Tanshinone IIA attenuates estradiol-induced polycystic ovarian syndrome in mice by ameliorating FSHR expression in the ovary.

Tanshinone IIA (TSIIA) is a major component of Salvia miltiorrhiza, a Chinese herb that exhibits a therapeutic effect on polycystic ovary syndrome (PCOS). The present study replicated PCOS via the neonatal treatment of estradiol in mice. Estrous cycles, body and ovarian weight, serum levels of testosterone and estradiol were determined. Histological examination of ovaries was performed. The mRNA and protein levels of aromatase luteinizing hormone receptor and follicle-stimulating hormone (FSHR) in ovaries and granule cells were assayed by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. TSIIA was revealed to reverse all disorders induced by estradiol treatment, including prolonged estrous cycles, increased body and ovarian weight, increased atretic cyst-like follicles and decreased corpus luteum, large antral follicles and preovulatory follicles. These improvements in PCOS as a result of TSIIA treatment are likely due to the revised testosterone/estradiol balance, as TSIIA reversed the decrease in aromatase mRNA, the enzyme that converts androgen to estrogen. As the expression of aromatase is regulated by the FSH pathway, TSIIA-mediated elevation in FSHR expression may lead to the upregulation of aromatase. Therefore, TSIIA revises the balance of androgen and estrogen by rescuing the reduced expression of FSHR and aromatase, thus attenuating murine PCOS. The current study aimed to further the application of natural drugs in the treatment of PCOS to confront the side effects of hormone drugs and expand the use of TSIIA.

Increased PIT1 and PIT2 Expression in Streptozotocin (STZ)-induced Diabetic Mice Contributes to Uptake of iAs(V).

OBJECTIVE: This study aimed to investigate the susceptibility of mice with streptozotocin(STZ)-induced diabetes mellitus (TIDM) to the uptake of pentavalent inorganic arsenic (iAsV) and the possible molecular mechanism. METHODS: TIDM was induced in mice by STZ. TIDM and normal mice were treated with 15.0 mg/kg Na2HAsO4·12H2O by intragastric administration. Then, the concentrations of arsenic in various tissues were measured by atomic fluorescence spectrometry. The gene expression levels of Pit1 and Pit2 were quantified by real-time RT-PCR, and their protein levels were detected by Western blotting in mouse heart, kidney, and liver tissues. RESULTS: The concentrations of arsenic in STZ-induced TIDM mouse tissues were higher at 2 h after intragastric administration of Na2HAsO4·12H2O. Compared with the levels in normal mice, PIT1 and PIT2, which play a role in the uptake of iAsV, were upregulated in the livers and hearts of TIDM mice. PIT1 but not PIT2 was higher in TIDM mouse kidneys. The upregulation of Pit1 and Pit2 expression could be reversed by insulin treatment. CONCLUSION: The increased uptake of iAsV in TIDM mouse tissues may be associated with increased PIT1 and/or PIT2 expression.

[Preparation antibodies against recombinant bovine IFN-gamma and development of sandwich ELISA for bovine IFN-gamma detection].

This study was aimed to establish ELISA for recombinant bovine IFN-gamma (BovIFN-gamma) detection and provide a new method for diagnosis of pathogenic infection. The total RNA was isolated from peripheral blood leucocytes cultured with PHA mitogen stimulation. Then bovine IFN-gamma (BovIFN-gamma) gene cDNA was amplified by RT-PCR and cloned into pET28a to obtain the expression plasmid designated as pETBovIFN-gamma. The pETBovlFN-gamma was further transformed into competent E. coli BL21 cells and a 18kD His-tagged protein as expected was expressed after IPTG induction. By using purified recombinant BovIFN-gamma as antigen and lymphocyte-hybridoma technique, four hybridoma cell lines which stably secreted monoclonal antibodies against rBovIFN-gamma were generated, designated as A7, A10, G6, and G10. The immunoglobin subset was identified as IgG1 . Western-blotting analysis and ELISA demonstrated that the monoclonal antibodies secreted by all the four hybridoma cell lines could react specifically to the recombinant BovIFN-gamma, but not irrelative proteins such as Ag85B, ESAT-6-CFP-10 and GM-CSF, suggesting that the four hybridoma cell lines were rBovIFN-gamma specific monoclonal antibodies. A sandwich ELISA was established by using A10 secreted monoclonal antibody and rabbit polyclonal antibodies against BovIFN-gamma, HRP labeled goat anti-rabbit IgG. The results indicated that the sensitivity was 2ng/mL. This sandwich ELISA to detect BovIFN-gamma paved the way to develop a sensitive method for specific infection detection such as bovine tuberculosis diagnosis.

[Blood leptin, orexins and NPY levels and their relations in obese children].

OBJECTIVE: To investigate the concentration levels of leptin, orexins and neuropeptide Y (NPY) in the blood of obese children, and to analysed the relationship between these substances. METHODS: RIA methods were used to measure the concentrations of leptin, orexinA, orexinB, and NPY in the blood of 98 obese children [BMI: male (29.24 +/- 1.87) kg/mZ, female (28.12 +/- 2.30) kg/m2] and in 104 normal children [BMI: male (20.49 +/- 1.95) kg/m2, female (19.59 +/- 1.51) kg/m2] as the control group. RESULTS: The leptin concentrations in obese children [male (26.00 +/- 14.66) ng/mL; female (33.59 +/- 14.63) ng/mL] were higher than those in the control group [male (6.65 +/- 44.49) ng/mL; female [10.48 +/- 5.52) ng/mL P < 0.013]. The concentrations of plasma orexinA in obes children [male (3.23 +/- 1.86) pg/mL; female (3.38 +/- 1.80) pg/mL] were lower than those in the control group [male (4.52 +/- 1.52) pg/mL; female (4.71 +/- 1.53) pg/mL P < 0.05]; Negative correlations between leptin and NPY were noted in the obesity group (r = -0.302) and the control group (r = -0.310, P < 0.01), while the slopes in the two groups were different (control group -2.969; obese group -0.809). A positive correlation between NPY and orexinA was noted (r = 0.207, P < 0.05). The fluctuation range of orexinA in obese children was markedly narrowed when compared with that in the control group. CONCLUSION: The concentration level of peripheral orexinA and leptin in the obese and non-obese children change inversely. The obesity in children correlates with the concentrations of orexinA, leptin, NPY as well as with their interactions.