Differential expression of myocardial heat shock proteins in rats acutely exposed to fluoride

Abstract

Acute fluoride (F^-) toxicity is known to cause severe cardiac complications and leads to sudden heart failure. Previously, we reported that increased myocardial oxidative damage, apoptosis, altered cytoskeleton and AMPK signaling proteins associated with energy deprivation in acute F^- induced cardiac dysfunction. The present study was aimed to decipher the status of myocardial heat shock proteins (Hsps-Hsp27, Hsp32, Hsp40, Hsp60, Hsp70, Hsp90) and heat shock transcription factor 1 (Hsf1) in acute F^--intoxicated rats. In order to study the expression of myocardial Hsps, male Wistar rats were treated with single oral doses of 45 and 90 mg/kg F^- for 24 h. The expression levels of myocardial Hsps were determined using RT-PCR, western blotting, and immunohistochemical studies. Acute F^--intoxicated rats showed elevated levels of both the transcripts and protein expression of Hsf1, Hsp27, Hsp32, Hsp60, and Hsp70 when compared to control. In addition, the expression levels of Hsp40 and Hsp90 were significantly declined in a dose-dependent fashion in F^--treated animals. Our result suggests that differential expression of Hsps in the rat myocardium could serve as a balance between pro-survival and death signal during acute F^--induced heart failure.

Keywords: Acute fluoride toxicity; Cardiac dysfunction; Hsf1; Hsps.

Fig. 1

Effect of acute F⁻ intoxication on myocardial mRNA expression of Hsf1 and Hsps in rats. mRNA expression and its densitometric analysis of myocardial Hsf1 and Hsps (Hsp27, Hsp32, Hsp40, Hsp60, Hsp70, and Hsp90) in control and F⁻-treated rats. GAPDH was used as loading control. The values represent mean ± SD (n = 3). *p < 0.05 significantly different from control, ⁺ p < 0.05 significantly different from 45 mg F⁻-treated rats (one-way ANOVA followed by Tukey’s multiple comparison test)

Fig. 2

Effect of acute F⁻ intoxication on myocardial protein expression of Hsf1 and Hsps in rats. Western blotting and its densitometric analysis of myocardial Hsf1 and Hsps (Hsp27, Hsp32, Hsp40, Hsp60, Hsp70, and Hsp90) in control and F⁻-treated rats. GAPDH was used as loading control. The values represent mean ± SD (n = 3). *p < 0.05 significantly different from control, ⁺ p < 0.05 significantly different from 45 mg F⁻ treated rats (one-way ANOVA followed by Tukey’s multiple comparison test)

Fig. 3

a Effect of acute F⁻ intoxication on myocardial immunohistochemical expression of Hsf1 and low molecular weight Hsps in rats. Myocardial sections were stained with antibodies specific for Hsf1, Hsp27, Hsp32, and Hsp40 in control and F⁻-intoxicated rats. Number of immunopostively stained cells in ten fields/section (×200) was quantified by three independent observers in blinded fashion. The bar diagram shows the quantification of positively stained cells of control and F⁻-treated rats. The values represent mean ± SD (n = 3). *p < 0.05 significantly different from control, ⁺ p < 0.05 significantly different from 45 mg F⁻-treated rats (one-way ANOVA followed by Tukey’s multiple comparison test). b Effect of acute F⁻ intoxication on myocardial immunohistochemical expression of high molecular weight Hsps in rats. Myocardial sections were stained with antibodies specific of Hs60, Hsp70, and Hsp90 (×200) in control and F⁻ intoxicated rats. Number of immunopostively stained cells in ten fields/section (×200) was quantified by three independent observers in blinded fashion. The bar diagram shows the quantification of positively stained cells of control and F⁻ treated rats. The values represent mean ± SD (n = 3). *p < 0.05 significantly different from control, ⁺ p < 0.05 significantly different from 45 mg F⁻-treated rats (one-way ANOVA followed by Tukey’s multiple comparison test)