RESUMEN
Epidermal cell models are critical for studying skin biology. The gold standard used by the scientific community has historically been primary cell cultures from discarded tissue, typically from neonates (foreskin). Although directly applicable to humans, this system suffers from multiple issues, including substantial donor-to-donor variability and a finite number of divisions in culture. As such, we have identified a faithful alternative called N/TERT2G cells. These cells show many of the characteristics of primary cells, including barrier formation, differentiation kinetics and/or protein expression, and pathogenesis. From our observations, N/TERT2G cells can serve as a reproducible and genetically manipulatable platform in studying skin biology.
RESUMEN
Microglia, the resident immune cells of the central nervous system (CNS), play multiple roles in maintaining CNS homeostasis and mediating tissue repair, including proliferating in response to brain injury and disease. Cranial irradiation (CI), used for the treatment of brain tumors, has a long-lasting anti-proliferative effect on a number of cell types in the brain, including oligodendrocyte progenitor and neural progenitor cells; however, the effect of CI on CNS-resident microglial proliferation is not well characterized. Using a sterile cortical needle stab injury model in mice, we found that the ability of CNS-resident microglia to proliferate in response to injury was impaired by prior CI, in a dose-dependent manner, and was nearly abolished by a 20 âGy dose. Similarly, in a metastatic tumor model, prior CI (20 âGy) reduced microglial proliferation in response to tumor growth. The effect of irradiation was long-lasting; 20 âGy CI 6 months prior to stab injury significantly impaired microglial proliferation. We also investigated how stab and/or irradiation impacted levels of P2Y12R, CD68, CSF1, IL-34 and CSF1R, factors involved in the brain's normal response to injury. P2Y12R, CD68, CSF1, and IL-34 expression were altered by stab similarly in irradiated mice and controls; however, CSF1R was differentially affected. qRT-PCR and flow cytometry analyses demonstrated that CI reduced overall Csf1r mRNA levels and microglial specific CSF1R protein expression, respectively. Interestingly, Csf1r mRNA levels increased after injury in unirradiated controls; however, Csf1r levels were persistently decreased in irradiated mice, and did not increase in response to stab. Together, our data demonstrate that CI leads to a significant and lasting impairment of microglial proliferation, possibly through a CSF1R-mediated mechanism.