Anti‐inflammatory effects of exendin‐4, a glucagon‐like peptide‐1 analog,on human peripheral lymphocytes in patients with type 2 diabetes

Aims/Introduction

Type 2 diabetes is characterized by dysregulation of immunity, oxidative stress and reduced incretin effects. Experimental studies suggest that glucagon‐like peptide (GLP‐1) might have immunomodulating effects. We hypothesize that GLP‐1 receptor agonist, exendin‐4, might reduce inflammatory response in type 2 diabetes.

Materials and Methods

Using peripheral blood mononuclear cells (PBMC) sampled from 10 type 2 diabetes and 10 sex‐ and age‐matched control subjects and supernatants from PBMC culture, the expression of phospho‐mitogen activated protein kinase (MAPK) signaling pathways in CD4+ T helper lymphocytes and monocytes was analyzed using flow cytometry. Cytokines/chemokines and superoxide anion before and after treatment with exendin‐4 were measured by cytometric bead array and chemiluminesence assay, respectively.

Results

Compared with control subjects, PBMC from type 2 diabetes patients showed activated MAPK (P38, c‐Jun NH2‐terminal protein kinase and extracellular signal‐regulated kinase) signaling pathway, elevated superoxide anion, increased pro‐inflammatory cytokines (tumor necrosis factor‐α, interleukin‐1β, interleukin‐6) and chemokines (CCL5/regulated on activation normal T‐cell expressed and secreted and CXCL10/interferon‐γ‐induced protein 10). These changes were attenuated by exendin‐4, possibly through the suppression of p38 MAPK.

Conclusions

These results suggest that exendin‐4 might downregulate pro‐inflammatory responses and reduce oxidative stress by suppressing MAPK signaling pathways in type 2 diabetes.     

Antibodies selected from combinatorial libraries block a tumor antigen that plays a key role in immunomodulation

We searched for cell-surface-associated proteins overexpressed on B cell chronic lymphocytic leukemia (CLL) to use as therapeutic antibody targets. Antibodies binding the immunosuppressive molecule CD200 were identified by cell panning of an antibody phage display library derived from rabbits immunized with primary CLL cells. B cells from 87 CLL patients exhibited 1.6- to 5.4-fold cell-surface up-regulation of CD200 relative to normal B cells. An effect of increased CD200 expression by CLL cells on the immune system was evaluated in mixed lymphocyte reactions. Addition of primary CLL but not normal B cells to macrophages and T cells downregulated the Th1 response, as seen by a 50-95% reduction in secreted IL-2 and IFN-gamma. Antibodies to CD200 prevented downregulation of the Th1 response in most B cell CLL samples evaluated, indicating abrogation of the CD200/CD200R interaction can be sufficient to restore the Th1 response. A disease-progression-associated shift of the immune response from Th1 to Th2 has been observed in numerous cancers. Because this cytokine shift is also believed to promote the induction of regulatory T cells, reverting the immune response to Th1 through direct targeting of the cancer cells may provide therapeutic benefits in CLL by encouraging a cytotoxic T cell response.     

Elimination of clonogenic tumor cells from HL-60, Daudi, and U-937 cell lines by laser photoradiation therapy: implications for autologous bone marrow purging

Laser photoradiation therapy was tested in an in vitro model for its efficacy in the elimination of non-Hodgkin's lymphoma cells. Results show that at 31.2 J/cm2 of laser light in the presence of 20 micrograms/mL of merocyanine 540 (MC540) there was greater than 5 log reduction in Burkitt's lymphoma (Daudi) cells. Similar tumor cell kill was obtained for leukemia (HL-60) cells at a laser light dose of 93.6 J/cm2. However, to obtain the same efficiency of killing for histiocytic lymphoma (U-937) cells, a higher dose of MC540 (25 micrograms/mL) was required. Clonogenic tumor stem cell colony formation was reduced by greater than 5 logs after laser photoradiation therapy. Under identical conditions for each cell line the percent survival for granulocyte-macrophage colony-forming units (CFU-GM, 45.9%, 40%, 17.5%), granulocyte/erythroid/macrophage/megakaryocyte (GEMM, 40.1%, 20.1%, 11.5%), colony-forming units (CFU-C, 16.2%, 9.1%, 1.8%), and erythroid burst-forming units (BFU-E, 33.4%, 17.8%, 3.9%) was significantly higher than the tumor cells. Mixing of gamma ray- irradiated normal marrow cells with tumor cells (1:1 and 10:1 ratio) did not interfere with the elimination of tumor cells. The effect of highly purified recombinant interferon alpha (rIFN) on laser photoradiation therapy of tumor cells was also investigated. In the presence of rIFN (30 to 3,000 U/mL), the viability of leukemic cells was observed to increase from 0% to 1.5% with a concurrent decrease in membrane polarization, suggesting an increase in fluidity of cell membrane in response to rIFN. However, at higher doses of rIFN (6,000 to 12,000 U/mL) this phenomenon was not observed. The viability of lymphoma cells remained unaffected at all doses of rIFN tested. These results may have therapeutic relevance in patients undergoing interferon treatment who require bone marrow transplantation, as the complete elimination of tumor cells by marrow-purging procedures may be hampered by this increased survival in the presence of interferon.     

Murine thymocytes proliferate in direct response to interleukin-7

The ability of interleukin-7 (IL-7) to stimulate murine thymocyte proliferation was investigated. IL-7, either alone or in concert with lectin, induced proliferation of adult thymocytes as well as day 13 fetal and adult CD4-/CD8-thymocytes. The IL-7-induced proliferative response of unfractionated thymocytes could not be inhibited by antibodies to IL-2, or IL-4, IL-6, or the IL-2 receptor. In addition, IL-2, IL-4, and IL-6 were not produced by thymocytes activated with IL- 7, as judged by the absence of biologically active cytokine in IL-7- stimulated culture supernatants. IL-7 could act in concert with IL-2 and IL-4 or with IL-4 to enhance the proliferative response of thymocyte cultures. Thus, IL-7 may cause proliferation of thymocytes directly, not indirectly, through production of IL-2, IL-4, or IL-6. IL- 7 may then play a significant role in differentiation of T lymphocytes.