11 research outputs found

    Standardizing Umbilical Cord Mesenchymal Stromal Cells for Translation to Clinical Use: Selection of GMP-Compliant Medium and a Simplified Isolation Method

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    Citation: Smith, J. R., Pfeifer, K., Petry, F., Powell, N., Delzeit, J., & Weiss, M. L. (2016). Standardizing Umbilical Cord Mesenchymal Stromal Cells for Translation to Clinical Use: Selection of GMP-Compliant Medium and a Simplified Isolation Method. Stem Cells International, 14. doi:10.1155/2016/6810980Umbilical cord derived mesenchymal stromal cells (UC-MSCs) are a focus for clinical translation but standardized methods for isolation and expansion are lacking. Previously we published isolation and expansion methods for UC-MSCs which presented challenges when considering good manufacturing practices (GMP) for clinical translation. Here, a new and more standardized method for isolation and expansion of UC-MSCs is described. The new method eliminates dissection of blood vessels and uses a closed-vessel dissociation following enzymatic digestion which reduces contamination risk and manipulation time. The new method produced >10 times more cells per cm of UC than our previous method. When biographical variables were compared, more UC-MSCs per gram were isolated after vaginal birth compared to Caesarian-section births, an unexpected result. UC-MSCs were expanded in medium enriched with 2%, 5%, or 10% pooled human platelet lysate (HPL) eliminating the xenogeneic serum components. When the HPL concentrations were compared, media supplemented with 10% HPL had the highest growth rate, smallest cells, and the most viable cells at passage. UC-MSCs grown in 10% HPL had surface marker expression typical of MSCs, high colony forming efficiency, and could undergo trilineage differentiation. The new protocol standardizes manufacturing of UC-MSCs and enables clinical translation

    Univariate gradient statistic for a marginal cure rate model with high-dimensional covariates

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    Master of ScienceDepartment of StatisticsWei-Wen HsuCure rate models, also known as two-component mixture models, have been well established and widely used in the literature for analyzing the lifetime data of long-term survivors. Owing to the advancement of genomic technology, it is now of interest to identify the significant genes or microarrays that are highly associated with the survival outcome under the cure rate model framework. The identification procedure using these genomic data will involve the technique of variable selection for high-dimensional covariates. However, the cure rate model requires the modeling of the cure fraction and the survival function of the uncured individuals, which inevitably leads to a more complicated variable selection process. In this paper, we propose a gradient-statistic-based variable selection method under a marginal representation of the cure rate model. This marginal model can produce interpretable covariate effects on the overall survival response by relating the marginal mean hazard rate to high-dimensional covariates directly while regarding the cure fraction as a nuisance parameter. A univariate gradient score is then used iteratively to determine significant covariates. Coupled with the use of a False Discovery Rate approach, the top-ranked list of covariates can be easily obtained by controlling the family-wise error rate. The proposed method is evaluated by extensive simulations and illustrated with an application of the TCGA breast cancer dataset which contains more than 400,000 microarrays

    Costs of cold acclimation on survival and reproductive behavior in Drosophila melanogaster.

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    Fitness is determined by the ability of an organism to both survive and reproduce; however, the mechanisms that lead to increased survival may not have the same effect on reproductive success. We used nineteen natural Drosophila melanogaster genotypes from the Drosophila Genetic Reference Panel to determine if adaptive plasticity following short-term acclimation through rapid cold-hardening (RCH) affects mating behavior and mating success. We confirmed that exposure to the acclimation temperature is beneficial to survival following cold stress; however, we found that this same acclimation temperature exposure led to less efficient male courtship and a significant decrease in the likelihood of mating. Cold tolerance and the capacity to respond plastically to cold stress were not correlated with mating behavior following acclimation, suggesting that the genetic control of the physiological effects of the cold temperature exposure likely differ between survival and behavioral responses. We also tested whether the exposure of males to the acclimation temperature influenced courtship song. This exposure again significantly increased courtship duration; however, courtship song was unchanged. These results illustrate costs of short-term acclimation on survival and reproductive components of fitness and demonstrate the pronounced effect that short-term thermal environment shifts can have on reproductive success

    Survival following cold stress with and without the acclimation pre-treatment.

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    <p>A. Flies that were acclimated had higher survival than flies that did not receive the acclimation treatment prior to cold stress. B. Genotype significantly influenced survival following the basal cold tolerance and acclimation treatments. Each point and connecting line represents the change in a genotype‚Äôs average survival between the basal cold tolerance and acclimation treatments. This change (acclimation survival‚ÄĒbasal survival) is RCH capacity, one measure of phenotypic plasticity. Variation in change in survival between the two treatments led to genotype-specific variation in RCH capacity.</p

    Temperature treatments for the cold tolerance and mating assays.

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    <p>Flies were exposed to either (A) a basal cold tolerance treatment for 1 hour at -6¬įC or (B) an acclimation treatment for two hours at 4¬įC to induce RCH followed by -6¬įC for 1 hour to determine the level of cold tolerance for each genotype. C. Flies were exposed to either the acclimation treatment temperature (4¬įC) for 2 hours to induce RCH or were held at 25¬įC for the mating latency and song assays. Shading indicates the timing of lights on for experimental flies used in each assay.</p