Browsing by Subject "2B4"
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Item Expression and Function of Immune Receptors in Pediatric Acute Lymphoblastic Leukemia(2016-12-01) Powers, Sheila B.; Stephen O. Mathew; Porunelloor A. Mathew; W. Paul BowmanAcute lymphoblastic leukemia (ALL) is a cancer that mainly affects children around the age of five. Due to current treatments, 80-90% of children achieve long-term remission. However, because of the current treatments, which include chemotherapy and CNS-directed radiation, these children may experience late effects which impact growth and development. Even though survival rate is high, quality of life can be greatly reduced for some of the survivors. Natural Killer (NK) cells are cells of the innate immune system that are important in fighting cancer and virally-infected cells. They have been a subject of interest in ALL because ALL of the B cell lineage is particularly resistant to NK cell killing. NK cells get activated by their surface receptors and their ligands on target cells. In B cell ALL, the NK cells do not appear to get the proper activating signals and this has been determined to be one reason this cancer is able to thrive. Our lab has cloned three immune receptors, 2B4 (CD244), CS1 (CRACC, CD319) and LLT1 (CLEC2D), that are expressed on NK cells as well as other immune cells. These receptors have been shown to play an important role in NK cell activation. Two other receptors, NKp30 and NKp46, are well-known activating receptors that have also been implicated in ALL. In this project I compared the mRNA and surface protein receptor expression of immune receptors between healthy subjects and ALL subjects. Altered expression of immune receptors was observed in ALL subjects. In particular, a significant decrease in mRNA expression of 2B4, LLT1 and NKp30 was observed in ALL subjects at diagnosis compared to healthy subjects. mRNA expression of CS1 was increased significantly after chemotherapy treatment. In contrast, NKp46 mRNA expression was significantly increased in ALL subjects as compared to healthy subjects. Cell surface protein expression of CS1 was significantly upregulated on T cells and monocytes and LLT1 on NK cells of ALL subjects at diagnosis. Interestingly, NKp30 was overexpressed on B cells, T cells, monocytes and NKp46 was overexpressed on T cells and monocytes but not on NK cells of ALL subjects at diagnosis. I also detected a significant increase of soluble CS1and BAT-3 in ALL subjects at diagnosis between the ages of 1-11 yrs. Also, soluble CD48 was significantly increased in ALL subjects after chemotherapy treatments. Future mechanistic studies may shed more light in the immune dysfunction in ALL ultimately contributing to better treatment options for patients with pediatric ALL.Item Function and Regulation of the Natural Killer Cell Receptor 2B4 (CD244)(2005-05-01) Vaidya, Swapnil V.; Porunelloor A. Mathew; Richard Easom; Hriday DasThe purpose of these studies was to investigate two issues related to the natural killer (NK) cell receptor, 2B4 (CD244) – its in vivo function and transcriptional regulation. In previous in vitro studies, ligation of 2B4 with a monoclonal antibody enhanced the cytotoxicity of NK and CD8 T cells against various tumor cell lines, indicating that 2B4 is an activating receptor. To study the role of 2B4 in vivo, 2B4 deficient (2B4-/-) mice were used. The initial characterization of the 2B4-/- mice indicate a thymic developmental defect with an increase in the immature CD4-/CD8- population in the thyme of female but not male mice. NK cells from the 2B4-/- mice were impaired in activation by IL-2 as compared to wild type NK cells. These results suggest a role of 2B4 in lymphoid development. The in vivo role of 2B4 in tumor rejection was studied in a mouse tumor model in which melanoma cells were injected intravenously and pulmonary metastases enumerated 14 days later. The murine melanoma cell line, B16, was stably transfected with CD48, the counter-receptor for 2B4. Using CD48+ and CD48- B16 cells in tumor experiments indicated that 2B4 functioned as an inhibitory receptor. In addition, a gender-specific role of 2B4 in the rejection of B16 melanoma cells was discovered. 2B4-/- male mice cleared B16 cells more efficiently than wild type male mice, while female 2B4-/- mice were impaired in controlling tumor growth as compared to wild type female mice. In vitro and in vivo studies indicate a complex role for NK cells in the mechanism of this gender effect. Several studies have shown that the expression of 2B4 is upregulated during viral infections and under certain cytokine stimulation. Previously, it has been shown activator protein-1 (AP-1) plays an important role in the transcription of the 2B4 gene. In this study an Ets transcription factor was shown to upregulate the transcription of the gene. This element functions in an AP-1 dependent manner. Stimulation of surface 2B4 down-regulates its own expression by decreasing the activity of the Ets element in the 2B4 promoter. These studies identify a role of 2B4 in lymphoid development and tumor rejection in vivo. The gender-specific defect in 2B4 knock-out mice implicates its role in lupus. The transcriptional studies provide insights into the regulation of 2B4 gene.Item Molecular Basis for 2B4-CD48 Interactions(2001-08-01) Huynh, Van T.; Mathew, Porunelloor A.; Goldfarb, Ronald; Das, HridayHuynh, Van T., Molecular Basis for 2B4-CD48 Interactions. Master of Science, Molecular Biology and Immunology, August 2001, 93 pp., 3 tables, 19 illustrations, bibliography, 51 titles. Natural killer cells are lymphocytes that play a role against cancer and viral infections. 2B4 is a membrane glycoprotein expressed on natural killer cells. In the present study we characterized 2B4 from mice strains BALB/c, 129/svj and A.CA. Nucleotide and peptide analysis revealed that polymorphyic residues in 2B4 are located in the variable domain. My second project was to determine the amino acids involved in the binding between 2B4 and CD48. Twelve mutations were made in human 2B4 to disrupt their interaction. In the last part of the study, an attempt has been made to elucidate the role of tyrosine and threonine amino acids found in the novel tyrosine motifs (TxYxxI/V) that reside in the cytoplasmic domain.Item Molecular Regulation of Interferon Gamma in 2B4-Activated Natural Killer Cells: Functional Role in Tumor Rejection(2001-11-01) Johnson, Lori Ann; Mathew, Porunellor A.; Goldfarb, Ronald H.; Dimitrijevich, S. DanNatural killer cells are a third population of lymphocytes, distinct from T and B cells. NK cells are non-MHC-restricted cytotoxic effector cells which are effective against intracellular pathogens, virally-infected cells and tumor cells. 2B4 is a natural killer cell receptor originally identified in the mouse as a surface molecule involved in non-MHC-restricted killing and enhancement of IFN-γ secretion. The human and rat homologues of 2B4 have recently been cloned in our laboratory. Interferon gamma (IFN-γ) is a cytokine with potent anti-viral and anti-proliferative effects. In addition, this cytokine acts as a global immune regulator by regulating gene expression and serving to attract other immune cells. In this work, we establish the function of human 2B4 in a NK cell line, YT. We have shown that human 2B4 activation induces cytolytic function and enhances IFN-γ release in YT cells. Additionally we show that 2B4’s regulation of IFN-γ occurs at the transcriptional level, both through mRNA stability and increased promoter activity. We also demonstrate that several regions in the IFN-γ promoter respond to 2B4 activation and IFN-γ both separately and together in the rejection of metastatictumor cells in C57B7/6 mice. Our results confirm that both 2B4 and IFN-γ are critical in the rejection of metastatic tumor cells. Through the use of activating monoclonal antibodies, our studies indicate that 2B4’s anti-tumor activity is through IFN-γ as well as through cytolytic function of NK cells.Item The Functional Role of Human 2B4 (CD244) Isoforms in Natural Killer Cells(2007-05-01) Rao, Krithi K.; Porunelloor Mathew; Rance Berg; Harlan JonesRao, Krithi K., Functional role of human 2B4 (CD244) isoforms in natural killer cells. Master of Science (Immunology), July, 2007, 66 pp., 15 illustrations, bibliography. Natural killer (NK) cells are a subpopulation of lymphoctyes that play an important role against tumor metastasis and various viral and bacterial infections. NK cell functions are controlled by a balance between positive and negative signals through various receptors. We have identified, cloned, and characterized the 2B4 (CD244) receptor in mice and human. 2B4 is involved in killing cancer cells and virus-infected cells by NK cells. 2B4 is involved in killing cancer cells and virus-infected cells by NK cells. 2B4 is a counter-receptor for CD48 and recent findings show that 2B4-CD48 interactions plan an important role in NK, T and B cell functions. In humans, two isoforms of 2B4, h2B4-A and h2B4-B, are expressed that differ in the extracellular domain. In the present investigation, we have studied the functions of h2B4-A and h2B4-B. Our data demonstrate that these two isoforms differ in their binding affinity for CD48, resulting in differential cytolytic function as well as cytokine production by NK cells. Thus, differential expression of 2B4 isoforms by NK cells may regulate immune responses mediated through 2B4-CD48 interactions.