Search PubMed⌕ Search

Biomedical subjects

A Orfao

Publications and source records attributed to A Orfao.

At least 127 records · Page 7Linked to original sources
PubMed1997

Enzyme changes in zymogen granules and in pancreatic secretion throughout long-term CCK treatment.

Pancreatic enzyme storage and secretion were studied in rats treated twice daily with s.c. injections (5 micrograms/kg) of CCK-8 for 3, 7, and 15 days. Isolated zymogen granules were analyzed by flow cytometry to determine their FSC (forward scatter), SSC (side scatter), and amylase and trypsinogen contents. DNA content, pancreatic weight, and both basal and stimulated pancreatic secretion under i.v. CCK infusion (1.25 micrograms/kg/h) were also studies. Two subsets of zymogen granules were identified by flow cytometry in both control and CCK-treated rats on the basis of FSC and SSC parameters: Z1 (smaller and less complex) and Z2. Both subsets displayed a high degree of heterogeneity with respect to their enzyme content per zymogen granule. During the first 7 days of CCK treatment, hyperplasia and hypertrophy developed in the rats together with changes in the zymogen granules, reflected by a significantly decreased FSC, and increased SSC, and an increase in the mean trypsinogen/amylase ratio per granule. A rise in pancreatic enzyme secretion, especially of trypsin, was observed. After 15 days of CCK administration, a simultaneous decrease in amylase content and increase in trypsinogen content per zymogen granule was observed. A desensitization of the pancreas to CCK happened after 15 days of CCK administration, reflected by a reduction of all the pancreatic functions that had been increased at shorter CCK administration periods. Nevertheless, trypsinogen appeared resistant to desensitization because its secretion significantly increased in response to an i.v. infusion of CCK. CCK treatment displayed a differential packaging of the enzymes in individual zymogen granules; the trypsinogen/amylase ration was significantly higher in Z2 zymogen granules than in Z1 subset throughout the treatment.

Amylases↗
PubMed1997

Hydrocortisone induces an increase of amylase content in individual zymogen granules from rat pancreas.

This study was performed to evaluate the effects of different doses of hydrocortisone (1, 10 and 25 mg/kg/day) administered for 1, 3 and 8 days on pancreatic enzyme storage in rats. The enzyme content in both pancreas homogenates and in individual isolated zymogen granules (ZGs) was measured using standard biochemical assays and flow cytometry, respectively. Hydrocortisone did not alter the total amount of pancreatic DNA but increased the pancreas enzyme content in a time-dose-dependent way. Amylase activity was significantly increased after hydrocortisone administration at day +8 when 10 mg/kg/day was used, and from the first day of treatment when 25 mg/kg/day was administered. A significant increase in trypsin activity was also observed in response to 25 mg/kg/day of hydrocortisone but only from the third day of treatment onwards. As compared with control rats, chronic administration of either 1 or 10 mg/kg/day of hydrocortisone did not alter significantly either the size or the percentage of the two ZG subpopulations (Z1 and Z2) identified in the pancreas by flow cytometry; in addition, no significant changes were observed in the mean amylase content per individual granule, although its mean concentration increased in rats treated with 10 mg/kg/day for 3 and 8 days. Nevertheless, when 25 mg/kg/day of hydrocortisone were administered for 1 and 3 days, a significant increase in the proportion of Z1 ZGs was observed, which may be related to the formation of new and smaller ZGs. When a very high dose of hydrocortisone (25 mg/kg/day) was used, an overall increase in the pancreatic enzyme content related to an increase in the mean amylase content per individual ZG was observed; this effect was apparent from the first day of treatment in the Z1 subset of ZGs and from day +3 in the Z2 subpopulation. Only a high concentration of hydrocortisone was able to alter the enzyme storage process in individual zymogen granules, but they maintain a normal enzyme load at lower hydrocortisone doses.

Amylases↗
PubMed1997

Methimazole therapy in Graves' disease influences the abnormal expression of CD69 (early activation antigen) on T cells.

At present, the in vivo response of T, B and natural killer (NK) cells to antithyroid drug therapy remains largely unknown. In the present study, we have prospectively analyzed the in vivo effects of methimazole treatment on a large number of circulating T and NK cell subsets, some of them expressing cell surface activation antigens involved in the very early phase of the immune response, in a group of 17 hyperthyroid, untreated patients with Graves' disease (GD). As one of the first events during T cell activation is the expression of interleukin (IL) receptors, we also studied the binding of IL-2 and IL-6 to T cells. Patients with Graves' disease were sequentially studied at diagnosis/before treatment (day 0) and 7, 14, 30, 90 and 180 days after methimazole therapy. The results were compared with both a group of 19 age- and sex-matched control volunteers and a group of 20 untreated/euthyroid patients with Graves' disease in long-term remission. The combination of flow cytometry and three-color immunofluorescence revealed a clear (P < 0.01) decrease in the percentage of NK cells before and during the whole course of therapy with respect to both controls and patients with Graves' disease who were in long-term remission. Before therapy, a marked increase (P < 0.001) in the ratio of B to NK cells was also observed; thereafter, a slight decrease in this ratio was observed, although normal values were detected only in patients in long-term remission. Expression of the CD69 early activation antigen in the hyperthyroid untreated patients with Graves' disease was clearly increased (P < 0.01) with respect to both controls and patients with Graves' disease who were in long-term remission. This abnormal CD69 expression was found to be significantly reduced (P < 0.001) by methimazole therapy, and this represents a new effect of the drug. Expression of the low-affinity receptor for IL-2 (CD25)--another early T cell activation marker--was not altered in Graves' disease, but the binding of IL-2 and IL-6 to T cells exhibited a progressive and parallel increase during the first 30 days of therapy, decreasing thereafter. Our results show that methimazole therapy downregulates the abnormally high expression of the CD69 early activation antigen on T cells, being less effective on inducing changes in other T cell activation markers and in NK cells.

Adult↗
PubMed1997

Abnormalities of peripheral blood T lymphocytes and natural killer cells in alcoholic hepatitis persist after a 3-month withdrawal period.

Present information about the behavior of the different lymphoid subsets in alcoholic hepatitis (AH), including cells displaying cytotoxic activity, is scanty and contradictory. The aim of this study was to gain further insight into knowledge of the immunological abnormalities involved in AH and the possible role of ethanol (EtOH) consumption in these changes. We analyzed the distribution of a wide range of peripheral blood (PB) lymphoid subsets, both during active EtOH intake and after a 3-month withdrawal period, using multiple stainings with monoclonal antibodies and flow cytometry, as well as natural killer (NK) cytotoxic activity. AH patients entering the study were selected strictly; only those undergoing their first episode of AH with no other lesions at liver biopsy were enrolled. Regarding the alcohol intake period, the most striking finding was a significant increase of the absolute number of PB T cells affecting both CD4+ and CD8+ lymphocytes. These changes were associated with a higher expression of T-cell activation antigens, such as HLA DR and CD11c. Simultaneously, a significant increase in both NK cells (CD3-/CD56+) and the cytotoxic T cells coexpressing the CD3 and the CD56 molecules together with an increased NK cytotoxic activity were observed. By contrast, the CD19+/CD5+ B-cell subset was significantly decreased. No significant changes were observed with EtOH withdrawal except in CD5+ B lymphocytes, which returned to normal values. Our results show that, in AH patients, a significant expansion of both activated T cells and NK lymphocytes occurs in the PB, which is associated with an increased NK cytotoxic activity. Interestingly these abnormalities persist during the withdrawal period.

Adult↗
PubMed1997

Long-term blockade of cholecystokinin (CCK): effects of L 364,718 (a CCK receptor antagonist) on pancreatic enzyme storage and secretion.

To determine the effect of long-term blockade of cholecystokinin (CCK) on both pancreatic storage and secretion processes, L 364,718 (a CCK receptor antagonist) was administered to rats at 0.1 mg/kg/day for 3, 7, and 15 days. Zymogen granules were analyzed by flow cytometry to determine their light scattering properties-forward scatter and side scatter-as well as their amylase content measured by a specific antiserum. The mean number of zymogen granules per cell was counted on pancreatic sections using electron microscopy. DNA content, pancreatic weight, and enzyme secretion were also studied under both basal conditions and CCK infusion at a dose of 1.25 micrograms/kg/h, which is able to displace the CCK receptor antagonist. Two subpopulations of zymogen granules (Z1 and Z2) were identified on the basis of their light scattering parameters, in both control and L 364,718-treated rats. L 364,718 administered for 3, 7, and 15 days induced a significant reduction in the amylase content of individual zymogen granules, for both Z1 and Z2 zymogen granule subsets. In contrast, the number of zymogen granules per cell increased from day +3 of treatment onward, the highest values being detected at day +7. Hyperplasia was observed only at day +15. Basal enzyme secretion decreased significantly in rats treated with L 364,718 for 3 and 7 days but recovered to control values after 15 days of treatment. No significant differences in CCK-stimulated amylase secretion were observed between control and L 364,718-treated rats. At day +15 of L 364,718 treatment a significant increase in enzyme secretion was observed with respect to shorter treatment periods; this was associated with a significant increase in both the number of cells and the number of zymogen granules per cell. Our results indicate that chronic administration of L 364,718 induces a biphasic effect on pancreatic function. Interestingly, although enzyme secretion reached recovery after long-term treatment (15 days), the storage process is altered since the mean enzyme content in each individual zymogen granule remains significantly reduced.

Amylases↗
PubMed1997

Alterations in tumor necrosis factor-alpha, interferon-gamma, and interleukin-6 production by natural killer cell-enriched peripheral blood mononuclear cells in chronic alcoholism: relationship with liver disease and ethanol intake.

No previous studies have been reported on human alcoholism in which the pattern of cytokine secretion by natural killer (NK) cells is explored. The goal of the present study was to evaluate the role of NK cells in the production of cytokines in patients with chronic alcoholism, analyzing at the same time the possible relationship between cytokine production and both alcoholic liver disease and ethanol (EtOH) intake. A total of 30 chronic alcoholic patients-11 without liver disease [alcoholics without liver disease (AWLD) group] and 19 diagnosed of alcoholic liver cirrhosis-were included in this study. Twenty-five age- and sex-matched healthy volunteers were analyzed as controls. Production of interferon (IFN)-gamma, tumor necrosis factor-alpha (TNF-alpha), and interleukin (IL)-6 was performed on NK-enriched peripheral blood mononuclear cells (PBMC) after stimulation with IL-2 and IFN-alpha. In AWLD patients, the production of TNF-alpha was significantly reduced, compared with normal controls, under both IFN-alpha (p < 0.01) and IL-2 (p < 0.05) stimulation. In patients with cirrhosis, TNF-alpha production by PBMC enriched in NK cells varied depending on the EtOH intake status at the moment of evaluation. Accordingly, an increased concentration of this cytokine was detected in the supernatants of cirrhotic patients and active EtOH intake, particularly after IFN-alpha stimulation (p < 0.05); whereas, in patients with at least 1 year of alcohol withdrawal, TNF-alpha levels remained within normal range. The results on the production of IL-6 and IFN-gamma in AWLD and cirrhotic patients showed that only cirrhotic patients with a prolonged EtOH withdrawal period display abnormal production. Accordingly, in this group of patients, a significantly increased release of IL-6 was observed after both IFN-alpha and IL-2 stimulation (p < 0.01 and p < 0.05, respectively). By contrast, a lower IFN-gamma production (p < 0.005) was detected with respect to the control group. Our results point to the existence of an abnormal cytokine secretion by NK cells from chronic alcoholism patients, which depend on both the existence of liver disease and the status of EtOH intake.

Adult↗
PubMed1996

Long lasting immunological effects of ethanol after withdrawal.

The aim of the present study was to analyze on chronic alcoholic patients the effect of ethanol (EtOH) withdrawal on the immune system through the investigation of the distribution of PB lymphoid subsets, using multiple-stainings with monoclonal antibodies and flow cytometry. For this purpose a group of 20 patients with active alcoholism without liver disease, negative for hepatitis virus, and without malnutrition was analyzed and followed for 9 months after alcohol consumption had been discontinued. Twenty-five age- and sex-matched healthy volunteers were included in the study. The following panel of monoclonal antibodies combinations (FITC/PE/PerCP or PE-Cy5) was used: TCR alpha beta/CD3/HLA DR, CD25/CD56/CD3, TCR gamma delta/CD3/HLA DR, CD45RA/CD45R0/CD4, CD3/CD8, CD19/CD5, and CD3/CD11c. Analysis was performed on at least 1,500 events/tube at flow cytometry using the Lysys II software program. During the alcohol intake period, the most striking findings were a significant (P < 0.05) expansion of the CD8+ T-lymphocyte subset, which coexpresses the activation associated antigens HLA DR and CD11c, as well as a significant increase in both NK-cells (CD3-/CD56+) and the T-cell subset with NK activity coexpressing CD3 and CD56 (P < 0.05 and P < 0.01, respectively). In addition, a decrease in the CD5+ B-cells (P < 0.05), associated with reduced serum gamma-globulin levels, was also observed. During alcohol withdrawal, a rapid decrease towards normal values of activated CD8+/HLA DR+ and CD11c+ T-lymphocytes was observed as well as a normalization of CD19+/CD5+ B-cells and gamma-globulin serum levels; these changes might be directly related to EtOH suppression. Surprisingly, however, new immunological imbalances emerged in spite of the absence of alcohol intake. Thus, a progressive and significant expansion (P < 0.05) of CD4+ T-cells associated with an increased expression of the CD25 activation-related antigen and a preferential use of the CD45R0 isoform by CD4+ T-cells were observed. In parallel, there was an even more evident increase (P < 0.01) in the number of PB NK-cells. Our results show that EtOH consumption induces changes in the immune system, its effects persisting or even becoming more evident after suppression of EtOH intake for a 9 month period.

Alcoholism↗
PubMed1996

Multiparametric cell-cycle analysis of peripheral blood-activated lymphocyte subsets using staining based on the TdT method for incorporated BrdUrd.

This study describes a new method for the simultaneous assessment of the distribution of a cell population in the G0/G1, S, and G2/M cell-cycle phases by using multiparameter flow cytometry and single staining based on BrdUrd incorporation. Both the K562 cell line and PHA-stimulated peripheral blood lymphocytes (PBL) were analyzed. Cells were cultured in the presence of BrdUrd for 30 min prior to cell harvesting. Once collected, cells were exposed to ultraviolet light for 5 min and then fixed immediately in 70% ethanol (-20 degrees C) for at least 30 min. Once fixed, the cells were placed for 30 min at 37 degrees C in the presence of terminal deoxynucleotidyl transferase (TdT) and dUTP labeled with digoxigenin; they were then stained with FITC-labeled anti-digoxigenin. Our results show that G0/G1, s, and G2/M cell populations can be clearly discriminated according to FITC fluorescence and light-scatter parameters. In this way, S-phase cells can be identified by their FITC staining. From the cells which were negative for anti-digoxigenin-FITC antibody, two clear populations could be resolved in a forward scatter, side scatter, and fluorescence pulse-width three-dimensional plot; the values obtained for G0/G1 cells were lower than those obtained for G2/M cells in all three parameters. Multiparameter analysis of PBL stained for two surface antigens (CD3 and CD8) and for BrdUrd by direct or indirect TdT method permitted cell-cycle analysis of different subpopulations, including CD3+/CD8+, CD3+/CD8-, CD3-/CD8+, and CD3-/CD8-.

Bromodeoxyuridine↗
PubMed1996

Comparison of Vindelov et al. and bromodeoxyuridine/DNA double-staining flow cytometry methods for analysis of cell cycle distribution in rat thymocytes.

This study compares the cell cycle distribution in rat thymocytes obtained by means of bromodeoxyuridine (BrdUrd) labeling of S-phase cells and the analysis of the S-phase fraction obtained according to the technique of Vindelov et al. (Cytometry 3:332-338, 1983). The proportion of BrdUrd-labeled cells was analyzed in single cell suspensions of adult rat thymocytes after in vivo injection of BrdUrd and the results then compared with those obtained after measuring the cell DNA contents according to the Vindelov et al. method. The percentage of BrdUrd-positive cells was greater than the S-phase fraction obtained using the Vindelov et al. technique. By contrast, no major differences were observed between the percentage of BrdUrd-positive cells and the S-phase fraction obtained after analyzing the DNA histograms of the same data files with the RFIT mathematical model. The elimination of trypsin treatment used in the Vindelov et al. method did not alter the results, whereas the use of DNA denaturation with 2N HCl was shown to increase the percentage of S-phase rat thymocytes (calculated from DNA histograms) independently of whether trypsin treatment was used or not. However, the value of the S-phase fraction was not as great as that obtained after BrdUrd labeling. Thus when comparing BrdUrd-labeling and the Vindelov et al. technique, important differences in the percentage of S-phase adult rat thymocytes were observed. Selective G0/G1 cell loss during washing and centrifugation steps performed after the DNA denaturation used for BrdUrd detection was the main reason for these differences.

Animals↗
PubMed1996

Performance evaluation of the FACSCount System: a dedicated system for clinical cellular analysis.

Flow cytometers are instruments that can determine multiparameter data simultaneously and have a great potential in providing unique information about cells. The FACSCount System is designed as the first dedicated flow cytometer for the clinical laboratory. Its current configuration provides CD4, CD8, and CD3 absolute counts from 100 microliters of whole blood. Adapting the FACSCount System to the clinical setting are minimal sample handling, lysis free cell preparation, automated fluorescence gating, built-in calibrated reference beads, and appropriate error code reporting. Quality control checks ensure that reported CD4 counts, important for clinical follow-up and patient management, are accurate, precise, and reproducible across instruments over time.

Adolescent↗
PubMed1996

General concepts about cell sorting techniques.

Research involving cell analysis frequently requires isolation of certain cell types or subcellular components either as a final objective or as a preparative tool for further assays. At present, there are a high number of cell sorting methods that are suitable for being used in the clinical laboratory. These methods can be divided into two major groups: (1) bulk sorters and (2) single-cell-based sorters. This latter group mainly refers to fluorescence-activated cell sorting (FACS) by flow cytometry (FCM). In both cases, separation of cell subsets is based on their classification according to one or more cell characteristics. In bulk sorters, cell classification and sorting are usually achieved in a single step; by contrast, in FACS techniques, these two steps are independent sequential processes. In addition, bulk sorters generally use a single-cell characteristic to isolate cell subsets and have a higher throughput rate, as compared with FACS by FCM, where several parameters can be used simultaneously to classify cells for their further isolation. As a consequence of the mechanisms underlying these two cell sorting methods, the balance between cell purity and cell recovery on the sorted fraction are generally different, the single-cell-based methods usually providing both a higher purity and recovery. Thus, in practice, bulk separation methods are frequently used either as a preparative step for FCM-based cell sorting or for the enrichment of the sample in specific cell subsets, when a higher throughput rate is required; in contrast, FACS by FCM is selected for the isolation of cell subsets when a high purity and, especially, recovery of a specific subpopulation of cells present in a sample are needed.

Animals↗
PubMed1996

Astrocyte differentiation in primary culture followed by flow cytometry.

Astrocyte proliferation and differentiation in primary culture were followed by flow cytometry. The time-courses of the percentages of astrocytes in the different cell-cycle phases suggest that astrocytes proliferate during the first 10 days in culture thereafter reaching confluence. Two types of astrocytes are identified immunocytochemically: one growing in the bed monolayer, identified as type-1 astrocytes, and another growing on the top of the monolayer, identified as type-2 astrocytes. In addition, three populations identified as being formed of type-1, type-2 and putative progenitor cells, respectively, were followed by flow cytometry. Progenitor cells were the major type 2 h after plating (89%) but their percentage decreases sharply (to 16%) during the first 5 days in culture, with no ensuing changes. In contrast, the percentage of type-1 cells (11%) rapidly increased after plating reaching a maximum 5 days later (73%). Later, it decreased (to 47%) and was maintained thereafter. The percentage of type-2 cells was undetectable immediately after plating but increased from the 3rd to the 10th day with no further changes. Our results suggest that progenitor cells differentiate into type-1 astrocytes triggered by the culture medium but the differentiation of progenitor cells into type-2 astrocytes is brought about by some type-1-secreted factor. In this work we report a rapid and simple method for following the growth and differentiation of rat brain astrocytes in primary culture.

Animals↗
PubMed1996

Antigen phenotype and cytotoxic activity of natural killer cells in hemodialysis patients.

The antigen phenotype and cytotoxic activity of peripheral blood natural killer (NK) cells have been studied in 35 patients on hemodialysis, evaluating the influence exerted by different types of dialysis membranes and diverse clinical and biologic features of end-stage renal disease. Two-color immunofluorescent analyses with different monoclonal antibodies and flow cytometry were performed to identify the NK cells. Functional cytotoxic assays were simultaneously performed in the same patients. The results show that hemodialysis elicits higher proportions of CD3-/CD56+ and CD3-/CD16+ NK cells and a decrease in their cytotoxic activity compared with the control group. This alteration is related to the use of cuprophan membranes; biocompatible membranes would not significantly affect NK cells. The cuprophan membranes were also found to induce a higher degree of NK cell activation, measured as the number of CD16+/HLA-DR+ cells. Age appeared to be associated with NK cell function. Regarding age, in patients older than 65 years the change from cuprophan to biocompatible membranes was associated with a recovery of NK cytotoxic activity to normal levels. In summary, hemodialysis with cuprophan membranes induces an increased proportion of both NK cells and activated NK cells associated with decreased NK cytolytic activity.

Adult↗
PubMed1996

A randomized study comparing the effect of GM-CSF and G-CSF on immune reconstitution after autologous bone marrow transplantation.

Haemopoietic growth factors (HGFs) have been shown to accelerate recovery from severe neutropenia after autologous bone marrow transplantation (ABMT) but their effect on immune reconstitution is not well defined. The present study compares, through randomized trial, the in vivo effect of GM-CSF and G-CSF administration on the immune recovery of patients who underwent ABMT. For that purpose, we have sequentially analysed 14 different T, B and NK lymphoid cell subsets using appropriate dual staining during the first year following transplant (days +6, +17, +31, +66, +90, +120, +180, +360). 24 patients with lymphoproliferative disorders (20 lymphomas and four multiple myelomas) and who had undergone ABMT were included in the study. The median age was 43 years (range 22-62 years). All lymphoma patients were homogenously conditioned with BEAM. Our results show that both GM-CSF and G-CSF aid T-cell (CD3+/alpha beta) recovery though their contribution varies depending on the T-cell subset analysed. G-CSF contributed to a significantly faster recovery of CD8+ cells (P = 0.03). The CD8+ cell regeneration was produced mainly by activated cells (CD38+/HLA-DR+) which lacked the CD11b antigen. In contrast, GM-CSF favoured the regeneration of CD4+ cells (through both the CD45RO+ and CD45RA+ subset), leading to a higher CD4+:CD8+ ratio (P = 0.007). No statistically significant differences were detected in the three groups of patients as regards both the recovery of NK cells and NK activity. Furthermore, the use of HGF did not seem to exert a significant influence on the recovery of B lymphocytes. This recovery was based on the CD5+ subpopulation that showed a rapid rise after the first month. We suggest that G-CSF and GM-CSF not only influence myeloid recovery, but also regeneration of the immune system after ABMT.

Adult↗
PubMed1996

CD5+ B cells in Graves' disease: correlation with disease activity.

In the present paper the distribution of peripheral blood CD5+/CD19+ (CD5+B) and CD5-/CD19+ (CD5-B) B-lymphocytes in Graves' disease (GD) patients is analyzed in order to correlate them with disease activity, interleukin-2 (IL-2) and IL-6 binding to T and B cells as well as with anti-thyrotropin (TSH) receptor antibodies and the thyroid hormone serum levels. The combination of flow cytometry and 3-color immunofluorescence revealed a remarkable increase in the absolute numbers of CD5+ B cells in hyperthyroid-untreated GD patients (218 +/- 137 x 10(6)/l vs. 66 +/- 69 x 10(6)/l in healthy subjects, p < 0.01) that gradually fell to normal values once hyperthyroidism had been controlled by methimazole. However, relative numbers of CD5+ B cells persisted at a relatively stable but increased level in GD patients in long term remission of an average of 3.1 years. This was also confirmed in a follow-up study of a group of 12 newly diagnosed patients during the first 90 days of anti-thyroid drug therapy with methimazole. No correlation was observed between either CD5+ B cells or CD5- B cells and the serum levels of pathogenic anti-TSH receptor antibodies. Increased numbers of CD5+ B cells were related to the increased free thyroxine and total triiodothyronine serum levels. In addition, a strong correlation between both the absolute levels of B cells binding to exogenous IL-2 and IL-6 and the absolute number of CD5+ B cells in hyperthyroid GD patients (LR = 0.798, p < 0.001; LR = 0.569, p < 0.01, respectively) was observed. These results suggest that CD5+ B cells in GD are partly regulated by thyroid hormone serum levels as well as by IL-2 and IL-6 binding to B cells. Nevertheless, they are not involved, at least directly, in the production of anti-TSH receptor antibodies.

Adult↗
PubMed1996

Expression of the CD117 antigen (c-Kit) on normal and myelomatous plasma cells.

The surface expression of CD117 on plasma cells (PCs) from normal individuals and patients with multiple myeloma (MM) has been analysed using triple-stained cells for flow cytometry. In addition, the clinical significance of CD117 expression in MM patients and its possible value for the evaluation of minimal residual disease was explored. A total of 11 healthy volunteers and 56 untreated MM patients were included in the study. The expression of CD117 was analysed by flow cytometry, using simultaneous staining with the MAbs BB4, CD117 and CD38. Cell acquisition was performed in two consecutive steps using a live gate drawn on SSC/CD38 cells and a total of 300,000 events were acquired. For data analysis, the Paint-a-Gate Plus software (Becton Dickinson) was used. PCs were identified according to their strong reactivity for CD38 and their positivity for BB4, as well as by their light scatter distribution. Dilution experiments of CD117+ myelomatous PCs with normal bone marrow (BM) cells were performed in order to assess the sensitivity level of the technique for detection of CD117+ residual PCs. None of the PCs from normal BM samples showed reactivity for the CD117 antigen. In contrast, CD117 antigen was present in 18/56 MM patients (32%), the proportion of positive cells in these cases being as high as 92.1 +/- 9%. Therefore, within PC lineage the c-Kit antigen would be restricted to the myelomatous population and thus could be considered as a 'tumour-associated marker' for monitoring minimal residual disease in about one third of MM patients. Dilution experiments indicate that the detection limit with this marker would be 10(-4) (one myelomatous PC/10(4) normal BM cells). Upon comparing the clinical and haematological disease characteristics of CD117-positive and CD117-negative cases, no significant differences were found.

Adult↗
PubMed1996

Distribution of peripheral blood lymphoid subsets in alcoholic liver cirrhosis: influence of ethanol intake.

The aim of the present study was to investigate the effect of chronic ethanol (EtOH) consumption on the immune system in patients with alcoholic liver cirrhosis (ALC), as analyzed by the distribution of peripheral blood (PB-) T, B, and NK lymphoid subsets using multiple stainings with monoclonal antibodies and flow cytometry. For that purpose, we have analyzed a group of patients with ALC and active EtOH intake (ALCET group) which were re-evaluated 3 months after alcohol withdrawal. As controls, both ALC patients with at least 1 year of alcohol withdrawal (ALCAW group) and healthy subjects were used. Regarding the alcohol intake period, the most relevant findings were a significant activation of the PB T-cell compartment, and specifically of the TCR alpha beta + subset, as reflected by an increased expression of both the HLA DR and CD11c antigens as well as a significant increase of both the PB NK cells (CD3-/CD56+) and the cytotoxic T cells coexpressing the CD3 and CD56 molecules. In addition, a decrease of both the numbers of total B cells and their CD5+/CD19+ subset were observed. After a relatively short withdrawal period (3 months), the abnormalities of T, P, and NK cells disappeared. These findings suggest the existence of a close relationship between EtOH consumption and the abnormalities of the immune system observed during active alcoholism. Nevertheless, ALCAW individuals displayed marked alterations on the immunophenotypic profile, as reflected by a significantly decreased number of total T cells, due to reduced levels of the CD3+/TCR alpha beta+, CD4+, CD8+, and CD4+/CD45RA+ T-cell subsets. In addition, a significantly decreased number of total PB B cells was observed in this group of patients. Our results show that in patients suffering from ALC, the abnormalities of the immune system due to a direct effect of EtOH intake (or its metabolites) should be distinguished from the immunological alterations related to the liver disease itself.

Alcohol Drinking↗