| Objective: Observe the changes of gynecological Excess-cold syndrome model rats serum, uterine and ovarian tissue in HIF-1α, ET-1, VEGF and its relationship with reproductive endocrine. Study on the relationship between the occurrence of gynecological Excess-cold syndrome and hypoxia; Using Warming Meridian intervention to investigate the effect of the Modified decoction for Warming Meridian on reproductive function of gynecological cold syndrome model rats.Methods: Healthy clean grade female SD 60 rats, February age, weight 200±20g, adaptive feeding for 1 weeks, female SD rats were randomly divided into normal group, model group and treatment group with 20 rats in each group, according to the random numbertable method. Rat model of gynecology excess-cold syndrome was made by ice water immersion. From the first day, those rats of the treatment group were intragastric administration in dose of 3.51g/200g/day seperatively, qd for 2 weeks. The other two groups were given the same voluminal distilled water. After 2 weeks, weighed and sacrificed the rats by decapitation during anestrum. Get blood by decapitation, centrifugalization, separating serum, placed them respectively. Dissected the level of E2, P, T, FSH and LH by means of radio immunoassay. Dissected the serum level of HIF-1α, ET-1, VEGF by the means of ABC-ELISA. Dissected and got rats uterus and ovaries quickly. Fixed the right uterus and ovaries in 4% Polyoxymethylene, dehydrated with alcohol, transparented with dimethyl benzene, embedding, slice in 5μm, HE dyeing, then observed the construre of ovarian histomorpholory under light microscope. Using immunohistochemical method to observe the uterine tissue, expression of ovarian tissue HIF-1 alpha, ET-1, VEGF protein. The other uterus and ovaries were placed in labeled plastic freezer tube, then dissected the HIF-1α m RNA、VEGF m RNA by the means of RT-PCR.Results:1 General state of health: After being frozen, the model rats appeared such symptoms as chilling, im in clothing hari, preference for herd, twisting and less-movement, slow reaction, with dark purple in claws and tail and rotten stool. In treatment group, the above-mentioned symptoms improved to some different degree.2 Morphlogical observation of the uterus and ovaryuterus: The normal group rats uterine tissue of normal morphology, Intima thicker, a large number of glands in the lamina propria, Vascular rich, pipe diameter thinker. The endometrium of model group rats were significantly thinner, the number of gland less, lumen of blood vessel narrower. In treatment group, the above-mentioned symptoms improved to some different degree.Ovary: The normal group rats, the boundary between the cortex and medulla of clear, number of follicles in cortex, the dominant follicle is visible, Ovarian vascular rich and vascular morphological rules, While in model group, there can be seen less grades of follicles and the mature follicle rare, lumen of blood vessel narrower. In treatment group, the above-mentioned symptoms improved to some different degree.3 Change of contents of serum E2, P, T, FSH and LHCompared with normal group, in model group the serum E2, P, T, FSH and LH levels decreased, differences were significant(P<0.01). Compared with model group, in treatment group the serum E2, P, T, FSH and LH levels rose, differences were significant(P<0.05). Compared with normal group, in treatment group, the serum E2, P, T, FSH and LH levels decreased slightly, differences were not significant(P>0.05).4 Changes of serum HIF-1α, ET-1, VEGFCompared with normal group, in model group the content of HIF-1α is higher, differences were significant(P<0.01). Compared with model group, in treatment group the content of HIF-1α is lower, differences were significant(P<0.01). Compared with normal group, in treatment group the content of HIF-1α is lower, differences were significant(P<0.05)Compared with normal group, in model group the content of ET is higher, differences were significant(P < 0.01). Compared with model group, in treatment group the content of ET is lower, differences were significant(P<0.05). Compared with normal group, in treatment group the content of ET is lower, differences were significant(P<0.01)Compared with normal group, in model group the content of VEGF is higher, differences were not significant(P>0.05). Compared with model group, in treatment group the content of VEGF is lower, differences were not significant(P>0.05). Compared with normal group, in treatment group the content of VEGF is lower slightly, differences were not significant(P>0.05).5 Expression of HIF-1α、ET、VEGF proteins in uterus and ovariesUterus:Compared with normal group, in model group, the expression of HIF-1α、ET、VEGF proteins in uterus tissue increased, differences were significant(P<0.01). Compared with model group, in treatment group, the expression of HIF-1α 、 ET 、 VEGF proteins reduced, differences were significant(P<0.01).Ovary:Compared with normal group, in model group the expression of HIF-1α、ET proteins in uterus tissue increased, differences were significant(P<0.01). Compared with model group, in treatment group the expression of HIF-1α 、 ET proteins reduced, differences were significant(P < 0.01). Compared with normal group, in model group the expression of VEGF proteins in uterus tissue increased, differences were significant(P<0.01). Compared with model group, in treatment group the expression of VEGF proteins reduced, differences were not significant(P>0.05).6 Expression of HIF-1α m RNA、VEGF m RNA proteins in uterus and ovariesUterus: Compared with normal group, in model group the expression of HIF-1α m RNA increased, differences were significant(P<0.01). Compared with model group, in treatment group the expression of HIF-1α m RNA in ovarian tissue reduced, differences were significant(P<0.01). Compared with normal group, in model group the expression of VEGF m RNA increased, differences were significant(P < 0.05). Compared with model group in treatment group the expression of VEGF m RNA in Uterus tissue reduced, differences were not significant(P>0.05).Ovary: Compared with normal group, in model group the expression of HIF-1α m RNA、VEGF m RNA increased, differences were significant(P<0.01). Compared with model group, in treatment group the expression of HIF-1α m RNA、VEGF m RNA in ovarian tissue reduced, differences were significant(P<0.01).Conclusions:1 Gynecological Excess-cold syndrome model rats have the reproductive endocrine disorders, and uterus, ovarian morphological abnormalities, ovarian function decline.2 The uterine and ovarian tissue of gynecological Excess-cold syndrome model rats in a state of lack of hypoxia.3 Uterus, ovarian of the Gynecological Excess-cold syndrome model rats appeared morphological abnormalities, ovarian function declined and reproductive endocrine disorders, they are closely related to the local state of lack of oxgen.4 Warming meridian can improve the uterine and ovarian tissue of gynecological Excess-cold syndrome model rats oxygen deficiency state, corrected the tissue morphological abnormalities and recover the normal hemodynamics, maintain uterine and ovarian normal reproductive function... |
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