Introduction: Several factors can sensitize the skin, from external factors, such as extreme cold and heat, to the aesthetic procedures. This sensitization can cause damage, affecting the skin's protective barrier and unbalancing its microbiota. Simultaneous modulation of different biological processes is the key to ensure fast and efficient recovery. Hence, it is important to use multi-repair agents that help in the process of repairing sensitized skin, either by relieving symptoms or stimulating healing. Restoration and maintenance of the microbiota that was altered during such events is also essential for a good recovery. Therefore, a cosmetic formulation (CF) containing a combination of active ingredients that has a synergistic effect in the three stages of the healing process: panthenol, niacinamide, bisabolol, hyaluronic acid, tocopherol and a prebiotic molecule, α-glucan was developed to repair, soothe and moisturize sensitive, irritated and sensitized skin, balancing and protecting the skin microbiota. This research aimed to evaluate preclinical efficacy of the CF in corneodesmosin (CDSN) expression and the clinical efficacy in skin regeneration and microbiota maintenance and regeneration. Methods: To evaluate corneodesmosin expression analysis were performed with reconstructed human epidermis (RHE), subject to lactic acid 0,6%, treated with the CF or not and then collected to quantification using in situ immunofluorescent labeling and semi-quantitative image analysis. Cellular regeneration was evaluated by reflectance confocal microscopy. The facial images of the clinical study participants were obtained before the beginning of the study (before peeling procedure) and 14 days after the continuous use of the product and were evaluated the skin characteristics, among others, the thickness of the layers of the epidermis and organization of the keratinocytes. To assess the restoration and maintenance of the microbiota, metagenomics was studied. Samples were collected on the skin surface by sterile swab before the start of the study (before peeling procedure) and 14 days after the continuous use of the product in the selected participants (10 participants). Microbiota analysis was performed both quantitatively (abundance) and qualitatively (diversity). The microbiota data generated from each sample were compared against public databases to classify the species present. Results: Corneodesmosin was detected mainly at the surface of cells located within the granular layer. In response to the stress with lactic acid, the expression of CDSN strongly decreased (-72%). On the other hand, with 0.2% CF, a limited diminution of CDSN expression in response to lactic acid (-38% vs. -72% in untreated stressed RHEs) was observed.The cellular regeneration studies showed the recovery of the stratum corneum, restoring its initial thickness, an increase in the epidermal thickness (+12%), in the dermal papilla depth parameter (+20%), in the keratinocytes morphology parameter (+36%) and in the reflectance of the granular layer (+36%), as well as a significant reduction in the skin surface irregularity (-33%) 14 days after the peeling procedure associated with the use of CF.Maintenance of the proportions between the main phyla of the human skin resident microbiota, as well as the main species belonging to them, and an increase in alpha diversity in the microbiome was observed 14 days after the peeling procedure associated with the use of CF, with the increase of the species of the genus Streptococcus and Veillonella. Discussion: The data obtained by reflectance confocal microscopy analysis suggest that the use of CF promotes cell proliferation, in addition to an improvement in the pattern of the dermal-epidermal junction and a better organization of the keratinocytes, resulting in a better structuring of the basal layer and cellular renewal. In addition, the increase in the granular layer reflectance indicates deep hydration and greater nutrition of the region, while the significant reduction in the surface irregularity parameter represents a more cohesive and, therefore, more uniform stratum corneum, corroborating the increase in corneodesmosin expression after treatment with CF. The metagenomic study showed the maintenance and restoration of the skin microbiota after the use of CF, ensuring the return of the resident microbiota, in addition to bringing more diversity to the microbiota, showing also a postbiotic effect. Conclusion: The results obtained suggest that the cosmetic formulation containing panthenol, niacinamide, bisabolol, hyaluronic acid, tocopherol and a prebiotic molecule, α-glucan could restore the integrity of the skin barrier and the skin's microbiota of a injured skin.