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Biomedical subjects

A Weinreb

Publications and source records attributed to A Weinreb.

At least 19 recordsLinked to original sources
PubMed1999

Preparation of a diagnostic antigen of human melanoma based on lymphocyte activation as measured by intracellular fluorescein fluorescence polarization.

The intracellular fluorescein fluorescence polarization (IFFP) test indicates that peripheral blood lymphocytes (PBL) of cancer patients display stimulatory sensitivity to a short incubation with specific tumor protein extracts. In this work, a human lymphocyte activation melanoma antigen (LAMA) was purified from supernatant of a human melanoma cell line (L1M1), which could specifically stimulate lymphocytes of melanoma patients. The results showed a significant stimulation of lymphocytes from healthy donors after incubation with phytohaemagglutinin (PHA), while no stimulation was observed after incubation with LAMA. On the other hand, lymphocytes from melanoma patients showed a significant stimulation with LAMA, while generally showing minor or no stimulation with PHA. Melanoma specificity of LAMA was demonstrated by no response in lymphocytes from patients of lung, colon, or breast cancer. The purified fraction is therefore considered to be a shared tissue-specific antigen which may be useful in immunodiagnosis and immunotherapy of melanoma.

Antigens, Neoplasm↗
PubMed1998

Genetic determinants of autoimmune disease and coronary vasculitis in the MRL-lpr/lpr mouse model of systemic lupus erythematosus.

MRL-lpr/lpr (MRL/lpr) mice are a model of human autoimmune disease. They exhibit a number of characteristics of systemic lupus erythematosus, including anti-DNA Abs, anti-cardiolipin Abs, immune complex-mediated vasculitis, lymphadenopathy, and severe glomerulonephritis. Although the autoimmune disorder is mediated primarily by mutation of the Fas gene (lpr), which interferes with lymphocyte apoptosis, MRL/lpr mice also have other predisposing genetic factors. In an effort to identify these additional factors, we have applied quantitative trait locus (QTL) mapping using an intercross between MRL/lpr mice and the nonautoimmune inbred strain BALB/cJ. A complete linkage map spanning the entire genome was constructed for 189 intercross progeny, and genetic loci contributing to features of the autoimmunity were identified using statistical analytic procedures. As expected, the primary genetic determinant of autoimmune disease in this cross was the Fas gene on mouse chromosome 19, exhibiting a lod score of 60. In addition, two novel loci, one on chromosome 2 (lod score, 4.3) and one on chromosome 11 (lod score, 3.1), were found to contribute to levels of anti-DNA Abs. Interestingly, the chromosome 19 and chromosome 11 QTLs, but not the chromosome 2 QTL, also exhibited associations with anti-cardiolipin Abs (lod scores, 38.4 and 2.6). We further examined the effects of these QTLs on the development of coronary vasculitis in the F2 mice. Our results indicate that the QTLs on chromosomes 11 and 19 also control the development of vasculitis, demonstrating common genetic determinants of autoantibody levels and vasculitis.

Animals↗
PubMed1998

Mapping a gene for combined hyperlipidaemia in a mutant mouse strain.

Familial combined hyperlipidaemia (FCHL) is a common, multifactorial disorder associated with elevated levels of plasma triglyceride, cholesterol, or both. A characteristic feature is increased secretion of very low density lipoproteins (VLDL) and apolipoprotein B (apoB). Although FCHL is the most common cause of premature coronary artery disease (CAD), accounting for over 10% of cases, its aetiology remains largely unknown. One powerful approach to the dissection of complex genetic traits involves the use of animal models. We have identified a mouse strain, HcB-19/Dem (HcB-19), which exhibits hypertriglyceridaemia, hypercholesterolaemia and elevated levels of plasma apoB. Like FCHL patients, HcB-19 mice also exhibit increased secretion of triglyceride-rich lipoproteins, and their hyperlipidaemia becomes progressively more severe with age. It is likely that the hyperlipidaemia results from a mutation of a novel gene that arose during development of strain HcB-19. We mapped the hyperlipidaemia gene (Hyplip1) to the distal portion of mouse chromosome 3. This region is syntenic to human chromosome 1q21-q23, which has recently been shown to harbour a gene associated with FCHL in families from a Finnish isolate.

Age Factors↗
PubMed1997

Cell activation influences cell staining kinetics.

Stimulation of cells has so far been observed, among other methods, by the decrease of the intracellular fluorescein fluorescence polarization (IFFP). It is shown that the rate constant of leakage of the fluorescent marker out of the cells increases with stimulation much more significantly than the polarization decreases; thus it might provide a more sensitive method to observe cells stimulation. It is also shown that due to negligible leakage of the marker out of the cells shortly after initiation of the staining of the cell suspension, the fluorescein fluorescence polarization (FFP) of the cell suspension, is very close to IFFP.

Fluoresceins↗
PubMed1997

Carboxyfluorescein as a fluorescent probe for cytoplasmic effects of lymphocyte stimulation.

The application of carboxyfluorescein (CF), as an impermeable fluorescent probe for lymphocyte stimulation with phytohaemagglutinin (PHA), is investigated by following a decrease in the degree of fluorescence polarization. Since CF does not enter the mitochondria, the present results indicate that the measured effect of stimulation occurs in the cytoplasm. The results also reveal that the fluorescence yield of intracellular CF is smaller than that of extracellular CF. Moreover, the degree of fluorescence polarization of intracellular CF is inversely related to its concentration. Following cell disruption, fluorescence intensity increases and polarization decreases. These effects might indicate a weak or reversible association of intracellular CF with cytoplasmic proteins.

Cytoplasm↗
PubMed1997

Detection of melanoma by monitoring the intracellular fluorescein fluorescence polarization changes in lymphocytes.

The effectiveness of detecting melanoma by measuring the intracellular fluorescein fluorescent polarization (IFFP) of patients' SCM (structuredness of the cytoplasmic matrix)-responding lymphocytes was examined. SCM-responding lymphocytes from 46 melanoma patients and 32 healthy volunteers were labeled with fluorescein diacetate and challenged with different stimuli, and the resulting polarization was determined. The polarizations (P) obtained upon stimulation with nothing (P-0), encephalitogenic factor (P-EF), phytohaemagglutinin (P-PHA), or melanoma antigen (P-MEL), and the ratios RR(ef) (P-EF/P-PHA) and RR(mel) (P-MEL/P-PHA) were lower for SCM-responding lymphocytes from the patients as a group than for those of the controls. The specificity and sensitivity of the IFFP tests (using cutoff values) to detect melanoma were 90.6 and 73.9%, respectively. The IFFP tests may facilitate the discrimination between melanoma patients and healthy subjects, and may be used in follow-up of patients with melanoma.

Algorithms↗
PubMed1996

Lymphocyte fluorescence polarization measurements with the cellscan system: application to the SCM cancer test.

The SCM (Structuredness of Cytoplasmic Matrix) cancer test, a procedure based on detection of differences in lymphocyte activation between individuals with and without cancer, has remained controversial with inconsistent results reported by different authors. As originally described, the test includes two technically demanding steps, the first a lymphocyte separation procedure and the second a series of fluorescence polarization measurements. The Cellscan, a high-precision static cytometer system has been configured to perform the SCM test. The apparatus facilitates the polarization measurements and can analyze cells separated using simpler procedures than were originally described. Using methods and diagnostic criteria adapted for the Cellscan system, the SCM test correctly classified > 90% of patients with cancer and > 90% of individuals without cancer.

Adult↗
PubMed1996

Decrease of intracellular fluorescein fluorescence polarization (IFFP) in human peripheral blood lymphocytes undergoing stimulation with phytohaemagglutinin (PHA), concanavalin A (ConA), pokeweed mitogen (PWM) and anti-CD3 antibody.

In the present study we describe the induction of changes in intracellular fluorescein fluorescence polarization (IFFP) in lymphocytes undergoing activation with a variety of stimulants. These stimulants included the lectins phytohaemagglutinin (PHA), concanavalin (ConA), pokeweed mitogen (PWM) and anti-CD3 antibody. Changes in IFFP were detected in individual cells using the Cellscan apparatus. Our results show that by employing mitogenic concentrations of PHA, as revealed in a [3H]-thymidine incorporation assay, a decrease in the IFFP in human peripheral blood lymphocytes (PBL) occurred within 40 min. ConA and anti-CD3 affected similarly IFFP, whereas PWM, a B lymphocyte lectin, had no effect on IFFP at the concentrations employed. Kinetic analysis revealed that changes in IFFP occurred within 20-40 min after exposure to the stimulants and lasted for 24 h. Our results show that stimulants which activate CD3+ lymphocytes caused immediate changes in IFFP, in an enriched population of human PBL. The possible mechanisms involved in IFFP modulation following exposure to selected stimulants are discussed.

Antibody Specificity↗
PubMed1996

Adaptation of the cellscan technique for the SCM test in breast cancer.

The value of the SCM (Structuredness of Cytoplasmic Matrix) cancer test, a procedure based on the detection of differences in lymphocyte activation in the presence and absence of cancer, has remained controversial, with inconsistent results having been reported among investigators. The Cellscan, a high-precision static cytometer system, has been designed to perform the SCM test; the apparatus facilitates the polarisation measurements and can examine cells which have been separated by simpler procedures than were originally described. In this study, using methods and diagnostic criteria adapted for the Cellscan system in a hospital environment, the SCM test correctly classified over 90% (76/80) of patients with breast cancer and differentiated over 90% (72/73) of individuals without cancer.

Adult↗
PubMed1996

Effect of interleukin-1 alpha, interleukin-1 beta and tumor necrosis factor-alpha on the intracellular fluorescein fluorescence polarization of human lung fibroblasts.

In the present study we aimed to detect early intracellular changes in the cytoplasmic matrix induced in human, pulmonary-derived fibroblasts following exposure to interleukin (IL)-1 alpha, IL-1 beta and tumor necrosis factor-alpha. Such changes were detected by measuring intracellular fluorescein fluorescence polarization (IFFP) using the Cellscan apparatus. IFFP measurement was selected in our study since it has been shown to reflect the microviscosity of the cytoplasmic matrix. Significant reductions (> or = 5%) in the IFFP were induced in fibroblasts by all the cytokines employed. The effect of cytokines on IFFP was achieved at concentrations of 5-10 ng/ml of the cytokines. The reduction in IFFP, following stimulation with the cytokines, was detected as early as 20 min after exposure to the cytokines, lasted at least 40-60 min after exposure to IL-1 alpha and IL-1 beta, and was inhibited by vinblastine, an inhibitor of the polymerization of microtubules. Our results show that IFFP measurements by the Cellscan may reveal rapid intracellular changes occurring in the cytoskeleton components of activated cells.

Cytochalasin B↗
PubMed1996

Inhibition of mitogen-induced changes in intracellular fluorescein fluorescence polarization of human peripheral blood lymphocytes by colchicine, vinblastine and cytochalasin B.

We have previously reported that the exposure of human peripheral blood lymphocytes (PBL) to a variety of stimulants caused rapid changes in intracellular fluorescein fluorescence polarization (IFFP) in the activated cells. In the present study we further analyzed possible mechanisms responsible for the changes in IFFP in PBL exposed to phytohaemagglutinin (PHA) and anti-CD3 antibody. By employing several agents which are known to affect the polymerization of the cytoskeleton we showed that both cytochalasin B, which regulates the microfilaments structure, and vinblastine and colchicine, which affect the microtubules, completely abolished the changes induced in IFFP of human PBL by both PHA and anti-CD3. This effect was dose dependent and was noted at concentrations ranging from 10 to 100 microM of cytochalasin B and 10 microM of vinblastine and colchicine. The effect of these cytoskeleton modulators occurred within 20 minutes after the initiation of activation with PHA. Our results indicate that activation with PHA and anti-CD3 causes early changes in the microtubules and microfilaments components of the cytoskeleton. The possible application of IFFP measurement in analyzing early changes in the cytoskeleton following cell activation is discussed.

Colchicine↗
PubMed1996

Detection of colon cancer by monitoring the intracellular fluorescein fluorescence polarization changes in lymphocytes.

Structuredness of the cytoplasmic matrix (SCM)-responding lymphocytes from healthy donors can be differentiated from SCM-responding lymphocytes of patients with malignant disease on the basis of the change in the intracellular fluorescein fluorescence polarization (IFFP) induced by their exposure to various antigens. We have found that the overall sensitivity, specificity and positive and negative predictive values of the test were 92.0, 92.6, 96.3, and 84.7%, respectively. We demonstrated the capability of the test to distinguish between healthy people and colorectal cancer patients per stage of the disease. We also found a significant difference in IFFP values between Dukes' C patients and patients with metastatic disease, rendering the test potentially helpful in follow-up.

Adult↗
PubMed1995

Fluorescence polarisation changes in lymphocyte cytoplasm as a diagnostic test for breast carcinoma.

Lymphocytic cytoplasm from individuals with malignant disease, and from those without, differ in such a way as to be diagnostic both of malignancy generally and of specific types of cancer. Mitogenic stimulation of lymphocytes by phytohaemagglutinin (PHA) and antigenic stimulation by encephalitogenic factor (EF) and certain specific tumour-associated antigens, provokes changes in the structure of the cytoplasmic matrix (SCM) which are detectable upon fluorescence polarisation. The degree of change is quantifiable both by calculating the polarisation ration (PR, polarisation before and after stimulation) and the relative ratio (RRSCM, the ratio between the polarisation obtained after exposure to EF [PEF] and to the polarisation measured after exposure to PHA [PPHA]). A new tumour-associated antigen specific for breast cancer, CaBr, was tested for its diagnostic efficacy in comparison with that of EF, by prospectively testing blood samples from 138 consecutive women with suspicious breast masses. The previously known discriminatory power (sensitivity 60.7% and specificity 90.7%) of the polarisation-derived RRSCM was reconfirmed. However, the RR'SCM (the new ratio using CaBr instead of EF), was significantly more sensitive (77.4%; P < 0.01) and specific (94.4%) than the RRSCM in detecting breast cancers. The polarisation changes in the cytoplasmic matrix after stimulation by CaBr alone suggest the best discriminatory power (sensitivity 90.5% and specificity 94.4%) between cancerous and non-cancerous patients.

Adult↗
PubMed1995

Genetic factors in lipoprotein metabolism. Analysis of a genetic cross between inbred mouse strains NZB/BINJ and SM/J using a complete linkage map approach.

A genetic cross was constructed from two parental inbred strains of mice, NZB/BINJ and SM/J, which differ markedly in their plasma lipoprotein levels. Plasma lipid and apolipoprotein values were measured in 184 F2 progeny on a normal chow diet and on an atherogenic diet. Genetic markers were typed at 126 loci spanning all chromosomes except the Y. Statistical analysis revealed significant linkage or suggestive linkage of lipoprotein levels with markers on a number of chromosomes. Chromosome 1 markers were linked to levels of total cholesterol (lod 5.9) and high density lipoprotein (HDL) cholesterol (lod 8.1), chromosome 5 markers were linked to levels of total cholesterol (lod 6.7) and HDL cholesterol (lod 5.6), and chromosome 7 markers were linked to levels of total plasma triglycerides (lod 5.1) and free fatty acids (lod 5.6). Plasma apoAII levels were linked to the apoAII gene (lod score 19.6) and were highly correlated with plasma HDL cholesterol levels (r = 0.63, P = 0.0001), indicating that apoAII expression influences HDL cholesterol levels. Molecular studies suggested that structural differences in the apoAII polypeptide of the two strains may contribute to differences in clearance of the protein.

Amino Acid Sequence↗
PubMed1994

Apparatus for high-precision repetitive sequential optical measurement of living cells.

A system is described which permits the repetitive spectroscopic measurement of individual cells within a population of many cells, while the location of each cell is preserved during various manipulations of the cells and/or their surrounding medium. The central mechanical feature of the system is the cell carrier, a matrix of apertures in which the cells become trapped. The detector electronics operate in a preset photon counting mode, permitting the measurement of low and high light intensities with the same degree of precision. The present instrument configuration is specifically designed for the accurate and precise measurement of the polarization of fluorescence of probes within living cells for application to routine performance of the Cercek SCM test for cancer. With modifications, the apparatus can be applied to a wide range of other clinical and research tasks.

Cell Separation↗
PubMed1993

Multiple-scattering effects in transmission oximetry.

To develop an algorithm for the spectrophotometric determination of the oxygen saturation in blood, a model for the transmission of light in a scattering and absorbing medium is developed, taking into account effects of multiple scattering. The computed results obtained by a Monte Carlo simulating program agreed well with those found by experiment. The results were compared with those obtained by the commonly used algorithm of transmission oximetry (based on Lambert-Beer's law) and it was found that the calibration curves obtained by this method were strongly dependent on the haematocrit and thickness of the sample. These curves were less reliable the lower the saturation of oxygen.

Algorithms↗
PubMed1993

Detecting marker-QTL linkage and estimating QTL gene effect and map location using a saturated genetic map.

A simulation study was carried out on a backcross population in order to determine the effect of marker spacing, gene effect and population size on the power of marker-quantitative trait loci (QTL) linkage experiments and on the standard error of maximum likelihood estimates (MLE) of QTL gene effect and map location. Power of detecting a QTL was virtually the same for a marker spacing of 10 cM as for an infinite number of markers and was only slightly decreased for marker spacing of 20 or even 50 cM. The advantage of using interval mapping as compared to single-marker analysis was slight. "Resolving power" of a marker-QTL linkage experiment was defined as the 95% confidence interval for the QTL map location that would be obtained when scoring an infinite number of markers. It was found that reducing marker spacing below the resolving power did not add appreciably to narrowing the confidence interval. Thus, the 95% confidence interval with infinite markers sets the useful marker spacing for estimating QTL map location for a given population size and estimated gene effect.

Chromosome Mapping↗