As we approach 2022, the mattress industry is becoming strained by mattress brands names. Mattress Best For Health
Below are the 10 mattresses which have been deemed the best of the 2021 season.
Keep in mind that they are listed in no specific order.
So, if you’re searching for the most comfortable mattress there is no way to choose between these manufacturers.
- 0.1 Saatva Mattress
- 0.2 Nectar Mattress
- 0.3 Puffy Mattress
- 0.4 Dreamcloud Mattress
- 0.5 Awara Mattress
- 0.6 Nolah Signature
- 0.7 Eight Sleep POD
- 0.8 Brooklyn Aurora
- 0.9 Purple Mattress
- 0.10 Plushbeds Botanical Bliss
- 0.11 Sweetnight Mattress
- 0.12 Layla Mattress
- 0.13 Bear Pro Mattress
- 0.14 Loom & Leaf Mattress
- 0.15 Nest Bedding Mattress
- 1 Mattress Best For Health
- Saatva is an eco-friendly firm which uses environmentally friendly materials during their bed. In order to be a green option, Saatva has also cornered the market with their free White Glove delivery, further establishing themselves as the most prestigious mattress manufacturers that sell directly to consumers.
- Coil-on coil construction provides greater support and better body-contouring than conventional innersprings.
- You can choose from a variety of thickness, and the degree of firmness.
- The eco-friendly design includes recyclable steels, foams made from bio-based materials, and organic cotton in the cover.
- Free full-service shipping A 120-night sleep trial Flexible guarantee, and solid customer support.
- Nectar is possibly the most popular and hottest mattress that is available as at the time of writing. The incredible comfort of the mattress will not disappoint you. The support of Nectar is excellent and it doesn’t have too much sinkage.
- Some mattress companies offer a 10-year warranty, nevertheless the Nectar comes with a Forever Warranty.
- The top layer of Nectar is comprised from Quilted Gel Visco Elastic Memory Foam. In addition, it comes with a gel memory foam, along with Polyurethane Support Foam.
- While most brands offer a trial of their sleep, Nectar provides a 365 night sleep trial. ; Nectar offers you a whole year to try their service.
- A majority of people believe, they believe that Puffy Mattress is, without doubt, one of the most comfy mattresses available. People often compare it to enjoy sleeping on the cloud.Also people who suffer from back pain say that they feel relief thanks to Puffy’s Adaptive Flexible Cloud Technology. With Puffy, you receive both the opportunity to try a sleep trial for 101 nights, along with a lifetime assurance.
- Incredible mattress to your money.Very top-quality mattress. It can take up to two months before Puffy Mattress is shipped; since the mattress is built to specifications.
- Puffy’s top layer is CertiPUR-US Certified It is made of Cooling Cloud Relief Foam.The Cooling Cloud Relief foam layer is responsible for making the user feel as though they’re on cloud. cloud.The bottom layer is comprised of a Firm Core Support Foam that is also CertiPUR-US certified.
- Another great mattress choice, is the ever-popular DreamCloud mattress.Having a top-quality natural latex cover, DreamCloud offers a great bounce, as well as a more rounded the ability to contour.
- Some of the highlights of this mattress apart from the tufting are the handles that are stitched along the side with a price of 1,199 for a queen-sized bed and flax yarns and cashmere blend. For all of these features you can expect to pay upwards of $5,000.
- DreamCloud is a high-end hybrid mattress, offering an entire year-long sleep trial as opposed to the standard 100 night trial. New pocket coil technology permits the manufacturing costs to be lower and the savings are directly passed on to the client.
- Awara is an hybrid mattress, that combines innerspring support and a significant comfort system.
- A latex and latex hybrid, Awara Mattress pairs a 4-inch layer of natural, Rainforest Alliance-certified Dunlop latex with a 9-inch support core made of pocketed coils.
- The latex used in the Awara was certified as such by the Rainforest Alliance. This means that it meets the criteria on environmental sustainability, economic and social impact.
- Awara Also, it is GOLS (Global Organic Latex Standard) certified, which indicates that this latex comes from organic sources. This cover constructed out of New Zealand wool and organic cotton. The Best Mattress for Light Sleepers
- The Signature is Nolah’s most popular product. It is soft and supportive all at the same time, thanks to the brands exclusive AirFoam as well as high-density poly foam.
- The Signature 12 is a flippable mattress that offers two choices for firmness. Medium-firm one side and medium-soft on the other.
- Great mattress for couples due to its great motion transfer.
- Comes with a 120 night trial of sleep, as well as free shipping.
Eight Sleep POD
- By far the most advanced mattress on this list The Eight Sleep POD comes with all the features you’d expect from a smart bed mattress, and more.
- The POD comes with 5 layers, including an elongating foam topper that is incorporated inside the Active Grid for improved convenience and pressure-point relief.
- It likewise has ambient sensing unitsthat measure the level of humidity, temperature in the room and the local weather conditions to react intelligently
- This mattress does not only alter the temperature as your body’s temperature changes, but it also performs daily health monitoring of your Respiratory Rate and Resting Heart Rate along with Heart Rate Variability.
- The Aurora is a hybrid bed mattress consisting of a variety of foam layers and a layer of coil springs.
- The Aurora is available in three degree of firmness: soft medium, and company.
- At present, the Aurora is among the top selling mattresses and the most expensive offered.
- Most suitable for back sleepers and those who prefer hot sleep.
- Includes a 120-night sleep trial as well as a 10-year warranty. Mattress Best For Health
- Purple Mattress is one of the most well-known brands on the market.
- Purple mattress Purple mattress isn’t made from memory foam, as many mattresses are however, it is made from hyper-elastic polymer.
- Purple has a very different feel than the other mattress brand.
- Allows you to rest comfortably throughout the night while offering a highly responsive motion transfer.
- Purple mattress includes a 10 year limited warranty.
- The top layer consists of a hyper-elastic polymer gel grid. The lower layer is made of polyurethane foam.
Plushbeds Botanical Bliss
- PlushBeds is based in California The Botanical Bliss is their best selling mattress.
- Made from organic substances, including organic cotton covers as well as a layer that is made of organic New Zealand wool, and multiple layers of natural Dunlop latex, the mattress is sold in two firmness options: Medium and Firm. The Best Mattress for People who sleep lightly
- PlushBeds Botanical Bliss Mattress made of 100% latex is recommended by chiropractors and orthopedic specialists, thanks to its capacity to boost healthy spines.
- This mattress is great for those looking to sleep cool as well as enjoy more bounce on their mattress.
- The Botanical Bliss comes with a 100-night sleep trial along with free shipping.
- Sweetnight has a full line of mattresses and pillows, mostly using gel memory foam for their products.
- Some Sweetnight products can be purchased on Amazon, but not all of their products, or mattress sizes.
- Sweetnight has 5 mattress models to choose from, both memory foam and hybrid.
- Includes a 100 night sleep trial and free shipping.
- Available in both memory foam and hybrid.You can purchase Sweetnight in a hybrid or memory foam model.
- The double-sided Layla mattress that allows sleepers to choose between different firmness levels, depending on which side they choose to face up. The feeling of this product is different than any other you have really felt. This helps you to not only spring back really quickly, but also allows you to stay cool.
- The milder side of the mattress is made up of 3 inches of copper infused memory foam. It has a 3.5lb density, and is an extremely comfortable layer.Being a pretty thick layer of foam, you are able to sink in, as it provides pressure relief.
- Layla mattress has a 4 month sleep trial, along side a lifetime warranty. This is a huge selling point, because a 10 year guarantee is offered only by companies.
Bear Pro Mattress
- The medium-firmness makes the Bear Pro a specifically great option for combination sleepers. Those who turn at night should really feel specifically nice support when they are on their back and stomach and adequate convenience when they are on their side.
- While memory foam is known for capturing heat as well as can make sleepers get too hot throughout the evening, the Bear Pro’s gel memory foam will aid with cooling. The combination of the Celliant cover as well as cooling foam layers ought to keep most sleepers cool during the night.
- The Bear Pro mattress has 4 layers of foam and a softer feel than either of Bear’s other cushion offerings. In addition, both greatest differentiators are the copper-infused foam as well as the extra pillow supplied.
- The Bear Pro includes cost-free delivery and also returns, a 100-night sleep trial, and a 10-year guarantee. Funding is available as are numerous devices.
Loom & Leaf Mattress
- Created by Saatva, the Loom & Leaf mattress, is a memory foam mattress, that competes with luxury brands, such as TEMPUR-Pedic Loom & Leaf is one of very few online memory foam mattresses that’s not compressed and sent in a box.
- Loom & Leaf uses a gel chilled, high density high density memory foam since it is top layer. It is supported with a foundation layer comprised of high density polyfoam. This all adds up to a very supportive, pressure-relieving feel.
- With Loom & Leaf offering both firm and relaxed models, they cater to stomach sleepers of any weight.
- Loom & Leaf mattresses all come with a 120 night sleep trial, along with a 15 year guarantee. If you are ever unsatisfied for whatever reason, Loom & Leaf offers you a complete refund.
Nest Bedding Mattress
- Nest Bedding really holds the award for both the Best Rated and Top Ranked brand online. They really thrive from their client satisfaction. They offer a lifetime guarantee, in addition to a lifetime comfort guarantee.
- Nest Bedding comes in several different firmness choices. Almost too many. They consist of a latex, 2 memory foam and 3 hybrid choices in their lineup. To be sure, you can find the mattress that’s the best alternative for you.Therefore it doesn’t matter if you’re thicker, lighter, sleep on your side, or you deal with chronic pain. Nest can match your own preferences.
- They were one of online’s first brands to sell mattresses. Nest Bedding covers each kind of sleeper, by having SIX distinct mattresses in their lineup.
Support and Comfort Mattress Best For Health
The two most important aspects to consider when you are looking to buy a new mattress, is relaxation and support.
There are several things you should consider while you lay down. You want the sinkage and shape to be appropriate, for proper spine alignment. Also, you need mattresses that are able to support your entire body while you sleep at night.
It’s crucial to maintain the an optimal alignment from top the bottom while you sleep. This is necessary to make sure that you are getting an optimal rest and to ensure that you’re lessening back and neck discomfort.
While providing support for your body, a proper mattress has to provide relief from stress.
The areas of stress in your body are exactly what control you thrashing during the evening.
The memory foam mattress has made their place in the spotlight because they are not only being comfortable, but as both supportive.
Should You Choose Stiff or Soft?
This is on the same lines as when we discuss comfort and support. Just like a mattress should not be too stiff, it must never be too soft.
A mattress needs to have as much neutrality as is possible to be, in order to ensure that your spine and body remain in an upright position.Too excessive rigidity within mattresses will cause irritation to tension points, causing pinched nerves, in addition to the blood to stop flowing.
If you’re finding that a lot of the moments you wake up, experiencing the sensation of pins and needles all over your body, this is an indicator of a mattress that is too rigid. And if your mattress is too soft it can cause an sagging effect like a hammock.
When this occurs it is possible to end up suffering from back pain due to your spine being correctly aligned.
This is the reason it’s important to find a mattress that’s close to neutral. You want your mattress to be soft in the right places, but not too soft.
However, you also need your mattress to be supportive, but not too rigid..
A very often overlooked characteristics of a mattress is the breathability or fever neutrality of the mattress’s material.
Perhaps the most impressive feature of memory foam mattresses now is their capacity to be able to conform to your body.Irregardless the type of mattress, those that have fewer layers are known to cause what is known as “the toaster result”.
In essence, a mattress regulates your temperature, causing the body to be too hot.
Today, the majority of mattress manufacturers utilize modern technology to develop exceptional mattresses made of the finest materials.
Mattress manufacturers want their designs to sleep cool and warm, yet also provide warmth in the places it’s needed, and provide breathability.
Some businesses do a better job at this, over their competition. Mattress Best For Health
It’s no surprise, that if you are in search of a mattress of the highest quality is usually comes with a great price.
It’s a big investment to make, so when you purchase one it is essential to find an item that can last for many years.
It is essential to choose a mattress that’s been well designed, with a long span in mind.
If you plan to spend cash on a mattress which is brand new, look for one with at minimum a 10-year guarantee.
Mattress Best For Health
Quick Links to Other Best Mattresses:


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AIKEN, SC – Junior forward Josh Dollard scored seven of his game-high 31 points in the fourth overtime to lead second-seeded and 17th-ranked USC Aiken to a thrilling 90-87 win over third-seeded and 23rd-ranked Armstrong Atlantic State University, Saturday night (March 7) in the semifinals of the 2009 Peach Belt Conference Men’s Basketball Tournament in Aiken, S.C.
USCA (23-6) advances to face top-seeded and fifth-ranked Augusta State University in Sunday afternoon’s championship game at 4:30 p.m. in the USCA Convocation Center.
AASU (23-6) must now await the announcement of the NCAA Division II national tournament field on Sunday night (at 9:30 p.m.).
Dollard hit on 14-of-24 shots and also grabbed nine rebounds. Senior guard Kingsley Oguchi scored 22 points and hauled down seven rebounds.
Junior forward Dwight Tolbert paced the Pirates with 21 points, while T.T. Hall and Brian Moultrie added 17 each and senior 6-6 forward Calvin Wilson tallied 14 points and 14 rebounds. Hall played 58 of 60 minutes as did point guard Gabriel Robinson, who scored nine points and handed out eight assists.
The first four-overtime game in the history of the PBC Tournament featured 18 lead changes and 14 ties. AASU had the ball at the end of each extra session, but was unable to convert the game-winning shot, while USCA’s 6-7 senior forward Brian Egwuatu blocked potential game-winning attempts at the end of the first two overtime periods.
Dollard made one of two free throws with seven seconds left in the final overtime period, allowing the Pirates a chance to force extra session No. 5, but Tolbert’s three-point attempt from the right wing was just off the mark as the final horn sounded.
AASU led by as many as five points in the first half before carrying a 37-34 advantage into the locker room at halftime.
In the second stanza, the Pirates would lead by six (44-36) until USCA responded with a 13-2 run to go ahead 51-46. Tolbert drained two free throws with six ticks remaining to force the contest into the first overtime period, tied at 63-63. He intercepted a Pacer pass seconds later, but his half-court heave at the final horn was off the back iron.
Dollard scored all seven of the Pacers’ points in the last overtime period.
USCA shot 41.3 percent (33-of-80) for the game, including 10-of-30 three point attempts, while AASU shot 39 percent (30-of-77) and 7-of-21 from long distance. The Pacers made 14-of-22 free throws, while the Pirates were 20-of-31 at the stripe.

domain: sports


Paul McCartney will play live at the wedding of Prince William and Kate Middleton
England, January 6, 2011
Next April 29, the former Beatle Paul McCartney will give a concert at the wedding of Prince William and Kate Middleton. The musician will perform live at the gala reception at Buckingham Palace for Queen Elizabeth II, her grandson and his wife.
The Daily Star reported that “William and Kate have made her list of wedding and Paul McCartney is the best choice to play live”.
On the other hand, the flamboyant British musician said he “feels privileged” for the invitation.
The official source reported that “Prince William is following in the footsteps of his mother Princess Diana, by inviting celebrity friends to join the big wedding day. Paul McCartney has been invited to sing some songs and he feels very privileged”.
Authorities of the St James Palace gave details of the wedding of Prince William and Kate Middleton, for example, the route that the bride will carry out heading to the Westminster Abbey, where the ceremony will take place.
Palace spokesmen indicated that William and Kate will be married by the Archbishop of Canterbury and head of the Anglican Church of England, Dr. Rowan Williams.


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text:
EBT Crash: Obama Martial Law Dry Run Or ‘A Glitch’? (Video)
Monday, October 14, 2013
This story is a collaborative effort between Susan Duclos and Beforeitsnews reporter Live Free or Die
Was the recent EBT card debacle that shut down EBT transactions in 17 states across America a ‘glitch’ as been told to us by the MSM and TPTB or was it a ‘dry run’ for the collapse of society leading to Barack Obama declaring martial law across America? While Twitter exploded with warnings of martial law, the old adage was brought back: “Society is Always Three Meals Away From Savagery”. The video below clearly shows that this ‘glitch’ may have actually been a dry run for martial law.

domain: politics


text:
z-mode is a contemporary ladieswear fashion label created by Lithuanian born Victoria Rangayah. Victoria's debut SS11 collection, launched during London Fashion Week, focused upon dresses and contemporary tailoring with an ethical fashion bias.
Victoria spent five years in South Africa where she graduated from the London School of Fashion in 2006. After graduating she worked as a womenswear denim designer for a manufacturing company.
Having worked in a huge retail environment, Victoria wanted to create something of her very own.
Through Victoria's passion for feminine design and a strong desire for success z-mode was born.
Victoria's designs were selected for exhibition in Museum Africa, Johannesburg and her 'shoelace dress' was selected and showcased in 'Clothes Show Live - Fashion Theatre' 2010. In addition Victoria has understandably become the darling of many fashion magazines and bloggers.
Black & White Movies. Understated glamour and sophisticated silhouettes. Dressing up to feel special, whether for crowded party room or just one significant other. Contemporary beauty in true nostalgic style.

domain: artistic


History Lessons, With Popcorn, for America's UndergraduatesRoundup: Historians' Take
tags: history, movies
Zach Messitte is president of Ripon College in Wisconsin and a professor of political science. He teaches a class called "American Foreign Policy and the Movies."
The merits of the nine nominees for the Academy Award for best picture are being debated by cineastes around the world. But as a college professor, this year's Oscar nominees also reaffirm the growing importance of the movies in the classroom and how film can inspire young people to greater academic inquiry.
Although most of my students have never heard of the 1970s Abscam scandal, many have seen "American Hustle." The horrors of American slavery are brought back on screen, in "12 Years a Slave," for a generation of white undergraduates who see nothing extraordinary about having African Americans as roommates, teammates or teachers. As difficult as it is to succinctly explain the 2008 economic meltdown, "The Wolf of Wall Street" does a good job of playing up the excess and speculation that contributed to the global economic crisis. And when trying to convey the concept of failed states and why countries such as Somalia are real threats to U.S. national security, "Captain Phillips" is a way to begin a larger discussion about piracy, the perils of foreign aid and the Law of the Sea.
Historian Arthur Schlesinger Jr. said, "The fact that film has been
the most potent vehicle of the American imagination suggests all the
more strongly that movies have something to tell us, not just about the
surfaces but about the mysteries of American life." Today's movies also
tell us a lot about the changing nature of teaching....
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The nuclear receptor steroidogenic factor 1 (SF-1 [officially designated NR5A1]) is essential for fetal gonadal development, but its roles in postnatal ovarian function are less well defined. Herein, we have extended our analyses of knockout (KO) mice with markedly decreased SF-1 expression in granulosa cells. As described, these SF-1 KO mice had hypoplastic ovaries that contained a decreased number of follicles and lacked corpora lutea. In the present study, we showed that SF-1 KO mice exhibited abnormal estrous cycles, were infertile, and released significantly fewer oocytes in response to a standard superovulation regimen. Moreover, they had blunted induction of plasma estradiol in response to gonadotropins. The granulosa cell-specific SF-1 KO also significantly affected ovarian expression of putative SF-1 target genes. Consistent with their decreased follicle number, these mice had reduced ovarian expression of anti-müllerian hormone (Amh), which correlates with the reserve pool of ovarian follicles, as well as decreased gonadotropin-induced ovarian expression of aromatase (Cyp19a1) and cyclin D2 (Ccnd2). In contrast, perhaps because of their abnormal cyclicity, SF-1 KO ovaries had higher basal expression of inhibin-alpha. They also had decreased immunoreactivity for genes related to proliferation (Ccnd2 and Mki67 [also known as Ki67]) and increased expression of Cdkn1b, also known as p27, which inhibits cyclin-dependent kinases, arguing for a role of SF-1 in granulosa cell proliferation. These findings demonstrate that SF-1 has a key role in female reproduction via essential actions in granulosa cells.
Ovarian folliculogenesis is a complex process that starts during fetal life with the transformation of primordial germ cells into oocytes enclosed within primordial follicles. These primordial follicles sequentially give rise to primary, secondary, preantral, antral, and finally preovulatory (graafian) follicles that release their oocytes in response to the LH surge; the residual follicle cells then form the corpus luteum (CL). Throughout follicle maturation, sex steroids—whose biosynthesis is regulated by pituitary hormones (e.g., gonadotropins) and by growth factors that act in autocrine/paracrine fashion (e.g., insulin-like growth factors)—modulate the differentiation and growth of ovarian cells .
Steroidogenic factor 1 (SF-1 [officially designated NR5A1]) is a nuclear receptor that has essential roles in development of the fetal gonads in both sexes . SF-1 acts at multiple levels of the hypothalamic-pituitary-steroidogenic organ axis to regulate expression of genes that are important in steroid biosynthesis and sex differentiation. Knockout (KO) mice lacking SF-1 exhibit multiple endocrine abnormalities, including adrenal and gonadal agenesis, impaired expression of pituitary gonadotropins, and structural abnormalities of the ventromedial hypothalamic nucleus .
In both rodent and human ovaries, SF-1 is expressed in multiple somatic cell lineages, including granulosa cells, theca/interstitial cells, and the CL [3–5]. Because multiple ovarian cell types express SF-1, it has been difficult to define specific roles of SF-1 in each compartment. Proposed SF-1 target genes in granulosa cells include the following: anti-müllerian hormone (Amh), which is involved in regulation of follicular growth and development; aromatase (Cyp19a1), which is essential for estrogen biosynthesis by granulosa cells; and inhibin-alpha (Inha), which is important in the control of ovarian hormonal homeostasis [6–9].
In humans, SF-1 mutations have been described in a number of 46,XY subjects who presented with impaired sex differentiation, with or without adrenal insufficiency [10–15]. However, only one 46,XX subject with a heterozygous loss-of-function mutation of SF-1 has been reported, to our knowledge; this individual had adrenal insufficiency but apparently normal ovaries . Thus, the roles of SF-1 in female development and reproduction remain unclear. Moreover, the orphan nuclear receptor LRH-1 (officially designated NR5A2), which is more than 95% identical to SF-1 in its DNA-binding domain, also is expressed in rodent [3, 17] and human  granulosa cells, and studies [19, 20] have demonstrated that proposed ovarian SF-1 target genes also can be transactivated by LRH-1. These findings have raised questions about the relative roles of these two NR5A family members in the ovaries.
To explore specific roles of SF-1 within ovarian granulosa cells, we have used the Cre/loxP strategy to inactivate a conditional SF-1 allele in a cell-specific manner. We previously reported that female mice with granulosa cell-specific KO of SF-1 had hypoplastic ovaries that lacked CLs and failed to exhibit postnatal sex differentiation . In this article, we extend these studies to characterize the ovaries of these mice with respect to fertility, follicle number and distribution, and expression of markers of differentiated ovarian function.
All experiments involving mice were conducted under protocols approved by the Institutional Animal Care and Use Committee of University of Texas Southwestern. Animal handling was in accord with the Guide of Care and Use of Laboratory Animals.
The Amhr-2/Cre knockin allele  was used to drive Cre expression in granulosa cells of the ovary, resulting in mice with granulosa cell-specific KO of SF-1 as previously described . To generate mice with targeted disruption of a conditional Sf-1 allele in ovarian granulosa cells, mice heterozygous for the null Sf-1 allele (N) and the Amhr-2/Cre knockin allele were crossed with mice homozygous for the conditional floxed Sf-1 allele (F) . Mice were maintained at standard temperature and light conditions (12L:12D) and were fed ad libitum with phytoestrogen-free mouse chow (D10001; Research Diets, Inc., New Brunswick, NJ) to reduce possible effects of exogenous estrogens on the ovarian phenotype.
Genotyping was performed by PCR analysis of genomic DNA from tail snips. The primers used to detect Amhr-2/Cre, to distinguish genetic sex, and to delineate the wild-type (WT), N, and F Sf-1 alleles have been described . The PCR products were resolved by agarose gel electrophoresis, and product sizes were determined relative to size markers. Mice that were heterozygous for the F allele (F/+) and negative for the Amhr-2/Cre allele were designated WT, mice that carried the F and N Sf-1 alleles (F/N) but lacked the Amhr-2/Cre allele were designated SF-1 heterozygous (SF-1 Het), and mice with F/N alleles and positive for the Amhr-2/Cre allele (Cre/+) were considered KO.
To determine the stages of the estrous cycle, vaginal washes were collected for 21 consecutive days. Estrous phase was defined by the presence of epithelial and cornified enucleated cells within the specimen. To assess fertility, 8- to 10-wk-old SF-1 KO and control females were paired with one young fertile male mouse. Cages were monitored daily, and the number of litters and litter size were recorded.
Female mice at age 8–12 wk were injected s.c. with 5 IU of eCG (Sigma Chemical Co., St. Louis, MO). Blood samples were drawn, and ovaries were harvested for histological examination or RNA extraction at 48 h after treatment.
Mice were anesthetized, and blood was collected by cardiac puncture or retro-orbital bleeding into plastic centrifuge tubes containing EDTA (Sarstedt, Numbrecht, Germany); after centrifugation, the plasma was stored at −20°C until assays were performed. Samples were obtained both basally and at 48 h after eCG injection. Hormone assays for estrogen, progesterone, and testosterone were performed at the Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, on extracted serum samples. The intraassay coefficients of variation were less than 10% for all hormone assays. Assays for plasma LH and FSH were performed by the Ligand Assay and Analysis Core Laboratory, University of Virginia Health Science Center, Charlottesville.
After euthanasia, the ovaries and uterus were removed, cleaned of fat and mesentery, blotted on filter paper, and weighed. The tissues were then fixed overnight at 4°C in 4% paraformaldehyde and then embedded in paraffin. Sections (7 μm) were cut and stained by standard protocols with hematoxylin-eosin or used for immunohistochemical analyses.
Sections used for immunohistochemistry were washed with PBS and boiled for 20 min in 10 nM sodium citrate (pH 6.0) for antigen retrieval. Section were then incubated overnight at 4°C with primary antibodies, including the following at the indicated dilutions: goat anti-AMH (sc-6886, 1:5000 dilution; Santa Cruz Biochemicals, Santa Cruz, CA), rabbit anti-SF-1 (1:10000 dilution; a gift from Professor Ken Morohashi), rabbit anti-CYP11a1 (AB1244, 1:5000 dilution; Chemicon International, Inc., Temecula, CA), mouse anti-INHA (MCA951ST, 1:500 dilution; Serotec Ltd., Oxford, U.K.), rabbit anti-CYP19A1 (1:200 dilution; a gift from Professor Shinji Hayashi), rabbit anti-Ki67 (PRO229, 1:200 dilution; YLEM, Rome, Italy), rabbit anti-cyclin D2 (sc-181l, 1:200 dilution; Santa Cruz Biochemicals), and rabbit anti-p27 (sc-528, 1:2000 dilution; Santa Cruz Biochemicals). After washing, sections were incubated with secondary antibodies (Vectastain Elite ABC kit; Vector, Burlingame, CA) for 1 h at room temperature, washed, and developed.
Negative controls were prepared by omitting the primary and/or secondary antibody. No staining above the background was detected. Sections from at least three mice in each genotype were processed in parallel for comparison of immunostaining, and immunohistochemical studies were repeated three to five times to ensure reproducibility of results.
RNA was extracted from frozen ovaries using TRIZOL reagent (Invitrogen, Carlsbad, CA) according to the manufacturer's protocol. All RNA samples were purified using RNeasy Mini Kit (Qiagen, Studio City, CA). Reverse transcription was performed with 1 μg of tRNA using 250 U of Superscript II reverse transcriptase (Invitrogen) and random primers (Invitrogen). Ten microliters was used for each reaction. Quantitative PCR (qPCR) was performed on the ABI Prism 7900HP sequence detection system (Applied Biosystems, Foster City, CA) using commercially purchased primers (Invitrogen) with SYBR Green master mix (Applied Biosystems). All primers sets were validated to ensure efficient amplification of a single product before use in assays. Relative mRNA levels were calculated using the comparative ΔΔCt method (Applied Biosystems user bulletin 2). Comparisons were made with samples from cells harvested from WT ovaries using cyclophilin as the reference gene; expression levels in basal samples from WT ovaries were set to 1.
Primer pairs used for qPCR assays were designed as follows: SF-1 (Nr5a1) (forward), 5′ CCCTTATCCGGCTGAGAATT 3′; SF-1 (Nr5a1) (reverse), 5′ CCAGGTCCTCGTCGTACGA 3′; LRH-1 (Nr5a2) (forward), 5′ TGGGAAGGAAGGGACAATCTT 3′; LRH-1 (Nr5a2) (reverse), 5′ CGAGACTCAGGAGGTTGTTGAA 3′; Cyp19a1 (forward), 5′ GGAGAACAATTCGCCCTTTCT 3′; Cyp19a1 (reverse), 5′ CAAACTTCCACCATTCGAACAA 3′; Inha (forward), 5′ ATGTCTCCCAGGCTATCCTTTTC 3′; Inha (reverse), 5′ CGTATGTGTTGGGATGGCCGGA 3′; Cyp11a1 (forward), 5′ CGATACTCTTCTCATGCGAG 3′; Cyp11a1 (reverse), 5′ CTTTCTTCCAGGCATCTGAAC 3′; Ccnd2 (forward), 5′ ACACCGACAACTCTGTGAAGC 3′; Ccnd2 (reverse), 5′ GCCAGGTTCCACTTCAGCTTA 3′; Amh (forward), 5′ GCAGTTGCTAGTCCTACATC 3′; Amh (reverse), 5′ TCATCCGCGTGAAACAGCG 3′; Fshr (forward), 5′ TCTCTCAGAGTGATTCCCTG 3′; Fshr (reverse), 5′ AGCAAGTTTGGCTGTTATGAGG 3′; Lhcgr (forward) 5′ CTGAAAACTTGCCCTCAG 3′; Lhcgr (reverse) 5′ AATCGTAATCCCAGCCACTG 3′; cyclophilin (Ppia) (forward), 5′ TGGAGAGCACCAAGACAGACA 3′; cyclophilin (Ppia) (reverse), 5′ TGCCGGAGTCGACAATGAT 3′.
Ovaries were sectioned at 6 μm and stained with hematoxylin-eosin. The follicles in each ovary were counted serially in every tenth section through the entire ovary. Only healthy nonatretic follicles with visible oocyte nuclei were scored. Follicles were classified as primordial, primary, small preantral, or antral. Primordial follicles had a compact oocyte surrounded by a single layer of flattened (fusiform) granulosa cells, primary follicles had an enlarged oocyte surrounded by a single layer of cuboidal granulosa cells, and small preantral follicles had an enlarged oocyte surrounded by three or four layers of granulosa cells. Follicles with five or more layers of granulosa cells were considered antral follicles.
Mice aged 8–12 wk were treated s.c. with 5 IU of eCG (Sigma Chemical Co.), followed 48 h later by 5 IU s.c. of hCG (Sigma Chemical Co.). Mice were killed at 16–18 h after hCG treatment, and the oocytes were harvested from the oviducts and incubated in PBS with 10% BSA. After treatment with hyaluronidase (300 μg/ml; Sigma Chemical Co.) for 10 min at 37°C, oocytes were collected and counted manually under a Nikon microscope (Nikon, Natick MA).
Data are given as mean ± SEM. One-way ANOVA was applied to compare means among the groups. Appropriate post hoc, pairwise multiple comparisons were performed (least significant difference test) when applicable. The SPSS version 14.0 software package (SPSS Inc., Chicago, IL) was used for the statistical analysis, and P ≤ 0.05 was considered significant.
Estrous cycles in adult female mice were assessed in vaginal smears collected for 21 consecutive days. The WT and SF-1 Het mice had cyclical estrous cycles that lasted 4–5 days and progressed consecutively through the expected changes in cell content, with an estrous phase of approximately 2 days (Table 1). In contrast, the granulosa cell-specific SF-1 KO mice had markedly disordered estrous cycles that ranged from irregularly prolonged cycles (50%) to complete acyclicity (50%).
We also evaluated fertility by mating adult female granulosa cell-specific SF-1 KO mice with young WT male mice. In longer than 1 yr of controlled observation, no SF-1 KO mice delivered any pups; this contrasts with WT mice, which averaged 9 ± 1 litters per year, with 6.3 ± 1.8 pups per litter. However, we found in the uterus of one SF-1 KO mice a single embryo at early stage of development.
To determine if the infertility was due to a primary ovarian defect or was secondary to changes at other levels of the hypothalamic-pituitary-gonadal axis, we used a standard superovulation protocol and counted the number of oocytes released into the oviducts. As shown in Figure 1, WT females ovulated an average of 35.3 ± 2.9 oocytes per female, while SF-1 Het females ovulated an average of 23.3 ± 5.1 oocytes per female; a marked decrease was seen in the number of oocytes collected from mice with granulosa cell-specific KO of SF-1 (1.0 ± 0.7 oocytes per female). Most (80%) SF-1 KO females had no oocytes retrieved from the oviducts; two mice released three and four oocytes each, some of which were in immature stages of development (P < 0.001 compared with WT and SF-1 Het mice). These data show that SF-1 KO mice exhibit an impaired or absent response to gonadotropin-induced superovulation. The number of oocytes retrieved also differed significantly between SF-1 Het and WT mice (P ≤ 0.01), suggesting that SF-1 haploinsufficiency diminishes oocyte production to some degree without impairing fertility.
We next examined ovarian size and histology. Ovaries from SF-1 KO mice (1.79 ± 0.15 mg, n = 12) weighed significantly less than those from SF-1 Het mice (5.05 ± 0.50 mg, n = 6; P < 0.01) and from WT mice (7.39 ± 0.52 mg, n = 12; P < 0.001). The ovaries of adult WT and SF-1 Het mice contained follicles at all stages of development, including CLs (Fig. 2 and data not shown). In contrast and consistent with our previous study , sections of SF-1 KO ovaries showed the expected range of follicles up to the preovulatory stage but had no CL, suggesting a defect in the final steps of folliculogenesis and/or ovulation. Moreover, careful quantitation in serial sections of adult SF-1 KO ovaries revealed a reduction in the total number of follicles (Ikeda, unpublished results).
SF-1 in the ovary is expressed in both granulosa and theca/interstitial cells, which have complementary functions and differentially express some genes involved in the steroidogenic pathway. Theca cells synthesize androgen precursors under stimulation by LH, which the granulosa cells then convert to estradiol (E2) . To further assess the cell specificity of SF-1 inactivation and the resulting downstream effects, we used immunohistochemical analyses. We examined SF-1 immunoreactivity in ovarian sections from adult mice (age, 12 wk). As shown in Figure 2, WT mice expressed SF-1 in both granulosa and theca/interstitial cells of growing follicles and at a lower level in cells of the CL. In contrast, SF-1 KO mice had strong SF-1 immunoreactivity in the theca/interstitial cells of all follicles but only weak expression in granulosa cells of a few follicles. Although only semiquantitative, this residual expression in the immunohistochemical analyses presumably reflects cells in which the Amhr-2/Cre did not fully inactivate SF-1 expression. Nonetheless, SF-1 expression was silenced in most granulosa cells, consistent with the reproductive phenotypes seen in these mice.
We next compared the number of follicles in WT, SF-1 Het, and SF-1 KO mice at age 21 days, which corresponds to the time just before the initiation of puberty; we reasoned that evaluation at this age might provide insights into the initial ovarian reserve. Relative to WT mice, the 21-day-old SF-1 KO mice had significant reductions in follicles at all stages of development, whereas no differences were observed between SF-1 Het and WT mice (Table 2). Of note, the number of primordial follicles—the primordial follicle pool—was significantly reduced compared with that in WT mice. These data suggest that the granulosa cell-specific SF-1 KO mice have a decreased reserve of follicles before puberty that then persists into adulthood.
We next evaluated plasma levels of ovarian steroid hormones and gonadotropins (Table 3), including E2, progesterone, testosterone, FSH, and LH. Unexpectedly, the basal levels of the ovarian steroid hormones in the granulosa cell-specific SF-1 KO mice did not differ significantly from WT and SF-1 Het levels at the diestrous phase. The normal testosterone level was consistent with the known role of theca cells in ovarian androgen production; these cells do not express Amhr-2/Cre, and SF-1 should remain functionally intact. We further hypothesized that the apparently normal circulating E2 levels reflected not only ovarian E2 production but also E2 produced by other sites of aromatase expression such as adipocytes. Although the LH levels did not differ significantly among the three groups, plasma FSH was increased significantly in SF-1 KO mice relative to WT and SF-1 Het mice (P ≤ 0.01).
In an effort to uncover defects in ovarian E2 production, we measured the steroidogenic response to eCG stimulation. Female mice at age 8–12 wk were treated with eCG, and plasma samples were collected 48 h later. As summarized in Table 3, plasma E2 levels were stimulated by eCG in WT mice, whereas plasma progesterone levels were unaffected. Similar responses were seen in SF-1 Het mice. In contrast, the eCG-stimulated increase in E2 levels was markedly blunted in mice with granulosa cell-specific KO of SF-1 (P < 0.001 vs. WT and SF-1 Het mice). This relative defect in ovarian estrogen biosynthesis in these mice is consistent with their increased basal levels of FSH; both of these factors suggest a primary gonadal defect secondary to the absence of SF-1 that eventuates in premature ovarian failure. However, because large antral and preovulatory follicles are the major source of E2, we cannot exclude a contribution of the decreased number of these follicles toward the impaired eCG-stimulated synthesis of E2.
As a biomarker of E2 levels, we examined uterine weight and histology. At age 8–10 wk, uterine weights in WT mice (151 ± 10 mg, n = 12) and SF-1 Het mice (155 ± 12 mg, n = 6) did not differ significantly; mice of both genotypes had significantly increased uterine weights relative to SF-1 KO mice (48.8 ± 7.0 mg, n = 12; P < 0.001 vs. both groups). The uterine histology of WT and SF-1 Het mice revealed thick myometrial and stromal layers, ample endometrial glands, and multiple layers of luminal epithelial cells (Fig. 3, B and C). In contrast, the uterine histology of SF-1 KO mice showed reductions in epithelial, myometrial, and stromal layers. In particular, the glandular elements of the endometrial layer were less complex, with only a few scattered glands detected in the stroma. These findings collectively indicate that ovarian E2 production is inadequate to stimulate normal uterine differentiation in the granulosa cell-specific SF-1 KO mice.
The FSH-induced differentiation of ovarian granulosa cells during folliculogenesis activates a complex pattern of gene expression. To examine the effect of the granulosa cell-specific SF-1 KO during these processes and to permit more precise quantitation, we used qPCR assays to evaluate gene expression in the ovaries of 8- to 12-wk-old WT, SF-1 Het, and SF-1 KO mice before and after stimulation with eCG (Fig. 4).
Expression levels for many of the genes analyzed did not differ significantly among these mice. However, several differences are notable. Basal expression of SF-1 was decreased in SF-1 KO samples relative to WT samples (P < 0.05). This is consistent with the immunohistochemical analyses shown in Figure 2 and the fact that our inactivation strategy targets SF-1 expression in a significant population of ovarian cells. Following gonadotropin stimulation, SF-1 expression in RNA from whole ovaries of WT and SF-1 KO mice did not differ significantly, perhaps reflecting gonadotropin-induced SF-1 expression in the theca cells. In contrast, basal expression of LRH-1 in SF-1 KO ovaries was increased significantly relative to WT levels (P ≤ 0.05) and nonsignificantly relative to SF-1 Het levels; similar responses to gonadotropins were seen in each group. This finding may reflect a compensatory mechanism within granulosa cells to maintain function in the absence of SF-1.
The gene whose expression was most severely impaired by the SF-1 KO was anti-müllerian hormone (Amh). This well-characterized SF-1 target gene normally is expressed exclusively in granulosa cells, where it is proposed to have an important role in the recruitment of ovarian follicles . SF-1 KO ovaries had significantly decreased basal and gonadotropin-stimulated expression of Amh, strongly supporting the role of SF-1 in Amh expression in vivo..
Another gene whose expression was affected by the SF-1 KO is inhibin alpha (Inha). This target gene of SF-1 is induced by FSH and has been implicated as a negative regulator of stromal proliferation in the ovary . As expected, both WT and SF-1 Het mice showed significant induction of Inha expression after eCG treatment. In contrast, SF-1 KO mice showed increased basal expression of Inha but—unlike WT and SF-1 Het mice—did not show increased expression in response to eCG (P ≤ 0.01 vs. WT mice). This increased basal expression of Inha, coupled with high FSH levels, suggests that the negative regulation of FSH by ovarian inhibin is lost in SF-1 KO mice, a typical phenomenon of aging.
The expression of other genes in SF-1 KO mice differed significantly only after gonadotropin stimulation. For example, gonadotropin-stimulated expression of aromatase (Cyp19a1), which is selectively expressed in granulosa cells , was significantly decreased in SF-1 KO mice relative to SF-1 Het mice (P < 0.05) and was slightly reduced relative to WT mice. This is consistent with the proposed key role of SF-1 in aromatase expression and the impaired E2 production in granulosa cell-specific SF-1 KO mice in response to gonadotropins (Table 3). The expression of Cyp11a1 was increased in the SF-1 KO mice after eCG injection compared with WT and SF-1 Het mice. These data were in agreement with the results of the immunohistochemical analyses (discussed herein). The expressions of FSH receptor (Fshr) and LH receptor (Lhcgr) were also analyzed [25, 26]. No significant differences in Fshr expression were shown among the three groups. Instead, significantly higher expression of Lhcgr at the basal level (P = 0.02 vs. WT mice and P = 0.014 vs. SF-1 Het mice) and exaggerated increase after eCG injection compared with controls (P = 0.047 vs. WT mice) were seen in SF-1 KO mice. Similarly, the expression of cyclin D2, a marker of granulosa cell proliferation that normally is induced by FSH [27, 28], was significantly lower in SF-1 KO mice only after eCG treatment (P = 0.02 vs. WT mice). These data suggest underlying impairment in the proliferation of SF-1-deficient ovarian granulosa cells following gonadotropin stimulation (Fig. 4).
Collectively, these results suggest that the ovaries in granulosa cell-specific SF-1 KO mice do not respond properly to eCG stimulation both in terms of expression of certain critical genes (e.g., aromatase, Amh, and cyclin D2) and in terms of estrogen biosynthesis. This inability presumably reflects a primary ovarian defect that results from the absence of SF-1 expression in the granulosa cells.
We next used immunohistochemical analyses to evaluate ovarian expression of other genes of interest. In WT mice, AMH was expressed at high levels in granulosa cells of primary, secondary, and small preantral follicles, while ovaries in SF-1 KO mice contained only a few primary and secondary follicles that expressed AMH (Fig. 5). Also, CYP19A1 levels in WT mice seemed to be higher in granulosa cells of growing follicles compared with those in SF-1 KO mice. Although WT mice expressed CYP11A1 only in the theca cells and the CLs, ovaries from SF-1 KO mice also contained some large follicles that ectopically expressed CYP11A1 in granulosa cells before and after hormone treatment. This expression may reflect efforts to compensate for the absence of estrogen production in SF-1 KO mice and suggests that CYP11A1 transcription can be induced by SF-1-independent mechanisms under certain circumstances. In contrast to these other putative SF-1 target genes, the qPCR data suggested that basal expression of Inha was actually increased in SF-1 KO mice (Fig. 4). Consistent with this, SF-1 KO mice seemed to have higher expression of INHA in the immunohistochemical analyses (Fig. 5). Thus, these studies largely confirmed the conclusions drawn from the qPCR analyses of gene expression.
The reduced number of follicles and ovarian size in the granulosa cell-specific SF-1 KO mice suggested that these mice might have defects in cell proliferation in the ovaries. Therefore, we extended our immunohistochemistry analyses to examine the expression of CCND2 (cyclin D2), MKI67 (Ki67), and CDKN1B (p27). As shown in Figure 6, granulosa cell-specific KO of SF-1 was associated with considerably decreased expression of cyclin D2 and MKI67, while there was increased expression of CDKN1B in the granulosa cells of growing follicles. These findings suggest that impaired cell proliferation is an important component of ovarian hypoplasia seen in these mice.
Using the Cre/loxP system, we studied the roles of SF-1 in ovarian physiology in vivo, focusing specifically on its functions in granulosa cells. Consistent with our initial description , the granulosa cell-specific inactivation of SF-1 caused marked postnatal gonadal defects, including ovarian hypoplasia, altered uterine differentiation, and defects in both estrogen biosynthesis and fertility. Although these findings do not exclude important roles of LRH-1 in ovarian function, they define essential roles of SF-1 in granulosa cell function that cannot be replaced by LRH-1. In most respects, SF-1 Het mice exhibited no reproductive phenotype; they had mild ovarian hypoplasia with normal ovarian function and approximately 65% of the normal number of oocytes harvested after superovulation. Moreover, the gene expression profiles in SF-1 Het mice generally paralleled those in WT controls both basally and after eCG treatment, indicating that the presence of one SF-1 allele suffices for largely normal ovarian function.
SF-1 KO mice exhibited markedly reduced ovarian size, with a significantly decreased number of growing follicles and the absence of CLs (Fig. 2), suggesting that they are impaired in terminal stages of follicle differentiation and/or ovulation. A careful inspection of ovarian histology in young adult SF-1 KO mice showed a paucity of primary and secondary follicles, which led us to evaluate the number of follicles just before the initiation of puberty. At age 21 days, SF-1 KO mice already had a decreased number of follicles (Table 2). These data again suggest that SF-1 has important roles in ovarian development and follicular maturation. This defect was not apparent during fetal development of SF-1 KO ovaries up to Embryonic Stage 16.5 .
We also evaluated ovarian function by determining the plasma levels of ovarian steroid hormones both basally and after stimulation with gonadotropins (Table 3). The basal E2 level in SF-1 KO mice did not differ significantly from the levels in WT and SF-1 Het mice, but marked impairment was noted in the response to eCG. Moreover, basal plasma FSH revealed a significantly increased concentration in SF-1 KO mice compared with mice of other genotypes. These data together indicate that ovarian estrogen production is defective in mice with granulosa cell-specific KO of SF-1. Although developing ovarian follicles are the main source of E2, other tissues (e.g., adipose tissue) can produce estrogen. The normal basal E2 level in SF-1 KO mice may derive from these other sources.
Studies [29, 30] have demonstrated that AMH—which is produced by granulosa cells of the growing follicles—suppresses development of the primordial follicles. Thus, the FSH-dependent recruitment of small antral follicles in Amh KO mice was increased, and premature exhaustion of the primordial follicle reserve occurred . SF-1 has a key role in Amh transcription ; consistent with this, Amh transcripts were decreased in our qPCR analyses, and only a few follicles contained immunoreactive AMH (Fig. 5). Therefore, the decreased levels of AMH in the granulosa cell-specific SF-1 KO mice may contribute to the diminished follicle number observed.
Similarly, the reduced induction of Cyp19a1 expression in ovarian granulosa cells seen in the qPCR and immunohistochemical analyses likely contributes to the deficient E2 production. The hypoplastic uteri with impaired glandular differentiation were consistent with estrogen deficiency in the granulosa cell-specific SF-1 KO mice. We cannot exclude the possibility that these ovaries had few follicles and therefore had lower capability to produce E2 after gonadotropin induction.
Studies [7, 33] in female aromatase KO mice showed severely impaired E2 synthesis and follicle development, resulting in defective ovulation. The phenotype described herein resembles several aspects of aromatase KO mice but apparently was less pronounced in terms of hormone deficiencies and gonadal morphology. The attenuated phenotype in mice with granulosa cell-specific KO of SF-1 likely arises because they lack SF-1 and CYP19A1 only in ovarian granulosa cells, whereas aromatase KO mice lack CYP19A1 expression in all tissues. By avoiding any exogenous source of estrogen with a soy-free diet , we showed that our mice can still produce E2 but not at levels sufficient to induce ovulation. Moreover, we found that the ovarian hemorrhagic cysts previously described  occurred in only a few of these mice fed a phytoestrogen-free diet.
As a proposed target gene of SF-1 in ovarian cells, we also examined inhibin-alpha expression in the granulosa cell-specific SF-1 KO mice. This gene is transcriptionally activated by FSH in ovarian granulosa cells during follicular growth . Transient transfection assays using the inhibin-alpha promoter showed that both SF-1 and LRH-1 can increase inhibin-alpha promoter activity [20, 35]. A dynamic association/dissociation model has been proposed to explain the interactions of NR5A members and the inhibin-alpha promoter: at baseline, SF-1 is bound to the inhibin-alpha promoter, but FSH induces LRH-1 to replace SF-1 . In our investigations, increased basal expression of Inha was detected in SF-1 KO mice, localizing to granulosa and theca/interstitial cells. Therefore, we hypothesize that LRH-1 can replace SF-1 to induce basal Inha expression in granulosa cells. However, LRH-1 alone was unable to increase Inha expression after gonadotropin stimulation, as revealed by the reduced Inha transcript levels in SF-1 KO mice. These findings, together with the elevated FSH concentration, suggest that adult ovarian granulosa cells of SF-1 KO mice are unable to respond appropriately in terms of gene expression and steroid biosynthesis. A similar defect is seen in aging mice where the negative regulation of FSH by ovarian inhibin is lost, supporting an accelerated ovarian senescence phenotype in these mice. Chronically elevated expression of inhibin-alpha was reported in ovaries of acyclic aging female rats, and increased expression of INHA has been associated with lack of ovulation and loss of estrous cyclicity due to loss of estrogen secretion . The extent to which altered expression of Inha contributes to the phenotype seen in the granulosa cell-specific SF-1 KO mice remains to be determined.
The impaired induction of cyclin D2 transcripts by eCG, the lower immunoreactivity for cyclin D2 and MKI67, and the intense signal of CDKN1B suggest that SF-1 KO mice have an underlying defect in granulosa cell proliferation and follicle growth [27, 37]. Thus, changes in proliferation may be a major factor in their phenotype. We previously showed that male mice with Leydig cell-specific SF-1 KO had impaired somatic cell proliferation during fetal development , and evidence from other studies [38, 39] strongly implicates SF-1 as a key regulator of adrenocortical proliferation. By analogy, it is likely that SF-1 regulates other genes involved in granulosa cell proliferation; the identification of these genes will be an important target for future studies.
We thank Dr. Richard Behringer for providing the Amhr-2/Cre mice, Dr. Suria Pancharatnam for helpful discussions, Dr. Nancy R. Stallings for assistance with qPCR analyses, and Dr. Stefano Romeo for statistical analysis.
1Supported by grant R01 HD046743 from the National Institute of Health to K.L.P.

domain: biology


text:
Finch Farm in Rehoboth, MA chemical-free
E-mail [email protected]
Visit our website
196 Willard Ave
Seekonk, MA 02771
Finch Farm is a small, 1.5 acre mixed vegetable farm located in Rehoboth, Massachusetts. The farm is managed by Ian Augustine and Seamus Sullivan. We grow vegetables, cut flowers and culinary herbs using organic methods of weed and pest management. We are a twenty minute drive outside of Providence, Rhode Island, where we live. That being so, in our first season we hope to provide to the communities that surround us in both Rhode Island and Southeastern Massachusetts.
SEMAP regularly updates the Local Food Guide. Let SEMAP know if something is inaccurate.

domain: agronomy


Re: [PATCH RESEND] iwlwifi, Do not implement thermal zone unless ucode is loaded
From: Kalle Valo
Date: Wed Jul 13 2016 - 02:51:14 EST
Prarit Bhargava <prarit@xxxxxxxxxx> writes:
>> We implement thermal zone because we do support it, but the problem is
>> that we need the firmware to be loaded for that. So you can argue that
>> we should register *later* when the firmware is loaded. But this is
>> really not helping all that much because the firmware can also be
>> stopped at any time. So you'd want us to register / unregister the
>> thermal zone anytime the firmware is loaded / unloaded?
> You might have to do that. I think that if the firmware enables a feature then
> the act of loading the firmware should run the code that enables the feature.
> IMO of course.
But I suspect that the iwlwifi firmware is loaded during interface up
(and unloaded during interface down) and in that case
register/unregister would be happening all the time. That doesn't sound
like a good idea. I would rather try to fix the thermal interface to
handle the cases when the measurement is not available.


text:
South Korea’s dominant mobile-messaging service, Kakao Corp., and local Internet portal Daum Communications Corps. said Monday they will merge and establish a new company by the end of year in a bid to boost their presence in both web and mobile devices
[more…] With the all-stock deal, unlisted Kakao has dropped previous plans for an IPO by 2015 and turned itself into a serious challenger to Naver Corp 035420.KS, the country’s biggest Internet portal operator, which also owns rival mobile messenger operator Line.
The two companies said they had agreed that each Kakao share is worth 113,429 won. Daum will issue 43 million new shares to Kakao shareholders, including 33 million common shares.
“The synergies from the deal are obvious. The deal will give firepower to Kakao to aggressively expand its business here and overseas, while Daum will have a mobile messaging service as its new growth engine,” said Kim Mi-song, an analyst at Hyundai Securities.
Kakao Corp operates South Korea’s dominant mobile messenger service, Kakao Talk and it has some 145 million users globally, although it has been less successful than Line at expanding overseas.
The newly merged company will be called Daum Kakao.
Kakao reported 55.6 billion won of net profit last year on 211 billion won of revenue.

domain: news


Regional microclimate humidity of clothing during light work as a result of the interaction between local sweat production and ventilation
journal contributionposted on 2013-06-13, 10:40 authored by Hiroyuki Ueda, Yoshimitsu Inoue, Mitsuo Matsudaira, Tsutomu Araki, George HavenithGeorge Havenith
Purpose – The aim of this study is to explore the influence of the clothing ventilation in three body regions on the humidity of the local clothing microclimates under five work-shirts immediately after the onset of sweating in light exercise. Design/methodology/approach – The clothing microclimate ventilations were measured at chest, back and upper arm using a manikin. Separate wear trials were performed to determine the sweat production and the humidity of the clothing microclimate at the same locations as where the ventilation was measured during light exercise. Findings – Every shirt shows the greatest value of ventilation index (VI) for the chest and the smallest one for the upper arm. The values of VI differ remarkably at the chest among the five shirts. Comfort sensation became gradually worse as the time passed after starting exercise. There was no significant difference among the clothing conditions in mean values of rectal temperature, local skin temperatures, microclimate temperatures, microclimate relative humidities and local sweat rates at three regions over 10?min after the onset of sweating. A relationship was observed between the ratio of the mean moisture concentration in the clothing microclimate to the mean sweat rate at the chest and the back and the VI. Originality/value – The results suggest that clothing ventilation should be measured in different body regions in response to sweat rates in corresponding regions.
CitationUEDA, H. ... et al., 2006. Regional microclimate humidity of clothing during light work as a result of the interaction between local sweat production and ventilation. International Journal of Clothing Science and Technology, 18 (3-4), pp. 225 - 234.
- AM (Accepted Manuscript)
NotesThis article was published in the journal, International Journal of Clothing Science and Technology [© Emerald] and the definitive version is available at: http://dx.doi.org/10.1108/09556220610668473


Ashwiny Iyer Tiwari: Alcoholism is leading to abuse in the lockdown
The writer-filmmaker deliberates over the rise in domestic violence cases in the society amid the Covid-19 lockdown.
Filmmaker Ashwiny Iyer Tiwari confesses that though she thought twice before putting out a tweet against domestic violence cases in the lockdown period, it was something she feels strongly about.
Telling us what made her write it, the 40-year-old says, “I have seen that domestic abuse is a huge thing because of alcoholism. I saw it in the house of someone I know, it is a problem. The truth is, you are in a society where alcohol is a form of shying away from handling the situation. Right now there are so many stories about losing jobs, not having enough money to sustain, men find a way in alcohol, and when that stress is not managed correctly, it leads to abuse at home.”
Adding how it impacts children, she further tells us, “It leads to the child growing up in an environment seeing the man of the house do it. Therefore, I wrote ‘handle it calmly’, because if you do not, it leads to destruction and division of families and mental instability, of not just yourself, but everyone around you. You are going and putting the little money you can save for a rainy day, in alcohol. Then what about the family?”
Apart from expressing her thoughts on the crisis and the lockdown on social media, Tiwari has spent the past one-and-a-half months finishing scripts, and also her debut novel, nearing completion.
“I have also been cooking,” she informs us, “I got into that zone where you have children at home and you have to feed them, so that came as a default. After Panga released, there was so much happening I didn’t get time sit and write, so I did that. Then we are working on the screenplay of my biopic on Narayan and Sudha Murthy, which I will direct and is being co-written by my husband Nitesh Tiwari and other writers. By June, I should also finish my novel which I started writing after Bareilly Ki Barfi. It’s almost been two-and-a-half years.”


We reported in March that Apple was opening up its plug-in architecture in the latest version of its popular image workflow tool, Aperture 2.1. Per a report on DPReview today, PictureCode’s Noise Ninja noise reduction system is now available for download.
Available for some time as both a Photoshop plug-in and a stand-alone software package, Noise Ninja provides advanced noise reduction controls for cleaning up high-ISO shots.
Apple has added a batch of new third-party apps for Aperture of late, but the PictureCode addition is almost certainly the most exciting plug-in yet for Apple’s Lightroom alternative.
To check out the latest available plug-ins for Aperture, visit Apple’s Aperture downloads site.


text:
Editors note: News broke on Nov. 9 that The Black Crowes will reunite this summer to recreate the “Shake Your Money Maker” album on tour. Here are the dates: LINK
A Wall Street Journal article on Oct. 11 started to churn the waters on a possible reunion of The Black Crowes, which would mark the 30 year anniversary of their landmark album, “Shake Your Money Maker.”
The story is intriguing for a few reasons. First, the band has a deep song catalog, led by the aforementioned “Shake Your Money Maker,” which contains mega hits “Jealous Again,” “Hard To Handle,” “She Talks To Angels” and “Twice As Hard,” not to mention iconic Crowes tunes such as “Sister Luck,” “Seeing Things” and “Thick ‘N Thin.”
But the second and third records “Southern Harmony and Musical Companion” and “Amorica” are often tabbed as the die-hard fan favorites, with “Three Snakes and One Charm,” “By Your Side,” “Lions,” “Warpaint” and Before The Frost… Until The Freeze,” all consisting of stellar songs in their own right.
In addition to all of the great music though, are personality conflicts. Singer Chris Robinson and brother/guitar player Rich Robinson have had a long, widely aired feud off and on over the decades, resulting in hiatuses, and in 2015, Rich declaring the band was over once and for all because Chris wanted virtually all of the profits, eliminating the three-way share between founding members Chris, Rich and drummer Steve Gorman.
Speaking of Gorman, he just released a book about the history of the band from his perspective – “Hard To Handle: The Life And Death of The Black Crowes.” The book is entertaining and chock full of fireworks between the band members. Needless to say, the brothers are often painted in the book as spoiled brats, jealous and consumed by maximizing making money for themselves.
According to recent interviews from Gorman, he claims he has known about a reunion between the brothers, but hasn’t been asked to play. He doesn’t believe he will, primarily because he feels like a potential reunion is motivated by a good ‘ol fashioned money grab by the Robinsons (Shake Your Money Maker indeed) with them not being interested in paying him his share … and potentially being miffed about how they are characterized in his book.
That being said, this all went down a similar path in 2005 when the Crowes were on hiatus and came back … and it took an 11th hour phone call from Chris to Steve to get the band rightfully back together. Never say never, until the band is onstage. As it is, they all seem to be back in the news regularly these days, so the PR effect is bringing attention.
The biggest question is what are they going to do about a lineup, and will it be any good? Chris, of course, has spent most of the past decade in his psychedelic folk-rock jam outfit, Chris Robinson Brotherhood, while Rich has released several of his own solo records, played guitar in Bad Company and started a band with many former Black Crowes members – The Magpie Salute, which coincidentally just released its second full-length LP, “High Water II.”
CRB suddenly cut short its touring schedule after July, and Magpie hasn’t played since the winter, fueling further speculation a Crowes reunion is in the works. Magpie has former Crowes Marc Ford on guitar, as well as Sven Pipien on bass. A founding member was Eddie Harsch on keys, although Harsch died before the band really ever got a chance to get going. Chris also had his own band with former Crowes in it, As The Crow Flies, which played about a month’s long tour last year as well as a few festivals. That lineup contained former Crowes member Audley Freed on guitar, as well as Andy Hess on bass and Adam MacDougall on keys.
Of course, if Live Nation is at the helm with the makings of a 30 year anniversary tour on the table, that all sounds like big money for the brothers. My hope is that they don’t cut corners on the music. There have been many Black Crowes band members over the years, 17 I believe. Of course, the fans want to see a classic lineup, which the closest to that right now would be Chris, Rich, Steve Gorman on drums, Sven Pipien or Johnny Colt on bass, Marc Ford on guitar, and Adam MacDougall on keys, plus the girls singing harmonies.
The Crowes tried to make a go of it without Gorman on drums once before, in 2005, and Chris had to personally insist that Steve return, because the sound wasn’t right without him. That’s a real issue. There are other great drummers out there – Tony Leone did a very solid job with As The Crow Flies, and Joe Magistro (one-time drum tech for Gorman) played the Crowes songs well with Magpie but there is a sound that the hard hitting, but sticky Gorman brings, along with Rich’s open tuning mastery. It can’t be replicated.
My take: Give the fans what they want, make the music as great as possible, say a proper goodbye (if they don’t want to play with each other again), and people will pay to see it. The ensuing quality of the shows will justify everything.
— Jon Siembieda


text:
GREEN BAY, WI (WTAQ) - A Green Bay woman is facing federal indictment involving a more than $1 million fraud scheme.
After a lengthy investigation involving Brown County Sheriff’s Office and the Federal Bureau of Investigation, 48-year-old Lisa Lewis was charged with defrauding at least 13 elderly investors.
Lewis would pass herself off as an account representative with Fidelity Investments and Fidelity Brokerage services. She wasn’t even a registered broker or had affliaction with Fidelity.
Lewis convinced victims to transfer money to accounts and obtained their passwords allowing her access to those accounts. She eventually used the money for personal use.
Authorities say she wrongfully obtained fund in excess of $1 million.

domain: finance


text:
Published November 11th, 2011
“When I was eight or nine years old, I read about Scott and Amundsen and the “Race for the South Pole,” and from then on, I was hooked on Antarctic history and things Antarctic in general. Antarctica became a passion for me, even as I spent my life in the worlds of economics, finance, and business. Decades passed before I was able to visit Antarctica. When I at last did, in 1995, my trip took me to the Antarctic Peninsula Region, thousands of miles from where Scott and Amundsen had run their race. But over the years since I was a small girl, I had read far more Antarctic history, and I knew that I was going to a place rich in history and stories of fascinating events and people. What I hadn’t anticipated was how spectacularly beautiful it would be, how addictive I would find the ice, the landscape, and the wildlife, how emotionally I would react when I first saw Antarctic land. I have been back to Antarctica many times since 1995, both to the Peninsula Region and elsewhere, and every time I experience the same sense of awe and wonder at simply being there.
I have traveled more in the last 16 years to Antarctica than to anywhere else, so in a sense, “Antarctica” is my favorite place. But that’s a very large place, and there are certain locales that speak to me more than others. Since my intellectual focus is on history, the places important to the human story of Antarctica tend to be the ones that stay with me the most. But the landscape has also captured me, so my absolutely favorite places are those with both fascinating stories and wonderful surroundings. The coast of Victoria Land, Ross Island, South Georgia, the South Orkneys, Port Lockroy. . . . All are favorite places, ones that I want to return to over and over. But it is Deception Island—an amazing small doughnut-shaped island in the South Shetlands that is an active volcano—that most quickly comes to mind as a “favorite place.” Here there is a marvelous marriage of magnificent scenery, geology, and a deep history that stretches from the beginnings of Antarctic exploration up through today. Ruins of the abandoned whaling station and a British base, victims of the 1967–70 eruptions, are among the first things one sees after sailing through the narrow cleft in the island walls into the drowned crater that fills the island center. I feel ghosts around me as I walk about there—sealers from the 1820s; the first dedicated scientific expedition to Antarctica; whalers; the earliest Antarctic aviators; Argentine, British, and Chilean scientific bases. . . . On a sunny day, the hillsides, where they are not covered by glaciers, display a wonderful color palette of black and red lava. Steam rises from black sand beaches where one may also see the occasional penguin or fur seal. On the outside wall of the island, facing the sea, there is a huge chinstrap penguin rookery. This is a very special island, with a unique human and geological story, and it is indeed one of my favorite places in the world. “

domain: travel


text:
The pink hairdo debuted around the world Thursday (May 6) morning is courtesy of BTS' RM, who showed off his new hair color in a set of candid pictures.
BTS' official Twitter account shared four pictures of RM playing with his white Spitz named Rap Mon (nicknamed Monie) with a Korean caption that reads "Puppy is difficult" when translated to English. But the ARMY was having a slightly difficult time understanding his decision to return to the pastel color after he said he wouldn't last September.
During an interview with AskAnythingChat shortly after the release of BTS' first English-language single and Billboard Hot 100 No. 1 hit "Dynamite," RM (real name Kim Nam-joon) said he wouldn't dye his hair pink because it reminded him of his "dark side." But while promoting the K-pop supergroup's second English-language single "Butter" in his teaser clip, fans thought they had caught a glimpse of the pink hair while he was sauntering into the frame and posing in front of the camera.

domain: celebrity