scholarly journals STUDIES ON LYSOSOMES

1968 ◽  
Vol 37 (2) ◽  
pp. 394-411 ◽  
Author(s):  
Günter Brittinger ◽  
Rochelle Hirschhorn ◽  
Steven D. Douglas ◽  
Gerald Weissmann
Keyword(s):  
Acid Phosphatase ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Structural Characteristics ◽  
Gradient Centrifugation ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Hydrolase Activity ◽  
Cell Fractionation ◽  
Acid Hydrolase

Pure suspensions of human lymphocytes were separated from peripheral blood by means of nylon wool, homogenized in 0.34 M sucrose-0.01 M EDTA solution, and fractionated by differential centrifugation. The bulk of acid hydrolase activity was found to be concentrated in a 20,000 g x 20 min granular fraction, whereas nuclear, debris, and supernatant fractions contained lesser concentrations of hydrolases. Acid hydrolase activity present in the granular fraction showed appropriate "latency" as judged by its dose-dependent release into the 20,000 g x 20 min supernatant after exposure to membrane-disruptive agents such as streptolysin S, filipin, and lysolecithin. Heparin proved to be necessary in the suspending medium so that reproducible homogenization and cell fractionation could be obtained. Even excessive contamination of lymphocyte suspensions with platelets did not appreciably alter the acid hydrolase activity of lymphocyte homogenates or the distribution of enzymes in subcellular fractions. Discontinuous density-gradient centrifugation of a 500 g x 10 min supernatant, containing both acid hydrolase-rich organelles and mitochondria, resulted in partial resolution of hydrolase-rich organelles from mitochondria. Fine structural studies of the intact lymphocytes showed the presence of acid phosphatase-positive, membrane-bounded organelles. Electron microscopy of the "large granule" (20,000 g x 20 min) fraction of such lymphocytes demonstrated 80–90% mitochondria, 5–10% platelets, and 5–10% membrane-bounded acid phosphatase-positive structures. The data indicate the presence in human peripheral blood lymphocytes of acid hydrolase-rich granules which possess many of the biochemical and structural characteristics of lysosomes in other tissues.

Intracellular free magnesium in human lymphocytes and the response to lectins

1991 ◽  
Vol 80 (6) ◽  
pp. 539-547 ◽  
Author(s):  
L. L. Ng ◽  
J. E. Davies ◽  
M. C. Garrido
Keyword(s):  
Phaseolus Vulgaris ◽  
Peripheral Blood ◽  
Proliferative Response ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Blood Lymphocytes ◽  
Fluorimetric Method ◽  
Human Peripheral Blood Lymphocytes ◽  
Free Magnesium

1. Intracellular free [Mg2+] was measured in human peripheral blood lymphocytes using a fluorimetric method based on the dye furaptra. It was necessary to correct for the extracellular leakage of the dye by using either 10 mmol/l EDTA or 0.05 mmol/l Mn2+. 2. As the proliferative response of lymphocytes to mitogenic lectins has been linked to a dependence on extracellular Mg2+, the intracellular [Mg2+] was studied in lymphocytes stimulated with various mitogenic and non-mitogenic lectins. 3. Only lymphocytes treated with phytohaemagglutinin-L, a leucoagglutinin from Phaseolus vulgaris that binds to tri- and tetra-antennary complex glycoproteins, showed a marked increase in intracellular [Mg2+]. This effect was partially inhibited by N-acetylgalactosamine. The stimulation by different lectins of the incorporation of [3H]-thymidine into lymphocytes was not correlated to the changes in intracellular [Mg2+]. 4. The proliferative response of lymphocytes to lectins is therefore not wholly dependent on a rise in intracellular [Mg2+].

scholarly journals Phenotypically and functionally distinct subpopulations of human lymphocytes with T cell markers also exhibit different cytochemical patterns of staining for lysosomal enzymes

Blood ◽  
1984 ◽  
Vol 63 (5) ◽  
pp. 1067-1071 ◽  
Author(s):  
A Landay ◽  
LT Clement ◽  
CE Grossi
Keyword(s):  
T Cell ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Suppressor Cells ◽  
Peripheral Blood Lymphocytes ◽  
Acid Hydrolases ◽  
Cytochemical Localization ◽  
Cell Markers ◽  
Granular Lymphocytes

Abstract Human peripheral blood lymphocytes that express T cell markers, when reacted for the cytochemical localization of lysosomal acid hydrolases, display two major patterns of staining, i.e., dot-like and scattered granular. Previous attempts to fractionate T cells according to surface markers have yielded populations of cells with heterogeneous patterns of cytochemical staining. In this study, peripheral blood cells forming rosettes with sheep erythrocytes have been fractionated by sequential staining with two monoclonal antibodies, D12 and 2D2, followed by fluorescence-activated cell sorting. These reagents have been shown previously to recognize a subpopulation of cells capable of suppressing T cell proliferation. All of the cells positive for D12 and 2D2 stained for acid hydrolases with the scattered granular pattern, whereas the large majority of the cells negative for both markers stained with the dot-like pattern. It is concluded that suppressor cells within the E+ cell fraction have the cytochemical characteristics of large granular lymphocytes.

scholarly journals The Transfer RNA Methylases of Human Lymphocytes. I. Induction by PHA in Normal Lymphocytes

Blood ◽  
1971 ◽  
Vol 37 (3) ◽  
pp. 282-292 ◽  
Author(s):  
DAVID H. RIDDICK ◽  
ROBERT C. GALLO
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Transfer Rna ◽  
Normal Peripheral Blood ◽  
Blood Lymphocytes ◽  
Normal Human ◽  
Total Capacity ◽  
Normal Human Peripheral Blood

Abstract Assays were performed that measured both the rate and extent (or total capacity) of transfer RNA methylases in extracts of lymphocytes cultured in the presence and absence of phytohemagglutinin (PHA). The tRNA methylases of human peripheral blood lymphocytes undergoing blastogenesis in culture with PHA had a five- to sixfold increase in rate and a three- to sevenfold increase in extent of methylation of heterologous tRNA. These data suggest that PHA transformed lymphocytes not only contain elevated levels of tRNA methylases, but that the increase includes qualitatively different enzymes from those found in normal peripheral blood lymphocytes. Experiments in which lymphocytes were incubated for various times with PHA revealed that tRNA methylase induction occurred late in or after DNA synthesis and after morphologic transformation, but prior to mitosis. Rate and extent of tRNA methylation increased simultaneously. PHA induction of tRNA methylase activity was dependent on the synthesis of new RNA in lymphocytes cultured from 40 to 45 hours. The increase was not due to different levels of inhibitors or activators or preferential degradation of reaction components. The data suggest that quantitative and qualitative changes occur in the tRNA methylases of the normal human peripheral blood lymphocyte stimulated by PHA to undergo transition to an undifferentiated cell "in cycle." The possible significance of these findings to control of protein synthesis in PHA transformed lymphocytes is discussed.

scholarly journals Genotoxic effects of diazinon on human peripheral blood lymphocytes / Genotoksično djelovanje diazinona na limfocite ljudske periferne krvi

2015 ◽  
Vol 66 (2) ◽  
pp. 153-158 ◽  
Author(s):  
Fulya Dilek Gökalp Muranli ◽  
Martin Kanev ◽  
Kezban Ozdemir
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Blood Lymphocytes ◽  
Human Peripheral Blood Lymphocytes ◽  
Current Assessment ◽  
High Level

Abstract The aim of this study was to evaluate the genetic damage in human peripheral blood lymphocytes following 24 and 48- hour exposure to a commercial diazinon formulation Basudin 60EM® at concentrations between 0.01 and 40 μg mL-1. For this purpose we used the micronucleus (MN), fluorescence in situ hybridization (FISH), and alkaline single cell gel electrophoresis (comet) assay. Diazinon significantly increased the frequency of micronucleated cells compared to control. Forty-eight-hour exposure increased this frequency even at lower concentrations (0.01-10 μg mL-1). The FISH results revealed aneugenic effects at 10 μg mL-1. The comet assay also confirmed DNA damage at concentrations between 10 and 40 μg mL-1. Our findings have confirmed the genotoxic potential of diazinon and its cytotoxic effect on human lymphocytes. The increased DNA damage in our study raises concern about the current assessment of the health risk posed by this pesticide and calls for a high level of caution in agricultural and household use.

scholarly journals A NEW SURFACE MARKER ON T LYMPHOCYTES OF HUMAN PERIPHERAL BLOOD

1973 ◽  
Vol 138 (5) ◽  
pp. 1270-1275 ◽  
Author(s):  
Sten Hammarström ◽  
Ulla Hellström ◽  
Peter Perlmann ◽  
Marie-Louis Dillner
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Helix Pomatia ◽  
Peripheral Blood Lymphocytes ◽  
Immunofluorescent Staining ◽  
Positive Staining ◽  
Cell Fractionation ◽  
Binding Studies ◽  
Neuraminidase Treatment ◽  
Surface Receptors

Neuraminidase treatment of human peripheral blood lymphocytes uncovers cell surface receptors that bind purified A hemagglutinin from the snail Helix pomatia. No hemagglutinin was bound to untreated lymphocytes. Binding studies with 125I-labeled hemagglutinin suggested that the number of receptors on neuraminidase-treated lymphocytes was approximately 1·106/cell. The apparent association constant for hemagglutinin binding to lymphocytes, as calculated from Scatchard's plots, was 5–7·108 liters/mol. Immunofluorescent staining with FITC-conjugated hemagglutinin gave positive reactions with approximately 60% of the lymphocytes from normal donors. Positive staining was inversely related to the number of lymphocytes with Fc or complement receptors or with surface immunoglobulin, thus suggesting that See PDF for Structure the lymphocytes with receptors for Helix pomatia A hemagglutinin are T cells. Cell fractionation on columns charged with hemagglutinin indicate that these receptors may be used for separating subpopulations of human peripheral lymphocytes.

scholarly journals Cytotoxic, clastogenic and genotoxic effects of cis-tetraammine(oxalato)ruthenium(III) dithionate on human peripheral blood lymphocytes

2020 ◽  
Vol 42 ◽  
pp. e50517
Author(s):  
Manuela da Rocha Matos Rezende ◽  
Vivianne de Souza Velozo-Sá ◽  
Cesar Augusto Sam Tiago Vilanova-Costa ◽  
Elisangela Silveira-Lacerda
Keyword(s):  
Comet Assay ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Low Frequency ◽  
Peripheral Blood Lymphocytes ◽  
Negative Control ◽  
Blood Lymphocytes ◽  
Human Peripheral Blood Lymphocytes

There is a concern about stablishing the clinical risk of drugs used for cancer treatment. In this study, the cytotoxic, clastogenic and genotoxic properties of cis-tetraammine(oxalato)ruthenium(III) dithionite - cis-[Ru(C2O4)(NH3)4]2(S2O6), were evaluated in vitro in human lymphocytes. The mitotic index (MI), chromosomal aberrations (CA) and DNA damage by comet assay were also analyzed. The MTT test revealed that the ruthenium compound showed a slight cytotoxic effect at the highest concentration tested. The IC50 value for the compound after 24 hours of exposure was 185.4 µM. The MI values of human peripheral blood lymphocytes treated with 0.015, 0.15, 1.5 and 150 µM of cis-[Ru(C2O4)(NH3)4]2(S2O6) were 6.1, 3.9, 3.2 and 0.2%, respectively. The lowest concentration, 0.015 µM, did not show any cytotoxic activity. The CA values for the 0.015, 0.15 and 1.5 µM concentrations presented low frequency (1.5, 1.6 and 2.3%, respectively), and did not express clastogenic activity when compared to the negative control, although it was observed clastogenic activity in the highest concentration tested (150 µM). The results obtained by the comet assay suggest that this compound does not present genotoxic activity at lower concentrations. The results show that cis-[Ru(C2O4)(NH3)4]2(S2O6) has no cytotoxic, clastogenic or genotoxic in vitro effects at concentrations less than or equal to 0.015 µM. This information proves as promising in the treatment of cancer and is crucial for future trials.

scholarly journals Phenotypically and functionally distinct subpopulations of human lymphocytes with T cell markers also exhibit different cytochemical patterns of staining for lysosomal enzymes

Blood ◽  
1984 ◽  
Vol 63 (5) ◽  
pp. 1067-1071
Author(s):  
A Landay ◽  
LT Clement ◽  
CE Grossi
Keyword(s):  
T Cell ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Suppressor Cells ◽  
Peripheral Blood Lymphocytes ◽  
Acid Hydrolases ◽  
Cytochemical Localization ◽  
Cell Markers ◽  
Granular Lymphocytes

Human peripheral blood lymphocytes that express T cell markers, when reacted for the cytochemical localization of lysosomal acid hydrolases, display two major patterns of staining, i.e., dot-like and scattered granular. Previous attempts to fractionate T cells according to surface markers have yielded populations of cells with heterogeneous patterns of cytochemical staining. In this study, peripheral blood cells forming rosettes with sheep erythrocytes have been fractionated by sequential staining with two monoclonal antibodies, D12 and 2D2, followed by fluorescence-activated cell sorting. These reagents have been shown previously to recognize a subpopulation of cells capable of suppressing T cell proliferation. All of the cells positive for D12 and 2D2 stained for acid hydrolases with the scattered granular pattern, whereas the large majority of the cells negative for both markers stained with the dot-like pattern. It is concluded that suppressor cells within the E+ cell fraction have the cytochemical characteristics of large granular lymphocytes.

Cytotoxic Effects of Trimethyltin Chloride on Human Peripheral Blood Lymphocytes in vitro

1989 ◽  
Vol 8 (5) ◽  
pp. 349-353 ◽  
Author(s):  
B.B. Ghosh ◽  
G. Talukder ◽  
A. Sharma
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Sister Chromatid Exchanges ◽  
Blood Lymphocytes ◽  
Human Peripheral Blood Lymphocytes ◽  
Abnormal Cells ◽  
Trimethyltin Chloride

Trimethyltin chloride was found to induce cytotoxic damage in vitro in human peripheral blood lymphocytes. Two concentrations (0.5 μg and 1.0 μg) were added to lymphocytes from male and female subjects in mitogen stimulated and serum supplemented culture medium for 72 h. A dose-related increase of inhibition of replication index (P < 0.01) and cell division (P < 0.001) was observed. The frequencies of abnormal cells and chromosomal aberrations such as chromatid and chromosome breaks, dicentrics, triradial and quadriradial configurations were increased significantly (P < 0.001), as were micronucleus counts (P < 0.001) and sister chromatid exchanges (P < 0.001). Endoreduplication, an extremely rare spontaneous event in human lymphocytes, was observed in a few cases at all dose levels. The effects were more pronounced in lymphocytes obtained from habitual smokers.

scholarly journals IDIOTYPIC DETERMINANTS OF IMMUNOGLOBULIN M DETECTED ON THE SURFACE OF HUMAN LYMPHOCYTES BY CYTOTOXICITY ASSAYS

1972 ◽  
Vol 136 (3) ◽  
pp. 650-655 ◽  
Author(s):  
P. Wernet ◽  
T. Feizi ◽  
H. G. Kunkel
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Significant Proportion ◽  
Peripheral Blood Lymphocytes ◽  
Immunoglobulin M ◽  
Blood Lymphocytes ◽  
Cytotoxicity Assays ◽  
Human Peripheral Blood Lymphocytes ◽  
Surface Immunoglobulin

The complement-mediated cytotoxicity assay has been used to demonstrate surface immunoglobulin determinants on human peripheral blood lymphocytes. In two individuals with monoclonal serum IgM components, idiotypic antisera demonstrated similar components on a significant proportion of the peripheral blood lymphocytes.

scholarly journals Appearance of Hydrolase Rich Granules in Human Lymphocytes Induced by Phytohemagglutinin and Antigens

Blood ◽  
1967 ◽  
Vol 30 (1) ◽  
pp. 84-102 ◽  
Author(s):  
ROCHELLE HIRSCHHORN ◽  
KURT HIRSCHHORN ◽  
GERALD WEISSMANN
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Hydrolytic Enzyme ◽  
Total Activity ◽  
Peripheral Blood Lymphocytes ◽  
Streptolysin S ◽  
Stimulation Of

Abstract Purified human peripheral blood lymphocytes have been shown to develop acid hydrolase-rich granules between 24 and 48 hours after stimulation by phytohemagglutinin, prior to mitosis. This increase has been measured biochemically as a net increase in total activity of the lysosomal enzymes: acid β-glycerophosphatase, acid phenolpthalein phosphatase, and aryl sulfatase. The subcellular localization of acid hydrolytic enzyme activities has been investigated, and they have been shown to be concentrated in a large granule fraction of sucrose homogenates and to behave as if they were membrane-bounded, in that their activity could be released by lysolecithin. It has also been demonstrated by histochemical technics that stimulation of lymphocytes by antigen (PPD) and by streptolysin S, as well as by phytohemagglutinin, produced an increase in acid phosphatase activity. Chloroquine, an inhibitor of the response to phytohemagglutinin, has also been shown to inhibit the development of acid phosphatase activity. These results are interpreted to suggest that both specific and nonspecific stimulants of lymphocytes induce lysosome-like structures in premitotic cells.

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