| Objective:To observe the therapeutic efficacy of formula of warming yang,promoting blood circulation and diuress on podocyte cathepsin L of adriamycin-induced rats with minimal change nephropathy model and puromycin aminonucleoside-induced injury of mouse glomerular podocytes,and to explore the formula,s mechanisms of action of reducing urinary protein by the way of podocyte cathepsin L.Methods:1.One-time tail vein injection of 5.5mg/kg ADR induced nephropathy rats model similar to human minimal change nephrotic syndrome.Rats were randomly divided into model group,Chinese herb group,prednisone group and normal group.After 1 week,the Chinese herb group once a day by gastric gavage 3ml formula of warming yang,promoting blood circulation and diuress,prednisone group with 12mg/kg by gastric gavage prednisone suspensionml.Normal group and model group,daily fed with 3ml distilled water for 4 weeks.24 hours urinaryalbumin excretion were measured by SS turbidimetric method in rats(in front of the moding,moding after 1 week,5 week).The distribution and protein expression of synaptopodin were measured by immuneofluorescence.The distribution and protein expression of cathepsin L were measured by immunohistochemical.Besides,the influence of treatments on podocytes was examined in renal tissues by electron microscopy after the rats were sacrificed for 28 days treatment.2.Serum with the formula of warming yang,promoting blood circulation and diuress were obtained by the serum Pharmaeology method of traditional Chinese medicine.Mouse podocyte cells(MPCs) were subjected to PAN treatment alone(PAN group)or PAN plus dexamethasone(DEX group) or PAN plus serum containing traditional Chinese drugs at 10% or 20% concentration respectively(20% or 10% serum containing traditional Chinese drugs group) for 24 h.Meanwhile,Mouse podocyte cells plus cell culture or 20% serum containing traditional Chinese drugs as negative control.The expression of cathepsin L and synaptopodin in podocytes was detected by indirect cell immunofluorescence staining.FITC-conjugated phalloidin was used to stain F-actin.A cortical F-actin score index(CFS index) was designed to quantify the degree of cytoskeletal reorganization in cultured podocytes.Results:1.In model group,24-hour urinary protein was increased.The fusion and effacement of foot processes were observed,and the expression of synaptopodin significantly decreased and the expression of cathepsin L significantly increased.Compared with normal group(P<0.01).Treatment with prednisone or Chinese herb both partially ameliorated the reduction of the expression of synaptopodin and reduced the expression of cathepsin L and improved podocyte foot process effacement.Compared with model group(P<0.01).The formula of warming yang,promoting blood circulation and diuress reduces the expression of cathepsin L in adriamycin-induced rats with minimal change nephropathy model more clearly than prednisone.2.PAN significantly decreased the expression of synaptopodin and increased the expression of cathepsin L in mouse glomerular podocytes,F-actin arranged in disorder.There are cortical F-actin ring formations and CFS index is higher than normal control(P<0.01).Dexamethasone or serum containing traditional Chinese drugs in diffident concentration treatment up-regulated synaptopodin,down-regulated cathepsin L and improved PAN-induced change in CFS index. Conclusion:The formula of warming yang,promoting blood circulation and diuress attenuates the decreased expression of synaptopodin and the increased expression of cathepsin L and improves podocyte foot process effacement in adriamycin-induced rats with minimal change nephropathy model;at the same time,the serum contained the formula can up-regulated synaptopodin,down-regulated cathepsin L and alleviate PAN-induced cytoskeletal reorganization of podocytes.So it can stabilize the actin cytoskeleton,improve podocyte foot process effacement and reduce proteinuria by regulating cathepsin L. |
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