Effect of Huoxue Huayu Lishui Decoction on P65 and TAK1 of acute soft tissue injury in rats

Objective:To explore the mechanism of the action of Huoxue Huayu Lishui prescription on acute ankle joint injury.Methods:Forty-eight SPF rats were randomly divided into blank group,model group,Shujin Huoluo capsule group,low,medium and high dose groups for promoting blood circulation and removing blood stasis.The blank group was not modeled,and the other groups were modeled.Each group of 8 animals,the Shujin Huoluo capsule group was given 0.2g/ml mixed liquid by gavage,and the low,medium and high blood circulation and blood stasis Lishui prescription group was given 1.03g/ml,2.06g/ml,4.12g/ml decoction For the stomach,the normal group was given 1.5ml/100g,once a day for 1 week.The expression of NF-κB P65 and transforming growth factor kinase 1(TAK1)protein in the soft tissue around the ankle joint of rats was detected by sports protein immunoblotting method,and the mRNA expression of NF-κB P65 and TAK1 was detected by PCR technology.Results:One day after modeling,the expressions of P65 and TAK1 mRNA and protein in the model group,the positive drug group,and low,medium and high doses of Huoxue Huayu Lishui Recipe were significantly different from those of the blank control group(P<0.05).After taking the medicine for 7 days,compared with the model group,the expressions of P65 and TAK1 mRNA and protein in the Shujin Huoluo capsule group and the low,medium and high dose Huoxue Huayu Lishui Recipe group were all decreased(P<0.05).After 7 days of taking the medicine,compared with the Shujin Huoxue Capsule group,there was no difference in P65 protein in the low-dose Huoxue Huayu Lishui Recipe group.The expression of P65 protein in the middle and high groups and the Shujin Huoxue Capsule group were all decreased(P<0.05).Compared with the Shujin Huoxue Capsule group,the expressions of TAK1 protein,P65 and TAK1 mRNA in the low,medium,and high groups and the Shujin Huoxue Capsule group were all decreased(P<0.05).The results suggest that Huoxue Huayu Lishui Recipe can inhibit the expression of NF-κB P65 and TAK1 protein in ankle joint injury.Conclusion:Huoxue Huayu Lishui prescription may regulate the expression of NF-κB P65 and TAK1 in ankle joint injury of NF-κB rats,so as to improve the clinical symptoms of local swelling and pain in the ankle joint.

Effect of Huoxue Huayu Detoxification on GSK-3β and β-catenin levels in OSF patients and its mechanism

Objective:To explore the effects of Huoxue Huayu detoxification method on the levels of GSK-3βandβ-catenin in OSF patients and its mechanism.Methods:A total of 120 patients with oral submucosal fibrosis(OSF)treated in our hospital from May 2018 to December 2019 were selected and divided into routine treatment group and Huoxue Huayu detoxification group of 60 cases according to the lottery method.The routine treatment group was treated with triamcinolone acetonide injection,and the Huoxue Huayu detoxification group was treated with Huoxue Huayu detoxification.The lesion area,mouth opening degree,SFRP1 level in gingival crevicular fluid and saliva,TCM symptom score,TGF-β1,IL-6,GSK-3β,β-catenin expression and clinical effect were observed in the two groups.Results:After treatment,the lesion area of oral mucosa in the Huoxue Huayu detoxification group was smaller than that in the routine treatment group(P<0.05),and the mouth opening degree in the Huoxue Huayu detoxification group was larger than that in the routine treatment group(P<0.05);After treatment,the level of SFRP1 in gingival crevicular fluid and saliva and clinical effect in the Huoxue Huayu detoxification group were higher than those in the routine treatment group(P<0.05);the TCM syndrome score,TGF-β1,IL-6,GSK-3β,β-catenin expression in the Huoxue Huayu detoxification group were lower than those in the routine treatment group(P<0.05).Conclusion:By acting on Wnt/β-catenin signaling pathway,Huoxue Huayu detoxification method can regulate the expression of TGF-β1,IL-6,GSK-3β,β-catenin,improve the clinical symptoms of patients with oral submucous fibrosis,alleviate the body inflammation,alleviate the pain,with remarkable effect.

Shugan Huoxue Huayu Fang(疏肝活血化瘀方)attenuates carbon tetrachloride-induced hepatic fibrosis in rats by inhibiting transforming growth factor-β1/Smad signaling

OBJECTIVE:To investigate the potential mechanism by which Shugan Huoxue Huayu Fang(疏肝活血化瘀方,SGHXHYF)ameliorates liver fibrosis.METHODS:Liver fibrosis was induced in rats by intraperitoneal injection of carbon tetrachloride(CCl_(4))in peanut oil solution(40%,3 m L/kg body weight)twice a week for 8 weeks.A normal control group received the same volume of peanut oil alone.During weeks 5-8,the CCl_(4)-injected rat groups were administered saline(vehicle control),colchicine(0.1 mg/mL,1 mg/kg,positive control),or SGHXHYF(0.1 mg/mL;0.3,0.6 and 1.2 mg/kg)once daily by oral gavage.Rats were sacrificed 24 h after the last treatment.Blood samples were collected for measurement of serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP),albumin(ALB),collagenⅠ,and collagenⅢlevels.Liver samples were analyzed by histopathological staining,Masson's staining of extracellular matrix proteins,and immune-ohistochemical staining ofα-smooth muscle actin(α-SMA).TGF-β1/Smad protein and mRNA levels were analyzed by Western blot and quantitative reverse transcription-polymerase chain reaction analysis,respectively.In vitro experiments were also performed using rat hepatic stellate cells(HSCs).RESULTS:Compared with the control animals,CCl_(4)-exposed rats exhibited elevated serum levels of ALT,AST,ALP,collagenⅠ,and collagenⅢ;reduced serum levels of ALB;and increased collagen deposition andα-SMA expression in liver sections,reflecting liver fibrosis.CCl_(4) also increased expression of TGF-β1 and the activated(phosphorylated)forms of Smad2 and Smad3 but reduced expression of the negative regulator Smad7 in the liver.Notably,concomitant administration of SGHXHYF to CCl_(4)-exposed rats was found to significantly reverse or abolish the pro-fibrotic effects of CCl_(4) in the liver and reduced serum transferase levels.Analysis of HSCs in vitro confirmed that,mechanistically,SGHXHYF inhibited activation of the TGF-β1/Smad signaling pathway by downregulating phosphorylated Smad2 and Smad3 and upregulating Smad7 levels.CONCLUSION:SGHXHYF ameliorated CCl_(4)-induced liver fibrosis by inhibiting the TGF-β1/Smad signaling pathway.These findings suggest that SGHXHYF may have clinical utility for the treatment or prevention of liver fibrosis.

Huoxue Jiedu Huayu recipe(活血解毒化瘀方)alleviates contralateral renal fibrosis in unilateral ureteral obstruction rats by inhibiting the transformation of macrophages to myofibroblast

OBJECTIVE:To investigate the action and underlying mechanisms of Huoxue Jiedu Huayu recipe(活血解毒化瘀方,HJHR)against unilateral ureteral obstruction(UUO)-induced injury in the contralateral kidney.METHODS:Forty-eight male Sprague-Dawley rats weighing(200±10)g were used in this study and randomly assigned to 4 groups:a sham group,a UUO group,a UUO+eplerenone(EPL)group,and a UUO+HJHR group.The contralateral kidneys were harvested for further study 180 d after surgery.Histological analysis,immunohistochemistry and immunofluorescence were used to study the fibrosis of the contralateral kidneys obtained from UUO rats.Contralateral kidney damagerelated pathway proteins were detected by real-time polymerase chain reaction and Western blot analysis.RESULTS:HJHR significantly inhibited fibrosis of the contralateral kidney in UUO rats by attenuating the UUOinduced macrophage-to-myofibroblast transition(MMT)in the contralateral kidney.Moreover,HJHR attenuated fibrosis in the contralateral kidney of UUO rats by preventing MMT through the aldosterone/mineralocorticoid receptor/serum/glucocorticoid regulated kinase 1 pathway.CONCLUSIONS:Our findings suggest that HJHR may be a potential treatment for renal interstitial fibrosis of obstructive nephropathy.