The augmentation of tumor-specific immunity using haptenic muramyl dipeptide (MDP) derivatives

Summary A new haptenic compound, a muramyl dipeptide (MDP) derivative (designated as L4-MDP-ONB) cross-reactive with Bacillus Calmette Guerin (BCG) was synthesized. The cross-reactivity of L4-MDP hapten to BCG was demonstrated from the following evidence; (a) lymph node cells from BCG-primed C3H/HeN mice exhibited appreciable L4-MDP-specific proliferative responses to the in vitro stimulation of L4-MDP-modified syngeneic cells (L4-MDP-self); (b) inoculation of L4-MDP-self into footpads of BCG-primed C3H/HeN mice elicited ample delayed type-hypersensitivity (DTH) responses in vivo as measured by footpad swelling; and (c) BCG-primed mice contained L4-MDP-reactive helper T cell activity which functions to augment the generation of effector T cell responses to cell surface antigens. This crossreactivity between L4-MDP hapten and BCG as measured by the helper T cell activity was applied to enhanced induction of tumor-specific immunity. When BCG-primed C3H/HeN mice were immunized with L4-MDP-modified syngeneic X5563 tumor cells, these mice could generate augmented tumor-specific in vivo protective (tumor neutralizing) immunity as well as in vitro cytotoxic T cell responses. These results indicate the effectiveness of L4-MDP hapten in augmenting tumor-specific immunity. The present approach is discussed in the context of potential advantages of this new hapten for its future application to clinical tumor systems.     

Significance of exercise and bed rest in pregnancy--study on the lying postures of gravidas during sleep (2)

For the purpose of determining the most reasonable lying posture for pregnant women, we investigated the lying positions of both 247 non-pregnant women and 302 pregnant women during sleep. As for the rate of each position during the entire period of observation, 33.2% of the non-pregnant women were in the supine position, 41.2% in the lateral position, 18.4% in Sims' position and 7.1% in the prone position. In the pregnant group, the rate of supine position, simple lateral position and Sims' position was 34.2%, 52.2% and 12.7%, respectively, but the rate of the prone position was limited to 0.8%. All of the gravidas assuming the prone position were at less than 16 weeks of gestation. Non-pregnant women could sleep in a variety of positions, but pregnant women could assume the prone position during sleep only in the stage when the abdomen is not yet prominent or distended. Pregnant women were restricted significantly, either consciously or unconsciously, with progress in gestation.     

Effect of calcium and F-actin on the rate of SH2 modification in skeletal myosin

The rate constant of modification of a specific thiol group, SH₂, with N-ethylmaleimide (NEM) has been used to estimate the conformational change in the local area containing SH₂ (SH₂ region) of skeletal myosin as a structural probe. The rate of Mg²⁺-ATP-induced SH₂ modification of subfragment-1 (S-l) isozymes was regulated by Ca²⁺ in the pCa range below 6.4 and was not regulated in the pCa range above 6.4. No substantial difference between S-1 containing alkali light chain, A1, (S-1(A1)) and S-1 containing alkali light chain, A2, (S-1(A2)) was observed in the Ca²⁺-dependent rate of SH₂ modification. Due to the presence of this Ca²⁺ regulation in myosin (absence in S-1 isozymes) in the pCa range above 6.4, absence of 5,5-dithiobis-(2-nitrobenzoic acid) (DTNB) light chain in S-1 isozymes, and high affinity of Ca²⁺ for DTNB light chain, this Ca²⁺ regulation in the pCa range above 6.4 is possibly related to the Ca²⁺ binding to DTNB light chain. F-Actin, which is entirely free from tropomyosin and troponin, enhanced the rate of Mg²⁺-ATP-induced SH₂ modification of S-1 isozymes equally and of myosin, and reduced the Ca²⁺ sensitivity with an increase in F-actin concentration.     

O2 Regulation of Bacteriochlorophyll Synthesis in the Aerobic Bacterium Erythrobacter

This study examined the effect of oxygen on the bacteriochlorophyll(Bchl) synthetic activity of the aerobic marine bacterium Erythrobacter.The activity of the orange-pigmented strain E. longus OCh 101was highest at full atmospheric oxygen tension, while that ofthe pink-pigmented strain Erythrobacter sp. OCh 114 was lowat this tension and not observed in the absence of oxygen. (Received January 26, 1987; Accepted August 13, 1987)     

Cloning and nucleotide sequence of the penicillinase antirepressor gene penJ of Bacillus licheniformis.

The penicillinase antirepressor gene, penJ, of Bacillus licheniformis ATCC 9945a was cloned in Escherichia coli by using pMB9 as a vector plasmid. The penicillinase gene, penP, its repressor gene, penI, and penJ were encoded on the cloned 5.2-kilobase HindIII fragment of the recombinant plasmid pTTE71. The penJ open reading frame was composed of 1,803 bases and 601 amino acid residues (molecular weight, 68,388). A Shine-Dalgarno sequence was found 7 bases upstream from the translation start site. Since this sequence was located in the 3'-terminal region of the penI gene, penJ might be transcribed together with penI as a polycistronic mRNA from the penI promoter. Frameshift mutations of penJ were constructed in vitro from pTTE71, and the penJ mutant gene was introduced into B. licheniformis by chromosomal recombination. The transformant B. licheniformis U173 (penP+ penI+ penJ) turned out to be uninducible for penicillinase production, whereas the wild-type strain (penP+ penI+ penJ+) was inducible. Only when these three genes (penP, penI, and PenJ) were simultaneously subcloned in Bacillus subtilis did the plasmid carrier exhibit inducible penicillinase production, as did wild-type B. licheniformis. It was concluded that penJ is involved in the penicillinase induction. The regulation of penP expression by penI and penJ is discussed.     

Mutations in {beta}-Tubulin Block Transhyphal Migration of Nuclei in Dikaryosis in the Homobasidiomycete Coprinus cinereus

ß-Tubulins from fourteen benomyl-resistant strainsof the homobasidiomycete Coprinus cinereus, which carry thebenA, benB, benC or benD mutations, were analyzed by urea SDS-PAGEor isoelectric focusing and subsequent immunoblot analysis.Electrophoretic aberrations in a major ß-tubulin isotype,denoted ß1 were found in two strains, BEN154 and BEN215,both of which carry benomyl resistance mutations in benA ⁺ Theaberrations of ß1 in BEN154 and BEN215 cosegregatedwith benomyl resistance among the progeny of outcrosses of BEN154 and BEN215 to wild type, indicating that the ß1aberrations were caused by the benA mutations. Both the mutantand wild-type ß1 tubulins were present in the heterozygousdikaryons, BEN 154/wild-type and BEN215/wild-type, ruling outpost-translational modification as a possible cause for theaberrations in ß1. Thus, we conclude that benA isa structural gene for ß1. Transhyphal migration ofnuclei in dikaryosis was blocked in the mycelia of BEN 154 andin its progeny that carried benA (ß1 mutation), demonstratingthat microtubules are involved in the migration process. Nuclearmigration in dikaryosis seems to differ in terms of mechanism,at least in part, from the migration of tetrad nuclei from basidiainto prespores during formation of basidiospores and from themigration of nuclei from basidiospores into hyphae during germination,because a benA mutation blocked the former without affectingthe latter two processes. (Received May 19, 1989; Accepted August 30, 1989)     

Comparison of the inhibitions of proliferation of normal and psoriatic fibroblasts by 1 alpha,25-dihydroxyvitamin D3 and synthetic analogues of vitamin D3 with an oxygen atom in their side chain

The inhibitory effects of 1 alpha,25-(OH)2D3 and synthetic oxa-derivatives of vitamin D3 on growth of normal and psoriatic fibroblasts in culture were compared. Proliferation of normal fibroblasts was strongly inhibited by these new compounds in the following order: 22-oxa-1 alpha,25-(OH)2D3 greater than 22-oxa-1 alpha-(OH)D3 greater than 1 alpha,25-(OH)2D3 greater than 20-oxa-1 alpha,25-(OH)2D3. 22-Oxa-1 alpha,25-(OH)2D3 was about 10-times more inhibitory than 1 alpha,25-(OH)2D3. Proliferation of psoriatic fibroblasts was not inhibited by 1 alpha,25-(OH)2D3 at concentrations of up to 10(-6) M, but was suppressed by 10(-8)-10(-6) M 22-oxa-1 alpha,25-(OH)2D3 and 10(-6) M 22-oxa-1 alpha-(OH)D3. These results suggest that oxa-derivatives of vitamin D3, especially 22-oxa-1 alpha,25-(OH)2D3, should be useful in further studies on the cause and treatment of psoriasis.     

A spectrophotometric assay for the cyclization activity of cyclomaltohexaose (alpha-cyclodextrin) glucanotransferase

A cyclomaltohexaose (alpha-cyclodextrin) determination method which is both highly reproducible and selective is described. It involves the formation of an inclusion complex between the cyclodextrin and methyl orange under conditions of low pH (1.2) and low temperature (16 degrees C) and is useful for the assay of cyclodextrin glucanotransferase activity. The formation of the inclusion complex causes a decrease in absorbance of the methyl orange solution and this is monitored at a wavelength of 505 nm. The decrease in absorbance is linearly correlated with the cyclomaltohexaose concentration in the range of 0.25 optical density unit and 0.30 mM cyclomaltohexaose. The specificity of the test for cyclomaltohexaose is high, with only limited interference by linear oligosaccharides and other cyclodextrins: cyclomaltoheptaose and cyclomaltooctaose cause absorbance variations of 16 and 5%, respectively, of the response of maltohexaose. The formation of the complex is instantaneous and the complex is stable in time, provided the temperature is constant. The presence of methyl orange does not hinder enzymatic activity determination. The reaction is stopped by acidification and absorbance is measured at the fixed temperature of 16 degrees C. Possible interferences inherent to the composition of the sample itself can be suppressed by running appropriate controls and calculating a corrected optical density. This colorimetric method is simple and should be versatile in assaying diverse cyclomaltohexaose glucanotransferase enzymes.