Association study of ACE polymorphisms and systemic lupus erythematosus in Northern Chinese Han population.

Systemic lupus erythematosus (SLE) is an autoimmune disease, with multiple genetic and environmental factors involving in its etiology. Angiotensin converting enzyme (ACE) gene was reported to have important roles in the development and progression of SLE. In this study, a case-control study was carried out to investigate the effects of seven SNPs and I/D in ACE gene in the development of SLE in Northern China. Seven SNPs including A5466C, T3892C, A240T, C1237T, G2215A, A2350G and C3409T were genotyped by PCR-RFLP method, and I/D was examined by agarose gel electrophoresis followed PCR directly. 314 SLE patients were compared to 320 normal controls in the study. Data were analyzed by SPSS 13.0 and HaploView software. The frequency distribution of SNP A2350G and Alu I/D and five haplotypes (AAAACCCI, AGAACCTD, AAAATCTI, TAAATTTI and TAAATCTI) were demonstrated to be different between case and control groups significantly. Whereas other SNPs and haplotypes had no differences in two cohorts. The results revealed that variations of ACE gene had association with SLE, which indicated ACE gene may play an important role in pathogenesis of SLE in Northern Chinese Han population.

Network pharmacology-based approach uncovers the JAK/STAT signaling mechanism underlying paederia scandens extract treatment of rheumatoid arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is a common autoimmune disease. Paederia scandens (Lour.) Merr is a common folk remedy used in Hainan, China, to dispel the wind and dampness associated with RA. METHODS: The active components of Paederia scandens were extracted using network pharmacology. The potential targets of active components were used to determine activated pathways, and the in vitro effects of Paederia scandens extracts were verified in RA fibroblast-like synoviocytes (HFLS-RA). RESULTS: We identified 27 active components using ultra-high-performance liquid chromatography (UHPLC)-quadrupole time-of-flight (QTOF)-mass spectrometry (MS). Among the major target genes with high connectivity, IL-1ß, PI3K, TNF, and JAK2 are known to play key roles in RA development. High-affinity interactions were identified between active compounds in Paederia scandens extract and Janus kinase JAK 2, which are key components of the JAK-signal transducer and activator of transcription (STAT) signaling pathway. In HFLS-RA cells, Paederia scandens extract treatment reduced the mRNA levels of IL-6, IL-1ß, and IL-17. Paederia scandens extract treatment also significantly inhibited the phosphorylation of JAK 2 and STAT3, regulating cell proliferation. CONCLUSIONS: Based on these results, we confirmed that Paederia scandens has potential for application as a therapeutic and preventive food and acts through the modulation and suppression of JAK-STAT pathway activation to control the inflammatory response in RA.

[Effect of electroacupuncture on expression of CREB and its ability to bind to synaptic proteins in amygdala and hippocampus of rats with post-traumatic stress disorder].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the expression of cAMP-response element binding protein (CREB, a key protein for BDNF-TrkB signaling) and it's blinding ability to synaptic key protein in the amygdala and hippocampus of rats with post-traumatic stress disorder (PTSD), so as to lay a foundation for further study of the interaction mechanism between BDNF-TrkB signaling and synaptic plasticity. METHODS: Twenty-four male SD rats were randomly divided into blank, model and electroacupuncture (EA) groups, with 8 rats in each group. The PTSD model was established by psychological stress (bondage) and physiological stress (forced swimming and anesthesia). After modeling, EA (2 Hz/100 Hz, 1 mA) was applied to "Baihui"(GV20) "Shenting"(GB24) and bilateral "Shenshu"(BL23) for 20 min, once daily for 21 days. The behavioral changes (spontaneous locomotor within 30 min and contextual fear conditioning tests in 7 days) were detected by using a spontaneous locomotor detection box, and a conditioned fear response test chamber, respectively. The expression of CREB was detected by immunohistochemistry and Western blot, separately. The binding abilities of CREB to synaptic proteins (post synaptic density 95 [PSD95], synaptophysin [SYN] and growth-associated protein 43 [GAP43]) were verified by chromatin-immunoprecipitation (CHIP) technique. RESULTS: After modeling, the spontaneous locomotor distance, the expression levels of CREB and the binding ability of CREB to PSD95 protein in the amygdala and hippocampus were significantly decreased (P<0.01), and the percentage of freezing time significantly increased in the model group relevant to the blank group (P<0.01)． Following the intervention, the spontaneous locomotor distance, and the expression levels of CREB and the binding ability of CREB to PSD95 protein were considerably increased in the EA group relevant to the model group (P<0.05，P<0.01). No significant changes were found in the binding abilities of CREB to SYN and GAP43 after modeling and after EA intervention (P>0.05). CONCLUSION: EA can improve the motor activity in PTSD rats, which may be associated with its effect in increasing the binding ability of CREB to the synaptic key protein PSD95 to regulate the interaction between the synaptic plasticity and BDNF-TrkB signaling pathway of the amygdala and hippocampus.

[Mechanism of Shenqi compound recipe anti-earlier diabetic artherosclerosis in GK rats].

OBJECTIVE: To explore the mechanism of Shenqi compound recipe (SQCR) anti-earlier diabetic artherosclerosis in GK rats. METHOD: Four-month specefic pathogen free (SPF) GK rats were divided randomly according to blood glucose level into four groups: model group (5 mL x kg(-1) x d(-1) sterile water), ramipril group (positive control, 1 mg x kg(-1) x d(-1)), SQCR low dosage (0.72 g x kg(-1) x d(-1)) and SQCR high dosage group (2.88 g x kg(-1) x d(-1)) and Wistar rats as normal control group(5 mL x kg(-1) x d(-1) sterile water). GK rats took high-fat diet freely and meanwile were injected N-omega-nitro-L-arginine methyl ester (L-N-AME) intra-peritoneally with the dose of 10 mg x kg(-1) x d(-1) in order to induce earlier diabetic artherosclerosis, while normal control group took regular diet and were injected normal saline intra-peritoneally. In the experiment periods, each group was administrated correspondent substance respectively for 32 d. At the end, sampling blood by abdominal aorta and picking aorta on ice. Determined monocyte chemoattractant protein-1 (MCP-1) concentration by ELISA, messenger ribonucleic acid (mRNA) expression of MCP-1 and peroxisome proliferator-activated receptor gamma (PPARgamma) in aorta by reverse transcriptase PCR (RT-PCR). RESULT: Concentrations of MCP-1 in serum in SQCR low and high dosage groups and the mRNA expression of MCP-1 in SQCR high dosage group were all decreased significantly compared with model group (P < 0.05). The mRNA expression of PPARgamma in SQCR low and high dosage groups all increased compared with model group (P < 0.05 or P < 0.01). CONCLUSION: Inhibiting the mRNA and protein expression of MCP-1 and upregulating the mRNA expression of PPARgamma in aorta might be contribute to SQCR anti-earlier diabetic artherosclerosis in GK rats partly.

[Effects and mechanism of ShenQi compound recipe on inflammation maker in GK rats].

OBJECTIVE: To explore the effects and mechanism of ShenQi Compound Recipe on inflammation maker of type 2 diabetes mellitus in GK rats. METHODS: Rats were ranodmly divided into Model group, Ramipril group (positive control, 1 mg/kg x d), SQCR low dosage (0.72 g/kg x d), SQCR high dosage group (2.88 g/kg x d) and Wistar control group. Each group was administrated correspondent substance respectively for 32 days. Determined C-reactive protein (CRP) by ELISA and tumour necrosis factor (TNF)-alpha by radioimmunassay. The mRNA expression of nuclear factor (NF)-kappaB p65 in aorta was determined by real time RT-PCR, and activation of it using immunohistochemistry staining. RESULTS: Concentrations of CRP and TNF-alpha in serum and the expression of mRNA and activation of NF-kappaB were all decreased in SQCR low and high dosage groups compared with model group (P < 0.05 or P < 0.01). CONCLUSION: These results suggest that SQCR can decrease the level of inflammation maker in serum, which may be resulted from reducing the mRNA expression and activation of NF-kappaB.

Albiflorin Granule significantly decreased the cholesterol gallstone formation by the regulation of insulin transduction signal.

OBJECTIVE: To study the mechanism of insulin resistance in the cholesterol gallstone formation from insulin signal transduction pathway so as to reveal the possible mechanism and the effective role of Albiflorin Granule on preventing the cholesterol gallstones. METHODS: Serum triglycerides (TG), free fatty acid (FFA), and total cholesterol (TC) from different groups were measured and liver cells InsR, PKB, IKK-ß protein expression levels were detected by western blotting. RESULTS: Albiflorin significantly decreased the cholesterol gallstone formation rate, increased glucose infusion rate in gallstone guinea pigs and improved insulin resistance. Compared with the normal group, insulin receptor and PKB protein expression in GS group were significantly reduced. IKK-ß protein in the GS group increased significantly and Albiflorin could reduce IKK-ß protein expression in guinea pig liver cells. CONCLUSIONS: The model of insulin resistance in cholesterol gallstone guinea pig was successfully established, which plays an important role in the cholesterol gallstone formation. All aspects of insulin signaling pathway are involved in gallstone formation. Albiflorin can regulate various aspects of insulin signal transduction pathway to prevent the formation of gallbladder.

Effect of emodin on mobility signal transduction system of gallbladder smooth muscle in Guinea pig with cholelithiasis.

OBJECTIVE: To study the effect of emodin on protein and gene expressions of the massagers in mobility signal transduction system of cholecyst smooth muscle cells in guinea pig with cholesterol calculus. METHODS: The guinea pigs were randomly divided into 4 groups, such as control group, gall-stone (GS) group, emodin group and ursodeoxycholic acid (UA) group. Cholesterol calculus models were induced in guinea pigs of GS, emodin and UA groups by lithogenic diet, while emodin or UA were given to the corresponding group for 7 weeks. The histomorphological and ultrastructure change of gallbladder were detected by microscope and electron microscope, the content of plasma cholecystokinin (CCK) and [Ca2+]i were analyzed successively by radioimmunoassay and flow cytometry. The protein and mRNA of Gsα, Giα and Cap in cholecyst cells were determined by western blotting and real time polymerase chain reaction (RT-PCR). RESULTS: Emodin or UA can relieve pathogenic changes in epithelial cells and muscle cells in gallbladder of guinea pig with cholesterol calculus by microscope and transmission electron microscope. In the cholecyst cells of GS group, CCK levels in plasma and [Ca2+]i decreased, the protein and mRNA of GS were down-regulated, the protein and mRNA of Gi and Cap were up-regulated. Emodin significantly decreased the formative rate of gallstone, improved the pathogenic change in epithelial cells and muscle cells, increased CCK levels in plasma and [Ca2+]i in cholecyst cells, enhanced the protein and mRNA of Gs in cholecyst cells, reduced the protein and mRNA of Gi and Cap in cholecyst cells in guinea pig with cholesterol calculus. CONCLUSION: The dysfunction of gallbladder contraction gives rise to the disorders of mobility signal transduction system in cholecyst smooth muscle cells, including low content of plasma CCK and [Ca2+]i in cholecyst cells, abnormal protein and mRNA of Gs, Gi and Cap. Emodin can enhance the contractibility of gallbladder and alleviate cholestasis by regulating plasma CCK levels, [Ca2+]i in cholecyst cells and the protein and mRNA of Gs, Gi and Cap.

Association between CD14 SNP -159 C/T and gastric cancer: an independent case-control study and an updated meta-analysis.

PURPOSE: The association between CD14 -159C/T polymorphism and the susceptibility to gastric cancer (GC) has been reported. However, the results were inconclusive. In the present study, a case-control study and a meta-analysis were performed to assess the possible association between -159C/T in the CD14 gene and GC risk. PATIENTS AND METHODS: Relevant studies were searched in several databases including PubMed, Web of Science, EMBASE, Chinese National Knowledge Infrastructure database, and Wanfang database (last search was performed on December 30, 2015). In addition, a case-control study involving 164 GC cases and 169 controls was also performed in the analysis. Statistical analysis was performed by the software Revman5.3. RESULTS: A total of ten published studies and the present case-control study involving 2,844 GC and 3,983 controls were included for the meta-analysis. The analysis result indicated that the T allele of CD14 -159C/T polymorphism did not confer risk for GC (in our study: [P=0.93]; in the meta-analysis: T vs 2N odds ratio =1.28 and 95% confidence interval (CI) =0.95-1.24, [P=0.24]). However, we found a significant association in the recessive model (in our study: TT vs TC+CC [P=0.04]; in the meta-analysis: TT vs TC+CC odds ratio =1.12 and 95% CI =1.01-1.26, [P=0.04]). Furthermore, a subgroup analysis by ethnicity showed that TT genotype was significantly associated with GC in Asian (odds ratio =1.17 and 95% CI =1.02-1.34, [P=0.02]) but not in Caucasian. CONCLUSION: Our results highlight the TT genotype of CD14 -159C/T as a genetic susceptibility factor for gastric cancer, particularly, in Asians and population-based controls.

Pregnane receptor gene polymorphisms, pathogenic bacteria distribution and drug sensitivity, and TCM syndrome differentiation in patients with cholelithiasis.

OBJECTIVE: To investigate the distribution of pathogens and drug resistance in bile and the association between the pregnane X receptor (PXR) gene polymorphisms, traditional Chinese medicine (TCM) syndromes and the risk of cholesterol gallstone disease (CGD). METHODS: A total of 392 samples were enrolled in this study from January 2014 to February 2015, among which 192 patients were with CGD, and 200 samples were healthy. Strains were isolated and susceptibility testing was the disk diffusion method susceptibility testing. The patients were divided into hepatochlic hygropyrexia, stagnation of liver-qi, and the accumulation of damp. The PXR gene polymorphisms were analyzed by polymerase chain reaction-restriction fragment length polymorphism. The association between the PXR gene polymorphisms and the risk of CGD was examined by logistic regression analysis. RESULTS: A total of 192 cases were detected in 230 of bile culture pathogens, including Gram-negative bacteria 133 (57.83%), Gram-positive bacteria 76 (33.04%), and fungi 21 (9.13%). The top five pathogens were Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis, Candida albicans, and Enterococcus feces, of which 110 cases was of single infection, 48 cases of mixed infection of two strains, eight cases of mixed infection of three bacteria. Among 59 Escherichia coli, the yield extended-spectrum beta-lactamases had 40 (67.80%). The hepatochlic hygropyrexia was the most TCM syndrome, followed by stagnation of liver-qi, and the accumulation of damp was least. Different pathogens and the rs6785049 genotypes distributed differently in cholelithiasis patients with different TCM syndromes (P < 0.05). In hepatochlic hygropyrexia patients the Gram-negative bacteria was most. There was significant differences between CGD group and control group in rs6785049 (P < 0.001). Comparison with wild-type portable GG, GA genotype increased the risk of the occurrence of gallstones (OR = 0.40, 95%CI: 0.16-0.79); likewise, carrying the GA+AA genotype also increased the risk (OR = 0.38, 95%CI: 0.19-0.81). There was no significant differences in rs2276707, rs3814055 site polymorphic loci alleles in CGD group and control group. CONCLUSIONS: In the treatment of cholelithiasis, bile samples should be collected for bacterial culture and sensitivity test, and drugs should be strictly chosen based on the results. The rs6785049 polymorphisms in PXR gene may increase the risk of gallstones ontogeny, and gallstones can be early detected and prevented by detecting genotypes. rs6785049 polymorphisms in PXR gene may has relationship with TCM syndromes.

miR-143 regulates proliferation and apoptosis of colorectal cancer cells and exhibits altered expression in colorectal cancer tissue.

Colorectal cancer is one of the most commonly diagnosed cancers and a leading cause of death. Studies have shown that abnormal expression of microRNAs, small non-coding RNA molecules that regulate gene expression, is linked to the occurrence of cancer. This study sought to determine the role of microRNA-143 (miR-143) in colorectal cancer. Reduced levels of miR-143 expression were detected in colorectal cancer tissues compared to normal adjacent tissue. Transfection of artificially synthesized miR-143 mimics into SW-480 cells, a colorectal cancer cell line, resulted in increased levels of cell proliferation and apoptosis. Further, cells transfected with miR-143 mimics showed a reduction in the proportion of cells in S phase and an increase in the proportion of cells in G1 phase. The altered expression levels of miR-143 in colorectal cancer and its ability to affect the behavior of colorectal cancer cells suggest miR-143 could be used as a new target for the diagnosis and treatment of colorectal cancer.

Pharmacokinetic and pharmacodynamic analysis of ferulic acid-puerarin-astragaloside in combination with neuroprotective in cerebral ischemia/reperfusion injury in rats.

OBJECTIVE: To investigate the effects of the active ingredients combined therapy on inflammatory factors interleukin 1 beta (IL-1ß) and neuropeptide Y (NPY) based on pharmacodynamics in rats. METHODS: The animal model was built by transient middle cerebral artery occlusion (MCAO). The method for evaluating the concentrations of the FA-Pr-Al components in rat plasma was established by using HPLC and the expression levels of IL-1ß and NPY were determined by ELISA. A new mathematics method of the trend of percentage rate of change (PRC) was used to assess the correlation between pharmacokinetics (PK) and pharmacodynamics (PD). RESULTS: FA-Pr-Al in combination reduced neurological deficits, decreased infarct volume and inhibited the expression levels of IL-1ß and NPY (all P<0.05) compared with the model group. FA, Pr and Al all displayed two compartment open models in rats. Clockwise hysteresis loops were obtained by time-concentration-effect curves. IL-1ß and NPY level changes in the plasma followed an opposite trend to the plasma concentration tendency after Cmax was reached. Astragaloside's PRC value was significantly higher than those of FA and puerarin between 120 to 180 min. CONCLUSIONS: The pharmacokinetics of FA-Pr-Al in combination were closely related its pharmacodynamics in treating ischemia/reperfusion injury, and the components of FA-Pr-Al may have a synergistic pharmacological effect. Astragaloside may play a more pronounced role in regulating IL-1ßand NPY levels compared with puerarin or FA.

Effects of Porphyromonas gingivalis extracellular vesicles on human periodontal ligament fibroblasts.

The purpose of this study was to investigate the influencing mechanism of Porphyromonas gingivalis extracellular vesicles on human periodontal ligament fibroblasts to better understand the pathogenesis of periodontitis, the major cause of adult tooth loss. Human periodontal ligament fibroblasts were cultured and randomly assigned to a control group and an extracellular vesicles (ECV) group. The ECV group was exposed to isolated Porphyromonas gingivalis extracellular vesicles; the control group was not exposed. Western blotting was used to detect expression of matrix metalloproteinase 1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1), and RT-PCR was used to detect mRNA expression of alkaline phosphatase (ALP). When human periodontal ligament fibroblasts were processed by Porphyromonas gingivalis extracellular vesicles (ECV), protein expression levels of both MMP-1 and TIMP-1 were significantly higher than that of the control group (P<0.05). In contrast, ALP mRNA expression in human periodontal ligament fibroblasts processed by ECV was significantly lower than that of the control group (P<0.05). Porphyromonas gingivalis extracellular vesicles can up-regulate expression of MMP-1 and TIMP-1 protein and ALP mRNA of human periodontal fibroblasts.

Curcumin treatment alters ERK-1/2 signaling in vitro and inhibits nasopharyngeal carcinoma proliferation in mouse xenografts.

Curcumin, a plant phenol, has been used for centuries in traditional medicines for its anti-inflammatory and anti-neoplastic properties. The compound is believed to act on a range of proteins involved in cell cycle regulation. In this study, the effect of curcumin on ERK-1/2 pathway protein expression and on proliferation of nasopharyngeal carcinoma cells was investigated. CNE-2Z nasopharyngeal carcinoma cells were cultured with 10, 20, 40, or 80 µM curcumin for 24 h before proliferation was assessed by MTT colorimetry. Cell proliferation was increasingly inhibited as the concentration of curcumin increased (P<0.005). Additionally, Western blotting revealed that expression of p-ERK-1/2, MMP-9, and TIMP-1 was altered following curcumin treatment, also in a dose-dependent manner. Expression of p-ERK-1/2 and MMP-9 decreased, while expression of TIMP-1 increased (P<0.05). Finally, CNE-2Z cells were xenografted under the skin of 18 nude mice. Mice were treated with vehicle only (control), 24 mg/kg curcumin (low-dose group), or 50 mg/kg curcumin (high-dose group) every other day for 40 days beginning 24 h after xenografting. Compared to tumors from the control group, the volume and weight of xenograft tumors was significantly lower in both curcumin groups, with a higher magnitude of difference in the high-dose curcumin group (P<0.05). These results indicate that curcumin treatment can inhibit proliferation of nasopharyngeal carcinoma cells and alter expression of proteins in the ERK-1/2 signaling pathway. Therefore, curcumin warrants further investigation as a potential treatment for nasopharyngeal cancer.

Overexpression of microRNA-21 and microRNA-126 in the patients of bronchial asthma.

MicroRNAs are implicated in an increasing number of diseases and health complications, including asthma. Previous studies have suggested roles for microRNA-21 (miR-21) and microRNA-126 (miR-126) in asthma, although these relationships are incompletely understood. The aim of this study was to further assess the relationship between miR-21, miR-126 and the occurrence and treatment of allergic asthma. Quantitative real-time PCR analyzed expression levels of miR-21 and miR-126 in bronchial epidermal cells from asthma patients treated with (treatment group, n=19) or without (non-treatment group, n=16) inhaled corticosteroids or from non-asthmatic healthy individuals (normal group, n=12). Bronchial epidermal cells were cultured in vitro to determine if there was a relationship between IL-13 and miR-21 and miR-126 expression. Compared to the normal group, miR-21 and miR-126 expression was significantly upregulated in asthma patients regardless of treatment. miR-21 and miR-126 expression was significantly higher in the non-treatment group than the treatment group (P<0.05). In vitro, miR-21 and miR-126 expression increased in bronchial epidermal cells with increasing IL-13 concentration. Because upregulation of miR-126 and miR-21 were associated with occurrence and therapy of allergic asthma, they may be indices of therapeutic effect that are valuable for the evaluation for asthma treatment.

Association of APOE polymorphisms and insulin resistance with TCM syndromes in type 2 diabetes patients with macroangiopathy.

To investigate the association between apolipo-protein E (APOE) polymorphisms and insulin resistance and Traditional Chinese Medicine (TCM) syndromes in type 2 diabetes mellitus (T2DM) with macroangiopathy, 60 patients with T2DM macroangiopathy were enrolled and divided into three groups: dryness-heat due to deficiency of yin, Qi-Yin deficiency, and Yin-Yang deficiency, according to the TCM syndromes, with a control group of 20 healthy individuals. APOE genotype analysis was performed with polymerase chain reaction amplification and restriction fragment length polymorphism, and the results showed that the proportion of the ε4/4 and ε3/4 genotypes and frequencies of the ε4 and ε3 alleles were higher in the Qi-Yin deficiency group (P<0.05). Among the T2DM macroangiopathy patients, the E4 group had the largest number of cases, as well as a significantly longer disease course compared to the E2 group (P<0.05). The insulin resistance index (IRI), insulin action index and body mass index (BMI) of patients in the Yin-Yang deficiency group were significantly different from those of patients with dryness-heat due to deficiency of yin and Qi-Yin deficiency. Furthermore, correlation analysis of the BMI and IRI of patients in the Yin-Yang deficiency group revealed a correlation coefficient r=0.696 (P<0.01) and a typical correlation between them. In conclusion, the Qi-Yin deficiency in T2DM patients with macroangiopathy is associated with the APOE E4 and E3 genotypes. Thus, the APOE gene polymorphism can, to some degree, reflect the TCM syndrome types of T2DM patients with macroangiopathy. Insulin resistance plays an important role in the occurrence of T2DM macroangiopathy and is closely associated with the Yin-Yang deficiency according to the TCM differentiating types.

[Effect of pricking blood therapy at Ashi points on peripheral pain mediums in the acute gouty arthritis rat].

OBJECTIVE: To study the analgesic mechanism of pricking blood therapy at Ashi points in the acute gouty arthritis rat. METHODS: Forty rats were randomly divided into a blank group, a model group, an indomethacin group and a pricking blood group. Except the blank group, other groups were injected with sodium urate liquor into the ankle cavity to develop the acute gouty arthritis rat model, and the indomethacin group received gastric perfusion of indomethacin, and the pricking blood group were treated with pricking blood therapy at Ashi points. The peripheral pain mediums K+, NE, DA, (5-HT contents were determined. RESULTS: The K+, DA, 5-HT contents in the pricking blood group decreased significantly as compared with the model group (P < 0.01, P < 0.05); there was no significant difference in the content of NE between the pricking blood group and the model group. CONCLUSION: Pricking blood at Ashi points can effectively inhibit release of the peripheral pain mediums K+, DA and 5-HT.