ABSTRACT
Recent spatiotemporal report demonstrated that epidermal stem cells have equal potential to divide or differentiate, with no asymmetric cell division observed. Therefore, how epithelial stem cells maintain lifelong stem-cell support still needs to be elucidated. In mouse blood and bone marrow, we found a group of large cells stained strongly for eosin and containing coiled-tubing-like structures. Many were tightly attached to each other to form large cellular clumps. After sectioning, these large cell-clumps were composed of not cells but numerous small particles, however with few small "naked" nuclei. The small particles were about 2 to 3 µm in diameter and stained dense red for eosin, so they may be rich in proteins. Besides the clumps composed of small particles, we identified clumps formed by fusion of the small particles and clumps of newly formed nucleated cells. These observations suggest that these small particles further fused and underwent cellularization. E-cadherin was expressed in particle-fusion areas, some "naked" nuclei and the newly formed nucleated cells, which suggests that these particles can form epithelial cells via fusion and nuclear remodeling. In addition, we observed similar-particle fusion before epithelial cellularization in mouse kidney ducts after kidney ischemia, which suggests that these particles can be released in the blood and carried to the target tissues for epithelial-cell regeneration. Oct4 and E-cadherin expressed in the cytoplasmic areas in cells that were rich in protein and mainly located in the center of the cellular clumps, suggesting that these newly formed cells have become tissue-specific epithelial stem cells. Our data provide evidence that these large particle-producing cells are the origin of epithelial stem cells. The epithelial stem cells are newly formed by particle fusion.
Subject(s)
Stem Cells/cytology , Animals , Epithelial Cells/cytology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, TransgenicABSTRACT
We found a group of non-platelet RNA-containing particles (NPRCPs) in human umbilical cord blood. These particles can aggregate, fuse and become non-nucleated cells when cocultured with nucleated cells in vitro. The non-nucleated cells further differentiate into nucleated cells expressing octamer binding transcription factor 4 (OCT4). The NPRCPs are approximately 1-5 µm in diameter, have a thin bilayer membrane, contain short RNAs and microRNAs and express OCT4, sex-determining region Y 2 (SOX2) and DEAD box polypeptide 4 (DDX4). To confirm the function of NPRCPs in vivo, we examined the effects of tail vein-injected green fluorescent protein (GFP)-labelled NPRCPs on mouse kidneys damaged by prior ischaemia and reperfusion from Day 1 to Week 6. Within 1 day of injection of NPRCPs, immunofluorescence and immunohistochemistry revealed a large number of extravasated NPRCPs in the renal calyces, damaged glomeruli and duct tubules. During the course of regeneration, NPRCPs fused into large, non-nucleated cellular structures that further became large nucleated cells to regenerate multicellular kidney tubules. In addition, many NPRCPs became tiny nucleated cellular structures that further differentiated into interstitial cells in connective tissue. The extravasated NPRCPs also arranged themselves into non-cell glomerular structures before further regenerating into nucleated cells of the glomerulus. In conclusion, the results demonstrate that, via different patterns of differentiation, NPRCP-derived cells can regenerate mouse kidney tissue damaged by ischaemia.
Subject(s)
Adult Stem Cells/cytology , Cell Transdifferentiation , Cell-Derived Microparticles/transplantation , Kidney/physiology , RNA/metabolism , Regeneration , Reperfusion Injury/therapy , Adult Stem Cells/metabolism , Adult Stem Cells/ultrastructure , Animals , Cell Differentiation , Cell-Derived Microparticles/metabolism , Cell-Derived Microparticles/ultrastructure , Cells, Cultured , DEAD-box RNA Helicases/metabolism , Female , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Ischemia/physiopathology , Kidney/pathology , Mice , Mice, Inbred BALB C , Mice, Transgenic , MicroRNAs/metabolism , Octamer Transcription Factor-3/metabolism , Recombinant Proteins/metabolism , Reperfusion Injury/etiology , Reperfusion Injury/metabolism , Reperfusion Injury/pathology , SOXB1 Transcription Factors/metabolismABSTRACT
We found a group of non-platelet RNA-containing particles (NPRCP) in human umbilical cord blood. To understand the origin, characterization and differentiation of NPRCP, we examined cord blood-isolated NPRCP in vitro. The NPRCP range in size from < 1 to 5 µm, have a thin bilayer membrane and various morphological features, contain short RNA and microRNA and express octamer-binding transcription factor 4 (OCT4), sex-determining region Y 2 (SOX2) and DEAD box polypeptide 4 (DDX4). On coculture with nucleated cells from umbilical cord blood, NPRCP fuse to small, active, non-nucleated cells called 'particle fusion-derived non-nucleated cells' (PFDNC). The PFDNC are approximately 8 µm in diameter and are characterized by their twisting movement in culture plates. They can easily move into and out of nucleated cells and finally differentiate into mesenchymal-like cells. In addition, the larger non-nucleated cellular structures that are derived from the aggregation and fusion of multiple NPRCP can further differentiate into large stem cells that also release OCT4- and SOX2-positive non-nucleated small cells. Our data provide strong evidence that NPRCP can fuse into PFDNC, which further differentiate into mesenchymal-like cells. Multiple NPRCP also fuse into other types of large stem cells. We believe that stem cells are derived from NPRCP fusion. There is considerable potential for the use of NPRCP in clinical therapy.
