Inhibition of leukocyte adhesion by the in vivo and in vitro administration of cannabinoids

Cannabinoids have been shown to affect various aspects of arachidonic acid metabolism both in vivo and in vitro. Eicosanoid metabolites of arachidonate and related octadecanoate are believed to be involved in cell adhesion processes as agonists in some instances and as antagonists in other cases. This report shows data in which cannabinoids exhibit marked inhibitory effects on the adhesion of mouse peritoneal cells to polystyrene culture dishes. The effects could be seen by in vivo administration of the drugs as well as by direct exposure of the cells in vitro. The data suggest that this inhibition of adhesion is mediated by one or more products generated by stimulation of a lipoxygenase pathway.     

The ras-like protein p25rab3A is partially cytosolic and is expressed only in neural tissue.

An antipeptide antiserum has been developed against a sequence near the C terminus of the small guanine nucleotide-binding protein p25rab3A. This protein is the product of one of a large number of genes that show homology to the ras proto-oncogenes. Immunoblotting with the antiserum specifically detected a 25-kilodalton protein in brain membranes. This protein coeluted from a MonoQ high-resolution ion-exchange column with a 25-kilodalton GTP-binding protein at a salt concentration similar to that known to elute purified p25rab3A. Unlike p21ras, which is exclusively membrane bound, p25rab3A is present in both the cytosol and membrane fractions of rat brain. It was not detected in other tissues, although a band of slightly lower molecular weight was observed with skeletal muscle. Western blot (immunoblot) analysis of five regions of the rat brain indicated that p25rab3A is most abundant in the hypothalamus and hippocampus.     

Translocation of cytosolic acetylcholine into synaptic vesicles and demonstration of vesicular release

The rate of translocation of newly synthesized acetylcholine (ACh) from the presynaptic cytosol of Torpedo electric organ nerve terminals into synaptic vesicles and the extent to which ACh release from these neurons is mediated by a vesicular mechanism were investigated. For this purpose the compound 2(4-phenylpiperidino)cyclohexanol (AH5183), which inhibits the active transport of ACh into isolated cholinergic synaptic vesicles, was employed. Preincubation of purified Torpedo nerve terminals (synaptosomes) with AH5183 does not affect the intraterminal synthesis of [3H]ACh but results in a marked inhibition (85%) of its Ca2+-dependent K+-evoked release. By contrast, the evoked release of the endogenous nonlabeled ACh is not affected by this compound. When AH5183 is added during radiolabeling, it causes a progressively smaller inhibition of [3H]ACh release which is completely abolished if the drug is added after the preparation has been labeled. These findings suggest that most of the newly synthesized synaptosomal [3H]ACh (85%) is released by a vesicular mechanism and that some [3H]ACh (15%) may be released by a different process. The translocation of cytosolic [3H]ACh into the synaptic vesicles was monitored by determining the time course of the loss of susceptibility of [3H]ACh release to AH5183. It was found not to be coupled kinetically to [3H]ACh synthesis and to lag behind it. The nature of the intraterminal processes underlying this lag is discussed.     

The oral assimilation of radiomanganese by the mouse

The metabolism of orally administered radiomanganese was studied in mice. Assimilation of absorbed manganese (Mn) was determined using whole body counting techniques. When⁵⁴MnCl₂ was administered, 2.7% of the dose was retained after 10 d compared with 1.2% from the⁵⁴Mn-nitrilotriacetate (NTA) complex. However, this difference was accounted for by the rapid and persistent adsorption of the Mn onto the teeth of the lower jaw when fed as the ionic salt at pH 2.0 compared with the NTA-chelate fed at pH 9.0. Once corrected for the amount adsorbed onto the teeth, the biodistribution and relative specific activity of the assimilated radiomanganese into a variety of tissues were similar for both forms of the metal.     

Partial amino acid sequence of porcine elastase II. Active site and the activation peptide regions

The partial amino acid sequence of porcine elastase II, isolated from crude trypsin Type II, was determined. The enzyme consists of two polypeptide chains, a light chain composed of 11 residues, and a heavy chain (Mr = 23 500) with four intrachain disulfide bridges; the two chains are held together by one interchain disulfide bond. Elastase II was fragmented into several peptides by chemical cleavages with CNBr at the two methionine residues, 99 and 180 (chymotrypsinogen numbering), and with hydroxylamine at the peptide bond following DIP-Ser195. About 50% of the sequence was determined and the positions of 120 amino acids were located, including the light chain residues and most of the active site residues. The partial sequence shows 64% difference between porcine elastase II and elastase I and only 26% difference between porcine elastase II and bovine chymotrypsin B.     

Erythropoietin and megakaryocytopoiesis

To determine if erythropoietin influences megakaryocytopoiesis, the purified recombinant human hormone (rEpo) was added to serum-free liquid cultures of murine marrow. A dose-related increment in acetylcholinesterase (AchE) production was observed. To assess if increments in this relatively megakaryocyte-specific enzyme marker were mediated by a direct hormone-megakaryocyte interaction rather than via an accessory cell population, rEpo was added to cultures of isolated single megakaryocytes. A significant, dose-related increase in cell size was noted in the presence of the hormone, accompanied by a high probability of an increase in cellular DNA content. The data show that rEpo does directly influence some aspects of megakaryocytic maturation, although the physiologic significance of this effect remains unknown.     

Effect of an acute bout of exercise on glucose disposal in human obesity

The effect of acute exercise on insulin action has been studied in six obese (150-250% ideal body weight) non-insulin-dependent diabetics (OD), seven obese normoglycemics (ON), and six lean healthy controls (LC). Using a three-stage euglycemic clamp, the metabolic clearance rate (MCR) of glucose under increasing insulin concentrations was measured. The insulin dose-response curve was assessed on two separate occasions: 1) a base-line test and 2) 1 h after aerobic treadmill exercise at a steady-state heart rate of 150-160 beats/min. In the base-line test, under all insulin levels, glucose MCR was significantly lower in obese compared with lean individuals (P less than 0.01). Exercise increased glucose MCR at the highest hormonal concentrations applied to 124 and 134% of base line in OD and in ON, respectively (P less than 0.05); the insulin concentration required for one-half of the maximal clearance rate of glucose achieved in this study decreased from 200 to 130 and from 160 to 95 microU/ml in OD and ON, respectively (P less than 0.05). The changes in these parameters were insignificant in LC. It is suggested that acute exercise affected the insulin dose-response curve in OD and in ON but not in LC; although enhanced by exercise, glucose MCR remained significantly lower in both obese groups compared with control subjects. We concluded that insulin resistance, which accompanies extreme obesity, could be markedly decreased but not completely reversed by one bout of exercise.     

A two-domain mechanism for group A streptococcal adherence through protein F to the extracellular matrix.

Streptococcus pyogenes binds to the extracellular matrix (ECM) and a variety of host cells and tissues, causing diverse human diseases. Protein F, a S.pyogenes adhesin that binds fibronectin (Fn), contains two binding domains. A repeated domain (RD2) and an additional domain (UR), located immediately N-terminal to RD2. Both domains are required for maximal Fn binding. In this study, we characterize RD2 and UR precisely and compare their functions and binding sites in Fn. The minimal functional unit of RD2 is of 44 amino acids, with contributions from two adjacent RD2 repeats flanked by a novel 'MGGQSES' motif. RD2 binds to the N-terminal fibrin binding domain of Fn. UR contains 49 amino acids, of which six are from the first repeat of RD2. It binds to Fn with higher affinity than RD2, and recognizes a larger fragment that contains fibrin and collagen binding domains. Expression of UR and RD2 independently on the surface-exposed region of unrelated streptococcal protein demonstrates that both mediate adherence of the bacteria to the ECM. We describe here a mechanism of adherence of a pathogen that involves two pairs of sites located on a single adhesin molecule and directed at the same host receptor.