[Protective effect of radix sophorae flavescentis mixture on intestinal mucosa in mice infected with Cryptosporidium parvum].

OBJECTIVE: To investigate the protective effect of radix sophorae flavescentis (RSF) mixture on intestinal mucosa in mice infected with Cryptosporidium parvum. METHODS: Thirty BALB/c male mice were randomly divided into control group, infection group and RSF mixture treatment group. Mice of the posterior two groups were inoculated intragastrically with 1 x 10(5) C. parvum oocysts, immunosuppressed with dexamethasone (5 microg/ml) and gentamycin sulfate (40 microg/ml) in drinking water. At the 8th day post-infection, mice in RSF mixture treatment group were treated with 0.2 ml dose of RSF mixture twice a week (three-day intervals) for three weeks. The mice in infection group and RSF mixture treatment group were monitored for oocyst shedding in fecal pellets every two days after treatment. At 28 days after infection, experimental mice were sacrificed, jejunal tissue was removed for preparation of paraffin-embedded sections. The changes of CD3+, CD4+, CD8+ T lymphocytes and IgA plasmocytes in intestinal mucosa were determined by immunohistochemistry. In addition, jejunum of infected mice and treated mice were collected, and ultrastructural changes were observed under electron microscopy. RESULTS: Compared with infection group, the level of oocyst shedding was obviously lower and the time of the oocyst discharging was significantly shorter in RSF mixture treatment group. The proportion of CD3+, CD4+ T lymphocyte and CD4+/CD8+ T cell ratio in infection group (49.7% +/- 2.4%, 25.7% +/- 2.2%, 1.1 +/- 0.3) were significantly lower than that of treatment group (62.4% +/- 1.4%, 37.5% +/- 3.1%, 1.5 +/- 0.3) and control group (66.5% +/- 1.9%, 40.1% +/- 1.8%, 1.5 +/- 0.2) (P < 0.01). CD8+ T lymphocytes showed no significant difference in each group (P > 0.05). The number of IgA plasmocytes in treatment group (52.7 +/- 3.5) was significantly higher than that of control group (8.3 +/- 2.3) and infection group (33.7 +/- 2.6) (P < 0.01). After administration for three weeks, the damaged C. parvum parasites were seldom seen in mouse jejunum, and lysosomes appeared in large number, RSF mixture treatment improved mitochondrial structure and repaired microvilli. In infection group, mitochondria ridges were significantly broken and microvilli surrounding C. parvum oocysts were shed, resulting in the appearance of crater-like lesions on the surface, the oocyst wall and host cell membrane fused together. CONCLUSION: RSF mixture is effective against Cryptosporidium parvum. The damage of intestinal mucosa in infected mice can be repaired after treatment.

[Effect of Toxoplasma gondii infection on the embryonic neural stem cells in rats].

OBJECTIVE: To investigate the effect of Toxoplasma gondii infection on the proliferation, differentiation and migration of the embryonic neural stem cells (NSCs) in early pregnancy of rat. METHODS: Twelve pregnant Sprague-Dawley rats were randomly divided into control and infection groups. Rats in the infection group were each inoculated intraperitoneally with 1 x 10(5) T. gondii RH strain tachyzoites at day 1 (E1 day). Same amount of physiological saline was intraperitoneally injected for rats in control group. At E5 day, blood samples were taken from caudal vein and Giemsa staining of blood cells was performed to find T. gondii. At E9, E10 and E11 day, two rats in each group per time point were sacrificed and reverse transcription PCR (RT-PCR) was performed to detect B1 gene expression of T. gondii in amniotic fluid to confirm T. gondii infection. NSCs were cultured in vitro. The proliferation level was detected by methyl thiazolyl tetrazolium (MTT) assay. After differentiation culture of NSCs, the immunofluorescence assay was conducted to detect the expression of nestin, microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP) to calculate the ratio of NSCs which differentiated to neurons and astrocytes. The embryonic nerve tissues at E9, E10 and E11 day in each group were taken to make frozen sections. The immunofluorescence assay was carried out to detect the expression of neuronal cell adhesion molecule (NCAM) in the nerve tissues at different developmental stages. RESULTS: Both the results of blood smears and RT-PCR confirmed that the pregnant rats and embryos were all infected by T. gondii in infection group. The morphology of the cultured NSCs under microscope was consistent with the characteristics of the normal NSCs. In addition, the NSC biomarker nestin protein was stained positive. The MTT assay showed that the proliferation level was lower in infection group than that of the control, and statistical differences were found between the two groups at day 3 and 4 after passages (P < 0.05). The immunofluorescence staining of MAP2 and GFAP showed that the percentage of neuron differentiation was 15.15% (55/363) in control group and 8.73% (31/355) in infection group, respectively, with a statistical difference (P < 0.05), and the percentage of astrocyte differentiation was 53.35% (199/374) and 67.48% 249/369), respectively (P > 0.05). In both groups, NCAM protein was found expressed at E9, E10 and E11 day in embryo nerve tissues. The fluorescence became stronger with time. The expression level in control group was significantly higher than that in infection group (P < 0.01). CONCLUSION: T. gondii infection at early gestation may inhibit the proliferation, differentiation and migration of neural stem cells in rats.

[Effect of radix sophorae flavescentis (RSF) mixture on mast cells in jejunal mucosa of mice infected by Cryptosporidium].

OBJECTIVE: To elucidate the role of mast cells (MC) activation in the jejunal mucous membrane in the pathogenesis of cryptosporidiosis (CPS) and explore the mechanism of prevention and treatment of radix sophorae flavescetis(RSF) mixture on CPS. METHODS: A total of 30 healthy male BALB/c mice were randomly divided into a normal control group, CPS model control group and RSF mixture experimental group. The mice of CPS model were inoculated intragastrically with 1 x 10(5) Cryptosporidium oocyst (CSO). The mice in the RSF mixture experimental group were treated with inoculation of RSF mixture (0.2 ml doses) twice one week for three weeks continuously after CPS models were established. Pathological changes of the jejunal mucosa membrane were observed by a light microscope. The MCs were stained by toluidine blue, the number of mast cells was recorded and the changes of degranulation were observed. RESULTS: The HE staining showed inflammatory pathological changes in the jejunal mucosa membrane of the CPS model control group. After three-week treatment of RSF mixture, the small intestine epithelium was integrated on the whole. The toluidine blue stain showed the number of mast cell in submucosa and muscular layer of the jejunal mucous membrane increased significantly in the model control group (12.80 +/- 0.84) compared with those of the normal control group (1.60 +/- 0.89) (P < 0.01) and an obvious degranulation was seen in the CPS model control group. The number of mast cells of the mice in the RSF mixture experimental group decreased significantly (P < 0.01) and the number (2.00 +/- 0.71) and morphous were closed to the normal after administration for three weeks. CONCLUSIONS: MC activation is involved in the intestinal inflammatory response caused by Cryptosporidium. RSF mixture could decline the number of MC, inhibit the activation and degranulation of MC in the jejunal mucosa membrane of CPS mice to reduce inflammation and repair the damaged intestinal mucosa, which may realize the purpose of treatment of CPS.