Ryozo Kamimura

Characterization and species differences in gastric ghrelin cells from mice, rats and hamsters

Ghrelin is a newly identified gastric peptide hormone that has various important functions, including growth‐hormone release and appetite stimulation. Ghrelin‐immunoreactive cells (ghrelin cells) are characterized by X‐type endocrine cells in the rat stomach. In the present study, we analysed ghrelin cells in fundi of stomach from ICR mice and Syrian hamsters immunohistochemically, immunoelectron microscopically and morphometrically, and compared the results with those from Wistar rats. Immunohistochemistry revealed that ghrelin cells were sparsely distributed in the proper gastric glands in all species. The number of ghrelin cells per unit area in hamsters was significantly lower than that in rats. Immunoelectron microscopy detected ghrelin immunolabelling in granules in the X‐type endocrine cells. However, the diameter of granules in the hamsters was significantly smaller than that in the mice and rats. Gastric ghrelin contents were determined by radioimmunoassay, and levels in the hamsters were significantly lower than those in mice and rats. The results from mice were identical to those from rats. In conclusion, gastric ghrelin cells in mice and hamsters are characterized by X‐type endocrine cells, as has been observed in rats. However, the data indicated that gastric ghrelin production was lower in hamster than in mouse or rat.

HMGB1 release in co-cultures of porcine endothelial and human T cells

Abstract:  High mobility group box‐1 (HMGB1) protein, primarily from the nucleus, is released into the extracellular milieu either passively by necrotic or damaged cells, or actively by secretion from monocytes/macrophages. Extracellular HMGB1 acts as a potent inflammatory stimulator by promoting cytokine (for example, tumor necrosis factor‐α) production, and also has pro‐coagulant activity. The signaling pathway initiated by receptor for advanced glycation end‐product (RAGE), which is the HMGB1 receptor, also induces complement activation. Recent studies have implicated HMGB1 in acute cardiac allograft rejection, and have identified infiltrating T cells and other damaged cells as its main sources. HMGB1 blockade using the anti‐HMGB1 antibody HMGB1 box‐A (amino‐terminal region) and soluble RAGE rescues mice from acute rejection. We therefore studied the release of HMGB1 in co‐cultures of porcine aortic endothelial cells (PAEC) and human leukocytes. Human T cells, but not B cells, monocytes or neutrophils, stimulated significant HMGB1 release in culture with PAEC; this activity required cell–cell contact and was dose‐dependent, as determined by Western blotting. The released HMGB1 originated from both cell types, as immunofluorescent microscopy showed that it was present in the cytosol of PAEC in contact with T cells, and had disappeared from the T‐cell nuclei. These results demonstrate that direct interactions between PAEC and T cells might be a key factor in triggering HMGB1 release, which suggests that HMGB1 is associated with graft rejection in the early phase.

Hydrogen Improves Glycemic Control in Type1 Diabetic Animal Model by Promoting Glucose Uptake into Skeletal Muscle

Hydrogen (H2) acts as a therapeutic antioxidant. However, there are few reports on H2 function in other capacities in diabetes mellitus (DM). Therefore, in this study, we investigated the role of H2 in glucose transport by studying cultured mouse C2C12 cells and human hepatoma Hep-G2 cells in vitro, in addition to three types of diabetic mice [Streptozotocin (STZ)-induced type 1 diabetic mice, high-fat diet-induced type 2 diabetic mice, and genetically diabetic db/db mice] in vivo. The results show that H2 promoted 2-[14C]-deoxy-d-glucose (2-DG) uptake into C2C12 cells via the translocation of glucose transporter Glut4 through activation of phosphatidylinositol-3-OH kinase (PI3K), protein kinase C (PKC), and AMP-activated protein kinase (AMPK), although it did not stimulate the translocation of Glut2 in Hep G2 cells. H2 significantly increased skeletal muscle membrane Glut4 expression and markedly improved glycemic control in STZ-induced type 1 diabetic mice after chronic intraperitoneal (i.p.) and oral (p.o.) administration. However, long-term p.o. administration of H2 had least effect on the obese and non-insulin-dependent type 2 diabetes mouse models. Our study demonstrates that H2 exerts metabolic effects similar to those of insulin and may be a novel therapeutic alternative to insulin in type 1 diabetes mellitus that can be administered orally.

Hepatocyte growth factor improves the survival of rats with pulmonary arterial hypertension via the amelioration of pulmonary hemodynamics

Hepatocyte growth factor (HGF) is a multifunctional growth factor with mitogenic, anti-apoptotic and anti-fibrotic activities. In this study, we investigated the effect of administration of recombinant human HGF on pulmonary arterial hypertension. Pulmonary arterial hypertension was induced in rats by a single injection of monocrotaline (MCT) and recombinant human HGF (0.12 mg/day) was administered into the right ventricle cavity using osmotic pumps, which were implanted subcutaneously 21 days after MCT injection. Continuous intravenous delivery of recombinant human HGF for 14 days led to prolonged survival of animals suffering from severe MCT-induced pulmonary arterial hypertension. Although a bolus injection of recombinant human HGF did not affect pulmonary arterial pressure, a 14-day administration of recombinant human HGF attenuated the inflammatory cell infiltrate, matrix accumulation and vascular medial thickening. As a consequence, the pulmonary lumen was enlarged and the pulmonary arterial pressure was significantly reduced. Additionally, continuous administration of recombinant human HGF suppressed lung tissue expression of platelet-derived growth factor, which plays an important role in the development of pulmonary arterial hypertension. These results indicate that recombinant human HGF possibly has a great potential for improving symptoms and altering the clinical course of pulmonary arterial hypertension.

A new animal model for split liver transplantation using an infrahepatic IVC graft.

An animal model of split liver transplantation using pigs is described herein. The donor liver was divided into two grafts, the right graft consisting of the right medial and lateral segments with the caudate lobe, and the left graft consisting of the left lateral and medial segments. To make implantation easier, a distal part of the donors inferior vena cava (IVC) was isolated and attached to the left graft with an anastomosis between the orifice of the renal vein and the grafts hepatic vein. The left graft thereby contained a newly constructed retrohepatic IVC for anastomosis to the recipient. During the anhepatic phase, no conventional bypass procedure was used, but an abdominal aortic clamp in combination with general hypothermia was employed. Ten pigs were used as donors and a total 20 liver transplantations performed. No immunosuppressive drugs were given in this series. Ten of the 20 recipients survived for more than 2 days, the mean survival time being 4.7 days, with a range of 2–14 days. The mean survival time of the left grafts was much longer than that of the right grafts, although no technical problems such as kinking of the graft or occlusion of the hepatic vein were encountered in either. This model is the first report of split liver transplantation in animals. The advantages of using the infrahepatic IVC graft include stability of the graft and safe hepatectomy. This model will therefore be useful for the experimental study of split liver transplantation and may also be employed for clinical use in the future.

Abdominal cavity cooling for general hypothermia - An experimental and clinical study.

一般腹部手術時の重要臓器血流遮断を安全にかつ簡便に行うために腹腔内冷却低体温法を実験および臨床の両面で検討した. 実験では雑種成犬を開腹後腹腔内にiceslushをみたし肝, 腎などを中心に温度変化, 血流変化を観察した. 食道温30℃ までは肝, 門脈系の血流分布率が変化しないことからこの域を安全域と考えた. 実験的応用は腹部多臓器移植および肝同所性移植でおこない生存例を得て本法に問題がないことが判明した. 臨床例では, 腹部大血管手術, 肝外傷, 膵全剔門脈再建の計8例に使用し良好な成績が得られた. 腹腔内冷却低体温法は簡便, 安全で導入が早く応用の範囲は広いと考える.

Size disparity between donor and recipient in canine heart transplantation

To assess the feasibility and function of small donor hearts for large recipients, small donor hearts were orthotopically transplanted into large recipients. Thirty adult mongred dogs were used to form donor-recipient (D-R) combinations with the D-R heart weight ratio ranging from 0.47 to 1.78. In the D-R heart weight ratio of 0.8 or above (mean 1.11, Group I), the successful transplantation rate was 57 per cent, while it was 81 per cent in cases of less than 0.8 (mean 0.67, Group II) (no statistical difference). In 10 dogs (5 with D/R heart weight ratio ≧0.8, 5 with <0.8) a recovery rate in cardiac function of the transplanted small heart was studied 2 hours following transplantation. There was no significant difference, hemodynamically in the normal range of left atrial pressure (LAP) between the two groups. The close relationship between cardiac output (CO) and volume load in the range of 10 to 15 mmHg of LAP was transformed to a linear function in the over 0.8 group. It was technically and hemodynamically possible to transplant small dog hearts to larger recipients up to the D-R heart weight ratio of 0.47. The importance of an adequate heart rate for the increase of CO in transplanted smaller donor hearts was emphasized.