Fig 1: Recombinant human HSPB5 (0.01 mg/mL) and human HSPB1 (0.01 mg/mL) reduce stiffness of permeabilized cardiomyocytes of obese ZSF1 rats and abrogate the effect of oral GGA treatment. Stiffness was measured as the passive force (Fpassive). (a and b) SL‐Fpassive curves after HSP treatment for lean, vehicle‐treated obese and GGA‐treated obese ZSF1 rats after incubation with recombinant human HSPB5 (a) or human HSPB1 (b). A second order polynomial (quadratic function) was used for curve fitting. For comparison, the Fpassive curve of cardiomyocytes isolated from vehicle‐treated obese ZSF1 rats (dashed line, triangles) and the Fpassive curve of cardiomyocytes isolated from lean rats are shown in panels a and b. (c–f) comparison of HSP effects on Fpassive of cardiomyocytes in vehicle‐ and GGA‐treated rats. Data are expressed as mean ± SD. ‡ p < 0.0001, † p = 0.0001 and *p < 0.05 vs. baseline of the same group. N = 4–5 per group, with four cardiomyocytes tested per rat. Effects of recombinant sHSPs were established using a two‐way ANOVA with a Bonferroni's post hoc test to correct for differences between the groups.
Fig 2: Oral GGA treatment redistributes HSPB5 and HSPB1 from the cytosol to the myofilaments in the left ventricle of obese ZSF1 rats. (a and b) HSPB5 and HSPB1 levels were similar between myocardia of all groups in cytosolic fraction. (c and d) myofilament levels of HSPB5 and HSPB1 were higher in the myocardium of obese ZSF1 rats treated with GGA when compared to vehicle‐treated obese and lean ZSF1 rats. (e and f) There was a redistribution of HSPB5 and HSPB1 to the myofilaments (myofilament expression levels relative to cytosolic expression levels) in the GGA‐treated obese ZSF1 rats compared to lean and vehicle‐treated obese ZSF1 rats, although this was particularly strong for HSPB1. Data are expressed as mean ± SEM, n = 4–5 per group. A one‐way ANOVA with a Bonferroni's post hoc test was utilized to assess differences between the groups. (g and h) Representative images of confocal laser microscopy for HSPB5 (upper panel) and HSPB1 (lower panel) in LV sections from lean, obese and obese ZSF1 rats treated with GGA (200 mg/kg/day). Immunohistochemical visualization of cell membranes (WGA; green), nuclei (DAPI; blue), and HSPB5/HSPB1 (red) was achieved with confocal laser microscopy. Both HSPB5 and HSPB1 immunostaining primarily occurred in the vicinity of the myofilaments for all rats. Greater HSPB5 immunostaining in the vicinity of the myofilaments was observed in both vehicle‐treated and GGA‐treated obese ZSF1 rats, whereas prominent elevation of HSPB1 immunostaining was noted in the GGA‐treated obese ZSF1 rats. n = 4 per group.
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