eactive vimentin was decreased in treated (manage and 
SAA+ cells) kidneys with most tubule lining cells good for the epithelial marker pan-keratin (not shown). GFP+ donor cells did not stain with Antibiotic-202 biological activity anti-vimentin. Kidney sections have been counterstained with DAPI to stain nuclei blue. The graphs show pixel density of vimentin (x10 to facilitate visualization on one axis) and pankeratin. #p0.05 vs no cells, p0.05 vs manage cells
Exosome Uptake by PCK cells. Isolation of exosomes (exo) was confirmed by electron microscopy demonstrating vesicles of proper size following damaging stain (EM, A) and enrichment of CD63 on immunoblot (B). Exosomes from regular Sprague Dawley rats express fibrocystin (FPC, B). Internal exosome protein was fluorescently labeled green and RNA red prior to incubation of PCK cells together with the exosomes for 16 hours. In panel C are representative fluorescence microscopy photos demonstrating uptake on the exosomes by the major PCK renal cells. The “no exo” handle was incubated with exo-glow dye but no exosomes. PCR genotyping shows expression of each wild kind and mutant Pkhd1 mRNA inside the PCK cells exposed to exosomes (D). Wild type Pkhd1 is discovered in SD exosomes and SD cells (SD) and only mutant Pkhd1 in control PCK cells. Right after the addition in the exosome uptake inhibitor chlorpromazine along with the protein synthesis inhibitor cytochalasin D prior to exosomes, no wild type Pkhd1 was transferred towards the mutant PCK cells. Graphs demonstrate quantification of band densities representing CD63 (corrected for actin) or mutant and wild sort Pkhd1. MW, molecular weight markers, + good control (panel B)
The effect of exosomes on PCK cysts in culture. Representative confocal photos of cells stained with rhodamine phalloidin to label actin red and Hoescht to label nuclei blue demonstrate cysts in tubule cells from PCK rats grown in matrigel without exosome therapy (A, F, G). Neither PCK cells cultured with exosomes (B, C, E) from Sprague Dawley (SD) rats nor SD tubular cells (D, H) type cysts in matrigel. The higher energy photos (A-D, scale bar 20um) show orthogonal projections to demonstrate the clear presence or absence of cysts. Panel F, displaying a 3D reconstruction of various photos, also demonstrates the cystic nature with the structures formed by PCK cells (scale bar 20um). The reduced energy images (E, G, H, scale bar 50um) demonstrate tubular structures within the presence of exosomes, a number of cysts in one particular field of PCK cells plus the absence of cysts in SD cells, respectively. In PCK cells treated with exosomes (E), “tube-like” structures are observed. Quantification of cyst quantity in a total of 168 fields is presented within the graph (I). B exosomes are from SAA+ cells, A are from SAA- manage cells. Both A and B exosomes include wild form Pkhd1 (above).
Wild kind Pkhd1 in kidneys from PCK rats treated with SD exosomes. PCR genotype demonstrates mutant Pkhd1 in kidneys from handle and treated PCK rats. Wild form Pkhd1 was found in kidneys from PCK rats treated with SD exosomes (24h following renal pelvic injection) but not in untreated rats. -, damaging PCR handle; exo, exosomes; MW, molecular weight markers.
Neural crest cells (NCCs) are multipotent precursor cells which might be specified at the boundary involving neural plate and epidermis upon induction by growth aspects which include Wnts, BMPs, FGFs and retinoic acid (RA) [1]. Committed NCCs undergo an epithelial to mesenchymal transition (EMT) just before delaminating in the neural plate and migra