Background: Among anti-hypertension drugs, calcium (Ca2+) antagonists cause gingival overgrowth as a side effect. We previously discovered that this side effect was due to elevation of the calcium concentration in the...Background: Among anti-hypertension drugs, calcium (Ca2+) antagonists cause gingival overgrowth as a side effect. We previously discovered that this side effect was due to elevation of the calcium concentration in the cytosol ([Ca2+]i). Ca2+ entry through non-selective cation channels (NSCCs) and Ca2+ release from intracellular Ca2+ stores are involved in this [Ca2+]i elevation. Furthermore, we discovered that calcium-sensing receptors (CaSRs) participate in nifedipine-induced [Ca2+]i elevation. Transient receptor potential (TRP) channels have been identified as NSCCs. In the present study, we undertook experiments to determine if TRPV1 channels are present in gingival fibroblasts and to ascertain if nifedipine-activated NSCCs are TRPV1 channels. Methods Normal human gingival fibroblast Gin-1 cells were used. The [Ca2+]i was measured using a video-imaging analysis system with the Ca2+-sensitive fluorescent dye fura-2/AM. Results: The NSCC inhibitor SKF96365 significantly inhibited nifedipine-induced [Ca2+]i elevation. TRPV1 channel agonists such as capsaicin, olvanil and resiniferatoxin concentration-dependently elevated the [Ca2+]i. The TRPV1 channel activator anandamide concentration-dependently increased the [Ca2+]i. The TRPV1 channel antagonists capsazepine, AMG9810, iodoresiniferatoxin, ruthenium red, and SB366791 significantly inhibited nifedipine-induced [Ca2+]i elevation. Conclusion: These results suggest that Ca2+ entry through TRPV1 channels is involved in the nifedipine-induced [Ca2+]i elevation seen in gingival fibroblasts. We describe here a modified version of our ‘calcium trigger theory’.展开更多
目的:本研究利用药效团、分子对接及实时反转录PCR(RT-PCR)方法探究诃子制草乌通过瞬时受体电位香草酸1(TRPV1)介导的减毒机制。方法:采用Discovery Studio 4.0软件构建基于TRPV1激动剂分子共同特征的TRPV1激活药效团,虚拟筛选出草乌中...目的:本研究利用药效团、分子对接及实时反转录PCR(RT-PCR)方法探究诃子制草乌通过瞬时受体电位香草酸1(TRPV1)介导的减毒机制。方法:采用Discovery Studio 4.0软件构建基于TRPV1激动剂分子共同特征的TRPV1激活药效团,虚拟筛选出草乌中的潜在活性成分;对草乌成分及诃子制草乌中引入的诃子成分进行Autodock vina分子对接研究;根据结果选出代表性成分,采用实时反转录PCR验证是否可促进心肌细胞中TRPV1 mRNA表达。结果:药效团共筛选出草乌中包括毒性较大的双酯型生物碱乌头碱、新乌头碱及次乌头碱在内的13个成分,分子对接结果表明这13个成分及诃子制草乌引入的诃子成分均有潜力与TRPV1结合。RT-PCR实验中没食子酸和新乌头碱在浓度≥25μmol/L时,均可明显促进大鼠H9c2心肌细胞中TRPV1 mRNA表达的增加,随着给药浓度的增大表达量增加,新乌头碱促进TRPV1 mRNA表达的作用更强。药效团、分子对接结果表明诃子和草乌中均含有多种具备TRPV1激活潜力的成分。结论:实验证明新乌头碱、没食子酸均可在一定程度上激活TRPV1通道,可作为TRPV1的激动剂。根据结果推测,诃子可能会通过拮抗草乌中的活性成分与TRPV1的结合,从而降低草乌通过TRPV1介导的毒性;也可能是诃子制草乌在引入没食子酸等诃子成分后,可在安全剂量范围内使TRPV1通道快速脱敏从而实现减毒。展开更多
The transient receptor potential cation channel subfamily V member 1（TRPV1） provides the sensation of pain（nociception）. However, it remains unknown whether TRPV1 is activated after peripheral nerve injury, or whe...The transient receptor potential cation channel subfamily V member 1（TRPV1） provides the sensation of pain（nociception）. However, it remains unknown whether TRPV1 is activated after peripheral nerve injury, or whether activation of TRPV1 affects neural regeneration. In the present study, we established rat models of unilateral sciatic nerve crush injury, with or without pretreatment with AMG517（300 mg/kg）, a TRPV1 antagonist, injected subcutaneously into the ipsilateral paw 60 minutes before injury. At 1 and 2 weeks after injury, we performed immunofluorescence staining of the sciatic nerve at the center of injury, at 0.3 cm proximal and distal to the injury site, and in the dorsal root ganglia. Our results showed that Wallerian degeneration occurred distal to the injury site, and neurite outgrowth and Schwann cell regeneration occurred proximal to the injury. The number of regenerating myelinated and unmyelinated nerve clusters was greater in the AMG517-pretreated rats than in the vehicle-treated group, most notably 2 weeks after injury. TRPV1 expression in the injured sciatic nerve and ipsilateral dorsal root ganglia was markedly greater than on the contralateral side. Pretreatment with AMG517 blocked this effect. These data indicate that TRPV1 is activated or overexpressed after sciatic nerve crush injury, and that blockade of TRPV1 may accelerate regeneration of the injured sciatic nerve.展开更多
文摘Background: Among anti-hypertension drugs, calcium (Ca2+) antagonists cause gingival overgrowth as a side effect. We previously discovered that this side effect was due to elevation of the calcium concentration in the cytosol ([Ca2+]i). Ca2+ entry through non-selective cation channels (NSCCs) and Ca2+ release from intracellular Ca2+ stores are involved in this [Ca2+]i elevation. Furthermore, we discovered that calcium-sensing receptors (CaSRs) participate in nifedipine-induced [Ca2+]i elevation. Transient receptor potential (TRP) channels have been identified as NSCCs. In the present study, we undertook experiments to determine if TRPV1 channels are present in gingival fibroblasts and to ascertain if nifedipine-activated NSCCs are TRPV1 channels. Methods Normal human gingival fibroblast Gin-1 cells were used. The [Ca2+]i was measured using a video-imaging analysis system with the Ca2+-sensitive fluorescent dye fura-2/AM. Results: The NSCC inhibitor SKF96365 significantly inhibited nifedipine-induced [Ca2+]i elevation. TRPV1 channel agonists such as capsaicin, olvanil and resiniferatoxin concentration-dependently elevated the [Ca2+]i. The TRPV1 channel activator anandamide concentration-dependently increased the [Ca2+]i. The TRPV1 channel antagonists capsazepine, AMG9810, iodoresiniferatoxin, ruthenium red, and SB366791 significantly inhibited nifedipine-induced [Ca2+]i elevation. Conclusion: These results suggest that Ca2+ entry through TRPV1 channels is involved in the nifedipine-induced [Ca2+]i elevation seen in gingival fibroblasts. We describe here a modified version of our ‘calcium trigger theory’.
文摘背景:内脏高敏感被认为是肠易激综合征的主要病理生理机制之一。目的:探讨内脏高敏感大鼠结肠特异背根神经节(DRG)神经元上的TRPV1表达及其电生理特征。方法:20只10 d龄大鼠随机分为模型组和对照组,模型组以结直肠内灌注乙酸诱导内脏高敏感模型,对照组灌注等量0.9%NaC l溶液。以结肠壁注射DiI荧光染料逆行标记DRG神经元(即结肠特异DRG神经元),免疫荧光法检测DRG神经元上的TRPV1表达,膜片钳技术记录神经元电生理特征。结果:模型组结肠特异DRG神经元上的TRPV1表达阳性率明显高于对照组(46.1%对36.6%,P<0.01),平均阈电流值明显低于对照组[(57.80±1.32)pA对(73.45±4.51)pA,P<0.05],2倍阈电流刺激下动作电位频率明显高于对照组[(8.20±1.10)Hz对(4.54±0.66)Hz,P<0.05],差异均有统计学意义。模型组60 mm Hg结直肠扩张压力下的腹壁回撤反射(AWR)评分与TRPV1表达阳性率、2倍阈电流刺激下动作电位频率呈正相关(r=0.87和r=0.73,P<0.01),与阈电流值呈负相关(r=-0.81,P<0.01)。结论:结肠特异DRG神经元TRPV1表达上调、细胞兴奋性增加,可能是内脏高敏感形成的重要环节。
基金supported by the National Natural Science Foundation of China,No.81171178the Natural Science Foundation of Shanxi Province in China,No.2012011036-3Scientific Research Foundation of Shanxi Province of China for the Returned Overseas Chinese Scholars,No.2013011054-2
文摘The transient receptor potential cation channel subfamily V member 1（TRPV1） provides the sensation of pain（nociception）. However, it remains unknown whether TRPV1 is activated after peripheral nerve injury, or whether activation of TRPV1 affects neural regeneration. In the present study, we established rat models of unilateral sciatic nerve crush injury, with or without pretreatment with AMG517（300 mg/kg）, a TRPV1 antagonist, injected subcutaneously into the ipsilateral paw 60 minutes before injury. At 1 and 2 weeks after injury, we performed immunofluorescence staining of the sciatic nerve at the center of injury, at 0.3 cm proximal and distal to the injury site, and in the dorsal root ganglia. Our results showed that Wallerian degeneration occurred distal to the injury site, and neurite outgrowth and Schwann cell regeneration occurred proximal to the injury. The number of regenerating myelinated and unmyelinated nerve clusters was greater in the AMG517-pretreated rats than in the vehicle-treated group, most notably 2 weeks after injury. TRPV1 expression in the injured sciatic nerve and ipsilateral dorsal root ganglia was markedly greater than on the contralateral side. Pretreatment with AMG517 blocked this effect. These data indicate that TRPV1 is activated or overexpressed after sciatic nerve crush injury, and that blockade of TRPV1 may accelerate regeneration of the injured sciatic nerve.
