The role of miR‐128‐3p through MAPK14 activation in the apoptosis of GC2 spermatocyte cell line following heat stress

Abstract

Background MicroRNAs play a crucial role in the regulation of spermatogenesis. For example, miR‐128‐3p expression is known to decrease significantly after testicular hyperthermia, but the regulatory effect of this change on the spermatogenesis damage caused by heat stress remains unclear. Objectives This study aimed to verify whether the target gene of miR‐128‐3p is MAPK14, which affects spermatogenic cell proliferation and apoptosis under testicular hyperthermia. Materials and methods Mouse testis and GC2 spermatocyte cell line heat stress models were established. miR‐128‐3p expression before and after heat stress was analyzed by reverse transcription polymerase chain reaction. MAPK14 and p‐MAPK14 expression was detected by Western blot, and cell apoptosis was analyzed by Annexin V‐FITC/PI. Subsequently, miR‐128‐3p inhibitors and mimics were used to interfere with spermatocytes before and after heat stress, respectively, for correlation detection. Results Compared with the control group, the heat stress group showed decreased miR‐128‐3p expression, increased p‐MAPK14 expression, and decreased cell proliferation activity. In the GC2‐spd cell line in vitro, miR‐128‐3p inhibitors were found to upregulate p‐MAPK14 expression, reduce cell proliferation activity, and increase apoptosis, consistent with the results obtained in the heat treatment alone. Furthermore, miR‐128‐3p mimics transfected in the GC2 cells after heat stress reduced p‐MAPK14 expression, alleviated the decrease in cell proliferation, and decreased the apoptosis level. Conclusions The downregulation of miR‐128‐3p expression plays an important role in spermatogenesis damages after testicular hyperthermia, which is probably attributable to the activation of the MAPK signaling pathway. Downregulated miR‐128‐3p expression induces the apoptosis and inhibits the proliferation of spermatogenic cells by promoting MAPK14 phosphorylation.

Genome-Wide Association Studies, Runs of Homozygosity Analysis, and Copy Number Variation Detection to Identify Reproduction-Related Genes in Bama Xiang Pigs

“…We identified 18 genes by using GWAS. Of these, MAPK14 was the only gene that was not associated with a specific sex; MAPK14 promotes phosphorylation and is therefore involved in both male and female reproduction ( 31 ). It is critical for the heat-induced proliferation of spermatogenic cells.…”

Role of p38 MAPK Signalling in Testis Development and Male Fertility

“…p38 MAPK participates in germ cell apoptosis via various pathways. Activated intrinsic apoptotic pathways, p38 MAPK/ROS/BAX/caspases, induce apoptosis of germ cells or ejaculated spermatozoan as a result of heat stress, hormonal stimulation, PM2.5, diabetes mellitus, and other stimuli [ 53 , 78 , 102 – 108 ]. Activation of BAX leads to the subsequent initiation of mitochondria-dependent death processes.…”

“…As summarised in Figure 3, compelling evidence suggests that p38 MAPK, as a major regulator under oxidative stress, exacerbates the decline in male fertility. [53,78,[102][103][104][105][106][107][108]. Activation of BAX leads to the subsequent initiation of mitochondria-dependent death processes.…”

“…Orchitis [137][138] Varicocele [135] Aging [93][94][95] Heat stress [105,109,111,117,125] Diabetes mellitus [108,[123][124]132] Testicular torsion [139][140][141][142] Cryptorchid [136] Heavy metal [112,116,122,[129][130] Harmful xenobiotics* [107,[113][114][118][119][120][121][126][127][128]131,133] Figure 3: A summary of the stimuli which induce oxidative stress and activate the p38 MAPK pathway to injure testicular cells. * Harmful xenobiotics include PM2.5, organic compounds, pesticides, and plasticisers.…”

Exposure to elevated temperature affects the expression of PIWI‐interacting RNAs and associated transcripts in mouse testes

“…6 Select studies have also notably reported varying expression levels of specific transcripts exposed to hot temperatures. 7,8 However, the expression status of transcripts coding for proteins involved in noncoding RNA synthesis has not thoroughly been measured in testes when submitted to hot temperatures.…”