ABSTRACT
Objective To investigate the effect and mechanism of all-trans retinoic acid (ATRA) on the cyclosporin (CsA)-induced proliferation and apoptosis of glomerular mesangial cells in rat models. Methods The glomerular mesangial cells induced by different doses of CsA were treated with different doses of ATRA. MTT assay was carried out to detect cell proliferation. Hoechst 33258 fluorescent staining was adopted to observe the morphology of the apoptotic cells. Flow cytometry was conducted to detect the cellular apoptosis rate. Immunofluorescent staining was employed to quantitatively measure the expression level of mitochondria-derived pro-apoptotic Smac protein. Results Compared with the control group, administration of CsA at a dose of 0.5 μg/mL and above could suppress cellular proliferation, and use of CsA at a dose of 1.0 μg/mL and above could induce cellular apoptosis. The expression level of Smac protein was significantly up-regulated by CsA administration with a dose and time dependence (all P<0.05).Compared with the CsA group, combined administration of CsA and ATRA exerted a more significant inhibitory effect on cellular proliferation. Supplement of ATRA could significantly inhibit glomerular mesangial cellular apoptosis induced by CsA and down-regulate the expression of Smac protein with a dose dependence (both P<0.05). Conclusions CsA can inhibit cellular proliferation, induce cellular apoptosis and up-regulate the expression of Smac protein of glomerular mesangial cells. ATRA is capable of suppressing glomerular mesangial cellular apoptosis induced by CsA, which is probably mediated by the Smac signaling pathway.
ABSTRACT
Objective To investigate the influence of retinoic acid receptor (RAR-α,RAR-β and RAR-γ)-mediated all-trans retinoic acid (ATRA) on renal tissue cell proliferation and apoptosis in rats with diabetic nephropathy,and to analysze the possible mechanism. Methods Male SD rats were randomly divided into normal control group (group N,n=10) and diabetic model group (n=20).Diabetes was induced by streptozotocin(STZ) injection.After successful modeling,the model rats were randomly divided into diabetes group (group D,n=10) and ATRA treatment group (group T,n=10).Rats in group T received ATRA 10 mg·kg-1·d-1 by gavage from the 2nd day of successful modeling for 8 or 12 weeks,meanwhile group N and group D received same volume distilled water.In each group,5 rats were sacrificed respectively at the 8th week or the 12th week,then biochemical markers were measured and kidney pathology was examined.Apoptosis index(AI)of renal tissue cells of each group was tested by TUNEL.The expressions of RAR-α,RAR-β and RAR-γ in renal tissues were tested using indirect immunofluorescence.The expressions of type Ⅰ collagen and laminin as proliferation indicators,along with Smac and caspase-3 as the correlated factors of apoptosis in renal tissue of each group were tested by immunohistochemistry staining.The mRNA expressions of Smac and caspase-3 were tested using real-time fluorescence quantitative PCR. Results Compared with group N,24 h urine protein,serum creatinine,blood urea nitrogen,ratio of kidney weight/body weight increased significantly (P＜0.05,respectively) in group D,and further increased with observation time.Compared with the group D,24 h urine protein and ratio of kidney weight/body weight decreased in group T (P＜0.05,respectively).Compared with group D,the group T presented minor pathological changes.TUNEL assay indicated that compared with group N,the group D showed an obvious increase in renal cell apoptosis in time-dependent manner,and the group T showed a decrease compared with the group D (P＜0.01,respectively).Compared with group N,the expression of RAR-α and RAR-β positive cells number in group D were decreased (P＜0.01,respectively).Compared with group D,the expression of RAR-α and BAR-β positive cells number in group T increased (P＜0.01,respectively).Renal tissues of each group did not show expressions of RAR-γ.After 12 weeks,compared with group N,expressions of type-Ⅰ collagen,laminin,Smac and caspase-3 protein in the glomerular mesangial area and basement membrane of renal tissues in group D increased significantly (P＜0.01,respectively),and enhanced with time.Compared with the group D,expressions of type Ⅰ collagen,laminin,Smac and caspase-3 protein in group T decreased (P＜0.01,respectively).Compared with the group N,group D had an obvious increase in the mRNA expressions of Smac and caspase-3,and a significantly decrease in group T (P＜0.01,respectively). Conclusions ATRA may prevent the cell proliferation and apoptosis in diabetic renal tissue through its receptor-mediated pathway,and may protect rats against diabetic nephropathy.