Objective:To elucidate the underlying mechanism and effect of electroacupuncture(EA)on the neurogenic bladder following suprasacral spinal cord injury(ssCI).A rat model of detrusor hyperreflexia after SsCI was establi...Objective:To elucidate the underlying mechanism and effect of electroacupuncture(EA)on the neurogenic bladder following suprasacral spinal cord injury(ssCI).A rat model of detrusor hyperreflexia after SsCI was established to examine the urodynamics,detrusor muscle tissue morphology,the protein and mRNA expression levels of pituitary adenylate cyclase activating peptide(PACAP)and its receptor PAC1R,and cyclic adenosine monophosphate(cAMP)content in the detrusor muscle with a focus on the PACAPcAMP signaling pathway.Method:A total of 72 female SD rats were randomized into control group and sham operation group(n=12 per group)by using a random number table.The remaining 48 rats were established into the model of detrusor hyperreflexia after SsCI.After successful modeling,these rats were randomly assigned to model,EA,and EA+PACAP6-38 groups(n=12 per group).The unsuccessful modeled rats were used for exploratory observation.For the rats in EA group,"Ciliao(BL32)""Zhongji(CV3)",and"Sanyinjiao(SP6)"were needled and stimulated by EA.The PACAP receptor antagonist PACAP6-38 was administered intraperitoneally in the EA+PACAP6-38 group before EA,and EA was applied for seven consecutive days.After treatment,the urodynamics of the rats were analyzed,and hematoxylin and eosin staining was used to examine rat bladder detrusor tissue morphology.The expressions of PACAP-38 and PAC1R were detected by immunohistochemistry and Western blot.The mRNA expression levels of PACAP-38 and PAC1R were examined by RT-qPCR,while cAMP content was detected by ELISA.Results:(1)Compared with sham operation group,it was exhibited disarray in the transitional epithelium cells of the bladder in the modeled rats.The intercellular space was significantly widened,accompanied by inflammatory cell infiltration and noticeable tissue edema.Both the bladder initial pressure and leak point pressure of the rats were higher(P<0.01),whereas the maximum cystometric capacity and bladder compliance were lower(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,together with the cAMP content,were lower(P<0.05).(2)Compared with the model rats,the EA group showed reduced inflammatory response in the detrusor muscle tissue,with decreased monocyte infiltration and less severe tissue edema.The bladder smooth muscle cells exhibited increased integrity,and there was decreased cellular tissue edema,inflammatory cell infiltration,and fibroplasia.The bladder initial pressure and leak point pressure were lower(P<0.05),while the maximum cystometric capacity and bladder compliance were higher(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,along with the cAMP content,were higher(P<0.05).(3)Compared to the EA group,the EA+PACAP6-38 group showed a less organized arrangement of muscle fibers in the detrusor muscle tissue,larger intercellular space,monocyte infltration,and considerable tissue edema.The changes in bladder initial pressure and leak point pressure were not significant(P>0.05),while the maximum cystometric capacity and bladder compliance were lower(P<0.05).The changes in the protein and mRNA expressions of PACAP-38 within the detrusor muscle were not signifcant(P>0.05),whereas the protein and mRNA expressions of PAC1R were reduced(P<0.05),and the cAMP content within the detrusor muscle was lower(P<0.05).Conclusion:EA can ameliorate the uninhibited contractile condition of the detrusor muscle in the bladder following SSCI.By mediating the PACAP-cAMP signaling pathway,it reduces the pathological damage to the detrusor muscle,thereby improving bladder function.展开更多
Objective:To observe the effects of electroacupuncture on urodynamics and intramedullary apoptosis related factor B cell lymphoma gene-2(Bcl-2),Bcl-2 related protein X(Bax)and brain-derived neurotrophic factor(BDNF)in...Objective:To observe the effects of electroacupuncture on urodynamics and intramedullary apoptosis related factor B cell lymphoma gene-2(Bcl-2),Bcl-2 related protein X(Bax)and brain-derived neurotrophic factor(BDNF)in neurogenic bladder rats after supersacral spinal cord injury.Methods:A total of 60 female SD rats,SPF grade,were randomly selected,12 rats in the blank group and 12 rats in the sham operation group,and the remaining 36 rats were made models.Of these 36 rats,24 rats meeting the model standard were randomized into the model group and electroacupuncture group,with 12 rats in each group.The neurogenic bladder models after supracacral spinal cord injury were prepared by the Hassan Shaker spinal cord transection method,via combining the previous model study experience.At the end of spinal shock stage,"Dazhui(大椎GV14)""Zhongji(中极CV 3)""Ciliao(次髎BL 32)",and"Sanyinjiao(三阴交SP 6)"were selected for electroacupuncture.Observation on the histomorphological changes of bladder detrusor and spinal cord in rats was conducted by HE staining.The apoptosis rate of spinal cord cells was measured by TUNEL assay.The maximum bladder volume and compliance were measured by urodynamics test.The protein expressions of Bcl-2,Bax,and BDNF in spinal cord tissues were measured by Western blot assay.The expressions of Bcl-2,Bax,and BDNF mRNA were measured by RT-PCR assay.Results:(1)The results of HE staining showed that compared with the blank group and sham operation group,in the model group the bladder epithelial cell structure was destroyed,the detrusor muscle fibers proliferated,and there were severe bleeding changes,the spinal cord tissue was disordered,the structure was destroyed,the cell space was enlarged,and accompanied by a large number of inflammatory cell infiltration and cystic cavity formation.Compared with the model group,in the electroacupuncture group,the intact bladder epithelial cells increased,the detrusor muscle fiber proliferation decreased,the bleeding changes decreased,the complete structural area of spinal cord tissues increased,the cell space decreased,the levels were clearer,and the cystic cavity decreased.(2)The results of TUNEL assay indicated that the TUNEL positive rate of spinal cord cells in the model group was significantly higher than that in the blank group and sham group(both P<0.05).Compared with the model group,the TUNEL positive rate of spinal cord cells in the electroacupuncture group was significantly reduced(P<0.05).(3)The results of urodynamics test suggested that compared with the blank group and sham operation group,the maximum bladder volume and compliance of the model group were significantly reduced(all P<0.05).Compared with the model group,the maximum bladder volume and compliance were significantly increased in the electroacupuncture group(both P<0.05).(4)The results of Western blot assay indicated that,compared with the blank group and sham operation group,in the model group the protein contents of Bcl-2 and BDNF were significantly reduced(all P<0.05),while the protein content of Bax in the model group was significantly increased(both P<0.05).(5)The results of RT-PCR assay indicated that,compared with blank group and sham operation group,in the model group the intramedullary Bcl-2 and BDNF mRNA were significantly reduced(all P<0.05),and Bax mRNA was significantly increased(both P<0.05).Conclusion:Electroacupuncture at"GV14""CV3""BL32",and"SP6"can promote the repair of bladder detrusor,improve the maximum bladder volume and compliance of bladder,promote morphological repair of bladder detrusor,reduce spinal cord cell apoptosis rate,thus to restore the function of bladder.This is maybe related with the electroacupuncture’s two-way regulatory mechanism of increased expression of protein and gene of Bcl-2 and BDNF and decreased expression of protein and gene of Bax in spinal cord tissues.展开更多
Objective： To investigate the effect and mechanism of electroacupuncture（EA） in the intervention of neurogenic bladder（NB） caused by suprasacral spinal cord injury（SCI）.Methods： The total 60 female Sprague-Daw...Objective： To investigate the effect and mechanism of electroacupuncture（EA） in the intervention of neurogenic bladder（NB） caused by suprasacral spinal cord injury（SCI）.Methods： The total 60 female Sprague-Dawley（SD） rats were divided into blank group, sham operation group by adopting random number table method, with 12 rats in each group. The remaining rats were made into models through operation, and 24 successful model rats were randomly divided into model group and EA group, with 12 rats in each group. Since the 19 th day after modeling, EA was conducted at ＂Cìliào＂（次髎BL 32）, ＂Zhōngjí＂（中极CV 3）,＂Sānyīnjiāo＂（三阴交SP 6） and ＂Dàzhuī＂（大椎GV 14） in EA group, remaining the needle for 20 min every time. The intervention was performed for once a day, and7 consecutive days were needed. The rest of the three groups were only tied up for the same time. Urodynamics test was conducted after the intervention. The contents of PACAP-38 and PAC1 R of the spinal cord of the injury site were detected by Western blot.Results： Compared with the blank group. there were no significant differences in the bladder capacity,leak point pressure, bladder compliance and PACAP-38 and PAC1 R contents in spinal cord in sham operation group; compared with the sham operation group, the maximum bladder capacity and the bladder compliance in the model group were lower（both P〈0.05）, the leak point pressure was higher（P〈0.05）,the contents of PACAP-38 and PAC1 R were lower（both P〈0.05）; compared with the model group, the maximum bladder capacity and the bladder compliance in the EA group were higher（both P〈0.05）, the leak point pressure was lower（P〈0.05）, the contents of PACAP-38 and PAC1 R were higher（both P〈0.05）.Conclusion： EA at BL 32, CV 3, SP 6 and GV 14 can improve the bladder function of NB rats of suprasacral SCI, and its mechanism may be related to the increase expression of PACAP-38 and PAC1 R protein in the injured spinal cord tissue.展开更多
基金Supported by National Natural Science Foundation of China:82274666,82205255Natural Science Foundation of Hunan Province:2022JJ30036,2022JJ40312,20221140301+1 种基金Research Project of Education Department of Hunan Province:20C1432,21B0369Discipline of Integrated Traditional Chinese and Western Medicine of Hunan Province:2020ZXYJH23。
文摘Objective:To elucidate the underlying mechanism and effect of electroacupuncture(EA)on the neurogenic bladder following suprasacral spinal cord injury(ssCI).A rat model of detrusor hyperreflexia after SsCI was established to examine the urodynamics,detrusor muscle tissue morphology,the protein and mRNA expression levels of pituitary adenylate cyclase activating peptide(PACAP)and its receptor PAC1R,and cyclic adenosine monophosphate(cAMP)content in the detrusor muscle with a focus on the PACAPcAMP signaling pathway.Method:A total of 72 female SD rats were randomized into control group and sham operation group(n=12 per group)by using a random number table.The remaining 48 rats were established into the model of detrusor hyperreflexia after SsCI.After successful modeling,these rats were randomly assigned to model,EA,and EA+PACAP6-38 groups(n=12 per group).The unsuccessful modeled rats were used for exploratory observation.For the rats in EA group,"Ciliao(BL32)""Zhongji(CV3)",and"Sanyinjiao(SP6)"were needled and stimulated by EA.The PACAP receptor antagonist PACAP6-38 was administered intraperitoneally in the EA+PACAP6-38 group before EA,and EA was applied for seven consecutive days.After treatment,the urodynamics of the rats were analyzed,and hematoxylin and eosin staining was used to examine rat bladder detrusor tissue morphology.The expressions of PACAP-38 and PAC1R were detected by immunohistochemistry and Western blot.The mRNA expression levels of PACAP-38 and PAC1R were examined by RT-qPCR,while cAMP content was detected by ELISA.Results:(1)Compared with sham operation group,it was exhibited disarray in the transitional epithelium cells of the bladder in the modeled rats.The intercellular space was significantly widened,accompanied by inflammatory cell infiltration and noticeable tissue edema.Both the bladder initial pressure and leak point pressure of the rats were higher(P<0.01),whereas the maximum cystometric capacity and bladder compliance were lower(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,together with the cAMP content,were lower(P<0.05).(2)Compared with the model rats,the EA group showed reduced inflammatory response in the detrusor muscle tissue,with decreased monocyte infiltration and less severe tissue edema.The bladder smooth muscle cells exhibited increased integrity,and there was decreased cellular tissue edema,inflammatory cell infiltration,and fibroplasia.The bladder initial pressure and leak point pressure were lower(P<0.05),while the maximum cystometric capacity and bladder compliance were higher(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,along with the cAMP content,were higher(P<0.05).(3)Compared to the EA group,the EA+PACAP6-38 group showed a less organized arrangement of muscle fibers in the detrusor muscle tissue,larger intercellular space,monocyte infltration,and considerable tissue edema.The changes in bladder initial pressure and leak point pressure were not significant(P>0.05),while the maximum cystometric capacity and bladder compliance were lower(P<0.05).The changes in the protein and mRNA expressions of PACAP-38 within the detrusor muscle were not signifcant(P>0.05),whereas the protein and mRNA expressions of PAC1R were reduced(P<0.05),and the cAMP content within the detrusor muscle was lower(P<0.05).Conclusion:EA can ameliorate the uninhibited contractile condition of the detrusor muscle in the bladder following SSCI.By mediating the PACAP-cAMP signaling pathway,it reduces the pathological damage to the detrusor muscle,thereby improving bladder function.
基金National Natural Science Foundation of China:81473753,81874510"Domestic First-class Cultivation Discipline"of Integrated Traditional Chinese and Western Medicine in Colleges and Universities of Hunan Province。
文摘Objective:To observe the effects of electroacupuncture on urodynamics and intramedullary apoptosis related factor B cell lymphoma gene-2(Bcl-2),Bcl-2 related protein X(Bax)and brain-derived neurotrophic factor(BDNF)in neurogenic bladder rats after supersacral spinal cord injury.Methods:A total of 60 female SD rats,SPF grade,were randomly selected,12 rats in the blank group and 12 rats in the sham operation group,and the remaining 36 rats were made models.Of these 36 rats,24 rats meeting the model standard were randomized into the model group and electroacupuncture group,with 12 rats in each group.The neurogenic bladder models after supracacral spinal cord injury were prepared by the Hassan Shaker spinal cord transection method,via combining the previous model study experience.At the end of spinal shock stage,"Dazhui(大椎GV14)""Zhongji(中极CV 3)""Ciliao(次髎BL 32)",and"Sanyinjiao(三阴交SP 6)"were selected for electroacupuncture.Observation on the histomorphological changes of bladder detrusor and spinal cord in rats was conducted by HE staining.The apoptosis rate of spinal cord cells was measured by TUNEL assay.The maximum bladder volume and compliance were measured by urodynamics test.The protein expressions of Bcl-2,Bax,and BDNF in spinal cord tissues were measured by Western blot assay.The expressions of Bcl-2,Bax,and BDNF mRNA were measured by RT-PCR assay.Results:(1)The results of HE staining showed that compared with the blank group and sham operation group,in the model group the bladder epithelial cell structure was destroyed,the detrusor muscle fibers proliferated,and there were severe bleeding changes,the spinal cord tissue was disordered,the structure was destroyed,the cell space was enlarged,and accompanied by a large number of inflammatory cell infiltration and cystic cavity formation.Compared with the model group,in the electroacupuncture group,the intact bladder epithelial cells increased,the detrusor muscle fiber proliferation decreased,the bleeding changes decreased,the complete structural area of spinal cord tissues increased,the cell space decreased,the levels were clearer,and the cystic cavity decreased.(2)The results of TUNEL assay indicated that the TUNEL positive rate of spinal cord cells in the model group was significantly higher than that in the blank group and sham group(both P<0.05).Compared with the model group,the TUNEL positive rate of spinal cord cells in the electroacupuncture group was significantly reduced(P<0.05).(3)The results of urodynamics test suggested that compared with the blank group and sham operation group,the maximum bladder volume and compliance of the model group were significantly reduced(all P<0.05).Compared with the model group,the maximum bladder volume and compliance were significantly increased in the electroacupuncture group(both P<0.05).(4)The results of Western blot assay indicated that,compared with the blank group and sham operation group,in the model group the protein contents of Bcl-2 and BDNF were significantly reduced(all P<0.05),while the protein content of Bax in the model group was significantly increased(both P<0.05).(5)The results of RT-PCR assay indicated that,compared with blank group and sham operation group,in the model group the intramedullary Bcl-2 and BDNF mRNA were significantly reduced(all P<0.05),and Bax mRNA was significantly increased(both P<0.05).Conclusion:Electroacupuncture at"GV14""CV3""BL32",and"SP6"can promote the repair of bladder detrusor,improve the maximum bladder volume and compliance of bladder,promote morphological repair of bladder detrusor,reduce spinal cord cell apoptosis rate,thus to restore the function of bladder.This is maybe related with the electroacupuncture’s two-way regulatory mechanism of increased expression of protein and gene of Bcl-2 and BDNF and decreased expression of protein and gene of Bax in spinal cord tissues.
文摘Objective： To investigate the effect and mechanism of electroacupuncture（EA） in the intervention of neurogenic bladder（NB） caused by suprasacral spinal cord injury（SCI）.Methods： The total 60 female Sprague-Dawley（SD） rats were divided into blank group, sham operation group by adopting random number table method, with 12 rats in each group. The remaining rats were made into models through operation, and 24 successful model rats were randomly divided into model group and EA group, with 12 rats in each group. Since the 19 th day after modeling, EA was conducted at ＂Cìliào＂（次髎BL 32）, ＂Zhōngjí＂（中极CV 3）,＂Sānyīnjiāo＂（三阴交SP 6） and ＂Dàzhuī＂（大椎GV 14） in EA group, remaining the needle for 20 min every time. The intervention was performed for once a day, and7 consecutive days were needed. The rest of the three groups were only tied up for the same time. Urodynamics test was conducted after the intervention. The contents of PACAP-38 and PAC1 R of the spinal cord of the injury site were detected by Western blot.Results： Compared with the blank group. there were no significant differences in the bladder capacity,leak point pressure, bladder compliance and PACAP-38 and PAC1 R contents in spinal cord in sham operation group; compared with the sham operation group, the maximum bladder capacity and the bladder compliance in the model group were lower（both P〈0.05）, the leak point pressure was higher（P〈0.05）,the contents of PACAP-38 and PAC1 R were lower（both P〈0.05）; compared with the model group, the maximum bladder capacity and the bladder compliance in the EA group were higher（both P〈0.05）, the leak point pressure was lower（P〈0.05）, the contents of PACAP-38 and PAC1 R were higher（both P〈0.05）.Conclusion： EA at BL 32, CV 3, SP 6 and GV 14 can improve the bladder function of NB rats of suprasacral SCI, and its mechanism may be related to the increase expression of PACAP-38 and PAC1 R protein in the injured spinal cord tissue.
