s-nitro-n-acetylpenicillamine and Insulinoma

Coptidis rhizoma (CR) is a herb used in many traditional prescriptions against diabetes mellitus in Asia for centuries. Our purpose was to determine the protective effect and its action mechanism of CR on the cytotoxicity of pancreatic beta-cells. Nitric oxide (NO) is believed to play a key role in the process of pancreatic beta-cell destruction leading to insulin-dependent diabetes mellitus (IDDM). Exposure of RINm5F cells to chemical NO donor such as S-nitroso-N-acetylpenicillamine (SNAP) induced apoptotic events such as the disruption of mitochondrial membrane potential (Deltapsim), cytochrome c release from mitochondria, activation of caspase-3, poly (ADP-ribose) polymerase cleavage and DNA fragmentation. Also, exposure of SNAP led to LDH release into medium, one of the necrotic events. However, pretreatment of RINm5F cells with CR extract protected both apoptosis and necrosis through the inhibition of Deltapsim disruption in SNAP-treated RINm5F cells. In addition, rat islets pretreated with CR extract retained the insulin-secretion capacity even after the treatment with IL-1beta. These results suggest that CR may be a candidate for a therapeutic or preventing agent against IDDM.

Topics: Animals; Apoptosis; Cell Line, Tumor; Coptis chinensis; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Insulinoma; Islets of Langerhans; L-Lactate Dehydrogenase; Male; Membrane Potentials; Mitochondria; Necrosis; Nitric Oxide Donors; Pancreatic Neoplasms; Penicillamine; Rats; Rats, Sprague-Dawley

Pancreatic islets and insulinoma cells are particularly vulnerable to serious damage by cytotoxic nitric oxide (NO) and/or oxidative stress, most probably due to their low expression levels of antioxidant enzymes. This cellular damage has been regarded as one of major obstacles to success of encapsulated islet transplantation for the treatment of type 1 diabetes. As an approach to preventing NO induced damage, crosslinked hemoglobin (Hb-C) with poly(ethylene glycol) was co-encapsulated with rat islets or insulinoma cells (RINm5F) in alginate/poly(L-lysine)/alginate microcapsules. Hb-C effectively protected the cells from NO damage, generated by treating the cell microcapsules with S-nitroso-N-acetylpenicillamine (SNAP, a nitric oxide donor) at concentrations up to 400 microM, preserving higher viability and insulin secretion than a control group (no SNAP and no Hb-C). When the cells were incubated with SNAP without Hb, there was SNAP concentration dependent cellular damage, and a colorimetric TUNEL assay revealed a typical cell apoptosis sign, indicating DNA damages.

Topics: Animals; Cell Line, Tumor; Cell Survival; Cells, Cultured; Coated Materials, Biocompatible; Cross-Linking Reagents; Hemoglobin C; Insulinoma; Islets of Langerhans; Islets of Langerhans Transplantation; Male; Nitric Oxide; Pancreatic Neoplasms; Penicillamine; Rats; Rats, Sprague-Dawley

An excessive production of nitric oxide (NO) in response to cytokines has been shown to be the major cause of the destruction of islet beta-cells associated with type 1 (insulin-dependent) diabetes mellitus. The NO-induced beta-cell death is the typical apoptosis. In the present study, we show evidence that supports a tight link between NO, Ca2+, protease and apoptosis in beta-cells. Three different NO donors, SNAP, NOR3 and NOC7, induced apoptosis in a beta-cell line, MIN6 cells, in a concentration-dependent manner. SNAP at 200 microM increased cytosolic Ca2+ concentration ([Ca2+]i) and induced apoptosis. The SNAP-induced apoptosis was blocked by a Ca2+ chelator, BAPTA-AM, and by an inhibitor of a Ca2+-dependent protease, calpain. In conclusion, an excessive NO production induces apoptosis, wherein an increase in [Ca2+]i and resultant activation of calpain play a key role.

Topics: Animals; Apoptosis; Calcium; Cell Survival; Chelating Agents; Diabetes Mellitus, Type 1; Egtazic Acid; Insulinoma; Islets of Langerhans; Mice; Nitric Oxide; Nitric Oxide Donors; Pancreas; Pancreatic Neoplasms; Penicillamine; Tumor Cells, Cultured