A substance inhibiting blast transformation of murine spleen lymphocytes stimulated with various mitogens, such as LPS, PHA, and PWM, was obtained from yeast-form cells of Histoplasma capsulatum. This active substance was partially purified from the cell-free extract by DEAE-Sepharose CL-6B column chromatography. As a result of this partial purification, the inhibitory activity was 1.26μg/ml in terms of ID₅₀. Materials from H. capsulatum also inhibited blast transformation of murine spleen lymphocytes stimulated with the antigen PPD as well as mitogens LPS, PHA, and PWM. However, the con A-induced proliferative response was only slightly affected. A similar result was observed for the MLR. These inhibitory activities were abolished by heating at 70C for 30min. These results suggest that the heat-labile active substance produced by H. capsulatum might directly affect the lymphocytes, leading to inhibition of their blast transformation.

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The R-form lipopolysaccharide (LPS) from Klebsiella strain LEN-111 (O3-:K1-) forms a hexagonal lattice structure with a lattice constant of 14 to 15nm when it is precipitated by addition of two volumes of 10mM MgCl₂-ethanol. When the LPS was suspended in various buffers (50mM) at pH 2 to 12 for 24hr at 4C, at pH 2 and 3 pits of the hexagonal lattice structure markedly disappeared, at pH 4 to 8.5 the lattice structure was stable, and at pH 9 to 12 it tended to loosen somewhat. The LPS from which cations were removed by electrodialysis retained the ability of hexagonal assembly, although the lattice constant of the hexagonal lattice of the electrodialyzed LPS was large. The lattice structure of the electrodialyzed LPS was much more labile than that of the non-electrodialyzed LPS at alkaline pH levels and the former was completely disintegrated into ribbon-like structures when the LPS was suspended in 50mM Tris buffer at pH 7.7 or higher. However, the electrodialyzed LPS formed a hexagonal lattice structure in Tris buffer at pH 8.5 containing 0.1 to 100mM MgCl₂. The lattice constants of the hexagonal lattice formed by the electrodialyzed LPS at 10 or 100mM MgCl₂ were very similar to that of the lattice of the non-electrodialyzed LPS. From these results it is concluded that the lability of the hexagonal lattice structure of the electrodialyzed LPS at alkaline conditions is due to removal of Mg²⁺ by electrodialysis.

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The R-form lipopolysaccharide (LPS) from Klebsiella strain LEN-111 (O3-:K1-) forms a hexagonal lattice structure with a lattice constant of 14 to 15nm when it is precipitated by addition of two volumes of 10mM MgCl₂-ethanol. The stability of this hexagonal lattice structure in long-term incubation at 4C was investigated. The hexagonal lattice structure was stable for at least 220 days when the LPS was suspended in distilled water, but it had been disintegrated into a rough mesh-like structure when the LPS was suspended in 50mM tris (hydroxymethyl) aminomethane (Tris) buffer, pH 8.5, at 4C for 60 days. Half of the Mg bound to the LPS was released when the LPS was suspended in Tris buffer for 60 days, whereas Mg was not released when it was suspended in distilled water even for 220 days. By contrast, it was stable for at least 220 days in Tris buffer containing 5mM MgCl₂. The LPS suspended in Tris buffer for 60 days, at which time the structure had been disintegrated, could be restored to the original hexagonal lattice structure within 24hr by addition of 5mM MgCl₂. From these results it is concluded that the hexagonal lattice structure of the LPS retains long-range stability if Mg bound to the LPS is not released from the LPS.

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A proteinaceous antigen (PAg) was purified from the culture supernatant of Streptococcus mutans 6715 (serotype g) by ultrafiltration, ammonium sulfate precipitation, DEAE-Sephacel ion-exchange chromatography, Phenyl-Sepharose CL-4B hydrophobic chromatography, and subsequent Sephacryl S-300 gel filtration. A yield of 0.1mg of PAg was obtained from a liter of culture supernatant. The isoelectric point and molecular weight of PAg were pH 4.6 and 210, 000, respectively. It contained 35% sugar, which was identified as glucose by gas-liquid chromatography. Amino acid analysis revealed that PAg contains 28% acidic and 11% basic amino acid residues. PAg retained its antigenicity after heating at 80C for 10min in deionized water, or after treatment with 0.1M HCl or 0.1M NaOH at 37C for 1hr. Immunodiffusion and immunoelectrophoresis analyses revealed that PAg is serologically distinct from other cell-surface antigens such as serotype-specific polysaccharide and lipoteichoic acid. A cross-reaction between PAg and a protein antigen similarly prepared from serotype c S. mutans was observed in immunodiffusion tests.

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Previously, we showed that the antiserum raised in male (DBA/2Ha× BALB/c)F₁(DCF₁) mice (T cell-replacing factor [TRF]-low response animals) by immunizing them with activated B cells from BALB/c mice (TRF-high-responders) contained antibodies against putative TRF-acceptor site(s). We have now evaluated the hypothesis that neonatal treatment of mice with the above antiserum suppresses the development of B cells responsive to TRF. Male DCF₁ mouse anti-BALB/c B-cell antiserum or normal DCF₁ mouse serum as a control was injected into BALB/c mice within 24hr after birth. In the antiserum-treated mice, no augmented primary immunoglobulin M (IgM) antibody responses to sheep red blood cells (SRBC) were observed under the conditions in which markedly augmented IgM anti-SRBC responses were induced in control BALB/c mice, suggesting that development of B cells reacting with male DCF₁ mouse anti-BALB/c B-cell antiserum is suppressed by the neonatal treatment with the antiserum. Furthermore, the development of B cell activity responsible for helper factors derived from T cells, such as TRF, was markedly suppressed in the neonatally antiserum-treated mice, whereas activity of B cells capable of interacting directly with helper T cells through antigen-bridges was not significantly affected by the same treatment. Such suppression of the B cell activity could be induced only when the antiserum was administered within 48hr after birth. Moreover, neonatal treatment of mice with the antiserum induced suppressed responsiveness of B cells to a T-independent type 2 antigen, TNP-Ficoll. Neither serum-borne suppressive serum components nor suppressor cells were detected by the system employed. These results support the hypothesis that TRF responsive B cells constitute a subpopulation distinct from the other B cells capable of cooperating with helper T cells via cognate interaction.

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Contact sensitivity to trinitrophenyl (TNP) hapten was induced by subcutaneous (s.c.) administration of TNP-modified syngeneic spleen cells or epidermal cells (EC) (TNP-EC). Intraperitoneal (i.p.) inoculation of TNP-EC resulted in a comparable response, whereas i.p. administration of TNP-spleen cells or TNP-modified-ultraviolet (UV)-preirradiated EC (TNP-UV-EC) failed to induce TNP-contact sensitivity responses. The present study investigates the effect of UV-irradiation on the potential of EC for inducing the contact sensitivity response. Exposure of BALB/c mouse EC in vitro to 1600J/m² of UV-B before they were modified with TNP had no discernible effect on the Ia-positivity and viability of EC. Coexistence of TNP-UV-EC had no inhibitory effect upon the contact sensitivity response induced by TNP-EC via the i.p. route. The absence of suppressor cell generation was substantiated by the adoptive transfer of spleen cells from mice administered TNP-UV-EC i.p. to normal syngeneic mice. The effect of interleukin 1 (IL-1) or epidermal cell-derived thymocyte-activating factor (ETAF) in restoring the ability of TNP-UV-EC to induce contact sensitivity was examined. IL-1 or ETAF administered along with TNP-spleen cells i.p. induced a potent contact sensitivity response, whereas the same preparations of IL-1 or ETAF were unable to restore the contact sensitivity induction by TNP-UV-EC. The results are discussed in the context of UV-induced cell surface changes of the Langerhans cell population.

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