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Biomedical subjects

S Ramachandra

Publications and source records attributed to S Ramachandra.

At least 19 recordsLinked to original sources
PubMed1999

A rapid automated SSCP multiplex capillary electrophoresis protocol that detects the two common mutations implicated in hereditary hemochromatosis (HH).

Currently two mutations in the HFE gene are known to be associated with the manifestation of the autosomal recessive disorder hereditary hemochromatosis (HH). A single-base mutation resulting in Cys282Tyr appears to have a causative role in the development of the disease, and a point mutation resulting in His63Asp may also be involved. Recent observations with a fully automated capillary electrophoresis (CE) system (ABI Prism 310) suggested that this instrument could be used for the precise identification of known mutations based on single-strand conformation polymorphism (SSCP). Two DNA fragments, each specific for one of the HFE mutation sites and labeled with a different fluorophor, were coamplified and without further manipulation simultaneously analyzed by CE-SSCP. Wild-type samples showed a mobility pattern that was clearly distinguishable from homozygous Cys282Tyr, homozygous His63Asp, or a compound heterozygous sample. To evaluate the reliability of this system for the detection of both mutations, 20 samples were analyzed blind. All genotypes, which were called automatically, were in concordance with those obtained by a previously validated restriction fragment length polymorphism method. Thus, SSCP in combination with CE provides a fast and precise research tool for the simultaneous identification of the two common mutations implicated in HH.

Electrophoresis, Capillary↗
PubMed1998

Identification of known p53 point mutations by capillary electrophoresis using unique mobility profiles in a blinded study.

This study is part of an ongoing project at the National Institute of Standards and Technology (NIST) that generates a panel of DNA clones containing the most common mutations found in the human p53 tumor suppressor gene. This panel will be made available as a reference source for evaluation and testing for p53 mutations. Single strand conformation polymorphism (SSCP) analysis has found widespread acceptance as a tool for simply and rapidly screening for mutations, albeit with a detection rate that can be below 100%. We have begun to analyze mutations found in exon 7 of the p53 gene by SSCP using laser induced fluorescence capillary electrophoresis (LIF-CE). PCR fragments, containing single point mutations, were amplified from genomic DNA isolated from cell lines using primers labeled with two different fluorophores. This dual labeling approach allowed better traceability of mobility shifts as a function of the experimental conditions. While analyzing the clones H596, Colo320, Namalwa and wild type (reference samples) at different temperatures, ranging from 25 to 45 degrees C, it was observed that each mutation responded in a unique way to changes in temperature both in magnitude and direction of shifts relative to the wild type sample. In a blinded study, ten p53 exon 7 samples were matched automatically, using ABI PRISM Genotyper software, against the four reference samples. From these 10 samples, six were correctly identified as containing one of the reference mutations, two corresponded to wild type, and two were correctly identified as non-reference mutations. This approach should prove helpful in the rapid screening of target sequences that are known to bear a limited number of mutations.

Automation↗
PubMed1997

Video-assisted thoracoscopic closure of postpneumonectomy bronchopleural fistulas.

Despite improved surgical techniques and advances in medical-surgical treatment, postpneumonectomy bronchopleural fistulas remain an important cause of morbidity and a therapeutic challenge. Video-assisted thoracoscopic closure of these fistulas reinforced by transposition of bulky chest wall muscles or omentum to obliterate the residual space may lessen risks and complications of repeated thoracotomy in these often frail, debilitated, and compromised patients. We report our initial experience with video-assisted thoracoscopic debridement of the empyema cavity and closure of the postpneumonectomy bronchopleural fistula by transposing an entire pectoralis major muscle in one patient and by transposing a pedicled omentum in another patient.

Aged↗
PubMed1996

Requirement for increased IL-10 in the development of B-1 lymphoproliferative disease in a murine model of CLL.

Malignant B-1 cells derived from NZB mice, a murine model of spontaneous autoimmunity and B cell lymphoproliferative disease, produce significantly higher levels of IL-10 mRNA than normal B-1 or B cells. IL-10 may act as an autocrine growth factor for the expansion of malignant B-1 cells. In order to determine if elevated endogenous production of IL-10 was a required element for the malignant transformation of B-1 cells in NZB mice, backcross animals were studied for the linkage between elevated IL-10 expression and the presence of lymphoid malignancy. The phenotypes of aged (NZB x DBA/2)F1 x NZB animals were determined and a strong correlation was found between the elevated levels of IL-10 mRNA and the development of B-1 malignant clones. In contrast, an increased level of IL-10 message was not associated with elevated serum IgM or the presence of anemia or reticulocytosis which is mainly seen in response to autoantibody production. These results indicate that, at least in NZB, the autoimmunity and lymphoproliferation phenotypes are not linked genetically. IL-10 may enhance proliferation and the development of B-1 cell malignancy rather than antibody production by the B-1 cell subpopulation. Thus, IL-10 plays an important role in B-1 malignancies, and downregulation of IL-10 could be a likely site for intervention in B cell malignancies.

Animals↗
PubMed1996

Mapping human telomere regions with YAC and P1 clones: chromosome-specific markers for 27 telomeres including 149 STSs and 24 polymorphisms for 14 proterminal regions.

A YAC library enriched for telomere clones was constructed and screened for the human telomere-specific repeat sequence (TTAGGG). Altogether 196 TYAC library clones were studied: 189 new TYAC clones were isolated, 149 STSs were developed for 132 different TY-ACs, and 39 P1 clones were identified using 19 STSs from 16 of the TYACs. A combination of mapping methods including fluorescence in situ hybridization, somatic cell hybrid panels, clamped homogeneous electric fields, meiotic linkage, and BLASTN sequence analysis was utilized to characterize the resource. Forty-five of the TYACs map to 31 specific telomere regions. Twenty-four linkage markers were developed and mapped within 14 proterminal regions (12 telomeres and 2 terminal bands). The polymorphic markers include 12 microsatellites for 10 telomeres (1q, 2p, 6q, 7q, 10p, 10q, 13q, 14q, 18p, 22q) and the terminal bands of 11q and 12p. Twelve RFLP markers were identified and meiotically mapped to the telomeres of 2q, 7q, 8p, and 14q. Chromosome-specific STSs for 27 telomeres were identified from the 196 TYACs. More than 30,000 nucleotides derived from the TYAC vector-insert junction regions or from regions flanking TYAC microsatellites were compared to reported sequences using BLASTN. In addition to identifying homology with previously reported telomere sequences and human repeat elements, gene sequences and a number of ESTs were found to be highly homologous to the TYAC sequences. These genes include human coagulation factor V (F5), Weel protein tyrosine kinase (WEE1), neurotropic protein tyrosine kinase type 2 (NTRE2), glutathione S-transferase (GST1), and beta tubulin (TUBB). The TYAC/P1 resource, derivative STSs, and polymorphisms constitute an enabling resource to further studies of telomere structure and function and a means for physical and genetic map integration and closure.

Animals↗
PubMed1996

A comparative immunohistochemical study of mammary and extramammary Paget's disease and superficial spreading melanoma, with particular emphasis on melanocytic markers.

A comparative immunohistochemical study was performed on Paget's disease of the nipple (PDN), extramammary Paget's disease (EMPD) and cutaneous superficial spreading melanoma (SSM) using antibodies to S100, NK1-C3 and HMB45, cytokeratin (CAM 5.2) and c-erb B2 oncoprotein (21N). Conventional histochemical stains for intracytoplasmic mucin and melanin were also done. Of the 20 cases of PDN, positivity was seen in 12 with S100, 16 with NK1-C3, none with HMB45, 20 with CAM 5.2 and 19 with 21N. All 5 cases of EMPD were CAM 5.2 positive and HMB45, S100 and 21N negative. Three EMPD were NK1-C3 positive. All 10 cases of SSM were S100, NK1-C3 and HMB45 positive and all were CAM5.2 and 21N negative. Mucin was demonstrable in 11 cases of PDN and all of EMPD but none of SSM. Melanin was seen in 2 PDN, 3 EMPD and all SSM cases. Identification of mucin and melanin, therefore, proved an unreliable means of distinguishing these diseases. Immunohistochemistry for cytokeratin and HMB45 appear to be the most specific markers in differentiating Paget's disease and SSM. Antibodies to c-erb B2 may also be valuable in this situation.

Biomarkers↗
PubMed1995

Inflammatory pseudotumour of soft tissues: a clinicopathological and immunohistochemical analysis of 18 cases.

Pseudosarcomatous proliferation of myofibroblasts, fibroblasts and inflammatory cells have been reported in a wide variety of sites and are most commonly termed inflammatory pseudotumours. Despite increasing recognition, the occurrence of inflammatory pseudotumour in somatic soft tissue is poorly documented, such that these lesions are commonly mistaken for a variety of benign and malignant neoplasms. This report documents our experience of 18 cases of soft tissue inflammatory pseudotumour. All arose in adults (median age 54.5 years; range 28-83 years) and showed no sex predilection. Anatomical location was varied but the head and neck region and abdominal cavity were most common. They displayed a spectrum of histological appearances but all were characterized, to a greater or lesser extent, by an admixture of myofibroblasts and fibroblasts, most commonly arranged in short interwoven fascicles, together with a polymorphic inflammatory cell component, consisting principally of lymphocytes and plasma cells. In some cases xanthoma cells were prominent. Simple surgical excision appeared to be curative. The importance of recognizing the presence of a wide clinicopathological spectrum is emphasized and the differential diagnosis, especially from fibrohistiocytic neoplasms or Hodgkin's disease, is discussed.

Adult↗
PubMed1994

Videothoracoscopic wedge resection for peripheral pulmonary nodules.

BACKGROUND: This study was done to evaluate the use of the endoscopic multifire linear stapler for videothoracoscopic wedge resection (VTWR) of peripheral pulmonary nodules and to define the indications, advantages, and drawbacks of this minimally invasive technique. STUDY DESIGN: A case study review of 57 consecutive video-assisted thoracic operations for wedge resection of peripheral pulmonary nodules performed upon 55 patients admitted to a community hospital from June 1991 through July 1993 is presented. RESULTS: There were 44 malignant and 13 benign lesions. Of the malignant peripheral pulmonary nodules (PPN), there were 19 adenocarcinomas, ten squamous cell carcinomas, two undifferentiated large-cell carcinomas, three bronchoalveolar carcinomas, two carcinoid tumors, one neuroendocrine tumor, and seven metastatic carcinomas. The benign nodules included five hamartomas, two granulomas, one aspergilloma, one nodular amyloidosis, one Wegener's granulomatosis, one focal pulmonary infarct, and two interstitial fibroses. Videothoracoscopic wedge resection alone was performed upon 37 patients, 17 of whom had primary carcinoma of the lung; seven had metastatic lesions, and the remainder had benign disease. Of the 17 patients with primary carcinoma of the lung who had VTWR alone, eight patients had marked impairment of pulmonary function, six had significant co-morbid disease, two had peripheral carcinoid tumors, and one had bilateral metachronous carcinomas. Videothoracoscopic wedge resections with concomitant lobectomies were performed upon 20 patients with primary carcinoma of the lung, including one patient with bilateral synchronous carcinomas. Five of the patients with nodules ranging from 2 to 3 cm in diameter were found to have metastasis to regional nodes. None of the patients who had lobectomies for peripheral carcinomas less than 2 cm in diameter had regional nodal metastases. There was no perioperative mortality and no significant morbidity. CONCLUSIONS: Videothoracoscopic wedge resection is a useful alternative to traditional transthoracic resection for suspicious, undiagnosed PPN, for low grade malignant neoplasms, such as carcinoid, for peripheral metastatic lesions, for bilateral synchronous or metachronous tumors, for the occasional clinically localized peripheral small-cell carcinoma as a surgical adjunct to chemotherapy, and for small, peripheral, T-1, N-0, M-0 bronchial carcinomas in compromised patients at high risk with marginal pulmonary reserve. The procedure is effective, minimally invasive, and has potential advantages over conventional thoracotomy, including less postoperative pain and morbidity, shorter hospitalization period and convalescence, and an earlier return to work and normal activities.

Humans↗
PubMed1994

Laparoscopic resection of a giant mesenteric cyst.

Laparoscopic resection of a giant retroperitoneal, mesenteric cyst in a 38-year-old man who presented with abdominal distension and pedal edema is described. The diagnosis was made by abdominal computed tomography, which revealed a large cystic mass that was causing extrinsic compression of the inferior vena cava and the right ureter. The hospital course was uneventful, and the patient was discharged 2 days postoperatively. This new endoscopic approach offers a useful alternative to the traditional transabdominal excision of a mesenteric cyst and may have some theoretical advantages, including less postoperative pain and shorter convalescence.

Adult↗
PubMed1993

Laparoscopic resection of a splenic artery aneurysm.

Laparoscopic resection of a 2.5 cm splenic artery aneurysm in a 64-year-old woman is presented. The diagnosis was suspected on plain roentgenograms of the lumbosacral spine and confirmed by abdominal CT scan and selective splenic arteriography. The hospital course was uneventful and she was discharged 2 days postoperatively. This new endoscopic approach offers a useful alternative to the surgical management of splenic artery aneurysms, and may have some theoretical advantages, including less postoperative pain and shorter convalescence.

Aneurysm↗
PubMed1993

Laparoscopically assisted resection of intussuscepted Meckel's diverticulum.

We report a case of a Meckel's diverticulum with pancreatic heterotopia that had intussuscepted into the distal ileum, masquerading as a small bowel tumor and presenting with acute lower gastrointestinal hemorrhage. Laparoscopically assisted resection of the diverticulum was successfully accomplished.

Adult↗
PubMed1992

Regulatory aspects of clonally expanded B-1 (CD5+ B) cells.

B-1 (CD5+ B) cells appear early in ontogeny, produce mainly unmutated polyreactive antibodies, and are capable of self-renewal. B-1 cells clonally expand with age and are the malignant cell in chronic lymphocytic leukemia. In this report immunological analysis of B-1 malignancies in NZB mice, a murine model of chronic lymphocytic leukemia, is related to current information on B-1 cells. B-1 clones from NZB mice produce high levels of interleukin-10, detected at the RNA level by semi-quantitative polymerase chain reaction. In addition, the B-1 malignant clones in NZB mice and their hybrids, are negative for B220/6B2 expression, the B-specific antigenic form of CD45 which is a membrane-associated phosphatase involved in lymphocyte activation. Both the autocrine production by B-1 cells of interleukin-10 and altered CD45 expression may be responsible for the clonal expansion of these cells, as well as the accompanying T cell expansion. We report the establishment of an in vitro cytotoxic CD8+ T cell line derived from an NZB with a B-1 malignancy. The effect of B-1 cell-derived interleukin-10 on subsets of T lymphocytes may account for the immunoregulatory properties of B-1 cells. In addition, the NZB malignancies were also characterized for immunoglobulin variable region sequence and antigen specificity. The B-1 malignancies produced immunoglobulin derived from unmutated germline sequences with no N base substitutions. It appears that both the immunoglobulin and interleukin-10 produced by the B-1 malignant cell in NZB mice may have immunoregulatory properties. A study of B-1 malignancies may shed light on the immunoregulatory properties of non-clonally expanded normal B-1 cells.

Animals↗
PubMed1992

Closure of a genetic linkage map of human chromosome 7q with centromere and telomere polymorphisms.

We have constructed a 2.4-cM resolution genetic linkage map for chromosome 7q that is bounded by centromere and telomere polymorphisms and contains 66 loci (88 polymorphic systems), 38 of which are uniquely placed with odds for order of at least 1000:1. Ten genes are included in the map and 11 markers have heterozygosities of at least 70%. This map is the first to incorporate several highly informative markers derived from a telomere YAC clone HTY146 (locus D7S427), including HTY146c3 (HET 92%). The telomere locus markers span at least 200 kb of the 7q terminus and no crossovers within the physical confines of the locus were observed in approximately 240 jointly informative meioses. The sex-equal map length is 158 cM and the largest genetic interval between uniquely localized markers in this map is 11 cM. The female and male map lengths are 181 and 133 cM, respectively. The map is based on the CEPH reference pedigrees and includes over 4000 new genotypes, our previously reported data plus 29 allele systems from the published CEPH version 5 database, and was constructed using the program package CRI-MAP. This genetic linkage map can be considered a baseline map for 7q, and will be useful for defining the extent of chromosome deletions previously reported for breast and prostate cancers, for developing additional genetic maps such as index marker and 1-cM maps, and ultimately for developing a fully integrated genetic and physical map for this chromosome.

Centromere↗
PubMed1992

Peripheral nerve sheath differentiation in malignant soft tissue tumours: an ultrastructural and immunohistochemical study.

Thirteen soft tissue sarcomas with ultrastructural evidence of nerve sheath differentiation were investigated by immunohistochemistry. Three arose in a major nerve or nerve trunk and four patients had von Recklinghausen's neurofibromatosis. Ultrastructurally, 10 cases showed variable differentiation towards Schwann cells, two resembled perineurial cells and one tumour had features suggestive of both cell types. Immunostaining for S-100 protein was positive in eight Schwann cell tumours, negative in the other two Schwann cell tumours and negative in those with perineurial-like cells. No cases demonstrated epithelial membrane antigen, so that the existence of perineurial cells in malignant nerve sheath tumours remains immunohistochemically unsubstantiated; it may be that the perineurial-like cells are merely incompletely differentiated Schwann cells, with nerve sheath differentiation manifesting a continuous spectrum. Leu 7 was detected in four Schwann cell tumours, three of which were ultrastructurally well differentiated. Cytokeratin and desmin were demonstrated only in an undifferentiated pleomorphic area of one Schwann cell tumour. Electronmicroscopy can aid diagnosis by revealing nerve sheath differentiation in malignant soft tissue tumours without demonstrable S-100 protein.

Adolescent↗
PubMed1990

Immunohistochemical distribution of c-erbB-2 in in situ breast carcinoma--a detailed morphological analysis.

An immunohistochemical study of c-erbB-2 expression was carried out on in situ (non-invasive) breast carcinoma, using antibody 21N, raised to the intracytoplasmic domain of the c-erbB-2 oncogene product. Strong membrane staining was observed in 44 out of 74 (59 per cent) cases of ductal carcinoma in situ (DCIS), but none of 48 lobular carcinoma in situ (LCIS) lesions. A detailed comparative morphological evaluation using several different parameters, including histological subtypes, was performed within the DCIS group. The results showed that there was a significant correlation between c-erbB-2 expression and the presence of large cell size, periductal lymphoid cell infiltration, marked nuclear pleomorphism, multinucleation, and a high mitotic rate. Of these, cell size appears to be the most important predictor of c-erbB-2 status, followed by the presence of periductal lymphoid cell infiltration. These results indicate, firstly, that LCIS and DCIS are biologically (as well as histologically) different and, secondly, that a subgroup of DCIS, which is associated with c-erbB-2 over-expression, exists and appears to have distinct histological features. The subgroup of DCIS cases which over-express c-erbB-2 may be a biologically definable category with prognostic importance. These results may therefore have relevance to breast screening programmes, but a larger study incorporating clinical data would be necessary to correlate these findings with clinical outcome.

Analysis of Variance↗
PubMed1990

Endometrial stromal sarcomas.

The case histories of 15 patients with endometrial stromal sarcoma attending the Royal Marsden Hospital between 1973 and 1987 were reviewed. Eight patients were classified as having low-grade and seven as high-grade tumors using specific histological criteria. Those patients with low-grade tumors had a long relapse-free and overall survival in the absence of receiving specific therapy after initial surgery. Five are alive and disease free, one has shown a partial response to hormonal manipulation, and one is alive with disease on no therapy. Only one patient has died and this occurred 111 months postdiagnosis. Conversely, all seven patients with high-grade tumors relapsed with local disease and three have died. Thus, after surgical resection patients with low-grade tumors probably do not require treatment until relapse, whereas adjuvant radiotherapy should be given to those patients with high-grade tumors. On relapse, hormone therapy should be the treatment of first choice for patients with low-grade tumors, whereas chemotherapy is more appropriate for patients with high-grade tumors.

Adult↗