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Enhancement of Wound Healing by Conditioned Medium of Adipose-Derived Stromal Cell with Photobiomodulation in Skin Wound.

ABSTRACT:

Background and objectives

The objective of this study was to investigate whether conditioned medium from photobiomodulation (PBM) irradiated adipose-derived stromal cell (ASC) spheroids prior to implanting could stimulate angiogenesis and tissue regeneration to improve functional recovery of skin tissue in an animal skin wound model.

Methods and results

ASC were split and seeded on chitosan-coated 24 well plate at a density of 7.5×10⁴ cells/cm², and allowed to adhere at 37°C. Within 3 days of culture, ASC formed spheroids by PBM irradiation. Conditioned medium (CM) fractions were collected from the PBM-ASC to yield nor adipose-derived stromal cell spheroid (spheroid) and PBM-spheroid, respectively, centrifuged at 13,000 g at 4℃ for 10 min, and stored prior to use for ELISA, protein assay, or in vivo wound-healing assays. Phosphate-buffered saline, cultured CM from ASCs, PBM irradiation prior to implanting conditioned medium from ASC, cultured CM from ASC spheroid, and PBM-spheroid-CM (PSC) were transplanted into a wound bed in athymic mice to evaluate therapeutic effects of PSC in vivo. PSC enhanced wound closure in a skin injury model compared to PBS, CM, PBM-CM, and spheroid-CM. The density of vascular formations increased as a result of angiogenic factors released by the wound bed and enhanced tissue regeneration at the lesion site.

Conclusions

These results indicate that implant of PSC can significantly improve functional recovery compared to PBS, CM, PBM-CM, or spheroid-CM treatment. Implant of PSC may be an effective form of paracrine mediated therapy for treating a wound bed.

SUBMITTER: Park IS

PROVIDER: S-EPMC8138655 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:PurposeThis study aimed to investigate the effects of photobiomodulation (PBM) and conditioned medium (CM) derived from human adipose-derived stem cells (h-ASCs), both individually and in combination, on the maturation stage of an ischemic infected delayed healing wound model (IIDHWM) in type I diabetic (TIDM) rats.MethodsThe study involved the extraction of h-ASCs from donated fat, assessment of their immunophenotypic markers, cell culture, and extraction and concentration of CM from cultured 1 × 10^6 h-ASCs. TIDM was induced in 24 male adult rats, divided into four groups: control, CM group, PBM group (80 Hz, 0.2 J/cm2, 890 nm), and rats receiving both CM and PBM. Clinical and laboratory evaluations were conducted on days 4, 8, and 16, and euthanasia was performed using CO2 on day 16. Tensiometrical and stereological examinations were carried out using two wound samples from each rat.ResultsAcross all evaluated factors, including wound closure ratio, microbiological, tensiometrical, and stereological parameters, similar patterns were observed. The outcomes of CM + PBM, PBM, and CM treatments were significantly superior in all evaluated parameters compared to the control group (p = 0.000 for all). Both PBM and CM + PBM treatments showed better tensiometrical and stereological results than CM alone (almost all, p = 0.000), and CM + PBM outperformed PBM alone in almost all aspects (p = 0.000). Microbiologically, both CM + PBM and PBM exhibited fewer colony-forming units (CFU) than CM alone (both, p = 0.000).ConclusionPBM, CM, and CM + PBM interventions substantially enhanced the maturation stage of the wound healing process in IIDHWM of TIDM rats by mitigating the inflammatory response and reducing CFU count. Moreover, these treatments promoted new tissue formation in the wound bed and improved wound strength. Notably, the combined effects of CM + PBM surpassed the individual effects of CM and PBM.Supplementary informationThe online version contains supplementary material available at 10.1007/s40200-023-01285-3.

Project description:Adult stem cells have been extensively investigated for tissue repair therapies. Adipose-derived stem cells (ASCs) were shown to improve wound healing by promoting re-epithelialization and vascularization as well as modulating the inflammatory immune response. In this study, we used ex vivo human skin cultured in a six-well plate with trans-well inserts as a model for superficial wounds. Standardized wounds were created and treated with allogeneic ASCs, ASCs conditioned medium (ASC-CM), or cell culture medium (DMEM) supplemented with fetal calf serum (FCS). Skin viability (XTT test), histology (hematoxylin and eosin, H and E), β-catenin expression as well as inflammatory mediators and growth factors were monitored over 12 days of skin culture. We observed only a moderate time-dependent decrease in skin metabolic activity while skin morphology was preserved, and re-epithelialization occurred at the wound edges. An increase in β-catenin expression was observed in the newly formed epithelia, especially in the samples treated with ASC-CM. In general, increased growth factors and inflammatory mediators, e.g., hepatocytes growth factor (HGF), platelet-derived growth factor subunit AA (PDGF-AA), IL-1α, IL-7, TNF-α, and IL-10, were observed over the incubation time. Interestingly, different expression profiles were observed for the different treatments. Samples treated with ASC-CM significantly increased the levels of inflammatory cytokines and PDGF-AA with respect to control, whereas the treatment with ASCs in DMEM with 10% FCS resulted in significantly increased levels of fibroblast growth factor-basic (FGF-basic) and moderate increases of immunomodulatory cytokines. These results confirm that the wound microenvironment can influence the type of mediators secreted by ASCs and the mode as to how they improve the wound healing process. Comparative investigations with pre-activated ASCs will elucidate further aspects of the wound healing mechanism and improve the protocols of ACS application.