Acetylcholine released from cholinergic nerves contributes to cutaneous vasodilation during heat stress.

Nitric oxide (NO) contributes to active cutaneous vasodilation during a heat stress in humans. Given that acetylcholine is released from cholinergic nerves during whole body heating, coupled with evidence that acetylcholine causes vasodilation via NO mechanisms, it is possible that release of acetylcholine in the dermal space contributes to cutaneous vasodilation during a heat stress. To test this hypothesis, in seven subjects skin blood flow (SkBF) and sweat rate were simultaneously monitored over three microdialysis membranes placed in the dermal space of dorsal forearm skin. One membrane was perfused with the acetylcholinesterase inhibitor neostigmine (10 microM), the second membrane was perfused with the NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME; 10 mM) dissolved in the aforementioned neostigmine solution (l-NAME(Neo)), and the third membrane was perfused with Ringer solution as a control site. Each subject was exposed to approximately 20 min of whole body heating via a water-perfused suit, which increased mean body temperature from 36.4 +/- 0.1 to 37.5 +/- 0.1 degrees C (P < 0.05). After the heat stress, SkBF at each site was normalized to its maximum value, identified by administration of 28 mM sodium nitroprusside. Mean body temperature threshold for cutaneous vasodilation was significantly lower at the neostigmine-treated site relative to the other sites (neostigmine: 36.6 +/- 0.1 degrees C, l-NAME(Neo): 37.1 +/- 0.1 degrees C, control: 36.9 +/- 0.1 degrees C), whereas no significant threshold difference was observed between the l-NAME(Neo)-treated and control sites. At the end of the heat stress, SkBF was not different between the neostigmine-treated and control sites, whereas SkBF at the l-NAME(Neo)-treated site was significantly lower than the other sites. These results suggest that acetylcholine released from cholinergic nerves is capable of modulating cutaneous vasodilation via NO synthase mechanisms early in the heat stress but not after substantial cutaneous vasodilation.     

Association of human lymph node homing receptor (Leu 8) with the TCR/CD3 complex.

Cross-linking of the human homologue of the murine MEL-14 lymph node homing receptor (Selectin-1, LECAM-1, Leu 8) on both T and B cells results in modification of cell function. To investigate this phenomenon, we performed studies to determine if the Leu 8 molecule influences T cell activation via the TCR/CD3 complex. In initial studies, we treated T cells with immobilized anti-CD3 (OKT3 mAb) in the presence or absence of immobilized Leu 8 mAb. We found that although Leu 8 mAb alone had no effect on T cell proliferation, this antibody markedly augmented immobilized OKT3 mAb-induced proliferation. In further studies, we immunoprecipitated surface radioiodinated T cell lysates with OKT3 and Leu 8 mAb to determine if molecules in the TCR/CD3 complex associate with Leu 8 molecules. Although Leu 8 mAb immunoprecipitated only a single protein of approximately 80 kDa from T cell lysates treated with Nonidet P-40 under reducing condition, it coimmunoprecipitated additional proteins of 48, 42, 28, 24, and 22 kDa from T cell lysates treated with 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate. These additional proteins were identified as the alpha-, beta-, gamma-, delta-, and epsilon-chains of the TCR/CD3 complex by one-dimensional and two-dimensional diagonal SDS-PAGE. Densitometric scanning showed that, on average, 18% of the TCR/CD3 complex associates with Leu 8. In a final study, we showed by immunoblotting analysis using anti-zeta peptide antibody that Leu 8 mAb coimmunoprecipitates the zeta-chain of CD3. These results indicate that the human lymph node homing receptor homologue (Leu 8) participates in the activation of T cells, probably via its association with the TCR/CD3 complex.     

Localization of smg p25A/rab3A p25, a small GTP-binding protein, at the active zone of the rat neuromuscular junction.

smg p25A is a small G protein which has been suggested to regulate neurotransmitter release from the synapses. We investigated here the ultrastructural localization of this small G protein in the rat neuromuscular junction by an immunoperoxidase method. The results showed that smg p25A was distributed non-uniformly on the presynaptic plasma membrane and among the synaptic vesicles with the focal accumulation on the discrete presynaptic sites which corresponded to the active zones, the regions of the presynaptic plasma membrane specialized for the exocytosis of the synaptic vesicles. This unique distribution of smg p25A suggests that it plays an important role in the attachment and fusion of the synaptic vesicles with the active zones.     

Immunological quantitation and immunohistochemical localization of leukotriene A4 hydrolase in guinea pig tissues

We prepared a highly specific polyclonal antibody against leukotriene (LT) A4 hydrolase using a recombinant human enzyme. Using this antibody, we quantified LTA4 hydrolase protein content in the cytosols of guinea pig tissues. The enzyme protein content correlated well with the enzyme activity with a correlation coefficient of 0.87. However, the enzyme activity per mg of the enzyme in the cytosols was low, particularly in the liver and adrenal gland, compared with the specific activity of the purified enzyme. These observations suggest the presence of inhibitory substances and/or inactive enzymes in the cytosols of these tissues. To determine the cellular localization of LTA4 hydrolase in tissues other than blood cells, we carried out immunohistochemical examinations of guinea pig tissues. We identified epithelial cells in the tracheobronchial system and gastrointestinal tract, smooth muscle cells in the bronchi and aorta, vascular endothelial cells, and the intestinal plexus as novel cellular sources of the enzyme in the parenchyme of the tissue. Thus, LTA4 hydrolase was widely distributed in various types of parenchymal cells in the tissues, and this observation warrants further investigations on the biological activities of LTB4 in these cells and tissues.     

In vivo effects of human recombinant tumor necrosis factor alone and in combination with other biological response modifiers on human digestive organ cancer xenografts transplanted in nude mice

The present study was designed to evaluate the effect of rTNF alone or in combination with other BRMs on human digestive organ cancers. Six kinds of human digestive organ cancer xenografts (esophageal, stomach, colonic, pancreatic, bile duct, and liver cancers: EC-YO, GC-YN, CC-KK, PC-HN, BDC-SN and Li-7, respectively) were transplanted in nude mice, and rTNF was administered at 10³, 5 × 10³, or 10⁴U/head directly into the tumor 3 times a week for 2 weeks. EC-YO was the most sensitive to rTNF, and intratumoral administration of rTNF at 10³ U/head caused tumor regression. PC-HN, CC-KK and GC-YN were relatively sensitive to rTNF, and their growth was significantly inhibited by rTNF at 5 × 10³ U/head, however, the tumors regrew after treatment. Li-7 and BDC-SN were resistant to rTNF. The effects of rTNF in combination with recombinant interferon- (rIFN-), recombinant interleukin-2 (rIL-2), or streptococcal preparation OK-432 were assessed in mice transplanted with GC-YN. All combinations of rTNF at 5 × 10³ U/head and other BRMs were more effective than rTNF alone, and GC-YN tumors were completely regressed after treatment with a combination of rTNF and rIFN- or rTNF and OK-432. However in all cases, the combination of rTNF at 10³ U/head and any other BRM did not improve the effect. Furthermore, the adverse effects of the combinations were more serious than those of rTNF alone.TNF may still be a useful cytokine, because it can induce the regression of tumors. However, for its clinical application, a method should be developed to reduce its side effects.     

Phenylketonuria in a patient with cystinuria.

During routine screening procedures for amino-acid disorders by thin-layer chromatography, a 16-year-old boy was found to have phenylketonuria and cystinuria. A phenylalanine and a cystine loading were carried out. The patient was found to be homozygous for phenylketonuria and heterozygous for cystinuria type II. His father was heterozygous for phenylketonuria and cystinuria, while his mother proved to be heterozygous only for phenylketonuria.     

Murine thyroid follicular epithelial cells can be induced to express class II (Ia) gene products but fail to present antigen in vitro

To determine whether thyroid follicular epithelial cells (TFEC) might be involved in the induction of autoimmune thyroiditis, they were tested for their potential to express Ia antigens, and for their ability to present antigen in vitro. Results showed that Ia antigens, absent on normal TFEC, could be readily induced with interferon gamma, as detected by immunofluorescence. Maximal expression of Ia antigens in over 50% of TFEC was observed after 4 days of culture in the presence of IFN-gamma, and was quantitatively comparable to spleen cells by cytofluorometric analysis. Moreover, primary TFEC in culture secreted thyroglobulin (tg) and interleukin 1. However, TFEC consistently failed to stimulate various populations of T cells. These included lymph node cells sensitized to tg, a T-cell clone specific for azo-benzene-arsonate tyrosine (ABA), and a hybridoma specific for beef insulin. Likewise, Ia-positive TFEC did not stimulate T-cell hybridomas restricted to the class II alloantigen I-Ab, while stimulating a hybridoma specific for the class I alloantigen Kb. T-cell unresponsiveness could not be explained by inhibitory activity of TFEC, released either into the culture supernatant or exerted by cell contact. The data indicate that Ia-positive TFEC failed to serve as class II-restricted antigen-presenting cells (APC) in vitro and thus argue against a primary role for these cells in the inductive phase of thyroiditis.     

Instability and oscillatory behavior of membrane-chemical reaction systems

The stability problem of a chemical system consisting of reversible reactions is analyzed with the aid of computer calculations. The system is based on the model proposed by Edelstein and exhibits oscillations of chemical species. The analysis shows that the oscillatory character is of limit cycle type. The results are applied to the construction of a membrane-chemical reaction system, which shows characteristic instability behavior. This is useful as a model of cell division.