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Clonal Analysis (clonal + analysis)
Selected AbstractsRunx3 controls growth and differentiation of gastric epithelial cells in mammalsDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2006Hiroshi Fukamachi Runx3 is a transcription factor expressed by gastric epithelial cells. In the Runx3,/, mouse, gastric epithelia exhibited hyperplasia, and epithelial apoptosis was suppressed. By analyzing growth of the epithelial cells in primary culture, we found that Runx3,/, gastric epithelial cells are less sensitive to the growth-inhibitory and apoptosis-inducing activities of TGF-,, suggesting that Runx3 is a major growth regulator of gastric epithelial cells by regulating their response to TGF-,. We also found that Runx3 plays an important role in the control of gastric epithelial differentiation. When subcutaneously implanted into nude mice, Runx3,/, gastric epithelial cells formed tumors in which some cells differentiated into intestinal-type cells. Clonal analysis showed that gastric epithelial cells transdifferentiate into intestinal-type cells in the tumor. Considering that gastric epithelial differentiation is very stable, and that intestinal-type cells never differentiate in the mouse stomach, it is remarkable that gastric epithelial cells transdifferentiate into intestinal-type cells. We conclude that Runx3 is deeply involved in the control of both growth and differentiation of gastric epithelial cells. The role of Runx3 in the specification of gastric epithelial cells is discussed. [source] Clonal analysis of patterns of growth, stem cell activity, and cell movement during the development and maintenance of the murine corneal epitheliumDEVELOPMENTAL DYNAMICS, Issue 4 2002J. Martin Collinson Abstract Patterns of growth and cell movement in the developing and adult corneal epithelium were investigated by analysing clonal patches of LacZ -expressing cells in chimeric and X-inactivation mosaic mice. It was found that cell proliferation throughout the basal corneal epithelium during embryogenesis and early postnatal life creates a disordered mosaic pattern of LacZ+ clones that contrasts with patterns of proliferation and striping produced during the later embryonic stages of retinal pigmented epithelium development. The early mosaic pattern in the corneal epithelium is replaced in the first 12 postnatal weeks by an ordered pattern of radial stripes or sectors that reflects migration without mixing of the progeny of clones of limbal stem cells. In contrast to previous assumptions, it was found that maturation of the activity of limbal stem cells and the pattern of migration of their progeny are delayed for several weeks postnatally. No evidence was found for immigration of the progeny of stem cells until the 5th postnatal week. There are approximately 100 clones of limbal stem cells initially, and clones are lost during postnatal life. Our studies provide a new assay for limbal and corneal defects in mutant mice. © 2002 Wiley-Liss, Inc. [source] Evolution of multi-drug resistant hepatitis B virus during sequential therapy,HEPATOLOGY, Issue 3 2006Hyung Joon Yim Multi-drug resistant hepatitis B virus (HBV) has been reported in hepatitis B patients who received sequential antiviral therapy. In vitro studies showed that HBV constructs with mutations resistant to lamivudine and adefovir have marked reduction in sensitivity to combination of lamivudine and adefovir, whereas constructs with mutations resistant to either drug remain sensitive to the other drug. We conducted this study to determine whether mutations conferring resistance to multiple antiviral agents co-locate on the same HBV genome in vivo and to describe the evolution of these mutations. Sera from six patients who had been found to have multi-drug resistant HBV mutations to lamivudine + adefovir, lamivudine + hepatitis B immunoglobulin (HBIG), or lamivudine + entecavir on direct sequencing were cloned after nested polymerase chain reaction (PCR). Analysis of 215 clones from 11 samples with multi-drug resistant mutations on direct sequencing showed that 183 (85%) clones had mutations to both therapies on the same genome; 31 clones had lamivudine-resistant mutants only. Clonal analysis of serial samples from three patients showed progressive evolution from all clones with lamivudine-resistant HBV mutations only to mixtures of clones that have multi-drug resistant mutations and clones that have lamivudine-resistant HBV mutations only, and ultimately all clones having multi-drug resistant HBV mutations. In conclusion, mutations conferring resistance to multiple antiviral agents co-locate on the same viral genome, suggesting that combination therapy directed against mutants resistant to each treatment may not be adequate in suppressing multi-drug resistant HBV. De novo combination therapy may prevent the emergence of multi-drug resistant mutants. (HEPATOLOGY 2006;44:703,712.) [source] Tumor-infiltrating lymphocytes derived from human renal cell carcinoma: Clonal analysis of its characteristicsINTERNATIONAL JOURNAL OF UROLOGY, Issue 3 2008Tomoyuki Shimabukuro Aim: To assess the characteristics of activated tumor-infiltrating lymphocytes (TIL), we report the isolation, growth response, and functional analysis of a CD4 - CD8+ TIL-clone derived from human renal cell carcinoma (RCC). Methods: Bulk TILs were expanded from a human RCC and the lymphocytes were separated into a CD8+ enriched population. Subsequently, using the limiting dilution technique, a TIL clone was established and its growth response, phenotype and cytotoxic activity were analyzed. Results: A clone, T16-13, by day 94 numbering 1 × 107 cells, was harvested and characterized as a CD4 - CD8+ clone. On day 144, the cytotoxic activity of this clone against the autologous tumor was relatively high (2.3 ± 0.7 LU30/106 cells). Meanwhile, against allogeneic renal tumors, there was no cytotoxic activity (,0.1 LU30/106 cells). Conclusions: A TIL clone possessing modest autologous tumor-specific cytotoxicity can be isolated from human RCC. The characteristics analysis of various TIL clones may provide a better understanding of an RCC tumor microenvironment and may help to establish new modalities for the treatment of patients with metastatic kidney cancer. [source] Clonal analysis of cutaneous fibrous histiocytoma (dermatofibroma)JOURNAL OF CUTANEOUS PATHOLOGY, Issue 7 2002Pei Hui Background:, Dermatofibroma (DF) or cutaneous fibrous histiocytoma is a common benign fibrohistiocytic lesion involving the dermis and subcutis. Histologically, it is subclassified into fibroblastic and histiocytoid forms. Its histogenesis is controversial. While often referred to as a neoplastic process, definite evidence of neoplasia in DF has been lacking. Alternatively, some authorities have suggested that DF is a fibrosing inflammatory process. Diagnostically, the most important question faced is the distinction from dermatofibrosarcoma protuberans (DFSP). Misdiagnosis can occur, as the early phase of DFSP can simulate DF, particularly the deep and cellular forms of DF. Methods:, To address this issue, and to investigate whether DF is in fact a neoplasm, we evaluated 31 examples of DF of various histological types in female patients and assessed clonality by analyzing X-chromosome inactivation as indicated by the methylation status of the androgen receptor gene (HUMARA). Representative cases of DFSP were analyzed for comparison. Results:, Among the selected 31 cases of DF, 24 cases provided intact DNA and informative polymorphism at the AR alleles, including one case of recurrent deep fibrous histiocytoma. Among these 24 cases, randomly inactivated AR alleles were observed in 17 cases including a deep, recurrent fibroblastic DF. A non-random inactivation at AR alleles was observed in seven cases, of which six cases showed either typical histiocytoid form of DF (four cases) or mixed cell types with predominant histiocytoid cell type (two cases). One fibroblastic DF also showed a monoclonal pattern. HUMARA analysis of DFSP revealed non-random inactivation of polymorphic AR alleles. Conclusions:, These findings suggest that DF is a heterogeneous process. Monoclonal genotype was found in DFs with histiocytoid or mixed type with predominant histiocytoid features, suggesting that histiocytoid cells probably represent the neoplastic component. The fibroblastic form of DF may represent a reactive fibroblastic proliferation. Alternatively, it may represent a true neoplasm whose neoplastic cell type has been obscured by prominent reactive fibroblastic component. [source] Acral pseudolymphomatous angiokeratoma of children: immunohistochemical and clonal analyses of the infiltrating cellsJOURNAL OF CUTANEOUS PATHOLOGY, Issue 5 2002Yoshitaka Hagari Background:, Acral pseudolymphomatous angiokeratoma of children (APACHE) is a disorder characterized clinically by red nodules and histopathologically by a massive subepidermal lymphohistiocytic infiltrate. Although it was initially thought to be a vascular nevus, it has never been regarded as a pseudolymphoma. Case report: We report a 7-year-old-girl with small red nodules on the dorsum of the right foot and a 73-year-old man with asymptomatic brown-red nodules on the lower extremities. Results:, Histopathologic examination revealed a massive lymphohistiocytic infiltrate with plasma cells, some eosinophils, or a multinucleated giant cell immediately beneath the epidermis. Thick-walled vessels were observed in the infiltrate. These characteristics are identical to those of acral pseudolymphomatous angiokeratoma of children. The infiltrate was composed mainly of equal numbers of CD4+ or CD8+ T cells and equal numbers of B cells stained for , or , light chains. PCR amplification of rearranged immunoglobulin heavy chain genes or T-cell receptor , genes showed no evidence of clonality, suggesting that these infiltrates were polyclonal both for B and T cells. Conclusions:, Our data support the idea that this disorder represents a reactive process. The modified term ,papular angiolymphoid hyperplasia' would define this disorder more appropriately. [source] Characterization of the epithelial cell adhesion molecule (EpCAM)+ cell population in hepatocellular carcinoma cell linesCANCER SCIENCE, Issue 10 2010Osamu Kimura Accumulating evidence suggests that cancer stem cells (CSC) play an important role in tumorigenicity. Epithelial cell adhesion molecule (EpCAM) is one of the markers that identifies tumor cells with high tumorigenicity. The expression of EpCAM in liver progenitor cells prompted us to investigate whether CSC could be identified in hepatocellular carcinoma (HCC) cell lines. The sorted EpCAM+ subpopulation from HCC cell lines showed a greater colony formation rate than the sorted EpCAM, subpopulation from the same cell lines, although cell proliferation was comparable between the two subpopulations. The in vivo evaluation of tumorigenicity, using supra-immunodeficient NOD/scid/,cnull (NOG) mice, revealed that a smaller number of EpCAM+ cells (minimum 100) than EpCAM, cells was necessary for tumor formation. The bifurcated differentiation of EpCAM+ cell clones into both EpCAM+ and EpCAM, cells was obvious both in vitro and in vivo, but EpCAM, clones sustained their phenotype. These clonal analyses suggested that EpCAM+ cells may contain a multipotent cell population. Interestingly, the introduction of exogenous EpCAM into EpCAM+ clones, but not into EpCAM, clones, markedly enhanced their tumor-forming ability, even though both transfectants expressed a similar level of EpCAM. Therefore, the difference in the tumor-forming ability between EpCAM+ and EpCAM, cells is probably due to the intrinsic biological differences between them. Collectively, our results suggest that the EpCAM+ population is biologically quite different from the EpCAM, population in HCC cell lines, and preferentially contains a highly tumorigenic cell population with the characteristics of CSC. (Cancer Sci 2010) [source] Mixed primary culture and clonal analysis provide evidence that NG2 proteoglycan-expressing cells after spinal cord injury are glial progenitorsDEVELOPMENTAL NEUROBIOLOGY, Issue 7 2007Soonmoon Yoo Abstract NG2+ cells in the adult rat spinal cord proliferate after spinal cord injury (SCI) and are postulated to differentiate into mature glia to replace some of those lost to injury. To further study these putative endogenous precursors, tissue at 3 days after SCI or from uninjured adults was dissociated, myelin partially removed and replicate cultures grown in serum-containing or serum-free medium with or without growth factors for up to 7 days in vitro (DIV). Cell yield after SCI was 5,6 times higher than from the normal adult. Most cells were OX42+ microglia/macrophages but there were also more than twice the normal number of NG2+ cells. Most of these coexpressed A2B5 or nestin, as would be expected for glial progenitors. Few cells initially expressed mature astrocyte (GFAP) or oligodendrocyte (CC1) markers, but more did at 7 DIV, suggesting differentiation of glial precursors in vitro. To test the hypothesis that NG2+ cells after SCI express progenitor-like properties, we prepared free-floating sphere and single cell cultures from purified suspension of NG2+ cells from injured spinal cord. We found that sphere cultures could be passaged in free-floating subcultures, and upon attachment the spheres clonally derived from an acutely purified single cell differentiated into oligodendrocytes and rarely astrocytes. Taken together, these data support the hypothesis that SCI stimulates proliferation of NG2+ cells that are glial progenitor cells. Better understanding the intrinsic properties of the NG2+ cells stimulated by SCI may permit future therapeutic manipulations to improve recovery after SCI. © 2007 Wiley Periodicals, Inc. Develop Neurobiol, 2007. [source] Emergence of fluoroquinolone-resistant clonal group A: clonal analysis of Norwegian and Russian E. coli isolatesAPMIS, Issue 8 2010LINDA STRAND Strand L, Jenkins A, Grude N, Allum A-G, Mykland H-C, Nowrouzian FL, Kristiansen B-E. Emergence of fluoroquinolone-resistant clonal group A: clonal analysis of Norwegian and Russian E. coli isolates. APMIS 2010; 118: 571,77. We describe a study of urinary tract and intestinal isolates of Escherichia coli from Norway and Russia using automated ribotyping, single nucleotide polymorphism analysis for clonal group A (CgA) supplemented with phylogrouping, virulence gene profiling and resistance profiling. CgA comprised 19% of the Norwegian UTI isolates from 2001. Two highly multiresistant fluoroquinolone-resistant CgA isolates were found. Ribotypes clustered into four major and six minor groups (ribogroups). Fluoroquinolone-resistant isolates and phylogroups A and B1 were associated with ribogroup RA. Ribogroup RB predominated among Russian UTI isolates and was predominantly phylogroup A and depleted in P-fimbriae. Ribogroup RC predominated among Norwegian UTI isolates and was rich in virulence factors (S-fimbriae, haemagglutinin and haemolysin) and predominantly phylogroup B2 and D. Ribogroup RG was associated with CgA and predominantly phylogroup D. Ribogroups RD, RE and RF had too few members for statistical analysis. The correlation between ribotype and phylogenetic group was not as strong as reported in other studies. [source] Identification and characterisation of human endometrial stem/progenitor cellsAUSTRALIAN AND NEW ZEALAND JOURNAL OF OBSTETRICS AND GYNAECOLOGY, Issue 3 2006Caroline E. GARGETT Abstract The human endometrium is a dynamic, cyclically regenerating tissue. Because adult stem cells are present in other regenerative tissues, a clonal analysis of purified endometrial epithelial and stromal cells derived from hysterectomy tissue was undertaken, as a first attempt to identify and characterise endometrial stem/progenitor cells. Rare populations of epithelial and stromal cells were clonogenic. Only those cells initiating large clones, 0.09% of epithelial cells and 0.02% of stromal cells, are likely to be endometrial stem/progenitor cells. [source] Molecular characterisation and clonal analysis of group A streptococci causing pharyngitis among paediatric patients in Palermo, ItalyCLINICAL MICROBIOLOGY AND INFECTION, Issue 2 2006G. Lorino Abstract Group A streptococci (n = 123), isolated consecutively from paediatric patients with pharyngitis from Palermo, Italy, were analysed. The emm and sof genes were sequenced, the presence of the speA and speC genes was investigated, and the macrolide resistance phenotypes and genotypes were determined. A limited number of emm/sof genotypes was found, and the most prevalent types were different from those found in a previous study from Rome. Macrolide resistance was found in the most prevalent clones, suggesting that the spread of mobile antibiotic resistance genes among the fittest clones in the community was the main mechanism influencing macrolide resistance rates in different emm types. [source] | |||||||||||||