Characterization of suppressor T cells induced with the thymus-independent antigen polyvinylpyrrolidone coupled to syngeneic cells

CAF₁ mice injected iv with polyvinylpyrrolidone (PVP) coupled to syngeneic spleen cells (PVP-SC) and challenged several days later with 0.25 μg PVP produced fewer PVP-specific IgM plague-forming cells (PFC) than mice injected with Mock-SC. Both 10,000 and 360,000 MW PVP could induce unresponsiveness after coupling to SC. The unresponsiveness induced by PVP-SC was shown to be mediated, at least in part, by antigen-specific suppressor T cells (T_S). The PVP-specific T_S were I-J positive and belonged to the Lyt 1⁺ 2⁺ subset of T cells. The T_s precursors were sensitive to 20 mg/kg cyclophosphamide (Cy) and to antilymphocyte serum (ALS). Kinetics studies suggested that unresponsiveness induced by PVP-SC may be of two types since unresponsiveness in the intact animal appeared earlier and did not last as long as detectable T_S activity.     

Functional heterogeneity among the T-derived lymphocytes of the mouse: III. Helper and suppressor T-cells activated by concanavalin A

We have studied the properties of T-cells which when activated by concanavalin A (Con A) either suppress or help the in vitro humoral response of mouse spleen cells. Previously established criteria for the T-cell populations, T₁ and T₂ were applied. T₁ cells were defined by their short half-life (2–3 wk) after adult thymectomy (ATx) and their resistance to small doses of antithymocyte serum (ATS). T₂ cells were defined by their long half-life (~15 wk) and their high sensitivity to ATS. T-cells which could be activated by Con A to help the response to the thymus-dependent antigen, sheep red blood cells, were found mainly in the T₂ subpopulation. T-cells which could be activated by Con A to suppress the response to the thymus-independent antigen, trinitrophenyl-lipopolysaccharide (TNP-LPS), were found within both the T₁ and T₂ subpopulations.These results, our previous results, and those of others suggest that the T-cell responses to phytomitogens distinguish precursors committed to different functions, while the T₁ and T₂ classifications distinguish T-cells at different stages of maturation.     

Control of mitogen-induced transformation: characterization of a splenic suppressor cell and its mode of action.

The present study demonstrates that mouse spleen cells contain a population of glass wool adherent T lymphocytes which exhibit the capacity to suppress non-glass adherent lymphocyte responses to mitogens. These suppressor cells are stimulated by both low and high doses of PHA¹ and high doses of con A. The suppressor cell effect is observed when UNA, but not RNA or protein synthesis, is studied. This glass-adherent suppressor cell population is characterized as being the primary DNA synthesizing cells during the early (0–8 hr) stages of culture. Suppression still occurs when the suppressor cells are treated with mitomycin C, actinomycin D or cycloheximide. This implies that new macromolecular synthesis may not be required for suppression to occur. Suppression is blocked by inhibiting synthesis of prostaglandin and is mimicked by Bu₂cAMP. This suggests that mitogen activated suppressor cells regulate T cell responses via production of prostaglandin which modulates the concentration of intracellular cyclic nucleotide levels.     

Characterization of the la antigens involved in suppressor T cell generation in the rat

The WRC rat, an intra-class II recombinant strain (RT1.B ⁿ B ^a D _, ^a ), was used to study the relative roles of the two class II loci in mixed lymphocyte reaction (MLR) proliferation and suppressor T cell (Ts) generation. Both MLR proliferation and Ts generation were noted in cultures of WRC with DA (RTI ^{a) stimulator cells. In contrast, cultures of WRC with BN (RT1sun} ) stimulator cells proliferate but do not generate significant amounts of Ts. The data suggest that RT1.B incompatibility is important in the generation of Ts in the WRC rat. Suppressor cells generated in cultures of WF (RT1 ^u ) with WRC stimulator cells potently suppressed a WF+WRC_x test MLR, with less suppression when tested against either the WF+DA_x or WF+BN_x MLRs. The latter experiments suggest that Ts clones may be produced to either class II subregion, and therefore that MLR proliferation and Ts induction are not necessarily linked, but vary with particular genotypes. The current lack of other rat intra-class II recombinant strains precludes assignment of suppressor induction/activation to a single locus.     

Analysis of ultraviolet light-induced suppressor mutations in the strain of Escherichia coli K-12 AB1157: an implication for molecular mechanisms of UV mutagenesis

Summary Genetic analysis of histidine independent (His⁴) revertants induced by ultraviolet light in the his-4 E. coli strain AB1157 was carried out: 83% carried ochre (UAA) suppressor mutations and 17% carried back mutations to his ⁺ or (intragenic?) suppressors not detectably separable from his-4. Using the specialized transducing psu 2int ^– phage, which carries supE-supB, it was determined that 87% of the ochre suppressors mapped in the supE-supB region. We were able to deduce that 56% of these affected tRNA ₁ ^Gln by a CAATAA change in the tRNA gene while 31% affected tRNA ₂ ^Gln by TAGTAA change. Although we were unable to deduce the base substitution of the remaining 13%, the results indicated that most of the suppressor mutations are caused by a G:C to A:T transition.These results suggest that the high incidence of supE-supB region suppressor mutation in E. coli by UV would be a reflection of the general feature of UV mutagenesis; i.e. preferential induction of G:C to A:T transition in repairing nonparing DNA lesions.     

Mutant of the glutamine transfer RNA gene as UGA suppressor in Escherichia coli

Summary A UGA suppressor derived from a glutamine tRNA gene of Escherichia coli K 12 was isolated and characterized. Phages carrying the suppressor su⁺2_UGA could be obtained only from a hybrid transducing phage, h ⁸⁰ cI ₈₅₇psu ⁺2_oc, but not from the original transducing phage cI ₈₅₇psu ⁺2_oc. By DNA sequence analysis, it was found that the su ⁺2 UGA suppressor obtained has two mutations; one is in the anticodon (TTATCA), as expected, and the other (CT) is at the 7th position from the 3 end of tRNA ₂ ^Gln . The significance of these mutations and the lethal effect on phage of the increased amounts of UGA suppressor tRNAs are discussed.     

Effect of melphalan in vitro on induction of murine suppressor T cells by ConA

Summary The effect of treatment with melphalan in vitro on the activity of spleen cells from BALB/c mice was investigated. Incubation of spleen cells with 1.5–5 g melphalan/1×10⁷ inhibited subsequent mitogenic stimulation by ConA or PHA and the allogeneic response of BALB/c spleen cells against C57B1 target spleen cells. Incubation of spleen cells with ConA led to induction of suppressor T cells which when added to fresh cultures inhibited the allogeneic response. Preincubation of spleen cells with melphalan even at low concentrations (0.15–0.5 g 1×10⁷ cells) which do not directly affect mitogenic stimulation or allogeneic response partially inhibited the generation of suppressor T cells by ConA. Treatment with melphalan had no effect on already induced suppressor T cells as shown by incubation of spleen cells with melphalan (0.15–5 g/1×10⁷ cells) after incubation with ConA. Addition of cells treated with melphalan alone (without ConA) to fresh cultures led to an increase in the allogeneic response.     

Regulatory function of T lymphocytes in the immune response to polyvinyl pyrrolidone. I. Two categories of suppressor T cells.

The regulation of antibody response of mice to polyvinyl pyrrolidone (PVP) was investigated using three preparations of PVP (K90, K30, and K15) differing from each other in molecular weight. The immunogenicity of PVP was higher as the molecular weight increased. The depletion of thymus-derived cells resulted in the augmentation of anti-PVP response. On the other hand, the response of intact mice to the most immunogenic PVP (K90) was suppressed more or less by the injection of any preparation of PVP 4 days prior to K90. This was most pronounced when the smallest PVP (K15) was preinjected. The suppression, however, was not observed in thymectomized-irradiated-bone marrow reconstituted mice.These results indicated that anti-PVP response was regulated by two different categories of thymus-derived cells, that is, “intrinsic” and “induced” suppressor cells. The activity of the latter was transferrable, PVP-specific, and eliminated by anti-Thy 1 serum and complement. In addition, the mean affinity of anti-PVP plaque-forming antibodies was found to be reduced by the action of “induced” suppressor cells.     

Characteristics of DTH suppressor cells in mice infected with Candida albicans

Inoculation of 10⁸ C. albicans intraperitoneally into Balb/c mice at given dosage was reported to induce suppression of antigen-specific delayed-type hypersensitivity. Adoptive transfer of spleen cells into normal syngeneic mice pre-treated with Cyclophosphamide confirmed the existence of suppressor cells in mice. Such cells were sensitive to treatment with anti- serum and complement, non-adherent to Sephadex G-10. A pretreatment of the mice with Cyclophosphamide eliminated DTH suppression. Treatment with antimacrophage agents via intraperitoneal abrogated suppression only if being effected before inoculation of alive 10⁸ Candida albicans.It is concluded that the spleen suppressor cell is a T-lymphocyte whose precursor is Cyclophosphamide-sensitive, requiring the macrophage to be induced.     

<Emphasis Type="Italic">Azohydromonas riparia</Emphasis> sp. nov. and <Emphasis Type="Italic">Azohydromonas ureilytica</Emphasis> sp. nov. isolated from a riverside soil in South Korea

White and pale yellow coloured bacteria were isolated from the riverside soil, Daejeon, South Korea, and were designated UCM-11^T, UCM-F25, and UCM-80^T. We found that all strains were able to reduce nitrate, and the cells were aerobic and motile. The DNA G+C contents of UCM-11^T, UCM-F25, and UCM-80^T were between 68.9 to 71.2 mol% and the main ubiquinone was observed as Q-8. Based on16S rRNA gene sequences, strains UCM-11^T and UCM-F25 were found to closely match with Azohydromonas australica IAM 12664^T (98.48–98.55%), and the strain UCM-80^T was the closest match with Azohydromonas lata IAM 12599^T (98.34%). The presence of summed feature 3 (C_16:1 ω7c and/or C_16:1 ω6c), C_16:0, summed feature 8 (C_18:1 ω7c and/or C_18:1 ω6c) as well as twokinds of hydroxyfatty acids consisting of C_10:0 3-OH and C_12:0 2-OH, and branched fatty acids containing C_16:0 iso and C_17:0 cyclo were detected in all the strains. Phosphatidylethanolamine was a major polar lipid. DNA–DNA relatedness confirmed UCM-11^T, UCM-F25 and UCM-80^T as novel members of the genus Azohydromonas. Based on the morphological, physiological, biochemical and genotypic characteristics, we suggest that strains UCM-11^T, UCM-F25, and UCM-80^T represent novel species within the genus Azohydromonas. The names Azohydromonas riparia sp. nov., and Azohydromonas ureilytica sp. nov. are proposed for the type strains UCM-11^T (=KACC 18570^T =NBRC 111646^T) and UCM-80^T (=KACC 18576^T =NBRC 111658^T), respectively.     

Immune amplification of murine CD8 suppressor T cells induced via an immune-privileged site: quantifying suppressor T cells functionally

Background

CD8⁺ suppressor T cells exert antigen-specific suppression of the expression of hypersensitivity by activated T cells. Therefore, CD8⁺ suppressor T cells serve a major regulatory role for the control of active immunity. Accordingly, the number and/or activity of CD8⁺ suppressor T cells should be influenced by an immune response to the antigen. To test this hypothesis we used an adoptive transfer assay that measures the suppression of the expression of delayed-type hypersensitivity (DTH) by CD8⁺ suppressor T cells to quantify the antigen-specific suppression of DTH by these suppressor T cells.

Methods

Suppressor T cells were induced in the spleens of mice by the injection of antigen into the anterior chamber of an eye. Following this injection, the mice were immunized by the same antigen injected into the anterior chamber. Spleen cells recovered from these mice (AC-SPL cells) were titrated in an adoptive transfer assay to determine the number of AC-SPL cells required to effect a 50% reduction of antigen-induced swelling (Sw50) in the footpad of immunized mice challenged by antigen.

Results

Suppression of the expression of DTH is proportional to the number of AC-SPL cells injected into the site challenged by antigen. The number of AC-SPL cells required for a 50% reduction in DTH-induced swelling is reduced by injecting a cell population enriched for CD8⁺ AC-SPL cells. Immunizing the mice receiving intracameral antigen to the same antigen decreases the RSw50 of AC-SPL cells required to inhibit the expression of DTH.

Conclusions

The results provide the first quantitative demonstration that the numbers of antigen-specific splenic CD8⁺ suppressor T cells are specifically amplified by antigen during an immune response.     

In vitro expression of secondary antitumor immunity by in vivo tumor-sensitized T cells: Inhibition by tumor-induced suppressor T cells

Summary In vitro cultivation of memory immune cells from P815- or P388-immune mice with corresponding irradiated tumor cells induced generation of cytolytic T cells (CTL). The induction of CTL generation, as well as the cytolytic activity itself, was tumor-specific. The in vitro generation of CTL from P815- or P388-immune cells was suppressed by spleen cells from mice bearing corresponding progressive tumors (tumor size 15 mm). The tumor-induced suppressor cells suppressed the in vitro generation of CTL, but did not affect their cytolytic function. The suppression was tumor-specific and was mediated by Ly1⁺2^–L3T4⁺ T cells. Treatment of suppressor cell donors with cyclophosphamide or sublethal -radiation completely abolished the ability of their spleen cells to inhibit the in vitro CTL generation.     

<Emphasis Type="Italic">Hymenobacter agri</Emphasis> sp. nov., a novel bacterium isolated from soil

A bacterial isolate was recovered from a soil sample collected in Jeollabuk-do Province, South Korea, and subjected to polyphasic taxonomic assessment. Cells of the isolate, designated strain S1-2-1-2-1^T, were observed to be rod-shaped, pink in color, and Gram-stain negative. The strain was able to grow at temperature range from 10 to 30 °C, with an optimum of 25 °C, and growth occurred at pH 6–8. Comparative 16S rRNA gene sequence analysis showed that strain S1-2-1-2-1^T belongs to the genus Hymenobacter, with closely related type strains being Hymenobacter daeguensis 16F3Y-2^T (95.8% similarity), Hymenobacter rubidus DG7B^T (95.8%), Hymenobacter soli PB^T (95.7%), Hymenobacter terrenus MIMtkLc17^T (95.6%), Hymenobacter terrae DG7A^T (95.3%), and Hymenobacter saemangeumensis GSR0100^T (95.2%). The genomic DNA G+C content of strain S1-2-1-2-1^T was 63.0 mol%. The main polar lipid of this strain was phosphatidylethanolamine, the predominant respiratory quinone was menaquinone-7, and the major fatty acids were C_15:0 iso (27.3%), summed feature 3 (C_16:1 ω7c/C_16:1 ω6c) (16.5%), C_15:0 anteiso (15.3%), and C_16:0 (14.7%), supporting the affiliation of this strain with the genus Hymenobacter. The results of this polyphasic analysis allowed for the genotypic and phenotypic differentiation of strain S1-2-1-2-1^T from recognized Hymenobacter species. On the basis of its phenotypic properties, genotypic distinctiveness, and chemotaxonomic features, strain S1-2-1-2-1^T is considered to represent a novel species of the genus Hymenobacter, for which the name Hymenobacter agri sp. nov. is proposed. The type strain is S1-2-1-2-1^T (=KCTC 52739^T?=?JCM 32194^T).     

<Emphasis Type="Italic">Hymenobacter daeguensis</Emphasis> sp. nov. isolated from river water

A Gram-stain-negative, non-motile, non-spore-forming, rod-shaped, aerobic bacterial strain, designated 16F3Y-2^T, was isolated from the Han River, South Korea, and was characterized taxonomically using a polyphasic approach. Comparative 16S rRNA gene sequence analysis showed that strain 16F3Y-2^T belonged to the family Cytophagaceae in the phylum Bacteroidetes and was most closely related to ‘Hymenobacter terrae’ DG7A (98.01%), H. soli PB17^T (97.26%), H. glaciei VUG-A130^T (96.78%), H. antarcticus VUG-A42aa^T (96.72%), H. ruber PB156^T (96.61%), and H. saemangeumensis GSR0100^T (95.77%). The G+C content of the genomic DNA of strain 16F3Y-2^T was 62.9 mol%. The isolate contained MK-7 as the predominant respiratory quinone, and summed feature 3 (C_16:1 ω7c/C_16:1 ω6c; 35.5%), C_15:0 iso (16.9%), C_16:1 ω5c (10.9%), and C_15:0 anteiso (9.9%) as major fatty acids. The major polar lipid was phosphatidylethanolamine. Phenotypic and chemotaxonomic data supported the affiliation of strain 16F3Y-2^T with the genus Hymenobacter. However, strain 16F3Y-2^T exhibited relatively low levels of DNA-DNA relatedness with ‘H. terrae’ KCTC 32554 (44.1%) and H. soli KCTC 12607^T (24.3%), clearly indicating that the isolate constitutes a new genospecies. Strain 16F3Y-2^T could be differentiated from its phylogenetic neighbors on the basis of several phenotypic, genotypic, and chemotaxonomic features. Therefore, strain 16F3Y-2^T represents a novel species in the genus Hymenobacter, for which the name Hymenobacter daeguensis sp. nov. is proposed. The type strain is 16F3Y-2^T (=KCTC 52537^T =JCM 31654^T).     

<Emphasis Type="Italic">Flavihumibacter profundi</Emphasis> sp. nov., isolated from eutrophic freshwater sediment

A Gram-stain-positive, aerobic, non-motile, non-spore-forming, and rod-shaped bacterium, designated strain CHu64-6-1^T, was isolated from a 67-cm-long sediment core collected from the Daechung Reservoir at a water depth of 17-m in Daejeon, Republic of Korea. Comparative 16S rRNA gene sequence studies placed the new isolate in the class Sphingobacteriia, and the isolate is notably most closely related to Flavihumibacter sediminis CJ663^T (98.1% similarity), Flavihumibacter solisilvae 3-3^T (97.8%), Flavihumibacter petaseus T41^T (97.5%), Flavihumibacter cheonanensis WS16^T (97.4%), and Flavihumibacter stibioxidans YS-17^T (97.2%). The cells of strain CHu64-6-1^T formed yellow colonies on R2A agar and contained MK-7 as the only menaquinone, phosphatidylethanolamine, an unidentified phospholipid, and two unidentified aminolipids as the major polar lipids, and C_15:0 iso, C_17:0 iso 3-OH, C_15:1 iso G, and C_16:1ω5c as the major fatty acids (> 5%). The DNA G + C content of the genome was determined to be 46.5 mol%. The DNA-DNA hybridization values of strain CHu64-6-1^T with F. sediminis CJ663^T, F. solisilvae 3-3^T, F. petaseus T41^T, F. cheonanensis WS16^T, and F. stibioxidans YS-17^T were 12.4–33.2%. Based on the combined genotypic and phenotypic data, we propose that strain CHu64-6-1^T represents a novel species of the genus Flavihumibacter, for which the name Flavihumibacter profundi sp. nov. is proposed. The type strain is CHu64-6-1^T (= KCTC 62290^T = CCTCC AB 2018060^T).     

T1 and T2 lymphocytes in primary and secondary delayed type hypersensitivity of mice. I. Contribution in the response to sheep red blood cells and to allogeneic spleen cells.

Peripheral T lymphocytes can be subdivided into two populations (T₁ and T₂ cells) based upon the short life span of T₁ cells after adult thymectomy (ATx) and sensitivity of T₂ cells to treatment with anti-thymocyte serum (ATS) in vivo. The contribution of the T₁ and T₂ cells to primary and secondary delayed type hypersensitivity (DTH) to sheep red blood cells (SRBC) and to primary DTH to allogeneic cells was studied in mice. T₂ cells were found to account for the development of the state of primary DTH responsiveness after intravenous immunization with SRBC and after subcutaneous immunization with allogeneic cells. No clear cut evidence was found that in the presence of T₂ cells DTH related T effector cells were generated from T₁ cells. In mice selectively depleted for T₁ cells by ATx, the remaining T₂ cells were capable to generate SRBC-specific T memory cells, but not in numbers as large as in non-thymectomized mice. On the other hand, T₁ cells in mice depleted for T₂ cells by ATS treatment, could give rise to normal numbers of SRBC-specific T memory cells. Apparently T₁ cells can compensate for the absence of T₂ cells during generation of T memory cells, but T₂ cells cannot do so for the loss of T₁ cells. From the time curve showing the ATx-induced decline of the population of SRBC-specific T₂ cells, involved in primary DTH responsiveness, the half life was calculated to be 6 to 7 months.     

<Emphasis Type="Italic">Flavobacterium zaozhuangense</Emphasis> sp. nov., a new member of the family <Emphasis Type="Italic">Flavobacteriaceae</Emphasis>, isolated from metolachlor-contaminated soil

Strain ZZ-8^T, a Gram-negative, aerobic, non-spore-forming, non-motile, yellow-pigmented, rod-shaped bacterium, was isolated from metolachlor-contaminated soil in China. The taxonomic position was investigated using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain ZZ-8^T is a member of the genus Flavobacterium and shows high sequence similarity to Flavobacterium humicola UCM-46^T (97.2%) and Flavobacterium pedocola UCM-R36^T (97.1%), and lower (<?97%) sequence similarity to other known Flavobacterium species. Chemotaxonomic analysis revealed that strain ZZ-8^T possessed MK-6 as the major respiratory quinone; and iso-C_15:0 (28.5%), summed feature 9 (iso-C_17:1 w9c/C_16:0 10-methyl, 22.9%), iso-C_17:0 3-OH (17.0%), iso-C_15:0 3-OH (8.9%), iso-C_15:1 G (8.6%) and summed feature 3 (C_16:1 w7c/C_16:1 w6c, 5.7%) as the predominant fatty acids. The polar lipids of strain ZZ-8^T were determined to be lipids, a glycolipid, aminolipids and phosphatidylethanolamine. Strain ZZ-8^T showed low DNA–DNA relatedness with F. pedocola UCM-R36^T (43.23?±?4.1%) and F. humicola UCM-46^T (29.17?±?3.8%). The DNA G+C content was 43.3 mol%. Based on the phylogenetic and phenotypic characteristics, chemotaxonomic data and DNA–DNA hybridization, strain ZZ-8^T is considered a novel species of the genus Flavobacterium, for which the name Flavobacterium zaozhuangense sp. nov. (type strain ZZ-8^T?=?KCTC 62315 ^T?=?CCTCC AB 2017243^T) is proposed.     

Concomitant induction of cytotoxic and suppressor cells: modulation by theophylline

Previous studies in man have shown that T cells with suppressor activity were mainly found among a subset bearing Fc receptors for IgG (T_γ). Recently, we found that virus-induced cytotoxic effector cells were also found predominantly among T_γ cells. In the present studies, we present evidence that similar, possibly overlapping T-cell populations can mediate both suppressor and cytotoxic activities when sensitized in vitro with virus-infected cells. In fact, both activities are found within the positively selected T_γ subset, but not in the T_γ-depleted population; both activities are abolished by irradiation but not by treatment with mitomycin C; a 1-hr exposition to theophylline at the onset of sensitization enhances both cytotoxic and suppressor activities. The data suggest that development of antiviral cell-mediated immune responses in vivo may also be accompanied by a concurrent induction of nonspecific suppressor cells. Such suppressor activity may play a role in the depressed cellular immune responsiveness which is associated with several systemic virus infections.     

Inhibition of rat mixed lymphocyte cultures by suppressor macrophages.

Normal rat spleens contain suppressor cells which can inhibit proliferative and cytotoxic responses of lymphocytes to alloantigens in vitro. The suppressor cells are adherent, phagocytic, resistant to treatment with ATS and C, radioresistant, resistant to treatment with mitomycin C, apparently absent from the thymus, and found in very high concentrations in peritoneal exudates. These characteristics indicate that the suppressor cell is a macrophages and not a T cell. When suppressor cells were removed from spleen cell suspensions, strong in vitro proliferative and cytotoxic responses to alloantigens could consistently be observed.