IMMUNOHEMATOLOGY: Understanding loss of donor white blood cell immunogenicity after pathogen reduction: mechanisms of action in ultraviolet illumination and riboflavin treatment

BACKGROUND: Donor white blood cells (WBCs) present in transfusion products can lead to immune sequelae such as production of HLA antibodies or graft‐versus‐host disease in susceptible transfusion recipients. Eliminating the immunogenicity of blood products may prove to be of clinical benefit, particularly in patients requiring multiple transfusions in whom allosensitization is common. This study examines a method of pathogen reduction based on ultraviolet light illumination in the presence of riboflavin. In addition to pathogens, WBCs treated with this system are affected and fail to stimulate proliferation of allogeneic peripheral blood mononuclear cells (PBMNCs) in vitro. STUDY DESIGN AND METHODS: This study sought to determine the mechanisms regulating this loss of immunogenicity. Treated cells were examined for surface expression of a number of molecules involved in activation and adhesion, viability, cell‐cell conjugation, and ability to stimulate immune responses in allogeneic PBMNCs. RESULTS: Compared with untreated controls, ultraviolet (UV)‐irradiated antigen‐presenting cells showed slightly reduced surface expression of HLA Class II and costimulatory molecules and had more significant reductions in surface expression of a number of adhesion molecules. Furthermore, treated cells had a severe defect in cell‐cell conjugation. The observed loss of immunogenicity was nearly complete, with UV‐irradiated cells stimulating barely measurable interferon‐γ production and no detectable STAT‐3, STAT‐5, or CD3‐ε phosphorylation in allospecific primed T cells. CONCLUSION: These results suggest that defective cell‐cell adhesion prevents UV‐irradiated cells from inducing T‐cell activation.     

Integrated Functional, Gene Expression and Genomic Analysis for the Identification of Cancer Targets

The majority of new drug approvals for cancer are based on existing therapeutic targets. One approach to the identification of novel targets is to perform high-throughput RNA interference (RNAi) cellular viability screens. We describe a novel approach combining RNAi screening in multiple cell lines with gene expression and genomic profiling to identify novel cancer targets. We performed parallel RNAi screens in multiple cancer cell lines to identify genes that are essential for viability in some cell lines but not others, suggesting that these genes constitute key drivers of cellular survival in specific cancer cells. This approach was verified by the identification of PIK3CA, silencing of which was selectively lethal to the MCF7 cell line, which harbours an activating oncogenic PIK3CA mutation. We combined our functional RNAi approach with gene expression and genomic analysis, allowing the identification of several novel kinases, including WEE1, that are essential for viability only in cell lines that have an elevated level of expression of this kinase. Furthermore, we identified a subset of breast tumours that highly express WEE1 suggesting that WEE1 could be a novel therapeutic target in breast cancer. In conclusion, this strategy represents a novel and effective strategy for the identification of functionally important therapeutic targets in cancer.     

Serum growth factor analysis in dry eye syndrome

Background: To perform a comprehensive serum growth factor analysis in dry eye syndrome patients and to compare this with matched controls. Methods: Six female dry eye syndrome patients and six age‐ and gender‐matched controls were recruited. Whole blood was collected, allowed to clot and then centrifuged. Serum was extracted by using sterile technique. Enzyme‐linked immunosorbent assays were performed to quantify serum growth factor levels. Results: Levels of transforming growth factor‐beta 1 and 2 (TGF‐β1 and β2), nerve growth factor (NGF), insulin‐like growth factor‐1 (IGF‐1), epidermal growth factor (EGF), acidic and basic fibroblast growth factor (FGF), keratinocyte growth factor (KGF), hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), platelet‐derived growth factor‐AA, AB and BB (PDGF‐AA, AB and BB), brain‐derived neurotrophic factor (BDNF), neurotrophin‐3 (NT‐3) and glial cell line‐derived neurotrophic factor (GDNF) were quantified, and statistical analysis was performed by using the Mann–Whitney U‐test with the Bonferroni correction. Conclusions: No significant difference was found between serum growth factor levels in dry eye syndrome patients versus controls. Our study provides comprehensive analysis of serum growth factor levels in autologous serum eye drops produced from ocular surface disease patients. A knowledge of growth factor levels in serum may be important because of the increasing use of autologous serum eye drops in refractory ocular surface diseases and for an understanding of how topical serum may provide benefit.     

WNT5A expression increases during melanoma progression and correlates with outcome

PURPOSE: Wnt ligands play a major role in development and are important in cancer. Expression microarray analysis correlates one member of this family, WNT5A, to a subclass of melanomas with increased motility and invasion. There are no large studies of clinical samples primarily addressing the importance of WNT5A in melanoma progression or outcome. Therefore, this study aimed to assess the protein expression of WNT5A during melanoma progression and its effect on outcome. EXPERIMENTAL DESIGN: Expression of WNT5A was determined in a series of 59 primary melanomas with matched metastases. To provide a benchmark of progression against which to assess WNT5A, expression of p16(ink4a) was analyzed, as this has been previously well documented in melanoma. The effect of WNT5A protein expression on outcome was assessed in 102 melanomas. RESULTS: Cytoplasmic WNT5A showed a trend of increasing expression with melanoma progression (P = 0.013), whereas there was diminishing p16(ink4a) expression (P = 0.006). Nevi showed relatively strong WNT5A expression. Strong cytoplasmic WNT5A was an independent risk factor for reduced metastasis-free and overall survival in multivariate analysis (P = 0.001 and 0.003, respectively). CONCLUSION: Cytoplasmic WNT5A increases with melanoma progression and strong expression is associated with poor outcome.     

Physical activity and nutritional status of adolescents on Guam

The 1999 Safe & Drug Free Schools and Communities Youth Risk Behavior Study collected data from representative samples of both high school (n=590) and middle school students (n=643) on Guam in May, 1999. Results showed that the dietary and exercise habits of Guam's adolescents were sub-optimal. Out of the 643 middle school students surveyed, 26% consumed at least 3 meals per week at a fast food restaurant and 53.3% consumed at least 2 cans of sweetened soda per day. Guam adolescents had a very low intake of fruits and vegetables. As many as 75.3% of high school students reported consuming fruits and vegetables less than once a day. Students who practiced unsafe weight loss practices were significantly more likely to smoke cigarettes, and use illegal drugs. In addition, 26.9% of adolescents surveyed were considered 'overweight'--similar to the prevalence of overweight reported among U.S. Mainland adolescents. Finally, data from survey showed that adolescents on Guam spent more time watching television and less time performing various physical activities than adolescents in the U.S. mainland. These suboptimal practices put Guam's adolescents at increased risk for immediate and long-term health problems.     

Adenomatous polyp recurrence and physical activity in the Polyp Prevention Trial (United States)

Objective: To examine prospectively the association between physical activity and adenomatous polyp recurrence. Methods: Information on past year total physical activity was collected annually through an interview-administered questionnaire from the 1905 men and women enrolled in a randomized dietary intervention study, the Polyp Prevention Trial. Multiple logistic regression analysis was used to examine the association between physical activity and polyp recurrence in up to three years of follow-up from baseline colonoscopy. Results: There were no significant associations between moderate, vigorous, or total physical activity at the start of the trial and overall polyp recurrence in either men or women. Participants who reported consistent vigorous activity throughout the trial period had no significantly reduced risk of polyp recurrence compared to those who reported consistent sedentary activity (OR = 0.8, CI = 0.5–1.1). Consistent vigorous activity was also not significantly associated with either advanced or multiple polyps, nor with polyp recurrence at any specific anatomical location in the large bowel. Conclusions: These prospective data suggest that recent physical activity is not associated with polyp recurrence in a three-year period.