E. adenophorum induces Cell Cycle Arrest and Apoptosis of Splenocytes through the Mitochondrial Pathway and Caspase Activation in Saanen Goats.

Yajun He,Quan Mo,Zhicai Zuo,Lei Wu,Yancheng Zhou,Yan Qiao,Weihong Chen,Yanchun Hu,Junliang Deng,Biao Luo,Ruiguang Xu,Yahui Wei,Wei He

doi:10.1038/srep15967

Yajun He, Quan Mo + Show 11 more

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https://doi.org/10.1038/srep15967

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Abstract

The precise cytotoxicity of E. Adenophorum in relation to the cell cycle and apoptosis of splenocytes in Saanen goats remains unclear. In the present study, 16 Saanen goats were randomly divided into four groups, which were fed on 0%, 40%, 60% and 80% E. adenophorum diets. The results of TUNEL, DAPI and AO/EB staining, flow cytometry analysis and DNA fragmentation assays showed that E. adenophorum induced typical apoptotic features in splenocytes, suppressed splenocyte viability, and caused cell cycle arrest in a dose-dependent manner. However, westernblot, ELISA, qRT-PCR and caspase activity analyses showed that E. adenophoruminhibited Bcl-2 expression, promoted Bax translocation to the mitochondria, triggered the release of Cytc from the mitochondria into the cytosol, and activated caspase-9 and -3 and the subsequent cleavage of PARP. Moreover, in E. adenophorum-induced apoptosis, the protein levels of Fas, Bid, FasL and caspase-8 showed no significant changes. E. adenophorum treatment induced the collapse of ΔΨm. Moreover, these data suggested that E. adenophorum induces splenocyte apoptosis via the activation of the mitochondrial apoptosis pathway in splenocytes. These findings provide new insights into the mechanisms underlying the effects of E. adenophorum cytotoxicity on splenocytes.

Highlights

Eupatorium adenophorum (E. adenophorum) is an invasive weed and an important destructive exotic species, representing a major threat to the economy and ecology in some regions of the world[1]
The ingestion of freeze-dried E. adenophorum leaf powder as a diet supplement induces hepatotoxicity in mice as well as anorexia, rumination suspension and photosensitization in cattle[31] and chronic pulmonary disease in horses[2]. These observations provide a rationale for exploring E. adenophorum as a cause of splenic toxicity and an inducer of apoptosis in the splenocytes of Saanen goats
Flow cytometry is frequently used to monitor early apoptosis[32], and the flow cytometry results of the present study showed that the E. adenophorum increased the numbers of apoptotic splenocytes, indicating that E. adenophorum reduced the survival and inhibited the growth of the splenocytes through the induction of apoptosis and cell cycle arrest, suggesting that the splenocytes of Saanen goats are sensitive to E. adenophorum

Summary

Introduction

Eupatorium adenophorum (E. adenophorum) is an invasive weed and an important destructive exotic species, representing a major threat to the economy and ecology in some regions of the world[1]. This plant is indigenous to Mexico but has been introduced to places such as Hawaii, New Zealand, India, Nepal, and California[2]. Previous studies have demonstratedthat ingestion of the methanolic extract of E. adenophorum induces obvious atrophy of the spleen, indicating that E. adenophorum represents a potential threat to the immune system[15]. We investigated the cytotoxic effects of E. adenophorum on splenocytesin Saanen goats and detected apoptosis-inducing effects at both the cell andtissue levels, in addition to examining effects on cell cycle progression, to illuminate the potential mechanisms involved in E. adenophorum-induced spleen toxicity in goats

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