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The effect of heat stress on gene expression, synthesis of steroids, and apoptosis in bovine granulosa cells

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Cell Stress and Chaperones Aims and scope

Abstract

Summer heat stress (HS) is a major contributing factor in low fertility in lactating dairy cows in hot environments. Heat stress inhibits ovarian follicular development leading to diminished reproductive efficiency of dairy cows during summer. Ovarian follicle development is a complex process. During follicle development, granulosa cells (GCs) replicate, secrete hormones, and support the growth of the oocyte. To obtain an overview of the effects of heat stress on GCs, digital gene expression profiling was employed to screen and identify differentially expressed genes (DEGs; false discovery rate (FDR) ≤ 0.001, fold change ≥2) of cultured GCs during heat stress. A total of 1211 DEGs including 175 upregulated and 1036 downregulated ones were identified, of which DEGs can be classified into Gene Ontology (GO) categories and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The results suggested that heat stress triggers a dramatic and complex program of altered gene expression in GCs. We hypothesized that heat stress could induce the apoptosis and dysfunction of GCs. Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to evaluate the expression of steroidogenic genes (steroidogenic acute regulatory protein (Star), cytochrome P-450 (CYP11A1), CYP19A1, and steroidogenic factor 1 (SF-1)) and apoptosis-related genes (caspase-3, BCL-2, and BAX). Radio immunoassay (RIA) was used to analyze the level of 17β-estradiol (E2) and progesterone (P4). We also assessed the apoptosis of GCs by flow cytometry. Our data suggested that heat stress induced GC apoptosis through the BAX/BCL-2 pathway and reduced the steroidogenic gene messenger RNA (mRNA) expression and E2 synthesis. These results suggest that the decreased function of GCs may cause ovarian dysfunction and offer an improved understanding of the molecular mechanism responsible for the low fertility in cattle in summer.

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Acknowledgments

This research was supported by the National Natural Science Foundation of China (Grant No. 31501955) and the Fundamental Research Funds for the Central Universities (Grant No. KJQN201607).

Author contributions

Lian Li conceived and designed the experiments. Lian Li and Yu Sun performed the experiments. Lian Li analyzed the data. Jie Wu, Yu Sun, and Man Luo contributed reagents/materials/analysis tools. Lian Li and Genlin Wang wrote the paper. Yu Sun and Man Luo prepared the materials. Jie Wu helped analyze the data and in RT-PCR.

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Authors and Affiliations

  1. College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, People’s Republic of China

    Lian Li, Jie Wu, Man Luo, Yu Sun & Genlin Wang

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  1. Lian Li
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  2. Jie Wu
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  3. Man Luo
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  4. Yu Sun
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  5. Genlin Wang
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Correspondence to Lian Li or Genlin Wang.

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Ethics statement

In the present experiment, animal care and procedures were approved and conducted under the established standards of Nanjing Agricultural University, Nanjing, China.

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The authors declare that they have no competing interests.

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Table S1

Analytical flowchart to study bovine granulosa cells transcriptome by DGEs (XLS 175 kb)

Table S2

Primer information. (XLS 140 kb)

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Li, L., Wu, J., Luo, M. et al. The effect of heat stress on gene expression, synthesis of steroids, and apoptosis in bovine granulosa cells. Cell Stress and Chaperones 21, 467–475 (2016). https://doi.org/10.1007/s12192-016-0673-9

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  • DOI: https://doi.org/10.1007/s12192-016-0673-9

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