Soon-Youn Choi

Erythropoietin Decreases Renal Fibrosis in Mice with Ureteral Obstruction: Role of Inhibiting TGF-β–Induced Epithelial-to-Mesenchymal Transition

The inhibitory effects of recombinant human erythropoietin (rhEPO) were examined against (1) the progression of renal fibrosis in mice with complete unilateral ureteral obstruction and (2) the TGF-beta1-induced epithelial-to-mesenchymal transition (EMT) in MDCK cells. Unilateral ureteral obstruction was induced in BALB/c mice and rhEPO (100 or 1000 U/kg, intraperitoneally, every other day) or vehicle was administered from day 3 to day 14. Immunoblotting and immunohistochemistry revealed increased expressions of TGF-beta1, alpha-smooth muscle actin (alpha-SMA), and fibronectin and decreased expression of E-cadherin in the obstructed kidneys. In contrast, rhEPO treatment significantly attenuated the upregulation of TGF-beta1 and alpha-SMA and the downregulation of E-cadherin. MDCK cells were treated with TGF-beta1 (5 ng/ml) for 48 h to induce EMT, and the cells were then co-treated with TGF-beta1 and rhEPO for another 48 h. Increased expressions of alpha-SMA and vimentin and decreased expressions of zona occludens-1 and E-cadherin were observed after TGF-beta1 treatment, and these changes were markedly attenuated by rhEPO co-treatment. TGF-beta1 increased phosphorylated Smad-2 expression in MDCK cells, which was decreased by rhEPO co-treatment. In conclusion, rhEPO treatment inhibits the progression of renal fibrosis in obstructed kidney and attenuates the TGF-beta1-induced EMT. It is suggested that the renoprotective effects of rhEPO could be mediated, at least partly, by inhibition of TGF-beta1-induced EMT.

Impact of low glucose degradation product bicarbonate/lactate-buffered dialysis solution on the epithelial-mesenchymal transition of peritoneum.

Background: Epithelial-mesenchymal transition (EMT) is important in the development of peritoneal fibrosis. Glucose degradation products (GDPs) may induce EMT in human peritoneal mesothelial cells (HPMCs). Methods: The effects of individual GDPs and GDPs derived from peritoneal dialysis fluid (PDF) in both HPMCs and peritoneal membranes were evaluated. EMT was assessed with α-smooth muscle actin (α-SMA) and E-cadherin. Results: In vitro, α-SMA protein and mRNA levels increased in the presence of the GDPs (formaldehyde, glyoxal, methylglyoxal, and 3-deoxyglucosone), and E-cadherin decreased. Changes in the EMT markers were most prominent after exposure to 3-deoxyglucosone. Changes in both α-SMA and E-cadherin protein levels were less with low (L)-GDP bicarbonate/lactate-buffered PDF compared to high (H)-GDP PDF. In the rat model after 8 weeks’ PDF infusion, the α-SMA/E-cadherin mRNA ratio increased in the H-GDP group compared with the L-GDP group (p < 0.05). The peritoneum in the H-GDP group tended to be thicker (p = 0.052) and had more blood vessels than that in the L-GDP group (p < 0.05). Tissue staining for TGF-β1 decreased in the L-GDP group. Dual-stained cytokeratin and α-SMA-positive myofibroblasts in the submesothelial layer were more prominent in the H-GDP group. Conclusion: GDPs found in PDF induce EMT of HPMCs, which is associated with peritoneal fibrosis and vascularization. Conversely, L-GDP PDF reduces EMT and peritoneal fibrosis.

Effects of Low Glucose Degradation Products Peritoneal Dialysis Fluid on the Peritoneal Fibrosis and Vascularization in a Chronic Rat Model

Abstract:  In the present study, we examined the effects of a new peritoneal dialysis fluid (PDF) with a low level of low glucose degradation products (GDP) on the functional and structural stability of the peritoneal membrane (PM). Male Sprague‐Dawley rats were divided into three groups: group C (n = 8), without dialysate infusion; group P (n = 12), infused with low‐level GDP solution (4.25% Physioneal, pH 7.0–7.4); and group D (n = 12), infused with conventional solution (4.25% Dianeal, pH 5.2, adjusted to pH 7.0). In groups D and P, animals were infused through a permanent catheter with 25 mL of PDF, twice daily for 8 weeks. Lipopolysaccharide was added into the PDF immediately before infusion on days 8, 9 and 10 in the two dialysis groups. When compared with group P, group D showed a higher glucose mass transfer at weeks 6 and 8, D/P urea at week 8, TGF‐β1 at weeks 4 and 8, and VEGF level at week 8. The submesothelial matrix layer of the parietal peritoneum was significantly thickened in group D and the lectin‐stained blood vessels in this layer were well‐visualized in group D compared with group P. There were significantly more peritoneal blood vessels in group D than group P. The transforming growth factor‐β induced gene‐h3 (βig‐h3) and TGF‐β1 levels in the peritoneal effluent correlated with the submesothelial thickness, which correlated with the dialysate‐to‐plasma ratio (D/P) of protein and, inversely, with the rate of glucose transport (D/D0 glucose, where D is glucose concentration in the dialysate and D0 is glucose concentration in the dialysis solution before it is infused into the peritoneal cavity). The present study showed that low‐GDP PDF effectively attenuated the peritoneal vascularization and fibrosis related to conventional solution.

Phosphate-Induced Apoptosis in Human Peritoneal Mesothelial Cells in vitro

Background: It has been demonstrated that phosphate uptake through the type III sodium-dependent phosphate co-transporter, Pit-1, induced apoptosis of aortic vascular smooth muscle cells and endothelial cells in vitro. However, the apoptotic effects of high phosphate (HP) level in human peritoneal mesothelial cells (HPMCs) are not known. Methods: To examine whether Pit-1 is expressed in HPMCs, we checked the Western blot assay of immunoreactive Pit-1 and the transcription of Pit-1 by reverse transcriptase PCR. We treated several different phosphate concentrations (1–4 mM) and calcium concentrations (1.8 and 2.8 mM) on HPMCs to assess the effects of concentration. MTT, TUNEL assays, and flow cytometry analysis using Annexin V and propidium iodide were performed to identify cell death and apoptosis. Bax and Bcl-2 by Western blot and caspase-3 activity were evaluated by colorimetric assay. In addition, phosphonoformic acid (PFA) and pan-caspase inhibitor, Z-VAD-FMK, were given to prevent phosphate-induced apoptosis. Results: Pit-1 expression on HPMCs was demonstrated. Apoptosis in HPMCs significantly increased with a high concentration of phosphate in a dose- and time-dependent manner, and was enhanced in the presence of 2.8 mM calcium. HP concentrations significantly decreased the anti-apoptotic Bcl-2/Bax ratio and increased caspase-3 activity. The treatment with PFA and Z-VAD-FMK prevented cell death by HP. Conclusion: Phosphate uptake through Pit-1 induces apoptosis in HPMCs by a caspase-related mechanism.

Dipeptidyl peptidase-4 inhibitor gemigliptin protects against vascular calcification in an experimental chronic kidney disease and vascular smooth muscle cells.

Although dipeptidyl peptidase-4 inhibitors, a class of antidiabetic drugs, have various pleiotropic effects, it remains undetermined whether gemigliptin has a beneficial effect on vascular calcification. Therefore, this study was performed to evaluate the effect of gemigliptin on vascular calcification in a rat model of adenine-induced chronic kidney disease and in cultured vascular smooth muscle cells. Gemigliptin attenuated calcification of abdominal aorta and expression of RUNX2 in adenine-induced chronic kidney disease rats. In cultured vascular smooth muscle cells, phosphate-induced increase in calcium content was reduced by gemigliptin. Gemigliptin reduced phosphate-induced PiT-1 mRNA expression, reactive oxygen species generation, and NADPH oxidase mRNA expression (p22phox and NOX4). The reduction of oxidative stress by gemigliptin was associated with the downregulation of phospho-PI3K/AKT expression. High phosphate increased the expression of frizzled-3 (FDZ3) and decreased the expression of dickkopf-related protein-1 (DKK-1) in the Wnt pathway. These changes were attenuated by gemigliptin treatment. Gemigliptin restored the decreased expression of vascular smooth muscle cells markers (α-SMA and SM22α) and increased expression of osteogenic makers (CBFA1, OSX, E11, and SOST) induced by phosphate. In conclusion, gemigliptin attenuated vascular calcification and osteogenic trans-differentiation in vascular smooth muscle cells via multiple steps including downregulation of PiT-1 expression and suppression of reactive oxygen species generation, phospho-PI3K/AKT, and the Wnt signaling pathway.

Histopathological and scanning electron microscopy findings of retrieved porous polyethylene implants

Porous polyethylene (PPE) implants are biocompatible alloplastic materials commonly used for facial augmentation. However, the effect of sub-periosteal PPE application on the surrounding tissues has not been analyzed clearly. This report documents the case of a 22-year-old woman who underwent peri-alar augmentation with PPE to improve midface retrusion. Although no infection or inflammation occurred at the surgical site, the patient requested removal of the PPE implant for aesthetic reasons alone at 1 year after the surgery. The removed implant was subjected to histological and morphological evaluation using conventional histological staining and scanning electron microscopy (SEM). Histopathological staining revealed bone ingrowth into the pores of the implant near the boundary with the host bone. Little evidence of a foreign body reaction was observed. SEM revealed densely arranged collagen fibres and osteoblastic cells in the pores. Moreover, the outer surface of the PPE implant in contact with the periosteum showed fibrous tissue ingrowth, leading to tissue adhesion. These findings confirm bone ingrowth into the PPE pore structure in humans.