Search PubMed⌕ Search

Biomedical subjects

J R Shapiro

Publications and source records attributed to J R Shapiro.

At least 55 records · Page 3Linked to original sources
PubMed1993

Chromosome number and carmustine sensitivity in human gliomas.

BACKGROUND: Although some patients with malignant gliomas respond to treatment with chemotherapeutic agents like BCNU, tumor recurrence inevitably occurs, heralding the development of chemoresistance. Treating and/or preventing chemoresistance requires distinguishing newly developed resistance from the presence of intrinsically resistant cells in the primary tumor population. This study relates the chromosomal complements of freshly resected astrocytomas to the cells' chemosensitivity and ultimately to the patients' response to treatment. METHODS: The authors dissociated 31 freshly resected human gliomas (5 astrocytomas, 10 anaplastic astrocytomas, 16 glioblastomas multiforme) into single cells, and performed cytogenetic analysis and BCNU sensitivity testing using the colony-forming assay (CFA) on first division cells from these tumors. RESULTS: The major cytogenetic abnormalities involved the loss of a sex chromosome in all three classes of gliomas and the gain of chromosome 7 in anaplastic astrocytoma and glioblastoma multiforme; clonal marker chromosomes were observed in only anaplastic astrocytoma and glioblastoma multiforme with no common rearrangement observed among the tumors. The five astrocytomas were near-diploid (2n+/-, 35-57 chromosomes/cell), and all were resistant to BCNU. Seven of ten anaplastic astrocytomas were composed primarily of 2n+/- cells and were BCNU resistant. Three other anaplastic astrocytomas had a 39% or greater representation of 4n+/- cells (88-101 chromosomes/cell), and these tumors were sensitive to BCNU. Ten of 16 glioblastomas multiforme were composed predominantly of 2n+/- cells and were resistant to carmustine. Six other glioblastomas multiforme had at least 41% 3n+/- (58-87 chromosomes/metaphase) and 4n+/- cell populations and were sensitive to carmustine. Thus, gliomas demonstrating BCNU sensitivity were more than 60% hyperdiploid (60 or more chromosomes/metaphase) with 1 to 8 clonal marker chromosomes and multiple clonal populations involving complex karyotypic deviations. In contrast, all 22 resistant tumors were composed primarily of near-diploid cells. Only 4 of 22 tumors had a clonal marker, and the chromosome ploidy changes were less extensive. CONCLUSIONS: In freshly resected untreated human gliomas, BCNU is most effective against hyperdiploid cells that have extensive ploidy changes and chromosome rearrangement, whereas resistance to carmustine is characteristic of near-diploid populations with few ploidy changes and rearranged chromosomes. This observation was consistent for all three classes of gliomas.

Adolescent↗
PubMed1993

Defective pro alpha 2(I) collagen synthesis in a recessive mutation in mice: a model of human osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a heritable disorder of connective tissue associated with fractures, osteopenia, and short stature. OI results from mutations affecting the pro alpha 1 or pro alpha 2 gene of type I collagen. We describe a strain of mice with a nonlethal recessively inherited mutation (oim) that results in phenotypic and biochemical features that simulate moderate to severe human OI. The phenotype of homozygous oim mice includes skeletal fractures, limb deformities, generalized osteopenia, and small body size. Their femurs are smaller and demonstrate marked cortical thinning and fewer medullary trabeculae than those of wild-type mice. Breeding studies show the mutation is inherited in most crosses as a single recessive gene on chromosome 6, near the murine Cola-2 gene. Biochemical analysis of skin and bone, as well as isolated dermal fibroblast cultures, demonstrate that alpha 1(I) homotrimeric collagen accumulates in these tissues and is secreted by fibroblasts. Short labeling studies in fibroblasts demonstrate an absence of pro alpha 2(I) collagen chains. Nucleotide sequencing of the cDNA encoding the COOH-propeptide reveals a G deletion at pro alpha 2(I) nucleotide 3983; this results in an alteration of the sequence of the last 48 amino acids. The oim mouse will facilitate the study of type I collagen-related skeletal disease.

Amino Acid Sequence↗
PubMed1993

BCNU-resistant human glioma cells with over-representation of chromosomes 7 and 22 demonstrate increased copy number and expression of platelet-derived growth factor genes.

We used standard karyotypic analyses of first-division cells to identify a subpopulation of cells in primary malignant gliomas with over-representation of chromosomes 7 and 22. These cells are a minor subpopulation in the primary tumor but become the dominant population after treatment in vitro of the cells with the chemotherapeutic agent 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). The selection for a cell with this specific karyotypic abnormality suggests that these chromosomes contain genes important to the growth of BCNU-resistant cells. Southern blot hybridization analyses demonstrate an increased copy number of the genes encoding platelet-derived growth factor (PDGF) A-chain and B-chain, which have been mapped to chromosomes 7 and 22, respectively. Reverse transcription followed by polymerase chain reaction (RT-PCR) analysis demonstrates increased expression of these genes. In addition, these cells secrete a mitogenic factor that stimulates 3H-thymidine uptake in NIH 3T3 cells. This factor is sensitive to anti-PDGF antibodies and beta-mercaptoethanol, but not to anti-EGF antibodies. These data suggest that autocrine and/or paracrine mechanisms occur in human malignant gliomas, and that over-expression of PDGF may play a role in the growth of BCNU-resistant cells in these tumors.

Base Sequence↗
PubMed1993

BCNU-sensitivity in parental cells and clones from four freshly resected near-diploid human gliomas: an astrocytoma, an anaplastic astrocytoma and two glioblastomas multiforme.

We compared the BCNU sensitivity of 4 freshly resected tumors (astrocytoma WM, and malignant gliomas MK, MB, and AM) and their clones to their karyology. The majority of primary cells in all 4 tumors had near-diploid chromosome numbers (2n +/-) and all were resistant to concentrations of BCNU exceeding 10 micrograms/ml. Following in vitro cultivation, the cells from tumors WM and MB retained their 2n +/- modal chromosome number with little change in the complexity of the karyotype. In contrast, tumors MK and AM demonstrated a more unstable genome. The modal chromosome number of MK shifted from 45 to 86 and that of tumor AM from 45 to 90. Karyotyping demonstrated additional ploidy changes and new marker chromosomes in both tumors. The colony forming assay (CFA) performed on the in vitro cultivated cells demonstrated little change in the sensitivity to BCNU in tumors WM and MB, while tumors MK and AM exhibited greater than a one log cell kill at 10.0 micrograms/ml and 15.0 micrograms/ml BCNU, respectively. The modal chromosome number and BCNU sensitivity followed a similar pattern in the 30 clones that were isolated; 21 clones with near-diploid and pseudodiploid chromosome numbers were all resistant to BCNU doses at or greater than 10 micrograms/ml. In contrast, 9 clones isolated from the 3 malignant gliomas with 3n +/- and 4n +/- modal chromosome numbers were sensitive to this concentration of BCNU. The karyotypes of the hyperdiploid clones were more complex; they contained 5 or more ploidy changes and/or had marker chromosomes. These studies confirm the association of diploidy and BCNU-resistance in freshly resected malignant gliomas.

Astrocytoma↗
PubMed1993

Tumorigenic, invasive, karyotypic, and immunocytochemical characteristics of clonal cell lines derived from a spontaneous canine anaplastic astrocytoma.

Tumor cells from a spontaneously arising canine astrocytoma were isolated and cloned. Three clonally derived cell lines (DL3580 clone 1, DL3580 clone 2, and DL3580 clone 3) were developed and found to express glial fibrillary acidic protein (GFAP) as well as epidermal growth factor receptor (EGFR/c-erbB1). The cell lines were tumorigenic as subcutaneous xenografts or as intracranial implants in athymic mice, or both. Both the monolayer astrocytoma cells and the xenograft tumor cells from clone 2 were aneuploid, with a modal number of 84 chromosomes per metaphase; clones 1 and 3 were also aneuploid with modal numbers of 82 and 75/79, respectively. The histology of both the initial spontaneously occurring tumor in the dog and the intracranial astrocytoma in athymic mice demonstrated features of diffuse infiltration into normal brain. These newly developed canine glioma cell lines are karyotypically stable for 1 yr in culture and carry the same marker chromosomes as the parental lines. These glioma cell lines may serve as models for investigating mechanisms of glioma invasion into brain. Additionally, clonal cell lines with divergent properties isolated from the same tumor may assist in studies of the molecular basis of astrocytoma progression and heterogeneity.

Animals↗
PubMed1993

Recurrent erythema migrans despite extended antibiotic treatment with minocycline in a patient with persisting Borrelia burgdorferi infection.

Erythema migrans recurred in a patient 6 months after a course of treatment with minocycline for Lyme disease. Polymerase chain reaction on heparinized peripheral blood at that time demonstrated the presence of Borrelia burgdorferi-specific DNA. The patient was seronegative by Lyme enzyme-linked immunosorbent assay but showed suspicious bands on Western blot. Findings of a Warthin-Starry stain of a skin biopsy specimen of the eruption revealed a Borrelia-compatible structure. Reinfection was not believed to have occurred. Further treatment with minocycline led to resolution of the erythema migrans.

Aged↗
PubMed1993

The significance of morphologically viable glioma cells found at the time of operation after interstitial brachytherapy.

The significance of finding morphologically intact viable glioma cells in tumors treated with high-dose irradiation delivered by interstitial brachytherapy was examined. Freshly resected tissue was taken from 12 patients after (n = 8) or both before and after (n = 4) interstitial brachytherapy. All posttreatment tissue was taken from regions within a radius of 2.0 to 4.0 cm of the radioactive source. From each sample, monolayer cell culture was established. All untreated samples from primary tumors grew well and became established as cell lines within 1 to 3 weeks. In contrast, cells from treated tumors only formed small colonies of 50 to 100 cells each. These cells grew slowly and, within 14 to 21 days, degenerated. Neither the use of conditioned medium or cell extract from established glioma cell lines nor the application of growth factors (platelet-derived growth factor and/or epidermal growth factor) stimulated growth or lengthened survival. The only exception was tumor resected from approximately 4 cm from the nearest radioactive source and from which a viable cell line could be established (IRR). Cytogenetic analysis of tissue from one sample (IR) before source implantation and from another (IRR) after source implantation, both from the same patient, showed that cells IR and IRR were derived from the same stem cell. To establish the reason why cell IRR remained clonogenic despite high-dose irradiation, IRR cells were irradiated with gamma irradiation with a dose rate of approximately 1 Gy/min for 24 hours. This colony-forming assay showed that IRR cells were radiosensitive.(ABSTRACT TRUNCATED AT 250 WORDS)

Astrocytoma↗
PubMed1992

Expression of mutant alpha (I)-procollagen in osteoblast and fibroblast cultures from a proband with osteogenesis imperfecta type IV.

This study compares the synthesis of mutant type I collagen in cultured dermal fibroblasts and trabecular osteoblasts that were isolated from a patient with moderately severe osteogenesis imperfecta (type IV). Previous study of this patient's dermal fibroblasts revealed a 2000 dalton deletion located in cyanogen bromide peptide 4 of alpha 2(I)-collagen. The phenotype of the bone cell cultures was defined by a 3-4 day logarithmic phase doubling time, predominantly type I collagen production over type III and alkaline phosphatase activity 13.5 times dermal fibroblast levels. The current study revealed that both fibroblasts and osteoblasts synthesized a normal and a shortened alpha 2(I) chain, each as the product of separate alleles. Following pepsin treatment of the procollagens, a shortened alpha 1(I) chain was also seen in both cell types. Cyanogen bromide peptide mapping of osteoblast alpha-chains demonstrated the same deletions in the cyanogen bromide peptide 4 as observed in the fibroblast cyanogen bromide maps. PAGE analysis of oligonucleotide-specific cDNA that was reverse transcribed from RNA isolated from fibroblasts and osteoblasts also demonstrated the presence of two bands, one the normal size of alpha 2(I) cDNA and a second species that was smaller by 54 base pairs. Sequencing of polymerase chain reaction-amplified cDNA fragments revealed an in-frame deletion of exon 12. This finding was confirmed by the RNase protection method. Genomic DNA sequencing detected a T----G point mutation in the second position of the 5' splice donor site of intron 12. Therefore, in this patient with osteogenesis imperfecta there was no qualitative alteration in the osteoblast-specific expression of this mutant alpha 2(I)-collagen allele compared to dermal fibroblasts.

Alkaline Phosphatase↗
PubMed1992

Identification of estrogen receptor mRNA and the estrogen modulation of parathyroid hormone-stimulated cyclic AMP accumulation in opossum kidney cells.

The opossum kidney (OK) cell was used as a model to test the hypothesis that estrogen directly affects proximal renal tubular epithelial cells. To demonstrate the expression of estrogen receptor in OK cells, we developed an approach using reverse transcription and the polymerase chain reaction. Analysis of the DNA amplified with nested primers revealed the predicted size fragment and restriction enzyme digestion products. To demonstrate the functional effects of estrogen, OK cells at confluence were preincubated in serum-free medium for 7-10 days with or without 17 beta-estradiol. Bovine PTH(1-34) (bPTH(1-34)) then stimulated a dose-dependent intracellular accumulation of cAMP that was maximal after 1 min and then gradually declined. Cyclic AMP in the medium slowly increased over 60 min. Preincubation with 17 beta-estradiol did not affect cell proliferation as measured by total protein content but caused an inhibition of bPTH(1-34)-stimulated intracellular cAMP accumulation that was maximal at 10(-11) M 17 beta-estradiol (71 +/- 3% control, p less than .001). bPTH(1-34) also increased cAMP release into the medium, an effect maximal using 10(-10) M 17 beta-estradiol (118 +/- 3% control, p less than .001). Preincubation with the inactive isomer 17 alpha-estradiol caused no changes in cAMP accumulation or release. Coincubation with the antiestrogen tamoxifen blocked the effects of 17 beta-estradiol. Sodium-dependent phosphate transport was: (1) inhibited by 2-h incubations with 10(-8) or 10(-10) M bPTH(1-34) and not affected by preincubation with 17 beta-estradiol, and (2) not inhibited by a 20-min incubation with 10(-8) M bPTH(1-34) unless cells were preincubated with 10(-8) M 17 beta-estradiol, suggesting that any possible effects of estrogen on phosphate transport are not directly mediated by changes in cAMP. These studies demonstrate the presence of estrogen receptor mRNA in OK cells as well as direct and specific effects of physiologic concentrations of estrogen on cAMP accumulation in these cells. This system may be a good model for further study of estrogen and PTH effects on the kidney.

Aged↗
PubMed1992

In vitro karyotypic and immunophenotypic characterisation of primitive neuroectodermal tumours: similarities to malignant gliomas.

Monoclonal antibody (Mab) mediated immunotherapy of brain tumours requires the identification of tumour-restricted cell surface antigens. We have characterised four primitive neuroectodermal tumours, which included pineoblastoma, medulloblastoma and ependymoblastoma cultures, that demonstrated in vitro evidence of malignant behaviour (anchorage-independent growth and nu/nu xenograft tumour formation). The cytogenetic findings ranged from normal G-banded and Q-banded karyotypes through mixed near-diploid/hyperdiploid. These cultures resembled the cell surface immunophenotypic spectrum of malignant gliomas. They were distinguished from normal glia in vitro by the expression of restricted fetal mesenchymal, neuronal, myoblastic, melanocytic, epidermal, chondrocytic, lymphoid and epithelial antigens. Certain antigens appeared sufficiently represented among central nervous system (CNS) neoplasms to afford potential targets for Mab-mediated immunotherapy.

Adolescent↗
PubMed1992

Myocardial contrast echocardiography: influence of ischaemia and hyperaemia in an animal model.

To evaluate changes in myocardial contrast echocardiography during ischaemia and hyperaemia, contrast studies were performed in 16 open chest dogs. Time-intensity curves were generated using videodensitometry after contrast injections to demonstrate ischaemic and non-ischaemic areas of interest during a wide range of coronary blood flow levels. For each time-intensity curve, the peak contrast intensity (PCI), washout halftime (T1/2) and area under the curve (AUC) were calculated. PCI and AUC decreased significantly only with severe ischaemia (90% or more reduction in flow), and increased significantly with hyperaemia of more than 2.5 times baseline flow. Both ischaemia and hyperaemia were found to prolong the T1/2. There was only a moderate linear correlation between the magnitude of hyperaemia and myocardial contrast echocardiographic parameters. There was significantly less increase in myocardial contrast echocardiographic parameters during hyperaemia in segments supplied by a stenosed coronary artery.

Albumins↗
PubMed1992

An osteopenic nonfracture syndrome with features of mild osteogenesis imperfecta associated with the substitution of a cysteine for glycine at triple helix position 43 in the pro alpha 1(I) chain of type I collagen.

Mutations affecting the pro alpha 1(I) or pro alpha 2(I) collagen genes have been identified in each of the major clinical types of osteogenesis imperfecta. This study reports the presence of a heritable connective tissue disorder in a family with an osteopenic syndrome which has features of mild osteogenesis imperfecta but was considered idiopathic osteoporosis in the proband. At age 38, while still premenopausal, she was found to have osteopenia, short stature, hypermobile joints, mild hyperelastic skin, mild scoliosis, and blue sclerae. There was no history of vertebral or appendicular fracture. Hip and vertebral bone mineral density measurements were consistent with marked fracture risk. Delayed reduction SDS-PAGE of pepsin-digested collagens from dermal fibroblast cultures demonstrated an anomalous band migrating between alpha 1(I) and alpha 1(III). This band merged with the normal alpha-chains upon prereduction, indicating an unexpected cysteine residue. Cyanogen bromide peptide mapping suggested that the mutation was in the smaller NH2-terminal peptides. cDNA was reverse transcribed from mRNA and amplified by the polymerase chain reaction. A basepair mismatch between proband and control alpha 1(I) cDNA hybrids was detected by chemical cleavage with hydroxylamine:piperidine. The cysteine substitution was thus localized to alpha 1(I) exon 9 within the cyanogen bromide 4 peptide. Nucleotide sequence analysis localized a G----T point mutation in the first position of helical codon 43, replacing the expected glycine (GGT) residue with a cysteine (TGT). The prevalence of similar NH2-terminal mutations in subjects with this phenotype which clinically overlaps idiopathic osteoporosis remains to be determined.

Adolescent↗
PubMed1990

Reproducibility of quantitative myocardial contrast echocardiography.

To determine whether myocardial contrast echocardiography is quantitatively reproducible, repeated intracoronary injections of sonicated albumin (5%) were performed in eight open chest dogs. Paired injections were performed at baseline, during ischemia produced by ligation of a coronary artery, and during hyperemia induced by intravenous infusion of 0.75 mg/kg body weight of dipyridamole. Contrast washout curves were generated for the left anterior descending coronary artery territory (ischemic area) and left circumflex coronary artery territory (nonischemic area) by beat per beat analysis of frozen end-diastolic frames of left ventricular short-axis views. Peak contrast intensity, contrast washout half-time and area under the curve were derived from these curves. A total of 75 contrast washout curves were analyzed for the study of interinjection, intraobserver and interobserver reproducibility. The correlation coefficients between measurements obtained from paired injections of the echocardiographic contrast agent (interinjection reproducibility) ranged from 0.78 for peak contrast intensity to 0.87 for area under the curve. Percent error varied between 14.7% and 24.7%. The intraobserver variability in measurements was less than the interinjection variability, with a cumulative mean percent error of 17.8% and correlation coefficients of 0.72 (peak contrast intensity), 0.95 (area under the curve) and 0.96 (washout half-time). Interobserver correlation for all indexes was high (r = 0.92 to 0.96). It is concluded that peak contrast intensity, contrast washout half-time and the area under the curve derived from myocardial contrast washout curves can be measured reproducibly from videotapes. In addition, the variability between two injections attempted under identical conditions is greater than reader variability from videotapes.

Albumins↗
PubMed1990

Factor analysis of the immunophenotypes of astrocytomas and malignant gliomas: correlations with tumor grade and patient survival.

Our previous investigations correlated the degree of cytogenetic and immunophenotypic heterogeneity of cultured normal glia, astrocytomas and malignant gliomas. The possible significance was suggested by the statistical correlation of individual antigens with diagnosis and patient survival. The present study has established the patterns of covariation of titers of monoclonal antibody reactivity with a panel of cell surface antigens among normal glia (8), astrocytomas (4), anaplastic astrocytomas (12), mixed malignant gliomas (8) and glioblastomas (21). A mean aggregate titer across 43 antigens was computed for each culture and then subtracted from the observed individual titers. Factor analysis was performed to determine a small number of Factors, derived as the weighted average of the 43 mean-adjusted antigens, which accounted for a significant proportion of the covariation of immunophenotypic expression in the sample of 53 cultures. Clusters of antigens were found to independently segregate in their deviation from the aggregate phenotype. Adjusting for age and diagnosis, Factors 1 and 4 correlated with patient survival among recurrent and primary neoplasms, respectively. Factor 2 additionally discriminated between primary and recurrent gliomas. Factor 3 was associated with age at diagnosis. Factors 1 and 2 correlated with the histopathologic grade of glial tumor. Scatter plots of Factor 1 vs. 2 revealed the minimal immunophenotypic diversity of the normal glia. Astrocytomas were similar but not identical. Progressive divergence was evident between the immunophenotypes of anaplastic astrocytomas, mixed gliomas and glioblastomas. These data suggest that qualitative and quantitative differences in antigenic heterogeneity may identify stages in glial tumor progression.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1990

Intravenous contrast echocardiography with use of sonicated albumin in humans: systolic disappearance of left ventricular contrast after transpulmonary transmission.

The transmission of echocardiographic contrast medium and the cyclic changes in left ventricular videodensity during transpulmonary contrast echocardiography were investigated in nine adult volunteers with the use of intravenous injections of sonicated albumin (microbubble size 5.2 +/- 2.6 microns). Right and left ventricular and myocardial contrast were quantitated by videodensitometric analysis. The injections caused no symptoms, and no hemodynamic or electrocardiographic changes were observed. All injections resulted in right ventricular contrast. Mean peak right ventricular videodensity was 75 +/- 48 at end-diastole and 61 +/- 36 gray scale U/pixel at end-systole (p less than 0.05). Seventy-eight percent of injections resulted in left ventricular contrast with a mean peak videodensity of 21 +/- 33 gray scale U/pixel. Early systole was associated with a rapid decrease in left ventricular contrast intensity with near total disappearance of contrast by end-systole (from 23 +/- 33 and 17 +/- 23 U/pixel at end-diastole to 6 +/- 10 and 3 +/- 2 at end-systole at the left ventricular base and apex, respectively; p less than 0.05). None of the injections resulted in myocardial contrast enhancement by visual or quantitative analysis. Thus, left ventricular contrast echocardiography can be achieved after intravenous injections of sonicated albumin. Transpulmonary left ventricular contrast echocardiography is associated with near total disappearance of contrast during systole. This may be secondary to the destruction of microbubbles by the high left ventricular systolic pressure. These findings may help explain the limited success of this technique thus far for myocardial perfusion imaging.

Adult↗