Elsevier

Biomedicine & Pharmacotherapy

Volume 102, June 2018, Pages 344-353
Biomedicine & Pharmacotherapy

Involvement of estrogen receptors in silibinin protection of pancreatic β-cells from TNFα- or IL-1β-induced cytotoxicity

https://doi.org/10.1016/j.biopha.2018.01.128Get rights and content

Abstract

Silibinin is a polyphenolic flavonoid that exhibits anticarcinogenic, anti-inflammatory and cytoprotective effects. The effect of silibinin on pancreatic islet β-cell is yet largely unknown in spite of well documented-hepatoprotective effects. Protecting the functional insulin-producing β-cells in the pancreas is a major therapeutic challenge in the patients with type 1 (T1DM) or type 2 diabetes mellitus (T2DM). This study reports the effect of silibinin on the rat pancreatic β-cell line, INS-1, damaged with pro-inflammatory cytokine, TNFα or IL-1β. Exposure to TNFα or IL-1β for 48 h caused INS-1 cells to reduce the production of insulin as well as cell viability. These actions of TNFα or IL-1β are associated with suppression of the expression of estrogen receptors (ERs). Further study revealed that silibinin protected the suppression in the expression of both ERα and ERβ that were involved in insulin synthesis and release, respectively. Furthermore, evidence is obtained that silibinin may impede the loss of critical INS-1 cells by promoting autophagy and preventing apoptosis. Direct cytoprotective effects of silibinin on INS-1 cells suggest that silibinin may be therapeutically beneficial for diabetes.

Introduction

Diabetes is a chronic disease characterized by development of insulin resistance, pancreatic islet β-cell dysfunction, and ultimately to hyperglycemia [1]. Diabetes has been recognized as an immune-mediated disease in which pro-inflammatory cytokines play important roles, accompanying impairment of insulin signaling and selective destruction of insulin producing β-cells [2]. The inflammation-induced metabolic dysfunction in pancreatic islet is one of the key factors in the progression of diabetes [3]. Inflammation affects the pancreatic islet β-cells during the progression of diabetes. Insulitis in type 1 diabetes is caused by autoimmunity and in type 2 diabetes by metabolic stressors such as hyperglycaemia and elevated levels of free fatty acids [4]. Pro-inflammatory cytokines such as tumor necrosis factor-α (TNFα) and interleukin-1β (IL-1β) are crucial as mediators leading to β-cell destruction [5].

The prevalence of diabetes is lower in females [6], and consistently, the recent data have suggested that 17β-estradiol (E2) contributes to insulin production in diabetic patients and animals [7]. Moreover, females are protected from β-cell death and hyperglycemia in the induction of most rodent models of diabetes, whereas males develop overt insulin deficient diabetes [8]. Taken together, the information indicated an important role of estrogen receptors (ERs) for protection of β-cells [9,10]. Estrogen, acts on both estrogen receptors alpha (ERα) and beta (ERβ), important regulators of lipid metabolism and glucose homeostasis. It is well known that ligand-activated ERα promotes β-cell survival and enhances glucose-stimulated insulin biosynthesis [7,10,11], while activation ERβ seems to predominantly enhance glucose-stimulated insulin secretion [[11], [12], [13]]. ERα mediates the function mainly in nucleus, while ERβ is predominantly localized in cytoplasm, mitochondria in particular, regulating mitochondrial gene expressions and maintaining membrane potential as well as ATP production in a variety of cell types, such as endothelial cells and rat primary neurons [14,15].

Silibinin, a natural polyphenolic flavonoid, is a major bioactive component of silymarin isolated from the plant milk thistle Silybum marianum (L.) Geartn. Silibinin is known to have hepatoprotective and anti-carcinogenic effects on human hepatocellular carcinoma and human breast cancer MCF-7 cells [[16], [17], [18]]. Silibinin has a steroidal structure, and the structural similarity with phytoestrogens with a polyphenolic flavonoid skeleton appears to be a cause of silibinin in binding and activating estrogen receptors (ERs) on mammary cells [18,19]. Up-regulation of ERα and ERβ mRNA expression with silibinin as well as partial binding of silibinin to ERs is also reported [20]. Moreover, silibinin is shown to protect streptozotocin-induced diabetes [[21], [22], [23]] with a further possibility that silibinin may reverse hyperglycemia and repair damaged pancreatic β-cells [21]. It is considered that silibinin and 17β-estradiol (E2) may have the similar protective effect on cytokine-induced toxicity in pancreatic β-cells. Hence, we have used the rat pancreatic β-cell line, INS-1, widely used as a model system for β-cells, to examine the effect of silibinin on protection of TNFα- or IL-1β-induced cell damage. The roles of ERα and ERβ involved are discussed.

Section snippets

Reagents

Silibinin with a purity of 99% was purchased from Jurong Best Medicine Material (Zhenjiang, Jiangsu, China). The reagent was dissolved in dimethylsulfoxide (DMSO) as a stock solution. The concentration of DMSO was kept below 0.1% in all of the cell cultures so as not to induce any detectable effects on cell growth. E2,3-(4,5-Dimethyl-2-thiazolyl)-2, 5-diphenyltetrazolium bromide (MTT), propidium iodide(PI), monodansylcadaverine (MDC), 4, 6-diamino-2-phenyl indole (DAPI), RNase A, estrogen

Results

Effects of silibinin on viability and physiological function of the INS-1 cells treated with TNFα or IL-1β.

Given that inflammation-mediated pancreatic β-cell damage was a key inducer in the development of diabetes and that pro-inflammatory cytokines caused both β-cell dysfunction and death [[25], [26], [27]], we investigated the potential effect of silibinin on survival of INS-1 cells treated with TNFα or IL-1β.

To examine the cytotoxicity of silibinin on INS-1 cell, cells were prepared and

Discussion

Destruction of pancreatic islet β-cells is an important etiological factor in development and progression of T2DM [33]. This study demonstrates that silibinin protects INS-1 cells from apoptosis induced with pro-inflammatory cytokine, either TNFα or IL-1β, thereby helping INS-1 cells maintain viability and function.

Silibinin has been shown to have anti-diabetic activity in streptozotocin-induced diabetic mice [21,23]. Several mechanisms have been described for the effects, including modulations

Acknowledgments

This study was supported by Foundation of Liaoning Education Committee (51120427) and Doctoral Starting up Foundation of Liaoning Science and Technology Department (51120390).

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