Interleukin 6 (IL6) and p53 are linked by mutual regulatory mechanisms and are both upregulated in aging. The aim of this study was to evaluate the effects of aging and IL-6 on expression of p53 in the mouse heart. Male C57BL6/J wild-type (WT) and IL6 knock-out (IL6KO) mice at the age of 4-5 months (young adult) and 24-30 months (old) were used. Myocardial expression of proteins: p53, p21, Mdm2, and phospho-Akt/Akt was estimated using western blotting and expression of p53 and p21 mRNA using real-time-PCR. Expression of p53 protein was lower in IL6KO than in WT hearts. Aging caused significant upregulation of p53 protein level, however it was significantly higher in old WT than in old IL6KO hearts (p<0.05). Similar p53 mRNA levels in all groups implied IL6 influence on age-related proteasomal degradation of p53. Localization of p53 mainly in the extranuclear compartment and lack of p21 upregulation in aged hearts may suggest quenched transcriptional activity of p53 despite increased abundance of p53. We conclude that lack of IL6 attenuates expression of p53 protein in the hearts of young mice and diminishes its accumulation with aging by post-transcriptional mechanisms, however this is not related to altered phenotype of aging heart.
Our earlier studies demonstrated slower age-related memory decline in IL-6-deficient than in control mice. Therefore, in the present study we evaluated the effect of IL-6 deficiency and aging on expression of p53, connected with accumulation of age-related cellular damages, in hippocampus of 4and 24-month-old IL-6-deficient C57BL/6J (IL-6KO) and wild type control (WT) mice. The accumulation of p53 protein in hippocampus of aged IL-6KO mice was significantly lower than in aged WT ones, while p53 mRNA level was significantly higher in IL-6-deficient mice, what indicates that the effect was independent on p53 transcription. Presence of few apoptotic cells in hippocampal dentate gyrus and lack of changes in levels of pro-apoptotic Bax, antiapoptotic Bcl-2, as well as in p21 protein in aged animals of both genotypes, points to low transcriptional activity of p53, especially in aged WT mice. Because the amount of p53 protein did not correlate with the level of Mdm2 protein, its main negative regulator, other than Mdm2dependent mechanism was involved in p53 build-up. Significantly higher mRNA levels of autophagyassociated genes: Pten, Tsc2, and Dram1 in IL-6KO mice, in conjunction with significantly lower amount of Bcl-2 protein in 4-month-old IL-6KO mice, suggests that lack of IL-6/STAT3/Bcl-2 signaling could account for better autophagy performance in these mice, preventing excessive accumulation of proteins. Taken together, attenuated p53 protein buildup, absence of enhanced apoptosis, and transcriptional up-regulation of autophagy-associated genes imply that IL-6 deficiency may protect hippocampus from age-related accumulation of cellular damages.
The significance of plasminogen activation during the tympanic membrane (TM) healing is known mainly from studies performed on knock-out mice. In the previous study, we reported activation of genes coding proteins of plasminogen activation and inhibition system in rat’s TM perforation healing. The aim of the present study was the evaluation of protein products expressed by these genes and their tissue distribution using Western blotting and immunofluorescent method, respectively, during 10-day observation period after injury. Otomicroscopical and histological evaluation were employed to assess the healing process. The expression of urokinase plasminogen activator (uPA) and its receptor (uPAR) were significantly upregulated in the proliferation phase, with subsequent gradual attenuation during remodeling phase of healing process, when keratinocyte migration was weakening. The expression of plasminogen activator inhibitor type 1 (PAI-1) also showed the highest levels during the proliferation phase. The increase of tissue plasminogen activator (tPA) expression was observed during the whole observation period, with the highest activity during the remodeling phase. Immunofluorescence of these proteins was present mainly in migrating epithelium. Our study found that plasminogen activation (uPA, uPAR, tPA) and inhibitory (PAI-1) molecules form a well-structured regulatory system of the epithelial migration that is critical to the healing of TM after its perforation.
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