In our experience, keratinocytes cultured in feeder-free conditions and in commercially available defined and serum-free media cannot be as efficiently massively expanded as their counterparts grown in conventional bovine serum-containing medium, nor can they properly form a stratified epidermis in a skin substitute model. We thus tested a new chemically defined serum-free medium, which we developed for massive human primary keratinocyte expansion and skin substitute production. Our medium, named Surge Serum-Free Medium (Surge SFM), was developed to be used alongside a feeder layer. It supports the growth of keratinocytes freshly isolated from a skin biopsy and cryopreserved primary keratinocytes in cultured monolayers over multiple passages. We also show that keratin-19-positive epithelial stem cells are retained through serial passaging in Surge SFM cultures. Transcriptomic analyses suggest that gene expression is similar between keratinocytes cultured with either Surge SFM or the conventional serum-containing medium. Additionally, Surge SFM can be used to produce bilayered self-assembled skin substitutes histologically similar to those produced using serum-containing medium. Furthermore, these substitutes were grafted onto athymic mice and persisted for up to six months. In conclusion, our new chemically defined serum-free keratinocyte culture medium shows great promise for basic research and clinical applications.

The increasing need for tissue substitutes in reconstructive surgery spurs the development of engineering methods suited for clinical applications. Cell culture and tissue production traditionally require the use of fetal bovine serum (FBS) which is associated with various complications especially from a translational perspective. Using the self-assembly approach of tissue engineering, we hypothesized that all important parameters of tissue reconstruction can be maintained in a production system devoid of FBS from cell extraction to tissue reconstruction. We studied two commercially available serum-free medium (SFM) and xenogen-free serum-free medium (XSFM) for their impact on tissue reconstruction using human adipose-derived stem/stromal cells (ASCs) in comparison to serum-containing medium. Both media allowed higher ASC proliferation rates in primary cultures over five passages compared with 10% FBS supplemented medium while maintaining high expression of mesenchymal cell markers. For both media, we evaluated extracellular matrix production and deposition necessary to engineer manipulatable tissues using the self-assembly approach. Tissues produced in SFM exhibited a significantly increased thickness (up to 6.8-fold) compared with XSFM and FBS-containing medium. A detailed characterization of tissues produced under SFM conditions showed a substantial 50% reduction of production time without compromising key tissue features such as thickness, mechanical resistance and pro-angiogenic secretory capacities (plasminogen activator inhibitor 1, hepatocyte growth factor, vascular endothelial growth factor, angiopoietin-1) when compared to tissues produced in the control FBS-containing medium. Furthermore, we compared ASCs to the frequently used human dermal fibroblasts (DFs) in the SFM culture system. ASC-derived tissues displayed a 2.4-fold increased thickness compared to their DFs counterparts. In summary, we developed all-natural human substitutes using a production system compatible with clinical requirements. Under culture conditions devoid of bovine serum, the resulting engineered tissues displayed similar and even superior structural and functional properties over the classic FBS-containing culture conditions with a considerable 50% shortening of production time.

Radiotherapy for cancer treatment is often associated with skin damage that can lead to incapacitating hard-to-heal wounds. No permanent curative treatment has been identified for radiodermatitis. This study provides a detailed characterization of the dose-dependent impact of ionizing radiation on skin cells (45, 60, or 80 grays). We evaluated both early and late effects on murine dorsal skin with a focus on the healing process after two types of surgical challenge. The irradiated skin showed moderate to severe damage increasing with the dose. Four weeks after irradiation, the epidermis featured increased proliferation status while the dermis was hypovascular with abundant α-SMA intracellular expression. Excisional wounds created on these tissues exhibited delayed global wound closure. To assess potential long-lasting side effects of irradiation, radiodermatitis features were followed until macroscopic healing was notable (over 8 to 22 weeks depending on the dose), at which time incisional wounds were made. Severity scores and biomechanical analyses of the scar tissues revealed that seemingly healed irradiated skin still displayed altered functionality. Our detailed investigation of both the acute and chronic repercussions of radiotherapy on skin healing provides a relevant new in vivo model that will instruct future studies evaluating the efficacy of new treatments for radiodermatitis.

Tissue-engineered skin substitutes (TESs) are used as a treatment for severe burn injuries. Their production requires culturing both keratinocytes and fibroblasts. The methods to grow these cells have evolved over the years, but bovine serum is still commonly used in the culture medium. Because of the drawbacks associated with the use of serum, it would be advantageous to use serum-free media for the production of TESs. In a previous study, we developed a serum-free medium (Surge SFM) for the culture of keratinocytes. Herein, we tested the use of this medium, together with a commercially available serum-free medium for fibroblasts (Prime XV), to produce serum-free TESs. Our results show that serum-free TESs are macroscopically and histologically similar to skin substitutes produced with conventional serum-containing media. TESs produced with either culture media expressed keratin 14, Ki-67, transglutaminase 1, filaggrin, type I and IV collagen, and fibronectin comparably. Mechanical properties, such as contraction and tensile strength, were comparable between TESs cultured with and without serum. Serum-free TESs were also successfully grafted onto athymic mice for a six-month period. In conclusion, Surge SFM and Prime XV serum-free media could be used to produce high quality clinical-grade skin substitutes.

Long‐term diabetes often leads to chronic wounds refractory to treatment. Cell‐based therapies are actively investigated to enhance cutaneous healing. Various cell types are available to produce biological dressings, such as adipose‐derived stem/stromal cells (ASCs), an attractive cell source considering their abundancy, accessibility, and therapeutic secretome. In this study, we produced human ASC‐based dressings under a serum‐free culture system using the self‐assembly approach of tissue engineering. The dressings were applied every 4 days to full‐thickness 8‐mm splinted skin wounds created on the back of polygenic diabetic NONcNZO10/LtJ mice and streptozotocin‐induced diabetic K14‐H2B‐GFP mice. Global wound closure kinetics evaluated macroscopically showed accelerated wound closure in both murine models, especially for NONcNZO10/LtJ; the treated group reaching 98.7% ± 2.3% global closure compared to 76.4% ± 11.8% for the untreated group on day 20 (p= 0.0002). Histological analyses revealed that treated wounds exhibited healed skin of better quality with a well‐differentiated epidermis and a more organized, homogeneous, and 1.6‐fold thicker granulation tissue. Neovascularization, assessed by CD31 labeling, was 2.5‐fold higher for the NONcNZO10/LtJ treated wounds. We thus describe the beneficial impact on wound healing of biologically active ASC‐based dressings produced under an entirely serum‐free production system facilitating clinical translation.

Background: Immunoglobulin A vasculitis (IgAV) is a systemic small-vessel vasculitis characterized by palpable purpura, often accompanied by joint, gastrointestinal, and/or renal involvement. Although cutaneous manifestations are usually mild and self-limited, they can occasionally present as persistent, severe, or recurrent skin lesions. Objectives: To assess the efficacy and safety of colchicine in the treatment of complicated cutaneous manifestations of IgAV. Methods: We conducted a retrospective chart review of patients treated with colchicine for skin manifestation of IgAV at our vasculitis clinic. This was complemented by a search of the PubMed and MEDLINE databases to identify additional reported cases. Results: A total of 30 cases were included, comprising 13 from our institution and 17 from the literature. Indications for colchicine use were persistent purpura lasting more than 4 weeks (n = 18), severe skin involvement including bullae, ulcerations, or widespread purpura (n = 9), and recurrent lesions after a symptom-free interval of at least 1 month (n = 8). Colchicine led to clinical improvement in 90% of patients (27/30), with complete resolution in 67% (20/30). All 12 patients treated with colchicine monotherapy responded favorably. Doses ranged from 0.6 to 1.8 mg/day, and the time to response varied from a few days to 3 months. No adverse effects were reported. Conclusions: Colchicine appears to be a safe and effective treatment for persistent, severe, or recurrent cutaneous manifestations of IgAV. These findings support its use in clinical practice and highlight the need for prospective controlled studies.