Jung Yeon Ghee

Prolonged Endoplasmic Reticulum Stress Induces Apoptotic Cell Death in an Experimental Model of Chronic Cyclosporine Nephropathy

Background/Aims: Apoptosis contributes to cyclosporine (CsA)-induced renal cell death. This study tested the effects of CsA-induced endoplasmic reticulum (ER) stress on apoptotic cell death in an experimental model of chronic CsA nephropathy. Methods: CsA (15 mg/kg per day) was given to rats for 7 or 28 days. The ER stress response was evaluated with BiP expression, and the proapoptotic response was assessed with CHOP and caspase 12 expression. ER structure was evaluated by transmission electron microscopy, and apoptotic cell death was detected with TUNEL staining. Results: Short-term treatment of CsA for 7 days activated both the ER stress response (induction of BiP mRNA and protein) and the proapoptotic response (upregulation of caspase 12 and CHOP mRNAs and proteins). However, long-term treatment with CsA for 28 days decreased BiP and further increased CHOP. The imbalance between the two responses coincided with the timing of the appearance of apoptotic cell death and the disruption of the ER structure. Conclusion: Prolonged CsA-induced ER stress causes apoptotic cell death by depleting molecular chaperones and activating the proapoptotic pathway.

Effect of Sirolimus on Calcineurin Inhibitor-Induced Nephrotoxicity Using Renal Expression of KLOTHO, an Antiaging Gene

Background. The aim of this study was to observe the effect of sirolimus (SRL) on calcineurin inhibitor (CNI)-induced nephrotoxicity in the aging process by using renal expression of KLOTHO, an antiaging gene. Methods. Mice were treated with vehicle (VH; 1 mL/kg/day of olive oil), cyclosporine A (CsA; 30 mg/kg/day), or tacrolimus (FK; 1 mg/kg/day) with or without SRL (0.3 mg/kg/day) for 2 weeks. KLOTHO expression was evaluated by using reverse-transcriptase polymerase chain reaction, immunoblotting, and immunohistochemistry. Oxidative stress was evaluated by using immunohistochemistry and urinary excretion of 8-hydroxy-2′-deoxyguanosine (8-OHdG). The calcium metabolism was evaluated by using renal ectopic calcification, serum intact parathyroid hormone level, and renal fibroblast factor 23 (FGF23) expression. Results. Treatment with CsA or FK alone significantly decreased KLOTHO expression and increased urinary 8-OHdG excretion compared with VH treatment but SRL treatment did not. Treatment SRL+CsA or SRL+FK further decreased KLOTHO expression and increased urinary 8-OHdG excretion compared with treatment of CsA or FK alone. There was a strong correlation between KLOTHO expression and urinary 8-OHdG excretion (r=−0.893; P<0.001). Treatment of CsA or FK alone increased renal ectopic calcification and serum intact parathyroid hormone level and decreased renal FGF23 expression compared with VH treatment (P<0.05) but SRL treatment did not. Treatment with SRL+CNI aggravated these parameters compared with CNI alone. Conclusions. SRL accelerates the CNI-induced oxidative process by down-regulating the renal antioxidant KLOTHO expression in the kidney.

Influence of angiotensin II on expression of toll-like receptor 2 and maturation of dendritic cells in chronic cyclosporine nephropathy.

Background. Angiotensin (Ang) II plays an important role in immune regulation. We evaluate the influence of the renin-angiotensin system (RAS) in the innate immune response caused by cyclosporine A (CsA)-induced renal injury. Methods. Two separate studies were performed in Sprague Dawley rats. First, losartan (LSRT, 10 mg/kg per day) was concurrently administered with CsA (15 mg/kg per day) for 28 days. Second, AngII (435 ng/kg/min) was infused with or without LSRT for 14 days. Results. AngII blockade with LSRT decreased toll-like receptor (TLR) 2 mRNA and protein expression, expression of tumor necrosis factor (TNF)-&agr; mRNA, and expression of major histocompatibility complex class II antigen, which was upregulated in CsA-induced renal injury. The increased number of matured dendritic cells (DCs) in CsA-induced renal injury was also decreased by concomitant treatment of LSRT. Direct infusion of AngII increased TNF-&agr; mRNA, TLR2 mRNA, and protein and the number of DCs, compared with the control rat kidney. In contrast, concomitant treatment of LSRT decreased all parameters. Conclusion. AngII plays a pivotal role in activating the innate immune response in CsA-induced renal injury.

Influence of Sirolimus on Cyclosporine‐Induced Pancreas Islet Dysfunction in Rats

This study was performed to investigate the effect of sirolimus (SRL) on cyclosporine (CsA)‐induced pancreatic islet dysfunction in rats. Three separate studies were performed. First, diabetogenic effect of SRL was evaluated with three different doses (0.15, 0.3 and 0.6 mg/kg). Second, rats were treated with SRL (0.3 mg/kg) with or without CsA (15 mg/kg) for 4 weeks. Third, rats were treated with CsA for 4 weeks, and then switched to SRL for 4 weeks. The effect of SRL on CsA‐induced pancreatic islet dysfunction was evaluated by an intraperitoneal glucose tolerance test, plasma insulin concentration, HbA1c level, HOMA‐R index, immunohistochemistry of insulin and pancreatic beta islet cell mass. The SRL treatment increased blood glucose concentration in a dose‐dependent manner. The combined treatment with SRL and CsA increased blood glucose concentration, Hemoglobin A1c (HbA1c) level, HOMA‐R [fasting insulin (mU/mL) x fasting glucose (mmol/L)]/22.5] index and decreased plasma insulin concentration, immunoreactivity of insulin and pancreatic beta islet cell mass compared with rats treated with CsA. CsA withdrawal for 4 weeks improved pancreatic beta‐cell function and structure. However, conversion from CsA to SRL further increased blood glucose levels compared with the rats converted from vehicle to SRL. The results of our study demonstrate that SRL is diabetogenic and aggravates CsA‐induced pancreatic islet dysfunction.

Expression of ammonia transporters, Rhbg and Rhcg, in chronic cyclosporine nephropathy in rats.

Background/Aims: Cyclosporine (CsA)-induced renal injury causes renal tubular acidosis. The current study was performed to evaluate the influence of CsA-induced renal injury on the ammonia transporter family members, Rh B-glycoprotein (Rhbg) and Rh C-glycoprotein (Rhcg). Methods: Rats were treated daily for 1 or 4 weeks with vehicle (VH) or CsA. Induction of chronic CsA-induced nephropathy was confirmed by demonstrating impaired renal function and characteristic histopathology. Rhbg and Rhcg expression was evaluated with immunoblot, immunohistochemistry, real-time RT-PCR and electron microscopy. Results: CsA treatment for 4 weeks developed mild metabolic acidosis and decreased urinary ammonia excretion. Rhcg mRNA expression was unchanged in both the cortex and outer medulla, but Rhcg protein expression in the CsA group was significantly reduced in the cortex and outer medulla. There were no significant differences in Rhbg mRNA and protein expression between the CsA and VH group. Conclusion: Long-term treatment with CsA in rats results in decreased urinary ammonia excretion accompanied by decreased expression of Rhcg; these changes are likely to mediate the CsA-induced defect in ammonium excretion in the collecting duct.

Upregulation of hyaluronan and its binding receptors in an experimental model of chronic cyclosporine nephropathy.

Aim:  Hyaluronan (HA) is an important extracellular matrix (ECM) proteoglycan. The localization of HA and its binding receptors, CD44 and LYVE‐1, was evaluated in an experimental model of chronic cyclosporine A (CsA)‐induced nephropathy.

Early and delayed effects of AST-120 on chronic cyclosporine nephropathy

BACKGROUND Removal of uraemic toxins by AST-120 (Kremezin(®)) decreases the progression of chronic kidney disease by reducing oxidative stress. We performed this study to evaluate whether AST-120 has a similar effect on progression of cyclosporine (CsA)-induced renal injury. METHODS Two separate studies were performed in adult Sprague-Dawley rats. First, AST-120 was administered with CsA (15 mg/kg) for 4 weeks (early treatment). Second, AST-120 was administered to the rats for 3 weeks after treatment with CsA for 3 weeks (delayed treatment). Uraemic toxin and oxidative stress were evaluated with plasma indoxyl sulphate (IS) levels and urinary 8-OHdG excretion. The effects of AST-120 on CsA-induced renal injury were evaluated in terms of renal function, interstitial fibrosis, inflammation, and apoptotic cell death. RESULTS CsA treatment for 4 weeks showed 2-fold increase in plasma IS and urinary 8-OHdG levels compared with the VH group. Early treatment with AST-120 significantly decreased both parameters, and this was accompanied by improved renal function and decreased interstitial inflammation, fibrosis, and apoptotic cell death compared with those of rats that received CsA alone. Delayed treatment with AST-120 also decreased the plasma IS and urinary 8-OHdG levels, and reduced the progression of chronic CsA nephropathy. Furthermore, delayed AST-120 treatment decreased the epithelial-mesenchymal transition in chronic CsA nephropathy. CONCLUSIONS Removal of uraemic toxins with AST-120 treatment is effective in decreasing the progression of CsA-induced renal injury by reducing oxidative stress.

Infiltration of Nestin-Expressing Cells in Interstitial Fibrosis in Chronic Cyclosporine Nephropathy

Background. Nestin-expressing cells play a role in the repair process of injured tissues and organs. This study examined the nestin-expressing cells in interstitial fibrosis in experimental chronic cyclosporine A (CsA) nephropathy. Methods. Sprague Dawley rats were treated daily for 1 or 4 weeks with CsA (15 mg/kg) or vehicle (VH; olive oil, 1 mg/kg). Nestin mRNA expression was evaluated with reverse transcriptional-polymerase chain reaction, and nestin-expressing cells were detected immunohistochemically. Localization of nestin was performed with double labeling studies for vimentin, aquaporin 1, or calbindin D28K. Results. Nestin mRNA expression was not different between VH- and CsA-treated rat kidneys. Nestin-expressing cells were rarely observed in the cortex in the VH group, but CsA-induced renal injury caused an increase in nestin-expressing cells in the cortex in a time-dependent manner. Nestin-expressing cells in the CsA group were localized to the area of interstitial fibrosis, and the number of nestin-expressing cells well correlated with the score of interstitial fibrosis (r=0.898). Nestin-expressing cells did not express vimentin, aquaporin 1, or calbindin D28K. Conclusions. CsA-induced renal injury recruits nestin-expressing cells to injured areas, and these cells might be involved in reparative fibrosis in the progression of chronic CsA nephropathy.

Comparison of Early and Late Conversion of Sirolimus in Experimental Model of Chronic Cyclosporine Nephropathy

Sirolimus (SRL) is a promising drug for replacing calcineurin inhibitors. We performed this study to determine the optimal time of conversion from cyclosporine (CsA) to SRL in an experimental model of chronic CsA nephropathy. Three separate studies were performed. In the first study, SRL was given to rats with or without CsA for 4 weeks. In the second study, rats were treated initially with CsA for 1 week, and then switched to SRL (early conversion). In the third study, CsA was given for 4 weeks and then replaced by SRL for 4 weeks treatment of CsA (late conversion). The influence of SRL on CsA-induced renal injury was evaluated by assessing renal function, histopathology (interstitial inflammation and fibrosis), and apoptotic cell death. Combined CsA and SRL treatment significantly impaired renal function, increased apoptosis, and interstitial fibrosis and inflammation compared with CsA or SRL treatment alone. Early conversion to SRL did not change renal function, histopathology, or apoptosis compared with early CsA withdrawal. By contrast, late conversion to SRL significantly aggravated these parameters compared with late CsA withdrawal. In conclusion, early conversion from CsA to SRL is effective in preventing CsA-induced renal injury in a setting of CsA-induced renal injury.