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PubMed1996

Clonal analysis of meningiomas.

Meningiomas are primary brain tumors arising from meningothelial cells. They usually grow slowly and are surgically easy to separate from the brain. A recent clonal analysis of meningiomas, using methylation-sensitive restriction fragment length polymorphisms, suggested a monoclonal origin. Using the same technique but with a highly informative X chromosome probe (M27 beta), we found that 17 (85%) of the 20 meningiomas analyzed were informative. Of the 17 informative tumors, 8 (47%) were monoclonal, 3 (18%) had loss of heterozygosity on the X chromosome, and, unexpectedly, 6 (35%) had a polyclonal pattern. Samples from two areas of one tumor showed a monoclonal pattern and loss of heterozygosity, respectively, on the X chromosome. A review of the histopathological and radiological features of the 17 informative tumors did not help to distinguish the clonal from the polyclonal tumors. We conclude that meningiomas are heterogeneous in clonal composition.

Adult↗
PubMed2006

Liver cell adenoma: a case report with clonal analysis and literature review.

We report a case of liver cell adenoma (LCA) in a 33-year-old female patient with special respect to its clonality status, pathogenic factors and differential diagnosis. The case was examined by histopathology, immunohistochemistry and a clonality assay based on X-chromosomal inactivation mosaicism in female somatic tissues and polymorphism at androgen receptor focus. The clinicopathological features of the reported cases from China and other countries were compared. The lesion was spherical, sizing 2 cm in its maximal dimension. Histologically, it was composed of cells arranged in cords, most of which were two-cell-thick and separated by sinusoids. Focal fatty change and excessive glycogen storage were observed. The tumor cells were round or polygonal in shape, resembling the surrounding parenchymal cells. Mitosis was not found. No portal tract, central vein or ductule was found within the lesion. The tumor tissue showed a positive reaction for cytokeratin (CK) 18, but not for CK19, vimentin, estrogen and progesterone receptors. Monoclonality was demonstrated for the lesion, confirming the diagnosis of an LCA. Clonality analysis is helpful for its distinction from focal nodular hyperplasia.

Adenoma, Liver Cell↗
PubMed1987

Clonal analysis using recombinant DNA probes from the X-chromosome.

It has been demonstrated that restriction fragment length polymorphisms of X-chromosome genes can be used in conjunction with methylation patterns to determine the clonal composition of human tumors. In this report, we show that several X-chromosome probes can be used for such analyses. In particular, probes derived from the hypoxanthine phosphoribosyltransferase gene and the phosphoglycerate kinase gene could be used for clonal analysis in over 50% of American females. The X-inactivation patterns observed with these probes were found to accurately reflect clonality in more than 95% of 92 tumors tested.

Alleles↗
PubMed1996

Clonal analysis of focal nodular hyperplasia of the liver.

Recent evidence suggests that focal nodular hyperplasia of the liver (FNH) may represent a hyperplastic response to a vascular malformation, but the precise etiology remains unclear. We performed a clonal analysis of ten FNHs from nine patients by patterns of X chromosome inactivation. DNA isolated from paraffin-embedded specimens was subjected to polymerase chain reaction amplification for a highly polymorphic region of the human androgen receptor gene (HUMARA). Predigestion of tumor DNA with the methylation-sensitive, restriction enzyme HpaII allowed for selective amplification of the methylated (inactivated) allele. Of the nine patients analyzed, seven were heterozygous for the HUMARA polymorphism and informative for analysis. One informative patient had two lesions, for a total of eight FNHS. Amplification of lesional DNA after HpaII digestion demonstrated clonality in six of the eight informative cases. Paired tissue samples from different lesional areas were available in four of the six FNHs with evidence of clonality. In three of the four cases, DNA extracted from the two tissue samples showed both evidence of clonality and an identical pattern of X chromosome inactivation. In the remaining case, one sample showed evidence of clonality whereas the other was nonclonal. Three hepatic adenomas from two informative patients were also analyzed for comparative purposes, all of which showed evidence of clonality after HpaII digestion. The current study illustrates that most cases of FNH show a uniform pattern of X chromosome inactivation consistent with clonality.

Adenoma↗
PubMed1985

A clonal analysis of the requirement for the trithorax gene in the diversification of segments in Drosophila.

The requirement for the trithorax gene during imaginal cell proliferation has been analysed by clonal analysis. Clones carrying the allelic Regulator of bithorax and induced by mitotic recombination at different developmental stages. The results indicate that the trithorax gene is required at least until the beginning of the third larval instar to ensure the correct differentiation of head, thoracic, and abdominal structures, and are consistent with a role for the gene in maintaining the appropriate levels of Antennapedia and bithorax complex expression.

Abdomen↗
PubMed1993

Phenotypic and clonal analysis of T lymphocytes in childhood immune thrombocytopenic purpura.

In an attempt to identify and characterize T-lymphocyte immunoregulatory abnormalities in immune thrombocytopenic purpura (ITP), we have performed phenotypic and clonal analysis on peripheral T lymphocytes from 23 children with ITP. Quantitation of lymphocyte subpopulations showed that children with acute ITP had higher numbers of CD45RA+ and lower numbers of CD45RO+ T cells than children with chronic ITP or controls, but these differences may be age related. Analysis of T-cell receptor variable beta gene usage identified 2 boys with chronic ITP and elevated numbers of V beta 8+ T cells. Eight T-cell clones were established (6 CD4+, 4B4+ helper-inducer lines and 2 CD8+ lines) that showed in vitro proliferation against allogeneic platelets. The addition of autologous antigen-presenting cells enhanced the proliferation of six clones, but not for two clones that coexpressed natural killer (NK) markers. Four of seven positive clones also had measurable interleukin (IL)-2 secretion following platelet stimulation, providing further evidence for T-cell reactivity. Our results provide the first evidence that patients with ITP may have platelet-reactive T lymphocytes identifiable at the clonal level, supporting the hypothesis that autoreactive peripheral T lymphocytes may mediate or participate in the pathogenesis of this disorder.

Acute Disease↗
PubMed1999

Clonal analysis of adenosquamous carcinoma of the lung.

Adenosquamous carcinoma of the lung is a subset of pulmonary carcinomas, and comprises less than 4% of lung carcinomas. Its histogenesis remains unclear. The clonality of adenosquamous carcinoma from four female patients was analyzed to determine whether the clonality between the squamous cell and adenocarcinomatous components coincides. Each lesion was precisely microdissected from methanol-fixed sections. Adjacent normal lung tissue was collected as a normal control. DNA was extracted for clonal analysis based on an X-chromosome-linked polymorphic marker, the human androgen receptor gene (HUMARA). HUMARA was found to be amplified with or without previous digestion by the methylation-sensitive restriction endonuclease HpaII. All four cases were informative. Squamous cell and adenocarcinomatous components showed identical monoclonal patterns in all four patients. In one case, only the squamous cell carcinomatous component showed loss of heterozygosity of the HUMARA locus. The results suggest that the squamous cell and adenocarcinomatous components originate from the same cell.

Aged↗
PubMed1990

Clonal analysis of chronic myeloproliferative disorders using X-linked DNA polymorphisms.

Restriction fragment length polymorphisms of the X-chromosome genes phosphoglycerate kinase and hypoxanthine phosphoribosyl transferase were used to study clonality in peripheral blood leukocytes from 48 women with chronic myeloproliferative disorders (c-MPD). A total of 50% of patients were heterozygous for one or both of the polymorphic loci. These included 17 cases with polycythemia vera, four patients with essential thrombocythemia (ET), and three cases with idiopathic myelofibrosis (IMF). A clear-cut monoclonal X-inactivation pattern was observed in 17 of 24 cases including all IMF patients. Only one patient with PV exhibited a nonclonal composition of her leukocytes, while six cases demonstrated a predominantly clonal pattern in peripheral blood cells. Among the latter category reckoned three of four ET patients. Cell separation analyses were performed in one ET and three PV patients. In all four cases a monoclonal pattern of the granulocyte fraction could be established, while T lymphocytes of these patients were of nonclonal origin. These data suggest that the vast majority of c-MPDs arise from multipotent hematopoietic stem cells. Moreover, this type of clonal analysis might be of help in discriminating between primary MPD and reactive processes.

Adult↗
PubMed2002

Polymerase chain reaction-based clonality analysis in thyroid lymphoma.

A previous patho-epidemiological study indicated that thyroid lymphoma (TL) evolves among active lymphoid cells into chronic lymphocytic thyroiditis (CLTH), a thyroid-specific autoimmune disease. In this study, clonality of B-cells in the CLTH and TL lesions was analyzed using polymerase chain reaction (PCR)-based method on surgically resected samples from 10 cases of TL; 7 mucosa-associated lymphoid tissue (MALT) lymphoma and 3 diffuse large B-cell lymphoma (DLBCL). CLTH lesions coexisted in all the cases with MALT lymphoma, but not in the three DLBCL cases. In cases of MALT lymphoma, the lymphomatous and CLTH areas were separately microdissected from each section and analyzed for clonality. In the cases of DLBCL, the whole specimens were used for clonality analysis. CLTH lesions showed smear in 6 samples, two bands in one, and more than three (oligoclonal pattern) in 2. MALT lymphoma lesions showed single or two bands (monoclonal pattern) in 4, oligoclonal pattern in 4, and smear in one. DLBCL showed monoclonal pattern in two and oligoclonal pattern in one. One common band was present among two separate MALT lesions in one case, but no common bands were found in the remaining six cases. These findings suggested the clonal evolution of B-cell from polyclonal to monoclonal proliferation to take place in the continuum of lymphoproliferative lesions into autoimmune thyroiditis.

Aged↗
PubMedDate not supplied

[Clonal analysis of the development of the pigment epithelium of the eye in chimeric or/or----AKR mice].

The development of cell clones was studied in the retinal pigment epithelium of chimaeric mice or/or----AKR. The clonal analysis suggests that on the 13th day of gestation the cells in the retinal pigment epithelium are distributed almost randomly while in the adults they are grouped in small coherent clones. In the retinal pigment epithelium the AKR cell predominated over the or/or cells. The interaction of mutant and normal clones during the eye development leads, in most cases, to the normalization of eyes in chimaeras. Cases of microphthalmia and asymmetry in distribution of clones suggest irregular and random distribution of these clones in the embryo.

Animals↗
PubMed2005

A computational method for inferring growth parameters and shape changes during development based on clonal analysis.

We describe a method for estimating growth parameters in various regions of a developing organ undergoing cell divisions, along with the corresponding changes in organ shape. Growth parameters are computed by coupling clonal analysis with a growth model, allowing a wide range of developmental stages to be covered. The method was applied to the development of dorsal petal lobes of Antirrhinum majus. The resulting description of growth patterns and shape changes is consistent with direct observations using scanning electron microscopy. This method can potentially be applied to other organs, and opens the way to comparative studies of growth and gene expression patterns.

Antirrhinum↗
PubMed1999

Immunohistochemical and clonal analysis of minute pulmonary meningothelial-like nodules.

The histogenesis of meningothelial-like nodule or so-called minute pulmonary chemodectoma remains unclear, with various immunohistochemical analyses giving inconsistent results. We performed an immunohistochemical and clonal analysis of minute pulmonary meningothelial-like nodules. Thirty-one histologically defined meningothelial-like nodules in 14 cases were stained immunohistochemically. One case had multiple lesions with brown pigment granules, which were positively stained with Berlin blue method, indicating the presence of hemosiderin. All meningothelial-like nodules were positive for vimentin and epithelial membrane antigen (EMA), but not for S-100 protein, chromogranin A, or synaptophysin. Five of 13 cases (13 of 28 lesions) were positive for CD68 by KP-1. Ten cases (24 lesions) stained for CD68 by PG-M1 were weakly positive. All lesions were negative for lysozyme, myosin, actin, keratin, and melanoma-associated antigen. Alveolar macrophages were intensely positive for CD68 and lysozyme in all examined cases. We analyzed the clonality of 11 minute pulmonary meningothelial-like nodule lesions in two female cases based on an X-chromosome-linked polymorphic marker, the human androgen receptor gene (HUMARA). The HUMARA was found to be amplified with or without prior digestion by the methylation-sensitive restriction endonuclease HpaII. Six of 11 lesions showed monoclonal expansion. Five lesions in a multiple case showed different patterns of monoclonality. Our findings showed that minute pulmonary meningothelial-like nodules have meningothelial-like and phagocytic characteristics but no muscular phenotype. Furthermore, some minute pulmonary meningothelial-like nodules may show monoclonal expansion, whereas others are polyclonal. Our data indicate that minute pulmonary meningothelial-like nodules are reactive rather than neoplastic.

Biomarkers↗
PubMed1991

In vitro clonal analysis of progenitor cell patterns in dorsal root and sympathetic ganglia of the quail embryo.

The possible presence of pluripotent cells in dorsal root (DRG) and sympathetic ganglia (SG) of the quail embryo has been investigated by in vitro clonal analysis. Both types of ganglia originate from the neural crest. At, or soon after, initiation of emigration from the neural tube, the neural crest appears as a mixed population of pluripotent cells and cells with more restricted developmental capacities. In the present study it was determined that pluripotent cells and precursor cells with restricted developmental potentials are also present in early DRG and SG, and that their proportions change with progressing age of the embryo. As in the neural crest, cells in one class are at least tripotent, able to give rise to pigment cells, sensory neurons, and cells in the sympathoadrenal lineage. Cells in the other class appear to have lost the melanogenic potential, but generate cells in the sensory neuron and sympathoadrenal lineages. In addition, there are two types of cells that seem to be committed to the melanogenic and sensory neuron lineages, respectively. Apparently committed melanogenic cells within the DRG are not detected after the first third of embryonic development, whereas precursor cells that are at least bipotent and generate both types of neurons persist in both DRG and SG at least through the first half of embryonic development. Neurogenic cells that are apparently committed to the sensory neuron or sympathoadrenal lineages were observed in the appropriate type of ganglion only, suggesting that location-specific cues influence the choice of phenotype generated by pluripotent neural crest cells.

Animals↗
PubMed2000

Diffuse leiomyomatosis of the uterus: a case report with clonality analysis.

Diffuse uterine leiomyomatosis is a rare condition distinguished from the common uterine leiomyomata by involvement of the entire myometrium by innumerable, ill-defined, often small and confluent, histologically benign smooth-muscle nodules. Fourteen cases have been previously described in the literature. We report a case of diffuse leiomyomatosis in a 39-year-old woman. Several microscopic foci of the process were microdissected for clonality analysis. All samples showed a non-random X-chromosome inactivation pattern, and thus were consistent with a monoclonal neoplastic population of cells. However, in different foci of tumor, different X chromosomes were inactivated, supporting the independent origin of neoplastic clones and rejecting the possibility of a single clonal origin of all tumor cells. The results of the molecular analysis suggest that diffuse uterine leiomyomatosis may be an exuberant example of diffuse and uniform involvement of the entire myometrium by multiple leiomyomata. HUM PATHOL 31:1429-1432.

Adult↗
PubMed2005

Clonal analysis of HIV-1 variants in proviral DNA during treatment interruption in patients with multiple therapy failures.

BACKGROUND: Treatment interruption (TI) in antiretroviral-treated patients on virological failure (VF) often results in a shift from resistant to wild-type HIV-1 in plasma. A clonal analysis was set out to determine the importance of archiving of resistant HIV-1 variants during TI and its relationship with the occurrence of a VF after treatment resumption. METHODS: A fragment of the pol gene was cloned and sequenced from peripheral blood mononuclear cells DNA sampled at the end of TI. Clonal sequences were compared to bulk plasma sequences determined before TI, after TI and at VF. RESULTS: Four patients were enrolled; all had a complete reversion to wild-type HIV-1 at the end of TI. In two patients with subsequent VF, archiving of minority resistant variants was detected in proviral DNA. Archived resistant variants were found to be phylogenetically linked to sequences detected before TI and at VF, suggesting the re-expansion of resistant HIV-1 from archived quasi-species. CONCLUSION: In patients having a TI in the context of VF, archiving of resistant HIV-1 variants in proviral DNA can be involved into the mechanisms of further VF after treatment resumption.

Anti-HIV Agents↗
PubMed1990

Clonal analysis of segmental and compartmental homoeotic transformations in polycomb mutants of Drosophila melanogaster.

We describe two new effects of Polycomb mutations on the determination of compartments in the wing of Drosophila. Ventral and posterior wing compartments are transformed partially to their dorsal and anterior counterparts. Although these new phenotypes are most strongly expressed in lethal pharate adults heteroallelic for Pc2 and a new allele PcT2, they are also found regularly but with low expressivity as dominant phenotypes of all the other Pc alleles we tested. Several different intersegmental homoeotic transformations caused by Polycomb have previously been described, leading to the hypothesis that Polycomb regulates the activity of certain selector genes normally active in specific segments. We now show that the degree of expression of the inter- and intrasegmental transformations are highly correlated in a range of different Pc genotypes, and that more than one determinative decision can be affected in a single compartment. This suggests that the wild-type Pc product may act as a general regulator of several different selector genes so as to influence both early embryonic and later determinative decisions in the imaginal discs. To test this idea we used clonal analysis to look at the effects of Pc on clonal restrictions at the dorsoventral and anterioposterior compartment boundaries, and its time of action with respect to each phenotype.

Alleles↗
PubMedDate not supplied

Establishment and application of a novel T cell clonality analysis using single-strand conformation polymorphism of T cell receptor messenger signals.

To identify the existence of antigen specific T cell responses and to follow the changes of these reactions, it is considered useful to evaluate whether certain T cells clonally accumulate in the lymphocyte population. For this purpose, we have established a novel method to analyze T cell clonality using a combination of reverse transcriptase-polymerase chain reaction of T cell receptor beta chain transcripts and single-strand conformation polymorphism (SSCP). Using this method, we obtained a smear-like pattern of electrophoresed DNA from the heterogeneous T cell population. On the other hand, a single T cell clone exhibits a band in the appropriate VP amplification and an accumulated T cell clone in a heterogeneous lymphocyte population is identified as a band in the background smear pattern. If a lymphocyte population was stimulated by an antigen either in vitro or in vivo, several distinct bands were found to be generated in the background smear. Thus, the dynamic changes of T cell clonal responses could be monitored with this method. Analyses of several immunological disorders, including autoimmune diseases, malignant disorders, and transplantations, revealed the involvement of antigen-specific T cell immune responses in these disorders. Furthermore, taking advantage of the reproducible mobility of a band of SSCP gel, we are now able to compare identities of the accumulated T cell clones in different samples without the need for nucleotide sequencing of each clone. Such information can thus elucidate the occurrence of uniform or stable immunological reactions in the host and also suggests that these reactions play an important role in vivo. Therefore, taken together, the above described novel T cell clonality analysis is considered to be useful in studying the T cell immune responses in various fields of immunology.

Autoimmune Diseases↗
PubMed1983

Clonal analysis of T cell unresponsiveness to alloantigens induced by neonatal injection of F1 spleen cells into parental mice.

Alloantigen-specific T cell precursors present in spleen cell populations obtained from parental mice that were rendered tolerant by the neonatal injection of F1 spleen cells were quantitated by clonal analysis, i.e., limiting dilution techniques. The results demonstrate that the frequencies for cytolytic as well as TCGF-producing (helper) and MAF-producing T cell precursors were dramatically reduced in tolerant spleen cell populations. The unresponsive state was antigen specific and could not be transferred in vitro in cell mixing experiments. Therefore, clonal deletion (or inactivation) would appear to represent a major mechanism for the observed tolerant state in this model system.

Animals↗