Proliferation of Submesothelial Mesenchymal Cells during Early Phase of Serosal Thickening in the Rabbit Bladder Is Accompanied by Transient Keratin 18 Expression

doi:10.1006/excr.1996.0088

Experimental Cell Research

Volume 223, Issue 2, 15 March 1996, Pages 327-339

https://doi.org/10.1006/excr.1996.0088 Get rights and content

Abstract

Partial outlet obstruction of the rabbit bladder induces serosal thickening and smooth muscle (SM) hypertrophy. Within thickened serosa, submesothelial (mesenchymal) cells differentiate into SM cells after 30 days of obstruction [S. Buoroet al. Lab. Invest.69, 589–602, 1993]. Here, we show that submesothelial cells transiently express keratin (K) 18 but not K8 soon after obstruction. We investigated a possible relationship between keratin expression and cell proliferation/differentiationin vivoandin vitro.The results of this study indicate that expression of K18 is spatiotemporally related to the pattern of cell proliferation with respect to the localization of an elastic membrane which divides the thickened serosa into an “extrinsic” and an “intrinsic” region. Moreover, K18 is not present in bladder mesenchyma during early development, indicating that its expression in the adult is not attributable to a dedifferentiation process. However, simultaneous K18, K8, and desmoplakin (DP) expression can be induced in normal and thickened serosa upon treatment with bromo-deoxyuridine. Our results indicate that K18 is a marker of proliferating mesenchymal cells in rabbit serosa, whereas the combined expression of K18, K8, and DP might be related to the hypothesized alterations in the stability of gene expression. A model is proposed in which keratin-containing submesothelial cells can act as a “transit” cell phenotype involved in both regenerating mesothelial cells and formation of SM cells.

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Cited by (14)

Bladder Dysfunction and Altered Somatic Sensitivity in PACAP<sup>-/-</sup> Mice
2010, Journal of Urology
Citation Excerpt :
The increased bladder mass in PACAP−/− mice was consistent with cystometry and urine spot data demonstrating increased VV and fewer but significantly larger urine spots. PACAP−/− mice also showed increased lamina propria thickness, as previously noted in animal models of outlet obstruction.23 Reasons underlying the increased lamina propria thickness in PACAP−/− mice may include tissue remodeling of the afferent and efferent nerves24 in the suburothelial plexus and detrusor,25 myofibroblasts and/or vasculature,26 as described in animal models of outlet obstruction and potentially influenced by changes in bladder nerve growth factor content.24,27
PACAP and receptors are expressed in micturition pathways. Studies show that PACAP has a role in detrusor smooth muscle contraction to facilitate adenosine triphosphate release from urothelium and PACAP antagonism decreases cyclophosphamide induced bladder hyperreflexia.
PACAP contributions to micturition and somatic sensation were studied in PACAP knockout (PACAP^−/−), litter mate heterozygote (PACAP^+/−) and WT mice by conscious cystometry with continuous intravesical saline or acetic acid (0.5%) instillation, urination patterns, somatic sensitivity testing of hind paw and pelvic regions with calibrated von Frey filaments, and morphological bladder assessments.
PACAP^−/− mice had an increased bladder mass with fewer but larger urine spots. In PACAP^−/− mice the lamina propria and detrusor smooth muscle were significantly thicker but the urothelium was unchanged. PACAP^−/− mice had increased bladder capacity, voided volume and intercontraction interval with significantly increased detrusor contraction duration and large residual volume. WT mice responded to acetic acid (0.5%) with a decrease in voided volume and intercontraction interval but PACAP^+/− and PACAP^−/− mice did not respond. PACAP^−/− mice were less responsive to somatic stimulation. PACAP^+/− mice also had bladder dysfunction, and somatic and visceral sensory abnormalities but to a lesser degree.
PACAP gene disruption contributes to changes in bladder morphology and function, and somatic and visceral hypoalgesia.
Establishment and characterization of atypical fibroblasts from human adult liver contributing to hepatocyte cord-like arrangement
2008, Cell Biology International
Citation Excerpt :
It is well accepted that studying fibroblasts in culture on rigid substrates closely replicates the mechanical stress that stimulates in vivo the neo-synthesis of ASMA, which serves slow down cell motility and acquire smooth muscle cell-like contractility (Rönnov-Jessen and Peterson, 1996; Tomasek et al., 2002; Hinz and Gabbiani, 2003). On the other hand, CK expression in cells of mesenchymal origin is associated with cell proliferation and motility (Pampinella et al., 1996; Corcoran and Ferretti, 1997). Moreover, transfection with both human CK8 and CK18 in mouse L fibroblasts, which normally express only vimentin, increased migratory and invasive properties (Chu et al., 1993).
We report the phenotypic and functional characterization of fibroblasts established in culture from the non-parenchymal epithelial cell populations of adult human livers. Human liver fibroblasts (hLF) expressed mesenchymal antigens vimentin, alpha-smooth muscle actin, collagen, fibronectin, CD73, CD90, CD105, and CD166 together with non-mesenchymal antigens cytokeratins 8 and 18, glial fibrillary acidic protein, and nestin. Mixed cell lineage-specific protein expression was not associated with stem-like cell properties. Coculturing hepatocytes onto confluent hLF showed that they survived and maintained metabolic activity such as albumin, glycogen, and urea production. Moreover, hepatocytes formed cord-like arrangements resembling those established in vivo. Hepatocyte arrangement depended on cell-to-cell contact and the tissue origin of fibroblasts. Time-lapse video imaging of cocultured cells showed that hepatocyte arrangement was coordinated by the stretching and shortening of underneath hLF. Our data suggest that hLF may represent resident fibroblasts of the adult human liver, which could assume guiding functions for hepatic epithelial cells.
Mesothelial progenitor cells and their potential in tissue engineering
2004, International Journal of Biochemistry and Cell Biology
The mesothelium consists of a single layer of flattened mesothelial cells that lines serosal cavities and the majority of internal organs, playing important roles in maintaining normal serosal integrity and function. A mesothelial ‘stem’ cell has not been identified, but evidence from numerous studies suggests that a progenitor mesothelial cell exists. Although mesothelial cells are of a mesodermal origin, they express characteristics of both epithelial and mesenchymal phenotypes. In addition, following injury, new mesothelium regenerates via centripetal ingrowth of cells from the wound edge and from a free-floating population of cells present in the serosal fluid, the origin of which is currently unknown. Recent findings have shown that mesothelial cells can undergo an epithelial to mesenchymal transition, and transform into myofibroblasts and possibly smooth muscle cells, suggesting plasticity in nature. Further evidence for a mesothelial progenitor comes from tissue engineering applications where mesothelial cells seeded onto tubular constructs have been used to generate vascular replacements and grafts to bridge transected nerve fibres. These findings suggest that mesothelial cell progenitors are able to switch between different cell phenotypes depending on the local environment. However, only by performing detailed investigations involving selective cell isolation, clonal analysis together with cell labelling and tracking studies, will we begin to determine the true existence of a mesothelial stem cell.
Bromodeoxyuridine induces keratin protein synthesis at a posttranscriptional level in human lung tumour cell lines
1999, Differentiation
Keratin intermediate filaments are formed in epithelial cells in a cell- and tissue-specific manner, but much remains unknown regarding the mechanisms which control the synthesis of these proteins. We examined the effect of the differentiation modulation agent, bromodeoxyuridine (BrdU), on two human keratin-negative (by immunocytochemistry) lung cell lines, DLKP and H82, and showed immunohistochemically that treatment with 10 μM BrdU over 7 days induced K8 and K18 protein synthesis in both lines. Immunoprecipitation and Western blot analyses revealed low levels of K8 and K18 proteins in untreated cell homogenates. These levels increased following treatment with BrdU for 7 days. K8 and K18 mRNAs were detected by Northern blot and reverse transcriptase polymerase chain reaction analyses in both lines before BrdU treatment, but no increase in mRNA levels was observed in either cell line over 21 days of treatment. This suggests, firstly, that keratin synthesis is normally blocked at a posttranscriptional level in DLKP and H82 cells, and secondly, that BrdU can reverse this block. A549 is a human lung cell line which contains K8 and K18 proteins. Treatment with BrdU increased K8 and K18 protein levels in these cells. No corresponding increase in K8 mRNA levels occurred, while an apparent increase in K18 mRNA levels was detected. HL-60 is a leukaemic cell line of haematopoietic rather than epithelial lineage which contains K8 and K18 mRNA transcripts prior to BrdU treatment, but does not contain keratin proteins. Again, K8 and K18 mRNA levels remained unchanged during BrdU treatment. However, neither K8 nor K18 proteins were detected following treatment, although BrdU is known to alter expression of other genes in HL-60 cells. BrdU thus appears to act at a posttranscriptional level and in an epithelial-specific manner to reverse a block in keratin synthesis in keratin-negative lung cancer cells and increase synthesis in keratin-positive lung cancer cells. This may represent a regulatory step in early lung development or a mechanism whereby tumour cells downregulate expression of a differentiated phenotype.
Time-dependent remodeling of the bladder wall in growing rabbits after partial outlet obstruction
1997, Journal of Urology
We asked whether a urethral constriction gradually developed during growth would give rise to a structural remodeling of the bladder wall distinct from that of the mature rabbits in terms of cellular response.
We examined the serosa and detrusor muscle in immature rabbits whose urethra was obstructed at 30 days postnatal and studied 7 to 30 days after partial outlet obstruction. Morphometry, bromo-deoxyuridine (BrdU) incorporation, Western blotting and immunocytochemical staining with a panel of monoclonal antibodies specific to selected cytoskeletal, cytocontractile and membrane-related proteins unique to non-muscle and smooth muscle cells (SMC) were used to analyze the effects of obstruction on the differentiation pattern.
In comparison with results in adult obstructed bladders, we have found that in growing rabbits: (1) the cell conversion from fibroblasts to SMC, occurring within the ‘extrinsic' region of serosal thickening, takes place earlier; (2) newly formed SMC are localized exclusively to the thickened serosa, and can group in bundles depending on the density of the regional innervation; (3) the peak level of BrdU incorporation is more elevated than in the adult bladder wall; and (4) change in the phenotypic profile of SMC of detrusor muscle is delayed.
These data indicate that the basic features of structural remodeling in the two models are similar, though partial outlet obstruction produced in growing animals accelerates the fibroblast conversion to SMC and their spatial, differentiation-specific arrangement in the serosa. The late phenotypic changes in obstructed detrusor muscle correlate with the decline of the DNA synthesis level after an initial burst and strongly suggest that newly formed SMC in the serosa do not derive from pre-existing SMC.
Desmoplastic crosstalk in pancreatic ductal adenocarcinoma is reflected by different responses of panc-1, MIAPaCa-2, PaTu-8902, and CAPAN-2 cell lines to cancer-associated/normal fibroblasts
2021, Cancer Genomics and Proteomics

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Regular ArticleProliferation of Submesothelial Mesenchymal Cells during Early Phase of Serosal Thickening in the Rabbit Bladder Is Accompanied by Transient Keratin 18 Expression

Abstract

Regular Article
Proliferation of Submesothelial Mesenchymal Cells during Early Phase of Serosal Thickening in the Rabbit Bladder Is Accompanied by Transient Keratin 18 Expression