 1st 2006
Complaints of persistent memory loss in otherwise well-functioning individuals after recovery from a psychiatric illness through electroconvulsive therapy (ECT) are best viewed as a conversion reaction or a somatoform disorder. The Camelford experience is a model for the complaints of ECT's profound personal memory losses.

domain: psychology


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domain: artistic


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A blog about psycholinguistics with posts each Thursday by Jessica Love, a psychologist and science writer at Northwestern University. Her most recent article “Reading Fast and Slow” appeared in our Spring 2012 issue.
Do baby sign language courses really work?
by Jessica Love | Thursday, January 22, 2015
On the psychology of artistic style
by Jessica Love | Thursday, January 15, 2015
We understand entropy surprisingly early in life
by Jessica Love | Thursday, January 08, 2015
When we expect to hear a rhyme and don’t
by Jessica Love | Thursday, December 18, 2014
by Jessica Love | Thursday, December 11, 2014
by Jessica Love | Wednesday, December 03, 2014
The search for the perfect title
by Jessica Love | Thursday, November 20, 2014
Verb meanings are slipperier than noun meanings
by Jessica Love | Thursday, November 13, 2014
Two studies suggest yes and no
by Jessica Love | Thursday, November 06, 2014
On our unspoken preferences for ordering adjectives
by Jessica Love | Thursday, October 30, 2014
THIS WEEK’S ARCHIVE PICK
by John Kaag
W. S. Merwin, one of the most celebrated and prolific poets of his generation, died last Friday at the age of 91. A former U.S. poet laureate and the winner of two Pulitzer Prizes, Merwin was also known for his work as a conservationist. After moving to Hawaii in the 1970s, he began the decades-long project of restoring different plant species to the former pineapple plantation where he lived. Merwin was notoriously difficult to contact, but in the fall of last year, John Kaag managed to interview him over the phone. Merwin, he reports, repeated the same lines throughout their conversation: “The time of wisdom cannot be measured, and, for me, wisdom is the garden. There is no time in the garden.” In an essay for the Scholar, Kaag explores this relationship between gardening and the passage of time, and how it is reflected in Merwin’s life and poetry.

domain: literature


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- The Colorado Rockies learned the hard way about the importance of guardrails when commenting during a crisis. In a game earlier this week, broadcasters for the Miami Marlins claimed a Coors Field fan screamed a racial slur that was caught by its microphones, and the Rockies validated that before looking into it by posting to social media that it was “disgusted at the racial slur by a fan … .” The next morning, it became clear the fan had actually yelled “Dinger,” the name of the Rockies’ mascot, in an attempt to get a photo. Media quickly backtracked and blamed the Rockies for legitimizing the story with its social media post (as evidenced by this post from 9News’ Nicole Vap).
- Trust in government isn’t exactly high these days, and the Colorado Department of Transportation (CDOT) reinforced some of the worst suspicions of the tin-foil-hat crowd when it lied about Independence Pass being closed recently due to a mudslide. Media uncovered emails this week showing back-and-forth among CDOT officials about how to reduce traffic on the congested pass, and claiming a fictitious mudslide was the consensus winner. CDOT officials quickly realized two things: it is bad policy to lie, and that discussing it via email was a mistake. Officials reversed their decision after a day and discoverable email discussions were not part of that process .
- The Royal Family and Prince Andrew are in the spotlight once again this week because of Andrew’s affiliation with disgraced pedophile/sex criminal Jeffrey Epstein. A New York woman filed a lawsuit against Andrew this week alleging that he raped and sexually abused her when she was a minor. Andrew’s previous denials only made the situation worse and the Royal Family has remained silent on the lawsuit, signaling that they understand what a dumpster fire this entire situation is. Here’s hoping he becomes Prince Andrew, Duke of York Prison before this is all over.
- Nostalgia purveyor Garrison Keillor hasn’t been heard from much after he was booted from his home on Minnesota Public Radio in 2017 due to allegations of inappropriate behavior with a female assistant. But this week, he came roaring back, making headlines for equating the U.S. Social Security Administration with the Nazi SS after he got caught in a bureaucratic snafu trying to obtain a replacement Medicare card.
- The chile wars between Colorado (Pueblo chile peppers) and New Mexico (Hatch chile peppers) heated up this week when The Pueblo Chieftain ran syndicated content from its parent company, Gannett, that was pro-Hatch chile. Chieftain News Director Luke Lyons issued a formal apology distancing the newspaper from the story, saying in part, “The article … was not meant to cause harm or to infer that the Hatch chile was superior. … We apologize for the harm and offense the story has caused.” It’s not clear whether journalism purists are more offended that Lyons would apologize for content or that he confused “infer” for “imply” in his apology.
So, who won the week? I’m going with “Tiana,” a somewhat anonymous Tokyo Olympics volunteer. She paid for a taxi out of her own pocket when Jamaican hurdler Hansle Parchment took a wrong bus and mistakenly ended up at the aquatics venue. Thanks to her generous help, Parchment made it to the track stadium in time and subsequently won a gold medal in the men’s 110 meters hurdle event. Parchment was able to track down Tiana afterward to thank her and repay the fare, and Jamaica’s minister of tourism has now invited Tiana for an all-expenses-paid trip to the Caribbean island.

domain: news


Elderly people who are terminally ill have a ''duty to die and get out of the way'' instead of trying to prolong their lives by artificial means, Gov. Richard D. Lamm of Colorado said Tuesday.
People who die without having life artificially extended are similar to ''leaves falling off a tree and forming humus for the other plants to grow up,'' the Governor told a meeting of the Colorado Health Lawyers Association at St. Joseph's Hospital.
''You've got a duty to die and get out of the way,'' said the 48-year-old Governor. ''Let the other society, our kids, build a reasonable life.''
Some groups of the elderly immediately denounced Mr. Lamm for the statements. Reactions From Elderly
Dorothy Minkel, 75 years old, who has lobbied the Legislature for more than a decade on issues of the elderly, said, ''It is an insult for anyone to suggest we have a duty to die.''.
Another lobbyist who represents elderly groups, Robert Robinson, 68 years old, said, ''To say they have a duty to die and get out of the way brings me back 40 years ago when a person in Germany not only advocated that but carried it through.''
At a Senior Day gathering today, one elderly woman told Mr. Lamm, ''I hate you for what you have said.'' But others at the meeting in the First Baptist Church applauded the Governor after hearing his explanation. At the meeting with representatives of senior citizens groups, part of planned Senior Day activities at the Colorado capital, the Governor said, ''We are really approaching a time of almost technological immortatily when the machine and the tubes and the drugs and the heart pacemakers . . . literally force life on us.
''I believe we really should be very careful in terms of our technological miracles that we don't impose life on people who, in fact, are suffering beyoind the ability for us to help.'' In Support of Options
A member of the audience, Ferne Taylor of Denver, drew applause when she said, ''When I can't digest my food, when I can't breathe on my own, when my heart can't beat on its own, it could just be that God is trying to tell me something.''
Mr. Lamm, when asked, said he did not have any particular age in mind for the terminally ill to ''get out of the way'' and that the decision should be made by the ill person in consultation with doctors and family members.
The Governor said Tuesday that the costs of treatment that allows some terminally ill people to live longer was ruining the nation's economic health.
Steve Mehlman, a spokesman for the American Association of Retired Persons in Washington, D.C., said cutting off treatment to the terminally ill was no way to stem the rising cost of hospital care. ''It is not the elderly's fault,'' Mr. Mehlman said, ''We're the victims of health care inflation, not the cause.''
The Governor's comment was the latest in a series of controversial remarks by Mr. Lamm, who is in his third term as Governor. He recently angered Nevada officials by saying that one in eight women under the age of 45 in Las Vegas was a prostitute.


Security researchers discovered a security flaw in the NVIDIA Tegra X1 processor that is used by the Nintendo Switch console. The vulnerability makes it possible to run homebrew software on the Switch, and opens up the possibility of piracy. Direct physical access to the console is needed for exploitation, so the risk is minimal.
The flaw is described as unpatchable, as it is inside the boot and power management ROM of the Nvidia Tegra X1 used in the Switch. Users only need to initiate recovery mode on the console and the exploit can be triggered via a malformed packet sent during a routine USB device status check. Thence up to 65,535 bytes of extra data can be executed without question.
Nintendo will likely issue an update to mitigate the attack.


Yes…you read it right. Hitler was also pursuing the huge strategic potential that emanates out of a simple physics formula- E=mc*c.
He had an active nuclear program run at the kaiser Wilhem Institut in Berlin to develop the Nazi Atom bomb…no prizes for guessing which city would have replaced Hiroshima in the annals of history. India would have got her independence (at least from the British) by default. But jokes apart, Hitler had come very close to getting this prized weapon of mass destruction…but never got a finished product, ready for use….else world history would have been so different.
So, how was the Nazi A-Bomb plan derailed. Well, its a legend fit for a classic Hollywood war movie…and they did make a few based on the incidents narrated below.
British knew about Hitler’s plan and felt that the best way was to disrupt the supply of indispensable raw materials which are required for producing weapon grade uranium. One such material in those times was Deuterium Oxide or Heavy Water. It was being produced at a ‘Norsk Hydro’ plant in Vermock, Norway, hidden in a fjord, well protected by geographical features and only seven stories of reinforced concrete. Early in the war, with the available bombers, it was not possible to knock it out by bombing from air. It needed boots on ground.
The British launched two major commando operations to scuttle the heavy water production. First one failed miserably when the glider carrying commando group, crashed, killing the whole team. In those days, transport aircraft or bomber would tug a glider which was basically an aeroplane without engines. These were mainly used as troop carriers.
Second attack carried out by specially trained Norwegian resistance fighters (similar to the likes of Azad and Bhagat Singh in India) was successful as the team got inside the facility and blew up the huge containers which stored the Heavy Water. The team was able to escape too.
But the Germans were able to scale up the production in quick time. By then, the Allied had bombers capable of hitting the Norsk Hydro facility and the US Air Force bombers attacked the plant in late 1943. Although it didn’t damage the plant but still scared the Nazis. They decided to cut the losses and take whatever Heavy water had been produced by then, back to Germany. The trip included a ferry ride through Lake Tinn. That’s where the saboteurs saw their opportunity and planted a plastic explosive device, to be triggered by an alarm clock…set to explode when the ferry would be over the deepest part of the lake. Fortunately for the Allies, the bomb worked and SF Hydro, the ferry sank in 450 meters of water…so did the dream of Hitler to obtain a nuclear bomb.


Multiple guns fired at a Freeport bar
FREEPORT, Ill. (WIFR) - Freeport police say multiple guns were reportedly fired early Sunday at an area bar. Around 12:30 a.m., police arrived at Benchwarmers, 2143 W. Galena Avenue. Multiple people reportedly left the bar when officials arrived, and police have not specified if anyone is in custody. The guns...
Winnebago corrections officer dies just three weeks following her retirement
ROCKFORD, Ill. (WTVO) — The Winnebago County Sheriff’s Office is mourning the passing of corrections officer Ruth Raxter, three weeks after her retirement. According to the sheriff’s office, Raxter died Friday, April 19th. “Ruth began her career with the Winnebago County Sheriff’s Office in 2001 as a Corrections Officer and recently retired on March 29, […]
Multiple ambulances called to scene of serious crash on Whitman Street in Rockford
Police have closed the westbound lanes of Whitman Street beneath the North Second Street overpass for a crash.
Dixon man charged with fleeing police, DUI after high-speed chase
OGLE COUNTY, Ill. (WTVO) — Police in Ogle County arrested Andrew Gonzales, 22, of Dixon, after he allegedly fled police in a high-speed chase while high on drugs and alcohol. According to police, a deputy attempted to stop a blue Genesis in the 500 block of W. Flagg Road around 4:21 a.m. on Tuesday morning. […]
Police: Rockford mother hits child with a broom handle over broken plate
ROCKFORD, Ill. (WTVO) — Rockford Police arrested Brittany Lucas, 30, for allegedly hitting a child with a broom handle causing bruising. On April 3rd, the victim was at school and told the gym teacher she could not participate in class because her side hurt. She explained that she was hit by her mother. The school […]
Rockford Walmart stabbing death lawsuit filed in Chicago. Here's why
The decision to file a civil lawsuit over the fatal stabbing of a Rockford Walmart employee in Cook County instead of Winnebago County probably came down to a number of factors, legal experts said. For starters, it's allowed under the state's venue law. Since the defendants — Walmart and and its security company Brosnan...
Rockford Vietnam War veteran surprised with exterior home makeover
ROCKFORD, Ill. (WIFR) - A Vietnam War veteran known for helping others gets a helping hand of his own. Through a giveaway with Randy Palmeri & Son Contracting, Nick Parnello or “Sgt. Nick” will receive a new roof and siding, free of charge. Parnello served in the U.S. Army as a sergeant and co-founded the Vietnam Veterans Honor Society and Operation Fallen Flags. Parnello was also instrumental in the creation of the LZ Peace Memorial on Guilford Road in Rockford.
Few storms possible this afternoon, low severe threat
Temperatures have been on the up and up, peaking in the upper 60s Monday afternoon. With Monday’s 60° weather however came a rather robust southwesterly wind, gusting up to 35 mph at times. It’s because of this warm wind that temperatures are starting out much more comfortable this morning, with most sitting in the low to mid 50s.
Tracking another cold night tonight across the Rockford region
ROCKFORD, Ill. (WIFR) - Frost advisories have been issued across the stateline starting at 2 a.m. until 8 a.m. This is because temperatures are expected to drop close to freezing again tonight with mostly clear skies. So, as you are heading out the door Monday morning, you’ll need a jacket and some pants.
Taser used to arrest Rockford man after traffic stop
ROCKFORD, Ill. (WIFR) - Rockford police officers shocked a suspect with a Taser Sunday evening after a traffic stop. Jerome Garrett, 36, of Rockford is accused of armed habitual criminal, unlawful possession of a firearm by a felon, possession of a firearm by a felon, possession of a firearm by a gang member and resisting arrest.
Woman injured in shootout as gunmen exchange dozens of rounds at Rockford party rental
ROCKFORD, Ill. (WTVO) — A 20-year-old woman was shot as gunmen exchanged between 50 to 75 shots at a 20th Street building that had been rented for a party, witnesses said. According to Rockford Police, during the gathering, gunfire was exchanged between suspects from the inside and outside of the building, located 1416-1418 block of […]
Rockford firefighters rescue man from burning home
Rockford firefighters rescued a man hanging out of a second-floor window of a burning residence Monday afternoon. Firefighters responded about 4:25 p.m. to 317 Howard Ave. after neighboring residents reported smoke coming from the two-story house as well as an elderly man being trapped in the building on the second floor.
Rockford police arrest 2 after man shot in the leg
ROCKFORD, Ill. (WIFR) - Two men face gun charges after police say shots were fired during an argument early Saturday. James Jones, 23, is charged with aggravated discharge of a weapon. Victor Lloyd, 28, faces charges of aggravated discharge of a weapon, aggravated battery with a firearm and unlawful use of a weapon by a felon.


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The discovery of cranial movement stemmed from the osteopathic principle that structure and function are interrelated.
Exploring Cranial Movement and CST
In the 1920s, osteopathic student William Sutherland postulated that the cranial sutures were designed to allow movement, and if they didn’t allow cranial movement this could cause symptoms. Sutures in the skull are like expansion joints in a bridge. They allow a small amount of movement but prevent shearing. In this case the movement would be between the different cranial bones.
In the 1970’s, Dr. John Upledger, an osteopathic physician, witnessed the rhythmic movements of the dura during a spinal surgery in which he was assisting. The dura is a tough membrane that encases the brain and spinal cord, and is attached to the inside of the skull, the upper neck, the sacrum and the tailbone. Thanks to Dr. Upledger’s initial research and subsequent work, we now know that the soft tissues of the body normally move in concert with the rhythmic motion of the cranial bones.
A recent randomized controlled study with cross-over experimental design has demonstrated effectiveness of CST in the treatment of migraine. Although the sample size was small (n=20), the symptoms identified were similar to those patients diagnosed with concussion.
Robert Harris, RMT, of the Cranial Centre in Toronto, among others, have taken the principles and techniques of CST and developed them beyond what has been taught by Dr. Upledger at a basic level.
The Effects of Trauma on Connective Tissues
It is important to know that the delicate connective tissues of the body are very sensitive to trauma and may distort this rhythm. Several traumatic incidents may leave the body unable to adapt and compensate, creating symptoms. Sometimes these symptoms are far removed from where the connective tissue cause is. The connective tissue relates not just to the cranial bones and dura, but surrounds every tissue in the body down to the smallest cells. It is involved in both the soft and hard tissues of the body.
Also, due to the meandering nature of this connective tissue, as well as its electrical conductivity, the presentation of symptoms may be unusual. This may lead the untrained practitioner to believe that the patient is malingering. If symptoms or objective findings do not change within a reasonable period of time, there may be a connective tissue answer.
Manual Approach to Craniosacral Therapy
Craniosacral therapy has developed into a manual (using the hands) approach designed to identify restrictions, and gently restore motion and function to the entire body. It can often be the solution by itself. Sometimes treatment will simply allow the patient to get back on track so that they can be progressed with more conventional approaches or return to their exercise program.
In Brian’s clinical experience, he found that CST has relieved headaches, spinal pain and TMJ (jaw) pain, as well as patients’ symptoms from fibromyalgia, acute or recurrent injuries—athletic or otherwise—and those from whiplash or head injury/concussion, which are sometimes not helped with other methods of treatment.
So what does a session actually feel like? The most common “side effect” is a feeling of deep relaxation. This relaxation is deeply therapeutic and an important component of tissue healing. It feels very much like meditation or waking up from a deep and restful sleep. Robert calls it a specifically directed mini yoga session for the soft tissues of the body. CST is one of the osteopathic approaches Brian uses to assist clients on their path towards wellness.
*The above information was gathered from course material and clinical experience.

domain: health


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B7-H1/PD-L1, a member of the B7 family of immune-regulatory cell-surface proteins, plays an important role in the negative regulation of cell-mediated immune responses through its interaction with its receptor, programmed death-1 (PD-1) 1,2. Overexpression of B7-H1 by tumor cells has been noted in a number of human cancers, including melanoma, glioblastoma, and carcinomas of the lung, breast, colon, ovary, and renal cells, and has been shown to impair anti-tumor T-cell immunity3-8.
Recently, B7-H1 expression by pancreatic adenocarcinoma tissues has been identified as a potential prognostic marker9,10. Additionally, blockade of B7-H1 in a mouse model of pancreatic cancer has been shown to produce an anti-tumor response11. These data suggest the importance of B7-H1 as a potential therapeutic target. Anti-B7-H1 blockade antibodies are therefore being tested in clinical trials for multiple human solid tumors including melanoma and cancers of lung, colon, kidney, stomach and pancreas12.
In order to eventually be able to identify the patients who will benefit from B7-H1 targeting therapies, it is critical to investigate the correlation between expression and localization of B7-H1 and patient response to treatment with B7-H1 blockade antibodies. Examining the expression of B7-H1 in human pancreatic adenocarcinoma tissues through immunohistochemistry will give a better understanding of how this co-inhibitory signaling molecule contributes to the suppression of antitumor immunity in the tumor's microenvironment. The anti-B7-H1 monoclonal antibody (clone 5H1) developed by Chen and coworkers has been shown to produce reliable staining results in cryosections of multiple types of human neoplastic tissues4,8, but staining on paraffin-embedded slides had been a challenge until recently13-18. We have developed the B7-H1 staining protocol for paraffin-embedded slides of pancreatic adenocarcinoma tissues. The B7-H1 staining protocol described here produces consistent membranous and cytoplasmic staining of B7-H1 with little background.
21 Related JoVE Articles!
The Use of Reverse Phase Protein Arrays (RPPA) to Explore Protein Expression Variation within Individual Renal Cell Cancers
Institutions: University of Edinburgh, University of St Andrews, University of Edinburgh, University of Edinburgh, Western General Hospital, University of Edinburgh, Queen Mary University of London.
Currently there is no curative treatment for metastatic clear cell renal cell cancer, the commonest variant of the disease. A key factor in this treatment resistance is thought to be the molecular complexity of the disease 1
. Targeted therapy such as the tyrosine kinase inhibitor (TKI)-sunitinib have been utilized, but only 40% of patients will respond, with the overwhelming majority of these patients relapsing within 1 year 2
. As such the question of intrinsic and acquired resistance in renal cell cancer patients is highly relevant 3
In order to study resistance to TKIs, with the ultimate goal of developing effective, personalized treatments, sequential tissue after a specific period of targeted therapy is required, an approach which had proved successful in chronic myeloid leukaemia 4
. However the application of such a strategy in renal cell carcinoma is complicated by the high level of both inter- and intratumoral heterogeneity, which is a feature of renal cell carcinoma5,6
as well as other solid tumors 7
. Intertumoral heterogeneity due to transcriptomic and genetic differences is well established even in patients with similar presentation, stage and grade of tumor. In addition it is clear that there is great morphological (intratumoral) heterogeneity in RCC, which is likely to represent even greater molecular heterogeneity. Detailed mapping and categorization of RCC tumors by combined morphological analysis and Fuhrman grading allows the selection of representative areas for proteomic analysis.
Protein based analysis of RCC8
is attractive due to its widespread availability in pathology laboratories; however, its application can be problematic due to the limited availability of specific antibodies 9
. Due to the dot blot nature of the Reverse Phase Protein Arrays (RPPA), antibody specificity must be pre-validated; as such strict quality control of antibodies used is of paramount importance. Despite this limitation the dot blot format does allow assay miniaturization, allowing for the printing of hundreds of samples onto a single nitrocellulose slide. Printed slides can then be analyzed in a similar fashion to Western analysis with the use of target specific primary antibodies and fluorescently labelled secondary antibodies, allowing for multiplexing. Differential protein expression across all the samples on a slide can then be analyzed simultaneously by comparing the relative level of fluorescence in a more cost-effective and high-throughput manner.
Cancer Biology, Issue 71, Bioengineering, Medicine, Biomedical Engineering, Cellular Biology, Molecular Biology, Genetics, Pathology, Oncology, Proteins, Early Detection of Cancer, Translational Medical Research, RPPA, RCC, Heterogeneity, Proteomics, Tumor Grade, intertumoral, tumor, metastatic, carcinoma, renal cancer, clear cell renal cell cancer, cancer, assay
Minimally Invasive Establishment of Murine Orthotopic Bladder Xenografts
Institutions: University of British Columbia.
Orthotopic bladder cancer xenografts are the gold standard to study molecular cellular manipulations and new therapeutic agents in vivo
. Suitable cell lines are inoculated either by intravesical instillation (model of nonmuscle invasive growth) or intramural injection into the bladder wall (model of invasive growth). Both procedures are complex and highly time-consuming. Additionally, the superficial model has its shortcomings due to the lack of cell lines that are tumorigenic following instillation. Intramural injection, on the other hand, is marred by the invasiveness of the procedure and the associated morbidity for the host mouse.
With these shortcomings in mind, we modified previous methods to develop a minimally invasive approach for creating orthotopic bladder cancer xenografts. Using ultrasound guidance we have successfully performed percutaneous inoculation of the bladder cancer cell lines UM-UC1, UM-UC3 and UM-UC13 into 50 athymic nude. We have been able to demonstrate that this approach is time efficient, precise and safe. With this technique, initially a space is created under the bladder mucosa with PBS, and tumor cells are then injected into this space in a second step. Tumor growth is monitored at regular intervals with bioluminescence imaging and ultrasound. The average tumor volumes increased steadily in in all but one of our 50 mice over the study period.
In our institution, this novel approach, which allows bladder cancer xenograft inoculation in a minimally-invasive, rapid and highly precise way, has replaced the traditional model.
Medicine, Issue 84, Bladder cancer, cell lines, xenograft, inoculation, ultrasound, orthotopic model
A Matrigel-Based Tube Formation Assay to Assess the Vasculogenic Activity of Tumor Cells
Institutions: University of Massachusetts, University of Massachusetts, University of Massachusetts.
Over the past several decades, a tube formation assay using growth factor-reduced Matrigel has been typically employed to demonstrate the angiogenic activity of vascular endothelial cells in vitro1-5
. However, recently growing evidence has shown that this assay is not limited to test vascular behavior for endothelial cells. Instead, it also has been used to test the ability of a number of tumor cells to develop a vascular phenotype6-8
. This capability was consistent with their vasculogenic behavior identified in xenotransplanted animals, a process known as vasculogenic mimicry (VM)9
. There is a multitude of evidence demonstrating that tumor cell-mediated VM plays a vital role in the tumor development, independent of endothelial cell angiogenesis6, 10-13
. For example, tumor cells were found to participate in the blood perfused, vascular channel formation in tissue samples from melanoma and glioblastoma patients8, 10, 11
. Here, we described this tubular network assay as a useful tool in evaluation of vasculogenic activity of tumor cells. We found that some tumor cell lines such as melanoma B16F1 cells, glioblastoma U87 cells, and breast cancer MDA-MB-435 cells are able to form vascular tubules; but some do not such as colon cancer HCT116 cells. Furthermore, this vascular phenotype is dependent on cell numbers plated on the Matrigel. Therefore, this assay may serve as powerful utility to screen the vascular potential of a variety of cell types including vascular cells, tumor cells as well as other cells.
Cancer Biology, Issue 55, tumor, vascular, endothelial, tube formation, Matrigel, in vitro
Thermal Ablation for the Treatment of Abdominal Tumors
Institutions: University of Wisconsin-Madison, University of Wisconsin-Madison.
Percutaneous thermal ablation is an emerging treatment option for many tumors of the abdomen not amenable to conventional treatments. During a thermal ablation procedure, a thin applicator is guided into the target tumor under imaging guidance. Energy is then applied to the tissue until temperatures rise to cytotoxic levels (50-60 °C). Various energy sources are available to heat biological tissues, including radiofrequency (RF) electrical current, microwaves, laser light and ultrasonic waves. Of these, RF and microwave ablation are most commonly used worldwide.
During RF ablation, alternating electrical current (~500 kHz) produces resistive heating around the interstitial electrode. Skin surface electrodes (ground pads) are used to complete the electrical circuit. RF ablation has been in use for nearly 20 years, with good results for local tumor control, extended survival and low complication rates1,2
. Recent studies suggest RF ablation may be a first-line treatment option for small hepatocellular carcinoma and renal-cell carcinoma3-5
. However, RF heating is hampered by local blood flow and high electrical impedance tissues (eg, lung, bone, desiccated or charred tissue)6,7
. Microwaves may alleviate some of these problems by producing faster, volumetric heating8-10
. To create larger or conformal ablations, multiple microwave antennas can be used simultaneously while RF electrodes require sequential operation, which limits their efficiency. Early experiences with microwave systems suggest efficacy and safety similar to, or better than RF devices11-13
Alternatively, cryoablation freezes the target tissues to lethal levels (-20 to -40 °C). Percutaneous cryoablation has been shown to be effective against RCC and many metastatic tumors, particularly colorectal cancer, in the liver14-16
. Cryoablation may also be associated with less post-procedure pain and faster recovery for some indications17
. Cryoablation is often contraindicated for primary liver cancer due to underlying coagulopathy and associated bleeding risks frequently seen in cirrhotic patients. In addition, sudden release of tumor cellular contents when the frozen tissue thaws can lead to a potentially serious condition known as cryoshock 16
Thermal tumor ablation can be performed at open surgery, laparoscopy or using a percutaneous approach. When performed percutaneously, the ablation procedure relies on imaging for diagnosis, planning, applicator guidance, treatment monitoring and follow-up. Ultrasound is the most popular modality for guidance and treatment monitoring worldwide, but computed tomography (CT) and magnetic resonance imaging (MRI) are commonly used as well. Contrast-enhanced CT or MRI are typically employed for diagnosis and follow-up imaging.
Medicine, Issue 49, Thermal ablation, interventional oncology, image-guided therapy, radiology, cancer
Enhancement of Apoptotic and Autophagic Induction by a Novel Synthetic C-1 Analogue of 7-deoxypancratistatin in Human Breast Adenocarcinoma and Neuroblastoma Cells with Tamoxifen
Institutions: University of Windsor, Brock University.
Breast cancer is one of the most common cancers amongst women in North America. Many current anti-cancer treatments, including ionizing radiation, induce apoptosis via DNA damage. Unfortunately, such treatments are non-selective to cancer cells and produce similar toxicity in normal cells. We have reported selective induction of apoptosis in cancer cells by the natural compound pancratistatin (PST). Recently, a novel PST analogue, a C-1 acetoxymethyl derivative of 7-deoxypancratistatin (JCTH-4), was produced by de novo synthesis and it exhibits comparable selective apoptosis inducing activity in several cancer cell lines. Recently, autophagy has been implicated in malignancies as both pro-survival and pro-death mechanisms in response to chemotherapy. Tamoxifen (TAM) has invariably demonstrated induction of pro-survival autophagy in numerous cancers. In this study, the efficacy of JCTH-4 alone and in combination with TAM to induce cell death in human breast cancer (MCF7) and neuroblastoma (SH-SY5Y) cells was evaluated. TAM alone induced autophagy, but insignificant cell death whereas JCTH-4 alone caused significant induction of apoptosis with some induction of autophagy. Interestingly, the combinatory treatment yielded a drastic increase in apoptotic and autophagic induction. We monitored time-dependent morphological changes in MCF7 cells undergoing TAM-induced autophagy, JCTH-4-induced apoptosis and autophagy, and accelerated cell death with combinatorial treatment using time-lapse microscopy. We have demonstrated these compounds to induce apoptosis/autophagy by mitochondrial targeting in these cancer cells. Importantly, these treatments did not affect the survival of noncancerous human fibroblasts. Thus, these results indicate that JCTH-4 in combination with TAM could be used as a safe and very potent anti-cancer therapy against breast cancer and neuroblastoma cells.
Cancer Biology, Issue 63, Medicine, Biochemistry, Breast adenocarcinoma, neuroblastoma, tamoxifen, combination therapy, apoptosis, autophagy
Quantitative Analysis of Autophagy using Advanced 3D Fluorescence Microscopy
Institutions: University of California, Davis , University of California, Davis , University of Tromsø, University of California, Davis , University of California, Davis , University of California, Davis .
Prostate cancer is the leading form of malignancies among men in the U.S. While surgery carries a significant risk of impotence and incontinence, traditional chemotherapeutic approaches have been largely unsuccessful. Hormone therapy is effective at early stage, but often fails with the eventual development of hormone-refractory tumors. We have been interested in developing therapeutics targeting specific metabolic deficiency of tumor cells. We recently showed that prostate tumor cells specifically lack an enzyme (argininosuccinate synthase, or ASS) involved in the synthesis of the amino acid arginine1
. This condition causes the tumor cells to become dependent on exogenous arginine, and they undergo metabolic stress when free arginine is depleted by arginine deiminase (ADI)1,10
. Indeed, we have shown that human prostate cancer cells CWR22Rv1
are effectively killed by ADI with caspase-independent apoptosis and aggressive autophagy
. Autophagy is an evolutionarily-conserved process that allows cells to metabolize unwanted proteins by lysosomal breakdown during nutritional starvation4,5
. Although the essential components of this pathway are well-characterized6,7,8,9
, many aspects of the molecular mechanism are still unclear - in particular, what is the role of autophagy in the death-response of prostate cancer cells after ADI treatment? In order to address this question, we required an experimental method to measure the level and extent of autophagic response in cells - and since there are no known molecular markers that can accurately track this process, we chose to develop an imaging-based approach, using quantitative 3D fluorescence microscopy11,12
Using CWR22Rv1 cells specifically-labeled with fluorescent probes for autophagosomes and lysosomes, we show that 3D image stacks acquired with either widefield deconvolution microscopy (and later, with super-resolution, structured-illumination microscopy) can clearly capture the early stages of autophagy induction. With commercially available digital image analysis applications, we can readily obtain statistical information about autophagosome and lysosome number, size, distribution, and degree of colocalization from any imaged cell. This information allows us to precisely track the progress of autophagy in living cells and enables our continued investigation into the role of autophagy in cancer chemotherapy.
Cellular Biology, Issue 75, Biochemistry, Molecular Biology, Medicine, Cancer Biology, Biophysics, Chemical Biology, Proteins, Microscopy, Fluorescence, autophagy, arginine deiminase, prostate cancer, deconvolution microscopy, super-resolution structured-illumination microscopy, live cell imaging, tumors, autophagosomes, lysosomes, cells, cell culture, microscopy, imaging, visualization
Use of Shigella flexneri to Study Autophagy-Cytoskeleton Interactions
Institutions: Imperial College London, Institut Pasteur, Unité Macrophages et Développement de l'Immunité.
is an intracellular pathogen that can escape from phagosomes to reach the cytosol, and polymerize the host actin cytoskeleton to promote its motility and dissemination. New work has shown that proteins involved in actin-based motility are also linked to autophagy, an intracellular degradation process crucial for cell autonomous immunity. Strikingly, host cells may prevent actin-based motility of S. flexneri
by compartmentalizing bacteria inside ‘septin cages’ and targeting them to autophagy. These observations indicate that a more complete understanding of septins, a family of filamentous GTP-binding proteins, will provide new insights into the process of autophagy. This report describes protocols to monitor autophagy-cytoskeleton interactions caused by S. flexneri in vitro
using tissue culture cells and in vivo
using zebrafish larvae. These protocols enable investigation of intracellular mechanisms that control bacterial dissemination at the molecular, cellular, and whole organism level.
Infection, Issue 91, ATG8/LC3, autophagy, cytoskeleton, HeLa cells, p62, septin, Shigella, zebrafish
A Mouse Tumor Model of Surgical Stress to Explore the Mechanisms of Postoperative Immunosuppression and Evaluate Novel Perioperative Immunotherapies
Institutions: Ottawa Hospital Research Institute, University of Ottawa, University of Ottawa, The Second Hospital of Shandong University, University of Tabuk, Ottawa General Hospital.
Surgical resection is an essential treatment for most cancer patients, but surgery induces dysfunction in the immune system and this has been linked to the development of metastatic disease in animal models and in cancer patients. Preclinical work from our group and others has demonstrated a profound suppression of innate immune function, specifically NK cells in the postoperative period and this plays a major role in the enhanced development of metastases following surgery. Relatively few animal studies and clinical trials have focused on characterizing and reversing the detrimental effects of cancer surgery. Using a rigorous animal model of spontaneously metastasizing tumors and surgical stress, the enhancement of cancer surgery on the development of lung metastases was demonstrated. In this model, 4T1 breast cancer cells are implanted in the mouse mammary fat pad. At day 14 post tumor implantation, a complete resection of the primary mammary tumor is performed in all animals. A subset of animals receives additional surgical stress in the form of an abdominal nephrectomy. At day 28, lung tumor nodules are quantified. When immunotherapy was given immediately preoperatively, a profound activation of immune cells which prevented the development of metastases following surgery was detected. While the 4T1 breast tumor surgery model allows for the simulation of the effects of abdominal surgical stress on tumor metastases, its applicability to other tumor types needs to be tested. The current challenge is to identify safe and promising immunotherapies in preclinical mouse models and to translate them into viable perioperative therapies to be given to cancer surgery patients to prevent the recurrence of metastatic disease.
Medicine, Issue 85, mouse, tumor model, surgical stress, immunosuppression, perioperative immunotherapy, metastases
Renal Capsule Xenografting and Subcutaneous Pellet Implantation for the Evaluation of Prostate Carcinogenesis and Benign Prostatic Hyperplasia
Institutions: University of Wisconsin-Madison, University of Rochester School of Medicine & Dentistry, University of Wisconsin-Madison.
New therapies for two common prostate diseases, prostate cancer (PrCa) and benign prostatic hyperplasia (BPH), depend critically on experiments evaluating their hormonal regulation. Sex steroid hormones (notably androgens and estrogens) are important in PrCa and BPH; we probe their respective roles in inducing prostate growth and carcinogenesis in mice with experiments using compressed hormone pellets. Hormone and/or drug pellets are easily manufactured with a pellet press, and surgically implanted into the subcutaneous tissue of the male mouse host. We also describe a protocol for the evaluation of hormonal carcinogenesis by combining subcutaneous hormone pellet implantation with xenografting of prostate cell recombinants under the renal capsule of immunocompromised mice. Moreover, subcutaneous hormone pellet implantation, in combination with renal capsule xenografting of BPH tissue, is useful to better understand hormonal regulation of benign prostate growth, and to test new therapies targeting sex steroid hormone pathways.
Medicine, Issue 78, Cancer Biology, Prostatic Hyperplasia, Prostatic Neoplasms, Neoplastic Processes, Estradiol, Testosterone, Transplantation, Heterologous, Growth, Xenotransplantation, Heterologous Transplantation, Hormones, Prostate, Testosterone, 17beta-Estradiol, Benign prostatic hyperplasia, Prostate Cancer, animal model
Experimental Metastasis and CTL Adoptive Transfer Immunotherapy Mouse Model
Institutions: Medical College of Georgia.
Experimental metastasis mouse model is a simple and yet physiologically relevant metastasis model. The tumor cells are injected intravenously (i.v) into mouse tail veins and colonize in the lungs, thereby, resembling the last steps of tumor cell spontaneous metastasis: survival in the circulation, extravasation and colonization in the distal organs. From a therapeutic point of view, the experimental metastasis model is the simplest and ideal model since the target of therapies is often the end point of metastasis: established metastatic tumor in the distal organ. In this model, tumor cells are injected i.v into mouse tail veins and allowed to colonize and grow in the lungs. Tumor-specific CTLs are then injected i.v into the metastases-bearing mouse. The number and size of the lung metastases can be controlled by the number of tumor cells to be injected and the time of tumor growth. Therefore, various stages of metastasis, from minimal metastasis to extensive metastasis, can be modeled. Lung metastases are analyzed by inflation with ink, thus allowing easier visual observation and quantification.
Immunology, Issue 45, Metastasis, CTL adoptive transfer, Lung, Tumor Immunology
Optimization of High Grade Glioma Cell Culture from Surgical Specimens for Use in Clinically Relevant Animal Models and 3D Immunochemistry
Institutions: Henry Ford Hospital.
Glioblastomas, the most common and aggressive form of astrocytoma, are refractory to therapy, and molecularly heterogeneous. The ability to establish cell cultures that preserve the genomic profile of the parental tumors, for use in patient specific in vitro
and in vivo
models, has the potential to revolutionize the preclinical development of new treatments for glioblastoma tailored to the molecular characteristics of each tumor.
Starting with fresh high grade astrocytoma tumors dissociated into single cells, we use the neurosphere assay as an enrichment method for cells presenting cancer stem cell phenotype, including expression of neural stem cell markers, long term self-renewal in vitro
, and the ability to form orthotopic xenograft tumors. This method has been previously proposed, and is now in use by several investigators. Based on our experience of dissociating and culturing 125 glioblastoma specimens, we arrived at the detailed protocol we present here, suitable for routine neurosphere culturing of high grade astrocytomas and large scale expansion of tumorigenic cells for preclinical studies. We report on the efficiency of successful long term cultures using this protocol and suggest affordable alternatives for culturing dissociated glioblastoma cells that fail to grow as neurospheres. We also describe in detail a protocol for preserving the neurospheres 3D architecture for immunohistochemistry. Cell cultures enriched in CSCs, capable of generating orthotopic xenograft models that preserve the molecular signatures and heterogeneity of GBMs, are becoming increasingly popular for the study of the biology of GBMs and for the improved design of preclinical testing of potential therapies.
Medicine, Issue 83, Primary Cell Culture, animal models, Nervous System Diseases, Neoplasms, glioblastoma, neurosphere, surgical specimens, long-term self-renewal
Modeling Spontaneous Metastatic Renal Cell Carcinoma (mRCC) in Mice Following Nephrectomy
Institutions: Roswell Park Cancer Institute, Sunnybrook Research Institute.
One of the key challenges to improved testing of new experimental therapeutics in renal cell carcinoma (RCC) is the development of models that faithfully recapitulate early- and late-stage metastatic disease progression. Typical tumor implantation models utilize ectopic or orthotopic primary tumor implantation, but few include systemic spontaneous metastatic disease that mimics the clinical setting. This protocol describes the key steps to develop RCC disease progression stages similar to patients. First, it uses a highly metastatic mouse tumor cell line in a syngeneic model to show orthotopic tumor cell implantation. Methods include superficial and internal implantation into the sub-capsular space with cells combined with matrigel to prevent leakage and early spread. Next it describes the procedures for excision of tumor-bearing kidney (nephrectomy), with critical pre- and post- surgical mouse care. Finally, it outlines the steps necessary to monitor and assess micro-and macro-metastatic disease progression, including bioluminescent imaging as well provides a detailed visual necropsy guide to score systemic disease distribution. The goal of this protocol description is to facilitate the widespread use of clinically relevant metastatic RCC models to improve the predictive value of future therapeutic testing.
Medicine, Issue 86, Spontaneous metastasis, orthotopic, nephrectomy, renal cell carcinoma, RCC, necropsy, kidney, bioluminescence, sub-capsular
Analysis of Nephron Composition and Function in the Adult Zebrafish Kidney
Institutions: University of Notre Dame.
The zebrafish model has emerged as a relevant system to study kidney development, regeneration and disease. Both the embryonic and adult zebrafish kidneys are composed of functional units known as nephrons, which are highly conserved with other vertebrates, including mammals. Research in zebrafish has recently demonstrated that two distinctive phenomena transpire after adult nephrons incur damage: first, there is robust regeneration within existing nephrons that replaces the destroyed tubule epithelial cells; second, entirely new nephrons are produced from renal progenitors in a process known as neonephrogenesis. In contrast, humans and other mammals seem to have only a limited ability for nephron epithelial regeneration. To date, the mechanisms responsible for these kidney regeneration phenomena remain poorly understood. Since adult zebrafish kidneys undergo both nephron epithelial regeneration and neonephrogenesis, they provide an outstanding experimental paradigm to study these events. Further, there is a wide range of genetic and pharmacological tools available in the zebrafish model that can be used to delineate the cellular and molecular mechanisms that regulate renal regeneration. One essential aspect of such research is the evaluation of nephron structure and function. This protocol describes a set of labeling techniques that can be used to gauge renal composition and test nephron functionality in the adult zebrafish kidney. Thus, these methods are widely applicable to the future phenotypic characterization of adult zebrafish kidney injury paradigms, which include but are not limited to, nephrotoxicant exposure regimes or genetic methods of targeted cell death such as the nitroreductase mediated cell ablation technique. Further, these methods could be used to study genetic perturbations in adult kidney formation and could also be applied to assess renal status during chronic disease modeling.
Cellular Biology, Issue 90,
zebrafish; kidney; nephron; nephrology; renal; regeneration; proximal tubule; distal tubule; segment; mesonephros; physiology; acute kidney injury (AKI)
Modeling Astrocytoma Pathogenesis In Vitro and In Vivo Using Cortical Astrocytes or Neural Stem Cells from Conditional, Genetically Engineered Mice
Institutions: University of North Carolina School of Medicine, University of North Carolina School of Medicine, University of North Carolina School of Medicine, University of North Carolina School of Medicine, University of North Carolina School of Medicine, Emory University School of Medicine, University of North Carolina School of Medicine.
Current astrocytoma models are limited in their ability to define the roles of oncogenic mutations in specific brain cell types during disease pathogenesis and their utility for preclinical drug development. In order to design a better model system for these applications, phenotypically wild-type cortical astrocytes and neural stem cells (NSC) from conditional, genetically engineered mice (GEM) that harbor various combinations of floxed oncogenic alleles were harvested and grown in culture. Genetic recombination was induced in vitro
using adenoviral Cre-mediated recombination, resulting in expression of mutated oncogenes and deletion of tumor suppressor genes. The phenotypic consequences of these mutations were defined by measuring proliferation, transformation, and drug response in vitro
. Orthotopic allograft models, whereby transformed cells are stereotactically injected into the brains of immune-competent, syngeneic littermates, were developed to define the role of oncogenic mutations and cell type on tumorigenesis in vivo
. Unlike most established human glioblastoma cell line xenografts, injection of transformed GEM-derived cortical astrocytes into the brains of immune-competent littermates produced astrocytomas, including the most aggressive subtype, glioblastoma, that recapitulated the histopathological hallmarks of human astrocytomas, including diffuse invasion of normal brain parenchyma. Bioluminescence imaging of orthotopic allografts from transformed astrocytes engineered to express luciferase was utilized to monitor in vivo
tumor growth over time. Thus, astrocytoma models using astrocytes and NSC harvested from GEM with conditional oncogenic alleles provide an integrated system to study the genetics and cell biology of astrocytoma pathogenesis in vitro
and in vivo
and may be useful in preclinical drug development for these devastating diseases.
Neuroscience, Issue 90, astrocytoma, cortical astrocytes, genetically engineered mice, glioblastoma, neural stem cells, orthotopic allograft
Initiation of Metastatic Breast Carcinoma by Targeting of the Ductal Epithelium with Adenovirus-Cre: A Novel Transgenic Mouse Model of Breast Cancer
Institutions: Wistar Institute, University of Pennsylvania, Geisel School of Medicine at Dartmouth, University of Pennsylvania, University of Pennsylvania, University of Pennsylvania.
Breast cancer is a heterogeneous disease involving complex cellular interactions between the developing tumor and immune system, eventually resulting in exponential tumor growth and metastasis to distal tissues and the collapse of anti-tumor immunity. Many useful animal models exist to study breast cancer, but none completely recapitulate the disease progression that occurs in humans. In order to gain a better understanding of the cellular interactions that result in the formation of latent metastasis and decreased survival, we have generated an inducible transgenic mouse model of YFP-expressing ductal carcinoma that develops after sexual maturity in immune-competent mice and is driven by consistent, endocrine-independent oncogene expression. Activation of YFP, ablation of p53, and expression of an oncogenic form of K-ras was achieved by the delivery of an adenovirus expressing Cre-recombinase into the mammary duct of sexually mature, virgin female mice. Tumors begin to appear 6 weeks after the initiation of oncogenic events. After tumors become apparent, they progress slowly for approximately two weeks before they begin to grow exponentially. After 7-8 weeks post-adenovirus injection, vasculature is observed connecting the tumor mass to distal lymph nodes, with eventual lymphovascular invasion of YFP+ tumor cells to the distal axillary lymph nodes. Infiltrating leukocyte populations are similar to those found in human breast carcinomas, including the presence of αβ and γδ T cells, macrophages and MDSCs. This unique model will facilitate the study of cellular and immunological mechanisms involved in latent metastasis and dormancy in addition to being useful for designing novel immunotherapeutic interventions to treat invasive breast cancer.
Medicine, Issue 85, Transgenic mice, breast cancer, metastasis, intraductal injection, latent mutations, adenovirus-Cre
Induction of Invasive Transitional Cell Bladder Carcinoma in Immune Intact Human MUC1 Transgenic Mice: A Model for Immunotherapy Development
Institutions: University of California, Davis, University of California, Davis, Merck KGaA, Darmstadt, Germany.
A preclinical model of invasive bladder cancer was developed in human mucin 1 (MUC1) transgenic (MUC1.Tg) mice for the purpose of evaluating immunotherapy and/or cytotoxic chemotherapy. To induce bladder cancer, C57BL/6 mice (MUC1.Tg and wild type) were treated orally with the carcinogen N-butyl-N-(4-hydroxybutyl)nitrosamine (OH-BBN) at 3.0 mg/day, 5 days/week for 12 weeks. To assess the effects of OH-BBN on serum cytokine profile during tumor development, whole blood was collected via submandibular bleeds prior to treatment and every four weeks. In addition, a MUC1-targeted peptide vaccine and placebo were administered to groups of mice weekly for eight weeks. Multiplex fluorometric microbead immunoanalyses of serum cytokines during tumor development and following vaccination were performed. At termination, interferon gamma (IFN-γ)/interleukin-4 (IL-4) ELISpot analysis for MUC1 specific T-cell immune response and histopathological evaluations of tumor type and grade were performed. The results showed that: (1) the incidence of bladder cancer in both MUC1.Tg and wild type mice was 67%; (2) transitional cell carcinomas (TCC) developed at a 2:1 ratio compared to squamous cell carcinomas (SCC); (3) inflammatory cytokines increased with time during tumor development; and (4) administration of the peptide vaccine induces a Th1-polarized serum cytokine profile and a MUC1 specific T-cell response. All tumors in MUC1.Tg mice were positive for MUC1 expression, and half of all tumors in MUC1.Tg and wild type mice were invasive. In conclusion, using a team approach through the coordination of the efforts of pharmacologists, immunologists, pathologists and molecular biologists, we have developed an immune intact transgenic mouse model of bladder cancer that expresses hMUC1.
Medicine, Issue 80, Urinary Bladder, Animals, Genetically Modified, Cancer Vaccines, Immunotherapy, Animal Experimentation, Models, Neoplasms Bladder Cancer, C57BL/6 Mouse, MUC1, Immunotherapy, Preclinical Model
Generation of Subcutaneous and Intrahepatic Human Hepatocellular Carcinoma Xenografts in Immunodeficient Mice
Institutions: University Health Network, University Health Network, University Health Network.
experimental models of hepatocellular carcinoma (HCC) that recapitulate the human disease provide a valuable platform for research into disease pathophysiology and for the preclinical evaluation of novel therapies. We present a variety of methods to generate subcutaneous or orthotopic human HCC xenografts in immunodeficient mice that could be utilized in a variety of research applications. With a focus on the use of primary tumor tissue from patients undergoing surgical resection as a starting point, we describe the preparation of cell suspensions or tumor fragments for xenografting. We describe specific techniques to xenograft these tissues i) subcutaneously; or ii) intrahepatically, either by direct implantation of tumor cells or fragments into the liver, or indirectly by injection of cells into the mouse spleen. We also describe the use of partial resection of the native mouse liver at the time of xenografting as a strategy to induce a state of active liver regeneration in the recipient mouse that may facilitate the intrahepatic engraftment of primary human tumor cells. The expected results of these techniques are illustrated. The protocols described have been validated using primary human HCC samples and xenografts, which typically perform less robustly than the well-established human HCC cell lines that are widely used and frequently cited in the literature. In comparison with cell lines, we discuss factors which may contribute to the relatively low chance of primary HCC engraftment in xenotransplantation models and comment on technical issues that may influence the kinetics of xenograft growth. We also suggest methods that should be applied to ensure that xenografts obtained accurately resemble parent HCC tissues.
Medicine, Issue 79, Liver Neoplasms, Hepatectomy, animal models, hepatocellular carcinoma, xenograft, cancer, liver, subcutaneous, intrahepatic, orthotopic, mouse, human, immunodeficient
Live Imaging of Drug Responses in the Tumor Microenvironment in Mouse Models of Breast Cancer
Institutions: Watson School of Biological Sciences, Cold Spring Harbor Laboratory, University of Oslo and Oslo University Hospital.
The tumor microenvironment plays a pivotal role in tumor initiation, progression, metastasis, and the response to anti-cancer therapies. Three-dimensional co-culture systems are frequently used to explicate tumor-stroma interactions, including their role in drug responses. However, many of the interactions that occur in vivo
in the intact microenvironment cannot be completely replicated in these in vitro
settings. Thus, direct visualization of these processes in real-time has become an important tool in understanding tumor responses to therapies and identifying the interactions between cancer cells and the stroma that can influence these responses. Here we provide a method for using spinning disk confocal microscopy of live, anesthetized mice to directly observe drug distribution, cancer cell responses and changes in tumor-stroma interactions following administration of systemic therapy in breast cancer models. We describe procedures for labeling different tumor components, treatment of animals for observing therapeutic responses, and the surgical procedure for exposing tumor tissues for imaging up to 40 hours. The results obtained from this protocol are time-lapse movies, in which such processes as drug infiltration, cancer cell death and stromal cell migration can be evaluated using image analysis software.
Cancer Biology, Issue 73, Medicine, Molecular Biology, Cellular Biology, Biomedical Engineering, Genetics, Oncology, Pharmacology, Surgery, Tumor Microenvironment, Intravital imaging, chemotherapy, Breast cancer, time-lapse, mouse models, cancer cell death, stromal cell migration, cancer, imaging, transgenic, animal model
Monitoring Tumor Metastases and Osteolytic Lesions with Bioluminescence and Micro CT Imaging
Institutions: Caliper Life Sciences.
Following intracardiac delivery of MDA-MB-231-luc-D3H2LN cells to Nu/Nu mice, systemic metastases developed in the injected animals. Bioluminescence imaging using IVIS Spectrum was employed to monitor the distribution and development of the tumor cells following the delivery procedure including DLIT reconstruction to measure the tumor signal and its location.
Development of metastatic lesions to the bone tissues triggers osteolytic activity and lesions to tibia and femur were evaluated longitudinally using micro CT. Imaging was performed using a Quantum FX micro CT system with fast imaging and low X-ray dose. The low radiation dose allows multiple imaging sessions to be performed with a cumulative X-ray dosage far below LD50. A mouse imaging shuttle device was used to sequentially image the mice with both IVIS Spectrum and Quantum FX achieving accurate animal positioning in both the bioluminescence and CT images. The optical and CT data sets were co-registered in 3-dimentions using the Living Image 4.1 software. This multi-mode approach allows close monitoring of tumor growth and development simultaneously with osteolytic activity.
Medicine, Issue 50, osteolytic lesions, micro CT, tumor, bioluminescence, in vivo, imaging, IVIS, luciferase, low dose, co-registration, 3D reconstruction
An Orthotopic Model of Murine Bladder Cancer
Institutions: Tulane University, Tulane University.
In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and subsequent cell adhesion. After their bladders are transurethrally catheterized and drained, animals are again catheterized to permit insertion of a platinum wire into bladders without damaging the urethra or bladder. The catheters are made of Teflon to serve as an insulator for the wire, which will conduct electrical current into the bladder to create a burn injury. An electrocautery unit is used to deliver 2.5W to the exposed end of the wire, burning away extracellular layers and providing attachment sites for carcinoma cells that are delivered in suspension to the bladder through a subsequent catheterization. Cells remain in the bladder for 90 minutes, after which the catheters are removed and the bladders allowed to drain naturally. The development of tumor is monitored via ultrasound. Specific attention is paid to the catheterization technique in the accompanying video.
Medicine, Issue 48, Bladder tumor, orthotopic, mouse, ultrasound
In vivo Bioluminescent Imaging of Mammary Tumors Using IVIS Spectrum
Institutions: Caliper Life Sciences.
4T1 mouse mammary tumor cells can be implanted sub-cutaneously in nu/nu mice to form palpable tumors in 15 to 20 days. This xenograft tumor model system is valuable for the pre-clinical in vivo
evaluation of putative antitumor compounds.
The 4T1 cell line has been engineered to constitutively express the firefly luciferase gene (luc2). When mice carrying 4T1-luc2 tumors are injected with Luciferin the tumors emit a visual light signal that can be monitored using a sensitive optical imaging system like the IVIS Spectrum. The photon flux from the tumor is proportional to the number of light emitting cells and the signal can be measured to monitor tumor growth and development. IVIS is calibrated to enable absolute quantitation of the bioluminescent signal and longitudinal studies can be performed over many months and over several orders of signal magnitude without compromising the quantitative result.
Tumor growth can be monitored for several days by bioluminescence before the tumor size becomes palpable or measurable by traditional physical means. This rapid monitoring can provide insight into early events in tumor development or lead to shorter experimental procedures.
Tumor cell death and necrosis due to hypoxia or drug treatment is indicated early by a reduction in the bioluminescent signal. This cell death might not be accompanied by a reduction in tumor size as measured by physical means. The ability to see early events in tumor necrosis has significant impact on the selection and development of therapeutic agents.
Quantitative imaging of tumor growth using IVIS provides precise quantitation and accelerates the experimental process to generate results.
Cellular Biology, Issue 26, tumor, mammary, mouse, bioluminescence, in vivo, imaging, IVIS, luciferase, luciferin

domain: biology


text:
Cowboys and Aliens Was on Tonight!
I remember watching Cowboys and Aliens in the movie theater the first time around. I had very high expectations, and it was somewhat of a letdown. Tonight thanks to HBO, I had a chance to watch it a second time. This time I didn’t find as many flaws, and enjoyed it a lot more than I did the first time around.
Daniel Craig, who plays Jake Lonergan, is an outlaw that’s been captured by the aliens. The only problem is that he has escaped and lost his memories of what happened and who he is. Not knowing that he was running around in a town where the family that he’d stolen from lived. Of course the Clayborne family kind of owns this town, having all the money. Paul Dano played Piercy, the son that was kind of a reject of the family and seemed always get himself into trouble using and living on his father’s reputation.
Olivia Wilde played Ella Swenson, a mysterious woman who seemed to always ask just the right questions at just the wrong time. As it turns out she’s an alien from another race whose home family has been wiped out by the same aliens invading Earth. She’s been chasing them around and followed them to earth in an attempt to destroy them.
The chemistry between Olivia and Daniel Craig bleeds through on the screen and leaves you with the impression that there’s something more than just their hatred for some aliens that are visiting Earth in an attempt to collect all the gold and kill all the inhabitants.
The main battle scene at the very end, again, leaves me with the feeling that many more people died than were available in total. Putting that all aside, I again find the movie quite enjoyable entertaining and having a good mix of excitement, cowboys, aliens, sci-fi, and technology, combined with good old-fashioned gunfights.
I give the movie an 8 out of 10. I might just catch it again here the next few weeks to see if there’s anything else that I missed. I suggest if you like cowboy movies, and you’re sci-fi fan, this is a must see.


In Windows NT-based operating systems, the System Idle Process is the system idle task: it measures how much idle capacity the CPU has at any given time. The process runs in the background and constantly monitors processing bandwidth, occupied memory and the Windows virtual paging file.
So basically, no. That's how much CPU power is unused.
I always used to wonder about that years ago, but its totally self explanatory when you think about it. If you said "oh raffaz is idle", you would be meaning that i was sitting on me arse doing nothing
the idle proccess is to take up all the time that your cpu isnt being used because your cpu must always be receiving threads otherwise it will have to reboot.
i guess this is kinda out there but if your an overclocker and you freeze your system that is why hitting the reset button wont work. the cpu has to completely restart. when you hit the reset button all it does is rerun the startup through the BIOS settings but the cpu is still reciving threads throughout this whole process


Obviously, issues didn’t work out for Savannah Chrisley and Colton Underwood on their date. Since then, Colton has moved on and is engaged to Jordan C Brown. The two seem actually pleased collectively and revealed they had been a pair not long after Colton came out as gay. “Growing up in sports, I was taught that homosexual is wrong and gay is unhealthy and football players usually are not gay,” Underwood stated. “By the time I realized that I was gay, I didn’t need to be homosexual.
Underwood obtained his begin within the franchise by vying for Becca Kufrin’s coronary heart on The Bachelorette. Kufrin was good associates with Booth after they each competed on a season of The Bachelor, and Booth advised Kufrin she sustained a relationship with Underwood prior to Kufrin becoming the lead. Ultimately, Booth’s discomfort with Kufrin relationship Underwood led Kufrin to send Underwood home. For those who may not bear in mind, both Tia and Becca competed for Arie Luyendyks Jr.’s coronary heart last season. While Tia did not make it as far as Becca, obviously, she still made all of it the best way to hometown dates. Plus, she was a fan favorite and many individuals even thought she could be the Bachelorette this season.
Savannah chrisley talks relationship ‘bachelor’ colton underwood
However, if Colton needs to upend our expectations of him, get pleasure from a profitable and entertaining season, and even — GASP — find true love on the end of it, I’d advise him to do the following things. When Colton and Tia finally broke up after a drawn-out relationship-that-should-have-never-been, I was prepared for a detox. ” I yelled each Monday night to nobody specifically. But identical to Gretchen Wieners needed everyone to hop aboard the “fetch” train, several episodes of BIP were an aggressive push to make fans swoon over Tia and Colton like they had been the new Ross and Rachel or one thing. He frolicked with go to this site Tia for a sizzling second before his appearance on the Bachelorette. And that second must have been actually, really hot (albeit without any type of vaginal intercourse) since Tia principally went into heat every time she noticed him.
Underwood posted a video montage of photos, set to “This Is What Falling in Love Feels Like” by JVKE, on Instagram. “Jordan and I are really looking forward to and leaning into the nontraditional route of it,” he said. “2021 was probably the most transformative yr of my life. Starting 2022 off with my greatest pal, teammate and now fiancé is something I by no means thought was going to be attainable.” At the end of February 2022, Underwood told PEOPLE that the couple had gotten engaged during a weekend getaway to Big Sur, California. After struggling together with his sexuality for years, Underwood mentioned he was lastly feeling good about himself. “Now that I’ve been out to myself for a year, it is the healthiest and happiest I’ve ever felt,” he shared.
Savannah chrisley and colton underwood’s relationship life now
It was straightforward for me to hide in plain sight behind a football masks and searching and fishing and the things that this world tells us is ‘masculine’ and ‘manly,'” he stated, adding, “I would have carried out something to see a gay football participant.” He admitted Randolph dumping him left him in “a dark place” because he realized it will be his last straight relationship. Several followers have advised Taylor they are so joyful she finally found love.
They could look severe, however the jury is somewhat uncertain about whether or not a much bigger commitment is in store for them. “Well, I did it…I turned 40. More grateful than ever for pals like these and nights like this. Thanks for all of the birthday love ❤,” Brown captioned a submit on Instagram that included a candy video of him and Underwood kissing. In an Instagram Story, Underwood posted a candy birthday tribute to Brown. He shared a couple of photos of himself and Brown, including considered one of them sitting with Underwood’s canine, Zooka. In the picture, the place Underwood’s hand is on Brown’s thigh, he wrote, “love.”
The bachelorette season 14, episode three recap: a skeleton of a man
This wasn’t the last of Booth and Underwood’s saga, although. The exes appeared on the same season of Bachelor in Paradise, and each fans and manufacturing had been hoping sparks would fly between the 2 of them. Underwood and Booth went residence on Bachelor in Paradise heartbroken and single.
In January 2021, Booth posted about shopping for a house as a single girl, too — so it appears like she’s still not seeing anyone. The Bachelor in Paradise debacle is behind Colton Underwood and Tia Booth now. The Bachelor Nation member had additionally just got out of a relationship on the time with Olympic gymnast Aly Raisman. We do not know how Becca’s season ends, however suffice it to say that Colton still may have an opportunity. Later that yr, Underwood sparked romance rumors with Jordan C. Brown after they were spotted having fun with a tropical getaway together. They made their relationship Instagram official in December 2021.


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African Violets are known as a difficult plant to grow and maintain indoors. However, they are beautiful plants that bloom all year long, they also live for years and are inexpensive to buy. They also do not require alot of care. Here is a few simple tips that may help you.
- Moderately Easy
- First you will want to pick a pot that will give the roots a little growing room, but not to much. Make sure the pot has holes in the bottom for drainage. Plant the violet in good potting soil with a mixture of sand. If the potting soil that you purchase does not have any sand content then you will want to purchase a small bag of sand and mix a tablespoon with the potting soil. Layer some rocks or gravel on the bottom of the pot for drainage. The biggest mistake most people make is not having good drainage; Violets cannot be drenched with water. If they are drenched in water they will rot quickly. When you start planting you will want to cover the roots to the base of the plant.
- When you water your African Violet you do not want to soak it. When it is warm weather you will want to water it more frequently, But only about 1/4 cup of water at a time. Never let water get on the leaves; they will rot.
- Put your African violet in light they need light to bloom, but not in direct sunlight. Put them in a window with sheer draperies or on a table in a window. Direct, hot sunlight will. burn the leaves.
- To start a new plant just pinch off a leaf from a healthy plant. Buy a small pot, three inches tall, and plant by placing the leaf firmly in the soil. You will want to leave at least one inch of the stem and leave standing up straight. Use my instructions above to care for it.
Tips & Warnings
Don’t soak the plant
Do not put in direct sunlight

domain: agronomy


How to stop more councils going bust
Local government reorganisation is not a subject designed to quicken the pulses of the politically ardent, neither is it often the subject of political radicalism, nor the platform of the radical public sector reform that we are oft told we need. Yet it should do, or be, all of those things.
After all it’s the central not the local state that has run up huge debts and structural deficits, and it’s the local not central government that has absorbed without collapsing (about which more shortly) the largest proportion by far of cuts to public expenditure.
Since the market is increasingly failing to provide for the middle (think of housing and the young) as well as the working classes, it is no accident that the repristinated


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When we are all awaiting the start of reservations for the new Apple Watch Series 7, several stores already show the official configurations in terms of colors and straps available. The main novelty of this Apple watch is the largest and most resistant screen ever created by Apple. Obviously, other important new features are also added and one of them is that in this model the black or space gray color disappears from the catalog, yes, this color is replaced by the so-called “Midnight” that we see on the left of the upper image.
On official reseller websites like Ktuin we already have some of the color combinations of origin that these Apple Watch Series 7 will wear. It is evident that the user will be able to modify the combinations of straps and colors to their liking, but initially these will come like this, as shown on the web.
Each user is a world and to be able to choose between the different Apple Watch case color combinations, whether to choose between aluminum or steel, the choice of the Apple Watch Nike Edition or Hermès They offer endless combinations that will undoubtedly delight those who are thinking of buying this new watch model.
The key now is to see the number of watches that Apple launches at the beginning of the reservations and it is that everything indicates that the stock will not be too high in the first hours and weeks after starting sales. We will see the advance of these Apple Watch Series 7 and above all the number of available models they offer along with delivery times.

domain: design


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Excerpt from 7.5 Secrets
Secret 2: Using exactly the right word can make your readership explode.
Too often bloggers use the first cool headline that comes to mind. Sometimes it doesn’t even make sense or tell the reader what the blog is about! That’s like labeling a can of peas as “Guess What!” But even good titles can be improved by using a more powerful word or phrase, one that is proven to work.
The difference in the number of Views your blog gets can be monumental, literally adding another zero or even two zeroes to your normal views! The results of Dr. Witt’s analysis presented below will give you specific words and phrases that did just that!
The Power Of Specific Words In A Title
Let’s look at specific words and phrases here and later at the use of Number & Symbols, Promises to Readers, Title Length, and accompanying Picture. Keep in mind this list is not definitive as it only draws on the words actually used in Dave’s titles. As you learn more from you own blog’s performance, add to these lists.
We looked at 22 different words, some of them closely related, like “success” and “successful.” You can view all of the word in tables in the Tables section of this book.
Note: We recognize blog titles are often accompanied by a few lines of text, and that text also plays a role in creating a View. The title, however, plays the key role as it summarizes (or should) “what’s in it for me,” the key question the reader wants answered.
The first sentence, by contrast, generally introduces the topic, which the headline has already provided. This is the 5 Second Rule©. In this book we will focus on the headline, but make some comments about the opening sentence when useful to you. You should carefully craft your opening sentence, too, to make it appealing.
Here are the words we reviewed and their ratings. If you want to see the Table data, click here: [Link to Table #1]
#1: “Success.” and variants “Successful,” “Succeed”
Success is a top performing word, showing a 400% difference between its presence in the top and bottom 25% of titles, which means that while not foolproof, it gives you a distinct advantage. The reason? One obvious reason is that we all want to be successful.
But a more subtle reason is that we all want to think of ourselves as having the skills to succeed because that makes us special. Reading about “success skills” makes us believe we are growing our skills (even if we’ve forgotten them within the hour!)
To learn more, read additional content in the Deeper Approach section of this book. [Deeper Approach Link #2]
#2: “Inspire” and variants “Inspiration” & “Inspiring” -
This is the only other five star word we found in Dave’s titles, showing a 300% difference between top and bottom titles. As with “Success,” it shows that paired with the right promise, it can be a powerhouse in getting clicks. One of Dave’s best performing titles, “25 Quotes to Inspire You to Become a Better Leader” (http://linkd.in/1BmLCVF), got over 877,000 Views! Also, “The 15 Most Inspiring Videos of All Time” (http://linkd.in/1xlmfCy) 215,513 Views, and the lowest performer “What Inspires People” (http://linkd.in/1DhaEEN) still got 15,650 Views. To learn more, read Deeper Approach.
#3: “You” and “Yours” -
We are usually our most important concern. So anything which promises a desirable benefit for us is attractive. That’s why “You” and “Yours” are high-impact words in a title. Of course, like the words above, the impact also depends on the promise. “Ways YOU can Win at Cricket” will have a small click-through rate in the U.S. no matter how big the word “You” is in the title. The “you” word must be accompanied by a need or desire of your readers.
#4: “Hot” Words -
Hot Words are our term for words which trigger a small, but important emotional response. The best examples of Hot Words, neither of which appear in these titles, are Free and Sex. The old joke that if you put them together, you’ve got a winning headline is actually not far from the truth. We tend to respond emotionally, that is with perked-up interest, to emotional words.
The "Hot" words identified in Dave’s list are below. . . Imagine how much more you will learn by reading the entire book! -- just click Add to Cart.


Soft V Henley
Soft is an understatement, this henley top is so cozy! It will be one you reach for all the time this fall and winter!
Mannequin Bust 33” Waist 23” Hip 34” Torso Length 26” wearing size S
Self: 80% Rayon 15% Polyester 5% Spandex
Contrast: 80% Rayon 16% Polyester 4% Spandex
Hand wash cold
Do not bleach
Do not dry clean


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The skies are still. The roads are quiet. At times it looks like the entire world is hiding indoors, guarding itself from the spread of Covid-19. But while great swathes of us are staying at home, there are others who are out there, who keep our world spinning.
I address this to you: the key worker. You are being asked to do things you have likely never even considered doing before. You are being told that your continued functioning is critical for our survival as a society.
- What the symptoms of coronavirus are, according to the WHO and NHS
- How the UK lockdown rules work
- What we know about when the UK might end lockdown
- The key statistics from deaths to testings that could end the pandemic
- Captain Tom Moore: how to support WW2 veteran’s 100th birthday Walk for the NHS fundraiser
There’s a solid chance you’re scared. I am, and I haven’t left the house in a month. There’s a good chance that your mind is spinning and your emotions are running wild. How much of an effect this is having on you is probably down to who you are, what you have done before.
For doctors and nurses who are used to working in fast-paced, high-adrenaline environments, it’s likely that they will have some measure of preparation for this. That this situation at least roughly fits within what they imagine themselves and their lives to look like.
For others, perhaps health workers who have suddenly found themselves on the front lines, or those in grocery stores and delivering our crucial goods, the change will be extreme.
Being faced with a situation that you perceive to be beyond your experience or expertise is inherently stressful. It requires greater cognitive resources and more emotional processing than to be in a situation that we are used to, that we have trained for.
Out of the ordinary
In ordinary times, I would be saying to you: “Take time for yourself, eat, sleep, cuddle with loved ones.” But these are not ordinary times.
So many frontline workers have been forced to isolate themselves from their families in order to protect them, suppressing our natural desires for social interactions. We feel we need our tribe around us, and never more so than in a threatening situation. Our attachment system screams out then for the people who make us feel safer. And so our coping mechanism is lost, our comfort removed.
When you are doing your job, there’s a good chance that you are too busy to think about much else, other than the next patient, the next customer. This is, after all, why you are so important to this society that you have to venture out into it. There’s a good chance that the role you play gives you focus, motivation, a sense of purpose.
Managing the come down
There is an equally good chance that, once that job is done, the psychological cost of your role begins to hit. You’ve been running on adrenaline and cortisol. There will always be a come down. Whether you are exhausted or agitated, weepy or angry, just know that it’s okay. This is all a normal part of a deeply abnormal situation.
You may be finding you have trouble sleeping, that you’re irritable, distractible. Did you know that you are more likely to have a car accident in the aftermath of a major trauma? Be aware of your movements while going about household chores. The distractibility can make you more likely to have domestic accidents too.
And when this ends – because this WILL end – you will come out of it with a new understanding of just how much you are capable of achieving.
Research tells us that for the vast majority of people, disasters, whilst being stressful and painful, can also be transformative, allowing them a new appreciation of themselves, their own strengths and capabilities, and a greater sense of warmth for those around them. Priorities will often change and possibilities appear where before there were none.
This period is a testing, difficult one. But the outcome of it can be quite extraordinary.
Loved ones will be hurting too
For those of you who are not the key workers, but rather the one who loves them: you may also be facing an entirely different kind of torture.
We want to be with those we love when the world is terrifying. It is a deeply human need. And you cannot.
When they return to you, they may seem distant, or snappy. In all likelihood, what you are seeing is their attempt to process their experiences. And the most important part of that processing? It’s you, their social support.
Encourage them to take practical steps to help them feel better. Research tells us that keeping a journal, in which we dissect the emotional aspects of our experience, can make a major difference in helping us grow from trauma.
Remind them to breathe, to focus on the moment before them. We will make it through this experience one moment at a time.
Remind them of their resilience and of their strength. They may not want to talk. That’s fine. Allow them to have you as their place of safety. Simply being there for them is more valuable than you can possibly know. And most importantly, to the key workers and to those who love them, thank you. For all that you do.
Emma Kavanagh’s latest novel, the thriller The Devil You Know, will be published on 28 May (Orion, £8.99)