• Hepatic gene expression profiling reveals key pathways involved in leptin-mediated weight loss in ob/ob mice.

      Sharma, Ashok; Bartell, Shoshana M; Baile, Clifton A; Chen, Bo; Podolsky, Robert H.; McIndoe, Richard A; She, Jin-Xiong; Center for Biotechnology and Genomic Medicine; Department of Medicine; Department of Pathology (2010-09-02)
      BACKGROUND: Leptin, a cytokine-like protein, plays an important role in the regulation of body weight through inhibition of food intake and stimulation of energy expenditure. Leptin circulates in blood and acts on the brain, which sends downstream signals to regulate body weight. Leptin therapy has been successful in treating leptin deficient obese patients. However, high levels of leptin have been observed in more common forms of obesity indicating a state of leptin resistance which limits the application of leptin in the treatment of obesity. If the central effect of leptin could be by-passed and genes which respond to leptin treatment could be regulated directly, new therapeutic targets for the treatment of obesity may be possible. The purpose of this study was to identify genes and subsequent pathways correlated with leptin-mediated weight loss. METHODOLOGY/PRINCIPAL FINDINGS: WE UTILIZED MICROARRAY TECHNOLOGY TO COMPARE HEPATIC GENE EXPRESSION CHANGES AFTER TWO TYPES OF LEPTIN ADMINISTRATION: one involving a direct stimulatory effect when administered peripherally (subcutaneous: SQ) and another that is indirect, involving a hypothalamic relay that suppresses food intake when leptin is administered centrally (intracerebroventricular: ICV). We identified 214 genes that correlate with leptin mediated weight loss. Several biological processes such as mitochondrial metabolic pathways, lipid metabolic and catabolic processes, lipid biosynthetic processes, carboxylic acid metabolic processes, iron ion binding and glutathione S-transferases were downregulated after leptin administration. In contrast, genes involved in the immune system inflammatory response and lysosomal activity were found to be upregulated. Among the cellular compartments mitochondrion (32 genes), endoplasmic reticulum (22 genes) and vacuole (8 genes) were significantly over represented. CONCLUSIONS/SIGNIFICANCE: In this study we have identified key molecular pathways and downstream genes which respond to leptin treatment and are involved in leptin-mediated weight loss. Many of these genes have previously been shown to be associated with obesity; however, we have also identified a number of other novel target genes. Further investigation will be required to assess the possible use of these genes and their associated protein products as therapeutic targets for the treatment of obesity.
    • The IL-10 and IFN-gamma pathways are essential to the potent immunosuppressive activity of cultured CD8+ NKT-like cells.

      Zhou, Li; Wang, Hongjie; Zhong, Xing; Jin, Yulan; Mi, Qing-Sheng; Sharma, Ashok; McIndoe, Richard A; Garge, Nikhil; Podolsky, Robert H.; She, Jin-Xiong; et al. (2008-09-05)
      BACKGROUND: CD8+ NKT-like cells are naturally occurring but rare T cells that express both T cell and natural killer cell markers. These cells may play key roles in establishing tolerance to self-antigens; however, their mechanism of action and molecular profiles are poorly characterized due to their low frequencies. We developed an efficient in vitro protocol to produce CD8+ T cells that express natural killer cell markers (CD8+ NKT-like cells) and extensively characterized their functional and molecular phenotypes using a variety of techniques. RESULTS: Large numbers of CD8+ NKT-like cells were obtained through culture of na??ve CD8+ T cells using anti-CD3/anti-CD28-coated beads and high dose IL-2. These cells possess potent activity in suppressing the proliferation of na??ve responder T cells. Gene expression profiling suggests that the cultured CD8+ NKT-like cells and the na??ve CD8+ T cells differ by more than 2-fold for about 3,000 genes, among which 314 are upregulated by more than 5-fold and 113 are upregulated by more than 10-fold in the CD8+ NKT-like cells. A large proportion of the highly upregulated genes are soluble factors or surface markers that have previously been implicated in immune suppression or are likely to possess immunosuppressive properties. Many of these genes are regulated by two key cytokines, IL-10 and IFN-gamma. The immunosuppressive activities of cells cultured from IL-10-/- and IFN-gamma-/- mice are reduced by about 70% and about 50%, respectively, compared to wild-type mice. CONCLUSION: Immunosuppressive CD8+ NKT-like cells can be efficiently produced and their immunosuppressive activity is related to many surface and soluble molecules regulated by IL-10 and IFN-gamma.
    • Influence of common variants in FTO and near INSIG2 and MC4R on growth curves for adiposity in Africanâ and Europeanâ American youth

      Liu, Gaifen; Zhu, Haidong; Dong, Yanbin; Podolsky, Robert H.; Treiber, Frank A.; Snieder, Harold; Center for Biotechnology and Genomic Medicine (2011-06-5)
      Electronic supplementary material: The online version of this article (doi:10.1007/s10654-011-9583-4) contains supplementary material, which is available to authorized users.
    • Intratumoral Convergence of the TCR Repertoires of Effector and Foxp3+ CD4+ T cells

      Kuczma, Michal; Kopij, Magdalena; Pawlikowska, Iwona; Wang, Cong-Yi; Rempala, Grzegorz A.; Kraj, Piotr; Center for Biotechnology and Genomic Medicine; Department of Biostatistics and Epidemiology; GHSU Cancer Center (2010-10-26)
      The presence of Foxp3+ regulatory CD4+ T cells in tumor lesions is considered one of the major causes of ineffective immune response in cancer. It is not clear whether intratumoral Treg cells represent Treg cells pre-existing in healthy mice, or arise from tumor-specific effector CD4+ T cells and thus representing adaptive Treg cells. The generation of Treg population in tumors could be further complicated by recent evidence showing that both in humans and mice the peripheral population of Treg cells is heterogenous and consists of subsets which may differentially respond to tumor-derived antigens. We have studied Treg cells in cancer in experimental mice that express naturally selected, polyclonal repertoire of CD4+ T cells and which preserve the heterogeneity of the Treg population. The majority of Treg cells present in healthy mice maintained a stable suppressor phenotype, expressed high level of Foxp3 and an exclusive set of TCRs not used by naive CD4+ T cells. A small Treg subset, utilized TCRs shared with effector T cells and expressed a lower level of Foxp3. We show that response to tumor-derived antigens induced efficient clonal recruitment and expansion of antigen-specific effector and Treg cells. However, the population of Treg cells in tumors was dominated by cells expressing TCRs shared with effector CD4+ T cells. In contrast, Treg cells expressing an exclusive set of TCRs, that dominate in healthy mice, accounted for only a small fraction of all Treg cells in tumor lesions. Our results suggest that the Treg repertoire in tumors is generated by conversion of effector CD4+ T cells or expansion of a minor subset of Treg cells. In conclusion, successful cancer immunotherapy may depend on the ability to block upregulation of Foxp3 in effector CD4+ T cells and/or selectively inhibiting the expansion of a minor Treg subset.
    • Lack of an association of miR-938 SNP in IDDM10 with human type 1 diabetes

      Mi, Xiaofan; He, Hongzhi; Deng, Yangxin; Levin, Abert M; She, Jin-Xiong; Mi, Qing-Sheng; Zhou, Li; Center for Biotechnology and Genomic Medicine (2011-10-20)
      MicroRNAs (miRNAs) are a newly discovered type of small non-protein coding RNA that function in the inhibition of effective mRNA translation, and may serve as susceptibility genes for various disease developments. The SNP rs12416605, located in human type 1 diabetes IDDM10 locus, changes the seeding sequence (UGU[G/A]CCC) of miRNA miR-938 and potentially alters miR-938 targets, including IL-16 and IL-17A. In an attempt to test whether miR-938 may be a susceptibility gene for IDDM10, we assessed the possible association of the miR-938 SNP with T1D in an American Caucasian cohort of 622 patients and 723 healthy controls by TaqMan assay. Our current data do not support the association between the SNP in miR-938 and type 1 diabetes.
    • Lack of correlation between the levels of soluble cytotoxic T-lymphocyte associated antigen-4 (CTLA-4) and the CT-60 genotypes.

      Purohit, Sharad; Podolsky, Robert H.; Collins, Christin; Zheng, Weipeng; Schatz, Desmond; Muir, Andy; Hopkins, Diane; Huang, Yi-Hua; She, Jin-Xiong; Center for Biotechnology and Genomic Medicine (2005-11-24)
      BACKGROUND: Cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) plays a critical role in downregulation of antigen-activated immune response and polymorphisms at the CTLA-4 gene have been shown to be associated with several autoimmune diseases including type-1 diabetes (T1D). The etiological mutation was mapped to the CT60-A/G single nucleotide polymorphism (SNP) that is believed to control the processing and production of soluble CTLA-4 (sCTLA-4). METHODS: We therefore determined sCTLA-4 protein levels in the sera from 82 T1D patients and 19 autoantibody positive (AbP) subjects and 117 autoantibody negative (AbN) controls using ELISA. The CT-60 SNP was genotyped for these samples by using PCR and restriction enzyme digestion of a 268 bp DNA segment containing the SNP. Genotyping of CT-60 SNP was confirmed by dye terminating sequencing reaction. RESULTS: Higher levels of sCTLA-4 were observed in T1D (2.24 ng/ml) and AbP (mean = 2.17 ng/ml) subjects compared to AbN controls (mean = 1.69 ng/ml) with the differences between these subjects becoming significant with age (p = 0.02). However, we found no correlation between sCTLA-4 levels and the CTLA-4 CT-60 SNP genotypes. CONCLUSION: Consistent with the higher serum sCTLA-4 levels observed in other autoimmune diseases, our results suggest that sCTLA-4 may be a risk factor for T1D. However, our results do not support the conclusion that the CT-60 SNP controls the expression of sCTLA-4.
    • Loss of Jak2 selectively suppresses DC-mediated innate immune response and protects mice from lethal dose of LPS-induced septic shock.

      Zhong, Jixin; Yang, Ping; Muta, Kenjiro; Dong, Robert; Marrero, Mario; Gong, Feili; Wang, Cong-Yi; Center for Biotechnology and Genomic Medicine; Vascular Biology Center (2010-03-16)
      Given the importance of Jak2 in cell signaling, a critical role for Jak2 in immune cells especially dendritic cells (DCs) has long been proposed. The exact function for Jak2 in DCs, however, remained poorly understood as Jak2 deficiency leads to embryonic lethality. Here we established Jak2 deficiency in adult Cre(+/+)Jak2(fl/fl) mice by tamoxifen induction. Loss of Jak2 significantly impaired DC development as manifested by reduced BMDC yield, smaller spleen size and reduced percentage of DCs in total splenocytes. Jak2 was also crucial for the capacity of DCs to mediate innate immune response. Jak2(-/-) DCs were less potent in response to inflammatory stimuli and showed reduced capacity to secrete proinflammatory cytokines such as TNFalpha and IL-12. As a result, Jak2(-/-) mice were defective for the early clearance of Listeria after infection. However, their potency to mediate adaptive immune response was not affected. Unlike DCs, Jak2(-/-) macrophages showed similar capacity secretion of proinflammatory cytokines, suggesting that Jak2 selectively modulates innate immune response in a DC-dependent manner. Consistent with these results, Jak2(-/-) mice were remarkably resistant to lethal dose of LPS-induced septic shock, a deadly sepsis characterized by the excessive innate immune response, and adoptive transfer of normal DCs restored their susceptibility to LPS-induced septic shock. Mechanistic studies revealed that Jak2/SATA5 signaling is pivotal for DC development and maturation, while the capacity for DCs secretion of proinflammatory cytokines is regulated by both Jak2/STAT5 and Jak2/STAT6 signaling.
    • Molecular cloning and characterization of the mouse Acdp gene family.

      Wang, Cong-Yi; Yang, Ping; Shi, Jing-Da; Purohit, Sharad; Guo, Dehuang; An, Haiqian; Gu, Jian-Guo; Ling, Jennifer X; Dong, Zheng; She, Jin-Xiong; et al. (2004-05-11)
      BACKGROUND: We have recently cloned and characterized a novel gene family named ancient conserved domain protein (ACDP) in humans. To facilitate the functional study of this novel gene family, we have cloned and characterized Acdp, the mouse homologue of the human ACDP gene family. RESULTS: The four Acdp genes (Acdp1, Acdp2, Acdp3 and Acdp4) contain 3,631 bp, 3,244 bp, 2,684 bp and 2,743 bp of cDNA sequences, and encode deduced proteins of 951, 874, 713 and 771 amino acids, respectively. The mouse Acdp genes showed very strong homologies (>90%) in both nucleotide and amino acid sequences to their human counterparts. In addition, both nucleotide and amino acid sequences within the Ancient Conserved Domain (ACD) are highly conserved in many different taxonomic species. Particularly, Acdp proteins showed very strong AA homologies to the bacteria CorC protein (35% AA identity with 55% homology), which is involved in magnesium and cobalt efflux. The Acdp genes are widely expressed in all tissues tested except for Acdp1, which is only highly expressed in the brain with low levels of expression in kidney and testis. Immunostaining of Acdp1 in hippocampus neurons revealed a predominant localization on the plasma membrane. CONCLUSION: The Acdp genes are evolutionarily conserved in diverse species and ubiquitously expressed throughout development and adult tissues suggesting that Acdp may be an essential gene. Acdp showed strong homology to bacteria CorC protein and predominantly localized on the plasma membrane. These results suggest that Acdp is probably a family of proteins involved in ion transport in mammalian cells
    • ParaKMeans: Implementation of a parallelized K-means algorithm suitable for general laboratory use.

      Kraj, Piotr; Sharma, Ashok; Garge, Nikhil; Podolsky, Robert H.; McIndoe, Richard A; Center for Biotechnology and Genomic Medicine (2008-04-28)
      BACKGROUND: During the last decade, the use of microarrays to assess the transcriptome of many biological systems has generated an enormous amount of data. A common technique used to organize and analyze microarray data is to perform cluster analysis. While many clustering algorithms have been developed, they all suffer a significant decrease in computational performance as the size of the dataset being analyzed becomes very large. For example, clustering 10000 genes from an experiment containing 200 microarrays can be quite time consuming and challenging on a desktop PC. One solution to the scalability problem of clustering algorithms is to distribute or parallelize the algorithm across multiple computers. RESULTS: The software described in this paper is a high performance multithreaded application that implements a parallelized version of the K-means Clustering algorithm. Most parallel processing applications are not accessible to the general public and require specialized software libraries (e.g. MPI) and specialized hardware configurations. The parallel nature of the application comes from the use of a web service to perform the distance calculations and cluster assignments. Here we show our parallel implementation provides significant performance gains over a wide range of datasets using as little as seven nodes. The software was written in C# and was designed in a modular fashion to provide both deployment flexibility as well as flexibility in the user interface. CONCLUSION: ParaKMeans was designed to provide the general scientific community with an easy and manageable client-server application that can be installed on a wide variety of Windows operating systems.
    • Physical interaction and functional coupling between ACDP4 and the intracellular ion chaperone COX11, an implication of the role of ACDP4 in essential metal ion transport and homeostasis.

      Guo, Dehuang; Ling, Jennifer X; Wang, Mong-Heng; She, Jin-Xiong; Gu, Jianguo; Wang, Cong-Yi; Center for Biotechnology and Genomic Medicine; Department of Physiology (2008-01-16)
      Divalent metal ions such as copper, manganese, and cobalt are essential for cell development, differentiation, function and survival. These essential metal ions are delivered into intracellular domains as cofactors for enzymes involved in neuropeptide and neurotransmitter synthesis, superoxide metabolism, and other biological functions in a target specific fashion. Altering the homeostasis of these essential metal ions is known to connect to a number of human diseases including Alzheimer disease, amyotrophic lateral sclerosis, and pain. It remains unclear how these essential metal ions are delivered to intracellular targets in mammalian cells. Here we report that rat spinal cord dorsal horn neurons express ACDP4, a member of Ancient Conserved Domain Protein family. By screening a pretransformed human fetal brain cDNA library in a yeast two-hybrid system, we have identified that ACDP4 specifically interacts with COX11, an intracellular metal ion chaperone. Ectopic expression of ACDP4 in HEK293 cells resulted in enhanced toxicity to metal ions including copper, manganese, and cobalt. The metal ion toxicity became more pronounced when ACDP4 and COX11 were co-expressed ectopically in HEK293 cells, suggesting a functional coupling between them. Our results indicate a role of ACDP4 in metal ion homeostasis and toxicity. This is the first report revealing a functional aspect of this ancient conserved domain protein family. We propose that ACDP is a family of transporter protein or chaperone proteins for delivering essential metal ions in different mammalian tissues. The expression of ACDP4 on spinal cord dorsal horn neurons may have implications in sensory neuron functions under physiological and pathological conditions.
    • Reduced Serum Vitamin Dâ Binding Protein Levels Are Associated With Type 1 Diabetes

      Blanton, Dustin; Han, Zhao; Bierschenk, Lindsey; Linga-Reddy, M.V. Prasad; Wang, Hongjie; Clare-Salzler, Michael; Haller, Michael; Schatz, Desmond; Myhr, Courtney; She, Jin-Xiong; et al. (2011-10-16)
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    • The Role of Endoplasmic Reticulum Stress in Autoimmune-Mediated Beta-Cell Destruction in Type 1 Diabetes

      Zhong, Jixin; Rao, Xiaoquan; Xu, Jun-Fa; Yang, Ping; Wang, Cong-Yi; Center for Biotechnology and Genomic Medicine (2012-02-14)
      Unlike type 2 diabetes which is caused by the loss of insulin sensitivity, type 1 diabetes (T1D) is manifested by the absolute deficiency of insulin secretion due to the loss of β mass by autoimmune response against β-cell self-antigens. Although significant advancement has been made in understanding the pathoetiology for type 1 diabetes, the exact mechanisms underlying autoimmune-mediated β-cell destruction, however, are yet to be fully addressed. Accumulated evidence demonstrates that endoplasmic reticulum (ER) stress plays an essential role in autoimmune-mediated β-cell destruction. There is also evidence supporting that ER stress regulates the functionality of immune cells relevant to autoimmune progression during T1D development. In this paper, we intend to address the role of ER stress in autoimmune-mediated β-cell destruction during the course of type 1 diabetes. The potential implication of ER stress in modulating autoimmune response will be also discussed. We will further dissect the possible pathways implicated in the induction of ER stress and summarize the potential mechanisms underlying ER stress for mediation of β-cell destruction. A better understanding of the role for ER stress in T1D pathoetiology would have great potential aimed at developing effective therapeutic approaches for the prevention/intervention of this devastating disorder.
    • A statistical framework for integrating two microarray data sets in differential expression analysis.

      Lai, Yinglei; Eckenrode, Sarah E; She, Jin-Xiong; Center for Biotechnology and Genomic Medicine (2009-02-11)
      BACKGROUND: Different microarray data sets can be collected for studying the same or similar diseases. We expect to achieve a more efficient analysis of differential expression if an efficient statistical method can be developed for integrating different microarray data sets. Although many statistical methods have been proposed for data integration, the genome-wide concordance of different data sets has not been well considered in the analysis. RESULTS: Before considering data integration, it is necessary to evaluate the genome-wide concordance so that misleading results can be avoided. Based on the test results, different subsequent actions are suggested. The evaluation of genome-wide concordance and the data integration can be achieved based on the normal distribution based mixture models. CONCLUSION: The results from our simulation study suggest that misleading results can be generated if the genome-wide concordance issue is not appropriately considered. Our method provides a rigorous parametric solution. The results also show that our method is robust to certain model misspecification and is practically useful for the integrative analysis of differential expression.
    • TRPM8 mechanism of autonomic nerve response to cold in respiratory airway.

      Xing, Hong; Ling, Jennifer X; Chen, Meng; Johnson, Richard D; Tominaga, Makoto; Wang, Cong-Yi; Gu, Jianguo; Center for Biotechnology and Genomic Medicine (2008-06-18)
      Breathing cold air without proper temperature exchange can induce strong respiratory autonomic responses including cough, airway constriction and mucosal secretion, and can exacerbate existing asthma conditions and even directly trigger an asthma attack. Vagal afferent fiber is thought to be involved in the cold-induced respiratory responses through autonomic nerve reflex. However, molecular mechanisms by which vagal afferent fibers are excited by cold remain unknown. Using retrograde labeling, immunostaining, calcium imaging, and electrophysiological recordings, here we show that a subpopulation of airway vagal afferent nerves express TRPM8 receptors and that activation of TRPM8 receptors by cold excites these airway autonomic nerves. Thus activation of TRPM8 receptors may provoke autonomic nerve reflex to increase airway resistance. This putative autonomic response may be associated with cold-induced exacerbation of asthma and other pulmonary disorders, making TRPM8 receptors a possible target for prevention of cold-associated respiratory disorders.