| Objectives1 Through a systematic review of domestic and international literature on co-morbid mood disorders in polycystic ovary syndrome(PCOS),advances in Chinese medicine treatment of PCOS with kidney deficiency and liver depression,and the correlation between PCOS and ovarian granulosa cell apoptosis,aimed to better understand the pathogenesis and treatment of PCOS and provide a basis for the selection and updating of treatment protocols.2 To observe the effects of the tutor’s formula "Bushen Jieyu Tiaochong Formula" on ovulation,clinical symptoms and auxiliary examination indexes,as well as on emotional symptoms and quality of life of the PCOS patients with kidney deficiency and liver depression,in order to investigate the intervention effects of Bushen Jieyu Tiaochong Formula on both clinical symptoms and psychological status of the PCOS patients with kidney deficiency and liver depression.3 A rat model of PCOS combined with chronic mild unpredictable stress(CUMS)was established using the combination of letrozole and chronic mild unpredictable stress(CUMS)method.The model was used as the research object,and the Western medicine Diane-35 was used as the positive control.The therapeutic effect of Bushen Jieyu Tiaochong Formula on the model rats was confirmed by evaluating the pathological status of PCOS and chronic stress;and based on the expression levels of SIRT1/P53 signaling pathway related proteins in ovarian/hippocampal tissue and the results of ovarian apoptosis related detection,explore the target and mechanism of Bushen Jieyu Tiaochong Formula in treating PCOS combined with chronic psychological stress.Methods1 Clinical researchUsing a self-control study method before and after treatment,46 patients with PCOS who met the diagnosis of kidney deficiency and liver depression were selected and given 3 consecutive courses of treatment(1 menstrual cycle as 1 course)with the Bushen Jieyu Tiaochong Formula,to observe the improvement of ovulation,and to observe the changes of clinical symptoms such as TCM symptoms score,BMI,hirsutism and acne before and after treatment,to detect the changes of FSH,LH,LH/FSH and FT,changes in sex hormone levels,the ovarian volume under ultrasound and other examination indexes,and assessed changes in emotional symptoms and quality of life such as Symptom Check list 90(SCL-90)scores and PCOS health-related quality of life questionnaire revised(MPCOSQ)scores.2 Experimental researchBy comparing the PCOS model constructed by letrozole alone,the chronic psychological stress model constructed by CUMS alone,and the PCOS combined with chronic psychological stress model constructed by letrozole combined with CUMS,the differences in the pathological states of PCOS such as ovarian tissue morphology,serum sex hormone levels,and chronic stress pathology such as rat behavior,hippocampal tissue morphology,and blood list amine neurotransmitter levels were evaluated.To evaluate the morphological function of the ovary/hippocampus in the rat model of PCOS combined with chronic psychological stress established by letrozole combined with CUMS method.An efficacy study was conducted by using the animal model of PCOS combined with chronic psychological stress.In addition to the blank group,the model group,the Bushen Jieyu Tiaochong Formula group and the western medicine group were treated with letrozole combined with CUMS to construct a rat model of PCOS combined with chronic psychological stress,and after the model was successfully established,the rats were treated with corresponding drugs.After treatment,the pathological status of PCOS was assessed by ovarian histology,serum sex hormone levels such as FSH,LH,LH/FSH and FT,and the pathological status of chronic stress in rats was assessed by the behavior of rats such as the open field test and forced swimming test,hippocampal histology,and blood levels of amine neurotransmitters such as 5-HT,DA and NE.An animal model of PCOS combined with chronic psychological stress was applied to investigate the mechanism.The expression of SIRT1,P53 and acetylated P53(Acetyl-P53)proteins in the ovaries and hippocampus of rats in the blank group,model group,Chinese herbal medicine Bushen Jieyu Tiaochong Formula group and western medicine Diane-35 group were detected by Western blot method.The apoptosis of ovarian granulosa cells was detected by applying TUNEL staining and Western blot to detect the expression of cysteine proteinase-3(Caspase-3)protein.Results1 Clinical research(1)Research 1①Ovulation rate of cycle:A total of 124 menstrual cycles were recorded in 46 patients during the treatment process,resulting in a total of 72 ovulation cycles,with a ovulation rate of 58.06%.The first course of treatment consisted of 46 menstrual cycles,resulting in 17 ovulation cycles,with an ovulation rate of 36.96%;The second course of treatment consisted of 42 menstrual cycles,resulting in 28 ovulation cycles,with an ovulation rate of 66.67%;After treatment,the third course of treatment consisted of 36 menstrual cycles,resulting in 27 ovulation cycles with an ovulation rate of 75%.②Traditional Chinese Medicine Syndrome Score:After treatment,the patient’s menstrual cycle score improved significantly compared to before(P<0.05).After treatment,the menstrual volume,color,and quality scores improved,but the difference was not statistically significant(P>0.05);Among various traditional Chinese medicine syndromes of kidney deficiency and liver depression,the scores of lumbosacral pain,decreased libido,irritability,or emotional depression after treatment showed significant improvement compared to before treatment,and the difference before and after treatment was statistically significant(P<0.05);After-treatment,the scores of shin soreness,knee tenderness,heel pain,dizziness,tinnitus,frequent nocturia,breast distension,chest and rib discomfort,or belching and sighing improved compared to before treatment,but the difference was not statistically significant(P>0.05).③PCOS related indicators:After treatment,the patient’s BMI significantly decreased(P<0.05),and the acne score significantly decreased(P<0.05);After treatment,the FT level significantly decreased compared to before treatment(P<0.05),while the LH and LH/FSH levels decreased,but the difference was not statistically significant(P>0.05);Although the ovarian volume of patients decreased to a certain extent after treatment,the difference was not statistically significant(P>0.05).(2)Research 2①SCL-90 scale scores:after 3 cycles of treatment,the total scale score,total mean score,and total number of positive items of the SCL-90 scale were significantly lower than before treatment,and the difference was statistically significant(P<0.05);as for the SCL-90 scale factors,all factor scores decreased compared with before treatment among which the factor scores of somatization,obsessive-compulsive symptoms,depression,hostility,paranoia psychoticism,and state of sleeping and eating were significantly lower than before treatment(P<0.05),and the factor scores of interpersonal sensitivity and anxiety decreased,but did not reach statistical significance(P>0.05).②MPCOSQ scale scores:after 3 cycles of treatment,the scores of total scores,acne problems,weight problems,emotional problems,menstrual concomitant problems and menstrual predictive problems on the MPCOSQ scale were significantly higher than before treatment,and the difference was statistically significant(P<0.05);while the scores of body hair problems and infertility problems before and after treatment were not significantly different(P>0.05).2 Experimental studies(1)Experiment 1①Pathological status of PCOS:Histopathological sections of rat ovaries showed that the ovaries of rats in the PCOS and PCOS+CUMS groups were enlarged in size and irregular in morphology,and a large number of cystic dilated follicles were seen in them,with a reduced number of granulosa cell layers and loosely arranged structures,suggesting ovarian polycysticlike changes,with more obvious pathological changes in rats in the PCOS+CUMS group.Serum sex hormone levels in rats showed that serum LH,LH/FSH and FT levels were significantly higher in both PCOS and PCOS+CUMS groups(P<0.05).②Chronic stress pathology:the results of the open field test and forced swimming test showed that the rats in the CUMS group and the PCOS+CUMS group had significantly fewer horizontal locomotor frames in the open field test(P<0.05)and significantly longer hovering immobility in the swimming test(P<0.05).Histopathological sections of the CA1 region of the rat hippocampus showed that the neuronal cells in the CA1 region of the hippocampus in the CUMS and PCOS+CUMS groups were loosely arranged and disorganized,with different sizes and wrinkled morphology,and some of the cells had deep cytoplasmic staining or blurred nucleoli or lysed chromatin in the nucleus.The serum levels of amine neurotransmitters in rats showed that the serum levels of 5-HT,DA and NE were significantly lower in the CUMS and PCOS+CUMS groups(P<0.05).(2)Experiment 2①Changes in the pathological status of PCOS after drug intervention:histopathological sections of rat ovaries showed that the number of growing follicles and atretic follicles in the model group increased significantly(P<0.05)and the number of atretic follicles in the Western medicine group increased significantly(P<0.05)compared with the blank group;the number of growing follicles in the ovaries of rats in the Chinese medicine group decreased significantly(P<0.05)compared with the model group.The serum sex hormone levels of rats showed that compared with the model group,the serum LH and FT of rats in the Chinese medicine group and western medicine group were significantly decreased(P<0.05),and there was a trend of decrease in LH/FSH,but it was not statistically significant(P>0.05).②Changes of chronic stress pathological state after drug intervention:the results of the open field test and forced swimming test showed that compared with the pre-treatment period,the number of horizontal motor frames of rats in the Chinese medicine group increased significantly(P<0.05)and the duration of forced swimming immobility decreased significantly(P<0.05)after treatment,and the duration of forced swimming immobility of rats in the western medicine group decreased,but the difference was not significant(P>0.05);in the comparison between groups after treatment In the comparison between the groups after treatment,the number of movements of rats in the Chinese medicine group was significantly increased compared with that in the model group(P<0.05),and the forced swimming hovering immobility time of rats in the Chinese medicine group and the western medicine group was reduced,but the difference was not significant(P>0.05).The histopathological sections of CA1 area of hippocampus in rats showed that the neuronal cells in CA1 area of hippocampus in Chinese medicine group and western medicine group showed significant improvement in pathological changes.The serum levels of amine neurotransmitters in the rats showed that the serum levels of 5-HT,DA and NE in the rats of the Chinese medicine group were significantly higher than those in the model group(P<0.05),and the NE levels in the rats of the western medicine group were significantly higher(P<0.05),while the changes of 5-HT and DA were not significant(P>0.05).(3)Experiment 3①Changes in the SIRT1/P53 pathway in ovarian and hippocampal tissues after drug intervention:In ovarian tissues,compared with the blank group,the SIRT1 level in the model group rats was significantly reduced(P<0.05),while the Acetyl-P53 level was significantly increased(P<0.05);Compared with the model group,the level of SIRT1 in the traditional Chinese medicine group significantly increased(P<0.05),while the level of SIRT1 in the western medicine group increased(P>0.05).The level of Acetyl-P53 in both groups significantly decreased(P<0.05).Although the P53 level in the model group increased,there was no significant difference between the four groups(P>0.05).In the hippocampal tissue,compared with the blank group,the SIRT1 level of the model group rats was significantly reduced(P<0.05),while the levels of P53 and Acetyl-P53 were both increased(P>0.05);Compared with the model group,the level of SIRT1 in the traditional Chinese medicine group significantly increased(P<0.05),while the level of SIRT1 in the western medicine group increased(P>0.05);The levels of P53 and Acetyl-P53 in both the traditional Chinese medicine group and the Western medicine group decreased compared to the model group,but there was no significant difference between the four groups(P>0.05).②Changes in apoptosis of ovarian granulosa cells after drug intervention:TUNEL staining showed that the apoptosis rate of ovarian granulosa cells in the blank group was the lowest;The number of apoptotic cells in the model group significantly increased,and the apoptosis rate significantly increased compared to the blank group(P<0.05);Compared with the model group,the apoptosis rate of ovarian granulosa cells in the traditional Chinese medicine group was significantly reduced(P<0.05),while the apoptosis rate of ovarian granulosa cells in the western medicine group was slightly reduced,but there was no statistical significance(P>0.05).The expression level of Caspase-3 protein in the ovaries of the model group rats showed a significant increase compared to the blank group(P<0.05);Compared with the model group,the levels of Caspase-3 in the traditional Chinese medicine group and the Western medicine group were significantly reduced(P<0.05).Conclusions1 Clinical research shows that the Bushen Jieyu Tiaochong Formula can improve the symptoms,signs and serum sex hormone levels of PCOS patients with kidney deficiency and liver depression,as well as improve the emotional state and quality of life of patients,and finally achieve the purpose of treating PCOS.2 The experimental research showed that the rat model of PCOS combined with chronic psychological stress established by letrozole and CUMS method could show both reproductive function and psychosomatic abnormalities,and the modeling effect was better than that of the single method,and the pathological state was more obvious.The formula can upregulate the expression of SIRT1 in the ovary/hippocampus of PCOS combined with CUMS rats,enhance its deacetylation effect on P53,reduce the content of acetylated P53 in the ovary/hippocampus,and improve the morphology and function of ovary/hippocampus by regulating the SIRT1/P53 signaling pathway;It can also down-regulate ovarian Caspase-3 protein expression level mediated by SIRT1/P53 pathway,inhibit the activation of apoptosis-related pathway,and then delay ovarian granulosa cell apoptosis. |
PDF Full Text Request