| 1. Objectives and significanceGout (gout) is a result of purine metabolic disorder, produce too much uric acid and (or) the uric acid excretion decreased, serum uric acid concentration increase led to the deposition of urate crystals in soft tissue caused by a group of metabolic diseases. The clinical characteristics for hyperuricemia, urate deposition caused by repeated attacks of acute, chronic arthritis and soft tissue injury, uric acid nephrolithiasis caused by gouty nephropathy. In recent years, due to changes in life style, the incidence rate was increased. China’s general population with hyperuricemia prevalence rate is about10%, gout prevalence of approximately0.58%. The acute gouty arthritis in adult male is the most common form of inflammatory arthritis, and show the young tendency.Gout is divided into primary and secondary two classes. Primary gout its etiology and pathogenesis is still not fully known, but it must have the following two basic conditions:(1) the formation of uric acid excessive purine metabolism affected by some enzymes in the regulation, research shows that primary gout and genetic defects leading to a lack of these enzymes or activity related to changes in.(2) the uric acid excretion decreased glomerular filtration reduction cause hyperuricemia is mainly seen in chronic kidney disease caused by renal failure, renal excretion of uric acid and threshold increase. Reason unknown. Renal tubular excretion of uric acid reduction and some urate transporter related, their involvement in the proximal convoluted tubules of uric acid salt active production and absorption, the abnormalities and gene variation. Secondary gout in chronic nephropathy induced by renal dysfunction, reduced excretion of uric acid (lactic acidosis in diabetic ketoacidosis, etc.), blood disease, tumor chemotherapy drugs causing nucleic acid conversion increases, the formation of uric acid increase or certain drugs such as thiazide diuretics, furosemide, low-dose acetylsalicylic acid; cyclosporin A, ethambutol, ethanol, lactic acid and other metabolites can inhibit renal tubular excretion of uric acid resulted in elevated uric acid.Acute gouty arthritis due to uric acid sodium (MSU) micro crystals in joints surrounding tissue deposition caused by acute inflammatory response, its pathogenesis and the relief mechanism is not very clear. Hyperuricemia in MSU crystal formation and deposition in cartilage, synovial membrane and joint peripheral, usually without inflammatory manifestation. In some factors (such as cold, trauma and alcohol), crystallization from falling into the joint cavity, causing inflammatory cascade reaction.Traditional Chinese medicine is also" gout " in the name of the Yuan Dynasty, Zhu Danxi put forward clearly" gout" disease", gout,100go limbs pain, books that the white tiger calendar day wind syndrome is also. Rate of sputum, fever, rheumatism, blood deficiency". Doctors will bi syndrome in offspring of Tongbi, or arthritis and Bi are called gout, arthritis or white tiger. In medicine, gout etiology and pathogenesis of congenital deficiency of spleen and kidney overstrain, diet, injury, loss of cold-dampness Yang, filled, and voiced block, and the cold evil, from the outside, the moisture of Sheng, because inside, belongs to this standard is false, the actual situation mixture. Congenital diet on spleen and kidney, with close relationship. Spleen governing ascending lucidity descending turbidity, kidney main transpiration gasification, rich food delicious, damage the spleen and stomach, spleen fails, ascending the clear and descending turbid lost division; renal failure caused by lack of warmth, gasification, are filled with blood run cold, poor flow, main and collateral channels of joints, arrested, more and more, main and collateral channels closed resistance, resulting in limb joint pain, blood stasis and damp turbidity long stalemate, due to cold and coalescence, attached to the joints, metaplasia gout nodules, then stiffness into stone, induced joint deformity, long patient kidney, kidney-qi deficiency, Yang micro due to renal failure.Heat shock protein (HSP), also known as stress proteins, biological cell stressor such as heat, ischemia, hypoxia, trauma, hunger stimulation occurred when the heat shock responsehighly conserved protein. HSP can be rapid, short-term adjustment of the cell survival function of the stress process, protect the cells to resist damage, and helps the cells to restore normal structure and function. HSP70is one of the most conservative and most important family, most abundant in most organisms, the most sensitive in the stress response as well as in clinical practice. HSP70mainly has the following major biological functions: molecular chaperone role to prevent apoptosis and promote cell proliferation, immune roleto and make biological get heat resistance.Recent studies have shown that HSP70may participate in the immune response and inhibit the inflammatory response in the damaged cell surface expression can help the immune system to recognize clear cytotoxic cells. And suppression protect cells and tissues from reactive oxygen species and cytokines from inflammatory injury in rats. HSP70can help to antigen-presenting cells and antigen presenting to T cells involved in cellular immunity, inhibit the expression of IL-1, TNF-alpha and other inflammatory mediators; HSP70through impede the P38mitogen-activated protein kinase mediated signaling pathway, inhibition of IKB phosphorylation and degradation in the cytoplasm, thereby inhibiting NF-κB from the release of the transfer of cytoplasm to the nucleus, inhibition of NF-κB activation, inhibition of the inflammatory mediators TNF-a, IL-6, etc., reduce the inflammatory response; the expression of HSP70has some influence on the development of gouty arthritis inflammationIn the treatment of GA, there are obvious advantages of Chinese medicine, With characteristic of overall treatment, less side effects, clinical efficacy, and can be long-term use. Gout-Qing Granules is a preparation from a Chinese herbal formula which has been used in clinical treatment for many years,functioning as clearing sputum, promoting blood circulation to remove meridian obstruction, regulating qi to alleviate pain. It has good clinical efficacy for Gouty arthritis.The formula constitutes of Pseudobulbus Cremastrae seu Pleiones, Rhizoma Polygoni Cuspidati, Rhizoma Smilacis Glabrae, Radix et Rhizoma Rhei, Radix Salviae Miltiorrhiae, Radix Acanthopanacis Bidentatae, Glycyrrhiza,etc. Previous pharmacological studies have shown that TongFengNing granules have significant pain, inhibit inflammation, but their specific anti-inflammatory mechanism is not clear.This topic by using TongFengNing to cure acute gouty arthritis in rats to detect the expression of heat shock protein gene and protein synthesis in acute gouty arthritis and explore the role of heat shock protein in inflammatory process, clear whether TongFengNing by the effects of heat shock protein expression and anti-inflammatory effects of acute gouty arthritis to provide the experimental basis for the further development TongFengNing, but also for the heat shock protein induction agent against acute gouty arthritis.2Methods and projects2.1. Composition of Traditional Chinese medicine compound Tongfengning: Atractylodes, Cortex Phellodendri, hyssop, coix seed, tuckahoe, dioscorea, hairy holly clematis, giant knotweed.2.2Acute gouty arthritis rat model:Inject sodium urate crystal solution in intra-articular to induct the acute arthritis model.2.3Experimental methods:50male Wistar rats were randomly divided into the blank group, model group, western medicine group, high dose of Chinese medicine group, low dose of Chinese medicine group.10rats in each group. Traditional Chinese medicine in high dose group30g/kg/day and in low dose15g/kg/day, western medicine compared with0.09mg/kg/day of colchicine, the blank group, model group gavage ns,2times a day, once every5days, and medication third days make model, model oflH measuring rat right ankle articular circumference, molding after24hours of observation of rat gait, molding after48when measured right ankle circumference, kill and sample blood to detect the amount of IL-1, IL-10; take synovial, extraction of RNA, HE staining and immunohistochemical staining.2.4. The detection index:2.4.1General situation:rat manner, gait, active degree.2.4.2Pathology:pathology of synovial2.4.3HSP70synthesis detection:Using LSAB immunohistochemical labeling method.2.4.4HSP70mRNA expression detection:Using real-time quantitative polymerase chain reaction (RT-PCR)2.4.5Serum IL-1, IL-10:ELISA assay detection.2.5Statistics:UsingSPSS13.0statistical analysis software, measurement data to show, the SPSS13.0package processing data among multiple groups, were compared using univariate analysis of variance, within groups before and after compared using a paired t test. P<0.05for the presence of significant difference, P<0.01as having significant.3. Results3.1The general conditions:Throughout the experimental period, the normal rats shiny hair, eyes, and quick response, eating normal; colchicine group, Chinese medicine gout drink group, the rats in model group were given in the inflammatory after the onset of inflammatory swelling of the ankles, with not enough land, three foot gait, activity severely limited. After the treatment of colchicine group, Chinese medicine benzbromarone groups of above symptoms improved markedly, the rats in model group were no significant improvement in symptoms.3.2Tongfengning’s effects on rat pathological morphology in rats:normal rat synovium normal, model rats of synovial lining cells was markedly thickened, site of injury and surrounded by inflammatory cell stacks, polymorphonuclear leukocytes, mononuclear cells, foreign body giant cell and inflammatory cell infiltration; neovascularization is visible in many locations, vascular cavity markedly dilated congestive; gout Decoction group and colchicine group, its inflammatory leukocyte infiltration, vascular dilatation were lower than the model group.3.3HSP70expression:Levene test for homogeneity of variance (F=0.078, P=0.989), using the method of LSD test. The group with significant difference (F=74.525, P=0.000). The expression of HSP70minimum blank group, model group was slightly higher than that of the blank group but there was no statistical significance (P=0.195); traditional Chinese medicine Western medicine group, low dose, high dose group were significantly higher than that of traditional Chinese medicine in model group (P=0.000, P=0.000, P=0.000), with statistical significance; traditional Chinese medicine Western medicine group, high dose group expression higher than that of traditional Chinese medicine in low dose group (P=0.000, P=0.000), with statistical significance; traditional Chinese medicine Western medicine group, high dose group expression difference does not have statistical significance (P=0.418).3.4The expression of HSP70mRNA:Levene test for homogeneity of variance (F=1.040, P=0.395), using the method of LSD test. The group with significant difference (F=35.778, P=0.000). The expression of HSP70mRNA minimum blank group, model group was slightly higher than that of the blank group (P=0.123), but no statistical significance; western medicine traditional Chinese medicine, Chinese medicine group, low dose and high dose group were higher than those in the model group (P=0.000, P=0.000, P=0.000), with statistical significance; traditional Chinese medicine Western medicine group, high dose group expression higher than that of traditional Chinese medicine in low dose group (P=0.001, P=0.003), with statistical significance; traditional Chinese medicine Western medicine group, high dose group expression difference does not have statistical significance (P=0.604).3.5Serum IL-1, IL-10beta:serum IL-lbeta, IL-10index among every group:1. IL-lbeta content:Levene test variances (F=8.109, P=0.000), using Welch method to approximate the variance analysis. The group IL-1beta content is compared, the difference was statistically significant (F=341.297, P=0.000). Comparison between two two group, model group was significantly higher than that in normal group and Western medicine group, low dose group, high, traditional Chinese medicine (P=0.000, P=0.000, P=0.000, P=0.000). Western medicine and Chinese medicine group was significantly lower than that of traditional Chinese medicine high low dose group (P=0.000, P=0.000), the difference was statistically significant. Traditional Chinese medicine Western medicine group and high dose group was no significant difference (P=1.000). The content of IL-10:Levene test variances (F=4.938, P=0.002), using Welch method to approximate the variance analysis. The group IL-10content, the difference was statistically significant (F=47.969, P=0.000). Comparison between two two group, model group was significantly lower than the blank group, western medicine group, low dose group, Chinese medicine high (P=0.000, P=0.000, P=0.000, P=0.000); traditional Chinese medicine Western medicine group and high dose group were higher than that of traditional Chinese medicine in low dose group (P=0.017, P=0.011), differences statistical significance. Traditional Chinese medicine Western medicine group and high dose group was no significant difference (P=1.000).4.Conclusion1. HSP70as a protective factor can reduce the acute gouty arthritis inflammatory injury;2. The treatment of Tongfengning on acute gouty arthritis is clear and effective, and inducing the high expression of HSP70is one of the important roles in the treatment of acute gouty arthritis... |
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