EVIDENCE THAT THE L-ASPARAGINASE OF GUINEA PIG SERUM IS RESPONSIBLE FOR ITS ANTILYMPHOMA EFFECTS : II. LYMPHOMA 6C3HED CELLS CULTURED IN A MEDIUM DEVOID OF L-ASPARAGINE LOSE THEIR SUSCEPTIBILITY TO THE EFFECTS OF GUINEA PIG SERUM IN VIVO

Cells of the original line of lymphoma 6C3HED, which regularly prove susceptible to the effects of guinea pig serum in vivo, were cultured in Eagle''s medium devoid of L-asparagine; after a latent period of 2 or more weeks, during which time the cell population declined markedly, some of the cells began to proliferate, and thereafter continued vigorous growth. On implantation into mice the proliferating cells were found, however, to have completely and permanently lost their susceptibility to the effects of guinea pig serum. By contrast, when cultures of the original line of 6C3HED cells were prepared in Eagle''s medium to which L-asparagine was added in a concentration of 20.0 mg/liter or more, they proliferated vigorously from the beginning; after long periods of growth in the enriched medium in vitro they remained susceptible to the effects of guinea pig serum upon test in vivo. Other amino acids, purines, and pyrimidines were unable to substitute for L-asparagine in this relation. Furthermore, a variant subline of 6C3HED cells which had become insensitive to guinea pig serum under in vivo conditions did not require L-asparagine for growth in tissue culture. It seems plain from the findings as a whole, that in 6C3HED cells, L-asparagine dependence in vitro is associated with the in vivo character of guinea pig serum sensitivity, and conversely L-asparagine independent variants are insusceptible to the effects of guinea pig serum. The implications of the findings complement those of a companion paper in which direct evidence is provided that the L-asparaginase of guinea pig serum is responsible for its antilymphoma effects.     

EVIDENCE THAT THE L-ASPARAGINASE OF GUINEA PIG SERUM IS RESPONSIBLE FOR ITS ANTILYMPHOMA EFFECTS : I. PROPERTIES OF THE L-ASPARAGINASE OF GUINEA PIG SERUM IN RELATION TO THOSE OF THE ANTILYMPHOMA SUBSTANCE

A number of the properties of the L-asparaginase present in guinea pig serum have been examined and shown to be indistinguishable from those of the agent responsible for inhibiting cells of lymphoma 6C3HED in vivo. The patterns of instability of the enzyme to changes in temperature and pH were found to parallel closely those of the antilymphoma agent. L-Asparaginase activity was essentially absent from the serum of newborn guinea pigs and this failed to inhibit 6C3HED cells. On separating guinea pig serum proteins by salt precipitation, electrophoresis, and chromatography on DEAE cellulose, antilymphoma activity was found only in fractions which contained L-asparaginase.     

STUDIES ON ANTIGENICITY : THE RELATIONSHIP BETWEEN IN VIVO AND IN VITRO ENZYMATIC DEGRADABILITY OF HAPTEN-POLYLYSINE CONJUGATES AND THEIR ANTIGENICITIES IN GUINEA PIGS

The enzymatic degradation of fluorescein conjugates of poly-L-lysine, poly-D-lysine, and exhaustively succinylated poly-L-lysine by aqueous extracts of spleens from "responder" (guinea pigs which can develop immune responses to hapten-poly-L-lysine conjugates) and "non-responder" guinea pigs was investigated. The in vivo degradation of H³-tagged dinitrophenyl conjugates of these synthetic polyamino acids was also studied by measuring urinary excretion of radioactive low molecular weight degradation products of these conjugates after their intraperitoneal injection. It was found that both responder and non-responder guinea pigs can degrade succinylated and unsuccinylated poly-L-lysine conjugates into small molecular fragments, but they cannot degrade hapten-poly-D-lysine conjugates. These studies demonstrate that in addition to the known requirements for antigenicity of macromolecules, i.e. the presence of antigenic determinants, and their capacity to be degraded by immunological tissues, the resulting degradation products must undergo certain additional, as yet unidentified, specific metabolic steps in order to induce an immune response.     

Genetic control of the immune response. A selective defect in immunologic (IgG) memory in nonresponder mice

The kinetics of antibody formation after immunization with the synthetic polypeptide poly-L(Tyr, Glu)-poly-D, L-Ala--poly-L-Lys [(T, G)-A--L] in aqueous solution were studied in genetically high (H-2^b) and low (H-2^k) responder strains of mice. During the 1st wk after immunization both strains developed brisk primary responses consisting of IgM antibody. With subsequent antigen challenge, only the high responder mice showed immunological memory, producing high titers of IgG antibody. In contrast, the low responder mice continued to make a persistent low level of IgM antibody and appeared unreactive to secondary or tertiary antigen challenge. These data are consistent with the hypothesis that the immune response-1 gene [controlling response to (T, G)-A--L] exerts its effect on the immune response at the time of switchover from IgM to IgG antibody production.     

Anticyanobacterial effect of l-lysine on Microcystis aeruginosa

Cyanobacterial blooms can cause serious environmental problems and threaten aquatic organisms and human health. It is therefore essential to effectively control cyanobacterial blooms in aquatic ecosystems. In the present study, the anticyanobacterial effect of l-lysine on Microcystis aeruginosa was examined. The results showed that the growth of M. aeruginosa (>90%) was effectively inhibited by l-lysine at dosages of 5.0, 6.5, and 8.0 mg L⁻¹ after 3 d treatment. The content of superoxide anion radicals, MDA content and SOD activity in M. aeruginosa cells increased after 1 d of treatment with l-lysine (3.0, 5.0, 6.5, and 8.0 mg L⁻¹), revealing that l-lysine induced oxidative stress in the cyanobacterial cells. The chlorophyll-a and protein contents in M. aeruginosa treated with l-lysine (3.0, 5.0, 6.5, and 8.0 mg L⁻¹) decreased after 2 d, indicating damage of the photosynthetic system by l-lysine treatment. Additionally, the production of exopolysaccharide by M. aeruginosa also increased and the expression of polysaccharide synthesis genes was upregulated by 3.0 mg L⁻¹l-lysine after 3 d of treatment. In response to the algicidal effects of l-Lysine, M. aeruginosa upregulated exopolysaccharide synthesis. Electron microscopic observations demonstrated that the cell membrane of M. aeruginosa was broken down during treatment with l-lysine (≥3.0 mg L⁻¹). Our results revealed that the effects of l-lysine on M. aeruginosa cells were comprehensive, and l-lysine is therefore an efficient anticyanobacterial reagent.

l-lysine had an anticyanobacterial effect on Microcystis aeruginosa, which involved growth inhibition, physiological and metabolic disturbance, and cell membrane damage.     

DIPLOID HETEROZYGOUS HYBRIDS FROM MATINGS BETWEEN ESCHERICHIA COLI AND SALMONELLA TYPHOSA

An Hfr strain of E. coli K-12 has been shown to mate at low frequency with a number of strains of S. typhosa. The hybrids, selected as lactose positives, retained all other antigenic and biochemical manifestations of the S. typhosa parent. A Lac⁺ hybrid, S. typhosa strain 643L⁺ was remated with the E. coli Hfr and plated on minimal media containing L-arabinose, D-xylose, L-rhamnose, or L-fucose as the sole carbon sources. Hybrids of the remated strain appeared at high frequency on the plates containing L-arabinose, and could be detected at lower frequencies on plates containing D-xylose, L-rhamnose, and L-fucose. Those selected for D-xylose or L-rhamnose utilization possessed the attributes of segregating diploid heterozygotes being highly unstable and continually segregating a cultural form typical of the S. typhosa parent. The unstable type exhibited most of the biochemical characteristics of the E. coli parent including the ability to produce indol, and also reacted with antiserum to both the E. coli and S. typhosa parent strains, owing to the acquisition of a thermolabile antigen from the E. coli parent. The parent and hybrid strains were examined in detail for changes in patterns of phage susceptibility and virulence. Acquisition of susceptibility to a number of the T phages, a characteristic of the E. coli parent, was observed in one of the hybrid types. A decrease in virulence of the diploid hybrid form of S. typhosa in the mouse virulence test was found.     

Selective extraction of trivalent actinides using CyMe4BTPhen in the ionic liquid Aliquat-336 nitrate

The extraction of Am(iii), Cm(iii) and Eu(iii) by 2,9-bis(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-1,2,4-benzotriazin-3-yl)-1,10-phenanthroline (CyMe₄BTPhen) from nitric acid solution was studied using the ionic liquid Aliquat-336 nitrate ([A336][NO₃]) as diluent. Results show a high selectivity of the solvent for Am(iii) and Cm(iii) over Eu(iii), but rather slow extraction kinetics. The kinetics of CyMe₄BTPhen were largely improved by the addition of 0.005 mol L⁻¹N,N,N′,N′-tetra-n-octyl-diglycolamide (TODGA) as a phase transfer reagent and by the use of 1-octanol as co-diluent. The addition of the phase transfer catalyst and co-diluent did not compromise the selectivity towards the actinide/lanthanide separation and thus this four-component system can be successfully applied to separate Am(iii) and Cm(iii) from the lanthanides.

Improving the kinetics of selective An(iii) extraction from nitric acid feed solutions into an ionic liquid based solvent: combining CyMe₄BTPhen with TODGA in Aliquat-336 nitrate.     

Requirement for alpha-L-fucose on the macrophage membrane receptor for MIF

α-L-fucose abolishes the activity of guinea pig migration inhibitory factor (MIF) on the macrophages. Other sugars such as α-D-glucose, β-D-galactose, α-L-rhamnose, methyl-α-D-mannoside, and N-acetyl-β-D-glucosamine had no effect. Theabolition of MIF activity by α-L-fucose was reversible. When macrophages were incubated with α-L-fucosidase, a glycosidase which splits terminal α-L-fucose from oligosaccharides, the macrophages no longer responded to MIF. On the other hand, MIF incubated with α-L-fucosidase was still active. These experiments strongly suggest that α-L-fucose comprises an essential part of a macrophage membrane receptor for MIF.     

N-Acetyl-l-leucine-polyethylenimine-mediated miR-34a delivery improves osteogenesis and bone formation in vitro and in vivo

Oral bone defects are difficult to treat. Recently, endogenous miR-34a was shown to be involved in bone anabolism. Clinical application of such microRNAs requires the inherent instability of microRNAs to be overcome by an efficient delivery system. In this study, we employed N-acetyl-l-leucine-modified polyethylenimine (N-Ac-l-Leu-PEI) as an miR-34a carrier and evaluated its delivery ability, transfection efficiency, cytotoxicity and whether it enhanced osteogenic differentiation and bone formation in vitro and in vivo. Stable N-Ac-l-Leu-PEI/miR-34a nanocomplexes were synthesized at a mass ratio of 4 and had a small size (190.34 nm), a low zeta potential (21.1 mV), a high transfection efficiency (69.39%) and no cytotoxicity in MG63 cells. N-Ac-l-Leu-PEI-mediated miR-34a delivery in vitro promoted ALP activity and expression of osteogenic differentiation markers, Runx2, SP7 and ColI to higher levels than those produced by Lipofectamine 2000-mediated delivery. N-Ac-l-Leu-PEI also achieved delivery of miR-34a in vivo to a local cranial bone defect area with miR-34a retaining the ability to initiate significant new bone formation 12 weeks post-implantation. This demonstrates the potential for N-Ac-l-Leu-PEI as a gene therapy vehicle for the regeneration of bone defects.

We employ N-acetyl-l-leucine-modified polyethylenimine as an miR-34a carrier and evaluate its delivery ability, transfection efficiency, cytotoxicity and whether it enhances osteogenic differentiation and bone formation in vitro and in vivo.     

Positive effects of concomitant heavy metals and their reduzates on hexavalent chromium removal in microbial fuel cells

Cr(vi) laden wastewaters generally comprise a range of multiple heavy metals such as Au(iii) and Cu(ii) with great toxicity. In the present study, cooperative cathode modification by biogenic Au nanoparticles (BioAu) reduced from aqueous Au(iii) and in situ Cu(ii) co-reduction were investigated for the first time to enhance Cr(vi) removal in microbial fuel cells (MFCs). With the co-existence of Cu(ii) in the catholyte, the MFC with carbon cloth modified with nanocomposites of multi-walled carbon nanotubes blended with BioAu (BioAu/MWCNT) obtained the highest Cr(vi) removal rate (4.07 ± 0.01 mg L⁻¹ h⁻¹) and power density (309.34 ± 17.65 mW m⁻²), which were 2.73 and 3.30 times as high as those for the control, respectively. The enhancements were caused by BioAu/MWCNT composites and deposited reduzates of Cu(ii) on the cathode surface, which increased the adsorption capacity, electronic conductivity and electrocatalytic activity of the cathode. This study provides an alternative approach for efficiently remediating co-contamination of multiple heavy metals and simultaneous bioenergy recovery.

The cooperative cathode modification by BioAu from Au(iii) and in situ Cu(ii) co-reduction enhanced Cr(vi) removal and bioelectricity generation in MFCs.     

Study on mechanism of elemene reversing tumor multidrug resistance based on luminescence pharmacokinetics in tumor cells in vitro and in vivo

While elemene (ELE) can reverse tumor multidrug resistance (MDR), the mechanisms for ELE reversing MDR remain unclear. Numerous studies have suggested that the efflux functionality of ATP-binding cassette (ABC) transporters, not their quantity, is more relevant to tumor MDR. However, no appropriate methods exist for real-time detection of the intracellular drug efflux caused by ABC transporters in vitro, especially in vivo, which hinders the examination of MDR reversal mechanisms. This study directly investigates the correlation between efflux functionality of ABC transporters and MDR reversal via ELE, using d-luciferin potassium salt (d-luc) as the chemotherapeutic substitute to study the intracellular drug efflux. Here, a luciferase reporter assay system combined with bioluminescence imaging confirmed that the efflux of d-luc from MCF-7/DOX^Fluc cells in vitro and in vivo was significantly reduced by ELE and when combined with Doxorubicin (DOX), ELE showed a synergistically anti-tumor effect in vitro and in vivo. Additionally, the luminescence pharmacokinetics of d-luc in MCF-7/DOX^Fluc cells and pharmacodynamics of the combined ELE and DOX in vivo showed a great correlation, implying that d-luc might be used as a probe to study ABC transporters-mediated efflux in order to explore mechanisms of traditional Chinese medicines reversing MDR.

The correlation between efflux functionality of ATP-binding cassette transporters and tumor multidrug resistance reversing via elemene was investigated using bioluminescence imaging (BLI) technology and luciferase reporter gene technology.     

Biosorption of Cr(vi) from aqueous solution using dormant spores of Aspergillus niger

Spores of Aspergillus niger (denoted as A. niger) were used as a novel biosorbent to remove hexavalent chromium from aqueous solution. The effects of biosorbent dosage, pH, contact time, temperature and initial concentration of Cr(vi) on its adsorption removal were examined in batch mode. The Cr(vi) uptake capacity increased with an increase in Cr(vi) concentration until saturation, which was found to be about 97.1 mg g⁻¹ at pH 2.0, temperature of 40 °C, adsorbent dose of 2.0 g L⁻¹ and initial concentration of 300 mg L⁻¹. Scanning electron microscopy, energy dispersive X-ray spectroscopy, field-emission transmission electron microscopy (FETEM), XPS and Fourier-transform infrared spectroscopy were applied to study the microstructure, composition and chemical bonding states of the biomass adsorbent before and after spore adsorption. The mechanisms of chromate anion removal from aqueous solution by the spores of A. niger were proposed, which included adsorption of Cr(vi) onto the spores followed by its reduction to Cr(iii). The reduced Cr(iii) was rebound to the biomass mainly through complexation mechanisms, redox reaction and electrostatic attraction. The removal of Cr(vi) by spores of A. niger followed pseudo-second-order adsorption kinetics. Monolayer adsorption of Cr(vi) was revealed by the better fitting of the Langmuir model isotherm rather than multilayer adsorption for the Freundlich model. The results indicated that A. niger spores can be used as a highly efficient biosorbent to remove Cr(vi) from contaminated water.

Spores of Aspergillus niger (denoted as A. niger) were used as a novel biosorbent to remove hexavalent chromium from aqueous solution.     

The fate of peptides pinocytosed by macrophages in vitro

A series of small peptides, such as might arise in the course of intralysosomal protein digestion, were screened for the ability to escape, intact, from mouse peritoneal macrophage lysosomes. Inability to penetrate lysosomal membranes was inferred from a peptide''s induction of lysosomal swelling, or vacuolization, in cultured macrophages. Two of the peptides tested, (D-Glu)₂ and (D-Ala)₃, induced vacuolization. Neither peptide was susceptible to hydrolysis by enzymes in macrophages or in the serum-containing culture medium. Their morphological effect was inhibited by parafluorophenylalanine, an inhibitor of pinocytosis. Once formed by either peptide, the vacuoles persisted for several hours in peptide-free medium. Quantitative studies of radioactively labeled (D-Glu)₂ confirmed the morphological evidence that (D-Glu)₂ is taken up by pinocytosis and stored, intact, in macrophage lysosomes. The majority of the peptides which failed to induce vacuolization—(L-Ala)₂, L-Ser·L-Ala, L-Val·L-Ala, L-Ala·L-Thr, Gly·D, L-Phe, L-Ala·D-His, (L-Ala)₃, (L-Glu)₂, and D-Leu·L-Tyr—were found to be susceptible to hydrolysis by cellular or serum peptidases. Their failure to induce vacuolization was attributed to their hydrolysis to subunits capable of penetrating lysosomal membranes. Some of the peptides which had failed to induce vacuolization—(D-Ala)₂, D-Ser·D-Ala, D-Val·D-Ala, Gly-D-Asn, D-Ala·D-Thr, and D-Arg·D-Val—were found to be indigestible. Except for the cytotoxic peptide D-Arg·D-Val, peptides in this category all had lower molecular weights and volumes than (Glu)₂ or (Ala)₃. It is inferred that these peptides are small enough to escape from macrophage lysosomes, while (Glu)₂ and (Ala)₃ are too large to escape intact. The implications of this inference for the mechanism of intracellular digestion of pinocytosed proteins are discussed.     

A simple and cost-effective paper-based and colorimetric dual-mode detection of arsenic(iii) and lead(ii) based on glucose-functionalized gold nanoparticles

We report a simple and cost-effective paper-based and colorimetric dual-mode detection of As(iii) and Pb(ii) based on glucose-functionalized gold nanoparticles under optimized conditions. The paper-based detection of As(iii) and Pb(ii) is based on the change in the signal intensity of AuNPs/Glu fabricated on a paper substrate after the deposition of the analyte using a smartphone, followed by processing with the ImageJ software. The colorimetric method is based on the change in the color and the red shift of the localized surface plasmon resonance (LSPR) absorption band of AuNPs/Glu in the region of 200–800 nm. The red shift (Δλ) of the LSPR band observed was from 525 nm to 660 nm for As(iii) and from 525 nm to 670 nm for Pb(ii). The mechanism of dual-mode detection is due to the non-covalent interactions of As(iii) and Pb(ii) ions with glucose molecule present on the surface AuNPs, resulting in the aggregation of novel metal nanoparticles. The calibration curve gave a good linearity range of 20–500 μg L⁻¹ and 20–1000 μg L⁻¹ for the determination of As(iii) and Pb(ii) with the limit of detection of 5.6 μg L⁻¹ and 7.7 μg L⁻¹ for both metal ions, respectively. The possible effects of different metal ions and anions were also investigated but did not cause any significant interference. The employment of AuNPs/Glu is successfully demonstrated for the determination of As(iii) and Pb(ii) using paper-based and colorimetric sensors in environmental water samples.

We report a simple and cost-effective paper-based and colorimetric dual-mode detection of As(iii) and Pb(ii) based on glucose-functionalized gold nanoparticles under optimized conditions.     

A comparative study on the coordination of diglycolamide isomers with Nd(iii): extraction,third phase formation,structure, and computational studies

A novel asymmetric diglycolamide N,N-dimethyl-N′,N′-dioctyl diglycolamide (L^II) was synthesized. The Nd(iii) extraction behavior from HNO₃ and loading capability of the solution of L^II in 40/60 (v/v)% n-octanol/kerosene were studied. Analyses by the slope method, ESI-MS, and FT-IR indicated that, similar to the previously studied isomer ligand N,N′-dimethyl-N,N′-dioctyl diglycolamide (L^I), 1 : 3 Nd(iii)/L^II complexes formed. Under the same experimental conditions, the distribution ratio and limiting organic concentration of L^II towards Nd(iii) were smaller than those of L^I, but the critical aqueous concentration of L^II was larger, which implies that L^II exhibited poorer extraction and loading capabilities towards Nd(iii) than L^I, and L^II has a tendency to be less likely to form the third phase. The quasi-relativistic density functional theory (DFT) calculation was performed to provide some explanations for the differences in their extraction behaviors. The electrostatic potential of the ligands indicated that the electron-donating ability of the amide O atoms in L^II displayed certain differences compared with L^I. This inhomogeneity in L^II affected the interaction between L^II and Nd(iii), as supported by QTAIM and bonding nature analysis, and it seemed to reflect in the extraction performance towards Nd(iii).

The inhomogeneous interactions of M–O_amide in the L^II ligand result in differences between the metal-ion extraction performances of two isomeric ligands.     

Adsorption of cadmium by live and dead biomass of plant growth-promoting rhizobacteria

Plant growth-promoting rhizobacteria (PGPR) have been extensively investigated in combination remediation with plants in heavy metal contaminated soil. However, being biosorbent, few studies of live and dead cells of PGPR have been undertaken. Meanwhile, the application of live or dead biomass for the removal of heavy metals continues to be debated. Therefore, this study uses living and non-living biosorbents of Cupriavidus necator GX_5, Sphingomonas sp. GX_15, and Curtobacterium sp. GX_31 to compare their Cd(ii) adsorption capacities by SEM-EDX, FTIR, and adsorption experiments. In the present study, whether the cells were living or dead and whatever the initial Cd(ii) concentration was, removal efficiency and adsorption capacity can be arranged as GX_31 > GX_15 > GX_5 (p < 0.05). However, removal efficiency in live and dead biosorbents was quite different and it greatly affected by the initial Cd(ii) concentrations. The dead cells exhibited a higher adsorption capacity than the live cells of GX_31. Nevertheless, for GX_5 and GX_15, the loading capacity of the non-living biomass was stronger than that of the living biomass at 20 mg L⁻¹ of Cd(ii), but the capacity was similar at 100 mg L⁻¹ of Cd(ii). Minor changes of spectra were found after autoclaving and it seemed that more functional groups of the dead biosorbent were involved in Cd(ii) binding by FTIR analysis, which also illustrated that the hydroxyl, amino, amide, and carboxyl groups played an important role in complexation with Cd(ii). Based on these findings, we concluded that the dead cells were more potent for Cd(ii) remediation, especially for GX_31.

Plant growth-promoting rhizobacteria (PGPR) have been extensively investigated in combination remediation with plants in heavy metal contaminated soil.     

An efficient mercapto-functionalized organic–inorganic hybrid sorbent for selective separation and preconcentration of antimony(iii) in water samples

This work reported on the application of mercapto-functionalized silica-supported organic–inorganic hybrid sorbent as a solid phase extraction (SPE) extractant for effective separation and preconcentration of Sb(iii) species in real water samples. The influences of pH, sorbent amounts, flow rates and the concentration of eluent on the adsorption and desorption of Sb(iii) species had been evaluated. The recovery of Sb(iii) species at pH 5 with 100 mg mercapto-functionalized hybrid sorbent at the flow rate of 5.0 mL min⁻¹ was greater than 95% without interference from all of metal ions tested. The trapped Sb(iii) species by extractant was then eluted with 5% HCl solution at the flow rate of 5.0 mL min⁻¹. The proposed procedure permitted large enrichment factors of about 200 and higher for 10 μg L⁻¹ of Sb(iii) species. The merits of analytical figures for the determination of Sb(iii) species were as follows: detection limit (3σ, n = 11), 2 ng L⁻¹; precision, 1.6% (n = 11) for 10 μg L⁻¹ of Sb(iii) species; the linear calibration curve presented in the concentration range of 1.0–200.0 μg L⁻¹. The validity of the proposed procedure was checked by the analysis of standard reference materials. Excellent agreement between the analytical results and the certified values (t-test at 95% confidence level) was found. The mercapto-functionalized hybrid sorbent as a SPE extractant was applied to the determination of Sb(iii) species in various water samples with satisfactory results.

This work reported on the application of mercapto-functionalized silica-supported organic–inorganic hybrid sorbent as a solid phase extraction (SPE) extractant for effective separation and preconcentration of Sb(iii) species in real water samples.     

Antigen-specific thymus cell factors in the genetic control of the immune response to poly-(tyrosyl, glutamyl)-poly-D, L-alanyl--poly-lysyl

The genetic control of the antibody response to a synthetic polypeptide antigen designated poly-L(Tyr, Glu)-poly-D,L-Ala--poly-L-Lys [(T, G)-A--L] has been studied in congenic high responder C3H.SW (H-2^b) and low responder C3H/HeJ (H-2^k) strains of mice. This response is controlled by the Ir-1 gene and is H-2 linked. The method employed was to study the ability of specifically primed or "educated" T cells of each strain to produce cooperative factors for (T, G)-A--L in vitro. Such factors have been shown to be capable of replacing the requirement for T cells in the thymus-dependent antibody response to (T, G)-A--L in vivo. The T-cell factors produced were tested for their ability to cooperate with B cells of either high or low responder origin by transfer together with bone marrow cells and (T, G)-A--L into heavily irradiated, syngeneic (for bone marrow donor) recipients. Direct anti-(T, G)-A--L plaque-forming cells were measured later in the spleens of the recipients. The results showed that (a) educated T cells of both high and low responder origin produced active cooperative factors to (T, G)-A--L, and no differences between the strains in respect to production of T-cell factors could be demonstrated; and (b) such factors, whether of high or low responder origin, cooperated efficiently with B cells of high responder origin only, and hardly at all with B cells of low responder origin. The conclusion was drawn that the cellular difference between the two strains lies in the responsiveness of their B cells to specific signals or stimuli received from T cells. As far as could be discerned by the methods used, no T-cell defect existed in low responder mice and the expression of the controlling Ir-1 gene was solely at the level of the B cells in this case.     

CELL INTERACTIONS BETWEEN HISTOINCOMPATIBLE T AND B LYMPHOCYTES : IV. INVOLVEMENT OF THE IMMUNE RESPONSE (Ir) GENE IN THE CONTROL OF LYMPHOCYTE INTERACTIONS IN RESPONSES CONTROLLED BY THE GENE

In the present study we have asked the question of whether F₁ carrier-primed T cells can serve as helper cells for either or both parental B cells when (a) the carrier molecule employed is under genetic control such that one parental strain is a responder and the other is a nonresponder, and (b) the determinant specificity of the parental B cells being assessed is not under genetic control and bears no relationship to the specificity of the carrier molecule. Utilizing the system of immune response gene control of responses to the terpolymer L-glutamic acid-L-lysine-L-tyrosine (GLT) to which A strain mice (H-2^a) are nonresponders, whereas BALB/c (H-2^d) and (BALB/c x A)F₁ hybrids (CAF₁) are responders, these studies demonstrate that GLT-primed T cells of CAF₁ donors can provide for responder BALB/c, but not for nonresponder A/J, the required stimulus for the anti-DNP responses of DNP-specific B cells of these respective parental strains to the DNP conjugate of GLT. The implications of these findings for Ir gene function in physiologic T-B cell interactions are discussed in detail.     

Carrier function in anti-hapten antibody responses. IV. Experimental conditions for the induction of hapten-specific tolerance or for the stimulation of anti-hapten anamnestic responses by "nonimmunogenic" hapten-polypeptide conjugates

Administration of nonimmunogenic 2,4-dinitrophenyl (DNP) conjugates of copolymers of D or L-glutamic acid and lysine (GL) induces hapten-specific tolerance in nonimmune and DNP-ovalbumin-primed strain 13 guinea pigs. This tolerant state is evidenced by depressed anti-DNP antibody synthesis in response to challenge with DNP-ovalbumin and by a diminished frequency of DNP-specific antigen-binding cells and of anti-DNP antibody-secreting cells. Such a nonimmunogenic compound (DNP-D-GL) will nevertheless elicit a DNP-specific anamnestic antibody response when administered at an appropriate time to DNP-ovalbumin-primed guinea pigs undergoing a graft-versus-host reaction. These experiments are discussed in terms of a two-cell theory of stimulation of antibody responses.