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PubMed1999

Characterization of serotypes and outer membrane protein profiles in enteroaggregative Escherichia coli strains.

Twenty-four Escherichia coli strains mainly isolated from children with diarrhea in São Paulo, and showing characteristics of enteroaggregative E. coli (EAEC), were characterized by serotyping and outer membrane protein (OMP) profiles. The relationship between these characteristics was evaluated, as well as the usefulness of OMP profiles in the clonal analysis of EAEC strains. All strains presented aggregative adherence to HeLa cells and were classified in two groups based on their interaction with the EAEC DNA probe. A diversity of serotypes and OMP profiles was observed in both groups studied. Although no significant correlation between serotypes and OMP profiles was observed, unique OMP profiles were identified in 80% of the probe-positive strains which were distributed in only 4 OMP profiles. This result may indicate the presence of a few clones in the probe-positive group. On the other hand, probe-negative strains seem to constitute a more diverse group. In general, the observed heterogeneity in serotypes and OMP profiles described in the present study suggest a great genetic diversity in EAEC isolates of either the same or different serotypes and in strains presenting the same EAEC markers identified in our community.

Bacterial Outer Membrane Proteins↗
PubMed2003

Stem cells and gastric cancer: role of gastric and intestinal mixed intestinal metaplasia.

All of the different types of stomach epithelial cells are known to be derived from a single progenitor cell in each gland. Similarly, cancers develop from single cells, based on data from clonality analysis in C3H/HeN<-->BALB/c chimeric mice. Using gastric and intestinal epithelial cell markers, intestinal metaplasia (IM) can be divided into two major types: a gastric and intestinal (GI) mixed type, and a solely intestinal (I) type. Ectopic expression of Cdx genes and down-regulation of Sox2 in isolated single GI mixed IM glands suggests abnormal differentiation of stem cells that can produce both gastric (G) and I type cells. Similarly, phenotypic expression of gastric cancer cells of each histological type can be clearly classified into G and I type epithelial cells. The heterogeneity of phenotypic expression of gastric cancer cells in individual cancers is assumed to reflect this intrinsic potential for differentiation in two directions. Gastric cancers at early stages, independent of the histological type, mainly consist of G type cells, and phenotypic shift from G to I type expression is clearly observed with progression. The data thus suggest IM may not be a preneoplastic change in gastric carcinoma, but rather that cells of the I type may appear independently in the gastric mucosa in IM and in gastric cancers. Intestinalization of gastric mucosa and cancer cells may represent a kind of homeotic transformation. Whether disturbance of the regulation of Sox2 and Cdx genes may be of importance to the biological behavior of gastric cancers should therefore be clarified in future studies.

Animals↗
PubMed1989

Circulating monoclonal B lymphocytes in multiple myeloma.

Peripheral blood mononuclear cells from 28 patients with multiple myeloma (MM) and nine patients with monoclonal gammopathy of unknown significance (MGUS) were studied by immunoglobulin gene analysis. Clonal immunoglobulin gene rearrangements in peripheral blood mononuclear cells (PBIGRA) were demonstrated in 10 of the 28 MM patients (36%). Bone marrow and peripheral blood mononuclear cells were studied simultaneously in five of these 10 patients, and identical gene rearrangements were demonstrated in both. The incidence of such gene arrangements was higher in patients with active disease (cases at presentation or relapsed = 10/19 [47%]) compared to remission status (0/9) and higher in untreated (47%) compared to treated patients (11%) (P less than 0.05). Patients with this phenomenon had higher serum calcium levels (P less than 0.001), and higher bone marrow plasma cell counts (P less than 0.05). Serum creatinine and beta 2-microglobulin were also higher but did not reach statistical significance. None of the patients with monoclonal gammopathy of uncertain significance had gene arrangements. Our findings confirm that circulating B lymphocytes are part of the malignant clone in MM and their presence correlates with high tumour volume.

Antibodies, Monoclonal↗
PubMed1992

Clonal lymphocytes are detectable in only some cases of MDS.

Clonal analysis of lymphocytes from patients with myelodysplastic syndrome (MDS) has been carried out using X-chromosome inactivation patterns detected by the probe M27 beta, and by polymerase chain reaction amplification of the immunoglobulin heavy chain gene hypervariable region, CDR3. Of 32 female patients heterozygous for M27 beta only seven (22%) demonstrate monoclonality of peripheral blood lymphocytes. 12 (37%) give unequivocal polyclonal results and the remaining cases give patterns of X-inactivation which cannot be interpreted either way. A study of 68 MDS patients showed five (7%) with a population of B-cells with a monoclonal rearrangement of CDR3 compared with none out of 60 normal individuals, none out of 15 with B-non Hodgkin lymphoma (B-NHL) in remission and 19 out of 25 (75%) of cases of B-chronic lymphocytic leukaemia (B-CLL). Monoclonal lymphocytes were found by both techniques in only two females with MDS. We conclude that the presence of polyclonal lymphocytes is a common finding in patients with MDS.

Adult↗
PubMed1994

Monoclonal Epstein-Barr virus genomes but lack of EBV-related protein expression in different types of gastric carcinoma.

Thirty-nine resection specimens of gastric carcinomas have been investigated for the presence of EBV RNA sequences using a highly sensitive non-radioactive in situ hybridization technique. Transcribed EBER sequences were found in seven (18%), including four cases of undifferentiated carcinoma with prominent lymphoid infiltration and three gastric adenocarcinomas. In the positive tumours all, or nearly all, tumour nuclei were distinctly labelled. No positive signals could be detected in the non-dysplastic epithelial cells or the reactive inflammatory infiltrate. Clonality analysis using specific probes to the variable tandem repeat region of the EBV yielded single episomal bands in all four cases tested, two of which were undifferentiated carcinomas and two adenocarcinomas. By means of immunohistology, no expression of the EBV-related proteins LMP or EBNA-2 was present in tumour cells of positive cases in in situ or blotting attempts. Our results suggest that infection of gastric carcinomas by the EB virus occurs early in tumourigenesis but, in contrast to nasopharyngeal carcinomas, does not result in the expression of EBV-specific proteins.

Adenocarcinoma↗
PubMed2006

Intracolony variation of bacterial gut microbiota among castes and ages in the fungus-growing termite Macrotermes gilvus.

The fungus-growing termites Macrotermes cultivate the obligate ectosymbiontic fungi, Termitomyces. While their relationship has been extesively studied, little is known about the gut bacterial symbionts, which also presumably play a crucial role for the nutrition of the termite host. In this study, we investigated the bacterial gut microbiota in two colonies of Macrotermes gilvus, and compared the diversity and community structure of bacteria among nine termite morphotypes, differing in caste and/or age, using terminal restriction fragment length polymorphism (T-RFLP) and clonal analysis of 16S rRNA. The obtained molecular community profiles clustered by termite morphotype rather than by colony, and the clustering pattern was clearly more related to a difference in age than to caste. Thus, we suggest that the bacterial gut microbiota change in relation to the food of the termite, which comprises fallen leaves and the fungus nodules of Termitomyces in young workers, and leaves degraded by the fungi, in old workers. Despite these intracolony variations in bacterial gut microbiota, their T-RFLP profiles formed a distinct cluster against those of the fungus garden, adjacent soil and guts of sympatric wood-feeding termites, implying a consistency and uniqueness of gut microbiota in M. gilvus. Since many bacterial phylotypes from M. gilvus formed monophyletic clusters with those from distantly related termite species, we suggest that gut bacteria have co-evolved with the termite host and form a microbiota specific to a termite taxonomic and/or feeding group, and furthermore, to caste and age within a termite species.

Age Factors↗
PubMed2005

From clones of cells to cloned genes and the proteinpaedia.

This brief description follows the association of the author with Ivan Lefkovits from 1971 until this volume. Sketches of our mutual interests are included. Time periods in California, Basel and Texas are described. Decisions about preparing new tools for clonal analysis are elucidated, and experimental approaches leading to the x-omic revolution are described.

B-Lymphocytes↗
PubMed1979

Prenatal monitoring for the Hunter syndrome: the heterozygous female fetus.

An abnormal level of 35S-sulfate labeled mucopolysaccharides was found in cultured amniotic fluid cells from a pregnancy, at risk for the Hunter syndrome, with a female fetal karyotype. Subsequent prenatal analyses suggested heterozygosity for the X-linked Hunter syndrome, and this was confirmed by clonal analysis of fibroblasts of the child after birth. The possible implications of abnormal biochemical results in association with a female karyotype in the prenatal diagnosis of the Hunter syndrome are discussed.

Amniocentesis↗
PubMed2006

Runx3 controls growth and differentiation of gastric epithelial cells in mammals.

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-beta, suggesting that Runx3 is a major growth regulator of gastric epithelial cells by regulating their response to TGF-beta. 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.

Animals↗
PubMed1997

Epstein-Barr virus (EBV)-induced CD30+ natural killer cell-type malignancy resembling malignant histiocytosis: malignant transformation in chronic active EBV infection associating hypogammaglobulinemia.

A 27-year-old male suffered from Epstein-Barr virus (EBV)-related liver dysfunction with persistent hypogammaglobulinemia. IgG titers to EBV antigens were significantly high, while other hepatitis markers were negative. Liver biopsy disclosed active intralobular inflammation. Two years later, he manifested persistent fever, leukopenia, effusions and hypoproteinemia, and his general condition worsened progressively. The peripheral blood small lymphocytes predominantly expressed natural killer (NK)-like phenotypes (CD2+, CD7+, CD16+, CD56+). Hepatosplenomegaly and marked elevation of serum lactic dehydrogenase were observed. He died of respiratory failure at the age of 29. At autopsy, the liver (2190 g), spleen (860 g), small bowel and mesenteric lymph nodes showed massive infiltration of large atypical lymphoid cells in close association with hemophagocytic histiocytes. Involvement was mildly noted also in the bone marrow, lungs, gall-bladder and kidneys. The atypical cells belonged to CD30+ activated NK-type cells expressing CD2, cytoplasmic CD3 epsilon, CD7, CD45RO, CD56, HLA-DR and HLA-DQ. T cell receptors (TCR), surface CD3, CD4, CD5 and CD8 were not expressed. Epstein-Barr virus-related small nuclear RNA (EBER1) and Epstein-Barr virus-associated nuclear antigen 1 were detected in the nuclei of a significant number of atypical cells, while EBV-related latent membrane protein-1 was negative. EBER1 was also identified in the nuclei of non-neoplastic small lymphocytes at both biopsy and autopsy. Monoclonal integration of the EBV genome into the lymphoma cells was shown by Southern blot analysis. Clonal rearrangement of TCR was undetectable. Roles of chronic active EBV infection in the development of NK cell-type malignancy resembling malignant histiocytosis are discussed.

Adult↗
PubMed2006

Identification and characterisation of human endometrial stem/progenitor cells.

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.

Endometrium↗
PubMed2002

Clonality structure in Candida dubliniensis.

Multilocus enzyme electrophoresis was performed on 76 European strains of Candida dubliniensis. Ten of the 20 enzyme-encoding loci were polymorphic, giving rise to 10 electrophoretic types within the sample studied. Investigation of the population genetics of a subset of 36 strains from HIV-infected patients in London showed the existence of strong heterozygote deficits and excesses associated with significant linkage disequilibria between pairs of loci. These findings, together with the predominance of multilocus genotypes, strongly suggest that C. dubliniensis is mainly (if not totally) clonal. Analysis of genotypes of a larger number of strains should confirm this conclusion and improve our understanding of the epidemiology of this pathogen.

AIDS-Related Opportunistic Infections↗
PubMed1993

T cells in reactive arthritis.

T cells appear to play a major role in the development, maintenance and also resolution of reactive arthritis (ReA). Recent advances in understanding the processes involved in T cell activation now allow us to examine the peripheral blood and synovial fluid T cell responses to given "arthritogenic" microorganisms in terms of antigen specificity, epitope identification, cytokine secretion patterns, HLA restriction and the role of different T cell subsets in ReA. Peripheral blood bulk proliferation and limiting dilution studies provide evidence that the peripheral T cell response against arthritis-associated gram-negative bacteria is decreased in patients developing immunological sequelae such as ReA after gastrointestinal infection. Using clonal analysis of synovial fluid CD4+ T cells it has been shown that a polyclonal rather than an oligoclonal response to a variety of bacterial antigens is induced at the site of synovitis and that these CD4+ T cells produce a Th1-type of cytokine. 65 kD heat shock protein may represent one of the possible linkages of anti-infectious and autoimmune reactions. Furthermore, a spectrum of killer cells is present in the synovial fluid of patients with ReA. This spectrum of cytotoxic T cells includes antigen-specific, class I-restricted alpha beta-TCR+CD8+ lymphocytes, antigen-specific, apparently non-MHC-restricted alpha beta-TCR+CD8+ lymphocytes and gamma delta-TCR+ cells with braod cytolytic activity directed against bacteria-infected target cells. HLA-B27-restricted Yersinia- or Salmonella-specific synovial fluid CD8+ T cells may provide the missing link between genetic disposition (HLA-B27) and extra-articular infection with arthritogenic bacteria in these patients.

Arthritis, Reactive↗
PubMed1998

The PS integrins are required for a regulatory event during Drosophila wing morphogenesis.

The PS1 and PS2 integrins are required for morphogenesis of the adult Drosophila wing. Clonal analysis experiments have shown that both integrins are necessary to maintain adhesion between the dorsal and ventral wing epithelia. We have found that early in wing morphogenesis, the integrins are also required for a regulatory event, and this may explain why PS1 and PS2 must be expressed on opposite surfaces of the wing at the onset of pupariation. Overexpression of integrin subunits during this early phase can lead to separation of dorsal and ventral surfaces, and we present evidence here that this dominant phenotype (the Blistermaker phenotype) results from a gain of integrin function, as opposed to negative interference from free integrin subunits. A possible model for an integrin signaling requirement in the wing is discussed.

Animals↗
PubMed2005

A clonogenic bone marrow progenitor specific for macrophages and dendritic cells.

Macrophages and dendritic cells (DCs) are crucial for immune and inflammatory responses and belong to a network of cells that has been termed the mononuclear phagocyte system (MPS). However, the origin and lineage of these cells remain poorly understood. Here, we describe the isolation and clonal analysis of a mouse bone marrow progenitor that is specific for monocytes, several macrophage subsets, and resident spleen DCs in vivo. It was also possible to recapitulate this differentiation in vitro by using treatment with the cytokines macrophage colony-stimulating factor and granulocyte-macrophage colony-stimulating factor. Thus, macrophages and DCs appear to renew from a common progenitor, providing a cellular and molecular basis for the concept of the MPS.

Animals↗
PubMed2000

Stem cells in epithelial tissues.

Most, if not all, epithelial tissues contain stem cells. They are responsible for normal tissue renewal or for regeneration following damage. Our present knowledge of their properties is limited and is mainly derived from studies of cell kinetics and from clonal analysis.

Animals↗
PubMed2025

Engraftment and persistence of HBB base-edited hematopoietic stem cells in nonhuman primates.

Sickle cell disease (SCD) is caused by a single nucleotide change in the &#x3b2;-globin gene that adenine base editors can convert to the nonpathogenic Makassar &#x3b2;-globin variant. Here, we evaluated the long-term efficiency and off-target editing potential of autologous Makassar base editing in three rhesus macaques as a step toward human translation. Base editing of CD34+CD90+ hematopoietic stem cells (HSCs) at the Makassar locus reached greater than 60% efficiency using a bystander nucleotide as a proxy for the sickle cell target in cells from healthy macaques. No impact on myeloid and erythroid colony formation was seen, and clonal analysis revealed that >90% of HSCs were edited, >20% with biallelic editing. After transplantation of autologous gene-edited HSCs, all three macaques rapidly recovered neutrophils, red blood cells, and platelets with stable editing of 25.6%, on average, observed across nucleated blood cells. Similarly, the bone marrow stem cell compartment maintained over 20% of cells harboring mono- or biallelic edits. Off-target editing was assessed at over 900 candidate sites, with editing observed at eight sites, but no selection for or impact of these edits was observed throughout engraftment. These data support further translation of base editing of autologous HSCs for the treatment of patients with SCD.

Animals↗
PubMed2005

Environmental contamination with vancomycin-resistant enterococci from hospital sewage in Portugal.

Vancomycin-resistant enterococci (VRE) were detected in samples of sewage obtained downstream of hospitals of the Porto area in Portugal, and in samples from the Douro Estuary. Clonal analysis, Tn1546 typing, and presence of putative virulence traits indicate the clinical origin of these isolates. This observation highlights the importance of hospital sewage in the VRE contamination of the environment.

Anti-Bacterial Agents↗