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Objectives:A traditional Chinese medicine“Tian Gui Recipe (TGR)”has beenused to effectively treat clomiphene resistant anovulatory disease and obesity,especiallyin polycystic ovary syndrome (PCOS) cases with hyperinsulinemia.The effect of TGRon obesity and anovulation was investigated in androgen-sterilized rats (ASR).Methods:Female SD rats at the age of 9 days were divided into 3 groups.GroupASR (n=15):anovulation was demonstrated at the age of 70 days by vaginal smear inrats while 1.25 mg of testosterone propionate was injected subcutaneously on its 9thneonatal day.Group A+H (n=25).TGR were administered to ASR at the age of 80days for 3 weeks.Rats with regular estrous cycle and ovulation after herbal treatmentwere included in this group.Group C (n=15):normal ovulated rats were recruited.Around the age of 112 days or on proestrous day,all rats were sacrificed under anesthe-sia.Serum leptin,testosterone (T),estrogen (E_2),follicular stimulating hormone(FSH),luteinizing hormone (LH) levels were measured with radioimmunoassay(RIA).Double immunofluorescent staining combined with confocal laser scanning mi-croscope and dual in situ hybridization were carried out on sections through areas of ar-cuate nucleus (ARC) to determine whether estrogen receptor (ER) immunoreactivity(IR) or long form of leptin receptor (OB-R_b) mRNA were expressed in neuropeptide Y(NPY) immuno- and mRNA-containing neurons,and their relationship to proopiome-lanocortin (POMC) mRNA-containing neurons.Immunohistochemistry and in situhybridization were used to observe the levels of ER-,NPY-,gonadotrophin releasinghormone (GnRH)-IR and gene expression levels of NPY,OB-R_b,and POMC in ARC.Meanwhile,the criteria of energy state,including daily food intake,retroperitoneal fatdepot pad and body weight,were measured and evaluated.Values of immunohis-tochemistry and in situ hybridization were expressed in mean optic density (MOD).Results:Seventeen out of the 25 rats (68%) in Group A+H were ovulated afterherbal treatment.ASR characterized with significantly high metabolic rate,energy im-balance and obesity (P<0.01,P<0.01,P<0.01).The mean serum leptin,T,E_2 levelsof ASR were significantly higher than that of Group C and Group A+H (P<0.01,P<0.01),while being contraryto that of FSH,LH levels (P<0.01,P<0.01).ERs wereproved distributing in NPY-containing neurons,OB-R_b coexpressed in NPY-synthesiz-ing neurons,and both NPY- and POMC- mRNA expressing neurons overlapped inARC.Compared to Group C,levels of NPY mRNA and NPY,POMC mRNA expres-sions increased (P<0.01,P<0.01,P<0.01),OB-R_b mRNA,ER and GnRH im-munoreactive expression decreased significantly in the hypothalamus of ASR (P<0.01, P<0.01,P<0.01).These dramatic neuroendocrine changes became normal in ovulatedASR after the herbal treatment (all P>0.05).Conclusion:The elevated peripheral estrogen caused by high testosterone and leptinlevels in ASR may down-regulate their corresponding receptors oriented on hypothalam-ic NPY-containing neurons respectively,therefore challenge the NPY mRNA and NPY,POMC mRNA overexpression.The overexpression of NPY,POMC gene transcriptionand translation stimulate food intake,promote the deposit of adipocyte tissue,and in-hibit GnRH expression and GnRH/FSH,LH release,which contribute to the occurrenceof obesity and anovulation in ASR.“TGR”may reverse these abnormal neuroendocrinecascades by lowering levels of testosterone,estrogen and leptin.
Objectives: A traditional Chinese medicine“Tian Gui Recipe (TGR)has beenused to effectively treat clomiphene resistant anovulatory disease and obesity,especiallyin polycystic ovary syndrome (PCOS) cases with hyperinsulinemia.The effect of TGRon obesity and anovulation was investigated in androgen -sterilized rats (ASR).Methods:Female SD rats at the age of 9 days were divided into 3 groups.GroupASR (n=15):anovulation was demonstrated at the age of 70 days by vaginal smear inrats while 1.25 mg of testosterone propionate Was injected subcutaneously on its 9thneonatal day. Group A+H (n=25).TGR were administered to ASR at the age of 80days for 3 weeks.Rats with regular estrous cycle and ovulation after herbal treatmentwere included in this group.Group C ( n=15):normal ovulated rats were recruited.Around the age of 112 days or on proestrous day,all rats were sacrificed under anesthe-sia.Serum leptin,testosterone (T),estrogen (E_2),follicular stimulating hormone (FSH) , luteinizing hormone (LH) l Evels were measured with radioimmunoassay (RIA). Double immunofluorescent staining combined with confocal laser scanning mi-croscope and dual in situ hybridization werecared out sections through areas of ar-cuate nucleus (ARC) to determine whether estrogen receptor (ER) immunoreactivity ( IR) or long form of leptin receptor (OB-R_b) mRNA were expressed in neuropeptide Y (NPY) immuno- and mRNA-containing neurons, and their relationship to proopiome-lanocortin (POMC) mRNA-containing neurons. Immunohistochemistry and in situhybridization were Used to observe the levels of ER-,NPY-,gonadotrophin releasinghormone (GnRH)-IR and gene expression levels of NPY,OB-R_b,and POMC in ARC.Meanwhile,the criteria of energy state,including daily food intake,reperiperitoneal fatdepot Pad and body weight,we average and evaluated.Values of immunohis-tochemistry and in situ hybridization were expressed in mean vegetation density (MOD).Results:Seventeen out of the 25 rats (68%) in Group A+H were ovulated afterhErbal treatment.ASR characterized with significant high metabolic rate,energy im-balance and obesity (P<0.01,P<0.01,P<0.01).The mean serum leptin,T,E_2 levelsof ASR were significantly higher than that of Group C and Group A+H (P<0.01, P<0.01), while being contrary to that of FSH, LH levels (P<0.01, P<0.01). ERs were proved distributing in NPY-containing neurons, OB-R_b coexpressed in NPY-synthesiz -ing neurons,and both NPY- and POMC- mRNA expressing neurons overlapped inARC.Compared to Group C,levels of NPY mRNA and NPY,POMC mRNA expres-sions increased (P<0.01,P<0.01,P<0.01),OB -R_b mRNA, ER and GnRH im-munoreactive expression decreased significantly in the hypothalamus of ASR (P<0.01, P<0.01, P<0.01). Thesese major neuroendocrine changes became normal in ovulated ASR after the herbal treatment (all P>0.05) .Conclusion:The elevated peripheral estrogen caused by high testosterone and leptinlevels in ASR may down-regulate their corresponding receptors oriented on hypothalam-ic NPY-containing neurons Respectively,therefore challenge the NPY mRNA and NPY,POMC mRNA overexpression.The overexpression of NPY,POMC gene transcriptionand translation stimulate food intake,promote the deposit of adipocyte tissue,and in-hibit GnRH expression and GnRH/FSH,LH release,which contribute To the occurrence of obesity and anovulation in ASR.”TGR"may reverse these abnormal neuroendocrinecascades by lowering levels of testosterone,estrogen and leptin.