Effect of icariside II and metformin on penile erectile function, glucose metabolism, reaction oxygen species, superoxide dismutase, and mitochondrial autophagy in type 2 diabetic rats with erectile dysfunction

Abstract

Background: Icariside II (ICAII) is a flavonoid isolated from herb Epimedium that has been shown to improve erectile function in rats. However, ICAII's underlying mechanism remains unclear.

Methods: Type 2 diabetes mellitus erectile dysfunction (T2DMED) rats were induced by single intraperitoneal injection of 25 mg/kg streptozotocin (STZ) and fed a high-fat, high-sugar, and high-calorie diet for 8 weeks. In the control and T2DMED groups, rats were administered with normal saline; in the metformin (MET) group, rats were administered with MET at 0.2 g/kg/day; and in the ICAII + MET group, rats were administered with ICA II at 10 mg/kg/day and MET for 0.2 g/kg/day. The deposition of advanced glycation end products (AGEs), expression of receptor for AGEs (RAGEs), reactive oxygen species (ROS), and superoxide dismutase (SOD) activity, and corpus cavernosum smooth muscle cells (CCSMCs) mitochondrial autophagy were measured. We also evaluated the expression of LC3-II/I, Beclin-1, P70S6K, PI3K, AKT, mTOR, phospho-AKT, phospho-mTOR, and phospho-P70S6K.

Results: ICAII and MET can improve erectile function, and decrease the levels of fasting plasma glucose (FPG), hemoglobin A1c (HbA1c), and AGEs in rats with T2DMED. Furthermore, ICAII and MET can decrease excessive CCSMC mitochondrial autophagy and the level of RAGE and oxidant stress in rats with T2DMED. ICAII and MET may enhance signaling via the PI3K-AKT-mTOR pathway in order to reduce the excessive mitochondrial autophagy of CCSMCs.

Conclusions: ICAII may effectively improve penile erectile function via decreasing excessive CCSMCs mitochondrial autophagy, RAGE, and oxidant stress. Furthermore, ICAII may enhance signaling via the PI3K-AKT-mTOR pathway in order to reduce excessive CCSMC mitochondrial autophagy.

Keywords: Advanced glycation end products (AGEs); icariside II (ICAII); mitochondrial autophagy; oxidative stress; type 2 diabetes mellitus erectile dysfunction (T2DMED).

Figure 1

ICAII could improve penile erectile function in T2DMED rats, with no obvious effect on MAP. Comparison of the number of mating rats, number of erectile rats, erection frequency, and erection rate between the control group, ICAII + MET group, MET group, and the T2DMED group ($\overline{x}\pm s$). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. MAP, arterial blood pressure; ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction.

Figure 2

Comparison of the ICP/MAP experimental of control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. AUC, area under the curve; ICP, intracorneal pressure; MAP, mean arterial pressure; ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction.

Figure 3

Comparison of the TAP experiment between the control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). TAP, tail artery pressure; ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction.

Figure 4

ICAII + MET could improve glucose metabolism by decreasing the absolute values of FPG, HbA1c, and AGEs in rats with T2DMED. Comparison of the FPG, HbA1c, and AGEs between the control group, ICAII + MET group, MET group, and the T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲, P<0.05; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group; ^※, P<0.05; ^※※, P<0.01, MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; FPG, fasting plasma glucose; HbA1c, hemoglobin A1c; AGEs, advanced glycation end products.

Figure 5

ICAII + MET could improve oxidative stress by decreasing the RAGE and ROS values, and increasing the SOD values, in rats with T2DMED. Comparison of the RAGE, ROS, and SOD values between the control group, MET group, T2DMED group, and ICAII + MET group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲, P<0.05; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group; ^※, P<0.05, MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; ROS, reactive oxygen species; SOD, superoxide dismutase; RAGE, receptor for advanced glycation end products.

Figure 6

Comparison of the MTT growth curves between the CCSMCs in the control group, MET group, T2DMED group, and ICAII + MET group ($\overline{x}\pm s$, n=16). ^★, P<0.05, control group vs. T2DMED group; ^▲, P<0.05, ICAII + MET group vs. T2DMED group. OD, optical density; ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; CCSMCs, corpus cavernosum smooth muscle cells.

Figure 7

ICAII could improve mitochondrial autophagy in rats with T2DMED. Comparison of the mitochondrial autophagosomes of CCSMCs and the autophagosome/CCSMC area ratio between the control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. MDC, monodansylcadaverine; ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; TEM, transmission electron microscopy.

Figure 8

ICAII could reduce the CCSMCs’ excessive autophagy. Comparison of autophagy related protein expression between the control group, ICAII + MET group, MET group, and T2DMED group. ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; CCSMCs, corpus cavernosum smooth muscle cells.

Figure 9

ICAII could enhance the PI3K-AKT-mTOR signaling pathway. Comparison of PI3K-AKT-mTOR signaling pathway related protein expression between the control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. the T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; mTOR, mammalian target of rapamycin.

Figure 10

Comparison of autophagy related mRNA expression between the control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction.

Figure 11

Comparison of the PI3K-AKT-mTOR signaling pathway related mRNA expression between the control group, ICAII + MET group, MET group, and T2DMED group ($\overline{x}\pm s$, n=16). ^★★, P<0.01, control group vs. T2DMED group; ^▲▲, P<0.01, ICAII + MET group vs. T2DMED group. ICAII, icariside II; MET, metformin; T2DMED, type 2 diabetes mellitus erectile dysfunction; mTOR, mammalian target of rapamycin.