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.     

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.     

Helicobacter pylori stimulates inducible nitric oxide synthase expression and activity in a murine macrophage cell line

BACKGROUND & AIMS: Helicobacter pylori uniquely colonizes the human stomach and produces gastric mucosal inflammation. High-output nitric oxide production by inducible nitric oxide synthase (iNOS) is associated with immune activation and tissue injury. Because mononuclear cells comprise a major part of the cellular inflammatory response to H. pylori infection, the ability of H. pylori to induce iNOS in macrophages was assessed. METHODS: H. pylori preparations were added to RAW 264.7 murine macrophages, and iNOS expression was assessed by Northern blot analysis, enzyme activity assay, and NO2- release. RESULTS: Both whole H. pylori and French press lysates induced concentration-dependent NO2- production, with peak levels 20-fold above control. These findings were paralleled by marked increases in iNOS messenger RNA and enzyme activity levels. iNOS expression was synergistically increased with interferon gamma, indicating that the H. pylori effect can be amplified by other macrophage-activating factors. Studies of lipopolysaccharide (LPS) content and polymyxin B inhibition of LPS suggested that the H. pylori effect was attributable to both LPS- dependent and -independent mechanisms. CONCLUSIONS: iNOS expression in macrophages is activated by highly stable H. pylori products and may play an important role in the pathogenesis of H. pylori-associated gastric mucosal disease. (Gastroenterology 1996 Dec;111(6):1524-33)     

A second tumor necrosis factor receptor gene product can shed a naturally occurring tumor necrosis factor inhibitor.

An inhibitor of tumor necrosis factor (TNF) has been isolated from the human histiocytic lymphoma cell line U-937 that is capable of inhibiting both TNF-alpha and TNF-beta. Protein sequencing has verified that it is distinct from a previously described TNF inhibitor that is a soluble fragment of a TNF receptor molecule (TNFrI). The cDNA sequence of this second TNF inhibitor clone suggests that it is also a soluble fragment of a TNF receptor. Expression of this cDNA sequence in COS-7 cells verified that it encodes a receptor for TNF-alpha (TNFrII) that can give rise to a soluble inhibitor of TNF-alpha, presumably through proteolytic cleavage. The extracellular domain of TNFrII has significant homology with that of TNFrI and these two receptors share a striking conservation of cysteine residue alignment with the extracellular domain of the nerve growth factor receptor. These three receptor molecules are therefore members of a family of polypeptide hormone receptors.