[Electroacupuncture at "Zusanli"(ST36) and "Xuanzhong"(GB39) improves inflammatory reactions and reduces levels of serum MMP-3 and MMP-9 in adjuvant arthritis rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) and "Xuanzhong" (GB39) on joint inflammatory reactions and serum matrix metalloproteinase-3 (MMP-3) and MMP-9 contents in rats with adjuvant arthritis (AA), so as to explore its mechanism underlying improvement of AA. METHODS: Forty male SD rats were randomly divided into control, model, acupoint and non-acupoint groups (n=10 in each group). The arthritis model was established by hypodermic injection of complete Freund's adjuvant (0.1 mL) into the bilateral footpads. EA (2 Hz, 3 V) was applied to bilateral ST36 and GB39 or two non-acupoints (5 mm left to ST36 and GB39) for 15 min, once every other day for a total of 8 times. The arthritis index score was evaluated according to the severity of local erythema and swelling of the ankle joint, plantar joint, toe joint and foot metacarpal joint (0-4 points). The inflammatory conditions of the ankle joint were observed by H.E. staining, and the contents of serum MMP-3 and MMP-9 were assayed by ELISA. RESULTS: The arthritis index score and serum concentrations of MMP-3 and MMP-9 were significantly increased in the model group relevant to the control group (P<0.01), and obviously decreased after EA intervention on the 18th day (P<0.01). The therapeutic effect of acupoint EA was notably superior to non-acupoint EA in down-regulating the arthritis index score and serum MMP-3 and MMP-9 concentrations (P<0.01). Under light microscope, marked proliferation of the synovial cells, inflammatory cell infiltration and increase of newly blood vessels were observed in the ankle joint of the model group, which was relatively milder in the acupoint group. CONCLUSION: acupoint EA intervention can significantly alleviate the inflammatory reaction of AA rats, which may be related to its effects in reducing the levels of serum MMP-3 and MMP-9.

Effect of electroacupuncture at the ST36 and GB39 acupoints on apoptosis by regulating the p53 signaling pathway in adjuvant arthritis rats.

p53 and mouse double minute 2 homolog (MDM2) serve key regulatory roles in the apoptosis of synovial cells. The present study aimed to investigate the effects of electroacupuncture (EA) at the 'Zusanli' (ST36) and 'Xuanzhong' (GB39) acupoints on apoptosis in an adjuvant arthritis (AA) rat model. A total of 40 male Sprague­Dawley rats were randomly divided into Control, AA, AA + EA and AA + sham EA groups (n=10 rats in each group). Rats in all the groups, with the exception of the control group, were injected with Complete™ Freund's adjuvant into the bilateral hindlimb footpad to establish the AA model. Rats in the AA + EA group were treated with EA at the ST36 and GB39 acupoints. Rats in the AA + sham EA group were treated with percutaneous electrical stimulation at a position of 5 mm away from the ST36 and GB39 acupoints. The arthritis index scores and hindlimb paw volumes of the rats in each group were recorded. Subsequently, pathological changes in the synovial tissue were evaluated by hematoxylin and eosin (H&E) staining, and the apoptotic rate of the synovial cells was detected by TUNEL staining. In addition, the expression levels of the apoptosis­associated proteins, Bax, phorbol­12­myristate­13­acetate­induced protein 1 (Noxa) and p53 upregulated modulator of apoptosis (PUMA), were determined by western blot analysis. The expression of both the gene and protein of p53 and MDM2 in synovial tissue was detected by reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and western blot analysis, respectively. The results indicated that the arthritis index scores and hindlimb paw volumes upon EA stimulation were significantly decreased compared with those of the AA group (P<0.05). H&E staining revealed that the synovial inflammation of EA stimulation was significantly decreased compared with the AA group (P<0.05). The TUNEL assay results indicated that the apoptotic rate of synovial cells in the AA + EA group was significantly increased compared with that in the AA group (P<0.05). Furthermore, an increased expression of proapoptotic proteins was confirmed by the increased expression levels of Bax, Noxa and PUMA in the AA + EA group. The results of RT­qPCR and western blot analysis demonstrated that, compared with the AA group, EA stimulation led to a marked increase in p53 (P<0.05) and a significant decrease in MDM2 (P<0.05) gene and protein expression. Taken together, these results demonstrated that EA performed on the ST36 and GB39 acupoints led to a significant amelioration in AA injury of model rats, by regulating the p53 signaling pathway and inducing apoptosis.

Genome-wide trait-trait dynamics correlation study dissects the gene regulation pattern in maize kernels.

BACKGROUND: Dissecting the genetic basis and regulatory mechanisms for the biosynthesis and accumulation of nutrients in maize could lead to the improved nutritional quality of this crop. Gene expression is regulated at the genomic, transcriptional, and post-transcriptional levels, all of which can produce diversity among traits. However, the expression of most genes connected with a particular trait usually does not have a direct association with the variation of that trait. In addition, expression profiles of genes involved in a single pathway may vary as the intrinsic cellular state changes. To work around these issues, we utilized a statistical method, liquid association (LA) to investigate the complex pattern of gene regulation in maize kernels. RESULTS: We applied LA to the expression profiles of 28,769 genes to dissect dynamic trait-trait correlation patterns in maize kernels. Among the 1000 LA pairs (LAPs) with the largest LA scores, 686 LAPs were identified conditional correlation. We also identified 830 and 215 LA-scouting leaders based on the positive and negative LA scores, which were significantly enriched for some biological processes and molecular functions. Our analysis of the dynamic co-expression patterns in the carotene biosynthetic pathway clearly indicated the important role of lcyE, CYP97A, ZEP1, and VDE in this pathway, which may change the direction of carotene biosynthesis by controlling the influx and efflux of the substrate. The dynamic trait-trait correlation patterns between gene expression and oil concentration in the fatty acid metabolic pathway and its complex regulatory network were also assessed. 23 of 26 oil-associated genes were correlated with oil concentration conditioning on 580 LA-scoutinggenes, and 5% of these LA-scouting genes were annotated as enzymes in the oil metabolic pathway. CONCLUSIONS: By focusing on the carotenoid and oil biosynthetic pathways in maize, we showed that a genome-wide LA analysis provides a novel and effective way to detect transcriptional regulatory relationships. This method will help us understand the biological role of maize kernel genes and will benefit maize breeding programs.

[Clinical effect evaluation of acupuncture combined with medication for prevention of endometriosis recurrence after surgery].

OBJECTIVE: To evaluate the effect and safety of acupuncture combined with Jiawei Mojie tablet for the prevention of endometriosis recurrence after surgery. METHODS: One hundred and six patients after the conservative surgery against endometriosis were randomly divided into a western medication group and an acupuncture and medication group(a combination group), 53 cases in each one. In the western medication group, gestrinone was applied orally. In the combination group, acupuncture and Jiawei Mojie tablet with oral administration were adopted. Xuehai (SP 10), Sanyinjiao (SP 6) and Guanyuan (CV 4) were mainly used, and acupoints based on syndrome differentiation were combined. After the surgery and when the menstruation was end, 3 to 5 days later the preventive treatment was used successively for 6 months. Patients were followed up after treatment and in 3 months, 6 months, and 18 months (two years after surgery) after treatment. The recurrence [Abdominal/perineal ultrasonic examination, serum cancer antigen 125 (CA125) test], menstruation score, endometriosis health profile-5 (EHP-5) and the safety index of serum alanine aminotransferase (ALT) were compared between the two groups. RESULTS: Eight patients (five cases in the western medication group and three cases in the combination group) were rejected because of dropping, and six ones (two cases in the western medication group and four cases in the combination group) discontinued. When patients were followed up in 18 months after treatment, the suspected recurrence rate through ultrasonic examination was 2.0% (1/50), and the unusual rate of serum CA125 text was 4.0% (2/50) in the combination group, which were apparently lower than 18.8% (9/48) and 25.0% (12/48) in the western medication group (both P < 0.01). After treatment and 3-month, 6-month, and 18-month following up, the menstruation scores of the combination group were obviously lower than those of the western medication group (P < 0.01, P < 0.05). After 18 months, the positive rate of EHP-5 in the combination group was 0 (0/50), which was markedly lower than 18.8% (9/48) in the western medication group (P < 0.01). After treatment and 3-month, 6-month, and 18-month following up, the unusual rates of serum ALT text in the combination group were lower than those in the western medication group (P < 0.01, P < 0.05). After treatment, the unusual rate of serum ALT text was not significantly different in the combination group compared with that before treatment (P > 0.05), and after treatment and 3-month, 6-month following up, the unusual rates of serum ALT text in the western medication group were remarkablely higher than those before treatment (P < 0.01, P < 0.05). After 18-month following up, the unusual rate of serum ALT text in the western medication group was not statistically significant compared with that before treatment (P > 0.05). CONCLUSION: Acupuncture combined with Jiawei Mojie tablet can effectively prevent endometriosis recurrence after surgery, and improve menstruation condition and life quality, which are less injurious for liver than gestrinone.

[Effects of Electroacupuncture on Joint Function in Rheumatoid Arthritis Patients of Liver- and Kidney-Yin Deficiency Type].

OBJECTIVE: To compare the effects between electroacupuncture (EA) plus western medicine and simple western medicine in improving clinical symptoms and local joint function of rheumatoid arthritis (RA) patients with yin deficiency of Liver and Kidney. METHODS: A total of 68 RA patients of yin deficiency of Liver and Kidney were equally randomized into EA+medication group and medication group (n=34 in each group). Both groups were given once-a-week methotrexate (7.5 mg/time) and once-a-day leflunomide (10 mg/time), while EA+medication group was additionally treated by EA at bilateral Ganshu (BL 18), Shenshu (BL 23), Xuanzhong (GB 39), Zusanli (ST 36), Taichong (LR 3), Hegu (LI 4) 3 times/week. The treatment lasted for 12 weeks. The visual analogue scale (VAS, for assessing rest pain), swollen joint count (SJC), tender joint count (TJC), patient's global assessment (PGA), physician's global assessment (PhGA), traditional Chinese medicine (TCM) symptom scoring, 28 joints activity index (disease activity score, DAS 28), American College of Rheumatology 20 (ACR 20, i.e. 20% of clinical improving rate), and health assessment questionnaire (HAQ) were assessed and erythrocyte sedimentation rate (ESR), serum C-reactive protein (CRP) levels were examined for comparison. RESULTS: Statistical differences were observed in before-after-treatment comparisons in both groups in reducing rest pain, SJC, TJC, serum CRP content, PGA and PhGA, HAQ scoring and DAS 28 (P<0.05, P<0.01). The ESR in the medication group and TCM symptom scoring of the EA+medication group were also significantly decreased after the treatment (P<0.05).The effects of the EA+medication group were superior to those of the medication group in reducing rest pain, SJC, TJC, TCM symptom and HAQ score, and the total effetive rate (P<0.05,90.9% vs 66.67%). Adverse reactions as pharyngeal obstruction sensation, anorexia, abdominal distension, etc. can be reduced by EA therapy coordinated with western medicine. CONCLUSIONS: EA is effective in relieving symptom and joint function in RA patients with yin deficiency of Liver and Kidney.

Electroacupuncture attenuates collagen-induced arthritis in rats through vasoactive intestinal peptide signalling-dependent re-establishment of the regulatory T cell/T-helper 17 cell balance.

OBJECTIVE: Imbalance between T-helper 17 (Th17) cells and regulatory T (Treg) cells is causally linked to the development of rheumatoid arthritis (RA). In this study, we tested the hypothesis that electroacupuncture (EA) confers therapeutic benefits in RA through activation of vasoactive intestinal peptide (VIP)-dependent signalling and restoration of the Th17/Treg cell balance. MATERIALS AND METHODS: A collagen-induced arthritis (CIA) model was induced in Sprague-Dawley rats by injection of bovine type II collagen in incomplete Freund's adjuvant on day 0 and day 7. Three days after the second injection, EA was given at acupuncture points GB39 and ST36 three times per week for 4 weeks. To block VIP signalling, [D-P-Cl-Phe(6)-Leu(17)]-VIP, a VIP receptor antagonist, was administered intraperitoneally 30 min before EA. Inflammatory and pathological responses in the joint were assessed. Synovial VIP receptor mRNA levels and Treg and Th17 cell frequencies in the spleen were determined. RESULTS: EA significantly reduced the severity of CIA, as evidenced by reduced paw volumes, arthritis scores and inflammation scores. EA significantly increased mRNA expression of the VIP receptor VPAC1 and led to an elevation in CD4(+)FOXP3(+) Treg cell frequency and a reduction in CD4(+)IL17(+) Th17 cell frequency. Pre-injection of a VIP receptor antagonist significantly reversed EA-induced expansion of Treg cells, but did not alter the frequencies of Th17 cells. CONCLUSIONS: EA exerts anti-inflammatory effects in a collagen-induced rat model of arthritis. These effects appear to be mediated through activation of VIP signalling and re-establishment of the Th17/Treg cell balance.

Electroacupuncture inhibits inflammation reaction by upregulating vasoactive intestinal Peptide in rats with adjuvant-induced arthritis.

Acupuncture is emerging as an alternative therapy for rheumatoid arthritis (RA). However, the molecular mechanism underlying this beneficial effect of acupuncture has not been fully understood. Here, we demonstrated that electroacupuncture at acupoints Zusanli (ST36), Xuanzhong (GB39); and Shenshu (BL23) markedly decreased the paw swelling and the histologic scores of inflammation in the synovial tissue, and reduced the body weight loss in an adjuvant-induced arthritis rat model. However, the electrical stimulation at nonacupoint did not produce any beneficial effects against the experimental arthritis. Most interestingly, the electroacupuncture treatment resulted in an enhanced immunostaining for vasoactive intestinal peptide (VIP), a potent anti-inflammatory neuropeptide, in the synovial tissue. Moreover, the VIP-immunostaining intensity was significantly negatively correlated with the scores of inflammation in the synovial tissue (r = -0.483, P = .0026). In conclusion, these findings suggest that electroacupuncture may offer therapeutic benefits for the treatment of RA, at least partially through the induction of VIP expression.

[Effects of acupuncture on the number and degranulation ratio of mast cells and expression of tryptase in synovium of rats with adjuvant arthritis].

OBJECTIVE: To observe the effects of acupuncture on synovial pathology, synovial mast cell degranulation and tryptase expression and to investigate the relationship between the functions of mast cells and effects of acupuncture on early adjuvant arthritis in rats. METHODS: Forty-six male Wistar rats were randomly divided into normal control group (n=16), untreated group (n=15) and acupuncture group (n=15). Adjuvant arthritis was induced by injection of 0.1 mL Freund's complete adjuvant in right hind limb footpad. Normal control group and untreated group received no acupuncture treatment, while rats in the acupuncture group were treated with sterilized disposable stainless steel needles inserted perpendicularly as deep as 2 to 3 mm at Xuanzhong (GB39), 6 mm at Shenshu (BL23) and 7 mm at Zusanli (ST36) for eight times (15 min each time) every two days. Setting the modeling day as the 0 day of the experiment, the body weight and paw volume of the rats were measured every three days from the 0 day. In the end, synovial tissues of the right hind ankles were sampled and made into paraffin sections. Then they were firstly stained with hematoxylin-eosin for observing synovial pathology to evaluate the effects of acupuncture on adjuvant arthritis, then stained with toluidine blue for observing the number and degranulation ratio of synovial mast cells and finally detected by immunohistochemical staining method to investigate the expression of tryptase in synovium. RESULTS: Compared with the untreated group, the body weight of rats in the acupuncture group was increased significantly (P<0.05), while the paw volume decreased obviously (P<0.01). Hematoxylin-eosin staining showed that acupuncture significantly inhibited inflammatory cell infiltration, synovial cell hyperplasia, and synovial fibroplasia in synovium of rats with adjuvant arthritis as compared with the untreated group (P<0.05). Toluidine blue staining showed that acupuncture could significantly diminish the numbers of total and degranulated mast cells in rats with adjuvant arthritis (P<0.01), which were significantly higher in the untreated group than in the normal control group (P<0.01). Showing by immunohistochemical staining, the expression of tryptase in synovium in the acupuncture group was decreased as compared with the untreated group (P<0.01). Analyzed by Spearman's bivariate correlation, the number of mast cells and degranulation ratio of mast cells were positively correlated with the pathological scores (r=0.837, P<0.01; r=0.634, P<0.01). CONCLUSION: Acupuncture can improve pathological condition of inflammatory synovium in rats with early adjuvant arthritis by inhibiting the function of synovial mast cells, which may play an important underlying role in the immunoregulation of acupuncture on adjuvant arthritis.