Effects of purine nucleoside phosphorylase deficiency on thymocyte development

• Published in: Journal of allergy and clinical immunology Vol. 128; no. 4; pp. 854 - 863.e1
• Main Authors: Papinazath, Taniya, MSc, Min, Wexian, MSc, Sujiththa, Suntharalingam, BSc, Cohen, Amos, PhD, Ackerley, Cameron, PhD, Roifman, Chaim M., MD, Grunebaum, Eyal, MD
• Format: Journal Article
• Language: English
• Published: New York, NY Mosby, Inc 01.10.2011; Elsevier; Elsevier Limited
• Subjects: Allergy and Immunology; Animals; Apoptosis; Apoptosis - immunology; Biological and medical sciences; Cell Line; Cell Proliferation; Cell Survival; Deoxyguanosine - immunology; DNA Fragmentation; Enzymes; fas Receptor - immunology; fas Receptor - metabolism; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Hematopoietic Stem Cells - enzymology; Hematopoietic Stem Cells - immunology; immune deficiency; Immunodeficiencies; Immunodeficiencies. Immunoglobulinopathies; Immunopathology; Kinases; Lymphocytes; Medical sciences; Membrane Potential, Mitochondrial - genetics; Membrane Potential, Mitochondrial - immunology; Mice; Mice, Knockout; Microscopy; Mitochondria; proliferation; Proteins; Purine nucleoside phosphorylase; Purine-Nucleoside Phosphorylase - genetics; Purine-Nucleoside Phosphorylase - immunology; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis; T-Lymphocytes - enzymology; T-Lymphocytes - immunology; thymocytes; Thymus Gland - enzymology; Thymus Gland - immunology; immune deficiency; Purine nucleoside phosphorylase; OP9 bone marrow stromal cells expressing the delta-like ligand 1; TCR; Single positive; apoptosis; dGK; Propidium iodide; DN; FACS; DP; PNP-KO; mice; T-cell receptor; dGuo; PNP; HSC; OP9-DL1; Deoxyguanosine triphosphate; SP; BrDU; Deoxyguanosine kinase; Double positive; proliferation; 5-Bromo-2-deoxyuridine; PNP deficient; TAT-PNP; dGMP; Purine nucleoside phosphorylase fused to TAT protein transduction domain; Adenosine deaminase; dGTP; Fluorescence-activated cell sorting; MMP; Deoxyguanosine monophosphate; Deoxyguanosine; Hematopoietic stem cell; PI; thymocytes; Double negative; ADA; Mitochondrial membrane potential; Cell proliferation; Immunopathology; Enzyme; Thymocyte; Transferases; Deficiency; Glycosyltransferases; Rodentia; Immune deficiency; Purine-nucleoside phosphorylase; Vertebrata; Mammalia; Immunology; Mouse; Cell death; Animal; Pentosyltransferases; Apoptosis
• ISSN: 0091-6749; 1097-6825
• DOI: 10.1016/j.jaci.2011.07.039

Background Inherited or acquired defects in purine nucleoside phosphorylase (PNP) impair purine metabolism, as well as the survival and function of T lymphocytes. However, the effects of PNP deficiency on thymocyte development are not well known. Objectives We sought to study thymocyte development in PNP-deficient (PNP-KO) mice. Methods Maturation, proliferation, and apoptosis were determined in thymocytes from PNP-KO mice and hematopoietic stem cells from these mice grown ex vivo into thymocyte-like cells. Results Reduced percentages of CD4+ CD8+ double-positive (DP) thymocytes with normal percentages of CD4− CD8+ and CD4+ CD8− single-positive thymocytes were found in the thymi of PNP-KO mice. Similarly, reduced DP-like thymocytes grew ex vivo from hematopoietic stem cells of PNP-KO mice. Thymi of PNP-KO mice contained increased apoptotic DP thymocytes. Increased apoptosis of PNP-deficient DP thymocytes occurred after exposure to deoxyguanosine (dGuo), although not after Fas ligation, and could be prevented by restoring PNP activity within the cells. In DP thymocytes from PNP-KO mice, dGuo caused mitochondrial membrane potential dissipation and induced release of cytochrome c from the mitochondria followed by nuclear DNA fragmentation. Inhibition of the caspase pathway prevented dGuo-induced nuclear DNA fragmentation but not mitochondrial membrane potential dissipation, indicating that PNP deficiency induces apoptosis that is initiated in the mitochondria of DP thymocytes. 5-Bromo-2-deoxyuridine incorporation demonstrated that PNP deficiency does not interfere with DP or single-positive thymocyte proliferation. Conclusions PNP is important for the survival of DP thymocytes. Accumulation of dGuo in cases of PNP deficiency leads to mitochondria-initiated apoptosis of DP thymocytes, which can be prevented by restoring PNP activity in the cells.