Role of biomechanics in the understanding of normal, injured, and healing ligaments and tendons

Ligaments and tendons are soft connective tissues which serve essential roles for biomechanical function of the musculoskeletal system by stabilizing and guiding the motion of diarthrodial joints. Nevertheless, these tissues are frequently injured due to repetition and overuse as well as quick cutting motions that involve acceleration and deceleration. These injuries often upset this balance between mobility and stability of the joint which causes damage to other soft tissues manifested as pain and other morbidity, such as osteoarthritis

An Engineered Viral Protease Exhibiting Substrate Specificity for a Polyglutamine Stretch Prevents Polyglutamine-Induced Neuronal Cell Death

BACKGROUND: Polyglutamine (polyQ)-induced protein aggregation is the hallmark of a group of neurodegenerative diseases, including Huntington's disease. We hypothesized that a protease that could cleave polyQ stretches would intervene in the initial events leading to pathogenesis in these diseases. To prove this concept, we aimed to generate a protease possessing substrate specificity for polyQ stretches. METHODOLOGY/PRINCIPAL FINDINGS: Hepatitis A virus (HAV) 3C protease (3CP) was subjected to engineering using a yeast-based method known as the Genetic Assay for Site-specific Proteolysis (GASP). Analysis of the substrate specificity revealed that 3CP can cleave substrates containing glutamine at positions P5, P4, P3, P1, P2', or P3', but not substrates containing glutamine at the P2 or P1' positions. To accommodate glutamine at P2 and P1', key residues comprising the active sites of the S2 or S1' pockets were separately randomized and screened. The resulting sets of variants were combined by shuffling and further subjected to two rounds of randomization and screening using a substrate containing glutamines from positions P5 through P3'. One of the selected variants (Var26) reduced the expression level and aggregation of a huntingtin exon1-GFP fusion protein containing a pathogenic polyQ stretch (HttEx1(97Q)-GFP) in the neuroblastoma cell line SH-SY5Y. Var26 also prevented cell death and caspase 3 activation induced by HttEx1(97Q)-GFP. These protective effects of Var26 were proteolytic activity-dependent. CONCLUSIONS/SIGNIFICANCE: These data provide a proof-of-concept that proteolytic cleavage of polyQ stretches could be an effective modality for the treatment of polyQ diseases

Paclitaxel and concomitant radiotherapy in high-risk endometrial cancer patients: preliminary findings

BACKGROUND: There is still much debate about the best adjuvant therapy after surgery for endometrial cancer (EC) and there are no current guidelines. Radiotherapy (RT) alone does not seem to improve overall survival. We investigated whether concomitant Paclitaxel (P) and RT gave better clinical results. METHODS: Twenty-three patients with high-risk EC (stage IIB, IIIA, IIIC or IC G3 without lymphadenectomy or with aneuploid tumor) underwent primary surgery and were then referred for adjuvant therapy. P was given at a dose of 60 mg/m2 once weekly for five weeks during RT, which consisted of a total radiation dose of 50.4 Gy. Three further weekly cycles of P at a dose of 80 mg/m2 were given at the end of RT. Overall survival and disease-free survival were calculated from the time of surgery. Patterns of failure were recorded by the sites of failure. RESULTS: A total of 157 cycles of P were administered both during radiotherapy and consolidation chemotherapy. Relapses occurred in five patients (21.7%). Median time to recurrence was 18.6 months (range 3–28). Survival rate for all the patients was 78.2%. Overall survival for the patients who completed chemo-radiation was of 81%. In this group median time to recurrence was 19.2 months (range 3–28). All recurrences were outside the radiation field. Mortality rate was 14.2%. CONCLUSION: This small series demonstrates pelvic radiotherapy in combination with weakly P followed by three consolidation chemotherapy cycles as an effective combined approach in high risk endometrial carcinoma patients

Cross-platform comparability of microarray technology: Intra-platform consistency and appropriate data analysis procedures are essential

BACKGROUND: The acceptance of microarray technology in regulatory decision-making is being challenged by the existence of various platforms and data analysis methods. A recent report (E. Marshall, Science, 306, 630–631, 2004), by extensively citing the study of Tan et al. (Nucleic Acids Res., 31, 5676–5684, 2003), portrays a disturbingly negative picture of the cross-platform comparability, and, hence, the reliability of microarray technology. RESULTS: We reanalyzed Tan's dataset and found that the intra-platform consistency was low, indicating a problem in experimental procedures from which the dataset was generated. Furthermore, by using three gene selection methods (i.e., p-value ranking, fold-change ranking, and Significance Analysis of Microarrays (SAM)) on the same dataset we found that p-value ranking (the method emphasized by Tan et al.) results in much lower cross-platform concordance compared to fold-change ranking or SAM. Therefore, the low cross-platform concordance reported in Tan's study appears to be mainly due to a combination of low intra-platform consistency and a poor choice of data analysis procedures, instead of inherent technical differences among different platforms, as suggested by Tan et al. and Marshall. CONCLUSION: Our results illustrate the importance of establishing calibrated RNA samples and reference datasets to objectively assess the performance of different microarray platforms and the proficiency of individual laboratories as well as the merits of various data analysis procedures. Thus, we are progressively coordinating the MAQC project, a community-wide effort for microarray quality control

Genetic Variation of the Human α-2-Heremans-Schmid Glycoprotein (AHSG) Gene Associated with the Risk of SARS-CoV Infection

Genetic background may play an important role in the process of SARS-CoV infection and SARS development. We found several proteins that could interact with the nucleocapsid protein of the SARS coronavirus (SARS-CoV). α-2-Heremans-Schmid Glycoprotein (AHSG), which is required for macrophage deactivation by endogenous cations, is associated with inflammatory regulation. Cytochrome P450 Family 3A (CYP4F3A) is an ω-oxidase that inactivates Leukotriene B4 (LTB4) in human neutrophils and the liver. We investigated the association between the polymorphisms of these two inflammation-associated genes and SARS development. The linkage disequilibrium (LD) maps of these two genes were built with Haploview using data on CHB+JPT (version 2) from the HapMap. A total of ten tag SNPs were selected and genotyped. In the Guangzhou cohort study, after adjusting for age and sex, two AHSG SNPs and one CYP4F3 SNP were found to be associated with SARS susceptibility: rs2248690 (adjusted odds ratio [AOR] 2.42; 95% confidence interval [CI] 1.30-4.51); rs4917 (AOR 1.84; 95% CI 1.02-3.34); and rs3794987 (AOR 2.01; 95% CI 1.10–3.68). To further validate the association, the ten tag SNPs were genotyped in the Beijing cohort. After adjusting for age and sex, only rs2248690 (AOR, 1.63; 95% CI, 1.30–2.04) was found to be associated with SARS susceptibility. The combined analysis of the two studies confirmed tag SNP rs2248690 in AHSG as a susceptibility variant (AOR 1.70; 95% CI 1.37–2.09). The statistical analysis of the rs2248690 genotype data among the patients and healthy controls in the HCW cohort, who were all similarly exposed to the SARS virus, also supported the findings. Further, the SNP rs2248690 affected the transcriptional activity of the AHSG promoter and thus regulated the AHSG serum level. Therefore, our study has demonstrated that the AA genotype of rs2268690, which leads to a higher AHSG serum concentration, was significantly associated with protection against SARS development

Aerobic exercise in adolescents with obesity: preliminary evaluation of a modular training program and the modified shuttle test

BACKGROUND: Increasing activity levels in adolescents with obesity requires the development of exercise programs that are both attractive to adolescents and easily reproducible. The aim of this study was to develop a modular aerobic training program for adolescents with severe obesity, with a focus on variety, individual targets and acquiring physical skills. We report here the effects on aerobic fitness from a pilot study. Furthermore, we examined the feasibility of the modified shuttle test (MST) as an outcome parameter for aerobic fitness in adolescents with severe obesity. METHODS: Fifteen adolescents from an inpatient body weight management program participated in the aerobic training study (age 14.7 ± 2.1 yrs, body mass index 37.4 ± 3.5). The subjects trained three days per week for 12 weeks, with each session lasting 30–60 minutes. The modular training program consisted of indoor, outdoor and swimming activities. Feasibility of the MST was studied by assessing construct validity, test-retest reliability and sensitivity to change. RESULTS: Comparing pretraining and end of training period showed large clinically relevant and significant improvements for all aerobic indices: e.g. VO(2 peak )17.5%, effect size (ES) 2.4; W(max )8%, ES 0.8. In addition, a significant improvement was found for the efficiency of the cardiovascular system as assessed by the oxygen pulse (15.8%, ES 1.6). Construct validity, test-retest reliability and sensitivity to change of the MST were very good. MST was significantly correlated with VO(2 peak )(r = 0.79) and W(max )(r = 0.84) but not with anthropometric measures. The MST walking distance improved significantly by 32.5%, ES 2.5. The attendance rate at the exercise sessions was excellent. CONCLUSION: This modular, varied aerobic training program has clinically relevant effects on aerobic performance in adolescents with severe obesity. The added value of our aerobic training program for body weight management programs for adolescents with severe obesity should be studied with a randomized trial. This study further demonstrated that the MST is a reliable, sensitive and easy to administer outcome measure for aerobic fitness in adolescent body weight management trials

Intra- and Inter-clade Cross-reactivity by HIV-1 Gag Specific T-Cells Reveals Exclusive and Commonly Targeted Regions: Implications for Current Vaccine Trials

The genetic diversity of HIV-1 across the globe is a major challenge for developing an HIV vaccine. To facilitate immunogen design, it is important to characterize clusters of commonly targeted T-cell epitopes across different HIV clades. To address this, we examined 39 HIV-1 clade C infected individuals for IFN-γ Gag-specific T-cell responses using five sets of overlapping peptides, two sets matching clade C vaccine candidates derived from strains from South Africa and China, and three peptide sets corresponding to consensus clades A, B, and D sequences. The magnitude and breadth of T-cell responses against the two clade C peptide sets did not differ, however clade C peptides were preferentially recognized compared to the other peptide sets. A total of 84 peptides were recognized, of which 19 were exclusively from clade C, 8 exclusively from clade B, one peptide each from A and D and 17 were commonly recognized by clade A, B, C and D. The entropy of the exclusively recognized peptides was significantly higher than that of commonly recognized peptides (p = 0.0128) and the median peptide processing scores were significantly higher for the peptide variants recognized versus those not recognized (p = 0.0001). Consistent with these results, the predicted Major Histocompatibility Complex Class I IC50 values were significantly lower for the recognized peptide variants compared to those not recognized in the ELISPOT assay (p<0.0001), suggesting that peptide variation between clades, resulting in lack of cross-clade recognition, has been shaped by host immune selection pressure. Overall, our study shows that clade C infected individuals recognize clade C peptides with greater frequency and higher magnitude than other clades, and that a selection of highly conserved epitope regions within Gag are commonly recognized and give rise to cross-clade reactivities