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D Shibata

Publications and source records attributed to D Shibata.

At least 109 records · Page 6Linked to original sources
PubMed1994

Anatomical distribution of HTLV-I proviral sequence in an autopsy case of HTLV-I associated myelopathy: a polymerase chain reaction study.

HTLV-I associated myelopathy (HAM) is a slowly progressive paraplegia of the lower extremities observed among HTLV-I carriers. An autopsy of a typical HAM case in which perivascular lymphocytic infiltration was not limited to the central nervous system was examined. Spinal dorsal roots, salivary gland, lungs, liver and kidney showed non-specific, but unusual sporadic perivascular lymphocytic infiltration, which resembled the findings in the spinal cord. To investigate the anatomical distribution of HTLV-I provirus, the HTLV-I proviral sequences, tax and pol, were amplified from the formalin-fixed paraffin-embedded tissues of the autopsy case using polymerase chain reaction (PCR). By PCR, strong HTLV-I provirus signals were detected in the spinal cord, peripheral nerve, muscle, lungs and liver. Weak signals were detected in the medulla oblongata, optic nerve and lymph node, while the other organs, including the cerebrum, were negative. The data from this study show the specific distribution of HTLV-I provirus in the distinct organs of a HAM patient.

Aged↗
PubMed1994

Patterns of p53 mutations in squamous cell carcinoma of the lung. Acquisition at a relatively early age.

Squamous cell carcinoma (SQCC) of the lung is thought to arise after the accumulation of multiple mutations, including p53. To better characterize when p53 mutations are acquired, 37 SQCC of the lung were examined by polymerase chain reaction and single-strand conformation polymorphism analysis. Somatic p53 mutations were detected in nine tumors (24.3%). There were no significant differences in the stage, sex, or race between patients with or without p53 mutations. However, the patients with SQCC and p53 mutations were significantly (P = 0.0006) younger (mean age, 54.3 years) compared with patients without p53 mutations (mean age, 65). The topographical tissue distributions of the p53 mutations were examined by selective ultraviolet radiation fractionation. In all nine cases, the specific p53 mutant alleles were homogeneously present throughout the primary tumors, in all three examples with in situ carcinoma, and in all four cases with metastases. In one case, squamous metaplasia contiguous with the primary tumor also contained the same p53 mutation. Normal or hyperplastic and metaplastic or dysplastic epithelium not contiguous with the primary tumors lacked the specific p53 mutations. These findings suggest that p53 mutations are commonly acquired at a relatively early age, before the bulk of clonal expansion, and usually persist throughout the progression of SQCC of the lung.

Aged↗
PubMed1994

Further evidence that one of the earliest alterations in colorectal carcinogenesis involves APC.

Colorectal cancer is hypothesized to arise after the accumulation of multiple mutations in critical oncogenes or tumor suppressor genes. The relative timing of each mutation is unknown because the exact number and types of mutations differ between tumors. However, for every mutation except the first, tumor heterogeneity must exist until clonal dominance is reestablished. This principle was applied to mutant APC genes in eight colorectal adenomas. The APC mutations were homogeneously present throughout the adenomas, including those less than 1 cm in size, but absent from the normal polyp stalks. In one adenoma with APC and c-K-ras mutations, both mutations were simultaneously present in only a small discrete portion, suggesting that the c-K-ras mutation was acquired after the APC mutation. These findings suggest that when mutations in APC occur, they are usually one of the first events in colorectal carcinogenesis or provide such a strong selective advantage that intratumor heterogeneity is seldom observed.

Adenoma↗
PubMed1994

c-k-ras and p53 mutations occur very early in adenocarcinoma of the lung.

The topographical distribution of a mutation provides insight into past patterns of tumor evolution. This approach was applied to two loci commonly mutated in adenocarcinoma of the lung--p53 and c-K-ras. In 41 primary adenocarcinomas, c-K-ras codon 12 point mutations were detected in 8 (19.5%) tumors and p53 point mutations were detected in 10 (24.4%) tumors, with one tumor harboring both mutations. These mutations were only detected in malignant cells and with a homogeneous topographical distribution throughout 16 tumors, including metastasis. Intratumor heterogeneity was detected in only one tumor in which a small portion lacked the specific p53 mutation. Based on this topographical analysis, it is likely that when these mutations occur in adenocarcinoma of the lung, they are usually acquired during the very earliest phases of tumor formation before the bulk of clonal expansion, and in very small precursor lesions.

Adenocarcinoma↗
PubMed1993

Genetic heterogeneity of the c-K-ras locus in colorectal adenomas but not in adenocarcinomas.

BACKGROUND: Previous molecular genetics studies of colorectal cancer have identified multiple mutations in the c-K-ras gene (also known as KRAS2) in all phases of its development. Because of technical difficulty, prior studies rarely focused attention on the detailed distribution of c-K-ras mutations in multiple regions of the same primary tumor specimen. However, with recent development of the selective UV radiation fractionation method, characterization of c-K-ras mutations in multiple regions of the same primary tumor specimen can be performed. PURPOSE: Our purpose was to describe how c-K-ras mutations were distributed among cells obtained from multiple regions of the same primary tumor in an attempt to describe differences between early and late colorectal carcinogenesis. METHODS: Formalin-fixed, paraffin-embedded tissue blocks were obtained. Seven adenocarcinomas and seven adenomas were selected for the presence of mutant c-K-ras genes and histologic transitions between normal and neoplastic tissue. Tissue sections were prepared for analysis by the selective UV radiation fractionation method by placing thin, fixed tissue sections on a plastic slide with no coverslip. Under the microscope, small ink dots from a felt-tip pen were manually placed directly on relatively pure cell subpopulations. The slides were placed with the tissue side exposed to a UV transilluminator for 2-4 hours to inactivate the DNA present in the unprotected ("undotted") cells. Individual dots were cut out of the plastic slide into 2 x 2-mm squares and placed into microfuge tubes. The DNA was extracted and supernatant used for polymerase chain reaction (PCR) analysis. Mutations at c-K-ras codons 12 and 13 were detected. RESULTS: The selective UV radiation fractionation method and PCR analysis revealed that c-K-ras mutations never extended into normal mucosa and were present in all neoplastic cells regardless of phenotypes in all seven adenocarcinomas and three of the seven adenomas. Further examination of two carcinomas for p53 (also known as TP53) mutations or loss of heterozygosity demonstrated that these additional mutations were also present in all tumor cells, suggesting that a single transformed clone was responsible for the majority of growth. However, in four other adenomas, tumor heterogeneity was demonstrated, since c-K-ras mutations were detected only in discrete portions. CONCLUSIONS: Adenoma formation may include a stage in which multiple and genetically distinct neoplastic clones are present, while most carcinomas appear to have a homogeneous composition that may result from the successful progression of one of these clones.

Adenocarcinoma↗
PubMed1993

Ubiquitous somatic mutations in simple repeated sequences reveal a new mechanism for colonic carcinogenesis.

Spontaneous errors in DNA replication have been suggested to play a significant role in neoplastic transformation and to explain the chromosomal alterations seen in cancer cells. A defective replication factor could increase the mutation rate in clonal variants arising during tumour progression, but despite intensive efforts, increases in tumour cell mutation rates have not been unambiguously shown. Here we use an unbiased genomic fingerprinting technique to show that 12 per cent of colorectal carcinomas carry somatic deletions in poly(dA.dT) sequences and other simple repeats. We estimate that cells from these tumours can carry more than 100,000 such mutations. Only tumours with affected poly(dA.dT) sequences carry mutations in the other simple repeats examined, and such mutations can be found in all neoplastic regions of multiple tumours from the same patient, including adenomas. Tumours with these mutations show distinctive genotypic and phenotypic features. We conclude that these mutations reflect a previously undescribed form of carcinogenesis in the colon (predisposition to which may be inherited) mediated by a mutation in a DNA replication factor resulting in reduced fidelity for replication or repair (a 'mutator mutation').

Adult↗
PubMed1993

Epstein-Barr virus-associated non-Hodgkin's lymphoma in patients infected with the human immunodeficiency virus.

Lymphoproliferations associated with Epstein-Barr virus (EBV) commonly arise in settings of immune dysfunction, including human immunodeficiency virus (HIV) infection. In this study, EBV was associated with 39 of 59 (66%) HIV-related systemic lymphomas. Unlike the lymphoproliferations that arise in the setting of transplantation, the HIV-related lymphomas were monoclonal, as evaluated by Ig heavy chain rearrangements and EBV termini analysis, and associated (40%) with c-MYC rearrangements. Furthermore, analysis of multiple lymphoma tissues from one autopsy showed evidence that a single lymphoma clone was responsible for dissemination. The latent EBV nuclear antigen (EBNA-1) transcripts detected in the HIV-related lymphomas were characteristic of the pattern found in Burkitt lymphoma (g1 EBNA1) and not in transplant-related lymphoproliferations. However, unlike Burkitt lymphoma, EBV latent membrane-associated protein (LMP) transcripts were also detected, thereby constituting an EBV expression pattern (g1 EBNA1+, LMP+) not previously observed in B-cell lymphomas. These findings demonstrate a high frequency of EBV-associated lymphomas in the setting of HIV infection that are distinct from the lymphoproliferations that arise during iatrogenic transplant-associated immuno-suppression or in the general population. However, it is also apparent that HIV-related lymphomas are biologically heterogeneous, which may reflect the multiple mechanisms or steps necessary for eventual malignant transformation.

Antigens, Viral↗
PubMed1993

Purification and partial sequences of Aralia cordata cinnamyl alcohol dehydrogenase.

Cinnamyl alcohol dehydrogenase (CAD) (EC 1.1.1.195) from a dicot, Aralia cordata, was purified to homogeneity and its properties were characterized. The enzyme shows a preference for cinnamyl alcohols and cinnamyl aldehydes as substrates. The M(r) is estimated at 72,000. The enzyme is composed of two heterogeneous subunits of slightly different sizes, and it differs from the bean enzyme in the size of subunits. Partial amino acid sequencing of the purified enzyme was carried out both from the N-terminus and using selected peptides obtained by cyanogen bromide cleavage.

Alcohol Oxidoreductases↗
PubMed1993

Identification of mismatched fixed specimens with a commercially available kit based on the polymerase chain reaction.

Specimen mix-ups inevitably occur and have the potential for great harm. The ability to investigate mix-ups objectively and assign fixed tissues to patients correctly is unfortunately limited, as most such assays require fresh specimens. A commercial kit based on the polymerase chain reaction (PCR) can be applied to the molecular genetic analysis of fixed tissues. This kit, which can amplify and distinguish 21 different genotypes at a polymorphic human leukocyte antigen (HLA) locus, was applied to investigate 16 cases of potential specimen mismatches. The majority of tissues were small and essentially irreplaceable biopsy specimens, and four cases involved minute fragments of potential "floaters." Data were successfully obtained from all 16 cases despite the collection from several different hospitals and the small quantities of tissue. The assay required approximately 2 days for completion; therefore, data were returned within a clinically useful time period. This study provided evidence that molecular genetic assays based on the PCR can be applied to routinely obtained fixed-tissue specimens to investigate potential mismatches.

DNA Fingerprinting↗
PubMed1992

Epidemiological and biological study of acquired immunodeficiency syndrome-related lymphoma in the County of Los Angeles: preliminary results.

A population-based case control study of intermediate- and high-grade lymphoma in the County of Los Angeles, CA, was initiated in 1989. Human immunodeficiency virus (HIV)-positive lymphoma patients are compared to HIV-negative lymphoma patients, to HIV-positive controls with acquired immunodeficiency syndrome but without lymphoma, and to HIV-positive asymptomatic individuals. The HIV-negative lymphoma cases are compared to neighborhood controls, who are matched in terms of age, sex, race/ethnicity, and socioeconomic status. All cases are reviewed for pathology by a single group of pathologists. All cases and controls are studied for HIV, Epstein-Barr virus (EBV), and human herpesvirus 6 antigens and antibodies. Tissues from HIV-positive and -negative cases are studied for immunoglobulin gene rearrangement, presence of EBV and HIV, c-myc oncogene rearrangements, and karyotypic analysis. To date, with 294 lymphoma cases and 181 control cases interviewed, high-grade lymphoma has been diagnosed in 82% of the HIV-positive cases versus 40% of the HIV-negative cases (P = 0.001). Although elevated titers of EBV-viral capsid antigen were demonstrated in 82% of HIV-positive versus 50% of HIV-negative lymphoma cases, the geometric mean titer of EBV-viral capsid antigen is similar among HIV-positive lymphoma cases and HIV-positive controls. The geometric mean titer of human herpesvirus 6 antibodies was similar in HIV-positive and HIV-negative lymphoma cases and in the control populations. Monoclonality was demonstrated in all cases of lymphoma. EBV genome was demonstrated within lymphoma DNA in 68% of HIV-positive and 15% of HIV-negative lymphoma cases. Further study will be required to elucidate the full mechanisms of pathogenesis of the acquired immunodeficiency syndrome-related lymphomas.

Adolescent↗
PubMed1992

A pattern of accumulation of a somatic deletion of mitochondrial DNA in aging human tissues.

An assay that selectively amplifies a specific deletion of the mitochondrial genome has been used to study the extent of the deletion's accumulation in a variety of human tissues. The deletion occurs at much higher levels in nervous and muscle tissues than in all other tissues studied. The variation in deletion level between the same tissues in different persons of similar age appears to be less than the variation among tissues within an individual. Tests for artifactual explanations of the level differences were each negative. Three cellular parameters that are correlated with the level of the deletion are identified. The preferential accumulation of deleterious mitochondrial mutations in a restricted subset of aging human tissues may compound deficiencies of function in those tissues that accrue with age.

Adolescent↗
PubMed1992

cDNA cloning of rice lipoxygenase L-2 and characterization using an active enzyme expressed from the cDNA in Escherichia coli.

A full-length cDNA of rice lipoxygenase L-2 was cloned from 3-day-old seedlings. The identity of the clone was determined by amino acid sequencing of selected peptides of the purified enzyme and immunological characterization of an active enzyme that was produced from the cDNA in Escherichia coli by cultivation at 15 degrees C. The nucleotide sequence showed a strong bias toward G and C in the selection of nucleotides, especially at the third position of the codons (93% G/C). The complete amino acid sequence of the enzyme was deduced from the nucleotide sequence. The molecular mass of the enzyme was calculated to be 96,657 Da based on 865 amino acids. The amino acid sequence shares similarity with those of dicot lipoxygenases throughout the enzyme at a level of 50%. A hydropathy profile calculated from the amino acid sequence resembled those of dicot lipoxygenases, suggesting conservation of the secondary structure of these enzymes. The active enzyme, expressed in Escherichia coli, was characterized for pH dependence of the enzyme activity, intramolecular specificity, heat stability and Km. The enzyme had the same properties as the L-2 enzyme that was isolated from seedlings, but differed from the lipoxygenase L-3 isolated from mature plants.

Amino Acid Sequence↗
PubMed1992

Detection and localization of Epstein-Barr viral genomes in angioimmunoblastic lymphadenopathy and angioimmunoblastic lymphadenopathy-like lymphoma.

We studied 23 cases of angioimmunoblastic lymphadenopathy (AILD) and AILD-like lymphoma for evidence of Epstein-Barr virus (EBV) using the polymerase chain reaction (PCR) and in situ hybridization studies. EBV nucleic acid sequences were found by either PCR or in situ hybridization in 96% of the cases. There was a wide range in the number of EBV-positive cells among the different cases as detected by in situ hybridization. The EBV-positive cells most often possessed nuclei of intermediate to large size. Double-labeling immunohistochemistry/in situ hybridization studies demonstrated that most of the EBV-positive cells expressed the B-lineage antigen CD20 (as detected by L26), with a minority of the EBV-positive cells stained for the T-lineage associated antigen, CD43 (as detected by Leu 22). The abnormally high amounts of EBV found in AILD and AILD-like lymphoma may be a reflection of decreased immunocompetence in these patients. The presence of EBV-positive B cells may explain the presence of B-cell clones found by others as well as the paradoxical occurrence of B-cell lymphoma in a primary T-cell lymphoproliferative disorder.

Adult↗
PubMed1992

cDNA sequence and expression of a phosphoenolpyruvate carboxylase gene from soybean.

A full-length cDNA encoding a subunit of phosphoenolpyruvate carboxylase (PEPC) was isolated from a developing seed expression library of the C3 plant Glycine max. The corresponding mRNA is present at similar levels in leaf, stem, root and developing seed. Two potential start codons exist, and the activity of protein initiated from the first such codon could be subject to regulation by protein kinase. Sequence comparison shows a similar upstream start codon in the case of the Ppc2 gene from Mesembryanthemum crystallinum, previously assumed to lack the sequences necessary for phosphorylation. The soybean encoded protein tends to resemble other 'C3-type' PEPC proteins more closely than those implicated in C4 or crassulacean acid metabolism.

Amino Acid Sequence↗
PubMed1992

Appearance of new lipoxygenases in soybean cotyledons after germination and evidence for expression of a major new lipoxygenase gene.

The appearance and subsequent disappearance of lipoxygenase activity at pH 6.8 in germinated cotyledons of soybean (Glycine max [L.]) was shown using a variant soybean cultivar (Kanto 101) that lacks the two lipoxygenase isozymes, L-2 and L-3, that are present in dry seeds of a normal soybean cultivar (Enrei). Three new lipoxygenases, designated lipoxygenase L-4, L-5, and L-6, were purified using anionic or cationic ion exchange chromatography. The major lipoxygenase in 5-day-old cotyledons of the variant soybean was lipoxygenase L-4. Lipoxygenases L-5 and L-6 preferentially produced 13(S)-hydroperoxy-9(Z), 11(E)-octadecadienoic acid (13S-HPOD) as a reaction product of linoleic acid, whereas lipoxygenase L-4 produced both 13S-HPOD and 9(S)-hydroperoxy-10(E), 12(Z)-octadecadienoic acid. All three isozymes have pH optima of 6.5, no activity at pH 9.0, and preferred linolenic acid to linoleic acid as a substrate. Partial amino acid sequencing of lipoxygenase L-4 showed that this isozyme shares amino acid sequence homology with lipoxygenases L-1, L-2, and L-3 but is not identical to any of them. This indicates that a new lipoxygenase, L-4, is expressed in cotyledons.

Journal Article↗