| Objective:Progressive renal fibrosis represents the final injury pathway for all commonly encountered forms of renal disease that lead to end-stage renal failure. Local proliferation of macrophages and myofibroblasts are the early pathophysiologic events and play important roles in the renal fibrosis. A late study was shown that early combined MMF/angiotensin II antagonists treatment almost inhibited the progression of the renal fibrosis. MMF is a new immunosuppressant agent, but it needs to be observed its long-term effects and adverse events. In addition, it is expensive. Leflunomide is a very promising immunosuppressant agent. It inhibits lymphocyte function by inhibiting pyrimidine synthesis and is widely used to treated rheumatoid arthritis with minor side-effects. A study indicated that leflunomide can protect human renal function, which ameliorated chronic allograft nephropathy, but the machanism was unknown. Some studies show that leflunomide can inhibit local proliferation and infiltration of macrophages and lower proliferation of smooth muscle cells. Smooth muscle cells and myofibroblasts have many same characters. So leflunomide may also inhibit proliferation of myofibroblasts. Whether does leflunomide protect renal function by inhibiting proliferation of macrophages and myofibroblasts? So the level of CD68, PCNA, α-SMA, TGF-(1, histologic construction and renal function were studied from the renal cortex of 5/6 nephrectomy rats. In addition, by the medding trials in 5/6 nephrectomy rats, we analyzed the following questions: if leflunomide had a preventive effect on the development of renal fibrosis in 5/6 nephrectomy rats; if the protective effects related to inhibitation of proliferation of macrophages and myofibroblasts. Finally, the new pathway was advanced for preventing and curing human renal fibrosis.Methods:Male Sprague-Dawley rats weighing 200-220g were operated subtotal renal ablation. This surgery was done in a single surgical procedure by subcapsular removal of the right kidney and selective infarction of approximately two-thirds of the left kidney by ligation of two branches of the left renal artery, to leave approximately one sixth of the total kidney tissue mass. Sham operations consisted in laparotomy and manipulation of the kidneys and renal pedicle without destruction of renal tissue. First 40 rats were randomly divided into 2 groups: 30 rats had 5/6 nephrectomy, and 10 rats had a sham operation(Sham group,N=10). Nine rats in the group that had 5/6 nephrectomy died during surgery or in the following 48 hours. Three days after surgery, the surviving rats in the 5/6 nephrectomy group were randomly subdivided in two groups, one of which received LEF (5mg.kg-1.day-1by daily gastric gavage, LEF group, N=11) the other one (Nx group, N=10). Nx and Sham rats were received equal volume water by daily gastric gavage. All rats were allowed free access to food and water. Every 4 weeks after the treatment with leflunomide, the following studies were performed in every group. All of the items such as body weight, the amount of 24 hours urine protein, serum creatinine(Scr), liver enzyme were measured. At the four-week endpoint, five rats from each group were sacrificed. The remaining rats (5 rats from Sham group, 6 rats from LEF group, 5 rats from Nx group) were sacrificed eight weeks after surgery. Kidney weight were determined. The index about kidney hypertrophy (ratio of kidney weight to body weight) corrected by body weight were calculated. Immunohistochemical technique was used to estimate the expression of CD68, proliferating cell antigen (PCNA) and smoothe muscle actin (α-SMA) in renal cortex and the results were analyzed semi-quantitatively with the pathological image analysis system. The level of TGF- (1 in the renal cortex were examined by western blot. The renal tissue of three group rats was observed with light microscope after periodic acid-Schiff (PAS) and Masson staining. The experimental data was demonstrated in mean±standard deviation. The analysis of varianc...
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