Objective To study clinical significance and the difference of calcitonin receptor stimulating peptide-3 （CRSP-3） expression in placenta tissue between normal preg- nancies and patients with preeclampsia. Methods CR...Objective To study clinical significance and the difference of calcitonin receptor stimulating peptide-3 （CRSP-3） expression in placenta tissue between normal preg- nancies and patients with preeclampsia. Methods CRSP-3 expression position in placenta tissues of 50 cases with preeclampsia （preeclampsia group： 25 cases of mild preeclampsia and 25 cases of severe preeclampsia） and 30 cases of normal late pregnancy （control group） was detected using immunohistochemical methods. CRSP-3 mRNA and protein expression in placenta tissues was detected using Real-time PCR and Western blotting method. Results Syncytiotrophoblast and cytotrophoblast cells expressed CRSP-3 in placenta tissue of preeclampsia and normal pregnancy patients. The expression levels of CRSP- 3 mRNA and protein in placenta tissue ofpreeclampsia patients were higher than those in control group （P〈O.05）. The expression levels of CRSP-3 mRNA and protein in placenta tissue of severe preeclampsia patients were higher than those in control group （P〈0.01） significantly. The expression level of CRSP-3 mRNA and protein in placenta tissue of severe preeclampsia group was slightly higher than that in mild preeclampsia group, but there were no statistical differences between the two groups （P〉0. 05）. Conclusion The up-regulation of CRSP-3 in placenta of preeclamptic patients may be associated with the etiology of preeclampsia, but may have no relation with progress and degree of preeclampsia.展开更多
Numerous studies have demonstrated that endothelin-1 combines with endothelin receptor A, resulting in intense vasoconstriction. Although calcitonin gene-related peptide （CGRP） suppresses endothelin-1, CGRP and endo...Numerous studies have demonstrated that endothelin-1 combines with endothelin receptor A, resulting in intense vasoconstriction. Although calcitonin gene-related peptide （CGRP） suppresses endothelin-1, CGRP and endothelin receptor A exhibit direct biological effects on brain tissue. The present study analyzed CGRP and endothelin receptor A expression following subarachnoid hemorrhage in rabbits using immunohistochemistry. CGRP expression was significant at 5 days after model establishment, and endothelin receptor A expression was significant at 3 days after model induction. The perimeter of the basilar artery was measured to determine the amount of cerebral vasospasm. Analytical results revealed a significantly shortened basilar artery perimeter following subarachnoid hemorrhage. Changes in the basilar artery perimeter were negatively associated with endothelin receptor A expression, but positively correlated with CGRP expression in vessels. These results suggest that following subarachnoid hemorrhage, CGRP and endothelin receptor A expressions dynamically changed in brain vessels and tissues, although these changes were not synchronous. Changes in endothelin receptor A expression exhibited a significant effect on the occurrence and development of delayed cerebral vasospasm and delayed neuronal death, while CGRP relaxed vessels and protected nerves.展开更多
BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid...BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid receptors, over-excitation can cause neuronal apoptosis. Calcitonin gene related peptide has a strongly biological activity. On one hand, it can protect ischemic neurons through inhibiting the expression of NMDAR1 mRNA; on the other hand, it can play the protective effect through down-regulating the expression of NMDAR1 mRNA by exogenous calcitonin gene related peptide. OBJECTIVE: To observe the expression of NMDAR1 and the regulatory effect of calcitonin gene related peptide on the expression of NMDAR1 mRNA and protein in the cerebral cortex of rats with focal cerebral ischemia/reperfusion (I/R). DESIGN: Randomized controlled animal study. SETTING: China Medical University. MATERIALS: A total of 216 healthy male Wistar rats, general grade, weighing 250-280 g, were selected in this study. Twelve rats were randomly selected to regard as control group; meanwhile, other 204 rats were used to establish middle cerebral artery occlusion/reperfusion (MACO) models. The main reagents were detailed as follows: calcitonin gene related peptide (Sigma Company); calcitonin gene related peptide kit (Boster Company); antibody Ⅰ, Ⅱ and antibody β-actin Ⅰ, Ⅱ of NMDAR1 mRNA and chemiluminescence reagent (Santa Cruz Company, USA). METHODS: The experiment was carried out in the Laboratory of Neurobiology of China Medical University from August 2005 to June 2006. ① Right MCAO models of rats were established to cause focal ischemia and scored based on Zea Longa five-grade scale. If the scores were 1, 2 and 3 after wakefulness, the MACO models were established successfully and involved in the experiment. A total of 120 rats with successful modeling were randomly divided into I/R group and administration group with 60 in each group. All rats in the both groups were observed at five time points, including 6, 12, 24, 48 and 72 hours after reperfusion and after 2-hour ischemia, with 12 experimental animals at each time point. Six rats were prepared for detection of hybridization in situ, and the other 6 were used for Western blotting histochemical detection. Rats in the control group were opened their skin to separate common carotid artery and not treated with line and drugs. In addition, rats in the I/R group were treated with 1 mL saline at 2 hours after focal cerebral ischemia, and then, rats in the administration group were treated with 1 mL (1 g/L) calcitonin gene related peptide at 2 hours after focal cerebral ischemia. ② The expression of NMDAR1 mRNA was detected with hybridization in situ at various time points; moreover, the expression of NMDAR1 protein was measured with Western blotting method at various time points. The results were analyzed with Metamoph imaging analytical system. MAIN OUTCOME MEASURES: The expression of NMDAR1 mRNA and its protein in cortical neurons of rats at various time points. RESULTS: A total of 84 rats were excluded because of non-symptoms, exanimation or death; and then, 132 rats were involved in the final analysis. The expression of NMDAR1 mRNA and its protein in cortical neurons of rats in the control group was 0.205±0.001 and 0.184±0.001, respectively; after I/R, expression of NMDAR1 mRNA and its protein was up-regulated, especially, expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.245±0.003, 0.287±0.004, 0.354±0.008, 0.284±0.002 and 0.217±0.006, respectively; moreover, expression of protein at 6, 12, 24, 48 and 72 hours was 0.222±0.003, 0.261±0.028, 0.311±0.004, 0.259±0.013 and 0.210±0.008, respectively. There was significant difference between the two groups (0.205±0.001, P < 0.01). The expression was up-related in the former 24 hours, reached peak at 24 hours, down-regulated, and decreased to the level of control group at 72 hours. Except 72 hours, the expression of NMDAR1 mRNA and its protein was lower in administration group than that in I/R group at other four time points. In addition, the expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.223±0.005, 0.243±0.001, 0.292±0.002, 0.250±0.003 and 0.213±0.003, respectively; moreover, the expression of protein at 6, 12, 24, 48 and 72 hours was 0.216±0.006, 0.245±0.025, 0.276±0.003, 0.241±0.045 and 0.202±0.013, respectively. There was significant difference at various time points (P < 0.05). CONCLUSION: The expressions of NMDAR1 mRNA and its protein of peripheral cortical neurons are up-related in ischemic area after focal cerebral I/R. Meanwhile, exogenous calcitonin gene related peptide can protect cortical neurons through inhibiting expression of NMDAR1 mRNA and its protein after focal cerebral I/R.展开更多
Transient receptor potential channel A1 is one of the important transducers of noxious stimuli in the primary afferents, which may contribute to generation of neurogenic inflammation and hyperalgesia. The present stud...Transient receptor potential channel A1 is one of the important transducers of noxious stimuli in the primary afferents, which may contribute to generation of neurogenic inflammation and hyperalgesia. The present study was designed to investigate if activation of transient receptor potential channel A1 may induce calcitonin gene-related peptide release from the primary afferent neurons. We found that application of allyl isothiocyanate, a transient receptor potential channel A1 activator, caused calcitonin gene-related peptide release from the cultured rat dorsal root ganglion neurons. Knock- down of transient receptor potential channel A1 with an antisense oligodeoxynucleotide prevented calcitonin gene-related peptide release by allyl isothiocyanate application in cultured dorsal root ganglion neurons. Thus, we concluded that transient receptor potential channel A1 activation caused calcitonin gene-related peptide release in sensory neurons.展开更多
AIM:To determine the molecular mechanisms of Shugan decoction(SGD) in the regulation of colonic motility and visceral hyperalgesia(VHL) in irritable bowel syndrome(IBS).METHODS:The chemical compounds contained in SGD ...AIM:To determine the molecular mechanisms of Shugan decoction(SGD) in the regulation of colonic motility and visceral hyperalgesia(VHL) in irritable bowel syndrome(IBS).METHODS:The chemical compounds contained in SGD were measured by high-performance liquid chromatography.A rat model of IBS was induced by chronic water avoidance stress(WAS).The number of fecal pellets was counted after WAS and the pain pressure threshold was measured by colorectal distension.Morphological changes in colonic mucosa were detected by hematoxylin-eosin staining.The contents of tumor necrosis factor(TNF)-αin colonic tissue and calcitonin-gene-related peptide(CGRP)in serum were measured by ELISA.The protein expression of serotonin[5-hydroxytryptamide(5-HT)],serotonin transporter(SERT),chromogranin A(Cg A)and CGRP incolon tissue was measured by immunohistochemistry.RESULTS:SGD inhibited colonic motility dysfunction and VHL in rats with IBS.Blockers of transient receptor potential(TRP)vanilloid 1(TRPV1)(Ruthenium Red)and TRP ankyrin-1(TRPA1)(HC-030031)and activator of protease-activated receptor(PAR)4 increased the pain pressure threshold,whereas activators of PAR2and TRPV4 decreased the pain pressure threshold in rats with IBS.The effect of SGD on pain pressure threshold in these rats was abolished by activators of TRPV1(capsaicin),TRPV4(RN1747),TRPA1(Polygodial)and PAR2(AC55541).In addition,CGRP levels in serum and colonic tissue were both increased in these rats.TNF-αlevel in colonic tissue was also significantly upregulated.However,the levels of 5-HT,SERT and Cg A in colonic tissue were decreased.All these pathological changes in rats with IBS were attenuated by SGD.CONCLUSION:SGD alleviated VHL and attenuated colon motility in IBS,partly by regulating TRPV1,TRPV4,TRPA1,PAR2,5-HT,Cg A and SERT,and reducing CGRP and TNF-αlevel.展开更多
文摘Objective To study clinical significance and the difference of calcitonin receptor stimulating peptide-3 （CRSP-3） expression in placenta tissue between normal preg- nancies and patients with preeclampsia. Methods CRSP-3 expression position in placenta tissues of 50 cases with preeclampsia （preeclampsia group： 25 cases of mild preeclampsia and 25 cases of severe preeclampsia） and 30 cases of normal late pregnancy （control group） was detected using immunohistochemical methods. CRSP-3 mRNA and protein expression in placenta tissues was detected using Real-time PCR and Western blotting method. Results Syncytiotrophoblast and cytotrophoblast cells expressed CRSP-3 in placenta tissue of preeclampsia and normal pregnancy patients. The expression levels of CRSP- 3 mRNA and protein in placenta tissue ofpreeclampsia patients were higher than those in control group （P〈O.05）. The expression levels of CRSP-3 mRNA and protein in placenta tissue of severe preeclampsia patients were higher than those in control group （P〈0.01） significantly. The expression level of CRSP-3 mRNA and protein in placenta tissue of severe preeclampsia group was slightly higher than that in mild preeclampsia group, but there were no statistical differences between the two groups （P〉0. 05）. Conclusion The up-regulation of CRSP-3 in placenta of preeclamptic patients may be associated with the etiology of preeclampsia, but may have no relation with progress and degree of preeclampsia.
基金the National 863 Project of China,No. 2006AA02Z4Z4the National Natural Science Foundation of China,No. 30870844+1 种基金the New Century Excellent Talent Support Project of Ministry of Education,No. NCET-05-0831the "13115" Special Fund for Major Science and Technology Projects of Shaanxi Province,No. 2008ZDKG-66
文摘Numerous studies have demonstrated that endothelin-1 combines with endothelin receptor A, resulting in intense vasoconstriction. Although calcitonin gene-related peptide （CGRP） suppresses endothelin-1, CGRP and endothelin receptor A exhibit direct biological effects on brain tissue. The present study analyzed CGRP and endothelin receptor A expression following subarachnoid hemorrhage in rabbits using immunohistochemistry. CGRP expression was significant at 5 days after model establishment, and endothelin receptor A expression was significant at 3 days after model induction. The perimeter of the basilar artery was measured to determine the amount of cerebral vasospasm. Analytical results revealed a significantly shortened basilar artery perimeter following subarachnoid hemorrhage. Changes in the basilar artery perimeter were negatively associated with endothelin receptor A expression, but positively correlated with CGRP expression in vessels. These results suggest that following subarachnoid hemorrhage, CGRP and endothelin receptor A expressions dynamically changed in brain vessels and tissues, although these changes were not synchronous. Changes in endothelin receptor A expression exhibited a significant effect on the occurrence and development of delayed cerebral vasospasm and delayed neuronal death, while CGRP relaxed vessels and protected nerves.
文摘BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid receptors, over-excitation can cause neuronal apoptosis. Calcitonin gene related peptide has a strongly biological activity. On one hand, it can protect ischemic neurons through inhibiting the expression of NMDAR1 mRNA; on the other hand, it can play the protective effect through down-regulating the expression of NMDAR1 mRNA by exogenous calcitonin gene related peptide. OBJECTIVE: To observe the expression of NMDAR1 and the regulatory effect of calcitonin gene related peptide on the expression of NMDAR1 mRNA and protein in the cerebral cortex of rats with focal cerebral ischemia/reperfusion (I/R). DESIGN: Randomized controlled animal study. SETTING: China Medical University. MATERIALS: A total of 216 healthy male Wistar rats, general grade, weighing 250-280 g, were selected in this study. Twelve rats were randomly selected to regard as control group; meanwhile, other 204 rats were used to establish middle cerebral artery occlusion/reperfusion (MACO) models. The main reagents were detailed as follows: calcitonin gene related peptide (Sigma Company); calcitonin gene related peptide kit (Boster Company); antibody Ⅰ, Ⅱ and antibody β-actin Ⅰ, Ⅱ of NMDAR1 mRNA and chemiluminescence reagent (Santa Cruz Company, USA). METHODS: The experiment was carried out in the Laboratory of Neurobiology of China Medical University from August 2005 to June 2006. ① Right MCAO models of rats were established to cause focal ischemia and scored based on Zea Longa five-grade scale. If the scores were 1, 2 and 3 after wakefulness, the MACO models were established successfully and involved in the experiment. A total of 120 rats with successful modeling were randomly divided into I/R group and administration group with 60 in each group. All rats in the both groups were observed at five time points, including 6, 12, 24, 48 and 72 hours after reperfusion and after 2-hour ischemia, with 12 experimental animals at each time point. Six rats were prepared for detection of hybridization in situ, and the other 6 were used for Western blotting histochemical detection. Rats in the control group were opened their skin to separate common carotid artery and not treated with line and drugs. In addition, rats in the I/R group were treated with 1 mL saline at 2 hours after focal cerebral ischemia, and then, rats in the administration group were treated with 1 mL (1 g/L) calcitonin gene related peptide at 2 hours after focal cerebral ischemia. ② The expression of NMDAR1 mRNA was detected with hybridization in situ at various time points; moreover, the expression of NMDAR1 protein was measured with Western blotting method at various time points. The results were analyzed with Metamoph imaging analytical system. MAIN OUTCOME MEASURES: The expression of NMDAR1 mRNA and its protein in cortical neurons of rats at various time points. RESULTS: A total of 84 rats were excluded because of non-symptoms, exanimation or death; and then, 132 rats were involved in the final analysis. The expression of NMDAR1 mRNA and its protein in cortical neurons of rats in the control group was 0.205±0.001 and 0.184±0.001, respectively; after I/R, expression of NMDAR1 mRNA and its protein was up-regulated, especially, expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.245±0.003, 0.287±0.004, 0.354±0.008, 0.284±0.002 and 0.217±0.006, respectively; moreover, expression of protein at 6, 12, 24, 48 and 72 hours was 0.222±0.003, 0.261±0.028, 0.311±0.004, 0.259±0.013 and 0.210±0.008, respectively. There was significant difference between the two groups (0.205±0.001, P < 0.01). The expression was up-related in the former 24 hours, reached peak at 24 hours, down-regulated, and decreased to the level of control group at 72 hours. Except 72 hours, the expression of NMDAR1 mRNA and its protein was lower in administration group than that in I/R group at other four time points. In addition, the expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.223±0.005, 0.243±0.001, 0.292±0.002, 0.250±0.003 and 0.213±0.003, respectively; moreover, the expression of protein at 6, 12, 24, 48 and 72 hours was 0.216±0.006, 0.245±0.025, 0.276±0.003, 0.241±0.045 and 0.202±0.013, respectively. There was significant difference at various time points (P < 0.05). CONCLUSION: The expressions of NMDAR1 mRNA and its protein of peripheral cortical neurons are up-related in ischemic area after focal cerebral I/R. Meanwhile, exogenous calcitonin gene related peptide can protect cortical neurons through inhibiting expression of NMDAR1 mRNA and its protein after focal cerebral I/R.
基金supported by the Research Basis Formation Supporting Project for Private University
文摘Transient receptor potential channel A1 is one of the important transducers of noxious stimuli in the primary afferents, which may contribute to generation of neurogenic inflammation and hyperalgesia. The present study was designed to investigate if activation of transient receptor potential channel A1 may induce calcitonin gene-related peptide release from the primary afferent neurons. We found that application of allyl isothiocyanate, a transient receptor potential channel A1 activator, caused calcitonin gene-related peptide release from the cultured rat dorsal root ganglion neurons. Knock- down of transient receptor potential channel A1 with an antisense oligodeoxynucleotide prevented calcitonin gene-related peptide release by allyl isothiocyanate application in cultured dorsal root ganglion neurons. Thus, we concluded that transient receptor potential channel A1 activation caused calcitonin gene-related peptide release in sensory neurons.
基金Supported by Innovation Program of the Shanghai Municipal Education Commission,No.12YZ065National Natural Science Foundation of China,No.81072786,No.81473630 and No.81202665+2 种基金Longhua Medical Project,No.D-09High level Project of the University of Educational Commission of Shanghai,China,No.2008GSP19Shanghai Leading Academic Discipline Project,No.J50305
文摘AIM:To determine the molecular mechanisms of Shugan decoction(SGD) in the regulation of colonic motility and visceral hyperalgesia(VHL) in irritable bowel syndrome(IBS).METHODS:The chemical compounds contained in SGD were measured by high-performance liquid chromatography.A rat model of IBS was induced by chronic water avoidance stress(WAS).The number of fecal pellets was counted after WAS and the pain pressure threshold was measured by colorectal distension.Morphological changes in colonic mucosa were detected by hematoxylin-eosin staining.The contents of tumor necrosis factor(TNF)-αin colonic tissue and calcitonin-gene-related peptide(CGRP)in serum were measured by ELISA.The protein expression of serotonin[5-hydroxytryptamide(5-HT)],serotonin transporter(SERT),chromogranin A(Cg A)and CGRP incolon tissue was measured by immunohistochemistry.RESULTS:SGD inhibited colonic motility dysfunction and VHL in rats with IBS.Blockers of transient receptor potential(TRP)vanilloid 1(TRPV1)(Ruthenium Red)and TRP ankyrin-1(TRPA1)(HC-030031)and activator of protease-activated receptor(PAR)4 increased the pain pressure threshold,whereas activators of PAR2and TRPV4 decreased the pain pressure threshold in rats with IBS.The effect of SGD on pain pressure threshold in these rats was abolished by activators of TRPV1(capsaicin),TRPV4(RN1747),TRPA1(Polygodial)and PAR2(AC55541).In addition,CGRP levels in serum and colonic tissue were both increased in these rats.TNF-αlevel in colonic tissue was also significantly upregulated.However,the levels of 5-HT,SERT and Cg A in colonic tissue were decreased.All these pathological changes in rats with IBS were attenuated by SGD.CONCLUSION:SGD alleviated VHL and attenuated colon motility in IBS,partly by regulating TRPV1,TRPV4,TRPA1,PAR2,5-HT,Cg A and SERT,and reducing CGRP and TNF-αlevel.
