Evaluation of elevator shafts as a pathway for fungal spores and particles to enter a hospital housing immuno-compromised patients

Background. It is estimated that hospitals spend between 28 and 33 billion dollars per year as a result of hospital-acquired infections. (Scott, 2009) The costs continue to rise despite the guidance and controls provided by hospital infection control staff to reduce patient exposures to fungal spores and other infectious agents. With all processes and controls in place, the vented elevator shaft represents an unprotected opening from the top of the building to the lower floors. The hypothesis for this prospective study is that there is a positive correlation between the number of Penicillium/Aspergillus-like spores, Cladosporium, ascospores, basidiospores in spores/m3 as individual spore categories found in the hoistway vent of an elevator shaft and the levels of the same spores, sampled near-simultaneously in the outdoor intake of the elevator shaft. Specific aims of this study include determining if external Penicillium/Aspergillus-like spores are entering the healthcare facility via the elevator shaft and hoistway vents. Additional aims include determining levels of Penicillium/Aspergillus-like spores outdoors, in the elevator shafts, and indoors in areas possibly affected by elevator shaft air; and, finally, to evaluate whether any effect is observed due to the installation of a hoistway vent damper, installed serendipitously during this study. ^ Methods. Between April 2010 and September 2010, a total of 3,521 air samples were collected, including 363 spore trap samples analyzed microscopically for seven spore types, and polymerase chain reaction analyses on 254 air samples. 2178 particle count measurements, 363 temperature readings and 363 relative humidity readings were also obtained from 7 different locations potentially related to the path of air travel inside and near a centrally-located and representative elevator shaft. ^ Results. Mean Penicillium/Aspergillus-like spore values were higher outside the building (530 spores/m3 of air) than inside the hoistway (22.8 spores/m3) during the six month study. Mean values inside the hospital were lower than outside throughout the study, ranging from 15 to 73 spores/m3 of air. Mean Penicillium/Aspergillus-like spore counts inside the hoistway decreased from 40.1 spores/m3 of air to 9 spores/m3 of air following the installation of a back draft damper between the outside air and the elevator shaft. Comparison of samples collected outside the building and inside the hoistway vent prior to installing the damper indicated a strong positive correlation (Spearman's Rho=0.8008, p=0.0001). The similar comparison following the damper installation indicated a moderate non-significant inverse correlation (Spearman's rho = −0.2795, p=0.1347). ^ Conclusion. Elevator shafts are one pathway for mold spores to enter a healthcare facility. A significant correlation was detected between spores and particle counts inside the hoistway and outside prior to changes in the ventilation system. The insertion of the back draft damper appeared to lower the spore counts inside the hoistway and inside the building. The mold spore counts in air outside the study building were higher in the period following the damper installation while the levels inside the hoistway and hospital decreased. Cladosporium and Penicillium/Aspergillus -like spores provided a method for evaluating indoor air quality as a natural tracer from outside the building to inside the building. Ascospores and basidiospores were not a valuable tracer due to low levels of detection during this study. ^ Installation of a back draft damper provides additional protection for the indoor environment of a hospital or healthcare facility, including in particular patients who may be immunocompromised. Current design standards and references do not require the installation of a back draft damper, but evaluation of adding language to relevant building codes should be considered. The data indicate a reduction in levels of Penicillium/Aspergillus -like spores, particle counts and a reduction in relative humidity inside of the elevator shaft after damper installation.^

Nonlinear dimension reduction in pathway-based genome-wide association analysis

Pathway based genome wide association study evolves from pathway analysis for microarray gene expression and is under rapid development as a complementary for single-SNP based genome wide association study. However, it faces new challenges, such as the summarization of SNP statistics to pathway statistics. The current study applies the ridge regularized Kernel Sliced Inverse Regression (KSIR) to achieve dimension reduction and compared this method to the other two widely used methods, the minimal-p-value (minP) approach of assigning the best test statistics of all SNPs in each pathway as the statistics of the pathway and the principal component analysis (PCA) method of utilizing PCA to calculate the principal components of each pathway. Comparison of the three methods using simulated datasets consisting of 500 cases, 500 controls and100 SNPs demonstrated that KSIR method outperformed the other two methods in terms of causal pathway ranking and the statistical power. PCA method showed similar performance as the minP method. KSIR method also showed a better performance over the other two methods in analyzing a real dataset, the WTCCC Ulcerative Colitis dataset consisting of 1762 cases, 3773 controls as the discovery cohort and 591 cases, 1639 controls as the replication cohort. Several immune and non-immune pathways relevant to ulcerative colitis were identified by these methods. Results from the current study provided a reference for further methodology development and identified novel pathways that may be of importance to the development of ulcerative colitis.^

Identification and characterization of cellular genes differentially expressed in a subset of tumors with non -functional WT1

Wilms tumor (WT) or nephroblastoma is a genetically heterogeneous pediatric renal tumor that accounts for 6–7% of all childhood cancers in the U.S. WT1, located at 11p13, is the sole WT gene cloned to date. Additional genomic regions containing genes that play a role in the development of Wilms tumor include 11p15, 7p, 16q, 1p, 17q and 19q. This heterogeneity has made it extremely difficult to develop an understanding of the pathways involved in the development of WT, even in the 5–20% of tumors that show mutations at the WT1 locus. My research addresses this gap in our current comprehension of the development of WT. ^ I have used two complementary approaches to extend the current understanding of molecular changes involved in the development of WT. In order to minimize complexities due to genetic heterogeneity, I confined my analysis to the WT1 pathway by assessing those genetically defined tumors that carry WT1 mutations. WT1 encodes a zinc finger transcription factor, and in vitro studies have identified many genes that are potentially regulated in vivo by WT1. However, there is very little in vivo data that suggests that they are transcriptionally regulated endogenously by WT1. In one approach I assessed the role of WT1 in the in vivo regulation of PDGFA and IGF2, two genes that are strong contenders for endogenous regulation by WT1. Using primary tissue samples, I found no correlation between the level of RNA expression of WT1 with either PDGFA or IGF2, suggesting that WT1 does not play a critical role in their expression in either normal kidney or WT. ^ In a parallel strategy, using differential display analysis I compared global gene expression in a subset of tumors with known homozygous inactivating WT1 mutations (WT1-tumors) to the gene expression in a panel of appropriate control tissues (fetal kidney, normal kidney, rhabdoid tumor and pediatric renal cell carcinoma). Transcripts that are aberrantly expressed in this subset of Wilms tumors are candidates for endogenous transcriptional regulation by WT1 as well as for potentially functioning in the development of WT. By this approach I identified several differentially expressed transcripts. I further characterized two of these transcripts, identifying a candidate WT gene in the process. I then performed a detailed analysis of this WT candidate gene, which maps to 7p. Future studies will shed more light on the role of these differentially expressed genes in WT. ^

Role of janus tyrosine kinase -3 (JAK3) and signal transducer and activator of transcription (STAT5) pathway in T lymphocyte activation

T cell activation and expansion is essential for immune response against foreign antigens. However, uncontrolled T cell activity can be manifested as a number of lymphoid derived diseases such as autoimmunity, graft versus host disease, and lymphoma. The purpose of this research was to test the central hypothesis that the Jak3/Stat5 pathway is critical for T cell function. To accomplish this objective, two novel Jak3 inhibitors, AG490 and PNU156804, were identified and their effects characterized on Jak3/Stat5 activation and T cell growth. Inhibition of Jak3 selectively disrupted primary human T lymphocyte growth in response to Interleukin-2 (IL-2), as well as other γ c cytokine family members including IL-4, IL-7, IL-9, and IL-15. Inhibition of Jak3 ablated IL-2 induced Stat5 but not TNF-α mediated NF-κβ DNA binding. Loss of Jak3 activity did not affect T cell receptor mediated signals including activation of p56Lck and Zap70, or IL-2 receptor a chain expression. To examine the effects of Jak3/Stat5 inhibition within a mature immune system, we employed a rat heart allograft model of Lewis (RT1 1) to ACI (RT1a). Heart allograft survival was significantly prolonged following Jak3/Stat5 inhibition when rats were treated with AG490 (20mg/kg) or PNU156804 (80mg/kg) compared to non-treated control animals. This effect was synergistically potentiated when Jak3 inhibitors were used in combination with a signal 1/2 disrupter, cyclosporine, but only additively potentiated with another signal 3 inhibitor, rapamycin. This suggested that sequential inhibition of T cell function is more effective. To specifically address the role of Stat5 in maintaining T cell activity, novel Stat5 antisense oligonucleotides were synthesized and characterized in vitro. Primary human T cells and T-cell tumor lines treated with Stat5 antisense oligonucleotide (7.5 μM) rapidly underwent apoptosis, while no changes in cell cycle were observed as measured by FACS analysis utilizing Annexin-V-Fluorescein and Propidium iodide staining. Evidence is provided to suggest that caspase 8 and 9 pathways mediate this event. Thus, Stat5 may act rather as a negative regulator of apoptotic signals and not as a positive regulator of cell cycle as previously proposed. We conclude that the Jak3/Stat5 pathway is critical for γc cytokine mediated gene expression necessary for T cell expansion and normal immune function and represents an therapeutically relevant effector pathway to combat T cell derived disease. ^