Immune responses in vitro

Heber‐Katz, Ellen; Click, Robert E.

doi:10.1016/0008-8749(72)90067-6

Cited by 113 publications

(12 citation statements)

References 22 publications

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“…Whether the T-cell chains are associated in some noncovalent structure is uncertain. It is also uncertain whether the noncovalent linkage of Ig chains is related to the enhancing effect of sulfhydryl-containing reagents on the mixed lymphocyte reaction (18) and on the response of mouse spleen cells to immunization in vitro with sheep erythrocytes (19). There is an apparent discrepancy between our evidence for nondisulfide-linked structures and other evidence for a stable IgM monomer solubilized from the cell surface after iodination by the lactoperoxidase procedure (12).…”

Section: Resultscontrasting

confidence: 54%

Cell-Surface Immunoglobulin of Human Thymus Cells and Its Biosynthesis In Vitro

Moroz

Hahn

1973

Proc. Natl. Acad. Sci. U.S.A.

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Human thymus cells synthesize immunoglobulin in short-term culture, the nascent immunoglobulin appearing in both cytoplasmic and membrane fractions. Surface immunoglobulin was demonstrated by lactoperoxidase radioiodination of the cells. The demonstration of intracellular and surface immunoglobulin required procedures that minimize proteolytic degradation. Noncovalently linked, monomeric , chains and light chains were found in the cytoplasm and on the surface of the cells by means of acrylamide gel electrophoresis and by specific immunoprecipitation of the isolated chains.Mammalian bone marrow-derived lymphocytes (B cells) have membrane immunoglobulins (Ig) which are considered to be receptors for antigens and are indicative of the capacity of these cells to synthesize Igs (1-4). Thymus-derived lymphocytes (T cells) facilitate antibody production by B cells and are involved in cell-mediated immune reactions by means of specific antigen-binding receptors (5). Efforts to demonstrate membrane Ig on T cells have led to conflicting conclusions.In the present communication we report in evidenc shortterm tissue culture for synthesis by human thymus cells of IgM in the surface of these cells, and describe the immunochemical nature of the nascent cytoplasmic and surface immunoglobulin. Our findings emphasize the necessity for minimizing proteolytic degradation during the preparation of cellular and membrane fractions and suggest that variable proteolysis during solubilization of membrane components may account for conflicting reports on the presence or absence of surface Igs in T-cell membranes (6-11). MATERIALS AND METHODSIn Vitro Biosynthesis. Thymuses were obtained from 5-to 9-year-old children undergoing cardiac surgery. Cells were teased from the thymus, filtered through a stainless steel screen, and washed three times and resuspended in Eagle's medium containing 1:100 the standard amount of amino acids. Cell suspensions (5 X 107 cells per ml) were prein- Fig. 3, below), 50,000 K.I.U. of Trasylol (Bayer, 5 ml, 250,000 K.I.U., Kalikrein inactivation units) per liter and 0.2 M iodoacetamide were added to the STKM buffer. The cells were disrupted by homogenization at 40 and fractionated into "cytoplasmic fraction" and "membrane fraction" (13). (2)

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Section: Resultscontrasting

confidence: 54%

Cell-Surface Immunoglobulin of Human Thymus Cells and Its Biosynthesis In Vitro

Moroz

Hahn

1973

Proc. Natl. Acad. Sci. U.S.A.

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“…In 1970, Fanger and colleagues showed that cysteine, glutathione, or sulfite ion at mM concentrations in the presence of 20% fetal calf serum markedly enhanced the response of lymphocytes to transforming agents [11]. In 1972, Click and colleagues reported that 2-mercapto- ethanol (MER) at micromolar concentrations caused a 2- to 3-fold stimulation of antibody production [12] and T cell proliferation [13]. This finding was soon expanded to other immune systems and other sulfur compounds such as α-thioglycerol (TGL) [14].…”

Section: Sulfur As a Regulatory Agentmentioning

confidence: 99%

Thiosulfoxide (Sulfane) Sulfur: New Chemistry and New Regulatory Roles in Biology

Toohey¹,

2014

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The understanding of sulfur bonding is undergoing change. Old theories on hypervalency of sulfur and the nature of the chalcogen-chalcogen bond are now questioned. At the same time, there is a rapidly expanding literature on the effects of sulfur in regulating biological systems. The two fields are inter-related because the new understanding of the thiosulfoxide bond helps to explain the newfound roles of sulfur in biology. This review examines the nature of thiosulfoxide (sulfane, S0) sulfur, the history of its regulatory role, its generation in biological systems, and its functions in cells. The functions include synthesis of cofactors (molybdenum cofactor, iron-sulfur clusters), sulfuration of tRNA, modulation of enzyme activities, and regulating the redox environment by several mechanisms (including the enhancement of the reductive capacity of glutathione). A brief review of the analogous form of selenium suggests that the toxicity of selenium may be due to over-reduction caused by the powerful reductive activity of glutathione perselenide.

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“…Broome and Jeng, in 1972, reported a requirement for sulfhydryl compounds in certain murine leukemia cells in vitro (2,3). Other reports describe enhancement by sulfhydryl compounds of blast transformation (4,5), of antibody formation (6)(7)(8)(9), and of viability (2,8) in lymphocyte cultures. This report describes a study of the sulfhydryl dependence of some hematopoietic cells and the inhibitory effect of vitamin B12 compounds on these cells.…”

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confidence: 97%

Sulfhydryl dependence in primary explant hematopoietic cells. Inhibition of growth in vitro with vitamin B12 compounds.

Toohey¹

1975

Proc. Natl. Acad. Sci. U.S.A.

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Primary explants of P388, EL-4, and L1210 murine leukemia cells and of normal mouse bone marrow are shown to require sulfhydryl compounds for proliferation in vitro. Nine established cell lines show no stimulation by these compounds. Leukemia cells can lose the sulfhydryl dependence after various periods of adaptation to in vitro culture. Various sulfhydryl compounds have widely differing potencies in promoting in vitro proliferation of dependent cells. The effect appears to be specific for sulfhydryl groups in the reduced form. Vitamin B12 compounds inhibit the growth of sulfhydryl-requiring cells, apparently by catalyzing the oxidation of the sulfhydryl groups.Sulfhydryl compounds have received relatively little attention as essential nutrients for cells in vitro. In 1971, Park et al. (1) briefly described stimulation by cysteine and glutathione of plasmacytoma growth in vitro. Broome and Jeng, in 1972, reported a requirement for sulfhydryl compounds in certain murine leukemia cells in vitro (2, 3). Other reports describe enhancement by sulfhydryl compounds of blast transformation (4, 5), of antibody formation (6-9), and of viability (2,8) in lymphocyte cultures. This report describes a study of the sulfhydryl dependence of some hematopoietic cells and the inhibitory effect of vitamin B12 compounds on these cells. MATERIALS AND METHODSCell Culture. Eagle's minimum essential medium (MEM, ref. 10) was supplemented with 5-20% fetal calf serum (as indicated), 30 mg of glutamine/100 ml, 5 mg of asparagine/100 ml, 10 mg of serine/100 ml, and 11 mg of sodium pyruvate/100 ml. No antibiotics were added. In experiments involving a series of variable additives, a single homogeneous culture was used to pour all plates in the series. Variable additives were sterilized by filtration and placed in the plastic culture dishes before the culture was added.P388 murine leukemia cells (11) and L1210 murine leukemia cells (12) were obtained from peritoneal aspirates of DBA/2 mice which were inoculated 4-6 days previously with 106 cells. EI-4 murine leukemia cells (13) were obtained in a similar manner from C57 mice. Leukemia cells were cultured in 1.5-ml cultures in liquid medium in 35 X 10-mm Falcon plastic plates with an initial cell density of 1 X 105 cells per ml. After incubating at 370 in 5% CO2 for the indicated number of days (usually 3 days), the cells were suspended by swirling and diluted either 1 to 2 in trypan blue for counting in a hemocytometer or 1 to 800 in Isoton for counting in a Coulter counter.The For bone marrow culture, the medium was made at two times the normal concentration and mixed with an equal volume of 0.6% agar at 370 before adding fresh bone marrow cells obtained from the femurs of white Swiss Webster mice. Colony-stimulating factor was provided as an extract of gravid C3H or DBA/2 mouse uterus prepared by the method of Bradley et al. (21). Cultures of 1.5 ml were plated as a single layer in 35 X 10-mm Falcon plastic plates. Clones of more than 20 cells were counted as colonies on the seve...

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Immune responses in vitro

Cited by 113 publications

References 22 publications

Cell-Surface Immunoglobulin of Human Thymus Cells and Its Biosynthesis In Vitro

Cell-Surface Immunoglobulin of Human Thymus Cells and Its Biosynthesis In Vitro

Thiosulfoxide (Sulfane) Sulfur: New Chemistry and New Regulatory Roles in Biology

Sulfhydryl dependence in primary explant hematopoietic cells. Inhibition of growth in vitro with vitamin B12 compounds.

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