Herbs-partitioned moxibustion alleviates aberrant intestinal epithelial cell apoptosis by upregulating A20 expression in a mouse model of Crohn's disease

BACKGROUND A20 inhibits intestinal epithelial cell apoptosis in Crohn's disease, and herbspartitioned moxibustion(HPM) has been demonstrated to be an effective treatment for Crohn's disease. However, the mechanism by which HPM reduces intestinal epithelial cell apoptosis in Crohn's disease has not been thoroughly elucidated to date.AIM To elucidate whether HPM exerts its effects by upregulating A20 to affect intestinal epithelial cell apoptosis in a Crohn's disease mouse model.METHODS In this study, mice with A20 deletion in intestinal epithelial cells(A20 IEC-KO) were utilized to establish a Crohn's disease mouse model with 2,4,6-trinitrobenzenesulfonic acid(TNBS) administration, as well as wild-type mice. Mice were randomly divided into normal control(NC), model control(MC), mesalazine(MESA), and HPM groups. The morphology of the colonic mucosa was observed by hematoxylin-eosin staining, and serum endotoxin and apoptosis of epithelial cells were evaluated by enzyme-linked immunosorbent assay and terminal dUTP nick-end labeling assay accordingly. The protein expression levels of A20 and tumor necrosis factor receptor 1(TNFR1)-related signaling molecules were evaluated by Western blot, and co-expression of A20 and TNFR1-associated death domain(TRADD) and co-expression of A20 and receptor-interacting protein 1(RIP1) were observed by double immunofluorescence staining.RESULTS The intestinal epithelial barrier was noted to have an improvement in the HPM group of wild-type(WT) mice compared with that in A20 IEC-KO mice. Compared with A20 IEC-KO HPM mice, serum endotoxin levels and apoptosis percentages were decreased(P < 0.01), A20 expression levels were increased(P < 0.01), and expression of TNFR1, TRADDD, and RIP1 was decreased in the HPM group of WT mice(PTNFR1 < 0.05, PTRADD < 0.01, PRIP1 < 0.01). Both of the co-expression of A20/TRADD and A20/RIP1 showed a predominantly yellow fluorescence in the HPM group of WT mice, while a predominantly red fluorescence was noted in the HPM group of A20 IEC-KO mice.CONCLUSION Our findings suggest that HPM in treating Crohn's disease functions possibly via upregulation of the A20 expression level, resulting in downregulation of TNFR1,TRADD, and RIP1 to alleviate increased cell apoptosis in the intestinal epithelial barrier in Crohn's disease.

Herb-partitioned moxibustion alleviates colon injuries in ulcerative colitis rats

AIM To observe the effect of herb-partitioned moxibustion(HPM) on expression of colonic cytokines in ulcerative colitis(UC) rats.METHODS A UC rat model was established by protein immunization in combination with topical chemical stimulation.Rats in the HPM group(n = 8) received HPM at bilateral Tianshu(ST25) points.The gross injury and pathological scores of the colon were recorded.The expression profile of colonic cytokines was assayed using the protein microarray technique.Specific differential cytokines were selected and verified by ELISA.The corresponding Uni Prot Accessions of the differentially expressed cytokines were retrieved in the Uni Prot database.The pathways involved were analyzed with the help of the KEGG PATHWAY database.The DAVID database was used for functional cluster and pathway analysis.RESULTS HPM improved colon injuries in UC rats,manifested by accelerated repair of ulcers and alleviation of inflammation,and the gross injury and pathological scores both significantly decreased(P < 0.01).Fold change > 1.3 or < 0.77 was taken as the screening standard.There were 77 down-regulated and 9 up-regulated differentially expressed colonic cytokines in the HPM group compared with the model group,and expression of 20 differed significantly(P < 0.05).Twelve of the 20 significantly differentially expressed cytokines [β-catenin,interleukin-1 receptor 6(IL-1 R6),IL-1β,B7-1,nerve growth factor receptor,AMP-activated protein kinase-α1,neuropilin-2,orexin A,adipocyte differentiation-related protein,IL-2,Fas and Fas L] were up-regulated in the model group(n = 3,compared with the normal group) but downregulated in the HPM group(n = 3,compared with the model group).Functional cluster analysis showed that the differentially expressed colonic cytokines in the HPM group regulated apoptosis and protein phosphorylation.KEGG pathway analysis showed that 52 down-regulated and 7 up-regulated differentially expressed colonic cytokines in the HPM group had pathways.The pathways that interacted between the cytokines and their receptors accounted for the largest proportion(28 of the downregulated and 5 of the up-regulated cytokines).CONCLUSION HPM promotes the repair of colon injuries in UC rats,which is related to the regulation of several abnormally expressed cytokines.

Extracellular signal-regulated kinase, substance P and neurokinin-1 are involved in the analgesic mechanism of herb-partitioned moxibustion

Herb-partitioned moxibustion can effectively mitigate visceral pain, a major symptom in inflammatory bowel disease, but the analgesic lnechanism is still unclear. Moreover, extracellular signal-regulated kinase, substance P, and neurokinin-1 are involved in formation of central hyperalgesia. Thus, we postulated that the analgesic effect of herb-partitioned moxibustion may be associated with these factors. Accordingly, in this study, we established an inflammatory bowel disease visceral pain model in rat by enema with a mixed solution of 5% trinitrobenzenesulfonic acid and 50% ethanol. Bilateral Tianshu （ST25） and Qihai （CV6） points were selected for herb-partitioned moxi- bustion. Our results showed that herb-partitioned moxibustion improved visceral pain and down-regulated extracellular signal-regulated kinase, substance P, and neurokinin-1 protein and mRNA expression in dorsal root ganglia. These results indicate that down-regulation of extracellular signal-regulated kinase, substance E and neurokinin-1 protein and mRNA may be a central mechanism for the analgesic effect of herb-partitioned moxibustion.

Moxibustion inhibits interleukin-12 and tumor necrosis factor alpha and modulates intestinal flora in rat with ulcerative colitis

AIM: To investigate the effect of moxibustion on intestinal flora and release of interleukin-12 (IL-12) and tumor necrosis factor-α (TNF-α) from the colon in rat with ulcerative colitis (UC). METHODS: A rat model of UC was established by local stimulation of the intestine with supernatant from colonic contents harvested from human UC patients. A total of 40 male Sprague-Dawley rats were randomly divided into the following groups: normal (sham), model (UC), herb-partition moxibustion (HPM-treated), and positive control sulfasalazine (SA-treated). Rats treated with HPM received HPM at acupuncture points ST25 and RN6, once a day for 15 min, for a total of 8 d. Rats in the SA group were perfused with SA twice a day for 8 d. The colonic histopathology was observed by hematoxylin-eosin. The levels of intestinal flora, including Bifidobacterium, Lactobacillus, Escherichia coli (E. coli), and Bacteroides fragilis (B. fragilis), were tested by real-time quantitative polymerase chain reaction to detect bacterial 16S rRNA/DNA in order to determine DNA copy numbers of each specific species. Immunohistochemical assays were used to observe the expression of TNF-α and IL-12 in the rat colons. RESULTS: HPM treatment inhibited immunopathology in colonic tissues of UC rats; the general morphological score and the immunopathological score were significantly decreased in the HPM and SA groups compared with the model group [3.5 (2.0-4.0), 3.0 (1.5-3.5) vs 6.0 (5.5-7.0), P < 0.05 for the general morphological score, and 3.00 (2.00-3.50), 3.00 (2.50-3.50) vs 5.00 (4.50-5.50), P < 0.01 for the immunopathological score]. As measured by DNA copy number, we found that Bifidobacterium and Lactobacillus, which are associated with a healthy colon, were significantly higher in the HPM and SA groups than in the model group (1.395 ± 1.339, 1.461 ± 1.152 vs 0.045 ± 0.036, P < 0.01 for Bifidobacterium, and 0.395 ± 0.325, 0.851 ± 0.651 vs 0.0015 ± 0.0014, P < 0.01 for Lactobacillus). On the other hand, E. coli and B. fragilis, which are associated with an inflamed colon, were significantly lower in the HPM and SA groups than in the model group (0.244 ± 0.107, 0.628 ± 0.257 vs 1.691 ± 0.683, P < 0.01 for E. coli, and 0.351 ± 0.181, 0.416 ± 0.329 vs 1.285 ± 1.039, P < 0.01 for B. fragilis). The expression of TNF-α and IL-12 was decreased after HPM and SA treatment as compared to UC model alone (4970.81 ± 959.78, 6635.45 ± 1135.16 vs 12333.81 ± 680.79, P < 0.01 for TNF-α, and 5528.75 ± 1245.72, 7477.38 ± 1259.16 vs 12550.29 ± 1973.30, P < 0.01 for IL-12). CONCLUSION: HPM treatment can regulate intestinal flora and inhibit the expression of TNF-α and IL-12 in the colon tissues of UC rats, indicating that HPM can improve colonic immune response.

Moxibustion combined with acupuncture increases tight junction protein expression in Crohn's disease patients

AIM:To investigate the effect of herb-partitioned moxibustion combined with acupuncture on the expression of intestinal epithelial tight junction(TJ) proteins.METHODS:Sixty patients diagnosed with mild to moderate Crohn’s disease(CD)were allocated into the herb-partitioned moxibustion combined with acupuncture(HMA)group(n=30)or the mesalazine(MESA)group(n=30)using a parallel control method.There were 2 sets of acupoints used alternately for HMA treatment.The following points were included in Set A:ST25(Tianshu),RN6(Qihai),and RN9(Shuifen)for herb-partitioned moxibustion and ST36(Zusanli),ST37(Shangjuxu),LI11(Quchi),and LI4(Hegu)for acupuncture.The points for Set B included BL23(Shenshu)and BL25(Dachangshu)for herb-partitioned moxibustion and EX-B2 of T6-T1(Jiajixue)fo r acupuncture.The patients received the same treatment6 times a week for 12 consecutive weeks.The MESA group received 1 g of mesalazine enteric coated tablets4 times daily for 12 consecutive weeks.Intestinaltissues were stained and examined to compare the morphological and ultrastructural changes before and after the treatment session.Immunohistochemistry and in situ hybridization assays were used to detect the expression of intestinal epithelial TJ proteins zonula occludens-1(ZO-1),occludin,and claudin-1.The m RNA levels were also evaluated.RESULTS:After the treatment,both herb-partitioned moxibustion combined with acupuncture and mesalazine improved intestinal morphology and ultrastructure of CD patients;the patients treated with HMA showed better improvement.HMA significantly increased the expression of ZO-1(P=0.000),occludin(P=0.021),and claudin-1(P=0.016).MESA significantly increased the expression of ZO-1(P=0.016)and occludin(P=0.026).However,there was no significant increase in the expression of claudin-1(P=0.935).There was no statistically significant difference between the two groups for the expression of occludin and claudin-1(P>0.05).The HMA group showed a significant improvement in ZO-1 expression compared to the MESA group(2333.34±352.51 vs 2160.38±307.08,P=0.047).HMA significantly increased the expression of ZO-1 m RNA(P=0.000),occludin m RNA(P=0.017),and claudin-1 m RNA(P=0.017).MESA significantly increased the expression of ZO-1 m RNA(P=0.000),occludin m RNA(P=0.042),and claudin-1 m RNA(P=0.041).There was no statistically significant difference between the two groups in the expression of occludin and claudin-1 m RNA(P>0.05).However,the HMA group showed a significant improvement in ZO-1 m RNA expression compared with the MESA group(2378.17±308.77 vs 2200.56±281.88,P=0.023).CONCLUSION:HMA can repair intestinal epithelial barrier lesions and relieve inflammation by upregulating the expression of TJ proteins and their m RNAs.

Effect of herb-partitioned moxibustion on dopamine levels and dopamine receptor 1 expression in the colon and central nervous system in rats with Crohn’s disease

OBJECTIVE:To observe the effect of herb-partitioned moxibustion at the Tianshu (ST 25) and Qihai (CV 6) acupoints in rats with Crohn's disease,and explore the underlying mechanism from dopamine (DA) and dopamine receptor 1 (D1 R) in the colon,spinal dorsal horn and hypothalamus.METHODS:The rats were randomly divided into the normal,model (CD),herb-partitioned moxibustion (Mox) and mesalazine (Mesa) groups.Damage in the colons was scored and observed by hematoxylin and eosin staining.DA and D1R protein expression in the colonic mucosa were detected by immunohistochemistry.The concentrations of DA and D1R in the spinal dorsal horn and hypothalamus were measured by enzyme-linked immunosorbent assay,and D1R mRNA expression was evaluated by quantitative real-time polymerase chain reaction.RESULTS:In the colon,compared with the normal group,DA,D1 R protein expressions and D1 R mRNA expression were significantly higher in the model group,while decreased in the Mox group and the Mesa group.In the spinal dorsal horn and hypothalamus,compared with the normal group,the concentrations of DA and D1 R,and the D1R mRNA expressions were significantly higher in the model group,and decreased in the Mox group and the Mesa group.CONCLUSION:Herb-partitioned moxibustion at the Tianshu (ST 25) and Qihai (CV 6) acupoints relieved ulceration in CD rats,the underlying mechanism maybe relative with the regulation of DA and D1R in the colon,spinal dorsal horn and hypothalamus by moxibustion.

Moxibustion eases chronic inflammatory visceral pain through regulating MEK, ERK and CREB in rats

AIM To investigate the effects of herb-partitioned moxibustion(HPM) on phosphorylation of mitogen-activated extracellular signal-regulated kinase(MEK)1, extracellular signal-regulated kinase(ERK)1/2 and c AMP response element binding protein(CREB) in spinal cord of rats with chronic inflammatory visceral pain(CIVP), and to explore the central mechanism of HPM in treating CIVP.METHODS Male Sprague-Dawley rats were randomized into normal, model, HPM, sham-HPM, MEK-inhibitor and dimethyl sulfoxide(DMSO) groups. The CIVP model was established using an enema mixture of trinitrobenzene sulfonic acid and ethanol. HPM was applied at bilateral Tianshu(ST25) and Qihai(CV6) acupoints in the HPM group, while in the sham-HPM group, moxa cones and herb cakes were only placed on the same points but not ignited. The MEK-inhibitor and DMSO groups received L5-L6 intrathecal injection of U0126 and 30% DMSO, respectively. Abdominal withdrawal reflex(AWR), mechanical withdrawal threshold(MWT) and thermal withdrawal latency(TWL) were applied for the assessment of pain behavior. The colonic tissue was observed under an optical microscope after hematoxylin-eosin staining. Expression of phosphor(p)MEK1, p ERK1/2 and p CREB in rat spinal cord was detected using Western blotting. The levels of MEK, ERK and CREB m RNA in rat spinal cord were detected using real-time polymerase chain reaction. RESULTS Compared with the normal group, the AWR scores were increased significantly(P < 0.01) and the MWT and TWL scores were decreased significantly(P < 0.05) in the model, sham-HPM and DMSO groups. Compared with the model group, the AWR scores were decreased significantly(P < 0.01) and the MWT and TWL scores were increased significantly in the HPM and MEK-inhibitor groups(P < 0.05). Compared with the sham-HPM and DMSO groups, the AWR scores were decreased significantly(P < 0.01) and the MWT and TWL scores were increased significantly(P < 0.05) in the HPM and MEK-inhibitor groups. Compared with the normal group, the expression of p MEK1, p ERK1/2 and p CREB proteins and the levels of MEK, ERK and CREB m RNA in rat spinal cord were increased significantly in the model, sham-HPM and DMSO groups(P < 0.01 or < 0.05). Compared with the model group, the expression of p MEK1, p ERK1/2 and p CREB proteins and the levels of MEK, ERK and CREB m RNA in rat spinal cord were reduced significantly in the HPM and MEK-inhibitor groups(P < 0.01 or < 0.05). Compared with the sham-HPM and DMSO groups, expression of p MEK1, p ERK1/2 and p CREB proteins and the levels of MEK, ERK and CREB m RNA in rat spinal cord were reduced significantly in the HPM and MEK-inhibitor groups(P < 0.01 or < 0.05). CONCLUSION HPM down-regulates protein phosphorylation of MEK1, ERK1/2 and CREB, and m RNA expression of MEK, ERK and CREB, inhibiting activation of the MEK/ERK/CREB signaling pathway in the spinal cord of CIVP rats, which is possibly a critical central mechanism of the analgesic effect of HPM.

隔药灸对慢性炎性内脏痛大鼠mTOR-4E-BP1/S6RP信号通路的调控作用

目的:从mTOR-4E-BP1/S6RP信号通路角度,探讨隔药灸治疗慢性炎性内脏痛的中枢机制。方法:将雄性SD(Sprague Dawley)大鼠随机分为正常组、模型组、隔药灸组、mTOR抑制剂组。采用TNBS合50%乙醇灌肠的方法制备CIVP大鼠模型。隔药灸组采用隔附子饼灸“天枢”(双侧)“气海”;mTOR抑制剂组在L5~L6间鞘内注射雷帕霉素。采用腹壁撤回反射评分(Abdominal Withdrawal Reflex,AWR评分)观察各组大鼠痛行为变化;采用Western Blotting及RT-qPCR技术检测各组大鼠脊髓mTOR、4E-BP1、S6RP磷酸化蛋白及mRNA表达。结果:与正常组比较,模型组大鼠AWR评分在各个压力梯度下均明显升高(均P<0.01);脊髓mTOR、4EBP1、S6RP磷酸化蛋白及mRNA表达均明显增高(P<0.01或P<0.05)。与模型组比较,隔药灸组及mTOR抑制剂组大鼠经干预治疗后,AWR评分均显著下降(P<0.01,P<0.05);脊髓mTOR、4EBP1、S6RP磷酸化蛋白及mRNA表达均明显降低(P<0.01,P<0.05)。隔药灸组AWR评分、脊髓mTOR、4EBP1、S6RP磷酸化蛋白及mRNA表达与mTOR抑制剂组比较差异均无统计学意义(P>0.05)。结论:隔药灸能显著下调慢性炎性内脏痛大鼠脊髓mTOR、4E-BP1、S6RP的磷酸化蛋白和mRNA表达水平,抑制mTOR-4E-BP1/S6RP信号通路的过度激活,这可能是隔药灸发挥镇痛作用的重要中枢机制之一。

基于AMPK-ERK/mTOR信号通路探讨隔药灸对慢性炎性内脏痛大鼠的镇痛机制

目的:从腺苷酸活化蛋白激酶(AMP-activated Protein Kinase,AMPK)-细胞外信号调节激酶(Extracellular Signalregulated Kinase,ERK)/雷帕霉素靶蛋白(Mechanistic Target of Rapamycin,mTOR)信号通路角度,探讨隔药灸治疗慢性炎性内脏痛(Chronic Inflammatory Visceral Pain,CIVP)的中枢机制。方法:将雄性SD大鼠随机分为正常组、模型组、隔药灸组、AMPK激活剂组。采用三硝基苯磺酸(Trinitrobenzene Sulfonic Acid,TNBS)合50%乙醇溶液灌肠的方法制备CIVP大鼠模型。隔药灸组采用隔附子饼灸天枢穴(双侧)、气海穴进行治疗;AMPK激活剂组在L5~L6间鞘内注AMPK激活剂ATCAR。采用腹壁撤回反射(Abdominal Withdrawal Reflex,AWR)评分观察各组大鼠痛行为变化;采用蛋白质印迹法(Western Blotting,WB)及实时荧光定量(Real Time Quantitative,RT-q PCR)技术检测各组大鼠脊髓磷酸化AMPK(p-AMPK)、磷酸化ERK(p-ERK)、磷酸化mTOR(p-mTOR)蛋白及m RNA表达。结果:与正常组比较,模型组大鼠AWR评分在各个压力梯度下均明显升高(均P <0.01);脊髓p-AMPK蛋白及m RNA表达均明显降低(均P <0.01);脊髓p-ERK、p-mTOR蛋白及m RNA表达均明显增高(均P <0.01)。与模型组比较,隔药灸组及AMPK激活剂组大鼠经干预治疗后,AWR评分均显著下降(P <0.01,P <0.05);脊髓p-AMPK蛋白及m RNA表达均明显升高(均P <0.01);脊髓p-ERK、p-mTOR蛋白及m RNA表达均明显降低(P <0.01)。与AMPK激活剂组比较,隔药灸组AWR评分和脊髓p-AMPK、p-ERK、p-mTOR蛋白及m RNA表达差异均无统计学意义(P> 0.05)。结论:隔药灸能显著提高CIVP大鼠脊髓p-AMPK蛋白和m RNA表达水平,抑制AMPK下游p-ERK、p-mTOR蛋白及m RNA表达,进而抑制ERK及mTOR的过度激活,这可能是隔药灸发挥镇痛作用的重要中枢机制之一。

隔药灸脐对卵巢早衰模型大鼠Bcl-2、Bax蛋白和mRNA表达的影响

目的观察隔药灸脐对卵巢早衰模型大鼠Bcl-2和Bax表达的影响,探讨其对卵巢的保护机制。方法采用腹腔注射环磷酰胺建立大鼠卵巢早衰模型。48只雌性Wistar大鼠随机分为正常组、模型组、隔药灸脐组、西药组,每组12只。隔药灸脐组大鼠隔药灸"神阙",30 min/d,隔日1次,连续14 d;西药组大鼠予戊酸雌二醇灌胃,每日1次,连续14 d。采用免疫组化、Western blot和实时荧光定量PCR检测大鼠卵巢凋亡相关因子Bcl-2、Bax蛋白和mRNA的表达。结果模型组大鼠卵巢质量明显低于正常组(P<0.05)。Bcl-2、Bax蛋白主要表达在细胞胞浆中,隔药灸脐组和西药组较模型组卵泡颗粒细胞Bax表达范围明显缩小,Bcl-2表达范围明显增加。与正常组比较,模型组大鼠卵巢Bcl-2蛋白和mRNA表达明显降低(P<0.05,P<0.01),Bax蛋白和mRNA表达显著升高(P<0.05,P<0.01);与模型组比较,隔药灸脐组和西药组大鼠卵巢Bcl-2蛋白和mRNA表达显著升高(P<0.05,P<0.01),Bax蛋白和mRNA表达明显降低(P<0.05,P<0.01)。结论隔药灸脐可能通过上调卵巢早衰模型大鼠卵巢Bcl-2和下调Bax的表达,改善卵巢功能。

隔药灸结合针刺对克罗恩病患者肠黏膜TNF-α、TNFR1、TNFR2表达及肠上皮细胞凋亡的影响

目的观察隔药灸结合针刺对克罗恩病患者肠黏膜TNFα-、TNFR1、TNFR2含量以及肠上皮细胞凋亡的影响,探讨其对克罗恩病肠上皮屏障损伤的保护作用。方法将60例符合轻、中度克罗恩病诊断标准的患者分为治疗组和对照组(各30例),治疗组予以隔药灸结合针刺治疗,12次为1个疗程,疗程间休息3天,共治疗6个疗程;对照组予以常规西药美沙拉嗪口服,连续服用12周。采用克罗恩病活动指数评价两种治疗方法的临床疗效,同时观察患者治疗前后肠黏膜TNFα-、TNFR1、TNFR2含量及肠上皮细胞凋亡情况。结果治疗组和对照组临床总有效率分别为86.67%、63.33%;组间疗效比较,差异有统计学意义(P<0.05);两组治疗后肠黏膜TNFα-、TNFR1、TNFR2含量和肠上皮细胞凋亡率均明显降低(P<0.01);组间治疗后比较,肠黏膜TNFα-、TNFR1含量及肠上皮细胞凋亡率差异有统计学意义(P<0.05)。结论隔药灸结合针刺可能是通过降低克罗恩病肠黏膜异常增高的TNFα-、TNFR1、TN-FR2,从而抑制肠上皮细胞的异常凋亡,达到保护肠上皮屏障损伤和减轻肠道炎症反应之目的。

隔物灸调控缺氧诱导因子-1α表达抑制慢性萎缩性胃炎大鼠糖酵解的机制研究

目的:观察慢性萎缩性胃炎(CAG)大鼠胃黏膜组织缺氧诱导因子-1α(HIF-1α)及糖酵解关键酶表达变化,探究隔物灸对CAG的作用及可能机制。方法:将雄性Wistar大鼠随机分为正常组和造模组,自由饮用170 mg/L浓度MNNG联合法进行CAG大鼠造模。鉴定模型成功后,造模组大鼠随机分为模型组、隔药饼灸组、隔姜灸组和西药组,每组6只。隔药饼灸组、隔姜灸组均取中脘、气海穴进行隔物灸干预,西药组给予叶酸悬浊液灌胃。采用HE染色法观察胃窦组织病理学变化,实时荧光定量PCR(RT-qPCR)法检测胃黏膜组织信号转导及转录激活因子3(STAT3)、丙酮酸激酶M2(PKM2)和HIF-1α的mRNA表达水平,免疫组织化学法检测胃黏膜组织STAT3、HIF-1α蛋白表达,比色法和ELISA法测定胃黏膜组织LDH、PKM2活性。结果:与正常组比较,模型组大鼠胃黏膜组织固有腺体结构紊乱,腺体减少,出现萎缩、肠化、假幽门腺化生和(或)异型增生。模型组大鼠胃黏膜组织HIF-1α、STAT3、PKM2 mRNA表达上调(P<0.05),STAT3、HIF-1α蛋白表达增加(P<0.05),LDH、PKM2酶活性增加(P<0.05)。与模型组比较,隔药饼灸组、隔姜灸组、西药组固有腺体排列较规整,固有腺体萎缩、肠上皮化生、异型增生程度减轻;隔药饼灸和隔姜灸均能下调大鼠胃黏膜组织中HIF-1α、STAT3 mRNA与蛋白表达(P<0.05),降低LDH、PKM2酶活性(P<0.05)。结论:隔物灸中脘、气海穴可改善CAG大鼠胃窦组织形态学病变,修复胃黏膜损伤。抑制HIF-1α介导的异常糖酵解代谢水平可能是隔物灸法治疗CAG的效应机制之一。

背俞穴隔药灸配合天灸治疗变应性鼻炎的疗效观察

目的 观察背俞穴隔药灸配合天灸治疗变应性鼻炎的临床疗效。方法 将60例变应性鼻炎患者随机分成对照组和观察组,每组30例。对照组口服氯雷他定片,观察组采用背俞穴隔药灸配合天灸。观察两组治疗前后鼻部症状总评分(total nasal symptom score, TNSS)、鼻伴随症状总分(total non-nasal symptom score,TNNSS)、症状体征总分及鼻结膜炎生活质量调查问卷(rhino-conjunctivitis quality of life questionnaire,RQLQ)评分,并比较两组临床疗效。结果 两组治疗后TNSS、TNNSS、症状体征总分和RQLQ评分较治疗前降低(P<0.05);观察组治疗后TNSS、TNNSS、症状体征总分和RQLQ评分均低于对照组(P<0.05)。对照组总有效率为83.3%,观察组总有效率为93.3%,观察组高于对照组(P<0.05)。结论 背俞穴隔药灸配合天灸可以有效改善变应鼻炎的相关临床症状,提高变应性鼻炎患者生活质量,其疗效优于口服氯雷他定。

不同灸量隔药灸治疗原发性痛经寒凝血瘀证临床观察

目的探讨不同灸量隔药灸治疗原发性痛经寒凝血瘀证的临床效果。方法选择上饶市妇幼保健院妇科于2020年1月—2021年12月收治的原发性痛经寒凝血瘀证患者68例,根据采用的方法不同将其分为观察组(34例)与对照组(34例),观察组患者每次每穴隔药灸治4壮,对照组患者每次每穴隔药灸治8壮,确保对两组患者每个月经周期施灸7次,持续3个月,比较两组治疗前后中医证候评分、视觉模拟量表(VAS)评分、持续痛经时间。结果观察组患者临床治疗有效率为91.2%(31/34),高于对照组的73.5%(25/34),但差异无统计学意义(P>0.05)。两组患者治疗前后的VAS评分及疼痛持续时间均得到显著改善(P<0.05)。结论隔药灸可改善原发性痛经寒凝血瘀证患者的疼痛症状,缩短疼痛持续时间,有很好的疗效,而不同灸量对疗效的影响不大,文章建议应用4壮治疗。

隔药饼灸调节大鼠免疫抑制机制的转录组测序分析

背景:免疫抑制会导致机体免疫功能受损,加重病情。隔药饼灸能有效调节机体免疫功能,提高机体免疫力,但其调节机制目前仍不清楚。目的:基于转录组学的生物信息学技术对隔药饼灸干预的免疫抑制模型大鼠进行测序,探究隔药饼灸调控免疫的机制。方法:将24只SD大鼠随机分为空白组、模型组和隔药饼灸组,每组8只,模型组与隔药饼灸组采用腹腔注射环磷酰胺制备免疫抑制模型,35 mg/kg,连续3 d,造模结束后空白组与模型组不做干预,隔药饼灸组在大鼠中脘、神阙、关元及足三里穴位进行艾炷与药饼结合的隔药饼灸干预,连续10 d,1次/d,干预结束后次日取材。取各组大鼠外周血进行白细胞数量检测;采用Illumina测序平台进行RNA-seq,筛选差异基因,结合GO与KEGG数据库,对差异基因进行生物信息学相关分析。结果与结论:①与空白组比较,模型组白细胞数量显著减少(P<0.001)。②RNA-seq共筛选出模型组与空白组间差异基因3026个,其中1565个上调、1461个下调,隔药饼灸组与模型组间差异基因535个,其中280个上调、255个下调。③韦恩图分析获得模型组与空白组159个下调基因经隔药饼灸干预后上调,蛋白互作网络分析获得Oasl,Oas2,Isg15,Herc6,Mx2,Helz2,Mx1,Syk,Hspa1a及Ret等10个核心靶点;④GO与KEGG分析隔药饼灸调节机体的机制有免疫应答途径、病毒相关、血管新生和自身免疫系统等通路。⑤上述结果证实,实验筛选出隔药饼灸干预免疫抑制与Oasl,Oas2,Isg15,Herc6,Mx2,Helz2及Mx1靶点有明显关联,并且在免疫应答、病毒相关及血管新生等通路中发挥调控作用。

隔药灸对溃疡性结肠炎大鼠结肠NF-κB和STAT3活性的免疫调节机制

目的:观察隔药灸改善UC大鼠结肠炎症的效应,通过NF-κB通路和STAT3磷酸化参与免疫调节的作用机制。方法:将大鼠随机分成正常组、UC组和隔药灸组,每组8只。采用4%DSS诱导UC的大鼠模型,造模成功后采用隔药灸大鼠双侧天枢穴,每穴灸2壮艾炷,每天1次,共7次。通过比较组织病理学、血清学促炎症因子的蛋白浓度观察隔药灸干预UC大鼠的效应。采用Western Blot检测结肠组织NF-κB通路及STAT3活性探讨隔药灸改善UC大鼠结肠炎症反应的潜在机制。结果:UC组大鼠结肠组织病理学提示黏膜上皮表面溃疡形成,炎症反应明显,隔药灸能修复结肠上皮损伤,减轻UC大鼠结肠组织的炎症反应。与正常组比较,UC组大鼠血清IL-1β和IL-6蛋白浓度显著升高(P <0.05,P <0.01),p-STAT3磷酸化水平显著增加,pIκB-α和NF-κB p65蛋白表达均显著增加,IκB-α蛋白表达显著降低;与UC组比较,隔药灸组大鼠血清IL-1β和IL-6蛋白浓度显著降低(P <0.05),结肠组织NF-κB p65蛋白表达和pIκB-α磷酸化水平均降低,IκB-α蛋白表达增加。结论:隔药灸可能通过抑制NF-κB通路和STAT3磷酸化,减少了促炎症因子IL-1β和IL-6的释放,从而减轻了UC大鼠的结肠炎症反应。

隔药灸对克罗恩病大鼠结肠p38MAPK、ERK1/2及c-fos调节作用的研究

目的观察隔药灸对克罗恩(Crohns Disease,CD)大鼠结肠p38MAPK、ERK1/2、c-fos的影响,探讨隔药灸治疗CD的作用机制。方法将清洁级雄性SD大鼠随机分为正常组、模型组、隔药灸组和假灸组4组。采用TNBS合50%乙醇溶液灌肠制备大鼠CD模型。模型制备成功后,隔药灸组取天枢穴(双)、气海穴进行隔药饼灸治疗;假灸组取穴、操作与隔药灸组相同,但不点燃艾炷。治疗结束后,采用HE染色,光镜下观察大鼠结肠组织形态学变化;采用实时荧光定量PCR技术检测结肠p38MAPK mRNA表达;应用ELISA技术检测结肠p38MAPK、c-fos蛋白含量,Western blot技术检测结肠ERK1/2蛋白表达。结果与正常组比较,模型组大鼠结肠组织损伤严重,可见全壁性炎症、裂隙状溃疡,部分大鼠结肠可见纤维化;与模型组、假灸组比较,隔药灸组大鼠结肠形态结构改善,肠道炎症减轻;假灸组大鼠结肠损伤程度、炎症反应与模型组类似。与正常组比较,模型组p38MAPK mRNA表达增加(P <0.01),p38MAPK、ERK1/2、c-fos蛋白含量均增加(均P <0.05);与模型组比较,隔药灸组p38MAPK mRNA表达降低(P <0.05),p38MAPK、ERK1/2、c-fos蛋白含量均降低(均P <0.05);假灸组均无显著变化(均P> 0.05)。结论隔药灸能下调克罗恩病大鼠结肠p38MAPK、ERK1/2、c-fos的表达,改善炎症、促进肠道组织修复。

隔药饼灸对慢性萎缩性胃炎大鼠PTEN-AKT抑癌途径的作用机制研究

目的探索艾灸通过调控PTEN-AKT抑癌途径相关基因的表达,以达到修复慢性萎缩性胃炎(CAG)受损胃黏膜的可能机制。方法将52只雄性Wistar大鼠随机分为正常组和造模组,造模组采用N-甲基-N-硝基-N-亚硝基胍(MNNG)诱导剂结合夹尾刺激与饥饱失常法制备CAG大鼠模型,34周模型鉴定成功后,将造模大鼠随机分为模型组、隔药饼灸组和西药组,每组10只。隔药饼灸组取中脘和气海穴,每日1次,每周6次;西药组予叶酸悬浊液灌胃,每日1次,每周6次。干预4周后取材,采用HE染色法观察胃窦组织病理学变化,免疫组化法检测各组大鼠胃窦组织PTEN、p-AKT、Caspase-9、PIP2、MDM2的蛋白表达,RT-qPCR法检测各组大鼠胃窦组织PTEN、PI3KCA、Caspase-9、p53 mRNA的表达。结果与正常组比较,模型组可见固有腺体萎缩、肠上皮化生和异型增生;与模型组比较,隔药饼灸组和西药组固有腺体排列较规整,固有腺体萎缩、肠上皮化生、异型增生程度减轻。与正常组比较,模型组大鼠胃窦组织PTEN、Caspase-9蛋白和mRNA表达均显著降低(P<0.05),p-AKT、PIP2、MDM2蛋白表达均显著升高(P<0.05);与模型组比较,西药组和隔药饼灸组胃窦组织PTEN、Caspase-9蛋白表达和Caspase-9、PI3KCA mRNA表达均显著升高(P<0.05),p-AKT、PIP2、MDM2蛋白表达均显著降低(P<0.05),西药组胃窦组织PTEN的mRNA表达升高(P<0.05)。结论隔药饼灸中脘和气海穴可改善CAG大鼠的胃窦组织形态学变化,修复胃黏膜的损伤;隔药饼灸可能通过调控CAG大鼠胃窦组织PTEN-AKT抑癌途径有效控制CAG的胃黏膜损伤,是隔药饼灸治疗CAG和防治癌前病变的效应机制之一。

隔药饼灸对子宫内膜异位症裸鼠异位内膜组织mi-RNAs表达的影响

目的探讨隔药饼灸对子宫内膜异位症(EMs)裸鼠异位内膜组织mi-RNAs表达的影响。方法将16只SPF级BALB/c-nu小鼠14只,先分为两组。即正常组4只,模型组10只,模型组采用自体移植方法进行造模,造模成功后随机分成模型组和隔药饼灸组,每组4只。在治疗14 d后,采用Trizol法提取各组异位内膜组织,质控后采用RNA-seq高通量测序方法分析各组的差异表达mi-RNAs,筛选出差异表达mi-RNAs,结合生物信息学对其靶基因进行分析。结果模型组与正常组比较有差异表达的mi-RNAs共75个,其中上调有37个,下调有38个;隔药饼灸与模型组比较有差异表达的miRNAs共14个,其中上调5个,下调9个;经GO和KEGG分析,隔药饼灸影响mi-RNAs的靶基因主要与蛋白结合及激素代谢有关。结论 mi-RNAs在正常组、EMs模型组及隔药饼灸治疗异位内膜组织后的异常表达,提示mi-RNAs在EMs发生发展及隔药饼灸治疗中起重要调控作用。

隔药灸神阙穴治疗腹泻型肠易激综合征的疗效观察及其对血清5-HT、IL-10的影响

目的:观察隔药灸神阙穴治疗腹泻型肠易激综合征(IBS-D)患者的临床疗效及其对血清5-HT(5-羟色胺)、IL-10(白介素-10)的影响。方法:60例IBS-D患者以随机数字表法分为2组,每组各30例,观察组采取隔药灸神阙穴治疗,对照组给予双歧杆菌乳杆菌三联活菌片治疗,治疗4周后评估临床治疗效果,治疗前后分别采用胃肠症状自评量表(GSRS)、IBS症状严重程度量表(IBS-SSS)、IBS生活质量量表(IBS-QOL)进行评分,ELISA检测血清5-HT、IL-10含量。结果:观察组治疗4周后临床总有效率(93.3%)高于对照组(70.0%)(P<0.05);治疗后,2组患者GSRS评分、IBS-SSS评分较治疗前均降低(均P<0.05),IBS-QOL评分较治疗前均提高(P<0.05),血清5-HT水平明显降低(P<0.05),IL-10含量明显升高(P<0.05);观察组治疗后GSRS评分、IBS-SSS评分低于对照组(P<0.05),IBS-QOL评分高于对照组(P<0.05),血清5-HT水平低于对照组(P<0.05),IL-10高于对照组(P<0.05)。结论:IBS-D患者给予隔药灸神阙穴治疗临床疗效显著,有利于缓解胃肠道症状,减轻病情严重程度,提高患者生活质量;减轻肠道敏感性,抑制炎症反应是其部分作用机制。