In vivo multiphoton microscopy detects longitudinal metabolic changes associated with delayed skin wound healing.
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ABSTRACT:
SUBMITTER: Jones JD
PROVIDER: S-EPMC6242983 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Although there is growing evidence that cellular senescence influences wound healing, a clear understanding of how senescence can be beneficial and/or detrimental to wound healing is unknown. Wound healing may also be influenced by the baseline tissue senescence, which is elevated in aging and chronic wounds, both of which have significant healing delays. To study the effects of skin senescence on wound healing, we developed an elevated skin senescence model based on the subcutaneous transfer of irradiated fibroblasts into young 8-week-old wild-type C57BL/6 male mice. This senescent cell transfer significantly increased skin senescence levels compared to control transfers of non-irradiated fibroblasts. There was an increased presence of SA-β-Gal- and p21-positive senescent cells throughout the skin. Furthermore, the entire skin showed significantly elevated gene expression of senescence (p16, p21) and SASP markers (IL-6, MCP-1, MMP-3, MMP-9, and TGF-β). Subsequent wound healing in the skin with elevated senescence was markedly delayed and had similar kinetics to naturally aged 2-year-old mice. After the wounds had healed, the skin developed persistently elevated senescence. Our results demonstrate that states of elevated skin senescence can delay wound healing and result in sustained senescence after healing. Therefore, the accumulation of senescent cells in aged skin or chronic wounds may be a driver of delayed healing and can be considered a potential target to improve healing.
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Project description:Macrophages play essential roles throughout the wound repair process. Nevertheless, mechanisms regulating the process are poorly understood. MAFB is specifically expressed in the macrophages in hematopoietic tissue and is vital to homeostatic function. Comparison of the skin wound repair rates in macrophage-specific, MAFB-deficient mice (Mafbf/f::LysM-Cre) and control mice (Mafbf/f) showed that wound healing was significantly delayed in the former. For wounded GFP knock-in mice with GFP inserts in the Mafb locus, flow cytometry revealed that their GFP-positive cells expressed macrophage markers. Thus, macrophages express Mafb at wound sites. Immunohistochemical (IHC) staining, proteome analysis, and RT-qPCR of the wound tissue showed relative downregulation of Arg1, Ccl12, and Ccl2 in Mafbf/f::LysM-Cre mice. The aforementioned genes were also downregulated in the bone marrow-derived, M2-type macrophages of Mafbf/f::LysM-Cre mice. Published single-cell RNA-Seq analyses showed that Arg1, Ccl2, Ccl12, and Il-10 were expressed in distinct populations of MAFB-expressing cells. Hence, the MAFB-expressing macrophage population is heterogeneous. MAFB plays the vital role of regulating multiple genes implicated in wound healing, which suggests that MAFB is a potential therapeutic target in wound healing.
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Project description:ImportanceBasal cell carcinomas (BCCs) are diagnosed by clinical evaluation, which can include dermoscopic evaluation, biopsy, and histopathologic examination. Recent translation of multiphoton microscopy (MPM) to clinical practice raises the possibility of noninvasive, label-free in vivo imaging of BCCs that could reduce the time from consultation to treatment.ObjectivesTo demonstrate the capability of MPM to image in vivo BCC lesions in human skin, and to evaluate if histopathologic criteria can be identified in MPM images.Design, setting, and participantsImaging in patients with BCC was performed at the University of California-Irvine Health Beckman Laser Institute & Medical Clinic, Irvine, between September 2012 and April 2014, with a clinical MPM-based tomograph. Ten BCC lesions were imaged in vivo in 9 patients prior to biopsy. The MPM images were compared with histopathologic findings.Main outcomes and measuresMPM imaging identified in vivo and noninvasively the main histopathologic feature of BCC lesions: nests of basaloid cells showing palisading in the peripheral cell layer at the dermoepidermal junction and/or in the dermis.ResultsThe main MPM feature associated with the BCC lesions involved nests of basaloid cells present in the papillary and reticular dermis. This feature correlated well with histopathologic examination. Other MPM features included elongated tumor cells in the epidermis aligned in 1 direction and parallel collagen and elastin bundles surrounding the tumors.Conclusions and relevanceThis study demonstrates, in a limited patient population, that noninvasive in vivo MPM imaging can provide label-free contrast that reveals several characteristic features of BCC lesions. Future studies are needed to validate the technique and correlate MPM performance with histopathologic examination.
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