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| Author(s): |
Ana Carina Alves Pereira de Mira Geraldo
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Pirassununga. |
| Institution: | Universidade de São Paulo (USP). Faculdade de Zootecnica e Engenharia de Alimentos (FZE/BT) |
| Defense date: | 2013-08-02 |
| Examining board members: |
Evaldo Antônio Lencioni Titto;
Marcos Roberto Chiaratti;
José Bento Sterman Ferraz;
Ed Hoffmann Madureira;
Alfredo Manuel Franco Pereira
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| Advisor: | Evaldo Antônio Lencioni Titto; Joaquim Fernando Moreira da Silva |
| Abstract | |
The present study aimed to understand the effects triggered by high temperatures in heat shock proteins gene expression, HSPA1A and HSP90AA1, in cow lymphocytes from different breeds and origins. Part of the project was developed in Portugal, where 20 Limousin and 22 Mertolenga breed heifers were used and the second part was developed in Brazil with 11 Holstein Friesian and 20 Brahman heifers. The Holstein animals participated in Experiment I in which four blood samples were collected for subsequent in vitro heat shock, where each one of the blood samples was submitted to different temperatures: 40 ° C and 42 ° C (through water baths), 23 ° C (kept at room temperature) and 10 ° C (in refrigerator) for two hours. All animals from all breeds participated in Experiment II, where they were subjected to the heat tolerance test, where rectal temperature and respiratory rate were measured, and blood samples were collected (after shadow and after sun treatments). In all samples of both experiments was carried out the erythrocytes lysis so as to obtain the buffycoat. The RNA was isolated by the TRIzol method and RT-PCR performed with SuperScript III after digestion with DNase I. The real-time PCR apparatus took place in 7500 Real Fast Time, using TaqMan Gene Expression Assays for HSPA1A and HSP90AA1 target genes, ACTB and PPIA as endogenous genes. The ΔCt (Cttarget - Ctendogenous) were calculated as well as gene expression through the 2-ΔΔCt method. Statistical analysis was performed using linear mixed models, using the program R Project Software (version 3.0.1). In Experiment I it was attested the quality of the relationship for both endogenous with the mRNA-HSPA1A and mRNAHSP90AA1 through a pairwise regression analysis. As expected values, gene expression at a temperature of 23 °C were lower, followed by 10 °C (cold stress), 40 °C and 42 °C for the HSPA1A gene. In the case of HSP90AA1 the higher gene expression was found at 40 °C. In Experiment II, there was a slightly pronounced intra-race variation for respiratory rate values, allowing the assumption that the thermoregulatory effort of animals of the same breed may have been different. Regarding the rectal temperature, in Brahman breeds it was significantly different from the other breeds in the sun treatment. The differences observed were probably a result of different levels of stress to which the animals were subjected. The differences observed in ΔCt were not very expressive and significant differences were only observed in relative gene expression of Mertolenga breed between shade and sun treatments. We conclude that planned increasing temperature in bovine lymphocytes leads to different relative gene expression of HSPA1A and HSP90AA1. The mRNA-HSPA1A expression is greater in higher thermal stress, either by cold or by heat. The HSPA1A and HSP90AA1 relative gene expressions exhibited by animals with different thermolytic capabilities, are also different, existing an individual variability in relative gene expression of heat shock proteins. (AU) | |
| FAPESP's process: | 09/00462-9 - Thermotolerance in cows: cellular and physiological approaches |
| Grantee: | Ana Carina Alves Pereira de Mira Geraldo |
| Support Opportunities: | Scholarships in Brazil - Doctorate |