メチルグリオキサールによる腹膜線維化に対するリナグリプチンの効果

広島大学(Hiroshima University)博士(医学)Doctor of Philosophy in Medical Sciencedoctora

Linagliptin Ameliorates Methylglyoxal-Induced Peritoneal Fibrosis in Mice

Recent studies have reported increases of methylglyoxal (MGO) in peritoneal dialysis patients, and that MGO-mediated inflammation plays an important role in the development of peritoneal fibrosis through production of transforming growth factor-β1 (TGF-β1). Linagliptin, a dipeptidyl peptidase-4 inhibitor, exerts anti-inflammatory effects independent of blood glucose levels. In this study, we examined whether linagliptin suppresses MGOinduced peritoneal fibrosis in mice. Male C57/BL6 mice were divided into three groups: control, MGO injection plus saline, and MGO injection plus linagliptin (n = 6 per group). Peritoneal fibrosis was induced by daily intraperitoneal injection of saline containing 40 mmol/L MGO for 21 days. Saline was administered intraperitoneally to the control group. Linagliptin (10 mg/kg) or saline were administrated by once-daily oral gavage from 3 weeks before starting MGO injections. Immunohistochemical staining revealed that linagliptin suppressed expression of α-smooth muscle actin and fibroblast-specific protein-1, deposition of type I and III collagen, and macrophage (F4/80) infiltration. Peritoneal equilibration testing showed improved peritoneal functions in mice treated with linagliptin. Peritoneal injection of MGO increased plasma levels of glucagon-like peptide-1 (GLP-1) in mice, and a further increase was observed in linagliptin-treated mice. Although MGO increased plasma glucose levels, linagliptin did not decrease plasma glucose levels. Moreover, linagliptin reduced the TGF-β1 concentration in the peritoneal fluid of MGO-treated mice. GLP-1 receptor (GLP-1R) was expressed in monocytes/macrophages and linagliptin suppressed GLP-1R expression in MGO-injected mice. These results suggest that oral administration of linagliptin ameliorates MGO-induced peritoneal fibrosis.This work was supported by a grant from Ryokufukai and the Japanese Association of Dialysis Physicians

Application of Purified Botulinum Type A Neurotoxin to Treat Experimental Trigeminal Neuropathy in Rats and Patients with Urinary Incontinence and Prostatic Hyperplasia

Type A neurotoxin (NTX) of Clostridium botulinum was purified by a simple procedure using a lactose gel column. The toxicity of this purified toxin preparation was retained for at least 1 year at −30°C by supplementation with either 0.1% albumin or 0.05% albumin plus 1% trehalose. When purified NTX was used to treat 49 patients with urinary incontinence caused by either refractory idiopathic or neurogenic detrusor overactivity, 36 patients showed significant improvement in symptoms. These beneficial effects were also observed in cases of prostatic hyperplasia. The results obtained with NTX were similar to that of Botox. The effects of NTX on trigeminal neuralgia induced by infraorbital nerve constriction (IoNC) in rats were also studied. Trigeminal ganglion neurons from ipsilateral to IoNC exhibited significantly faster onset of FM4-64 release than sham-operated contralateral neurons. Intradermal injection of NTX in the area of IoNC alleviated IoNC-induced pain behavior and reduced the exaggerated FM4-64 release in trigeminal ganglion neurons

High-dose Dexamethasone Therapy as the Initial Treatment for Idiopathic Thrombocytopenic Purpura: Protocol for a Multicenter, Open-label, Single Arm Trial

Standard therapy for idiopathic thrombocytopenic purpura (ITP) has not been established. We are conducting a multicenter, prospective trial to determine the efficacy and safety of short-term, high-dose dexamethasone therapy in ITP patients aged 18-80 years with platelet counts of <20, 000 /μL, or with <50, 000/ μL and bleeding symptoms. The primary endpoints of this trial are the proportion of responses (complete plus partial response) on day 180 (day 46+180) after the completion of the 46-day high-dose dexamethasone therapy. The results of this investigation of the effectiveness and safety of this regimen will be essential for the establishment of standard therapy for ITP

Inhibition of H3K9 methyltransferase G9a ameliorates methylglyoxal-induced peritoneal fibrosis

<div><p>Activity of H3K9 histone methyltransferase G9a is reportedly induced by transforming growth factor-β1 (TGF-β1) and plays an important role in the progression of cancer and fibrosis. In this study, we investigated whether inhibition of G9a-mediated H3K9 methylation attenuates peritoneal fibrosis in mice and human peritoneal mesothelial cells (HPMCs). Nonadherent cells of peritoneal dialysis (PD) patients were isolated from PD effluent to examine expression of G9a. Peritoneal fibrosis was induced by peritoneal injection of methylglyoxal (MGO) in male C57/B6 mice for 3 weeks. BIX01294, a G9a inhibitor, was administered by subcutaneous injection. Effects of BIX01294 on MGO-induced pathological and functional changes in mice were evaluated by immunohistochemistry and a peritoneal equilibration test. HPMCs were isolated from human omentum, and the inhibitory effect of BIX01294 on TGF-β1-induced fibrotic changes was investigated in the HPMCs by western blotting. G9a was upregulated in nonadherent cells of human PD effluent, the peritoneum of MGO-injected mice, and TGF-β1-stimulated HPMCs. BIX01294 significantly reduced the submesothelial zone thickness and cell density in MGO-injected mice. Immunohistochemical staining revealed that BIX01294 treatment decreased not only mono-methylation of H3K9 (H3K9me1), but also the number of mesenchymal cells, accumulation of collagen, and infiltration of monocytes. In addition to the pathological changes, BIX01294 reduced the level of TGF-β1 in peritoneal fluid and improved peritoneal functions. Furthermore, BIX01294 inhibited TGF-β1-induced fibrotic changes along with suppression of H3K9me1 in HPMCs. Therefore, inhibition of H3K9 methyltransferase G9a suppresses peritoneal fibrosis through a reduction of H3K9me1.</p></div

BIX01294 inhibits monocyte/macrophage infiltration, TGF-β1, and H3K9me1 in mice with peritoneal fibrosis.

<p>(A) Immunohistochemical staining shows typical CD68 expression in peritoneal tissue of control mice, MGO-injected mice, and MGO-injected mice treated with BIX01294 (original magnification, ×200). (B) Graph indicates the number of CD68-positive cells in the three groups of mice. (C) Immunohistochemical staining shows typical TGF-β1 expression in peritoneal tissue of control mice, MGO-injected mice, and MGO-injected mice treated with BIX01294 (original magnification, ×200). (D) Graph indicates the number of TGF-β1-positive cells in the three groups of mice. (E) Two-color immunohistochemical staining shows that most CD68 cells (brown) are immunoreactive for TGF-β1 (blue-gray) (arrows). (F) TGF-β1 protein levels in mouse PD effluent were quantified by ELISA. (G) Immunohistochemical staining shows typical H3K9me1 levels in peritoneal tissue of control mice, MGO-injected mice, and MGO-injected mice treated with BIX01294 (original magnification, ×200). (H) Graph indicates the number of H3K9me1-positive cells in the three groups of mice. Data are expressed as the mean ± SE. Statistical analysis were performed by analysis of variance followed by Tukey’s post-hoc test. *<i>P</i> < 0.05, n = 5 mice per group.</p

Linagliptin suppresses peritoneal cell density and thickening in methylglyoxal (MGO)-injected mice.

<p>(a, b) Representative light microscopy of peritoneal tissues on day 21 (a: hematoxylin-eosin staining; b: Masson’s trichrome staining; original magnification: ×200) in control mice, MGO-injected mice treated with saline, and MGO-injected mice treated with linagliptin. (c, d) The thickness of the submesothelial compact zone increased along with its cellularity until day 21 in mice treated with saline, whereas cell density and zone thickening were suppressed in mice treated with linagliptin. Bars indicate the compact zone area. Statistical analyses were performed using analysis of variance (ANOVA) followed by Tukey’s <i>post-hoc</i> test. *<i>P</i> < 0.05.</p