| 1.ObjectiveThrough the "measured by time" research method,by observing and comparing the seasonal variation of the content of endocrine markers in the hypothalamus-pituitary-adrenal axis(HPA axis)between the normal group and the model group(Pinealectomy)to further explore the biological mechanism of ' kidney corresponding with winter and governing storage ',and to explore the innate character and significance of the Kidney in TCM and the role of pineal gland in the process of seasonal variations.At the same time,the subject hope to provide experimental basis for clinical treatment of seasonal visceral diseases related to seasonal changes.2.Methods2.1 Animal ExperimentThe healthy male Syrian golden hamster(120g±20g)was provided by Beijing Weitong lihua Experimental Animal Technology Co.,Ltd.They were purchased in the first five weeks of the spring equinox,summer solstice,equinox,and winter solstice,and randomly divided into normal group and model group with 8 rats in each group.The model group performed pinealectomy.The normal group and the model group were reared to the vernal equinox,summer solstice,autumnal equinox,and winter solstice on the same feeding conditions,moved to the dark room at 08:00 in the evening,and began to collect materials at 09:00 in the evening with 5w red light.The serum and hypothalamus of golden hamsters in the normal and model groups were removed.Pituitary glands and adrenal glands were used to measure the levels of endocrine markers by Enzyme linked immunosorbent assay(ELISA).2.2 Indicator detection2.2.1 Effects of seasonal changes on endocrine secretion of hypothalamus-pituitary-adrcnal Axis(1)Four seasons changes in serum corticotropin-releasing hormone(CRH),adrenocorticotropic hormone(ACTH),corticosterone(CORT),aldosterone(ALD),epinephrine(E),norepinephrine(NE),adrenal medullary hormone(ADM)effects of hormone secretion;(2)Effects of seasonal changes on corticotropin-releasing hormone(CRH)secretion in hypothalamus(3)Effect of seasonal changes on secretion of adrenocorticotropic hormone(ACTH)in pituitary(4)Effects of seasonal changes on secretion of adrenaline(E),norepinephrine(NE)and adrenal medullary hormone(ADM)in adrenal2.2.2Pineal regulation of the hypothalamus-pituitary-adrenal axis endocrine(1)The seasonal effects of melatonin(MT)and melatonin receptor(MR)on the hypothalamic-pituitary-adrenal axis endocrine.(2)The seasonal effects of pinealectomy on the hypothalamic-pituitary-adrenal axis endocrine.2.3 Statistical methodsAll data were expressed as mean±standard deviation(x±s)and analyzed using SPSS 20.0 statistical software;One-way analysis of variance or non-parametric test of K independent samples was used between groups;In the case of pairwise comparison,an independent sample t-test or non-parametric test of 2 independent samples is used.If P<0.05,it is statistically significant.3.Results3.1 Seasonal changes of cell signal transducing substance content in pineal gland and HPA glandular axis tissue3.1.1 Seasonal changes in cell signal transducing substance content in pineal tissue.? The seasonal changes of MT content in pineal gland tissues of normal golden hamsters were summer>autumn>winter>spring,and there were significant differences between summer and spring,autumn and winter(P<0.05).? The seasonal changes of MR contents in the pineal gland tissues of normal golden hamsters were spring>summer>autumn>winter.3.1.2 Seasonal changes in cellular signal transducing substance content in HPA axis.3.1.2.1 Seasonal changes of cell signal transducing substance content in hypothalamus.?The seasonal changes in MT content in the hypothalamus of the normal golden hamster were winter>summer>autumn>spring,and there were significant differences between the spring and winter(P<0.05);The seasonal changes of MT content in the hypothalamus tissue of the model group were significantly different in summer>autumn>winter>spring,and in spring and summer(P<0.05).? The seasonal changes of MR content in hypothalamus of the normal golden hamster were summer>winter>autumn>spring,and there were significant differences in the three seasons of spring,summer and winter(P<0.05);The changes of MR contents in hypothalamus tissue in the model group during the four seasons winter>autumn>summer>spring.? The four-year trend of CRH content in hypothalamus of normal golden hamster was summer>autumn>winter>spring,and there were significant differences between the four seasons(P<0.001).The seasonal changes of CRH content in the hypothalamus of the model group were winter>summer>autumn>spring,and there were differences between the four seasons(P<0.001).3.1.2.2 Seasonal changes of cell signal transducing substance content in pituitary.? The seasonal change trend of MT content in the pituitary tissues of normal golden hamsters was summer>spring>winter>autumn,and there were significant differences between spring,summer and winter(P<0.01).The seasonal change trend of MT content in the pituitary tissue of the model group was summer>autumn>spring>winter,and there was a very significant difference between the four seasons(P<0.001).?The change of MR contents in the pituitary tissue of the golden hamster in the normal group was summer>autumn>spring>winter;In the model group,the seasonal changes of the MR contents in the pituitary tissue were winter>summer>autumn>spring.? The seasonal changes of ATCH in the pituitary tissues of the normal golden hamster were summer>spring>winter>autumn,and significant differences were observed in the summer,autumn and winter(P<0.001).The seasonal changes of ATCH content in the pituitary tissue of the model group were autumn>summer>winter>spring.3.1.2.3 Seasonal changes of cell signal transducing substance content in adrenal gland.? The seasonal changes of MT content in the adrenal gland tissue of normal golden hamsters were winter>summer>autumn>spring,and there were significant differences between spring and winter(P<0.05).The seasonal changes of MT content in the adrenal glands of the model group were summer>spring>autumn>winter.? The seasonal changes of MR content in the adrenal gland tissue of normal golden hamsters were spring>winter>autumn>summer,and there were significant differences between the four seasons(P<0.05).The seasonal change trend of MR content in the adrenal tissue of the model group was autumn>summer>winter>spring,and there were differences between spring,summer and winter(P<0.001).? The seasonal change trend of ALD content in the adrenal gland tissue of the normal golden hamster was summer>a1utumn>winter>spring,and there were significant differences between the four seasons(P<0.001);The seasonal changes of ALD content in the adrenal glands of the model group were significantly different in summer>autumn>winter>spring,and significantly different between spring,summer and winter(P<0.001).? The CORT content in the adrenal glands of the normal golden hamster showed a seasonal change trend of summer>autumn>winter>spring,and there was a statistically significant difference between spring and summer(P<0.05).The four-year trend of CORT content in adrenal tissue of golden hamster in model group was summer>autumn>winter>spring,there was a statistically significant difference between spring,summer and winter(P<0.05).? The seasonal changes of E content in the adrenal gland tissue of the normal golden hamster were summer>autumn>spring>winter;The seasonal changes of E content in the adrenal glands of the model group were summer>spring>winter>autumn.?The seasonal changes of NE content in the adrenal gland tissue of the golden hamster in the normal group were summer>autumn>spring>winter;The seasonal changes of NE content in the adrenal glands of the model group were autumn>summer>winter>spring.?The four-year trend of ADM content in adrenal gland tissue of golden hamsters in the normal group was autumn>spring>winter>summer.The four-year trend of ADM content in the adrenal glands of the model group was summer>autumn>winter>spring,and there were differences between spring and summer(P<0.05).3.1.2.4 Seasonal changes of cell signal transducing substance content in serum.?The seasonal changes of MT content in serum of golden hamsters in the normal group were significantly different in winter>autumn>spring>summer,summer and winter(P<0.05);The seasonal variation trend of MT content in the serum of the model group was winter>autumn>spring>summer,and there were significant differences between the spring,autumn and winter(P<0.05).? The seasonal changes of CRH in serum of golden hamsters in the normal group were spring>autumn>winter>summer,and there were significant differences between the four seasons(P<0.05).The CRH content in the serum of the model group varied from spring to winter and autumn to summer in the four seasons,and there were significant differences between the four seasons(P<0.05).? The changes of ACTH content in golden hamster serum in normal group were spring>winter>summer>autumn,and statistically significant in spring,summer and autumn(P<0.05).The seasonal changes of serum ACTH levels in the model group were spring>autumn>winter>summer,and statistically significant in spring and summer(P<0.05).The seasonal changes of serum ACTH levels in the model group were spring>autrumn>winter>summer,and statistically significant in spring and summer(P<0.05).? The four-year trend of CORT content in the serum of normal golden hamsters was autumn>winter>summer>spring.The CORT content in the serum of the model group had a seasonal change trend of spring>autumn>summer>winter.?The seasonal changes of serum E content in golden hamsters in the normal group were winter>summer>spring>autumn,and there were significant differences between autumn and winter(P<0.01).The seasonal changes of serum E content in golden hamsters of model group were spring>autumn>summer>winter,and winter and winter seasons were different(P<0.05).?The seasonal changes of NE content in golden hamster serum in the normal group were spring>winter>summer>autumn,and there were differences between spring and autumn(P<0.01).The seasonal variation trend of NE content in serum of model group was spring>winter>summer>autumn,and there was a difference between spring and autumn(P<0.05).?The four-year trend of ADM content in serum of normal golden hamsters was spring>autumn>summer>winter;The trend of serum ADM content in the model group during the four seasons was summer>spring>autumn>winter,and there were significant differences between spring,summer and winter(P<0.05).4.Conclusion(1)"kidneys governing storage" are closely related to the endocrine regulation of the hypothalamus-pituitary-adrenal axis.(2)The seasonal regulation of hypothalamic-pituitary-adrenal axis hormones by pineal melatonin is an important neuroendocrine mechanism of "kidney corresponding with the winter,kidneys governing storage". |
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