Ameliorating Effect of β-Carotene on Gene Expression Alteration in Spermatozoon Cultured With Titanium Oxide

Abstract

Introduction: In the present study, we assessed the effects of β-carotene on Titanium oxide Nanoparticle (TNP) induced mouse Spermatozoon Stem Cells (SSCs) apoptosis, at molecular level. Methods: After isolation from cryptorchid mouse testis and characterization, spermatogonial stem cells were divided into four groups. In the control group, spermatogonial cells were cultured in α-MEM supplemented with 2% BSA (Bovine Serum Albumin). β-Carotene (BC) group was composed of control culture condition supplemented with 1 µg/ml β-carotene. TNP group comprised control culture condition supplemented with 1 µg/ml titanium oxide (TiO2). Ultimately the last group contained control culture condition supplemented with both 1 µg/ml BC and 1 µg/ml TiO2for three days. After that, spermatozoon viability was evaluated by MTT (3-[4,5-dimethylthiazolyl-2]-2,5-diphenyltetrazolium bromide) assay, apoptotic and necrotic indices with Annexin V/PI kit and gene expression of CASP3 and MAPK14 using qRT-PCR method. Results: TiO2 could significantly decrease viability of the cultured spermatozoon in TNP group compared to the control group. In BC group, we determined increased frequency of live spermatozoon compared to TNP or control group. Expression of apoptotic related genes significantly increased in TNP group. Spermatozoon induced by titanium oxide might be useful in clinical procedures. Measurement of apoptosis index using Annexin V/PI method also showed significant increase in apoptotic index of germ cells in TiO2 treated spermatozoon (P<0.05). Conclusion: Expression of apoptotic related genes in cultured spermatozoon could efficiently be decreased by β-carotene treatment. Application of BC had a potential protective effect in preventing apoptosis in germ cells and might be useful in clinic.

Keywords

• titanium oxide
• β-carotene
• spermatozoon
• gene expression