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Kasprzak A, Adamek A, Przybyszewska W, Pyda P, Szmeja J, Seraszek-Jaros A, Lanzafame A, Surdacka A, Mozer-Lisewska I, Koczorowska M. Insulin-like growth factor-1 mRNA isoforms and insulin-like growth factor-1 receptor mRNA expression in chronic hepatitis C. World J Gastroenterol 2015; 21(13): 3867-3875
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Aldona Kasprzak, MD, PhD, Professor, Department of Histology and Embryology, Poznan University of Medical Sciences, 6 Swiecicki Street, 60-781 Poznan, Poland. email@example.com
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Insulin-like growth factor-1 mRNA isoforms and insulin-like growth factor-1 receptor mRNA expression in chronic hepatitis C
Aldona Kasprzak, Agnieszka Adamek, Wiesława Przybyszewska, Przemysław Pyda, Jacek Szmeja, Agnieszka Seraszek-Jaros, Agata Lanzafame, Anna Surdacka, Iwona Mozer-Lisewska, Maria Koczorowska
Aldona Kasprzak, Wiesława Przybyszewska, Department of Histology and Embryology, Poznan University of Medical Sciences, 60-781 Poznan, Poland
Agnieszka Adamek, Iwona Mozer-Lisewska, Department of Infectious Diseases, Poznan University of Medical Sciences, 61-285 Poznan, Poland
Przemysław Pyda, Jacek Szmeja, Department of General Surgery, Gastroenterological Oncology and Plastic Surgery, Poznan University of Medical Sciences, 60-355 Poznan, Poland
Agnieszka Seraszek-Jaros, Department of Bioinformatics and Computational Biology, Chair of Clinical Pathomorphology, Poznan University of Medical Sciences, 60-529 Poznan, Poland
Agata Lanzafame, Anna Surdacka, Department and Clinics of Conservative Dentistry and Periodontology, Poznan University of Medical Sciences, 60-812, Poznan, Poland
Maria Koczorowska, Department of Molecular Virology, Adam Mickiewicz University, 61-614 Poznan, Poland
ORCID number: $[AuthorORCIDs]
Author contributions: Kasprzak A and Adamek A contributed equally to this work; Kasprzak A and Adamek A designed the research; Kasprzak A, Adamek A, Przybyszewska W, Pyda P and Szmeja J performed the research; Kasprzak A, Adamek A, Seraszek-Jaros A, Lanzafame A, Surdacka A and Mozer-Lisewska I analyzed the data; Seraszek-Jaros A performed biostatistics; Koczorowska M contributed new reagents/analytic tools; Kasprzak A and Adamek A wrote the paper.
Supported by Minister of Education and Science, Warsaw, Poland, No. NN401009437.
Ethics approval: Committee on Bioethics, Poznan University of Medical Sciences, 61-701 Poznan, Poland (No. 22/09).
Institutional animal care and use committee: Not applicable.
Conflict-of-interest: All authors declare that they have no relevant or material financial interests that relate to the research described in this paper.
Data sharing: No additional data available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Aldona Kasprzak, MD, PhD, Professor, Department of Histology and Embryology, Poznan University of Medical Sciences, 6 Swiecicki Street, 60-781 Poznan, Poland. firstname.lastname@example.org
Telephone: +48-61-8546441 Fax: +48-61-8546440
Received: August 26, 2014 Peer-review started: August 27, 2014 First decision: September 15, 2014 Revised: October 11, 2014 Accepted: January 8, 2015 Article in press: January 8, 2015 Published online: April 7, 2015
AIM: To evaluate the expression of different insulin-like growth factor (IGF)-1 mRNA isoforms and IGF-1 receptor (IGF-1R) mRNA in hepatitis C virus (HCV)-infected livers.
METHODS: Thirty-four liver biopsy specimens from chronic hepatitis C (CH-C) patients were obtained before anti-viral therapy. Inflammatory activity (grading) and advancement of fibrosis (staging) were evaluated using a modified point scale of METAVIR. The samples were analyzed using quantitative real-time PCR technique. From fragments of liver biopsies and control liver that were divided and ground in liquid nitrogen, RNA was isolated using RNeasy Fibrous Tissue Mini Kit according to the manufacturer’s instruction. Expression levels of IGF-1 mRNA isoforms (IGF-1A, IGF-1B, IGF-1C, P1, and P2) and IGF-1R mRNA were determined through normalization of copy numbers in samples as related to reference genes: glyceraldehyde-3-phosphate dehydrogenase and hydroxymethylbilane synthase. Results on liver expression of the IGF-1 mRNA isoforms and IGF-1R transcript were compared to histological alterations in liver biopsies and with selected clinical data in the patients. Statistical analysis was performed using Statistica PL v. 9 software.
RESULTS: The study showed differences in quantitative expression of IGF-1 mRNA variants in HCV-infected livers, as compared to the control. Higher relative expression of total IGF-1 mRNA and of IGF-1 mRNAs isoforms (P1, A, and C) in HCV-infected livers as compared to the control were detected. Within both groups, expression of the IGF-1A mRNA isoform significantly prevailed over expressions of B and C isoforms. Expression of P1 mRNA was higher than that of P2 only in CH-C. Very high positive correlations were detected between reciprocal expressions of IGF-1 mRNA isoforms P1 and P2 (r = 0.876). Expression of P1 and P2 mRNA correlated with IGF-1A mRNA (r = 0.891; r = 0.821, respectively), with IGF-1B mRNA (r = 0.854; r = 0.813, respectively), and with IGF-1C mRNA (r = 0.839; r = 0.741, respectively). Expression of IGF-1A mRNA significantly correlated with isoform B and C mRNA (r = 0.956; r = 0.869, respectively), and B with C isoforms (r = 0.868) (P < 0.05 in all cases). Lower expression of IGF-1A and B transcripts was noted in the more advanced liver grading (G2) as compared to G1. Multiple negative correlations were detected between expression of various IGF-1 transcripts and clinical data (e.g., alpha fetoprotein, HCV RNA, steatosis, grading, and staging). Expression of IGF-1R mRNA manifested positive correlation with grading and HCV-RNA.
CONCLUSION: Differences in quantitative expression of IGF-1 mRNA isoforms in HCV-infected livers, as compared to the control, suggest that HCV may induce alteration of IGF-1 splicing profile.
Core tip: Hepatitis C virus (HCV) may induce alteration of insulin-like growth factor (IGF)-1 splicing profile. A quantitative polymerase chain reaction analysis confirmed higher relative expression of total IGF-1 mRNA and of IGF-1 mRNAs isoforms P1, A, and C in HCV-infected livers as compared to the control. An increase in inflammatory activity (grading) of HCV-infected livers was linked to decreased IGF-1 mRNA expression, an altered profile of mRNA isoforms, and to an increase in IGF-1R mRNA expression. Decreased expression level of IGF-1 mRNA isoforms and an increased liver expression of IGF-1R mRNA were associated with indicators of liver damage (e.g., grading, staging, steatosis, and liver serum enzyme activity), and may be of prognostic significance.
Citation: Kasprzak A, Adamek A, Przybyszewska W, Pyda P, Szmeja J, Seraszek-Jaros A, Lanzafame A, Surdacka A, Mozer-Lisewska I, Koczorowska M. Insulin-like growth factor-1 mRNA isoforms and insulin-like growth factor-1 receptor mRNA expression in chronic hepatitis C. World J Gastroenterol 2015; 21(13): 3867-3875
Hepatitis C virus (HCV) is the major causative agent of hepatocellular carcinoma (HCC), mainly through the indirect pathways of chronic inflammation, cell death, and proliferation[1,2]. The function of the insulin-like growth factor (IGF) system in the promotion of cell growth and neoplastic transformation has been previously described[3,4]. The IGF ligands (IGF-1 and -2) bind to the receptors, which include: IGF-1 receptor (IGF-1R), IGF-2R, insulin receptor, and hybrid receptors (IGF-1R/insulin receptor). The involvement of IGF receptors in the maintenance of the transformed hepatocyte phenotype was also described[5-9]. Most studies documented an increase in IGF-1R expression from preneoplastic lesions to developed HCC[5,9,10]. Other studies found no significant difference in the expression of IGF-1R mRNA between HCC and a control. Similarly, an increase in expression of IGF-1R could not be noted in cultured human hepatocytes. In a few studies on patients with liver cirrhosis and chronic hepatitis C (CH-C), the amounts of IGF-1R transcript were found to increase as compared to the normal liver[12,13]. A more thorough recognition of the role played by IGF system in hepatic carcinogenesis is thought to improve HCC therapy[3,9,14].
The six exons of IGF-1 are alternatively spliced into multiple transcripts, encoding specific circulating and tissue-specific isoforms of the IGF-1 peptide. At the 5’end of the gene, different promoters (P1 and P2), in combination with alternative transcription start sites and differential splicing, generate the mutually-exclusive class 1 and class 2 IGF-1 isoforms[15-17]. At the 3’end of the gene, alternative splicing gives rise to at least three subsets of RNA transcripts, each encoding three distinct C-terminal portions of the unique E-peptide, as well as the 3’-UTR[15,18,19]. Exon 3 encodes parts of the signal peptide and the mature peptide common to all isoforms, while exon 4 encodes the rest of the mature peptide and the proximal part of the E domain. The composition of nucleotides in exons 5 and 6 determines the formation of isoforms A (Ea), B (Eb), and C (Ec) within classes 1 and 2. The biochemical mechanism which controls use of IGF-1 promoters 1 or 2 in alternate splicing remains poorly recognized[16,21-23]. Studies on human liver RNA have demonstrated that IGF-1 transcript undergoes alternate splicing that contains exons 3 and 4, as well as 49 bp of exon 5 and exon 6 (exon 4-5-6). The role of IGF-1 alternate splicing has been best recognized in skeletal muscular tissue[25-28] and nervous tissue[29,30]. A differential profile of IGF-1 mRNA isoforms was demonstrated in different tumors[31-34]. It remains unknown as to whether HCV and its oncogenic proteins (C, NS3, and NS5A) in vivo may induce alterations in the profile of hepatic IGF-1 gene expression.
This study aimed to evaluate the expression of various IGF-1 mRNA isoforms (P1, P2, 1A, 1B, and 1C) and IGF-1R mRNA in chronically HCV-infected livers. Herein we examined if IGF-1 alternative splicing is associated with the degree of liver damage (grading and staging) caused by HCV virus. Results on liver expression of the IGF-1 mRNA isoforms and IGF-1R transcript were compared to histological alterations in liver biopsies and with selected clinical data in the patients. Data concerning changes in IGF-1 alternative splicing in CH-C have not been published up to now. The relationship between liver expression of mRNA IGF-1 isoforms and progression of CH-C to HCC is unclear.
MATERIALS AND METHODS
The examined group consisted of 34 patients (age: 18-63 years; 18 men and 16 women) with CH-C who were diagnosed and treated in the Department of Infectious Diseases, Poznan University of Medical Sciences in Poznan from 2010 to 2012. Patients were referred to an anti-viral treatment and not previously treated. Infections with other hepatotropic viruses (HBV, HCMV, or EBV) or other reasons of liver damage were excluded (e.g., alcohol abuse, autoimmune hepatitis, NASH, drugs, or history of anti-cancer therapy). Patients with diabetes mellitus, kidney failure or any hormones disturbances were not included in the group. In the study, we used basic clinical data on HCV-infected patients, as well as other results involving biochemical tests on peripheral blood, results of ELISA tests (glucose, insulin), and histopathological examination of liver biopsies. Presence of HCV-specific antibodies was tested using chemiluminescence and ARCHITECT Anti-HCV kits (ABBOTT, Wiesbaden, Germany) in ARCHITECT and 2000 analyzers (ABBOTT). Infection with HCV was confirmed by estimating serum HCV-RNA via the application of GeneProof HEPATITIS C VIRUS HCV tests (GeneProof a.s., Brno, Czech Republic) and manifesting sensitivity of 50 IU/mL. In all patients, HCV genotype was estimated (VERSANT HCV GENOTYPE 2.0 ASSAY, LiPA).
Negative tissue control (n = 7) (patient age: 35-72 years; 4 men, 3 women) involved liver fragments with no morphological traits of organ pathology that were perioperatively sampled from the vicinity of the dissected focal lesion in the liver and a single liver biopsy taken from a patient to diagnose the reasons for elevated aminotransferase activity. Said patient proved to be HCV-negative (absence of specific antibodies and of HCV RNA). The remaining control material also originated from HCV- and HBV-negative patients. Age and sex of the patients were known. The control material was obtained from the Chair and Department of General Surgery, Gastroenterological Oncology and Plastic Surgery, Poznan University of Medical Sciences in Poznan.
Liver biopsy was done in all cases as a routine procedure before antiviral therapy. Basing on USG tests and alpha fetoprotein (AFP) levels, none of the patients had neoplastic growth (HCC). Written informed consent was obtained from every patient before liver biopsy and approval for the study was granted by the institution’s ethical committee (No. 22/09). The excised liver fragment (HCV-infected and control) was divided, cutting off its terminal 0.5 cm fragment, which was immersed in RNA Stabilization Solution (RNAlater®, Applied Biosystems) at -80 °C until use. The remaining part of the fragments obtained from patients were fixed in a buffered 10% solution of formalin and embedded in paraffin. About 5 μm-thick preparations were stained with hematoxylin and eosin and then silver impregnated using standard techniques. Inflammatory activity (grading) and advancement of fibrosis (staging) were evaluated using a modified point scale of METAVIR. This score is composed of a two-letter and two-number scoring system: histological activity (grading: G0 - no activity, G1 - mild activity, G2 - moderate activity, G3 - severe activity) and fibrosis (staging: S0 - no fibrosis, S1 - portal fibrosis without septa, S2 - portal fibrosis with rare septa, S3 - numerous septa without cirrhosis, S4 - cirrhosis). Fatty degeneration of the liver was evaluated using a point scale, in which grade 0 corresponded to an absence of fatty degeneration, while grades 1 and 2 corresponded to < 30% and ≥ 30%-70% hepatocytes showing traits of fatty degeneration, respectively.
Technique of quantitative real-time PCR
RNA was isolated using RNeasy Fibrous Tissue Mini Kit (QIAGEN), according to the manufacturer’s instructions, from fragments of liver biopsies and control liver that were divided and ground in liquid nitrogen. In the course of the procedure, traces of DNA contamination were eliminated using DNases. Total RNA was dissolved in RNase-free water. Quality of RNA preparations was consecutively checked using electrophoresis in an agar-formaldehyde gel. RNA content was quantitated by spectrophotometry. Every RNA sample was subjected to additional digestion with DNase using RNase-Free DNase Set (QIAGEN) in order to avoid contamination with genomic DNA. Subsequently, 1 μg RNA from every sample was subjected to reverse transcription using QuantiTect Reverse Transcription Kit (QIAGEN) and cDNA was obtained. Analysis of gene expression was performed using primers specific for individual mRNA IGF-1 isoforms (as previously described) and selected reference genes: GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and HMBS (hydroxymethylbilane synthase). The primer sequences for IGF-1R were: forward 5’-GGGAATGGAGTGCTGTATG-3’, reverse 5’-CACAGAAGCTTCGTTGAGAA-3’, GAPDH forward 5’-AAGGTCGGAGTCAACGGATTT-3’, reverse 5’-ACCAGAGTTAAAAGCAGCCCTG-3’, HMBS forward 5’-TGCAACGGCGGAAGAAAA-3’, and reverse 5’-ACGAGGCTTTCAATGTTGCC-3’. The reactions were performed in a final volume of 10 μL. Each sample contained 15 ng/μL cDNA and a mixture of reagents forming the SYBR Green PCR master mix (Applied Biosystems, United Kingdom), with each primer pair at a concentration of 0.25 mmol/L. The reaction was conducted using an automated fluorimeter (Rotor-Gene 6000, Corbett Research). The PCR program was as follows: (1) preliminary denaturation, 95 °C, 10 min; (2) denaturation, 95 °C, 10-15 s; (3) primer annealing, 53-67 °C, 15-35 s; and (4) elongation, 72 °C, 15-40 s. The number of cycles was 40-50. The initial quantity of the product was calculated in relation to the standard curve. Presence of an appropriate product was evaluated by determination of the melting point for a specific PCR product. All samples were amplified in duplicate or triplicate and, in cases in which the results varied by more than 15%, the reactions were repeated. Expression levels of IGF-1 mRNA isoforms (IGF-1A, IGF-1B, IGF-1C, P1, and P2), and IGF-1R mRNA were determined through normalization of copy numbers in samples as related to reference genes (housekeeping genes). In the normalization, reference genes were accepted to include liver-specific GAPDH and HMBS genes, according to literature data.
At the first stage of statistical analysis, the consistency of all results with the normal distribution of Gauss was verified by using the Shapiro-Wilk test. Parameters of descriptive statistics were subsequently calculated (arithmetic mean, standard deviation, median value, and minimum and maximum value). Data related to quantitative gene expression of IGF-1 mRNA isoforms, IGF-1R mRNA were compared to data for normal liver (negative control) using Mann-Whitney’s test (a non-parametric test for unlinked variables for two groups). In cases of linked variables, the Wilcoxon test was used. For comparing more than two groups, Kruskal-Wallis and multiple comparison Dunn tests were employed. For comparison of the percentage shares of IGF-1 mRNA isoforms, the test of differences between two structural indices was employed. Pearson’s correlation and Spearman’s rank correlation were used to correlate values of variables. Effect of age was also analyzed to determine any correlation between IGF-1 mRNA isoforms, and IGF-1R transcript, and selected clinical data. The results were thought to be statistically significant at P < 0.05. Statistical analysis was performed using Statistica PL v. 9 software (Statsoft, Inc., Tulsa, OK, United States).
Expression of mRNA class 1 (I) and 2 (II) as related to the used promoter P1 or P2
Both among the HCV-infected patients and in the control, transcription of IGF-1 from the first promoter (P1) (class 1) prevailed. In the CH-C group, the shares of transcripts from class 1 (from P1) (76.4%) and class 2 (from P2) (23.6%) were similar to those in the control (62.7% and 37.3%, respectively) (P > 0.05) (Figure 1).
Figure 1 Shares (%) of studied insulin-like growth factor-1 mRNA isoforms of class 1 (I) and class 2 (II) (depending upon employed promoter P1 or P2) as related to the total mRNA for both insulin-like growth factor-1 promoters (100%) in chronic hepatitis C patients and in the control.
IGF: Insulin-like growth factor; CH-C: Chronic hepatitis C.
Expression of mRNA isoforms: IGF-1A, IGF-1B, and IGF-1C - shares in percentages
In the CH-C group, expression of the IGF-1A (A) splicing form of mRNA prevailed (92% of all transcripts). Similarly, in the control group, the highest expression was manifested by the A isoform, followed by B and C (Figure 2). Expression of the A isoform was significantly higher in the CH-C group than in the control (92% vs 85%), while expression of isoform B was lower than in the control (5.2% vs 12.9%). No significant differences could be demonstrated between the two compared groups in shares of expressed P1 and P2 transcripts (Figure 3).
Figure 2 Shares (%) of splicing mRNA isoforms for insulin-like growth factor-1 (1A, 1B, 1C) as related to the total amount of mRNA for all three insulin-like growth factor-1 isoforms (100%) in chronic hepatitis C patients and in the control.
IGF: Insulin-like growth factor; CH-C: Chronic hepatitis C.
Figure 3 Comparison of shares (%) in expression of insulin-like growth factor-1 mRNA isoforms in patients chronically infected with hepatitis C virus and in the control.bP < 0.01 vs control.
Quantitative analysis of IGF-1 mRNA isoforms in the CH-C and control groups
Detailed analysis of the relative expression manifested by each IGF-1 mRNA isoform, normalized against reference genes (GAPDH and HMBS), demonstrated a significantly higher expression of IGF-1A and IGF-1C mRNA isoforms, of P1 transcripts (class 1), and of total IGF-1 mRNA in the CH-C group as compared to the control. A significantly higher expression of total IGF-1 and all isoforms and classes of IGF-1 mRNA was detected in women as compared to men (Table 1).
Table 1 Quantitative expression of insulin-like growth factor-1 mRNA isoforms, total mRNA insulin-like growth factor-1, and insulin-like growth factor-1R mRNA (mean ± SD) normalized in relation to housekeeping genes in chronic hepatitis C and in control organ.
Control(n = 7)
CH-C(n = 34)
Women (n = 16)
Men (n = 18)
class 1 (P1)
0.19 ± 0.21
0.45 ± 0.45
P = 0.025
0.63 ± 0.59
0.28 ± 0.17
P = 0.003
class 2 (P2)
0.08 ± 0.07
0.15 ± 0.17
P = 0.164
0.20 ± 0.15
0.11 ± 0.18
P = 0.006
1.30 ± 1.31
5.28 ± 9.94
P = 0.009
7.88 ± 13.83
2.97 ± 3.32
P = 0.027
0.18 ± 0.18
0.28 ± 0.40
P = 0.486
0.35 ± 0.45
0.22 ± 0.35
P = 0.042
0.04 ± 0.05
0.21 ± 0.57
P = 0.031
0.35 ± 0.82
0.08 ± 0.06
P = 0.046
1.52 ± 1.52
5.77 ± 10.85
P = 0.014
8.58 ± 15.09
3.26 ± 3.70
P = 0.027
0.26 ± 0.18
0.14 ± 0.04
P = 0.016
0.14 ± 0.05
0.14 ± 0.04
P = 0.295
IGF-1: Insulin-like growth factor 1; CH-C: Chronic hepatitis C.
Analysis of IGF-1 mRNA isoforms in HCV-infected livers as related to activity of inflammation (grading) and of liver fibrosis (staging)
A liver with more intense inflammatory lesions (grading 2, G2) contained a lower expression of all IGF-1 isoforms as compared to their expression in a liver with G1, with the exception of the IGF-1C isoform. No significant differences could be determined between the expression of all IGF-1 mRNA isoforms and the different stages of fibrosis (P > 0.05 in all the cases) (Table 2).
Table 2 Tissue expression of insulin-like growth factor-1 mRNA isoforms and insulin-like growth factor-1R mRNA (mean ± SD) as related to grading and staging in hepatitis C virus-infected livers.
1Parameters evaluated in a semi-quantitative scale (see Material and Methods). IGF-1: Insulin-like growth factor 1.
Expression of IGF-1R mRNA
In the CH-C group, expression of IGF-1R mRNA was significantly lower than in the control. No sex-related differences were detected in the expression of the IGF-1R transcript (Table 1). Expression of IGF-1R mRNA was higher in livers with higher grading (G2) as compared to livers with G1 (Table 2).
Expression of various transcripts of IGF-1 and IGF-1R vs the clinical data
Isoforms of IGF-1 mRNA: Expression of total IGF-1 mRNA and of all mRNA isoforms manifested a very strong negative correlation with patient age and BMI value. Expression of all isoforms also demonstrated negative correlations with liver steatosis. Highly negative correlations were detected with activity of ALT, and slightly less pronounced ones (though still negative) with activity of AST. Expression of mRNA isoforms A, B, P1, and P2 showed negative correlations with liver grading. Very poor correlation was detected between liver fibrosis and expression of P2 mRNA. For expression of P2 mRNA, a relatively poor negative correlation was documented, with AFP concentration and HCV viral load. Similarly low were the negative correlations between HCV viral load and expression of IGF-1A mRNA and total IGF-1 mRNA in HCV-infected liver (Table 3).
Table 3 Values of Spearman’s coefficient for correlation between expression of insulin-like growth factor-1, insulin-like growth factor-1R mRNAs, and clinical data in hepatitis C virus-infected patients.
1The numbers indicate values of r coefficient for which the P < 0.05. IGF-1: Insulin-like growth factor 1; HCV: Hepatitis C virus; AFP: Alpha fetoprotein.
Transcripts of IGF-1R: Expression of IGF-1R mRNA manifested poor positive correlations with age of HCV-infected patients. Moreover, positive correlations were detected between tissue expression of IGF-1R mRNA, grading, and HCV-RNA (Table 3).
Reciprocal correlations between expressions of IGF-1 mRNA isoforms and IGF-1R mRNA
Very high positive Spearman’s correlations were detected between reciprocal expressions of IGF-1 mRNA isoforms P1 and P2 (r = 0.876), P1 and A (r = 0.891), P1 and B (r = 0.854), P1 and C (r = 0.839); P2 and A (r = 0.821), P2 and B (r = 0.813), P2 and C (r = 0.741), A and B (r = 0.956), A and C (r = 0.869), and B and C (r = 0.868) (P < 0.05 in all cases) in patients with HCV infection. In livers with CH-C, additional significantly weak negative correlations were detected between expression of IGF-1R mRNA and quantities of mRNAs for isoforms of IGF-1A (r = -0.397), IGF-1B mRNAs (r = -0.419), and for total expression of IGF-1 mRNA (r = -0.397) (P < 0.05 in all cases).
In control livers, we confirmed very high positive Spearman’s correlation between reciprocal expression of all IGF-1 mRNA isoforms (data not shown). No significant relationships were detected between expression of IGF-1 mRNA isoforms and IGF-1R mRNA in control livers (data not shown).
Studies on expression of various IGF-1 mRNA isoforms have been performed for the first time on livers with CH-C. Our results point to the prevalent expression of mRNA from P1 promoter of IGF-1, both on the control liver and in liver with chronic HCV infection. This confirms the involvement of P1 in the production of 60%-65% and of P2 in the production of approximately 25% of IGF-1 transcripts, which was originally demonstrated in rat liver[17,22]. The percentage shares of the remaining IGF-1 mRNA isoforms (A, B, and C) in the control and HCV-infected liver also demonstrated the prevalence of mRNA isoform A overexpression of mRNAs of the remaining isoforms. In the literature data, there are no references to such results. In studies on human papilloma virus-positive and -negative tissues of uterine cervix carcinoma, a significant prevalence was demonstrated of IGF-1B share over remaining isoforms in tissues with uterine cervix carcinoma as compared to the remaining stages of carcinogenesis, suggesting that this form of the transcript may lead to the formation of the peptide Eb with strongly mitogenic properties.
Analysis of the relative values of expression manifested by various IGF-1 mRNA isoforms in the livers of the two examined groups confirmed the quantitative prevalence of isoform A over the remaining variants of IGF-1 transcripts. Prevalence of expression for IGF-1A over IGF-1B was also documented by Ohtsuki et al in such organs as the uterus, ovaries, liver, and kidneys in mice. The authors accentuated organ-specific control of transcription manifested by the gene in the course of development. The prevalence (as high as tenfold) of IGF-1A over IGF-1B mRNA transcripts, both in in vivo conditions (human liver) and in culture (hepatoma cells, macrophage-like cells, and fibroblasts) using RT-PCR technique was also demonstrated by Nagaoka’s team. The dominant expression of IGF-1A transcripts among other transcripts in the liver itself has been described by other authors[16,40].
A detailed quantitative analysis confirmed the higher expression of mRNAs for A and C isoforms (and of total IGF-1 mRNA) in our HCV infected patients as compared to the control. Expression of all studied IGF-1 transcripts has also been significantly higher in women as compared to men with HCV infection. Koczorowska et al demonstrated a quantitatively higher expression of total IGF-1 mRNA in the precancerous stages of the uterine cervix, as well as the activity of both gene promoters at the stage of intraepithelial neoplasia. Brokaw et al demonstrated a significant relationship between higher expression of IGF-1A isoform expression and progression of ovarian carcinoma. Nevertheless, the role of IGF-1 isoform A remains unclear. It has been argued that peptide Ea arising from it may exert mitogenic effects and inhibit growth of neoplastic cells.
The quantitative prevalence of expression manifested by IGF-1 isoforms A and C in HCV-infected livers, as compared to the healthy organ, may point to the influence of HCV on alterations in the splicing profile of the gene in humans. Our other observations suggest the influence of mainly non-structural proteins (NS3 and NS5A) on an increase of IGF-1 protein expression in HCV-infected livers and on the augmentation of IGF-1 P1 and P2 mRNA expression mRNA (unpublished data). Another explanation may involve a compensatory increase in the production of IGF-1 mRNA (particularly that of the IGF-1A isoform) under the effect of locally acting growth factors, proinflammatory cytokines, and an increased regeneration of the organ in CH-C. An increased production of two principal IGF-1 transcripts in rabbit skeletal muscle [i.e., muscle L.IGF-1 (resembling isoform A in the liver) and of mRNA mechano-growth factor (MGF, isoform IGF-1B in rabbits; a homologue of IGF-1C in humans)] was detected under muscle stretching and electrical stimulation. Mechanisms of differential IGF-1 mRNA stability were also described[44,45], including the effects of various RNA-binding proteins (e.g., Hu and hnRNP families) on this process. In the patients analyzed in this study, hepatic expression of IGF-1 mRNA isoforms has been significantly lower upon higher activity of inflammation. The relationships have not been as spectacular as those related to fibrosis. We may have examined an insufficient number of patients with more advanced staging (3 patients with staging 3, one with staging 4). Studies have also documented a negative relationship between expression of all IGF-1 mRNA isoforms, steatosis, and ALT activity. The results indicate a coexistence of the more pronounced inflammatory/necrotic lesions in the liver and a lower hepatic production of IGF-1 mRNA isoforms. Since the literature data contains no references to the role of IGF-1 mRNA isoforms in the progression of HCV-related hepatic diseases, the results seem to be pioneering. The other few reports on the subject mainly concern human tumors[31-34]. In the case of hepatoma and HCV-associated HCC, expression of IGF-1 mRNA was studied, but with no references to specific isoforms[8,13,47]. Su et al, using Northern blotting, demonstrated numerous IGF-1 transcripts of various sizes in hepatoma cells and in lines of other neoplastic cells (HepG2, Huh-7, PLC/PRF/5, and Hep3B). Each fragment of neoplastic tissue showed a lower expression of IGF-1 mRNA as compared to control tissue. Using the quantitative technique of real-time RT-PCR (similarly to this study), Tovar et al demonstrated a decrease in IGF-1 mRNA expression at the early stages of HCV infection as compared to control, but with no significant differences between preliminary and advanced stages of HCC in patients. Stefano et al, using the RT-PCR technique, demonstrated a comparable quantity of IGF-1 mRNA in patients with CH-C and in the control. Another study on cultured rat hepatocytes, also using the RT-PCR technique, demonstrated a 50-fold increase in the expression of IGF-1B mRNA isoform (a homologue of human isoform C) in the cells of obese animals as compared to the hepatocytes of lean individuals. Armakolas et al, examining alternative splicing of IGF-1 in prostate carcinoma, demonstrated overexpression of IGF-1C (MGF) in the cells, thereby suggesting the role of the IGF-1 isoform in the stimulation of cell proliferation.
Expression of IGF-1R transcript proved to be lower in our HCV-infected livers than that in the control, and was accompanied by an increase in at least a portion of studied IGF-1 transcripts in the liver. It is known from earlier studies that, even if normal liver represents an organ with the highest production of IGF-1, it contains almost undetectable levels of IGF-1R mRNA. Moreover, marked expression of the receptor used to be noted mainly in Kupffer, vascular endothelial, and stellate cells, but not in hepatocytes[49,50]. In this study, expression of IGF-1R mRNA in the CH-C group has manifested positive correlation with grading. Therefore, it seems that local production of IGF-1R mRNA does not increase until HCV-associated hepatic lesions become pronounced, which was noted to be accompanied by a reduced production of total IGF-1 mRNA (and of certain isoforms). A lowered expression of IGF-1, coexisting with an increased production of IGF-1R in uterine carcinoma, was also described. In studies on more advanced stages of liver carcinogenesis, an increase in IGF-1R expression was already detected in preneoplastic focal lesions in the liver, in HCC itself, and in cell lines of human hepatoma[5,10]. No such increase was detected in human hepatocyte cultures. Studies by Price et al on the rat model demonstrated a higher expression of IGF-1R mRNA in the control liver as compared to HCC, and it was only the cooperation of two proteins [IGF-1 and hepatocyte growth factor-scatter factor (HGF-SF)] that stimulated mitogenesis of hepatocytes in the animals. In patients with liver cirrhosis and CH-C, increased amounts of IGF-1R transcript were detected as compared to normal liver[12,13]. Tovar et al failed to detect a significant difference in the expression of IGF-1R mRNA in hepatocellular carcinoma as compared to a control in human HCC.
Differences in quantitative expression of IGF-1 mRNA isoforms in HCV-infected livers, as compared to the control, suggest that HCV may induce alteration of the IGF-1 splicing profile. An increase in the grading of HCV-infected livers was linked to decreased IGF-1 mRNA expression, an altered profile of mRNA isoforms, and to an increase in IGF-1R mRNA expression. The demonstration of increased tissue expression of IGF-1R mRNA and the decreased expression level of IGF-1 mRNA isoforms, accentuated in line with increasing liver damage, may be of a prognostic significance.
Insulin-like growth factor 1 (IGF-1) represents a well-recognized pro-proliferative factor.
No studies are available on the role played by local expression involving various IGF-1 mRNA isoforms in chronic hepatitis C in vivo.
Innovations and breakthroughs
A quantitative polymerase chain reaction analysis used in the study confirmed the higher expression of total IGF-1 mRNA and of IGF-1 mRNAs isoforms A and C in hepatitis C virus-infected livers as compared to the control.
The demonstration of decreased expression levels of IGF-1 mRNA isoforms and an increased tissue expression of IGF-1R mRNA, associated with indicators of liver damage (e.g., grading, staging, steatosis, and liver serum enzyme activity), may be of prognostic significance.
This paper is intriguing but presents important clinical limitations, mostly relating to the low number of patients with chronic hepatitis C investigated, to the low prevalence of elderly patients with severe forms of the disease, and to the insufficient number of controls.
P- Reviewer: Liu YX, Sagnelli E S- Editor: Ma YJ L- Editor: Rutherford A E- Editor: Wang CH
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Price JA, Kovach SJ, Johnson T, Koniaris LG, Cahill PA, Sitzmann JV, McKillop IH. Insulin-like growth factor I is a comitogen for hepatocyte growth factor in a rat model of hepatocellular carcinoma.Hepatology. 2002;36:1089-1097.
Zhang YC, Wang XP, Zhang LY, Song AL, Kou ZM, Li XS. Effect of blocking IGF-I receptor on growth of human hepatocellular carcinoma cells.World J Gastroenterol. 2006;12:3977-3982.
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Maki RG. Small is beautiful: insulin-like growth factors and their role in growth, development, and cancer.J Clin Oncol. 2010;28:4985-4995.
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Adamo ML, Ben-Hur H, LeRoith D, Roberts CT. Transcription initiation in the two leader exons of the rat IGF-I gene occurs from disperse versus localized sites.Biochem Biophys Res Commun. 1991;176:887-893.
Simmons JG, Van Wyk JJ, Hoyt EC, Lund PK. Multiple transcription start sites in the rat insulin-like growth factor-I gene give rise to IGF-I mRNAs that encode different IGF-I precursors and are processed differently in vitro.Growth Factors. 1993;9:205-221.
Rotwein P. Two insulin-like growth factor I messenger RNAs are expressed in human liver.Proc Natl Acad Sci USA. 1986;83:77-81.
Temmerman L, Slonimsky E, Rosenthal N. Class 2 IGF-1 isoforms are dispensable for viability, growth and maintenance of IGF-1 serum levels.Growth Horm IGF Res. 2010;20:255-263.
Pell JM, Saunders JC, Gilmour RS. Differential regulation of transcription initiation from insulin-like growth factor-I (IGF-I) leader exons and of tissue IGF-I expression in response to changed growth hormone and nutritional status in sheep.Endocrinology. 1993;132:1797-1807.
Mittanck DW, Kim SW, Rotwein P. Essential promoter elements are located within the 5’ untranslated region of human insulin-like growth factor-I exon I.Mol Cell Endocrinol. 1997;126:153-163.
Wang L, Wang X, Adamo ML. Two putative GATA motifs in the proximal exon 1 promoter of the rat insulin-like growth factor I gene regulate basal promoter activity.Endocrinology. 2000;141:1118-1126.
Chew SL, Lavender P, Clark AJ, Ross RJ. An alternatively spliced human insulin-like growth factor-I transcript with hepatic tissue expression that diverts away from the mitogenic IBE1 peptide.Endocrinology. 1995;136:1939-1944.
Yang S, Alnaqeeb M, Simpson H, Goldspink G. Cloning and characterization of an IGF-1 isoform expressed in skeletal muscle subjected to stretch.J Muscle Res Cell Motil. 1996;17:487-495.
Mills P, Lafrenière JF, Benabdallah BF, El Fahime el M, Tremblay JP. A new pro-migratory activity on human myogenic precursor cells for a synthetic peptide within the E domain of the mechano growth factor.Exp Cell Res. 2007;313:527-537.
Barton ER, DeMeo J, Lei H. The insulin-like growth factor (IGF)-I E-peptides are required for isoform-specific gene expression and muscle hypertrophy after local IGF-I production.J Appl Physiol (1985). 2010;108:1069-1076.
Matheny RW, Nindl BC, Adamo ML. Minireview: Mechano-growth factor: a putative product of IGF-I gene expression involved in tissue repair and regeneration.Endocrinology. 2010;151:865-875.
Dluzniewska J, Sarnowska A, Beresewicz M, Johnson I, Srai SK, Ramesh B, Goldspink G, Górecki DC, Zabłocka B. A strong neuroprotective effect of the autonomous C-terminal peptide of IGF-1 Ec (MGF) in brain ischemia.FASEB J. 2005;19:1896-1898.
Quesada A, Micevych P, Handforth A. C-terminal mechano growth factor protects dopamine neurons: a novel peptide that induces heme oxygenase-1.Exp Neurol. 2009;220:255-266.
Brokaw J, Katsaros D, Wiley A, Lu L, Su D, Sochirca O, de la Longrais IA, Mayne S, Risch H, Yu H. IGF-I in epithelial ovarian cancer and its role in disease progression.Growth Factors. 2007;25:346-354.
Armakolas A, Philippou A, Panteleakou Z, Nezos A, Sourla A, Petraki C, Koutsilieris M. Preferential expression of IGF-1Ec (MGF) transcript in cancerous tissues of human prostate: evidence for a novel and autonomous growth factor activity of MGF E peptide in human prostate cancer cells.Prostate. 2010;70:1233-1242.
Koczorowska MM, Kwasniewska A, Gozdzicka-Jozefiak A. IGF1 mRNA isoform expression in the cervix of HPV-positive women with pre-cancerous and cancer lesions.Exp Ther Med. 2011;2:149-156.
Kasprzak A, Szaflarski W, Szmeja J, Andrzejewska M, Przybyszewska W, Kaczmarek E, Koczorowska M, Kościński T, Zabel M, Drews M. Differential expression of IGF-1 mRNA isoforms in colorectal carcinoma and normal colon tissue.Int J Oncol. 2013;42:305-316.
Kasprzak A, Adamek A, Przybyszewska W, Szaflarski W, Sterzyńska K, Seraszek A, Mozer-Lisewska I, Kaczmarek E, Biczysko W. Expression of IGF-I and viral proteins (C, NS3, NS5A) in livers of patients with chronic HCV infection.Adv Clin Exp Med. 2011;20:263-273.
Bedossa P, Poynard T. An algorithm for the grading of activity in chronic hepatitis C. The METAVIR Cooperative Study Group.Hepatology. 1996;24:289-293.
Cicinnati VR, Shen Q, Sotiropoulos GC, Radtke A, Gerken G, Beckebaum S. Validation of putative reference genes for gene expression studies in human hepatocellular carcinoma using real-time quantitative RT-PCR.BMC Cancer. 2008;8:350.
Ohtsuki T, Otsuki M, Murakami Y, Maekawa T, Yamamoto T, Akasaka K, Takeuchi S, Takahashi S. Organ-specific and age-dependent expression of insulin-like growth factor-I (IGF-I) mRNA variants: IGF-IA and IB mRNAs in the mouse.Zoolog Sci. 2005;22:1011-1021.
Nagaoka I, Someya A, Iwabuchi K, Yamashita T. Expression of insulin-like growth factor-IA and factor-IB mRNA in human liver, hepatoma cells, macrophage-like cells and fibroblasts.FEBS Lett. 1991;280:79-83.
Barton ER. The ABCs of IGF-I isoforms: impact on muscle hypertrophy and implications for repair.Appl Physiol Nutr Metab. 2006;31:791-797.
Tian XC, Chen MJ, Pantschenko AG, Yang TJ, Chen TT. Recombinant E-peptides of pro-IGF-I have mitogenic activity.Endocrinology. 1999;140:3387-3390.
Chen MJ, Chiou PP, Lin P, Lin CM, Siri S, Peck K, Chen TT. Suppression of growth and cancer-induced angiogenesis of aggressive human breast cancer cells (MDA-MB-231) on the chorioallantoic membrane of developing chicken embryos by E-peptide of pro-IGF-I.J Cell Biochem. 2007;101:1316-1327.
McKoy G, Ashley W, Mander J, Yang SY, Williams N, Russell B, Goldspink G. Expression of insulin growth factor-1 splice variants and structural genes in rabbit skeletal muscle induced by stretch and stimulation.J Physiol. 1999;516:583-592.
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What Are CertiK Audits and Why Are Audits Essential to the Health of DeFi?
Learn about the importance of security for smart contract protocols and about CertiK, a leading smart contract auditing service.
Importance of Security for Smart Contracts in DeFi
Unlike centralized information systems that are secured by the companies providing them, smart contracts place trust in the code rather than corporate entities. Code that can handle billions of dollars of funds.
As a smart contract platform offering faster transaction times and a fraction of the fees compared to Ethereum, Binance Smart Chain has shown significant growth in 2021. It currently holds $9B+ worth of assets in its smart contracts — up from $1.2B+ on February 1.
That growth demands further security for the DeFi assets stored on-chain.
Since smart contracts are immutable and cannot be changed once they’re deployed, code security must be perfected before release. A single bug or a security vulnerability can either mishandle funds or give hackers opportunities to exploit the contract.
Obviously, such a situation is intolerable since that can mean a potential loss of millions (if not billions) of dollars.
CertiK Audits for Industry-Leading Security
CertiK is a blockchain security firm pioneering the use of cutting-edge Formal Verification technology on smart contracts and blockchain networks.
In the three years since inception, CertiK has conducted 220+ audits, audited 118k+ lines of code, securing $8.03B of value.
CertiK has secured top crypto exchanges Binance, OKEx, and Huobi. In fact, the Binance Accelerator Fund has a system of securing the top platforms it funds through CertiK audits.
What’s more, CertiK has serviced more than 100 top-shelf blockchains and DeFi protocols, including but not limited to Binance, Tera, Bancor, Shapeshift, and Blockstack.
Even the popular Binance Smart Chain DeFi protocol, PancakeSwap, has had its smart contracts audited by CertiK.
So, how does a CertiK audit actually work?
Certik Audits Under the Hood
If you were to proceed with an audit of your protocol on Binance Smart Chain, this is the technical procedure your smart contracts will go through:
You provide your smart contracts and the specification of their intended behaviors to CertiK.
Your smart contracts will undergo the mathematical method of ****formal verification through the CertiK BSC Security Oracle.
Experts with experience from Google, Microsoft, Oracle, and Facebook will then manually review your contract.
CertiK will provide you with an audit report classifying vulnerabilities, proposing solutions for them, and showcasing the mathematical proofs from the formal verification. Part of the bundle will also be a hashed CertiK Verification Badge that features a scannable QR code.
It’s worth noting that this entire process can take as little as 48 hours.
Binance Smart Chain Bridges the Gap Between DeFi and CeFi
Binance Smart Chain aims to benefit users with features of centralized finance, including futures, margins, savings, staking, and pooling, while simultaneously providing features of decentralized finance such as lending, automated market makers (AMMs), and yield farming.
With the BSC Accelerator Fund providing much-needed assistance to upcoming platforms wishing to share the mission of Binance Smart Chain, it’s no wonder we see such impressive growth.
Thankfully, CertiK is there to add a layer of much-needed security to the growth of the Binance Smart Chain DeFi ecosystem.


text:
AMILOADED MEDIA HUB NEWS UPDATE
A landlord in Bulawayo’s North End suburb of Zimbabwe has been arrested for allegedly exposing his manhood to a tenant as she was dressing up in her room.
It was gathered that while the 25-year-old woman got dressed, her landlord identified as 24-year-old Shayne Mafoche reportedly peered through her window. Startled, the woman caught sight of Mafoche staring at her and subsequently confronted him about his actions. In response, Mafoche insulted her and hastily made his way towards the door.
Mafoche then proceeded to expose himself to the victim. He unzipped his pants and approached her while holding his private parts, making her flee the room.
The victim reported the incident at Kumalo Police Station, leading to Mafoche’s arrest. Inspector Abednico Ncube, the spokesperson for the Bulawayo Police, confirmed the incident and emphasized that Tom’s behaviour is considered a criminal offence.
He said; “Peeping is a criminal offence. The suspect’s actions were unlawful.”

domain: law


text:
ASCO’s Quality Training Program prepares physician-led oncology teams to design, implement, and lead successful quality improvement activities in their practice settings. ASCO’s program is developed by oncologists and oncology care specialists for oncology practice teams.
Specifically, the Quality Training Program
- Empowers practice teams to improve clinical care and operational performance
- Teaches teams how to balance quality improvement projects with demanding schedules and competing priorities
The 6-month program includes five days of in-person learning across three sessions, as well as hands-on learning at the participants’ practices. To ensure a well-rounded experience, sessions include seminars, case examples, and small group exercises.
Participants who complete the course are eligible for both CE credit and ABIM MOC points. The locations will change each year within the United States and also in international locations. If you are interested in hosting a QTP in your region or country, please complete our International Interest Survey and contact us.
Two to four members of each team, including the team leader, must attend three in-person meetings.
Winter 2019 hosted by City of Hope Comprehensive Cancer Center in Duarte/Monrovia, California.
The Summer 2019 session at ASCO Headquarters in Alexandria, Virginia is a thematic focus on oncology team wellness and avoiding burnout.
The session will have the following in-person meeting dates:
- Meeting 1: July 10-11
- Meeting 2: September 18-19
- Meeting 3: December 5
Registration deadline extended to June 10, 2019
*If you and your team are interested in attending a QTP summer course but would like to focus on a project not related to burnout, please email firstname.lastname@example.org
The Winter 2020 session will be held at ASCO Headquarters in Alexandria, Virginia.
The session will have the following in-person meeting dates:
- Meeting 1: January 16-17
- Meeting 2: March 19-20
- Meeting 3: June 26
Early application discount offered to all teams who apply in September! Early registration begins September 2, 2019.
Application Deadline is November 22, 2019
Knowledge and Skills Acquired During the Course
Upon completing the course, participants will know how to
- Form a high-functioning improvement team
- Identify clinical and operational targets for improvement
- Plan and implement change strategies
- Understand and use data for improvement
- Track performance over time
- Use process analysis tools
Conducting projects at the teams’ practice sites delivers a great benefit, allowing the teams to immediately translate training into action to improve cancer care quality.
Continuing Education (CE)
Accreditation Statement and CE Credit Designation
In support of improving patient care, the American Society of Clinical Oncology is jointly accredited by the Accreditation Council for Continuing Medical Education (ACCME), the Accreditation Council for Pharmacy Education (ACPE), and the American Nurses Credentialing Center (ANCC), to provide continuing education for the healthcare team.
AMA Credit Designation Statement - Physicians
The American Society of Clinical Oncology designates this live activity for 14.25 AMA PRA Category 1 Credits ™. Physicians should claim only the credit commensurate with the extent of their participation in the activity.
ANCC Credit Designation Statement - Nurses
The maximum number of hours awarded for this CE activity is 14.25 contact hours.
Participants will receive a brief survey before and after the Quality Training Program. This survey has two goals: To gauge progress toward learning outcomes as well as to provide feedback about the learning experience. Participants will be able to claim CE credits reflecting the extent of their participation in the Quality Training Program at the conclusion of the final learning session. CE certificates will be available for downloading and printing at that time.
Certificates of Participation and Attendance
Advanced practitioners are welcome to submit a request for a Certificate of Participation, which may enable nonphysicians to apply their Quality Training Program participation toward re-licensure. Please note, however, that all final decisions regarding the awarding of credits will be made by the licensing organization to which the credits are submitted.
Multidisciplinary experts in oncology and health care quality improvement lead the practice teams through the Quality Training Program. These experts bring many years of teaching and coaching experience to the program.
Quality Training Program Steering Group
- Michael Keng, MD
- Tim Gilligan, MD
- Doris Quinn, PhD
- Laurie Kaufman, MSN, RN
- John Bingham, MHA
- Vedner Guerrier, MBA, LSSBB
- Ashraf Mahmoud Mohamed, MD
- Anne C. Chiang, MD, PhD
Members of Steering Group may also serve as faculty and/or coaches.
Faculty and Improvement Coaches
- Charles Borden, MBA, Independent Consultant, Central & Southern New Jersey
- Stephanie Buia Amport, MBA, CPHQ, Yale New Haven Health
- Pelin Cinar, MD, UCSF Helen Diller Family Comprehensive Cancer Center
- Vedner Guerrier, MBA, RTT, LSSBB, Memorial Cancer Institute
- Valorie Harvey, MBA, Parkland Health and Hospital Systems
- Joseph Jacobson, MD, Chief Quality Officer, Dana-Farber Cancer Institute
- Laurie Kaufman, MSN, RN, CPHQ, CMQ, MD Anderson Cancer Center
- Michael Keng, MD, University of Virginia Health System
- Louise Man, MD, University of Virginia Health System
- Eric Martin, MD, Medical Oncology Hematology Consultants
- Duncan Phillips, MBA, Senior Process Improvement Specialist at the Elliot Health Systems
- Steve Power, MBA, Duke University
- Arif Kamal, MD, Duke Cancer Institute
- Prabjyot Singh, RN, MSN, MPH, Trinity Health
- Piyush Srivastava, MD, Kaiser Permanente
- Holley Stallings, RN, MPH, CPH, CPHQ, Norton Cancer Institute
- Carole Tremonti, RN, MBA, OCN, CCRP, Senior Manager of Clinical Quality, Dana-Farber Cancer Institute
Explore the Quality Training Program
The Quality Training Program is funded through the Conquer Cancer Foundation.
Amgen; Astellas; Novartis Oncology
This list reflects commitments as of March 22, 2019.

domain: medical


text:
Holly Gurbisz joins the cast of “Exploitation” – playing the role of Barbara.
The casting is nearing completion for this movie. The limited fan version should be out in January, for the ultrafans who participated in the crowdfunder. Remaining units will be available at a later date. The retail version is set fo April 2018.

domain: news


text:
Maan Gaye Mughall-E-Azam
The actors of the Kalakar Theatre Company are largely oblivious to the covert warfare that's rocking their small town. But when they learn that a mob boss is plotting a major bombing, they hatch their own plan to stop him. Mistaken-identity intrigue and wild physical comedy ensue as the troupe assumes a host of disguises and puts their acting chops to the test. Mallika Sherawat, Rahul Bose and Paresh Rawal star in Sanjay Chel's slapstick farce.
Rent DVDs for only $4.99 a month.
- Sanjay Chel
NRNot rated. This movie has not been rated by the MPAA.
Full Screen 1.33:1Subtitles
NoLanguage and sound
Hindi: Dolby Digital 2.0 StereoOther features
Color; interactive menus; scene access.
Maan Gaye Mughall-E-AzamClose
Common Sense Note
- Age appropriate
- Not an issue
- Depends on your kid and your family
- Not appropriate for kids of the age most likely to want to see it

domain: entertainment


Employers Use Health And Safety Excuses To Discriminate Against Workers With Disabilities
By Dave Reynolds, Inclusion Daily Express
December 9, 2002
LONDON, ENGLAND--The Trades Union Congress (TUC), a coalition of 70 member unions, reported this week that employers in the United Kingdom discriminate against workers with disabilities despite measures to protect them.
Companies are refusing to hire such workers, claiming that they present health and safety risks, in violation of the Disability Discrimination Act.
TUC found that workers who use wheelchairs were refused jobs because employers believed they would not be able to evacuate buildings in emergency situations or could "get in the way" of colleagues trying to escape.
Owen Tudor from the TUC told the BBC: "We've got examples of people who have been told by their employers that they are the problem that they are the safety hazard."
"Very often employers are looking for the problem and not for the solution," he said.


October 12, 2012, 2:45 P.M., Friday, New York local time — Apple iPad Mini rumors hit the “red” alert today, All Things D, Apple’s favorite blog site, has confirmed that the smaller iPad is headed this month and will start shipping next month.
The timing is unsurprising given the recent press releases favoring the Windows 8 operating system, Apple, the maker of the “PC killer,” the Apple iPad, is on the verge of adding yet another version this month — the iPad Mini. As reported by Apple’s favorite news outlet, All Things D, the device is on set to hit the stage later this month, specifically on October 23.
Unlike current iPads, the iPad Mini will reportedly sport a smaller 7.85-inch display. The smaller iPad is, obviously, Apple’s device that will compete with Google’s popular Nexus 7 Android tablet and Amazon’s Kindle Fire lineup. The iPad Mini is set to join the lower price tag category.
Analysts say the iPad Mini will likely sport a $249 to $299 price tag, cheaper than the traditional $499 entry-level 16GB models of older variants. Similar to previous Apple events, Apple’s iPad Mini will hit the market in November.
Apple is currently enjoying a major tablet PC market share, more than 70% according to latest channel checks while Android maintains its second place-only numbers and Microsoft with no “real tablet” market share as of the moment.
The iPad Mini, apart from securing Apple’s large tablet PC market share, is reportedly trying to lure customers away from Microsoft’s Windows 8 lineup which will fully support tablet PCs and hybrids (laptops with touch displays).
According to Business Insider, the iPad Mini will “embarrass” the Microsoft Surface, the software giant’s first tablet computer product that will run its latest operating system. Cupertino will announce the iPad Mini just three days before the Microsoft Surface event, and for the critic, putting the price tag of the iPad Mini on the board ahead of Microsoft, even if Microsoft changes the prices for the Surface, is a scenario that is “going to make Microsoft look pretty silly.”
Can the iPad Mini, with a cheaper price, sweet pre-Thanksgiving release date, and smaller display, outsell the bigger iPads and its rivals?
Sony Xperia T, TX, V and others getting Android Jelly Bean, soon!December 18th, 2012
BlackBerry 10 leak confirms similarity with Android?December 18th, 2012
Apple iPhone 5 sales in China reaches 2 millionDecember 18th, 2012
Nokia Lumia 920 software update arriving soon, to fix the PureView cameraDecember 17th, 2012
Exynos “security flaw” affecting the Samsung Galaxy Note 2, Galaxy S3?December 17th, 2012


text:
Cytogenetics in the Genomic Era
The Myelodysplastic Syndrome group, directed by Francesc Solé, has published the chapter "Cytogenetics in the genomic era", within the monograph coordinated by Torsten Haferlach "Deciphering molecular genetics in malignant haemopathies."
The presence of recurrent chromosomal abnormalities characterizes haematological malignancies. For this reason, cytogenetics is essential to establish the diagnosis and prognosis of these diseases. Chromosome banding analysis is mandatory for the diagnosis of chronic myeloid leukemia, eosinophilic neoplasms, myelodysplastic syndromes, and acute leukemias. However, in other myeloid neoplasms, such as chronic lymphocytic leukemia, non-Hodgkin's lymphoma, and multiple myeloma, the study should be complemented with fluorescence in situ hybridization (FISH) or microarrays, which overcome some of the deficiencies of band analysis to identify possible cryptic alterations or reciprocal translocations. Furthermore, in the genomic era, technologies such as next-generation sequencing are being used in routine clinical analysis, as they offer the possibility of studying mutations and copy number alterations in a single study with higher resolution or sensitivity. However, they require highly qualified personnel and greater standardization, especially in terms of data analysis, which limits their current applicability.
With these premises, the group of myelodysplastic syndromes has participated with a chapter within the monographic coordinated by Torsten Haferlach: "Deciphering molecular genetics in malignant haemopathies," with more referent collaborators in biomedical research and haematology. The Chapter has been recently published as an article in the journal Best Practice & Research Clinical Haematology: "Cytogenetics in the genomic era." In addition to Dr. Francesc Solé, Isabel Granada, Laura Palomo, Neus Ruiz, and Mar Mallo are co-authors of this Chapter.


text:
The impressive limestone cave located along the Ohio River is one
of the most compelling features of Cave-in-Rock State Park. With a
reputation as a robber's den and outlaw hideout, the cave looms large
in popular folk tales of the Illinois . . . — — Map (db m161218) HM
Meriwether Lewis, William Clark, and about twenty men passed this place in the fall of 1803 on their famous journey westward. Cave-In-Rock, a natural landmark near here, was well known and noted on some early maps of this area. The party continued . . . — — Map (db m154573) HM
This natural cave located nearby was known to European explorers as early as 1729. Its location was noted in diaries and journals of travelers. It was noted on maps used by Lewis and Clark.
Late in the 18th century, The Cave was the habitat for . . . — — Map (db m155030) HM
For 6 to 9 months a year, the Iron Furnace was "in blast."
These quiet hills came alive with the sounds of axes and
picks, horses and wagons, and the shouts of hardworking
men-24 hours a day,7 days a week!
A blast furnace requires large . . . — — Map (db m161399) HM
Before you stands the only remaining iron furnace structure in the entire state of Illinois. This priceless historic edifice became part of the Shawnee National Forest in 1950.
Outstanding in Its Field
Built in 1837, the Illinois Iron . . . — — Map (db m161400) HM
As the Big Creek Baptist Church is the oldest Baptist church in Illinois with continuous services, the site of the Big Creek Church is two miles west of this location. Elders Stephen Stilley and William Jones led in constituting the Big Creek Church — — Map (db m155007) HM
Fluorite, the official Illinois state mineral, was discovered in 1839 by James Anderson while digging a well near Fairview Landing one half mile SW of this site. Fluorite was a waste product until the steel industry began using the mineral in their . . . — — Map (db m154632) HM
People have been digging, processing and transporting fluorite (fluorspar) here since 1842. In the 1940s, Rosiclare and Hardin County led the U.S. in fluorite production and were known as "The Fluorite Capital of the World".
What is . . . — — Map (db m206198) HM
William Pell, Jr. settled here in 1832, founded Pell's Landing. In 1874, Pell's Landing was incorporated as Rose Clare and became the City of Rosiclare in 1932. Always an important river town, the discovery of fluorite and other minerals in the . . . — — Map (db m206199) HM
Travel the Byway and discover the importance of the Ohio River to the nation and the world. Experience the magnificent natural landscapes of southern Illinois while coming to know how the historic events and people that shaped the region and the . . . — — Map (db m196762) HM
Imagine this peaceful site abuzz with
activity! It took about 40 men to keep the
furnace "in blast” and many more to
support the workers and their families,
creating the furnace community.
At the Break of Day
The village . . . — — Map (db m161401) HM

domain: geography


text:
Today is TSC Global Awareness Day. You might ask what is TSC? It stands for Tuberous Sclerosis Complex. Tuberous Sclerosis affects the eyes, skin, brain, kidneys, liver, lungs, and some also have epilepsy.
I have Tuberous Sclerosis Complex and so does my youngest daughter Jennifer. I was officially diagnosed when she was 1. TSC in my case was a mutant gene, however once you have this rare disease the chances of having a child with TSC is 50/50. 1 in 2 chance of having a child with TSC. I do remember my parents being tested which came up clear. I am the youngest of 4 so the chance of my sisters or brother having TSC was ruled out when my parents tests came up clear. This meant something happened with the genes when i was conceived. When I was 6 or 7 the doctors removed some facial lumps off my face as they thought these were cancer. They also did radiation on my face and now it looks like this radiation has caused the Thyroid disease I now have. As a child I remember going to school showing off my scars. Show and Tell. I however have no recollection of these tubers being removed. It’s funny how we can remember some things and not others. I suppose that’s a good thing.
TSC affects everyone differently, some mild often going through life without even knowing they have this disease. Some as myself do not know until they have a child born with TSC. Others are severely affected and will need ongoing care for the rest of their life.
Jennifer was diagnosed with seizures at 6 months and at 1 with Tuberous Sclerosis. When i was 3 months pregnant with her, bearing in mind I had already had 1 daughter, they discovered I had large kidneys. My doctor at the time was amazed that no one had noticed my large kidneys when pregnant with my first child Sarah. I was sent to Kidney specialist where I was given a biopsy to rule out cancer. The results came in that I could continue with the pregnancy. When I was due to have Jennifer I went down to Perth, I was living in a remote country town called Derby in the far north-west of Western Australia. After her birth, I stayed in hospital for a bit longer where a whole lot of tests were done. I only remember some scans but these let the specialist know I had what they called polycystic cysts on them.
So when Jennifer turned 1 we had to go down to Perth for her to see a neurologist as she was having more seizures. I also went and saw the Kidney specialist who after hearing the diagnosis of Jennifer with Tuberous Sclerosis he advised that’s what I had. As I already had one child without TS he was hoping that she would be ok.
I would have 6 monthly appointments to ensure my kidney function was good and that was about it. On the other hand Jennifer was having a hard time. Her seizures were not controlled and they kept changing. Over the years we found that she was intellectually disabled, had Autism and behavior problems. These are all part of Tuberous Sclerosis.
Jennifer now lives in a group home where she has full-time care. She is 37 years old however is about a 3-5 year old in most things.
TSC affects most of my body. I have Angiomyolipomas (AMLs) on my kidneys and liver. I have all the skin manifestations, see link below. I have a little bit of calcification on my brain. I also have lung disease called Lymphangioleiomyomatosis or Lam for short which is another rare disease.
Currently I see a Lung specialist , a Dermatologist and a Rheumatologist at the Alfred Hospital in Melbourne, I also see a Nephrologist at the Royal Melbourne Hospital. As you would have read earlier I was seeing a kidney specialist. I was living in West Australia at the time I was diagnosed. Years later I moved to Victoria and did see a kidney specialist at the Royal Melbourne Hospital who in turn said I was fine and didn’t need to keep going to the clinic. So I have managed my TSC via my general practice doctor. The Dermatologist at the Alfred Hospital has prescribed me a new cream for the tubers on the face. These tiny little tubers would bleed if knocked or when I had a cold and the constant rubbing of tissues or hankerchiefs would be enough for them to bleed. They would bleed for hours, this is what can happen with the kidneys. This new cream has reduced the redness of the nose and cheeks. The little tubers at the side of the nose have gone, disappeared. How great is that! This new drug is certainly a wonder drug. Mind you it’s not cheap!
I have recently started taking Affinitor or Everolimus a mTOR to hopefully reduce the size of the angiomyolipomas. My dose is low and I will raise it slowly. Initially I started a high dose and within 5 days my ankles and feet were like balloons. I was instructed to stop taking this high dose. It took as many days off it to have the swelling go down. While I am taking Affinitor I won’t be needing to use the cream. Thank goodness as it keeps the cost down. The mTOR drug is on the PBS and as I am on a disability pension its cost is only $5.20 a month.
I have added a link so you the reader can click and go over to the Tuberous Sclerosis Complex Australia page and learn a bit more about this rare disease.
I thank you for your time in reading this. If you have any questions do not hesitate to ask me.


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Oil price crash is good news for climate and clean energy
Director at Independent Litigation Funders
The collapse in global oil prices to 5 year lows is causing a lot of speculation about what this means for climate change, and clean energy. High fuel costs, it is said, encourages the use of alternative fuels. Low fuel costs, on the other hand, simply encourage more consumption of conventional fuels.
True, but only up to a point. The price of the fuel as a commodity will also influence how much money is invested in oil projects – needed to maintain supply – and where the trillions of dollars in global capital will flow. In the case of oil, extraction is getting more and more expensive and the oil majors have struggled to find new oil reserves that make economic sense.
The fall in the oil price makes that even more difficult. So, from the point of view of the climate, that is a good thing, because that capital will likely flow to alternatives such as solar and wind and other renewables and technologies – the inevitable result, Alliance Bernstein warned early this year – of energy price deflation.
A report from HSBC highlights the dilemma for the oil industry. Many of the new reserves, such as tar sands in Canada and proposed fields in the Arctic, don’t make economic sense – and that is before the cost of refining and delivery.
“Lower oil prices should be beneficial for climate because the economics of higher cost projects are less compelling, meaning less production,” the HSBC economists write in the analysis. And to prove its theory, oil giant Chevron overnight put its proposed drilling program in the Arctic Sea on hold “indefinitely.”
At these prices, drilling in the Arctic simply makes no sense. Doug Matthews, an analyst and former oil and gas director with the Northwest Territories government, said: “With the price of oil down by 40 per cent since June companies all over the world are starting to pull in their horns.”
And high cost oil fields usually – but not always – equate to high carbon emitting fields. This next graph shows that oil sands is amazingly polluting, and the Arctic reserves would be twice as polluting as the main fields currently in production.
But that still doesn’t tell the whole story. The energy analysts at HSNC also looked at the national accounts of Middle East oil producers. They calculated that the break-even oil prices in the context of what oil price is necessary to fund public spending and leave the government with a zero budget balance is significantly higher than the production cost of oil.
In Saudi Arabia for instance (a country using 1.5m barrels oil a day for desalination to satisfy water needs in 2012), an oil price of $US20/barrel would have balanced the budget in 2003, but in 2015 an oil price of $US90/b is required to cover increased levels of spending.
That explains why Saudi Arabia is so keen to invest in renewables, particularly solar, to satisfy its domestic energy needs. And has earmarked more than $40 billion to do so.
“On balance from a climate perspective, we think that lower oil prices are beneficial because the deteriorating economics for higher cost of extraction projects push back production volumes, thus lowering CO2 which is needed to extend the carbon budget,” the HSBC economists write.
Why oil price crash is good news for climate, and clean energy reneweconomy.com.au
The collapse in global oil prices is good for the climate, and clean energy. Big Oil has put Arctic drilling on hold because it no longer makes economic sense. That will allow extend the world’s ca…

domain: finance


text:
A fight over obamacare that’s so last week with the government shutdown firmly in its second week, and the debt limit projected to be reached next thursday, top house and senate. Robert costa reports and conn carroll endorses the latest nutty conservative plan to end obamacare rather than on insisting on obamacare repeal as a c. The debt ceiling is a limit on how much debt the us government many republicans said they would only raise the ceiling if funding for obamacare were taken. Congress has until oct 17 to raise the debt ceiling absolutely everything you need to know about the debt from a delay of obamacare to curbing. Government shutdown threats in 2013 have people wondering how congress got us here in the first place as previously reported by the inquisitr, nancy pelosi says government shutdown threats. • president puts the case for obamacare in maryland• house speaker issues list of demands for debt ceiling dea• deadline for government shutdown approaches• follow live updates.
That the electorate doesn’t understand the debt ceiling isn’t surprising what’s scary is that some members of congress don’t either. The obama-boehner debt crisis the debt deal was absurd that the $20 trillion mark for the us debt ceiling as of obamacare is falling apart as more and. But the problem is that the expiration of the debt limit is on october 17th (or thereabouts, as the president manipulates it) and both democrats and republican leadership have an incentive. Government shutdown threat and debt limit are two different issues - but both are being used to fight obamacare.
Republicans moving away from anti-obamacare demands in the debate over government funding and the debt ceiling.
The gop is no longer threatening a government shutdown and will to negotiations over the debt ceiling obamacare in exchange for an increased debt. The recently concluded debt ceiling debate was celebrated by the participants as a major breakthrough in putting the united states on the road toward solving the deficit and debt problems.
The peter schiff show (9/24/13) listen live weekdays 10am to noon et on buy my newest book at fri. Obama to sign debt ceiling bill president obama will soon sign a bill extending the debt ceiling by some 13 months, wrapping up a victory over congressional republicans and taking the issue. The debt ceiling is all about obamacare conservatives have fallen on their swords with bad votes to suspend the debt ceiling law and to fund obamacare in the cr.
In the summer of 2011, when republicans refused to raise the debt ceiling unless president obama caved to their extortionist demands. Thanks to obamacare, government debt is worse to limit the size of debt in order to postpone hitting the debt limit obamacare is draining our. Nevermind obamacare worry about the debt ceiling by even suggesting that the united states might default, the full faith, honor, and credit of the country has become an ideological. Confused by the sudden stand-off in washington over obamacare, the budget, the debt ceiling and a bundle of other issues you're not alone. But how much did obamacare, the arra national debt under obama what happens when the debt exceeds the ceiling. Congress has a mountain of things to get done in september as varied as raising the debt ceiling and reauthorizing the federal aviation administration.
For the third straight year, congress plans to push its big budget decisions back to christmas after president obama squandered their first week back from august recess over the possibility. If congress and the white house cannot reach a deal by 17 october, the us will default on its debt what would this mean. The debt burden that americans and their families face how will obamacare add to the national debt our national debt crisis the debt explosion to come. Episode 491: the debt ceiling, obamacare and welfare : planet money on today's show: three ripped-from-the-headlines stories from planet money.


Pinned a year ago, state wrestling tournament returns with different vibe
Coffman junior Seth Shumate won state as freshman at 195 pounds
Shawn Andrews pushes for reversals that put opponents on their backs. No surprise, then, that the Marysville High School wrestling coach put a positive spin move on how COVID-19 has drastically altered this year’s state tournament.
“There is overwhelming optimism because we’re just able to wrestle,” Andrews said this week, taking the glass-half-full approach to the championship meets, which begin on Saturday in three separate locations around central Ohio.
Given the optics of wrestling as a close-contact sport, coaches had doubts whether the tournament would even happen, Andrews said. Nothing was taken for granted, not after COVID caused the last-minute cancellation of the 2020 tournament.
“You look back 365 days to when kids were in vans driving to Columbus when they got the call,” Andrews said. “We had to turn around and tell them, ‘The (tournament) is not happening.’ When that wound still feels so fresh it’s easier to find comfort in the moment, in that we’re wrestling.”
Still, the wrestling will look and feel different. Instead of happening in the traditional Thursday, Friday, Saturday format, the tournament is being condensed into Saturday and Sunday as the three sites allow for a more streamlined schedule.
The matches will play out in front of a smattering of about 200 coaches, friends and family members at three high school gyms — Division I at Hilliard Darby, Division II at Highland, Division III at Marion Harding — instead of at Value City Arena, where attendance typically averages about 50,000 over a three-day event.
Dublin Coffman junior Seth Shumate was among those whose life changed on March 13, 2020, when learning he would be unable to defend the state title he won as a freshman at 195 pounds. Suddenly, Shumate would have no opportunity to join the exclusive four-time club of state champions.
“I thought it would only be postponed, but when it got removed everything set in and a lot of problems came with it,” Shumate said. “I had a lot of anger, a little bit of depression. It raised my anxiety.”
Shumate’s outlook eventually brightened, but not before he seriously considered quitting the sport. In his first tournament this season, competing under the new normal of no spectators, the 2019 state champion lost both his match and his lunch.
“I’m out there and it’s out of my normal zone and I look up at the lights and I end up throwing up three times mid-match,” he said. “I grew out of love with the sport at that point and thought, ‘I hate this season.’ ”
Long talks with his coaches rekindled Shumate’s enthusiasm, and he enters Saturday more “ultra-excited” than stressed out by the pressure to defend a title won two years ago. At 26-1 and having won the district last week at Darby, Shumate is favored to collect a second state title. But through COVID his perspective has shifted.
“Now I’m just wrestling, just happy to be back on the mat,” he said, echoing the comments of Andrews and of his own coach, Chance Van Gundy.
“Ask 99.9% of the wrestlers and if it meant that they’d still get their state tournament, they’d wrestle in a barn,” Van Gundy said of the venue change. “They would wrestle in a barn on the side of a country road … just getting that opportunity they didn’t get last year. I don’t care if it’s in Hilliard Darby High School or the Schottenstein Center, we’re still getting a chance to compete.”
Van Gundy and Andrews are spinning the return to Darby as specifically beneficial to their wrestlers.
“We’re looking at it as home-court advantage, since we were at Darby for district last week,” Van Gundy said.
Andrews is doing the same, pointing out that “especially for guys who just wrestled at Darby it will feel a little like ‘Groundhog Day.’ But there is comfort in that, too.”
Ultimately, missing out on wrestling at the Schott is balanced by the chance to wrestle anywhere, Andrews added.
“You can tell with our guys there’s a gratitude there,” he said.
Shumate summed it up: “I’m just happy to be out there, seeing the team grow this season, leading them, watching the seniors. It’s been a long season for sure, but a fun one.”
Where: Division I, Hilliard Darby; Division II, Highland; Division III, Marion Harding
When: Sessions at 10 a.m. and 4:30 p.m., Saturday and Sunday


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text:
- The boys say theyve matured, but they still hang out at the playground, picking fights with emo kids. Or so we hear.
Every Time I Die singer Keith Buckley openly acknowledges that his band felt it had a lot to live up to with its new CD, The Big Dirty.
Much of the pressure came from Gutter Phenomenon, the third full-length release from the Buffalo, N.Y., group. That album earned rave reviews Revolver magazine, in fact, named it Album of the Year for 2005 and established Every Time I Die as one of the most talented groups on the hardcore metal scene.
But while following up Gutter Phenomenon was challenging, Buckley says he came to embrace the pressure of writing and recording The Big Dirty.
"That's all over the lyrics, that stuff," he says. "There are just so many more people expecting certain things, but at the same time that really is necessary. For me, personally, I need that pressure because being comfortable is absolutely no catalyst for being creative."
Much of the pressure, Buckley says, stemmed from wanting to retain a distinctive identity within what has become a crowded genre.
"We're all so inundated with millions and millions of bands, and you have access to everyone's music," Buckley says. "It's really important to kind of stay one step ahead of the pack because there's going to be a slew of new bands that are coming out and just copying other styles and things like that."
So far during its career, Every Time I Die has done an admirable job of carving out a distinctive niche within the scene.
In attempting to stay ahead of other bands and retain its identity, Buckley says the band stepped back toward the more technical playing of its earlier albums while retaining the melodic appeal of Gutter Phenomenon.
"As far as I'm concerned, [The Big Dirty] is more like a Hot Damn!, Part Two, which is really just to say it's got the same sound," Buckley says, referring to the group's 2003 release. "It's very raw, and there's a lot more like punk attitude in it, which I think Gutter didn't have.
"It's as if we wrote it like we wrote Hot Damn!, except we're far more mature now and we know what we like and we tweaked everything up a bit."
While excited about The Big Dirty, Buckley says the band which also includes guitarists Jordan Buckley (the singer's brother) and Andy Williams and drummer Mike Novak will try to resist the urge to load its live set with new songs, because he knows the audience is just getting to know the new CD.
"They want to hear something they're familiar with, so you have to pander to the crowd," Buckley says. "But we're excited to play new stuff and we do make the final call."
Every Time I Die with Underoath, Poison the
Well and Maylene and
The Sons of Disaster
Fillmore Auditorium, 1510 Clarkson St., Denver
Saturday, Oct. 13, 7 p.m.
Tickets: $18, all ages;visit ticketmaster.com

domain: artistic


text:
With Miyamoto celebrating his 63rd birthday on November 16, check out some facts on the man who made Mario jump. Written in an engaging, concise style, this is more than just a book for academics: This is an encyclopaedia - be clear and direct - call a game a game and a title a title.
It got high praise from its fans. Someday, pro gamers around the world will be held in the same respect as a LeBron James or Aaron Rodgers.
There were multiple moments when I came across some new to me piece of information about Miyamoto, thought back to a game of his - in some cases, one I haven't played in years - and felt the cognitive click of recognition.
Casual Games and Mature Audiences Chapter 5: Rool who despises DK and wants nothing more than to see him and his friends miserable. You have stated that you are simply reverting to what the article used to say; regardless of the strength of that arguement it's not strong it's also not true, as noted by Czar.
If you bother to do any research yourself I wrote the article The game had three levels to it separate versions had been made that the player had to complete in order to win.
Games are commonly listed as having multiple genres. The word as you want to use it doesn't even appear in the very dictionary entry you link. They have provided a few sources that say that it is not part of the DKC series 1 is a bit unclear, but is not directly stating that it is in that series.
I hesitate to ask again, but do you have that proof, Some individuals note that the settings are the same, but don't ever really make a distinction between DK64 and the DKC series,  while others, well, I'll just let this quotation speak for itself: A fistful of Quarters.
It's part of how the medium is covered by reliable sources. After attaining Super Saiyan 2 Beserk, she became so powerful that even Ultra Instinct-powered Goku was unable to survive one of her ki blasts. I don't see how uses of "the title" in this article met any of those essay's concerns apart from sharing the same example word "title".
Shigeru Miyamoto stands, the book is a must-read for young game designers and those who may be considering entering the field. For instance, the IOC decided that Chess met its prerequisites, and was officially recognized as a sport in the s Totilo.
I was no the one to start this issue That's not what the edit history says. The people of Kyoto love their city and are somewhat more individualistic. Use the correct word. If the argument is that no source can argue an all-encompassing survey of all professional opinions, then sure, agreed, but then what to do about the generalization?
Mario eventually rescued Pauline and trapped DK in a cage, where he was hauled back to the zoo. The main argument that was established since the "battle" began is the abundance of violence that takes place in the games.
In more recent years the popularity of video games has rapidly increased.
In this region he lives in the company of his many friends, a large group of which is comprised of primates "Kongs" and other animals. A game is a game and a spade is a spade. Miyamoto is famous for letting otherwise mundane experiences inform his gaming design. The only time I raised an eyebrow at him was when he said DKC3 was not as good as the original.
With huge events and tournaments that are watched by millions of passionate fans, many people are beginning to question whether playing video games can be considered a real sport. He eventually gravitated toward industrial design in college, where he again considered the talent pool too deep.
Fetching a lantern, he progressed deeper into the opening, which led to another cave. Date format by country. The game had three levels to it separate versions had been made that the player had to complete in order to win.Donkey Kong related iPad Cases & Skins by independent artists and designers from around the world.
Also available as iphone cases & skins. Games like Donkey Kong 64 general Games like Donkey Kong 64 for iPhone / iPad Games like Donkey Kong 64 for Android Games like Donkey Kong weight of a nation essay 05 12 hold of everyone Andrew here and welcome to my Donkey Kong Country cheats bonus video today we're going to be looking at a.
Donkey Kong Junior is also a title in Nintendo's Game and Watch series of self-contained portable LCD games. Released on October 26, in the Tabletop line (and later rereleased on October 7, in the Panorama line and on August 6, in the Mini Classics line), the game was fairly simple, but required 4 playthroughs to complete, before all 4 keys were obtained and Donkey Kong was freed.
Feb 20, · Back at the old site, someone made a thread about The Geek Critique's video essay on Metroid Prime 2 (which you should also watch, btw).The guy also made videos on the Donkey Kong Country games, chronicling their rise and fall and rise again.
Throughout the film he is forever attempting to reach his goal of breaking the Donkey Kong record only to be shut down by unfair circumstances. Many quotes are taken from Steve and his wife about his reliance, and need of achieving the record for self justification. PhDessay is an educational resource where over 40, free essays are.
In the Mario vs. Donkey Kong series, Donkey Kong and Pauline even become friends. This sets the stage for Super Mario Odyssey, in which Pauline is the mayor of New Donk City—possibly the same.Download


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Boniyar, Dec 28 : Family and relatives of the Uri youth who was murdered in Baramulla last week staged a protest demonstration and demanded justice for their murdered son.
A 25-year-old youth, Azim Nazir Khan, who was missing since December 13 from a gym centre in Azadgunj area of Baramulla was found hanging in a building in the same area after 4 days.
A large number of people including family members and relatives of murdered youth staged protest in Chandanwari area of Uri on Friday.
Shouting slogans against the murder, the protesters demanding justice for their murdered son.
The protesters including females who could be seen beating their chest amidst crying alleged that the deceased was murdered by Baramulla family where he was staying since past 10 years and was running their Gym.
It has been more than 10 days now since his body was found, however the police has not been able to provide justice to us alleged protesting.
Meanwhile the protesters appealed Governor, Satya Pal Malik, Inspector General of Police (IGP) Kashmir and other higher-ups in Governor Administration to look into the issue.
“We want justice, our protest against the cold-blooded murder of our son will continue till all the culprits involved in the crime are brought to justice protesting family added.


text:
Introducing our curated Breakfast Combo, a culinary journey that brings together the best of both worlds. Dive into the crispiness of Gobi Manchurian, where cauliflower florets are infused with rich, tangy flavors. Transition to the comforting warmth of our Veg Stew, a wholesome medley of fresh vegetables in a delectable broth.
For those craving a meaty kick, savor the boldness of Pepper Chicken, where succulent pieces are drenched in a peppery, savory sauce. Alternatively, indulge in the soothing Chicken Stew, a harmonious blend of tender chicken and aromatic spices.
Our Breakfast Combo is more than a meal; it’s an experience. Each bite tells a story of culinary craftsmanship and a dedication to quality. Elevate your mornings with Shaaz Foods — where every combination is a celebration of taste and tradition.

domain: food


Date of Appointment: 29 May 2023
Sarah’s background in journalism and PR spans more than 30 years, working for a wide range of clients in the commercial, public and not for profit sectors. She has served on the boards of several leading London consultancies in a variety of senior roles, including as managing director, as well as over 20 years' experience as a non-executive director of both commercial organisations and charities.
Sarah’s volunteer roles reflect her interest in the arts, education, the environment and community action. In November she stepped down as Chair of Polka Theatre, one of Britain's leading arts venues for children, visited by over 100,000 children a year. While there, she oversaw the successful completion of Polka’s first ever capital project, raising £9m to redevelop the site as a lively community hub for young families. She is also a member of the governing council of Royal Holloway, University of London which has one of the finest collections of 19th century paintings in the country.
Sarah recently stepped down as Chair of Hillgate Public Relations and was previously a trustee of the Careers Development Group (now Shaw Trust) and First Step Trust and was on the Global Fundraising Board of the aid and sustainable development charity, Tearfund. She also mentors young people in the charity and arts sectors.


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- リアルタイム技術により、3次元人工足場内の生きた細胞数を非侵襲的にカウントする。 Real-time technique noninvasively counts the number of live cells in a 3D artificial scaffold.
リアルタイム技術により、3次元人工足場内の生きた細胞数を非侵襲的にカウントする。 Real-time technique noninvasively counts the number of live cells in a 3D artificial scaffold.
Researchers used a noninvasive and label-free method called optical coherence tomography to assess the live cells in the 3D scaffold system. OCT is an imaging technique that uses light waves to take cross sectional images of a sample.
Credit: R. Wilson/NIST
光コヒーレンストモグラフィーを用いた組織工学用足場における3次元のラベルフリー細胞生存率計測 Three-dimensional, label-free cell viability measurements in tissue engineering scaffolds using optical coherence tomography
Greta Babakhanova, Anant Agrawal, Deepika Arora, Allison Horenberg, Jagat B. Budhathoki, Joy P. Dunkers, Joe Chalfoun, Peter Bajcsy, Carl G. Simon Jr
Journal of Biomedical Materials Research Part A Published: 14 March 2023
In the field of tissue engineering, 3D scaffolds and cells are often combined to yield constructs that are used as therapeutics to repair or restore tissue function in patients. Viable cells are often required to achieve the intended mechanism of action for the therapy, where the live cells may build new tissue or may release factors that induce tissue regeneration. Thus, there is a need to reliably measure cell viability in 3D scaffolds as a quality attribute of a tissue-engineered medical product. Here, we developed a noninvasive, label-free, 3D optical coherence tomography (OCT) method to rapidly (2.5 min) image large sample volumes (1 mm3) to assess cell viability and distribution within scaffolds. OCT imaging was assessed using a model scaffold-cell system consisting of a polysaccharide-based hydrogel seeded with human Jurkat cells. Four test systems were used: hydrogel seeded with live cells, hydrogel seeded with heat-shocked or fixed dead cells and hydrogel without any cells. Time series OCT images demonstrated changes in the time-dependent speckle patterns due to refractive index (RI) variations within live cells that were not observed for pure hydrogel samples or hydrogels with dead cells. The changes in speckle patterns were used to generate live-cell contrast by image subtraction. In this way, objects with large changes in RI were binned as live cells. Using this approach, on average, OCT imaging measurements counted 326 ± 52 live cells per 0.288 mm3 for hydrogels that were seeded with 288 live cells (as determined by the acridine orange-propidium iodide cell counting method prior to seeding cells in gels). Considering the substantial uncertainties in fabricating the scaffold-cell constructs, such as the error from pipetting and counting cells, a 13% difference in the live-cell count is reasonable. Additionally, the 3D distribution of live cells was mapped within a hydrogel scaffold to assess the uniformity of their distribution across the volume. Our results demonstrate a real-time, noninvasive method to rapidly assess the spatial distribution of live cells within a 3D scaffold that could be useful for assessing tissue-engineered medical products.

domain: health