针灸对CIA 大鼠的脾脏单个核细胞IL-17、IL-12 表达的影响

目的：探讨针刺、艾灸对胶原诱导型关节炎大鼠（CIA）的脾淋巴细胞增殖能力及脾脏单个核细胞IL-17、IL-12等表达的影响。方法：将Wister大鼠随机分成正常组、模型组、艾灸组、针刺组，每组8只。采用免疫学方法制备胶原诱导型关节炎模型大鼠。艾灸组、针刺组于初次免疫后第12天，选取双侧阳陵泉、昆仑穴分别施温和灸、针刺干预。治疗结束后，观察各组大鼠四肢关节炎症评分；应用MTT比色法观察针灸对脾淋巴细胞增殖能力的影响；采用实时定量PCR技术检测脾脏单个核细胞IL-17、IL-12、TNF-α、IL-6、和IFN-γ mRNA表达的变化。结果：与正常组比较，模型组大鼠关节炎症评分明显增加（P<0.01）；与模型组比较，艾灸组、针刺组大鼠关节炎症评分均明显低于模型组（P<0.01）。与正常组比较，模型组脾淋巴细胞的增殖能力增高（P<0.01），脾脏单个核细胞IL-17、IL-12、TNF-α、IL-6、IFN-γ mRNA表达显著增加（P<0.01）；与模型组比较，艾灸组脾淋巴细胞的增殖能力下降（P<0.01），针刺和艾灸均能显著降低脾脏单个核细胞IL-12、IL-17、TNF-α、IL-6、IFN-γ mRNA的表达（P<0.01）；与艾灸组比较，针刺组在降低脾脏单个核细胞的IFN-γ mRNA的表达上优于艾灸组（P<0.05）。结论：针刺和艾灸治疗对CIA大鼠四肢关节炎症均具有改善作用，可能与艾灸能有效降低脾淋巴细胞增殖能力，针刺和艾灸均能调节脾单个核细胞异常变化的炎症细胞因子mRNA表达水平有关。     

Comparative study of electroacupuncture and moxibustion in influencing Tianshu (ST 25) regions mast cells in visceral hyperalgesia rats

Objective

To evaluate and compare electroacupunctures (EA) with different parameters and moxibustion at different temperatures influencing the activation of mast cells (MC) in Tianshu (ST 25) regions of visceral hyperalgesia model rats.

Methods

Rats (except for model group) respectively accepted 1 mA or 3 mA EA or moxibustion at 43 °C or 4 °C to stimulate Tianshu (ST 25) points after randomization of the fifty visceral hyperalgesia model rats, and then were compared with that in model and normal groups. Number, degranulation numbers, degranulation rates in Tianshu (ST 25) regions MC of rats in each group were observed using toluidine blue staining. Abdominal withdrawl reflex (AWR) score was used to evaluate the rat visceral hyperalgesia reactions.

Results

Compared with the normal group and the model group, MC numbers (P<0.05, P<0.01, P<0.01, P<0.01), degranulation numbers and degranulation rates (P<0.01, P<0.01, P<0.05, P<0.01) of Tianshu (ST 25) MC in regions tissues in 43 °C and 4 °C moxibustion groups, and 1 mA and 3 mA EA groups all increased significantly. Compared with the model group, AWR scores were significantly lower in 43 °C and 4 °C moxibustion groups, and 1 mA and 3 mA EA groups under the stimulation of 20 mmHg, 40 mmHg, 0 mmHg or 80 mmHg colorectal distension (CRD) (P<0.05 in 1 mA and 3 mA EA groups under the stimulation of 20 mmHg, P<0.01 in the other groups). AWR scores in 43 °C and 4 °C oxibustion groups under the stimulation of 20 mmHg, 40 mmHg, 0 mmHg or 80 mmHg CRD were not significantly different from those in the normal group (all P>0.05); AWR scores in 1 mA EA group under the stimulation of 0 mmHg or 80 mmHg were significantly higher than that in the normal group (P<0.01); AWR score in 3 mA EA group under the stimulation of 0 mmHg was significantly higher than that in the normal group (P<0.01), and AWR scores in 3 mA EA group under the stimulation of 20 mmHg or 80 mmHg were also higher than that in the normal group (P<0.05). AWR scores were higher in 1 mA EA group under the stimulation of 40 mmHg or 80 mmHg than that in 4 °C moxibustion group (P<0.05); AWR score was higher in 3 mA EA group under the stimulation of 40 mmHg than that in 4 °C moxibustion group (P<0.05).

Conclusion

There are differences among EA of different parameters and moxibustion of different temperatures in activating on Tianshu (ST 25) regions MC of visceral hyperalgesia model rats, as well as in improving the visceral hyperalgesia reaction. The effect of 4 °C moxibustion is the most significant.

     

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

Objective

To 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.

Methods

Smoke 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.

Results

Compared 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).

Conclusion

Certain 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.

     

隔药灸对溃疡性结肠炎大鼠结肠黏膜MyD88及TRAF6表达的影响

目的：观察隔药灸对UC大鼠结肠黏膜MyD88与TRAF6表达的影响,探讨隔药灸治疗UC的效应机制。方法：采用免疫学合并局部刺激方法复制实验性溃疡性结肠炎大鼠模型。将40只SD大鼠随机分为正常组（NC）、模型组（MC）、隔药灸组（HMP）、维柳芬组（SASP）,每组10只。隔药灸组于双侧天枢穴、气海穴施以隔药灸治疗,2壮／穴,1次／d,共8d;维柳芬组以柳氮磺胺吡啶（SASP）溶液灌胃,2次／d,共8d;正常组、模型组大鼠同步饲养,做与各干预组相同的抓取固定。干预结束后,收集结肠组织,免疫组化法检测结肠黏膜MyD88、TRAF6的表达,半定量图像分析。结果：隔药灸可明显改善UC大鼠结肠黏膜损伤程度及病理学评分（P〈0．01）,且与维柳芬作用相当（P〉0．05）;模型大鼠结肠黏膜MyD88与TRAF5的表达较正常大鼠明显增高（P〈0．01）,隔药灸干预后,结肠黏膜MyD88与TRAF6的表达显著降低（P〈0．01）,作用与维柳芬相当（P〉0．05）。结论：调控TLR／MyD88／NF—KB信号转导通路中MyD88、TRAF6的表达是隔药灸调节溃疡性结肠炎结肠黏膜局部过激免疫炎症反应的作用途径之一。     

隔药灸对急性胃黏膜损伤家兔穴区线粒体及I型胶原蛋白的影响

目的：观察隔药灸对急性胃黏膜损伤家兔胃黏膜损伤指数、穴区局部线粒体结构、I型胶原蛋白表达的影响,从胃黏膜修复及腧穴局部能量代谢角度探讨隔药灸对急性胃黏膜损伤的起效机制。方法：新西兰兔随机分为正常组、模型组、隔药灸组和西药组。在成功制备急性胃黏膜损伤模型的基础上,隔药灸组取“后三里穴”进行治疗,西药组以硫糖铝灌胃治疗。各组治疗干预结束后,留取胃黏膜和穴区局部组织,采用Guth法对各组的胃黏膜损伤指数进行评分,透射电镜观察后三里穴位局部组织线粒体形态结构,免疫组化法检测穴区I型胶原蛋白的表达。结果：与正常组比较,模型组、西药组和隔药灸组家兔胃黏膜损伤指数均显著升高(均P<0.01);模型组穴区细胞线粒体空泡化,I型胶原蛋白表达降低（P<0.01）;隔药灸组仅有少数线粒体水肿;西药组线粒体数目少、出现肿胀,线粒体嵴不明显。与模型组比较,隔药灸组和西药组胃黏膜损伤指数显著下降(均P<0.01),I型胶原蛋白表达升高（P<0.01或P<0.05）。隔药灸组胃黏膜损伤指数低于西药组(P<0.01),I型胶原蛋白表达高于西药组,但差异无统计学意义（P>0.05）。结论：隔药灸能够改善急性胃黏膜损伤家兔“后三里”穴穴区线粒体形态、提高急性胃黏膜损伤家兔后三里穴局部组织的I型胶原蛋白的表达,这可能是其修复胃黏膜损伤的起效途径之一。     

表观遗传修饰与溃疡性结肠炎

背景:基于溃疡性结肠炎发病因素的研究发现,其发生、发展是一个相当复杂的过程,涉及易感性基因响应肠道微生物内环境及其他外环境因素的变化,既可引发对机体固有免疫、特异免疫的保护和耐受,又可引起慢性炎症。目的:分析近10年溃疡性结肠炎与表观遗传修饰的相关研究进展,探讨表观遗传修饰在溃疡性结肠炎疾病中的发生、发展相关性。方法:以下列检索词:溃疡性结肠炎,表观遗传,DNA甲基化,组蛋白修饰,miRNA,ulcerative colitis,epigenetic,DNA methylation,Histone modification,miRNA,检索2000年1月至2015年1月PubMed数据库及万方医学网相关文献,根据纳入排除标准,对纳入的28条文献进行归纳分析。结果与结论:溃疡性结肠炎的发病主要是易感基因与内、外环境因素相关,受表观遗传修饰的影响。表观遗传修饰主要包括DNA甲基化、组蛋白修饰、miRNA所介导的沉默。基于溃疡性结肠炎的发病因素研究表明表观遗传修饰影响基因和环境因素之间相互作用,可作用在机体先天免疫及特异性免疫,在溃疡性结肠炎的发生、发展中具有非常重要的作用。     

基于数据挖掘技术分析腹泻型和便秘型肠易激惹综合征针灸使用规律

目的:通过文献分析临床针灸治疗腹泻型(D-IBS)和便秘型肠易激惹综合征(C-IBS)的经脉、腧穴选择特点和规律。方法:检索中国知网、万方、维普、Pubmed、Web of Science等数据库,收集2000—2016年针灸治疗D-IBS和C-IBS的临床研究报道,提取其所使用的针灸方式、经脉和腧穴。运用描述性分析、关联性分析和聚类分析研究其内在联系与规律规律,同时观察针灸治疗D-IBS和C-IBS的经脉、腧穴各自的规律和它们之间的差异。结果:共纳入针灸治疗D-IBS文献56篇、针灸治疗C-IBS文献6篇,D-IBS的治疗主要用针刺,临床以胃经和任脉的腧穴为主,如天枢、足三里、中脘,关联性分析发现胃经—任脉—脾经的为最常见的经脉组合,腧穴以足三里—天枢—太冲为主要关联群。聚类分析经脉主要分成2个核心聚类群,2个核心群组,腧穴则可以分成5个聚类群,5个核心群组。而C-IBS的研究则主要以悬灸为主,胃经、膀胱经的腧穴应用最多,如天枢、足三里、大肠俞、上巨虚,关联性分析发现经脉组合与D-IBS相同,也是胃经—任脉—脾经组合支持度和置信度最高,而腧穴则是以足三里—天枢的配伍最为常见。聚类分析则发现经脉、腧穴均可以分成2个核心聚类群,2个核心群组。结论:通过分析针灸治疗D-IBS和C-IBS的临床研究,认为虽然两者在针灸方法的使用上略微存在差异,D-IBS针灸并用,C-IBS以灸为主。但是在经脉和腧穴选择上却存在高度相似性。     

针灸对IBS内脏痛大鼠结肠肌间神经丛神经元P2X_2受体表达的调节

目的观察针灸对肠易激综合征(IBS)内脏痛大鼠结肠肌间神经丛神经元嘌呤受体亚型P2X2表达的调节。方法应用直结肠球囊扩张刺激法制备慢性内脏高敏感IBS大鼠模型,随机分为正常组、模型组、电针组,电针组采用电针"天枢""上巨虚"穴进行治疗。治疗结束后,进行腹部撤回反射评分(AWR),应用免疫荧光法检测肠结肠肌间神经丛P2X2表达变化。结果模型大鼠AWR评分升高,电针治疗后降低;模型组大鼠结肠肌间神经丛神经元中P2X2表达与正常组大鼠相比异常增加,电针组显著降低。结论电针(天枢、上巨虚)能抑制结肠肌间神经丛神经元P2X2受体异常表达,改善IBS大鼠的内脏痛觉敏感状态。     

温和灸对溃疡性结肠炎大鼠天枢穴P2RX7的作用机理研究

目的：溃疡性结肠炎（UC）是艾灸治疗的有效病症,本研究从艾灸作用的始动环节与穴位响应密切相关的角度,探讨温和灸的对穴区P2X嘌呤受体7表达的影响。方法：将20只SD雄性大鼠随机分成正常组6只、模型组14只。采用自由饮用4%葡聚糖硫酸钠（DSS）法建立UC模型,连续饮用7天后,取2只大鼠进行模型鉴定,造模成功后将剩下的12只大鼠随机分成UC组、UC+温和灸组,第8天改为自由饮用1%DSS溶液,第8天开始采用温和灸天枢穴（双）干预7天,每天10min。疗程结束后,称量大鼠体质量、取双侧天枢穴皮肤,采用HE 染色观察病理组织学形态变化,采用免疫组化法对P2RX7 的蛋白检测,从分子水平观察温和灸对天枢穴皮肤的作用;采用qRT-PCR的方法检测P2RX₇在天枢穴皮肤中的表达。结果：与正常组比较,UC组体质量于第4天开始明显降低（P<0.05）,天枢穴P2RX₇基因的mRNA表达水平降低（P<0.05）,天枢穴皮肤中主要分布在真皮层皮脂腺中的P2RX₇明显减少（P<0.05）。与UC组比较,UC+温和灸组在第8天明显增加（P<0.05）,天枢穴P2RX₇基因mRNA表达水平增高（P<0.05）,天枢穴皮肤P2RX₇主要分布在皮脂腺中的明显增加（P<0.05）。温和灸干预后所增加体重与天枢穴P2RX₇免疫组化IOD值的相关系数r=0.823（P<0.05）。结论：温和灸能明显增加UC大鼠体重,其起效的机制可能与上调天枢穴位皮肤P2X嘌呤受体7密切相关。     

保健灸的适宜时机探要

保健灸与介入年龄阶段、四时之气、治未病等方面对保健灸的适宜时机选择进行探讨,结果发现,儿童阶段,步入衰老阶段,老年阶段是保健灸介入的适宜年龄阶段;夏季三伏、冬季三九及24节气是保健灸介入的适宜自然时机;疾病先兆期、疾病早期是保健灸治未病的适宜介入时机。