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In vitro culture of feline embryos increases stress-induced heat shock protein 70 and apoptotic related genes.

ABSTRACT: Developmental competence and quality of in vitro produced embryos has been demonstrated to be lower than in vivo derived embryos. This study aimed specifically to determine the effects of in vitro culture of feline embryos using various culture densities on developmental competence and expression of stress- and apoptotic-related genes in terms of heat shock protein 70 (HSP70) and apoptotic-related (BAX and BCL-2) gene expressions. In experiment 1, we characterized the inducible form of a feline-specific HSP70 mRNA sequence, as it has not been previously reported. The primers for feline HSP70 mRNA were synthesized and tested on heat-treated cat fibroblasts. In experiment 2, feline embryos were cultured at different culture densities (embryo:culture volume; 1:1.25, 1:5 and 1:20). The developmental competence was determined along with HSP70, BAX and BCL-2 transcript abundances using quantitative RT-PCR. In vivo derived embryos were used as a control group. A partial cat HSP70 mRNA sequence (190 bp) was characterized and exhibited high nucleotide identity (93 to 96%) with other species. Cleaved embryos cultured at high density (1:1.25) developed to blastocysts at a lower rate than those generated from lower densities. Irrespective of the culture densities used, in vitro cultured blastocysts showed increased levels of HSP70 and BAX transcripts compared with in vivo counterparts. Blastocysts derived from the highest culture density (1:1.25) showed higher levels of upregulation of HSP70 and BAX transcripts than those cultured at lower culture densities (1:5 and 1:20). In conclusion, increased levels of pro-apoptotic (BAX) and stress-response (HSP70) transcripts correlated with developmental incompetence of embryos cultured at high embryonic density, indicating that stress accumulated during in vitro embryo culture affected the fate for embryo development and quality.

SUBMITTER: Sananmuang T

PROVIDER: S-EPMC3934196 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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In vitro culture of feline embryos increases stress-induced heat shock protein 70 and apoptotic related genes.

Sananmuang Thanida T Phutikanit Nawapen N Nguyen Catherine C Manee-In Sukanya S Techakumphu Mongkol M Tharasanit Theerawat T

The Journal of reproduction and development 20130125 2

Developmental competence and quality of in vitro produced embryos has been demonstrated to be lower than in vivo derived embryos. This study aimed specifically to determine the effects of in vitro culture of feline embryos using various culture densities on developmental competence and expression of stress- and apoptotic-related genes in terms of heat shock protein 70 (HSP70) and apoptotic-related (BAX and BCL-2) gene expressions. In experiment 1, we characterized the inducible form of a feline- ...[more]

PMID: 23358310

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Project description:It has been established that the synthesis of heat shock protein 70 (Hsp70) is temperature-dependent. The Hsp70 response is considered as a cellular thermometer in response to heat stress and other stimuli. The variation in Hsp70 gene expression has been positively correlated with thermotolerance in Drosophila melanogaster, Caenorhabditis elegans, rodents and human. Goats have a wide range of ecological adaptability due to their anatomical and physiological characteristics; however, the productivity of the individual declines during thermal stress. The present study was carried out to analyze the expression of heat shock proteins in different tissues and to contrast heat stress phenotypes in response to chronic heat stress. The investigation has been carried out in Jamunapari, Barbari, Jakhrana and Sirohi goats. These breeds differ in size, coat colour and production performance. The heat stress assessment in goats was carried out at a temperature humidity index (THI) ranging from 85.36-89.80 over the period. Phenotyping for heat stress susceptibility was carried out by combining respiration rate (RR) and heart rate (HR). Based on the distribution of RR and HR over the breeds in the population, individual animals were recognized as heat stress-susceptible (HSS) and heat stress-tolerant (HST). Based on their physiological responses, the selected animals were slaughtered for tissue collection during peak heat stress periods. The tissue samples from different organs such as liver, spleen, heart, testis, brain and lungs were collected and stored at -70 °C for future use. Hsp70 concentrations were analyzed from tissue extract with ELISA. mRNA expression levels were evaluated using the SYBR green method. Kidney, liver and heart had 1.5-2.0-fold higher Hsp70 concentrations as compared to other organs in the tissue extracts. Similarly, the gene expression pattern of Hsp70 in different organs indicated that the liver, spleen, brain and kidney exhibited 5.94, 4.96, 5.29 and 2.63-fold higher expression than control. Liver and brain tissues showed the highest gene expression at mRNA levels as compared to kidney, spleen and heart. HST individuals had higher levels of mRNA level expression than HSS individuals in all breeds. The Sirohi breed showed the highest (6.3-fold) mRNA expression levels as compared to the other three breeds, indicating the better heat stress regulation activity in the breed.

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In vitro culture of feline embryos increases stress-induced heat shock protein 70 and apoptotic related genes.

Publications

In vitro culture of feline embryos increases stress-induced heat shock protein 70 and apoptotic related genes.

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