Effects of kidney-tonifying Chinese herbs on thymus regen-eration after rapamycin-induced degeneration in mice.

OBJECTIVES: To investigate the effect of a variety of kidney-tonifying Chinese medicines on thymus regeneration after rapamycin (RAPA)-induced acute degeneration in mice. METHODS: Forty-eight 8-week-old male BALB/c mice (19-23 g) were randomly divided into normal control group, model control group, negative control group, positive control group, the fructus of Cnidium monnieri (L.) Cuss. group, the fructus of Psoralea corylifolia (L.) group, the fructus of Rubus chingii Hu group, and the tuber onion seed group, with 6 mice in each group. Except for the normal control group, mice in the other groups received intraperitoneal injections of rapamycin (1 mg·kg－1·d－1) for 5 consecutive days followed by 14 h of starvation to induce acute thymus degeneration. After successful modeling, in treatment groups ethanol extract of the fructus of Cnidium monnieri (L.) Cuss. (0.78 g·kg－1·d－1), fructus of Psoralea corylifolia (L.) (0.39 g·kg－1·d－1), fructus of Rubus chingii Hu (0.78 g·kg－1·d－1) and the tuber onion seed(0.39 g·kg－1·d－1) intraperi-toneally injected once a day for 5 days; while the negative control group was given with equal volume of normal saline, and the positive control group was given with metformin (300 mg·kg－1·d－1) instead. The grip strength was measured with a grip tester 2 h after the last administration. The pathological changes of thymus were observed by hematoxylin-eosin (HE) staining. The structure and distribution of thymic epithelial cells were observed by multiple immunofluorescence method. The proportion of T cell subsets in thymus and peripheral blood was analyzed by flow cytometry. The level of T cell receptor excision circles (TREC) in the genomic DNA of mouse spleen mononuclear cells was detected by quantitative PCR for evaluation of thymic output function. The expression levels of thymus aging-related and function-related factors in thymus tissue were detected by quantitative reverse transcription PCR. The expression levels of key factors cyclin-dependent kinase inhibitor 1A (p21) and tumor protein 53 (p53) were verified by immunohistochemistry. RESULTS: After RAPA treatment, the mouse limb grip strength decreased (P<0.01), the level of TREC in spleen was increased, the proportion of CD4+CD8+ decreased (P<0.05), and the proportion of mature CD4 single positive T cells (CD3+TCR β+CD4+CD8－) was decreased. Compared with the negative control group, the limb grip strength of mice in the fructus of Psoralea corylifolia (L.) group, the fructus of Rubus chingii Hu group and the tuber onion seed group was significantly enhanced (all P<0.05), and the level of TREC in spleen of the mice in each administration group was reduced. Among Chinese herb medicine-treatment groups, the recovery of thymus function and tissue structure in the tuber onion seed group was most obvious. Further study showed that compared with the negative control group, the proportion of CD4 single positive cells (CD3+TCR β+CD4+CD8－) in the thymus of the tuber onion seed group was significantly increased (P<0.01), and the proportion of CD28+ T cell and CD8+CD28+ T cell in peripheral blood was significantly increased (all P<0.01), and the mRNA levels of IL-1α, IL-6, p21 and p53 in thymocytes were decreased. The results of immunohistochemistry further confirmed the decrease of p21 and p53 protein expression. In normal mice treated with tuber onion seed, the grip strength of the limbs of mice was enhanced (P<0.01), however, thymus output and change the distribution of T cell subsets were inhibited, and there was no significant effect on thymus weight and the expression of SIRT1, CXCL2 and p21. CONCLUSIONS: The tuber onion seed and other kidney-tonifying traditional Chinese medicines can accelerate the regeneration process of mouse thymus after acute degeneration induced by rapamycin in mice, and the tuber onion seed exhibits the most pronounced therapeutic effect.