SUMMARY:The increase of cell proliferation during early wound healing is thought to be regulated by a decrease of apoptosis.In contrast, the reduction of cellularity during final wound maturation may be controlled by an increase of apoptotic cell death. Herein we studied whether p53 is involved in wound healing-associated apoptosis and whether transient inhibition of p53 is effective to improve the early healing process of cutaneous wounds. Using intravital microscopic and immunohistochemical techniques in hairless mice, we demonstrated that in vivo inhibition of p53 by pifithrin-␣ (PFT-␣; 2.2 mg/kg ip) accelerates early epithelialization and neovascularization of cutaneous wounds by (i) promoting leukocyte recruitment, (ii) increasing cell proliferation, and (iii) reducing apoptotic cell death. We further show that final wound closure with down-regulation of cell proliferation is not inhibited by PFT-␣ treatment, indicating that transient blockade of p53 function does not affect the process of wound maturation. Western blot analysis revealed that PFT-␣ lowered nuclear but not cytoplasmic p53, implying that cytoplasmic retention of p53 mediates the antiapoptotic effects of PFT-␣. Furthermore, PFT-␣ significantly increased expression of proliferating cell nuclear antigen protein in whole extracts of cutaneous tissue and caused a rise in proliferation of wild-type, but not mutant, p53-expressing keratinocytes. From our study we conclude that transient inhibition of p53 supports the early cell proliferation required for rapid tissue repair and that this may represent an attractive approach in the treatment of delayed wound
Early graft dysfunction due to ischemia reperfusion injury remains a major clinical challenge in liver transplantation. Because apoptosis may contribute to graft dysfunction, we studied whether transient inhibition of p53 is capable of improving graft quality by reducing apoptotic cell death. Rat livers were harvested and stored for 24 hours or 48 hours in a 4°C solution containing either pifithrin-␣ (PFT-␣), a specific p53-inhibitor, or the vehicle dimethyl-sulfoxide. Storage was followed by 2-hour reperfusion with 37°C Krebs-Henseleit buffer in an isolated liver perfusion system. Besides caspase-3 activation, apoptosis was quantified using fluorescence microscopy and hematoxylin-eosin histology. Trypan blue allowed for assessment of cell membrane damage, indicating both secondary apoptosis and primary necrosis. Bile flow, oxygen consumption, K ؉ -excretion and enzyme release served as indicators of overall graft quality. Upon 2-hour reperfusion, livers developed procaspase activation as well as a mixture of apoptotic and necrotic cell death, representing necrapoptosis. In livers that had been stored for 48 hours, necrapoptotic injury was more pronounced compared with that after 24-hour storage. PFT-␣ effectively attenuated caspase activation as well as hepatocellular apoptosis and necrosis. Attenuation of both modes of cell death by PFT-␣ was associated with improved liver function, metabolism, and integrity. Experiments with the caspase inhibitor z-VAD-fmk confirmed that apoptosis is one mode of cell death in cold ischemia reperfusion. In conclusion, inhibition of p53-dependent apoptosis by PFT-␣ reduces hepatic preservation-reperfusion injury and improves primary organ function and metabolism. Fortification of the preservation solution with PFT-␣ may represent a promising and easily applicable approach to mitigate reperfusion injury in liver transplants.
Sonoelastography may be superior to other US modalities in elucidating different cervical lymph node biopsy helping to distinguish benign from malignant lesions. This may replace the lymph node biopsies in the future. Moreover, its use in the follow-up of patients with cervical malignancies may reduce the number of future biopsies. Further studies with more patients may be needed for a better assessment of results.
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