Comparative study of isolation, expansion and characterization of epithelial cells.

BACKGROUND AIMS: The human epithelial cells (EPCs) have been identified as the essential element for the regeneration of skin construct for burns, wounds and various tissue engineer-based products. METHODS: In this study, the isolation, expansion and characterization of EPCs from various sources such as juvenile foreskin (JSK), buccal mucosa (BM), penile skin (PS) and urothelium (UR) in serum-free and xeno-free EpiLife media were evaluated. RESULTS: The growth kinetics study revealed that EPCs from JSK and BM had notably higher growth rates compared with the others. Overall, the EPCs from all sources retained basic morphological characteristics and the functional characteristics such as Pan Cytokeratin (AE1/AE3). In addition, the cryopreservation stability of EPCs was accessed for post-thaw viability and found to be greater than 80% at 1 year of storage, but demonstrated reduced cell recovery (51%) at the second year in fetal bovine serum-free cryopreservation media. CONCLUSIONS: Our result suggests that the EPCs from four cell sources can be grown in feeder-free, serum-free and xeno-free systems using commercially available EpiLife medium without losing epithelial cell characteristics even after passage 4. However, its suitability for clinical application must be accessed by preclinical and clinical studies.

Inhibition of proliferation and migration of stricture fibroblasts by epithelial cell-conditioned media.

INTRODUCTION: Urethral stricture is characterized by urethral lumen narrowing due to fibrosis. Urethroplasty of the urethral stricture involves excision of scar, and may be followed by reconstruction of the urethra using split-thickness skin, buccal mucosa, urethral mucosa or, more recently, tissue-engineered grafts. The stricture wound healing process after urethroplasty is known to be mediated by an interaction between keratinocyte and fibroblasts; however, the underlying mechanisms are not studied in detail yet. We investigated the influence of epithelial cell-conditioned medium (ECCM) (obtained from confluent penile skin, buccal mucosa and urethral cell cultures) on the proliferation and migration of stricture fibroblasts using an in vitro scratch assay. MATERIALS AND METHODS: ECCM was collected from confluent primary epithelial cell cultures of three different human biopsies (penile skin, buccal mucosa and urethral mucosa), whereas stricture fibroblasts were isolated from human urethral stricture biopsies. The effect of ECCM on stricture fibroblasts' proliferation and migration into the scratch was observed using a standard in vitro scratch assay over a period of 3 days. Four experiments were performed independently using four stricture fibroblasts from four patients and ECCM was collected from 12 different patients' primary cell cultures. RESULTS: ECCM from primary epithelial cells cultures obtained from penile skin, buccal mucosa and urethra inhibited stricture fibroblasts' proliferation and migration in the in vitro scratch assay. CONCLUSION: These results demonstrate the ability of ECCM to inhibit the proliferation and migration of stricture fibroblasts and present it as an effective adjunct in urethroplasty, which may influence stricture wound healing and inhibit the recurrence of stricture.

Mesenchymal stem cells are prospective novel off-the-shelf wound management tools.

Chronic/non-healing cutaneous wounds pose a debilitating burden on patients and healthcare system. Presently, treatment modalities are rapidly shifting pace from conventional methods to advanced wound care involving cell-based therapies. Mesenchymal stem cells (MSCs) have come across as a prospective option due to its pleiotropic functions viz. non-immunogenicity, multipotency, multi-lineage plasticity and secretion of growth factors, cytokines, microRNAs (miRNA), exosomes, and microvesicles as part of their secretome for assisting wound healing. We outline the therapeutic role played by MSCs and its secretome in suppressing tissue inflammation, causing immunomodulation, aiding angiogenesis and assisting in scar-free wound healing. We further assess the mechanism of action by which MSCs contribute in manifesting tissue repair. The review flows ahead in exploring factors that influence healing behavior including effect of multiple donor sites, donor age and health status, tissue microenvironment, and in vitro expansion capability. Moving ahead, we overview the advancements achieved in extending the lifespan of cells upon implantation, influence of genetic modifications aimed at altering MSC cargo, and evaluating bioengineered matrix-assisted delivery methods toward faster healing in preclinical and clinical models. We also contribute toward highlighting the challenges faced in commercializing cell-based therapies as standard of care treatment regimens. Finally, we strongly advocate and highlight its application as a futuristic technology for revolutionizing tissue regeneration.