B cell immunoglobulin production is regulated by helper T cells through direct interaction and secreted cytokines. In the present study, we functionally analyzed CD27 in cord and peripheral blood B cells. Adult peripheral blood B cells were separated into CD27+ and CD27- cells, which differed in their morphology. Cord blood B cells did not express CD27, and CD27 expression on peripheral blood B cells increased with age. Only CD27+ B cells had the ability to produce immunoglobulin, which was increased by contact with a tumor necrosis factor-related transmembrane ligand, CD70. Adult peripheral blood CD27+ B cells can be further subdivided into two discrete subtypes: IgD- CD27+ and IgD+ CD27+ B cells. IgD- CD27+ B cells produce IgG, IgM and IgA, whereas IgD+ CD27+ B cells predominantly produce IgM. The addition of activated CD4+ CD45RO T cells expressing CD70 caused down-regulation of CD27 expression on activated B cells, and this down-modulation was completely blocked by anti-CD70 monoclonal antibody, indicating direct T-B cell contact via CD27/CD70. The triggering via CD27 and CD40 additively increased the immunoglobulin production under Staphylococcus aureus Cowan strain plus interleukin-2 stimulation. Taken together, our findings demonstrate that peripheral blood B cells are separated into subpopulations by CD27 and IgD expression and that CD27+ B cells produce large amounts of immunoglobulin by interaction with the CD70 molecule.

We generated transgenic mice carrying the mouse type 2 nitric-oxide synthase (NOS2) cDNA under the control of the insulin promoter. Western and immunohistochemical analyses revealed that NOS2 was expressed abundantly in transgenic islets but not in control islets. When islets were isolated and cultured, high levels of nitrite were released from the transgenic islets. In transgenic mice, the ␤ cell mass was markedly reduced without the infiltration of macrophages or lymphocytes, and extensive DNA strand breaks were detected in the islets by in situ nick translation. All the transgenic mice developed hypoinsulinemic diabetes by 4 weeks of age, and treatment with an inhibitor of NOS2, aminoguanidine (200 mg/kg body weight every 12 h), prevented or delayed the development of diabetes. The present study shows that the production of nitric oxide by ␤ cell NOS2 plays an essential role in the ␤ cell degeneration. Insulin-dependent diabetes mellitus (IDDM)1 is caused by the degeneration of insulin-producing ␤ cells in pancreatic islets (1-4). Nitric oxide (NO ⅐ ), first identified as a physiological signaling molecule, has been also shown to be a cytotoxic effector molecule when generated in high concentrations by type 2 NO ⅐ synthase (NOS2) (5). In the process of IDDM, activated macrophages produce NO ⅐ , which is thought to be cytotoxic to ␤ cells, and NO ⅐ , which is produced by ␤ cell NOS2 induced by macrophage-derived cytokines such as interleukin-1␤ (IL-1␤), is also thought to be involved in ␤ cell degeneration (6). Although many in vitro studies (7-13) suggest that NO ⅐ produced by cytokine-induced NOS2 can cause the degeneration of ␤ cells, no in vivo study has clearly demonstrated the pathological significance of NO ⅐ produced within ␤ cells in the development of IDDM, because an infiltration of macrophages in islets always occurred in animal models of IDDM (6,14). In this study, we produced transgenic mice expressing NOS2 constitutively in pancreatic ␤ cells and found that the transgenic mice developed severe IDDM without macrophage or lymphocyte infiltration in and around islets. EXPERIMENTAL PROCEDURESConstruction of Rat Insulin II Promoter/Mouse NOS2 Hybrid GeneIslets were isolated from ICR mice by the collagenase digestion method (15) and cultured for 12 h in the presence of 150 units/ml IL-1␤ (Sigma). A mouse NOS2 cDNA was cloned by polymerase chain reaction (PCR) of reverse-transcribed RNA from IL-1␤-stimulated islets. Primers used in the PCR reaction were 5Ј-TTCCCGGGAGCAGAAGTGCAAAGTCTCA-GAC-3Ј and 5Ј-AAAGATCTGGGCTGTCAGAGCCTCGTGGCTTT-3Ј; these sequences correspond to the nucleotides Ϫ25 to Ϫ1 and 3418 to 3444 of mouse NOS2 cDNA (16) and contain XmaI and BglII sites (underlined sequences), respectively. The cloned cDNA sequence was determined and was found to be exactly the same as the reported NOS2 sequence (16). To express NOS2 in ␤ cells, the 0.7-kbp BamHI-XmaI fragment of the rat insulin II promoter (17), the 3.5-kbp XmaI-BglII fragment of mouse NOS2 cDNA containing the entire coding region, and the 1....

Theophylline, in addition to its bronchodilator effect, is reported to have an antiinflammatory action that may account for its clinical effectiveness in the reduction of inflammatory cells in the airway. In bronchial asthma, such inflammatory cytokines as GM-CSF and IL-5 are upregulated and have been proposed to cause granulocyte infiltration (neutrophils and eosinophils) in the airway by inhibition of granulocyte apoptosis. We examined the abilities of theophylline to counteract the prolongation of human granulocyte survival caused by cytokines. Theophylline was shown to shorten granulocyte survival in a dose-dependent manner. Upon incubation with a therapeutical concentration of theophylline (0.1 mM; 18 g/ml), percentages of GM-CSF (10 ng/ml)-induced delayed apoptosis increased from 18 Ϯ 2% to 38 Ϯ 3% ( P Ͻ 0.02) in neutrophils and from 21 Ϯ 2% to 35

Cyclic ADP-ribose (cADPR) has been shown to be a mediator for intracellular Ca 2 ϩ mobilization for insulin secretion by glucose in pancreatic ␤ cells, and CD38 shows both ADP-ribosyl cyclase to synthesize cADPR from NAD ϩ and cADPR hydrolase to hydrolyze cADPR to ADP-ribose. We show here that 13.8% of Japanese non-insulin-dependent diabetes (NIDDM) patients examined have autoantibodies against CD38 and that the sera containing anti-CD38 autoantibodies inhibit the ADP-ribosyl cyclase activity of CD38 ( P Յ 0.05). Insulin secretion from pancreatic islets by glucose is significantly inhibited by the addition of the NIDDM sera with anti-CD38 antibodies ( P Յ 0.04-0.0001), and the inhibition of insulin secretion is abolished by the addition of recombinant CD38 ( P Յ 0.02). The increase of cADPR levels in pancreatic islets by glucose was also inhibited by the addition of the sera ( P Յ 0.05). These results strongly suggest that the presence of anti-CD38 autoantibodies in NIDDM patients can be one of the major causes of impaired glucose-induced insulin secretion in NIDDM.

B cells can differentiate into the antibody-secreting cells, plasma cells, whereas the crucial signals that positively control the entry into the pathway to plasma cells have been unclear. Triggering via CD27 by CD27 ligand (CD70) on purified peripheral blood B cells yielded an increase in the number of plasma cells in the presence of interleukin-10 (IL-10). Differentiation into plasma cells by a combination of IL-10 and CD70 transfectants occurred in CD27+ B cells but not in CD27− B cells. Moreover, addition of IL-2 to the IL-10 and CD70-transfect activation system greatly induced differentiation into plasma cells. In the presence of only IL-2, IL-4, or IL-6, CD70 transfectants did not promote differentiation into plasma cells. On the other hand, CD40 signaling increased the expansion of a B-cell pool from peripheral blood B cells primarily activated by IL-2, IL-10, and anti-CD40 monoclonal antibody (MoAb). Finally, CD27 signaling also rescued B cells from IL-10–mediated apoptosis. These data demonstrate that CD27 ligand (CD70) is a key molecule to prevent the IL-10–mediated promotion of apoptosis and to direct the differentiation of CD27+ memory B cells toward plasma cells in cooperation with IL-10.