Bronchioalveolar stem cells (BASCs) have been shown to be a regional stem cell population in the distal lung. They are CCSP (Clara cell secretory protein) and SPC (surfactant protein C) positive. They respond to bronchiolar and alveolar injuries. BASCs are thought to maintain the bronchiolar Clara cells and the alveolar type 2 cells. Previous studies showed that Clara cells contribute to the renewal of the bronchiolar epithelium after oxidant mediated damages. Importantly, after the depletion of Clara cells through administration of naphthalene, residual cells of terminal bronchioles were fully capable of epithelial renewal and restoration. These naphthalene-resistant Clara cells were present at the bronchioalveolar duct junction (BADJ). BASCs were isolated based on positive staining for CD34, a marker for skin epithelial cells and hematopoietic stem cells, and for Sca-1, a marker for stem cells. Others studies performed in adult mouse lung indicate that Sca-1pos cell fractions are predominantly representative of mesenchymal cell lineages. This highlights that Sca-1 is not a selectable marker for epithelial stem cells in the adult murine lung, and therefore the need of alternative functional assays for the identification and characterization of BASCs. In the present study, we characterized BACS by alternative methods. We identified by immunofluorescence the proposed BASC cell population co-expressing SPC, a marker for alveolar type 2 cells, and CCSP, a marker for bronchial epithelial Clara cells, that resides at the BADJ. BASCs were then identified by flow cytometry using double-fluorescent Cre reporter mice that express YFP and mCherry under the control of SPC and CCSP promoters; also based on an adjusted McQualter protocol for their expression of EpCAMhigh and CD24low. Clonogenic assays of EpCAMhigh and CD24low sorted cells cultured on feeders and matrigel showed BASCs as small colony-forming-units. The proliferative activity of BASCs after naphthalene injury and pneumonectomy was also determined. Based on their expression of EpCAMhigh-CD24low, we showed the evidence of BASCs as a potential progenitor/stem cells for distal murine lung. We further aim to establish in vivo differentiation assays for BASCs to address changes in stemness and proliferative activity during experimental lung disease and lung regeneration.Oct4, specifically expressed in embryonic stem cells, can also be detected in adult stem cells such as bone marrow-derived mesenchymal stem cells. A role for maintaining pluripotency and self-renewal of embryonic stem cells is ascribed to Oct4 as a pluripotency marker. Several studies suggest a role for Oct-4 in sustaining self-renewal capacity of adult somatic stem cells. Other scientists have produced the evidence that Oct-4 gene ablation in the somatic stem cells revealed no abnormalities in homeostasis or regenerative capacity. Data strongly argue that Oct-4, even if expressed at low levels in somatic cells, is dispensable for the self-renewal of somatic stem cells, for tissue homeostasis, and for the regeneration of tissue in the adult. The aim of this project was to trace Oct4 positive cells in adult mouse lung. We identified a distinct pulmonary Oct4 expressing cell compartment that belongs based on its localization to telocytes. The hypothesis is supported from the literature reporting that telocytes are located in the perivascular wall and extended by their telopodes to the peribronchial and alveolar spaces. They express Oct-4, vimentin, Sca-1, PDGFR-beta, C-kit and VEGF. We confirmed these results by immunofluorescence confoncal microscopy of adult wild type mouse lung. Flow cytometric analyses using Oct4-GFP reporter mice identified a population of Oct4-GFP positive cells. Oct4-GFP positive cells were sorted cultured and were growing with long extensions like telocytes. Laser capture microdissection and qRT-PCR were also established to support immunofluorescence data on mRNA level. Since our data confirmed the hypothesis in which Oct4 positive cells are telocytes, our next goal is first to study the role of telocytes in adult mouse lung and determine the function of Oct4 in these cells.
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