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La fuerza mecánica es importante para el modelado, el remodelado y la regeneración ósea; estimula a los osteocitos provocando una alteración en la producción y secreción de moléculas de señalización que regulan la actividad de los osteoblastos y los osteoclastos. El objetivo del presente estudio fue evaluar el efecto del medio condicionado de células osteocíticas de ratón estimuladas mecánicamente sobre la capacidad proliferativa y migratoria de células mesenquimales y células óseas. Para ello, se analizó la proliferación y migración de las células preosteoblásticas de ratón, células mesenquimales preadiposas humanas y macrófagos de ratón en presencia del medio condicionado de las células osteocíticas, tras 6 y 24 horas después de ser sometidas a un estrés mecánico de 10 dinas/cm2 por flujo de fluido (FF) durante 10 minutos. Se encontró que la migración de células preosteoblásticas aumentó significativamente en presencia de medios condicionados de células osteocíticas con respecto al grupo control estático (SC) (SC=12,63±5,44; FF6h=23,03±11,57; FF24h=29,72±15,76; p

Approximately 10 million individuals have blindness due to limbal stem cell (LSCs) deficiency, one of the most challenging problems in ophthalmology. To replenish the LSC pool, an autologous extraocular cell source is appropriate, thereby avoiding the risk of immune rejection, the need for immunosuppression and the risk of damaging the contralateral eye. In recent years, adipose-derived mesenchymal stem cells (ADSCs) have been a key element in ocular regenerative medicine. In this study, we developed a protocol for deriving human LSCs from ADSCs compatible with the standard carrier human amniotic membrane, helping provide a stem cell pool capable of maintaining proper corneal epithelial homeostasis. The best protocol included an ectodermal induction step by culturing ADSCs with media containing fetal bovine serum, transforming growth factor-β inhibitor SB-505124, Wnt inhibitor IWP-2 and FGF2 for 7 days, followed by an LSC induction step of culture in modified supplemental hormonal epithelial medium supplemented with pigment epithelium-derived factor and keratinocyte growth factor for 10 additional days. The optimal differentiation efficiency was achieved when cells were cultured in this manner over vitronectin coating, resulting in up to 50% double-positive αp63/BMI-1 cells. The results of this project will benefit patients with LSC deficiency, aiding the restoration of vision.

Historically, primordial germ cells (PGCs) have been a good model to study pluripotency. Despite their low numbers and limited accessibility in the mouse embryo, they can be easily and rapidly reprogrammed at high efficiency with external physicochemical factors and do not require transcription factor transfection. Employing this model to deepen our understanding of cell reprogramming, we specifically aimed to determine the relevance of Ca 2+ signal transduction pathway components in the reprogramming process. Our results showed that PGC reprogramming requires a normal extracellular [Ca 2+ ] range, in contrast to neoplastic or transformed cells, which can continue to proliferate in Ca 2+ -deficient media, differentiating normal reprogramming from neoplastic transformation. Our results also showed that a spike in extracellular [Ca 2+ ] of 1-3 mM can directly reprogram PGC. Intracellular manipulation of Ca 2+ signal transduction pathway components revealed that inhibition of classical Ca 2+ and diacylglycerol (DAG)-dependent PKCs, or intriguingly, of only the novel DAG-dependent PKC, PKCε, were able to induce reprogramming. PKCε inhibition changed the metabolism of PGCs toward glycolysis, increasing the proportion of inactive mitochondria. This metabolic switch from oxidative phosphorylation to glycolysis is mediated by hypoxia-inducible factors (HIFs), given we found upregulation of both HIF1α and HIF2α in the first 48 hours of culturing. PKCε inhibition did not change the classical pluripotency gene expression of PGCs, Oct4, or Nanog. PKCε inhibition changed the histone acetylation of PGCs, with histones H2B, H3, and H4 becoming acetylated in PKCε-inhibited cultures (markers were H2BacK20, H3acK9, and H4acK5K8, K12, K16), suggesting that reprogramming by PKCε inhibition is mediated by histone acetylation., Competing Interests: None., (AJSC Copyright © 2021.)

Objective: We aimed to investigate the functionality of human decellularized stromal laminas seeded with cultured human corneal endothelial cells as a tissue engineered endothelial graft (TEEK) construct to perform endothelial keratoplasty in an animal model of corneal endothelial damage., Methods: Engineered corneal endothelial grafts were constructed by seeding cultured human corneal endothelial cell (hCEC) suspensions onto decellularized human corneal stromal laminas with various coatings. The functionality and survival of these grafts with cultured hCECs was examined in a rabbit model of corneal endothelial damage after central descemetorhexis. Rabbits received laminas with and without hCECs (TEEK and control group, respectively)., Results: hCEC seeding over fibronectin-coated laminas provided an optimal and consistent endothelial cell count density and polygonal shape on the decellularized laminas, showing active pump fuction. Surgery was performed uneventfully as standard Descemet stripping automated endothelial keratoplasty (DSAEK). Corneal transparency gradually recovered in the TEEK group, whereas haze and edema persisted for up to 4 weeks in the controls. Histologic examination showed endothelial cells of human origin covering the posterior surface of the graft in the TEEK group., Conclusions: Grafting of decellularized stroma carriers re-surfaced with human corneal endothelial cells ex vivo can be a readily translatable method to improve visual quality in corneal endothelial diseases., Competing Interests: The authors haave declared that no competing interests exist.