Correction: Impact of Heat Stress on Cellular and Transcriptional Adaptation of Mammary Epithelial Cells in Riverine Buffalo (Bubalus Bubalis).
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ABSTRACT:
SUBMITTER: Kapila N
PROVIDER: S-EPMC5764454 | biostudies-literature | 2018
REPOSITORIES: biostudies-literature
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Correction: Impact of Heat Stress on Cellular and Transcriptional Adaptation of Mammary Epithelial Cells in Riverine Buffalo (Bubalus Bubalis).
Kapila Neha N Sharma Ankita A Kishore Amit A Sodhi Monika M Tripathi Pawan K PK Mohanty Ashok K AK Mukesh Manishi M
PloS one 20180111 1
[This corrects the article DOI: 10.1371/journal.pone.0157237.]. ...[more]
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Project description:The present study aims to identify the heat responsive genes and biological pathways in heat stressed buffalo mammary epithelial cells (MECs). The primary mammary epithelial cells of riverine buffalo were exposed to thermal stress at 42°C for one hour. The cells were subsequently allowed to recover at 37°C and harvested at different time intervals (30 min to 48 h) along with control samples (un-stressed). In order to assess the impact of heat stress in buffalo MECs, several in-vitro cellular parameters (lactate dehydrogenase activity, cell proliferation assay, cellular viability, cell death and apoptosis) and transcriptional studies were conducted. The heat stress resulted in overall decrease in cell viability and cell proliferation of MECs while induction of cellular apoptosis and necrosis. The transcriptomic profile of heat stressed MECs was generated using Agilent 44 K bovine oligonucleotide array and at cutoff criteria of ≥3-or ≤3 fold change, a total of 153 genes were observed to be upregulated while 8 genes were down regulated across all time points post heat stress. The genes that were specifically up-regulated or down-regulated were identified as heat responsive genes. The upregulated genes in heat stressed MECs belonged to heat shock family viz., HSPA6, HSPB8, DNAJB2, HSPA1A. Along with HSPs, genes like BOLA, MRPL55, PFKFB3, PSMC2, ENDODD1, ARID5A, and SENP3 were also upregulated. Microarray data revealed that the heat responsive genes belonged to different functional classes viz., chaperons; immune responsive; cell proliferation and metabolism related. Gene ontology analysis revealed enrichment of several biological processes like; cellular process, metabolic process, response to stimulus, biological regulation, immune system processes and signaling. The transcriptome analysis data was further validated by RT-qPCR studies. Several HSP (HSP40, HSP60, HSP70, HSP90, and HSPB1), apoptotic (Bax and Bcl2), immune (IL6, TNFα and NF-kβ) and oxidative stress (GPX1 and DUSP1) related genes showed differential expression profile at different time points post heat stress. The transcriptional data strongly indicated the induction of survival/apoptotic mechanism in heat stressed buffalo MECs. The overrepresented pathways across all time points were; electron transport chain, cytochrome P450, apoptosis, MAPK, FAS and stress induction of HSP regulation, delta Notch signaling, apoptosis modulation by HSP70, EGFR1 signaling, cytokines and inflammatory response, oxidative stress, TNF-alpha and NF- kB signaling pathway. The study thus identified several genes from different functional classes and biological pathways that could be termed as heat responsive in buffalo MEC. The responsiveness of buffalo MECs to heat stress in the present study clearly suggested its suitability as a model to understand the modulation of buffalo mammary gland expression signature in response to environmental heat load.
| S-EPMC5040452 | biostudies-literature
Project description:The primary mammary epithelial cells of riverine buffalo were exposed to thermal stress condition at 42C for one hour. The cells were subsequently allowed to recover at 37C and harvested at different time intervals (30 min to 48 h) along with control samples (un-stressed). The study has identified several genes from different functional classes and biological pathways which could be termed as heat responsive in buffalo MEC
2016-08-26 | GSE84486 | GEO
Project description:BackgroundThe objective of this study was to establish the buffalo mammary epithelial cell line (BuMEC) and characterize its mammary specific functions.MethodologyBuffalo mammary tissue collected from the slaughter house was processed enzymatically to obtain a heterogenous population of cells containing both epithelial and fibroblasts cells. Epithelial cells were purified by selective trypsinization and were grown in a plastic substratum. The purified mammary epithelial cells (MECs) after several passages were characterized for mammary specific functions by immunocytochemistry, RT-PCR and western blot.Principal findingsThe established buffalo mammary epithelial cell line (BuMEC) exhibited epithelial cell characteristics by immunostaining positively with cytokeratin 18 and negatively with vimentin. The BuMEC maintained the characteristics of its functional differentiation by expression of β-casein, κ-casein, butyrophilin and lactoferrin. BuMEC had normal growth properties and maintained diploid chromosome number (2n = 50) before and after cryopreservation. A spontaneously immortalized buffalo mammary epithelial cell line was established after 20 passages and was continuously subcultured for more than 60 passages without senescence.ConclusionsWe have established a buffalo mammary epithelial cell line that can be used as a model system for studying mammary gland functions.
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Project description:The Hox complex contains 39 genes clustered into four groups involved in cell differentiation and development. We cloned full-length sequence of Hoxc11 gene from water buffalo Bubalus bubalis, assessed its copy number, localized the same onto the chromosome 5, and studied its evolutionary conservation across the species. Northern hybridization of Hoxc11 showed a 2.2 kb band in the tissues analyzed. Real-Time PCR showed highest expression of Hoxc11 gene in lung followed by spleen, spermatozoa, and testis. Six interacting partners of this gene showed higher expression in spleen, lung, testis, and spermatozoa. During the early stages of development, Hoxc11 and its interacting partners both showed lower expression, which then became prominent during the age of 1-3 years, regressed drastically thereafter, and remained so until the animal's life time (∼ 20 years). The high expression of Hoxc11 and its interacting partners in spermatozoa and testis during the onset of puberty suggests its likely role in the differentiation of gonads and subsequent reproductive activities. Additional work on Hoxc11 especially, in the context of respiratory, immunological, and in/fertility in other species, including humans would be useful for establishing its broader biological significance towards the enrichment of functional and comparative genomics.
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Project description:Gene expression studies require appropriate normalization methods for proper evaluation of reference genes. To date, not many studies have been reported on the identification of suitable reference genes in buffaloes. The present study was undertaken to determine the panel of suitable reference genes in heat-stressed buffalo mammary epithelial cells (MECs). Briefly, MEC culture from buffalo mammary gland was exposed to 42 °C for one hour and subsequently allowed to recover at 37 °C for different time intervals (from 30 m to 48 h). Three different algorithms, geNorm, NormFinder, and BestKeeper softwares, were used to evaluate the stability of 16 potential reference genes from different functional classes. Our data identified RPL4, EEF1A1, and RPS23 genes to be the most appropriate reference genes that could be utilized for normalization of qPCR data in heat-stressed buffalo MECs.
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Project description:Polymorphism within the promoter region of bovine thyroglobulin has been reported to be associated with milk and meat quality. In this study, we investigated the genetic variation within thyroglobulin promoter region of swamp and riverine buffaloes using PCR-SSCP technique and sequencing, and also analyzing association of polymorphism with the milk production traits. The study revealed four conformational patterns, A, B, C, and D among 323 buffaloes of two riverine breeds and different swamp populations. The frequency of SSCP variant 'A' was found to be invariably high among all buffalo populations. Variant 'C' was found to be absent in pure swamp population and present with higher frequency among riverine dairy breeds Mehsana and Nili Ravi. Frequency of D variant was observed to be highest in buffalo population, representing riverine and hybrid types. Sequencing of three representative PCR products of each of the SSCP variants, revealed three polymorphic sites responsible, 33C > T, 176G > T and 221C > T, in the buffalo TG promoter region. Further, association studies of SSCP variants with various milk production and milk quality traits indicated significant effect on fat percentage in buffaloes belonging to Mehsana and Nili Ravi dairy breeds. The preliminary results also showed the substantial variations in the distribution of SSCP variants' frequencies across swamp and riverine buffaloes, two distinct populations being reared for meat and milk production, respectively.
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Project description:Heat stress has a detrimental effect on the physiological and production performance of buffaloes. Elucidating the underlying mechanisms of heat stress is challenging, therefore identifying candidate genes is urgent and necessary. We evaluated the response of buffaloes (n = 30) to heat stress using the physiological parameters, ELISA indexes, and hematological parameters. We then performed mRNA and microRNA (miRNA) expression profiles analysis between heat tolerant (HT, n = 4) and non-heat tolerant (NHT, n = 4) buffaloes, as well as the specific modules, significant genes, and miRNAs related to the heat tolerance identified using the weighted gene co-expression network analysis (WGCNA). The results indicated that the buffaloes in HT had a significantly lower rectal temperature (RT) and respiratory rate (RR) and displayed a higher plasma heat shock protein (HSP70 and HSP90) and cortisol (COR) levels than those of NHT buffaloes. Differentially expressed analysis revealed a total of 753 differentially expressed genes (DEGs) and 16 differentially expressed miRNAs (DEmiRNAs) were identified between HT and NHT. Using the WGCNA analysis, these DEGs assigned into 5 modules, 4 of which were significantly correlation with the heat stress indexes. Interestingly, 158 DEGs associated with heat tolerance in the turquoise module were identified, 35 of which were found within the protein-protein interaction network. Several hub genes (IL18RAP, IL6R, CCR1, PPBP, IL1B, and IL1R1) were identified that significantly enriched in the Cytokine-cytokine receptor interaction. The findings may help further elucidate the underlying mechanisms of heat tolerance in buffaloes.
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