Effect of “Spleen-Stomach Harmonizing” Needling on Insulin Resistance and Expression of Insulin Receptor Substrate-1, -2, and Glucose Transporter-4 in Insulin Resistance Type 2 Diabetes Rats
【Abstract】Objective To observe the effect of manual acupuncture stimulation of “Quchi” (LI 11), “Hegu” (LI 4), “Zusanli” (ST 36), etc. with “spleen-stomach harmonizing” technique on insulin resistance index (ISI) and the expression of insulin receptor substrate-1 (IRS-1), IRS-2 (two upstream key proteins) and glucose transporter-4 (GluT-4, downstream key protein) of the phosphatidylinositol 3-kinase (PI 3K)/Akt signaling pathway in insulin resistance type 2 diabetes (IRT 2D) rats, so as to explore its mechanism underling improvement of type 2 diabetes. Methods A total of 35 male Wistar rats were used in the present study. The IRT 2Dmodel was established by feeding the animal with high-fat and high-sugar diet for 8 weeks, and subsequent intraperitoneal injection (i.p.i.) of a low dose of streptozotocin (25 mg/kg). Using the stratified random sampling method, 20 successful IRT 2Drats were equally divided into model and acupuncture groups. Ten normal rats were used as the control group. The bilateral acupoints of LI 11, LI 4, ST 36, “Xuehai” (SP 10), “Fenglong” (ST 40), “Yinlingquan” (SP 9), “Diji” (SP 8), “Sanyinjiao” (SP 6) and “Taichong” (LR 3) were punctured with filiform needles, followed by manipulating the needles with “spleen-stomach harmonizing” technique. The treatment was given once daily for 4 weeks except the weekends. Fasting blood glucose and serum insulin levels were measured using ELISA. The quadriceps muscle of thigh of each rat was sampled to test the expression levels of IRS-1, IRS-2, and GluT-4 proteins and genes with Western blot and quantitative Real-time PCR, respectively. Results Following four-week acupuncture intervention, the modeling-induced remarkable increase of ISI, and considerable down-regulation of expression levels of IRS-1, IRS-2 and GluT-4 proteins and genes in the quadriceps femoris were significantly reversed relevant to the model group (P < 0.01), suggesting a possible activation of PI 3 K/Akt signaling after acupuncture administration. Conclusion Spleen-stomach regulation needling can improve insulin resistance and up-regulate the expression of IRS-1, IRS-2 and GluT-4 in the quadriceps femoris in type 2 diabetes rats.
【Keywords】 Type 2 diabetes; Hand acupuncture; Insulin resistance; Insulin receptor substrate; Glucose transporter type 4; Quadriceps femoris;
(Translated by WU XX)
 CHEN HZ, ZHONG NS, LU ZY. Internal Medicine [M]. Beijing: People’s Medical Publishing House, 2013: 736 (in Chinese).
 POELKENS F, LAMMERS G, PARDOEL E M, et al. Upregulation of skeletal muscle inflammatory genes links inflammation with insulin resistance in women with the metabolic syndrome [J]. Exp Physiol, 2013, 98 (10): 1485–1494.
 CHI M, YE Y, ZHANG X D, et al. Insulin induces drug resistance in melanoma through activation of the PI 3 K/Akt pathway [J]. Drug Des Devel Ther, 2014, 8: 255–262.
 ZHANG Y, HAI J, CAO M, et al. Silibinin ameliorates steatosis and insulin resistance during non-alcoholic fatty liver disease development partly through targeting IRS-1/PI 3K/Akt pathway [J]. Int Immunopharmacol, 2013, 17 (3): 714–720.
 GALAGOVSKY D, KATZ M J, ACEVEDO J M, et al. The Drosophila insulin-degrading enzyme restricts growth by modulating the PI 3Kpathway in a cell-autonomous manner [J]. Mol Biol Cell, 2014, 25 (6): 916–924.
 ZHANG ZL, CHEN H, JI XQ, et al. Characteristics of TCM syndromes of chronic complications of typeⅡ diabetes [J]. Chinese Acupuncture and Moxibustion, 2011, 52 (16): 1379–1383 (in Chinese).
 LI ZX, LAN DC, ZHANG HH, et al. Acupoints selection exploring for acupuncture treatment of insulin resistance based on data mining [J]. Acupuncture Research, 2016, 41 (6): 545–549 (in Chinese).
 ZHANG ZL, XUE L, JI XQ, et al. Clinical study on effects of acupuncture on insulin resistance in the patient of type Ⅱ diabetes [J]. Chinese Acupuncture and Moxibustion, 2002, 22 (11): 723–725 (in Chinese).
 ZHANG ZL, JI XQ, ZHANG YH, et al. Controlled study on the needling method for regulating the spleen and stomach for treatment of diabetic retinopathy [J]. Chinese Acupuncture and Moxibustion, 2006, 26 (12): 839–842 (in Chinese).
 ZHANG P, LIU ZF, WANG CM, et al. Observation on the therapeutic effect of needling method for harmonizing spleen-stomach on diabetic gastroparesis [J]. Chinese Acupuncture and Moxibustion, 2007, 27 (4): 258–260 (in Chinese).
 ZHANG ZL, JI XQ, ZHAO SH, et al. Randomized controlled study on effects of the needling method for regulating spleen-stomach on coronary heart disease complicated by type 2 diabetes mellitns complicated [J]. Chinese Acupuncture and Moxibustion, 2008, 28 (9): 629–633 (in Chinese).
 JI XQ, WANG CM, ZHANG P, et al. Effect of spleen-stomach regulation-needling on nerve conduction activity in patients with diabetic peripheral neuropathy [J]. Acupuncture Research, 2010, 35(6): 443–447 (in Chinese).
 WANG M, ZHANG ZL, JI XQ, et al. Randomized controlled clinical trial for analyzing effect of “spleen-stomach regulation needling” on xidative stress level in patients with diabetic nephropathy [J]. Acupuncture Research, 2015, 40 (5): 409–414 (in Chinese).
 WANG X, ZHANG ZL, YANG YQ, et al. Effects of acupuncture on insulin resistance and morphologic changes of islet β-cell in rat with type 2 diabetes mellitus [J]. Journal of Nanjing University of Traditional Chinese Medicine, 2016, 32 (1): 45–49 (in Chinese).
 Ministry of Science and Technology of the People's Republic of China. 关于善待实验动物的指导性意见 [Z]. Ministry of Science and Technology of the People’s Republic of China, 2006 (in Chinese).
 XIANG XS. 2型糖尿病大鼠模型的建立及其在辅助降血糖功能评价中的作用 [D]. Beijing: Chinese Center For Disease Control And Prevention. 2010 (in Chinese).
 WANG Z, YANG YX, XIANG XS, et al. Estimation of the normal range of blood glucose in rats [J]. Journal of Hygiene Research, 2010, 39 (2): 133–142 (in Chinese).
 LIN WZ, WANG P. Experimental Acupuncture Science (实验针灸学) [M]. Shanghai: Shanghai Scientific and technical publishers. 1999: 288 (in Chinese).
 LI GW. Evaluation of islet cell function [J]. Foreign Medical Sciences (Section of Endocrinology), 2003, 23 (3): 159–163 (in Chinese).
 ZHANG ZL. 五味斋医话医案 [M]. Tianjin: Tianjin Science and Technology Press. 2009: 243 (in Chinese).
 HOXHAI G, DISSANAYAKE K, MACKINTOSH C. Effect of IRS 4 levels on PI 3-kinase signaling [J]. PLoS One, 2013, 8 (9): 1–9.
 ZHANG J, NAKATSU Y, SHINJO T. Par 14 protein associates with insulin receptor substrate 1 (IRS-1) , thereby enhancing insulin-induce [J]. J Biol Chem, 2013, 288 (28): 20692–20701.
 BOURA-HALFON S, ZICK Y. Serine kinases of insulin receptor substrate proteins [J]. Vitam Horm, 2009, 80: 313–349.
 ZHANG J, SUN T Q. IRS-2 and its involvement in diabetes and aging [J]. Vitam Horm, 2009, 80: 389–407.
 BRYANT N J, GOVERS R, JAMES D E. Regulated transport of the glucose transporter GLUT 4 [J]. Nat Rev Mol Cell Biol, 2002, 3 (4): 267–277.