Cell culture media dependent in vitro dynamics and culture characteristics of adult caprine dermal fibroblast cells.

The enhanced availability of functional fibroblasts from precious tissue samples requires an ideal cell-culture system. Therefore, this study was designed to investigate the performance of caprine adult fibroblast cells (cadFibroblast) when cultivated in different culture media. The cadFibroblast cell lines from adult Barbari (Capra hircus) bucks were established and the effect of different media viz. DMEM/F-12 [with low-glucose (5.5 mM; DL) and high-glucose (30 mM; DH)], α-MEM [with low-glucose (5.5 mM; ML) and with high-glucose (30 mM; MH)], and fibroblast growth medium (FGM) were evaluated. Cells were then compared for growth characteristics and in-vitro dynamics through cellular morphology, proliferation, population-doubling time, double-immunocytochemistry, colony-forming units, wound healing, transwell migration, and differential expression of fibroblast-specific markers (FSP-1 and vimentin). The results of immunocytochemistry, transwell migration/invasion, and wound healing assays showed the superiority of DH over DL and other media tested. Whereas, similar effects of glucose supplementation and expression of FSP-1 were not observed in α-MEM. Transwell migration was significantly (p < 0.05) lower in FGM compared with other media tested. Overall, our results illustrate the media-dependent deviation in in-vitro dynamics and culture characteristics of cadFibroblasts that may be useful to develop strategies to cultivate these cells efficiently for research and downstream applications.

Genetic studies of heat stress regulation in goat during hot climatic condition.

Various direct and indirect environmental constraints have an impact on livestock performance. The physiological parameters, such as rectal temperature, heart rate, and respiratory rate, are the primary indicators of thermal stress. Under a stressed environment temperature humidity index (THI) had established as a vital measurement to identify the thermal stress in livestock. THI in association with climatic variations can define the environmental effect as stressful or comfortable for livestock. Goats are small ruminants that adapt to a wide range of ecological variations due to their anatomical and physiological characteristics. However, the productivity of animals declines at the individual level during thermal stress. Stress tolerance can be determined through genetic studies associated with at the cellular level using physiological as well as molecular approaches. Information on genetic association with thermal stress in goats is scanty, this severely affects their survival and hence productivity of livestock. The ever-increasing demand for food across the globe needs deciphering novel molecular markers as well as stress indicators that play a vital role in livestock improvement. This review represents an analysis of current knowledge of phenotypic differences during thermal stress and signifies the importance of physiological responses and their association at the cellular level in goats. The regulation of vital genes associated with thermal stress such as Aquaporins (AQP 0, 1, 2, 4, 5, 6, 8), aquaglyceroporins (AQP3, 7, 9, and 10) and super-aquaporins (AQP 11, 12); BAX inhibitors such as PERK (PKR like ER kinase), IRE 1(inositol-requiring-1); Redox regulating genes such as NOX; Transport of Na+ and K+ such as ATPase (ATP1A1) and several heat shock proteins have been implicated in heat-stress related adaptations have been elucidated. As these changes have a significant impact on production performance as well as on livestock productivity. Such efforts may help in the development of molecular markers and will assist the breeders to develop heat-tolerant goats with improved productivity.

Differential expression and regulation of HSP70 gene during growth phase in ruminants in response to heat stress.

Heat shock proteins regulate the physiological mechanism of heat stress adaptation at cellular level. The present investigation was carried out to analyse the HSP70 gene regulation in various growth stage in ruminants in peripheral blood mononuclear cells (PBMCs). The relationship between HSP gene expression and thermotolerance in age-specific manner in ruminants has not been analysed. Therefore m-RNA HSP70 expression level was examined in different age groups of Jamunpari goat during hot climatic conditions. The experiment was carried out in 32 animals of Jamunapari goat belonging to the age groups of 3-months, 9-months, 12-months, and adults (2-3 year). Total RNA was isolated from peripheral blood mononuclear cells. The physiological response such as rectal temperature (RT), respiration rate (RR) and heart rate (HR) was used as indicator to heat stress. Temperature Humidity Index (THI) was used as an indicator of severity of environmental stress. The THI range varied from 82.00-92.08 during experimental period. The m-RNA HSP70 expression level at 9-month age of animals was up-regulated and significantly higher than other age groups. It was observed that the level of HSP70 transcripts in PBMCs was highest at 9-month age group, and age-related decline in HSP70 expression was observed in adult age. Based on the physiological response, the contrasting heat-stress phenotypes were recognised as heat stress susceptible (HSS) and heat stress tolerant (HST) individuals and the expression of m-RNA HSP70 was analysed at different ages in response to chronic heat stress. The differential mRNA expression of HSS individuals at 3 and 9-month of age showed the highest fold expression than HST. Age and phenotype had significant effect (p < 0.01) on the crossing point (CP) value. The m-RNA HSP70 gene expression in different age groups was correlated with heat stress tolerance and this could be used as biomarker for breeders to analyse the HSP response in -vivo in ruminants.

Estimates of genetic parameters for linear body measurements and prediction of body weight in goat.

The objective of this study is to estimate genetic parameters for linear body measurements along with their correlation to live weight with a focus on devising a scale to predict live weights from body measurement. A total of 142,564 records on body measures and live weights were collected from 8701 Jamunapari goats. Genetic parameters were obtained for body length (L), height at withers (H) and heart girth (G) from birth to adult stage by univariate and multivariate analysis using the average information restricted maximum likelihood method. The best model for body measures at birth included the additive effect of animal and dam along with their covariance and maternal environment, whereas for traits measured later in life, the maternal environment was not significant. After accounting for the direct maternal correlation (ram ), the total heritability estimates for linear body measurements (L,H and G) at the preweaning and postweaning stages of growth ranged from 0.14 to 0.20. Significant genetic variability implies further scope for selection. The genetic correlations of live weight at birth, 3, 6, 9 and 12 months with corresponding L,H and G were high in magnitude indicating scope to select animals for higher weight using morphometric measurements. When weighing scales are unavailable in the field, prediction of weight using L and G was recommended [live weight = (0.291 × L) + (0.306 × G) - 16.8]. We recommend the use of body measurements in the Jamunapari goat breeding program owing to their high genetic correlation with corresponding live weights.

Differential expression and characterization of ATP1A1 exon17 gene by high resolution melting analysis and RT-PCR in Indian goats.

The investigation was carried out to analyse the genetic polymorphism and gene expression of ATP1A1 gene in four different Indian goat breeds by using high resolution melting (HRM) and real time-PCR. ATPase is electro-genic ion pump which is maintains the balance of sodium and potassium ions in animal cells. The transport of Na+& K+ is variable at cellular level during extreme hot period. Therefore, susceptible and tolerant animals were selected based on the physiological responses during hot period. Blood samples were collected from individuals, DNA was isolated. The 300 bp fragment of ATP1A1 gene was amplified by PCR and HRM genotyping was performed. The melting curves were analysed, differential temperature-shift plot showed three different genotypes in all the analysed samples. Out of the 135 samples, the distribution percentages were 55.56% (AA/blue), 33.33% (AC/red) and 11.11% (CC/green). The sequence variation revealed a SNP at 143rd position (A>C). The nucleotide diversity was 0.695 ± 0.403, 0.732 ± 0.424, 0.662 ± 0.433 and 0.687 ± 0.398 in Barbari, Jamunapari, Jakharna and Sirohi, respectively. The respiration rate (RR) was significantly different (P < 0.05) between AA and AC (t = 1.875, df = 38) genotype and heart rate (HR) was significantly different (P < 0.05) between AA and CC genotype. The relative expression pattern of ATP1A1 in SNP variants and non-variants animal tissues showed 19.09 and 6.93 fold higher than control (non-variant), respectively. Jamunapari showed higher fold value of ATP1A1 gene in comparison to Barbari, Jakharna and Sirohi. However, the heat stress-susceptible phenotype had significantly higher gene expression than stress-tolerant in all the breeds. The variation may be used as a marker for selection on the basis of physiological parameters and expression of ATP1A1 gene in goats indicating the specificity of expression in each tissue.