Prostaglandin E2 inhibits the activation of cloned T cell hybridomas

To minimize complicating interactions inherent in heterogeneous cell populations, we used a panel of cloned murine autoreactive (E8.A1) and antigen-specific (HEL.C10, HEL.B14) T cell hybridomas to examine the effect of prostaglandin E2 (PGE2) on T cell activation. These T cells secrete interleukin 2 (IL 2) when co-cultured with a cloned population of I region-matched stimulator cells (TA3), or with mitogenic signals in the absence of TA3 stimulator cells. Physiologic concentrations of PGE2 inhibited the induction of IL 2 secretion by the T cell hybridomas tested, when they were activated either by TA3 cells or by mitogenic signals. IL 2 production was inhibited in a dose-dependent manner by concentrations of PGE2 between 10(-7) and 10(-11) M, with 50% inhibition occurring at 10(-10) M. Pretreatment of the T hybridoma cells with 10(-7) M PGE2 for 1 hr before culture also resulted in marked inhibition of IL 2 secretion. Similar pretreatment of the TA3 cells did not affect their ability to activate the T cell hybridomas. PGE2 at 10(-8) M induced a 30-fold increase in cAMP levels within 25 min of addition to culture of the E8.A1 T cell hybridoma, but caused no significant elevation of cAMP levels in TA3 cells. The direct addition of dibutyryl cAMP (dcAMP) to cultures of E8.A1 cells resulted in marked inhibition of IL 2 secretion when stimulated by TA3 or by mitogenic signals, with an average of 80% inhibition occurring at 10(-4) M dcAMP. PGE2 and dcAMP also inhibited the growth of E8.A1 cells. Initially, cell growth was virtually halted, but began to recover between 24 and 48 hr after the addition of either PGE2 or dcAMP. Neither PGE2 nor dcAMP inhibited the division of TA3 cells. High affinity binding sites for PGE2 were detected in the E8.A1 T cell hybridomas with an apparent Kd of 7.6 X 10(-10) M, which is consistent with the functional data. No specific binding was detected in the TA3 stimulator cells. These findings suggest that the immunosuppressive effects of PGE2 are localized to the T cell, are receptor regulated, and may be mediated by the associated increase of cAMP levels in the T cell hybridomas.     

Evidence Inconsistent with the Blaauw Model of Phototropism

The Blaauw model of phototropism equates the inhibition of growthat the illuminated side of a unilaterally illuminated organwith the blue light inhibition of overall organ extension evidentwhen some shoots are exposed to uniform blue light However,a study of the growth responses of Avena coleoptiles exposedto omnilateral, equal bilateral, unequal bilateral and unilateralblue light has revealed some light induced growth rate changeswhich cannot be explained by the Blaauw model. The growth responsesof cells at the illuminated and shaded sides of phototropicallystimulated coleoptiles seem to depend on the existence of alight gradient across the whole organ rather than the absolutelevels of light at either side. Key words: Phototropism, Avena coleoptile, Blaauw hypothesis, Blue light, Growth inhibition     

Mechanism of the cerebrovascular effect of piracetam

Piracetam produces a more pronounced effect on cerebral circulation disturbed by hemorrhagic shock as compared with intact animals. Piracetam has a depressant effect on the nervous regulation of cerebral circulation by suppressing the reflex constriction of the vessels in both arterial systems of the brain. The cerebrovascular effects of piracetam are not mediated through the GABAergic bicuculline-sensitive mechanisms, which is supported by experiments where the drug exhibits its effects under the blockade of GABA receptors.     

DNA transposition – a major contributor to plant chromosome structure

Higher plant nuclear genomes contain many families of dispersed repeats that change during evolution. Recent evidence from studies on genetically defined transposable elements raises the possibility that many of the dispersed repeats are remnants of such elements. Transposition of DNA has also occurred between mitochondria, chloroplasts and nuclei, a fact that underlines the major role played by DNA transposition in determining the structure of plant genomes.     

Coordinate regulation of histone mRNAs during growth and differentiation of rat myoblasts

To determine whether histone genes are coordinately regulated, histone mRNA concentrations were measured in exponentially growing L6 myoblasts, S-phase synchronized myoblasts and in differentiating myoblasts. The levels of various histone mRNA subspecies declined rapidly and coordinately once myoblasts were given the signal to differentiate. mRNA levels were reduced on average to 1-5% of the amount observed in exponentially growing cells by 48 h after the signal to differentiate. The reductions occurred in concert with the cessation of DNA synthesis as the cells differentiated. Inhibition of DNA synthesis by treating myoblasts with Ara-C or hydroxyurea resulted in a histone mRNA half-life of 10-13 min for each of the histones examined. One example of non-coordinate regulation was observed however among the H4 mRNA subspecies in S-phase synchronized cells. The levels of two major subspecies of H4 mRNA increased coordinately in S-phase compared to levels observed in cells growing exponentially. A third subspecies of H4 mRNA on the other hand was found to decline by 50%. These studies suggest that the majority of histone mRNA subspecies are under coordinate control, although one exception has been noted among the subspecies of histone H4.     

Analysis of 6-mercaptopurine in human plasma with a high-performance liquid chromatographic method including post-column derivatization and fluorimetric detection

A relatively simple assay with improved reliability and sensitivity for measuring levels of 6-mercaptopurine in human plasma is presented. After extraction of the compound and the added internal standard with phenyl mercury acetate, samples were separated by ion-pair reversed-phase high-performance liquid chromatography. On-line the analytes were oxidized to fluorescent products and detected in a flow-fluorimeter. The within-day coefficient of variation was 3.8% at a concentration of 25 ng/ml. The lower detection limit was 2 ng/ml when 1.0 ml of plasma was used. Mercaptopurine concentration versus time curves of two subjects after a single oral dose of azathioprine are shown.     

Purification of a soluble, sodium-nitroprusside-stimulated guanylate cyclase from bovine lung

A soluble, sodium-nitroprusside-stimulated guanylate cyclase as been purified from bovine lung by DEAE-cellulose chromatography, ammonium sulfate precipitation, chromatography on Blue Sepharose CL-6B and preparative gel electrophoresis. Apparent homogeneity was obtained after at least 7000-fold purification with a yield of 3%. A single stained band (Mr 72000) was observed after gel electrophoresis in the presence of sodium dodecyl sulfate. The purified enzyme migrated as one band also under non-denaturing conditions in acrylamide gels (5-12%). The mobility of this band corresponded to an Mr of 145000. The enzyme sedimented on sucrose gradients with an S20, w of 7.0 S. Gel filtration yielded a Stokes' radius of 4.6 nm. These data suggest that the enzyme has an Mr of approximately 150000 and consists of two, presumably identical, subunits of Mr 72000. Sodium nitroprusside stimulated the purified enzyme 15-fold and 140-fold to specific activities of 8.5 and 15.7 mumol of cGMP formed min-1 mg-1 in the presence of Mn2+ and Mg2+, respectively. Formation of cGMP was proportional to the incubation time and to the amount of enzyme added. The stimulatory effect of sodium nitroprusside was half-maximal at about 2 microM, was observed immediately after addition and could be reversed either by dilution or by removal of sodium nitroprusside on a Sephadex G-25 column. The purified enzyme in the absence of catalase was stimulated by sodium nitroprusside, N-methyl-N'-nitro-N-nitrosoguanidine and 3-morpholino-sydnonimine and in the presence of catalase by sodium nitrite and sodium azide. In the presence of Mn2+ and sodium nitroprusside, the purified enzyme catalyzed the formation of cAMP from ATP at a rate of 0.6 mumol min-1 mg-1.     

Effects of serial anesthesia using ketamine or ketamine/medetomidine on hematology and serum biochemistry values in rhesus macaques (Macaca Mulatta)

Background  This study aimed at determining the cumulative effect of daily anesthesia, using two drug regimens, over hematological and biochemical parameters.
Methods  Blood samples were obtained from rhesus monkeys 20 minutes after intramuscular administration of ketamine or ketamine/medetomidine combination for three consecutive days and results were evaluated to determine their effect on hematological and serum biochemistry values. Statistical significance of drug, day, and interaction of these two variables were evaluated.
Results  Drug effect resulted in a dramatic increase of aspartate aminotransferase and creatine kinase values. Day effect resulted in decreases of RBC, HCT, Hgb, and alkaline phosphatase but an increase of other biochemical parameters evaluated. The drug/day interaction effect was found to be –significant for RBC, platelets, aspartate aminotransferase, alanine aminotransferase, and creatine kinase values.
Conclusion  The results of our study suggest a cumulative effect of serial anesthesia and should be an important consideration when interpreting hematology and serum biochemistry in rhesus macaques.     

Antibody-targeted photolysis: in vitro immunological, photophysical, and cytotoxic properties of monoclonal antibody-dextran-Sn(IV) chlorin e6 immunoconjugates.

A set of anti-melanoma immunoconjugates were prepared which contained chlorin e6: antibody molar ratios of 18.9:1, 11.2:1, 6.8:1, and 1.7:1. All immunoconjugates retained antigen binding activity regardless of the chromophore:antibody substitution ratio that was attained. In contrast, the ground-state absorption spectra of the immunoconjugates showed features which appeared to be dependent on the chromophore:antibody molar ratio. In addition, the quantum yield of singlet oxygen generated by the conjugated chromophores was observed to be significantly less than that observed with the unbound dye. Time-resolved absorbance spectroscopy of the chromophore excited triplet state indicated that the loss of singlet oxygen quantum yield resulted from diminished chromophore triplet yield. Analysis of data obtained from in vitro photolysis of target melanoma cells, in combination with that obtained from the immunochemical and photochemical studies, indicates that the observed immunoconjugate phototoxicity can be reasonably quantitatively represented by (1) the ability of the immunoconjugate to bind SK-MEL-2 cell surface antigen, (2) the amount of chromophore localized at the target cells by immunoconjugate binding, (3) the delivered dose of light at 634 nm, and (4) the singlet oxygen quantum yield of the antibody-bound photosensitizer. Though these data argue strongly for photolysis by the cumulative dosage of singlet oxygen at the cell membrane, nonetheless, the concurrent photoinduced release of other cytotoxic agents should not be ruled out.