Browsing by Subject "CD38"
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Item Harnessing Natural Killer Cells for Improved Therapeutic Potential(2021-08) Kim, HansolCellular immunotherapy provides durable control for infected and transformed cells. However, current Natural Killer (NK) cell therapy products are limited by effector persistence. To this end, three strategies to improve the efficacy of NK cell-based therapies are discussed. Killer immunoglobulin-like receptors (KIR) are developed during maturation of NK cells. It showed that KIR develops primarily between CD56brightCD94high and CD56dimCD94high stages. Major demethylation activities were observed during the development of KIR at its proximal promoter region. Ascorbic acid showed to facilitate the development of KIR in collaboration with Ten-eleven translocation (TET) enzymes. NK cells with adaptive immunological properties persist and expand in response to cytomegalovirus (CMV) infection. The levels of intracellular metabolites were analyzed and showed that adaptive NK cells have relatively higher levels of metabolites associated with glycolysis, purine, and pyrimidine metabolism. This supports the notion that adaptive NK cells have upregulated metabolic profiles, and have capacity to expand upon reactivation of CMV which is similar to the characteristics of memory T cells. To address challenges associated with inconsistencies of the manufactured product, and treatment cost, we developed a triple gene-edited induced pluripotent stem cell (iPSC) platform for broad patient-based adoptive cell therapy. First edit is to introduce non-cleavable CD16 which prevents reduced efficacy by antibody-dependent cellular cytotoxicity (ADCC). Second edit allows iPSC-derived NK cells, termed iNK, to persist without supplementation of exogenous IL-15 by introducing IL-15 receptor fusion. The last edit was to avoid daratumumab-induced fratricide by knocking out CD38 on the surface of the iNK cells. These engineered iNK cells persisted in vivo without supplementation of exogenous cytokine and could be combined with daratumumab for enhanced treatment of multiple myeloma. The gene-edited iNK cells exhibited metabolic features and gene expression profiles similar to those of adaptive NK cells which has broad off-the-shelf potential for the treatment of advanced cancer.Item Identification and characterization of novel ADP-ribosyl cyclase family members(2011-08) Hirte, Renee MarieCD38 and CD157 have been identified as mammalian forms of ADP-ribosyl cyclase (ADPRC). Recently novel membrane bound and cytosolic ADPRCs that are distinct from CD38 and CD157 have been identified in various tissues by our laboratory as well as others. Recently, evidence has indicated the presence of ADPRC in tissues lacking CD38. From this it is clear that there are multiple forms of mammalian ADPRC, many of which have not yet been identified or characterized. The overall goal of the research presented in this thesis was to identify and characterize the novel cytosolic cyclase(s) present in the heart cytosol and determine the mechanism(s) by which cytosolic cyclase(s) are regulated. Previously it has been shown that Aplysia ADPRC, CD38 and CD157 share approximately 30% sequence identity. There are two main features shared by each of these members of the ADPRC family including a conserved region near the center and ten conserved cysteine residues that can be perfectly aligned. We used a molecular approach to identify potential candidates for the cytosolic protein (or other yet unidentified ADPRCs) based on a conserved protein motif search of the mouse genome to identify proteins that shared the feature of the conserved cysteine residues. The existence of novel cyclases has implications for a broad range of cellular processes that are influenced by calcium signaling. Characterization and identification of novel ADPRCs may provide key insight into the function of the novel cyclase(s) as well as interaction and relationships to other members of the ADPRC family, which will further elucidate the complexities of calcium signaling.Item Microrna Regulation On The Expression Of CD38 And Other Asthma Related Genes In Human Airway Smooth Muscle Cells(2015-04) Dileepan, MythiliCD38 is a multifunctional enzyme that regulates intracellular calcium ([Ca++]i ) homeostasis. It is expressed in airway smooth muscle (ASM) cells where it elevates [Ca++]i through its enzymatic product cyclic ADPribose (cADPR) and increases ASM contractility. Increased expression of CD38 in the ASM cells derived from the asthmatic patients (AS-HASM) and attenuated airway hyperresponsiveness to contractile stimuli shown by CD38-/- mice implicate the importance of CD38 in asthma. The proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is considered to be an important mediator for airway pathology in asthma. The reason for the differential expression of TNF-alpha-induced-CD38 in AS-HASM cells, does not involve transcriptional regulation of CD38 which is through signaling pathways mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K), and transcription factors nuclear factor kappa-B (NF-kB) and AP-I. Thus I hypothesized that post-transcriptional regulation of CD38 by microRNAs account for the differential expression of TNF-alpha induced -CD38 in AS-HASM cells. Among the several potential microRNAs predicted for CD38 by microRNA target-predicting algorithms, I selected miR-140-3p and miR-708 for further studies, as these showed differential expression in the AS-HASM cells compared to those from healthy subjects. Overexpression of these either microRNAs in ASM cells inhibited the TNF-alpha induced expression of CD38 at messenger RNA (mRNA) and protein levels. Luciferase-reporter assays with a mutated 3'UTR of the CD38 transcript confirmed the specific target sites for both microRNAs. Transcript stability assays revealed that mRNA degradation is not the mechanism underlying regulation by microRNAs. Examination of the expression and activation levels of proteins in the upstream signaling pathways of CD38 revealed that miR-140-3p, by inactivating p38 MAPK and NF-kB, and miR-708, by inactivating c-Jun N-terminal kinase (JNK) MAPK and Akt by elevating the expression of their phosphatases MKP-1 and PTEN respectively, control the expression of CD38 indirectly. Further, we found that miR-708 downregulates the expression of many chemokines and inhibits the serum induced proliferation of human ASM cells. We conclude that both microRNAs have therapeutic potential in controlling asthma related symptoms through regulating the expression of CD38 and chemokines and controlling the proliferation of human ASM cells.Item Regulation of CD38 expression in human airway smooth muscle (HASM) cells.(2009-05) Jude, Joseph AntonyCD38 is a multifunctional enzyme-cum-receptor expressed in a variety of mammalian tissues including airway smooth muscles (ASM). The ADP-ribosyl cyclase activity of CD38 generates cyclic ADP-ribose (cADPR), a Ca2+ mobilizing agent in ASM cells. In vivo studies in mice showed that CD38 plays an important role in the development of airway hyperresponsiveness (AHR) following exposure to cytokines. In vitro studies demonstrated that a variety of cytokines, including the inflammatory cytokine TNF-alpha, induce CD38 expression in human ASM (HASM) cells. Studies also showed that the TNF-alpha-induced CD38 expression in HASM cells is mediated through both the transcriptional and post-transcriptional mechanisms and involves activation of mitogen-activated protein kinases (MAPKs) and transcription factors NF-kappaB and AP-1. The role of CD38 in the pathogenesis of AHR in human asthmatics is not known. The current studies demonstrate that HASM cells isolated from asthmatic patients show differentially elevated CD38 expression following TNF-alpha exposure compared to non-asthmatic HASM cells. Basal and TNF-alpha-induced activation of extracellular signal- regulated kinase (ERK) and p38 MAPK are elevated in asthmatic HASM cells, whereas the TNF-alpha-induced activation of c-jun N terminal kinase (JNK) is elevated in non-asthmatic HASM cells compared to asthmatic cells. The TNF-alpha-induced NF-kappaB activation is elevated in asthmatic HASM cells, indicating that the differentially elevated CD38 expression in asthmatic HASM cells is mediated through transcriptional mechanisms. The role of cross-talk between ERK and phosphatidylinositol 3-kinase/Akt (PI3K/Akt) pathways in TNF-alpha-induced CD38 expression is investigated in the current studies. Two pharmacological inhibitors of PI3K, LY294002 and wortmannin, show differential effects on the TNF-alpha-induced CD38 expression in HASM cells. Transient expression of PI3K catalytic subunit (p110) elevates CD38 expression while transfection with phosphatase tensin homolog (PTEN) attenuates the TNF-alpha-induced CD38 expression, suggesting that PI3K/Akt pathway mediates CD38 expression in HASM cells in a non-ERK dependant manner. The role of adenylate-uridylate-rich elements (AREs) of the CD38 mRNA 3' untranslated region (3'UTR) in CD38 mRNA stability is currently being investigated. Preliminary findings show that, in HASM cells, the RNA-binding proteins HuR and TIA-1 selectively bind to an ARE of CD38 3'UTR in response to TNF-alpha-exposure, suggesting a potential role for the CD38 mRNA AREs in the post-transcriptional regulation of CD38 expression in HASM cells.