Cili Zhou

Electro-acupuncture relieves visceral sensitivity and decreases hypothalamic corticotropin-releasing hormone levels in a rat model of irritable bowel syndrome

Previous studies into electro-acupuncture (EA) treatment of irritable bowel syndrome (IBS) have principally focused on the peripheral effects of EA in a rat model of IBS. It is not known whether EA exerts central effects in this rat model. We have examined the effects of EA on hypothalamic corticotropin-releasing hormone (CRH) levels in a rat model of IBS provoked by colorectal distension (CRD) and forelimb immobilization. EA was administered once daily to IBS model rats over a period of 7 d; untreated IBS rats and controls were also studied. The behavioral response to distension was rated according to the abdominal withdrawal reflex (AWR) score; hypothalamic CRH levels were measured by radioimmunoassay. We report that EA treatment significantly decreased visceral sensitivity to CRD in this rat model. In treated animals, EA also decreased hypothalamic CRH to control levels. Reduced hypothalamic CRH levels may mediate the beneficial effects of EA in this rat IBS model.

Mechanisms Underlying the Analgesic Effect of Moxibustion on Visceral Pain in Irritable Bowel Syndrome: A Review

Irritable bowel syndrome (IBS) is a functional bowel disorder that causes recurrent abdominal (visceral) pain. Epidemiological data show that the incidence rate of IBS is as high as 25%. Most of the medications may lead to tolerance, addiction and toxic side effects. Moxibustion is an important component of traditional Chinese medicine and has been used to treat IBS-like abdominal pain for several thousand years in China. As a mild treatment, moxibustion has been widely applied in clinical treatment of visceral pain in IBS. In recent years, it has played an irreplaceable role in alternative medicine. Extensive clinical studies have demonstrated that moxibustion for treatment of visceral pain is simple, convenient, and inexpensive, and it is being accepted by an increasing number of patients. There have not been many studies investigating the analgesic mechanisms of moxibustion. Studies exploring the analgesic mechanisms have mainly focused on visceral hypersensitivity, brain-gut axis neuroendocrine system, and immune system. This paper reviews the latest developments in moxibustion use for treatment of visceral pain in IBS from these perspectives. It also evaluates potential problems in relevant studies on the mechanisms of moxibustion therapy to promote the application of moxibustion in the treatment of IBS.

Mild Moxibustion Decreases the Expression of Prokineticin 2 and Prokineticin Receptor 2 in the Colon and Spinal Cord of Rats with Irritable Bowel Syndrome

It has been proven that prokineticin 2 (PK2) and its receptor PKR2 play an important role in hyperalgesia, while mild moxibustion can relieve visceral hypersensitivity in a rat model of irritable bowel syndrome (IBS). The goal of the present study was to determine the effects of mild moxibustion on the expression of PK2 and PKR2 in colon and spinal cord in IBS rat model, which was induced by colorectal distension using inflatable balloons. After mild moxibustion treatment, abdominal withdrawal reflex (AWR) scores were assessed by colorectal distension; protein and mRNA expression of PK2 and PKR2 in rat colon and spinal cord was determined by immunohistochemistry and fluorescence quantitative PCR. Compared with normal rats, the AWR scores of rats and the expressions of PK2/PKR2 proteins and mRNAs in colon and spinal cord tissue were significantly increased in the model group; compared with the model group, the AWR scores of rats and the expressions of PK2/PKR2 proteins and mRNAs in colon and spinal cord tissue were significantly decreased in the mild moxibustion group. These findings suggest that the analgesia effect of mild moxibustion may be associated with the reduction of the abnormally increased expression of the PK2/PKR2 proteins and mRNAs in the colon and spinal cord.

Moxibustion treatment restoring the intestinal epithelium barrier in rats with Crohn’s disease by down-regulating tumor necrosis factor alpha, tumor necrosis factor receptor 1, and tumor necrosis factor receptor 2

ObjectiveTo investigate whether moxibustion regulates tumor necrosis factor alpha (TNF-α), tumor necrosis factor receptor 1 (TNFR1), and TNFR2 in the intestinal mucosa and to explore whether moxibustion could be used by means of this mechanism, to repair the intestinal epithelium barrier disruption in Crohn’s disease (CD).MethodsThe CD rat models were established by trinitrobenzene sulfonic acid (TNBs), randomly divided into a model control (MC) group, an herb-partition moxibustion (HPM) group, a mild-warm moxibustion (MWM) group, and a salicylazosulfapyridine (SASP) group, and all were compared with a normal control (NC) group. The HPM and MWM groups were treated by moxibustion at Tianshu (ST25) and Qihai (RN6) for 14 days, and the SASP group obtained the SASP solution orally for the same period of time. The intestinal epithelium morphology and TNF-α, TNFR1, and TNFR2 contents were observed by the transmission electron microscopy and enzyme linked immunosorbent assay.ResultsThe severity of morphological changes in CD intestinal epithelium was obviously improved, and the levels of TNF-α, TNFR1, and TNFR2 in the intestinal mucosa all significantly decreased in the HPM and MWM groups. However, there were no significant differences between the HPM and MWM groups.ConclusionThe moxibustion therapies (HPM and MWM) could reduce intestinal inflammation and restore intestinal epithelium barrier disruption in CD, which might be due to down-regulating TNF-α, TNFR1, and TNFR2 in intestinal mucosa and improving intestinal epithelium morphology.

Comparison of Electroacupuncture and Moxibustion for Relieving Visceral Hypersensitivity in Rats with Constipation-Predominant Irritable Bowel Syndrome

Aim. To compare whether there is different effect between electroacupuncture (EA) and moxibustion (Mox) on visceral hypersensitivity (their analgesic effects) in constipation-predominant irritable bowel syndrome (C-IBS). Methods. EA at 1 mA and 3 mA and Mox at 43°C and 46°C were applied to the Shangjuxu (ST37, bilateral) acupoint in rats with C-IBS and normal rats. An abdominal withdrawal reflex (AWR) score was used to assess visceral hypersensitivity. Toluidine blue staining was used to assess mast cell (MC) activity in colon of rats. Immunochemistry was used to measure 5-HT and 5-HT4 receptor expression in the colon. Results. AWR scores in all EA (1 mA and 3 mA) and Mox (43°C and 46°C) treatment groups after colorectal distention (CRD) stimulation pressure of 20, 40, 60, and 80 mmHg were significantly lower than those of the model (MC) group (P all < 0.01). The MC counts and degranulation rates in the colon of all EA and Mox treatment groups and the MC group were significantly higher than those of the NC group (P all < 0.01). MC degranulation rates in the colon of all EA and Mox treatment groups were lower than those of the MC group (P all < 0.05). 5-HT expression in colon of all EA and Mox treatment groups was significantly lower than that of the MC group (P all < 0.01), and 5-HT4R expression in colon of both EA groups was significantly higher than that of the MC group (P both < 0.01). Conclusion. EA and Mox treatments may both ameliorate visceral hypersensitivity at different degree in rats with C-IBS, and EA treatment was better than Mox.

Suspended moxibustion at Tianshu (ST25) inhibits prokineticin 1 and prokineticin receptor 1 expression in the spinal cord of rats with chronic visceral hypersensitivity.

Suspended moxibustion can decrease the expression of prokineticin 1 and its receptor in colonic tissue from rats modeling chronic visceral hyperalgesia. This study aimed to verify if rat spinal cord prokineticin 1 and its receptor contribute to the analgesic effect of suspended moxibustion in a rat model of irritable bowel syndrome where rats display chronic visceral hypersensitivity. Results showed that suspended moxibustion at Tianshu (ST25) point significantly decreased visceral sensitivity to colorectal distention in a chronic visceral hyperalgesia rat model; also protein and mRNA expression of prokineticin 1 and prokineticin receptor 1 in the spinal cord of rats was significantly decreased. Experimental findings indicate that prokineticin 1 and prokineticin receptor 1 are involved in the analgesia using suspended moxibustion in rats with chronic visceral hyperalgesia.

Effect of moxibustion on CRF and CRFR1 expressions in hypothalamus of TNBS-induced experimental colitis rats

ObjectiveTo observe the effect of moxibustion on the protein and mRNA expressions of corticotropin-releasing factor (CRF) and corticotropin-releasing factor receptor 1 (CRFR1) in hypothalamus of trinitrobenzene sulfonic acid (TNBS)-induced experimental colitis rats, and to explore the central mechanisms of moxibustion in improving visceral pain and the pain-related emotions in experimental colitis rats.MethodsThirty-six Sprague-Dawley (SD) rats were randomly divided into a normal group (NG), a model group (MG), a herb-partitioned moxibustion group (HPMG) and a sham herb-partitioned moxibustion group (SHPMG). Except the NG, rats in the remaining three groups all received TNBS enema to establish experimental colitis models. The HPMG received herb-partitioned moxibustion (HPM) at bilateral Tianshu (ST 25) and Qihai (CV 6) for intervention; for the SHPMG, the herbal cakes and moxa cones were only placed on the acupoints but not ignited; rats in the MG and NG were only fixed in the same way as those in the HPMG but did not receive any treatment. At the end of the intervention, the abdominal withdrawal reflex (AWR) score, the open field test (OFT) score and the elevated plus maze (EPM) score were observed to measure the changes in visceral pain and pain-related emotions of the rats. The enzyme-linked immunosorbent assay (ELISA) was used to examine the expressions of CRF and CRFR1 proteins in hypothalamus; the fluorescence-based quantitative polymerase chain reaction (PCR) was used to detect the expressions of CRF and CRFR1 mRNAs in hypothalamus.ResultsCompared with the NG, the AWR score increased significantly and the OFT and EPM scores dropped significantly in the MG (all P<0.05), and the expressions of hypothalamic CRF and CRFR1 proteins and mRNAs increased significantly (all P<0.01). Compared with the MG and SHPMG, the AWR score dropped significantly and the OFT and EPM scores increased significantly in the HPMG (all P<0.01), and the expressions of hypothalamic CRF and CRFR1 proteins and mRNAs decreased significantly (all P<0.05). There were no significant differences between the MG and the SHPMG (all P>0.05).ConclusionHPM can down-regulate the abnormally increased expressions of CRF and CRFR1 proteins and mRNAs in hypothalamus of the TNBS-induced experimental colitis rats, which is plausibly one of its action mechanisms in mitigating visceral pain and the pain-related emotions in the experimental colitis rats.摘要目的观察艾灸对三硝基苯磺酸(TNBS)诱导的实验性结肠炎大鼠下丘脑促肾上腺皮质激素释放因子(CRF)及其受体1(CRFR1)蛋白及mRNA表达的影响, 探讨艾灸改善实验性结肠炎大鼠内脏痛及痛情绪的中枢机制。方法将36只Sprague-Dawley (SD)大鼠采用完全随机方法分为正常组、模型组、隔药灸组和假隔药灸组。除正常组外,其余三组大鼠均采用TNBS灌肠制备实验性结肠炎模型。隔药灸组采用双侧天枢、气海穴隔药灸治疗; 假隔药灸组仅在穴位上放置药饼和艾炷, 但不点燃艾炷; 模型组和正常组均不进行治疗, 只做与隔药灸组相同的固定。治疗结束后, 检测各组大鼠腹壁撤回反射(AWR)、旷场实验(OFT)和高架十字迷宫测试(EPM)评分, 观察各组大鼠内脏痛和痛情绪变化; 采用ELISA技术检测各组大鼠下丘脑CRF和CRFR1的蛋白含量; 应用荧光定量PCR技术检测各组大鼠下丘脑CRF和CRFR1 mRNA的表达。结果与正常组比较, 模型组大鼠AWR评分显著升高; OFT、EPM评分显著降低(均P<0.05); 下丘脑CRF和CRFR1蛋白及mRNA表达均显著升高(均P<0.01)。与模型组和假隔药灸组比较; 隔药灸组大鼠AWR评分显著降低; OFT、EPM评分显著升高(均P<0.01), 下丘脑CRF和CRFR1蛋白及mRNA表达均显著降低(均P<0.05)。假隔药灸组与模型组比较, 差异均无统计学意义(均P>0.05)。结论隔药灸能降低TNBS诱导的实验性结肠炎大鼠下丘脑异常增高的CRF和CRFR1蛋白及mRNA的表达, 该作用可能是其缓解实验性结肠炎大鼠内脏痛及痛情绪的作用机制之一。

The Role of Autophagy and Related MicroRNAs in Inflammatory Bowel Disease

Accumulating evidence demonstrates that microRNA- (miR-) mediated posttranscriptional regulation plays an important role in autophagy in inflammatory bowel disease (IBD), a disease that is difficult to manage clinically because of the associated chronic recurrent nonspecific inflammation. Research indicates that microRNAs regulate autophagy via different pathways, playing an important role in the IBD process and providing a new perspective for IBD research. Related studies have shown that miR-142-3p, miR-320, miR-192, and miR-122 target NOD2, an IBD-relevant autophagy gene, to modulate autophagy in IBD. miR-142-3p, miR-93, miR-106B, miR-30C, miR-130a, miR-346, and miR-20a regulate autophagy by targeting ATG16L1 through several different pathways. miR-196 can downregulate IRGM and suppress autophagy by inhibiting the accumulation of LC3II. During the endoplasmic reticulum stress response, miR-665, miR-375, and miR-150 modulate autophagy by regulating the unfolded protein response, which may play an important role in IBD intestinal fibrosis. Regarding autophagy-related pathways, miR-146b, miR-221-5p, miR-132, miR-223, miR-155, and miR-21 regulate NF-κB or mTOR signaling to induce or inhibit autophagy in intestinal cells by releasing anti- or proinflammatory factors, respectively.

Effect of moxibustion at Shenshu (BL 23) on the ethology, corticosterone and glucocorticoid receptor in aging rats

ObjectiveTo observe the effect of moxibustion on learning and memory abilities, corticosterone and glucocorticoid receptor (GR) in subacute aging rats.MethodsTwenty four Sprague-Dawley (SD) rats were randomly divided into a normal group, a model group and a moxibustion group, 8 rats in each group. Rats in the model group and the moxibustion group were subcutaneously injected with 25% D-galactose [125 mg/(kg·bw)] for 40 d continuous; rats in the normal group were injected with saline at the same position for 40 d continuous. Rats in the moxibustion group were given mild moxibustion at bilateral Shenshu (BL 23) at the same time of modeling; rats in the normal group and the model group were only identically grabbed without moxibustion for 40 d. The learning and memory abilities of rats were observed using the Morris water maze at the end of the experiment. Abdominal aorta blood and thymus were collected after water maze experiment. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum corticosterone level, and immunohistochemical method was used to detect the expression of thymus GR.ResultsCompared with the normal group, rats in the model group showed that a significantly longer escape latency time (P<0.01) on the third and the fourth days; number of times crossing the platform in 70 s significantly reduced (P<0.01); activity times in the fourth quadrant significantly decreased (P<0.05); serum corticosterone levels increased (P<0.01); thymus GR expression decreased (P<0.05). Compared with the model group, rats in the moxibustion group showed that the escape latency times were significantly shorter on the third, the fourth and the fifth days (P<0.01, P<0.05); number of times crossing the platform in 70 s significantly increased (P<0.05); activity times in the fourth quadrant significantly increased (P<0.05); serum corticosterone levels decreased (P<0.05); thymus GR expression increased (P<0.05).ConclusionMoxibustion could improve the learning and memory abilities of subacute aging rats, down-regulate serum corticosterone levels, and increase thymus GR content.摘要目的观察艾灸对亚急性衰老大鼠学习记忆能力和皮质酮及糖皮质激素受体(glucocorticoid receptor, GR)的影响。方法将24 只(Sprague-Dawley, SD)大鼠按随机数字表分为正常组、模型组和艾灸组, 每组8 只。模型组和艾灸组按每日125 mg/(kg·bw)于颈背部皮下注射25%的D-半乳糖, 连续40 d; 正常组大鼠同部位注射等量生理盐水, 连续40 d。艾灸组大鼠造模同时给予温和灸双侧肾俞, 正常组和模型组只做相同抓取, 不艾灸, 共40 d。实验结束, 采用Morris水迷宫实验观察大鼠学习记忆能力, 水迷宫实验结束后取腹主动脉血和胸腺, 酶联免疫吸附测定(enzyme-linked immunosorbent assay, ELISA)法检测血清皮质酮含量,免疫组化法检测胸腺GR表达情况。结果与正常组比较, 模型组大鼠逃避潜伏期时间在第3 天和第4 天显著延长(P<0.01), 70 s内穿越平台次数显著减少(P<0.01), 在第四象限的活动时间明显减少(P<0.05); 血清皮质酮含量升高(P<0.01); 胸腺GR表达下降(P<0.05)。与模型组比较, 艾灸组大鼠逃避潜伏期时间在第3 天、第4 天和第5 天显著缩短(P<0.01 或P<0.05), 70 s内穿越平台次数显著增多(P<0.05), 在第四象限的活动时间明显增多(P<0.05); 血清皮质酮含量降低(P<0.05); 胸腺GR表达升高(P<0.05)。结论艾灸可改善亚急性衰老大鼠学习记忆能力, 下调血清皮质酮含量, 上调胸腺GR含量。

Influence of moxa smoke on mitochondrial transmembrane potential and Bax/Bcl-2 in alveolar type II epithelial A549 cells

ObjectiveTo investigate the influence of moxibustion products on mitochondrial transmembrane potential (MTP) and mRNA expression of Bax/Bcl-2 in alveolar type II epithelial A549 cells, and to further explore influence of moxibustion products on the oxidative damage of A549 cells.MethodsSmoke and particles generated by moxibustion were collected using the filter box for gas sampling. The moxa smoke extract (MSE) was diluted sequentially to the final concentrations of 0.05 mg/mL, 0.1 mg/mL, 0.2 mg/mL, 0.3 mg/mL and 0.4 mg/mL using the cell culture medium, and A549 cells were then intervened by the above MSE solution. Cell MTP was detected by JC-1 staining. Fluorescence quantitative polymerase chain reaction (PCR) was used to detect Bax/Bcl-2 mRNA expression of A549 cells.ResultsCompared with cells in the normal control group, MTP was significantly decreased in cells of 0.3 mg/mL and 0.4 mg/mL MSE intervention groups (P<0.01); while MTP showed no significant changes in cells of 0.05 mg/mL, 0.1 mg/mL and 0.2 mg/mL MSE intervention groups (P>0.05); compared with cells in 0.05 mg/mL MSE intervention group, MTP was decreased significantly in cells of 0.1 mg/mL, 0.2 mg/mL, 0.3 mg/mL and 0.4 mg/mL MSE intervention groups (P<0.05 ); compared with cells in 0.1 mg/mL MSE intervention group, MTP was decreased significantly in cells of 0.4 mg/mL MSE intervention group (P<0.01). Bax mRNA expression of cells in each concentration of MSE intervention group all showed no significant difference compared to that in the normal control group; Bcl-2 mRNA expression of cells was reduced with the increase of MSE intervention concentration. Wherein, Bcl-2 mRNA expressions of cells in 0.4 mg/mL and 0.3 mg/mL MSE intervention groups were significantly reduced compared with that of cells in the normal control group (P<0.05); Bcl-2 mRNA expression of cells in 0.4 mg/mL MSE intervention group was significantly reduced compared to that in 0.05 mg/mL MSE intervention group (P<0.05).ConclusionCertain higher concentration of moxa smoke could reduce MTP and mRNA expression of the anti-apoptosis gene Bcl-2 in alveolar type II epithelial A549 cells. Oxidative damage may be the important mechanism of apoptosis caused by the high concentration of moxa smoke solution, and further studies are necessary on the specific mechanisms.摘要目的观察艾灸生成物对肺泡II型上皮细胞A549 线粒体膜电位(mitochondrial transmembrane potential, MTP)、Bax/Bcl-2 mRNA 表达的影响, 进一步研究艾灸生成物对A549 细胞的氧化损伤作用。方法采用气体采样 滤盒采集艾灸生成的烟气颗粒物, 用细胞培养液稀释后依次配制质量浓度为0.05 mg/mL、0.1 mg/mL、0.2 mg/mL、 0.3 mg/mL、0.4 mg/mL 的艾烟提取物(moxa smoke extract, MSE)溶液干预A549 细胞。采用JC-1 染色法检测细胞 MTP, 荧光定量聚合酶链反应(polymerase chain reaction, PCR)法检测细胞Bax/Bcl-2 mRNA 的表达。结果与正常 对照组细胞相比, 0.3 mg/mL 和0.4 mg/mL MSE 干预组细胞MTP 显著降低 (P<0.01), 0.05 mg/mL、0.1 mg/mL 和 0.2 mg/mL MSE 干预组细胞MTP 无显著变化(P>0.05); 与0.05 mg/mL MSE 干预组细胞相比, 0.1 mg/mL、 0.2 mg/mL、0.3 mg/mL 和0.4 mg/mL MSE 干预组细胞MTP 显著降低(均P<0.05); 与0.1 mg/mL MSE 干预组细胞比较, 0.4 mg/mL MSE 干预组细胞MTP 显著降低(P<0.01)。各浓度MSE 干预组细胞Bax mRNA 的表达与正常对照 组相比均无显著性差异; 细胞内Bcl-2 mRNA 表达随MSE 干预浓度的增加而减少, 其中0.4 mg/mL 和0.3 mg/mL MSE 干预组细胞与正常对照组细胞Bcl-2 mRNA 的表达相比显著降低(P<0.05), 0.4 mg/mL MSE 干预组细胞Bcl-2 mRNA 的表达与0.05 mg/mL MSE 干预组细胞内Bcl-2 mRNA 的表达相比显著降低(P<0.05)。结论艾烟升高到一定浓度时可以降低肺泡II型上皮细胞A549 细胞内MTP, 减少凋亡抑制基因Bcl-2 mRNA 的表达, 氧化损伤可能是 高浓度艾烟溶液引起细胞凋亡的重要机制, 具体机制仍需进一步研究。