Effects Of Yiqi Huoxue Tongluo Recipe Combined With Acupuncture On Nav1.7 And Nav1.8 Of The Dorsal Root Ganglion In Rats With Diabetic Neuropathic Pain

ObjectiveTo study the effect of Yiqi Huoxue Tongluo Recipe combined with Zusanli acupuncture on Nav1.7 and Nav1.8 in dorsal root ganglia of diabetes neuropathic pain rats and its mechanism.MethodsSPF-grade SD rats were fed adaptively for 1 week,fed with high fat and high sugar for 4 weeks,and a single low dose of streptozotocin(35mg/kg)was intraperitoneally injected to prepare the model of type 2 diabetes rats.After 2 weeks,the mechanical pain threshold(MWT)and heat shrinkable foot reaction time(TWL)of the rats were detected,and the DNP model rats were identified as those whose both decreased to below 85% of the basic value.Model DNP rats were randomly divided into a model group,acupuncture group(Acu),Yiqi Huoxue Tongluo Formula group(YHTL),and Yiqi Huoxue Tongluo Formula combined acupuncture group(Acu+YHTL).The model group was given physiological saline by gavage,the Acu group was given acupuncture at the "Zusanli" point of rats(acupuncture for 20minutes),the YHTL group was given Yiqi Huoxue Tongluo Formula by gavage(9g/kg),and the Acu+YHTL group was given acupuncture at the "Zusanli" point of rats combined with Yiqi Huoxue Tongluo Formula by gavage.Rats in each group were treated once a day,5 days a week,for 4 consecutive weeks.MWT and TWL were detected at 0,1,2,3,and 4 weeks,respectively.After the end of the last treatment,PGE2 and TNF in the rat serum were detected by ELISA-α 、 The content of IL-6.Take the L4-L6 dorsal root ganglia of rats,Western Blot was used to detect the protein expression of Nav1.7 and Nav1.8,RT-qPCR was used to detect the mRNA content of Nav1.7 and Nav1.8,and immunofluorescence staining was used to detect the fluorescence expression of Nav1.7 and Nav1.8 in the dorsal root ganglia of rats.Results1 General situation observationAfter 4 weeks of treatment,the rats in the CTL group were generally in good condition without any adverse symptoms.The rats in the Model group were mentally depressed,their eyes were dull,their movements were slow,their hair became hard and yellow,and they suffered from excessive drinking,eating,and urination.The rats in the Acu group and the medication group were slightly depressed,but their eyes were bright,their voluntary activities decreased,and their body weight increased slowly.The rats in the Acu+YHTL group were in good spirits,their eyes were bright,their responses were more flexible,and their state was better than that of the rats in the Acu group and the medication group.2 Effect of Yiqi Huoxue Tongluo Recipe combined with acupuncture on body weight of DNP ratsAfter 4 weeks of treatment,the overall body weight of the rats in the CTL group was the highest,while the rats in the Model group were the lowest.Compared with the Model group,the body weight of the rats in the Acu group,YHTL group,and Acu+YHTL group increased significantly(P<0.05).Compared with the Acu group and the medication group,the Acu+YHTL group had a more obvious increase in body weight(P<0.05).3 Effect of Yiqi Huoxue Tongluo Recipe combined with acupuncture on FBG in DNP ratsAfter 4 weeks of treatment,the FBG of the rats in the CTL group was still in the normal range,while the FBG of the rats in the Model group remained at a high level.Compared with the Model group,the FBG of the rats in the Acu group and Acu+YHTL group decreased again(P<0.05),FBG did not change in the YHTL group;compared with Acu group,FBG decreased more significantly in Acu+YHTL group.4 Effect of Yiqi Huoxue Tongluo Recipe combined with acupuncture on MWT and TWL of DNP ratsBehavioral results showed that before treatment,compared with the CTL group,the MWT and TWL of the Model group and the rest of the groups were all decreased(P<0.05).After 1 week of treatment,compared with the CTL group,the Model group and the Acu group,The MWT and TWL of the medication group and the Acu+YHTL group still decreased,but after 2,3 and 4 weeks of treatment,compared with the Model group,the MWT and TWL of the YHTL group,the Acu group,and the Acu+YHTL group increased,indicating that the three treatments Both methods can improve the DNP symptoms of rats,and compared with the YHTL group and the Acu group,the MWT,and TWL of the Acu+YHTL group increased more significantly(P<0.05).In the Acu+YHTL group,the MWT and TWL of the rats decreased first and then gradually increased within 4 weeks of treatment,and the effect was better than that of the medication group and the Acu group.5 Effect of Yiqi Huoxue Tongluo Recipe combined with acupuncture on the expression of PGE2,TNF-α,IL-6 in DNP ratsELISA results showed that after 8 weeks of administration,compared with the CTL group,the expression levels of PGE2,TNF-α,and IL-6 in the Model group were significantly increased(P<0.05);The expression levels of PGE2,TNF-α,and IL-6 in mice were significantly decreased(P<0.05),indicating that Yiqi Huoxue Tongluo Formula can reduce the inflammatory response of DNP rats.6 Effect of Yiqi Huoxue Tongluo Recipe combined with acupuncture on Nav1.7 and Nav1.8 protein expression in dorsal root ganglia of ratsThe results of Western blot,PCR,and immunofluorescence showed that after 4weeks of treatment,compared with the CTL group,the expression levels of Nav1.7and Nav1.8 proteins in the rat dorsal root ganglion tissues of the Model group and other groups were significantly increased(P<0.05);compared with the Model group,the expression levels of Nav1.7 and Nav1.8 proteins in the dorsal root ganglion tissues of the three treatment groups of the Acu group,the YHTL group and the Acu+YHTL group were significantly decreased(P<0.05),compared with the YHTL group and the Acu group,the expression levels of Nav1.7 and Nav1.8 proteins in the dorsal root ganglion tissue of rats in the Acu+YHTL group were significantly decreased(P<0.05).Conclusion(1)The expressions of Nav1.7 and Nav1.8 are up-regulated in DRGs of DNP model rats,and the up-regulation is related to pain,which proves that Nav1.7 and Nav1.8 are involved in the occurrence of diabetic neuropathic pain.(2)Yiqi Huoxue Tongluo Recipe combined with acupuncture can alleviate DNP by further inhibiting the expression of Nav1.7 and Nav1.8 and reducing the inflammatory response in rats,which may be one of the mechanisms for the better curative effect of the combination of acupuncture and medicine one.