Hsp70 induction and hsp70 gene polymorphisms as indicators of acclimatization under hyperthermic conditions.

Acclimatization is a process which occurs as a result of repeated mild increases in core temperature, which allows an organism to carry out increased work in the heat due to better heat dissipation (Moseley, 1997). In order to prevent the occurrence of heat illness, it is necessary for individuals who perform work in hot, humid environments to undergo acclimatization. Exposure to different types of stimuli, such as heat exercise and oxidative stress, results in the formation of proteins in the cells which are known as heat shock protein (HSP) (Powers et al, 2001). The main function of HSP is to act as molecular chaperones. Specifically, the 70 kDa HSP, known as Hsp70, play an important part in cellular protection and adaptation during and following exposure to stressful events. Two prominent members of the Hsp70 family are Hsp72, which is the inducible form of Hsp70, and Hsp73, which is the constitutively synthesized form of the protein. Acquisition of thermotolerance in vitro in Chinese hamster fibroblast cells is accompanied by increased Hsp72 levels during a heating episode, indicating that Hsp72 tends to target proteins which have been damaged by a stress situation more than Hsp73 (Li and Werb, 1982). A number of different hsp70 genes are found, which include hsp70-1, hsp70-2 (3¢utr and pst I) and hsp70-hom (Milner and Campbell, 1990). Both the hsp70-1 and hsp70-2 genes encode the primary heat-induced Hsp70 protein as an identical protein product (Hunt and Morimoto, 1985). Following heat shock, no increase in the hsp70-hom mRNA levels is observed (Milner and Campbell, 1990). The use of the Hsp70 protein and hsp70 gene polymorphisms as markers of acclimatization were investigated by subjecting twenty-two individuals to exercise in a hot, humid environment. These individuals were exposed to an initial heat stress, where they participated in a step test at an external work rate of 70 W, followed by participation in an acclimatization program which involved exercising at various combinations of 35 W and 70 W of the external work rate. After acclimatization, the individuals were exposed to a second heat stress, identical to the initial heat stress. The Hsp70 levels both before and after acclimatization were determined in response to heat stress in blood serum by means of the StressXpress ELISA Kit (Stressgen Biotechnologies) and in human monocytes by means of Western blots, using a mouse antiñHsp70 monoclonal primary antibody (Stressgen Biotechnologies) and a goat antiñmouse IgG (H+L) peroxidase conjugated secondary antibody. The hsp70 gene polymorphisms were determined by means of polymerase chain reaction (PCR), using primers specific to the hsp70-1, hsp70-2 (3¢utr), pst I and hsp70-hom genes, so that the genotype combinations for each individual were determined. Blood type was also assessed. It was found that the basal serum Hsp70 levels in individuals who exhibited the ability to acclimatize decreased in response to the acclimatization program, which allowed more Hsp70 to be induced in response to the second heat stress compared to the initial heat stress. The individuals who were unable to acclimatize showed increased basal serum Hsp70 levels in response to acclimatization, which prevented these individuals from inducing high Hsp70 levels in response to the second heat stress. The Hsp70 induced in the monocytes during this program followed the same pattern in both the individuals able to acclimatize and those who were unable to acclimatize, and can therefore not be used as a marker of acclimatization. For the female participants, the current menstrual phase of each woman had to be taken into account, as this had an affect on the core temperature and therefore influenced the division of the female participants into their respective groups. These were the group of individuals who demonstrated the ability to acclimatize or the group of individuals who were unable to acclimatize. The use of oral contraceptives also had to be taken into account, as this too had an influence on the core temperature and therefore also affected the division of the individuals into the group who demonstrated the ability to acclimatize or those who were unable to acclimatize. Cyclic changes during the menstrual cycle may have also changed the Hsp profile. Regarding the hsp70 gene polymorphisms, the A/A, P2 P2 and A1 A1 genotype combination was not present in any of the individuals who were unable to acclimatize, however six of the individuals who showed the ability to acclimatize possessed this genotype combination. The level of induced Hsp70 levels present in the serum of individuals able to acclimatize and the presence or absence of the A/A, P2 P2 and A1 A1 genotype combination therefore have the potential to be used as markers of acclimatization.Dr. M. Cronj