Conjunctival Reconstruction with Progenitor Cell-Derived Autologous Epidermal Sheets in Rhesus Monkey

Severe ocular surface diseases are some of the most challenging problems that the clinician faces today. Conventional management is generally unsatisfactory, and the long-term ocular consequences of these conditions are devastating. It is significantly important to find a substitute for conjunctival epithelial cells. This study was to explore the possibility of progenitor cell-derived epidermal sheets on denuded amniotic membrane to reconstruct ocular surface of conjunctiva damaged monkeys. We isolated epidermal progenitor cells of rhesus monkeys by type IV collagen adhesion, and then expanded progenitor cell-derived epidermal sheets on denuded amniotic membrane ex vivo. At 3 weeks after the conjunctiva injury, the damaged ocular surface of four monkeys was surgically reconstructed by transplanting the autologous cultivated epidermal progenitor cells. At 2 weeks after surgery, transplants were removed and examined with Hematoxylin-eosin staining, Periodic acid Schiff staining, immunofluorescent staining, scanning and transmission electron microscopy. Histological examination of transplanted sheets revealed that the cell sheets were healthy alive, adhered well to the denuded amniotic membrane, and had several layers of epithelial cells. Electron microscopy showed that the epithelial cells were very similar in appearance to those of normal conjunctival epithelium, even without goblet cell detected. Epithelial cells of transplants had numerous desmosomal junctions and were attached to the amniotic membrane with hemidesmosomes. Immunohistochemistry confirmed the presence of the conjunctival specific markers, mucin 4 and keratin 4, in the transplanted epidermal progenitor cells. In conclusion, our present study successfully reconstructed conjunctiva with autologous transplantation of progenitor cell-derived epidermal sheets on denuded AM in conjunctival damaged monkeys, which is the first step toward assessing the use of autologous transplantation of progenitor cells of nonocular surface origin. Epidermal progenitor cells could be provided as a new substitute for conjunctival epithelial cells to overcome the problems of autologous conjunctiva shortage

Kallikrein-related peptidase 6 regulates epithelial-to-mesenchymal transition and serves as prognostic biomarker for head and neck squamous cell carcinoma patients

Background: Dysregulated expression of Kallikrein-related peptidase 6 (KLK6) is a common feature for many human malignancies and numerous studies evaluated KLK6 as a promising biomarker for early diagnosis or unfavorable prognosis. However, the expression of KLK6 in carcinomas derived from mucosal epithelia, including head and neck squamous cell carcinoma (HNSCC), and its mode of action has not been addressed so far. Methods: Stable clones of human mucosal tumor cell lines were generated with shRNA-mediated silencing or ectopic overexpression to characterize the impact of KLK6 on tumor relevant processes in vitro. Tissue microarrays with primary HNSCC samples from a retrospective patient cohort (n = 162) were stained by immunohistochemistry and the correlation between KLK6 staining and survival was addressed by univariate Kaplan-Meier and multivariate Cox proportional hazard model analysis. Results: KLK6 expression was detected in head and neck tumor cell lines (FaDu, Cal27 and SCC25), but not in HeLa cervix carcinoma cells. Silencing in FaDu cells and ectopic expression in HeLa cells unraveled an inhibitory function of KLK6 on tumor cell proliferation and mobility. FaDu clones with silenced KLK6 expression displayed molecular features resembling epithelial-to-mesenchymal transition, nuclear β-catenin accumulation and higher resistance against irradiation. Low KLK6 protein expression in primary tumors from oropharyngeal and laryngeal SCC patients was significantly correlated with poor progression-free (p = 0.001) and overall survival (p < 0.0005), and served as an independent risk factor for unfavorable clinical outcome. Conclusions: In summary, detection of low KLK6 expression in primary tumors represents a promising tool to stratify HNSCC patients with high risk for treatment failure. These patients might benefit from restoration of KLK6 expression or pharmacological targeting of signaling pathways implicated in EMT