Nicotine Induces Podocyte Apoptosis through Increasing Oxidative Stress.

Xiqian Lan,Moin A Saleem,Seyedeh Shadafarin Marashi Shoshtari,Pravin C Singhal,Ashwani Malhotra,Rivka Lederman,Judith M Eng,Stuart E Dryer

doi:10.1371/journal.pone.0167071

Xiqian Lan, Moin A Saleem + Show 6 more

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https://doi.org/10.1371/journal.pone.0167071

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Abstract

BackgroundCigarette smoking plays an important role in the progression of chronic kidney disease (CKD). Nicotine, one of the major components of cigarette smoking, has been demonstrated to increase proliferation of renal mesangial cells. In this study, we examined the effect of nicotine on podocyte injury.MethodsTo determine the expression of nicotinic acetylcholine receptors (nAChR subunits) in podocytes, cDNAs and cell lysate of cultured human podocytes were used for the expression of nAChR mRNAs and proteins, respectively; and mouse renal cortical sections were subjected to immunofluorescant staining. We also studied the effect of nicotine on podocyte nephrin expression, reactive oxygen species (ROS) generation (via DCFDA loading followed by fluorometric analysis), proliferation, and apoptosis (morphologic assays). We evaluated the effect of nicotine on podocyte downstream signaling including phosphorylation of ERK1/2, JNK, and p38 and established causal relationships by using respective inhibitors. We used nAChR antagonists to confirm the role of nicotine on podocyte injury.ResultsHuman podocytes displayed robust mRNA and protein expression of nAChR in vitro studies. In vivo studies, mice renal cortical sections revealed co-localization of nAChRs along with synaptopodin. In vitro studies, nephrin expression in podocyte was decreased by nicotine. Nicotine stimulated podocyte ROS generation; nonetheless, antioxidants such as N-acetyl cysteine (NAC) and TEMPOL (superoxide dismutase mimetic agent) inhibited this effect of nicotine. Nicotine did not modulate proliferation but promoted apoptosis in podocytes. Nicotine enhanced podocyte phosphorylation of ERK1/2, JNK, and p38, and their specific inhibitors attenuated nicotine-induced apoptosis. nAChR antagonists significantly suppressed the effects of nicotine on podocyte.ConclusionsNicotine induces podocyte apoptosis through ROS generation and associated downstream MAPKs signaling. The present study provides insight into molecular mechanisms involved in smoking associated progression of chronic kidney disease.

Highlights

It is estimated that there are more than a billion cigarette smokers all over the world, and over one third of them above 15 years of age [1, 2]
We studied the effect of nicotine on podocyte nephrin expression, reactive oxygen species (ROS) generation, proliferation, and apoptosis
Nicotine enhanced podocyte phosphorylation of ERK1/2, JNK, and p38, and their specific inhibitors attenuated nicotine-induced apoptosis. nicotinic acetylcholine receptors (nAChRs) antagonists significantly suppressed the effects of nicotine on podocyte

Summary

Introduction

It is estimated that there are more than a billion cigarette smokers all over the world, and over one third of them above 15 years of age [1, 2]. Clinical reports have demonstrated that cigarette smoking plays important role in the progression of chronic kidney disease (CKD), and it worsens CKD in patients with diabetes, hypertension, polycystic kidney disease, and post kidney transplant [2,3,4]. Nicotine plays its effects via the activation of the nicotinic acetylcholine receptors (nAChRs) [2, 11]. Both in vitro and in vivo studies demonstrated that nAChRs expressed by mesangial cells contribute to the proliferation of mesangial cells in response to stimulation by nicotine [14, 15]. Cigarette smoking plays an important role in the progression of chronic kidney disease (CKD).

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