The effect of copper on frog skin. The role of sulphydryl groups

1. 1. Cu²⁺ at a concentration of 10⁻⁴ M, when applied to the external side of the frog skin produces an increase in the short-circuit current (I_sc).

2. 2. This effect was studied in skins of Rana temporaria adapted to cold (5°C) and room temperature (20°C), skins of Rana pipiens adapted to cold, and the results compared with those obtained previously with Rana ribibunda.

3. 3. The observed effect is less dependent upon the adaptation to cold than upon the functional state of the skin: skins with low short circuit currents have a bigger response to Cu²⁺ than skins with high I_sc.

4. 4. A species difference cannot be ruled out since skins of Rana ribibunda exhibiting high I_sc give good responses to Cu²⁺.

5. 5. 5,5′-dithiobis(2-nitrobenzoic acid), a sulphydryl-oxidizing reagent, produces an effect similar to that of Cu²⁺, and dithiothreitol an SH-reducing agent, reverses the effect of this ion.

6. 6. Cu²⁺ also induces an increase in the unidirectional K⁺ fluxes and unmasks a net outward potassium flux.

7. 7. The outward K⁺ flux induced by Cu²⁺ is sensitive to ouabain.

8. 8. It is concluded that Cu²⁺ increases the permeability of the external barrier of the frog skin to Na⁺ and K⁺, probably by reacting with SH groups.

Interactions of acetylcholine receptor and acetylcholinesterase with lipid monolayers

The interaction of acetylcholine receptor and acetylcholinesterase with lipid monolayers was followed by measuring changes in surface pressure.When injected into the subphase of a lipid monolayer, the proteins caused increases in surface pressure from 5 to 10 dynes/cm, indicating a penetration of protein into the monolayer. At pH values below the isoelectric point of the proteins the incorporation was improved. The same was observed when Ca²⁺ (2 mM) was added.The presence of the enzyme in the mixed film could be demonstrated by using diiso[³H]propyl fluorophosphate-labelled acetylcholinesterase as well as by measuring enzyme activity. Acetylcholine receptor was shown to be present in the mixed film by using a complex made of the receptor and α-[³H]neurotoxin.     

Chymotryptic heavy meromyosin from gizzard myosin: a proteolytic fragment with the regulatory properties of the intact myosin.

Arginine deiminase (EC 3.5.3.6) from $\text{Mycoplasma}\text{arthritidis}$ is a dimeric enzyme. Velocity centrifugation in 6 M guanidine HCl and peptide mapping of the BrCN fragments suggest that the subunits are identical. The reaction of one out of four sulfhydryl groups with 0.3 mM 5,5′-dithiobis-(2-nitrobenzoic acid) has a half-life of about 30 min in 2 M guanidine HCl at 15°, pH 8. The enzyme is irreversibly inhibited by 1 mM formamidinium ion within 1 min. Inactivation by this affinity label is resolvable into two concurrent first-order reactions in the presence of guanidinium ion; the fraction of enzyme which reacts at the faster rate is about 50%. These results are interpreted as evidence for two catalytic subunits which differ in conformation.     

Antimetabolic activities of 2-fluoro-L-histidine.

2-Fluoro-L-Histidine inhibits protein synthesis in various cell cultures, as measured by ³H-leucine incorporation. This histidine analog also inhibits the cytopathogenicity of a number of RNA and DNA viruses in primary and continuous cell cultures; it blocks the transformation of normal mouse (MO) cells by murine sarcoma virus, and partially suppresses the release of murine leukemia virus by a continuously infected mouse cell line (JLSV5). In human skin fibroblasts, it reduces the interferon-inducing capacity of poly(I)·poly(C). Inhibition of cell protein synthesis may be the common cause of the various effects. 4-Fluoro-L-histidine is essentially inert in all of the test systems examined.     

Evidence from ATP synthesis driven by a proton gradient in Methanosarcina barkeri.

Hepatoma Tissue Culture (HTC) cell nuclei were isolated from untreated cells and from cells treated with sodium butyrate to increase the levels of acetylated histone. Nuclei from sodium butyrate treated cells exhibited a dramatic increased rate of digestion with DNase I as compared to control cell nuclei. Micrococcal nuclease showed no preference for chromatin containing hyperacetylated histones.     

The action of the adenosylhomocysteine hydrolase inhibitor, 3-deazaadenosine,on phagocytic function of mouse macrophages and human monocytes

An inhibitor of adenosylhomocysteine hydrolase, 3-deazaadenosine, caused profound inhibition of phagocytosis of opsonized erythrocytes by mouse resident peritoneal macrophages $\text{in}\text{vitro}$. The inhibition was evident at concentrations as low as 2×10^?7M, and increased with increasing concentration and time of exposure to the analogue. It was not associated with detachment of the macrophage monolayers or with loss of cell viability. Although the inhibition was not reversible, progression of the functional impairment was interrupted by washing out the analogue. In striking contrast, phagocytic function of human blood monocytes was unaffected by 3-deazaadenosine.     

Bromoacetylcholine as an affinity label of the acetylcholine receptor from Torpedo californica

Bromoacetyl[methyl-³H]choline is a highly specific label for the reduced acetylcholine binding site on the acetylcholine receptor from $\text{Torpedo californica}$. Only one of two binding sites per receptor monomer is susceptible to labeling. The labeled site is on the α chain of the receptor.     

Enhancement of natural and antibody-dependent lymphocyte cytotoxicity by drugs which inhibit prostaglandin production by tumor target cells

Our previous observations suggested that the production of prostaglandins by tumor cells exposed to lymphocytes might constitute a mechanism by which the tumor cells Could subvert the effects of a cellular immune response directed against them. The present experiments tested this hypothesis by determining whether inhibition of prostaglandin production permitted enhanced expression of natural and antibody-dependent lymphocyte cytotoxicity against the target cells. Cell lines T₂₄ and HCV₂₉ were labelled with ⁵¹Chromium and incubated with purified lymphocytes obtained from venous blood of normal donors. Antiserum to T₂₄ and varying concentrations of inhibitors of prostaglandin synthetase (indomethacin, fenclozic acid, acetylsalicylic acid, and 2,6-xylenol) were added at the onset of incubation and assay tubes were incubated for varying times at 37 °C. In some experiments, lymphocytes or labeled target cells were preincubated with inhibitors and then washed prior to their addition to the assay tubes. Cytotoxicity was determined by measuring ⁵¹Chromium release and assessing any differences that might reflect the presence of the various drugs. Each prostaglandin synthetase inhibitor significantly enhanced both natural and antibody-dependent lymphocyte cytotoxicity. Enhancement appeared to reflect an effect on the target cells, presumeably by an inhibition of prostaglandin production. No increase in spontaneous ⁵¹Chromium release was apparent. The inhibitors did not appear to activate lymphocytes. This evidence supports the suggestion of a mechanism in which tumor cells may prevent the effect of a cellular immune response by producing inhibitory levels of prostaglandins. These results also suggest that manipulation of this mechanism can enhance the effectiveness of the lymphocyte response and may be a consideration in assessing lymphocyte/tumor cell interaction in vitro and in vivo.     

Evaluation of fasciotomy and vasodilator for treatment of frostbite in the dog

Severe freezing injury was produced in the hind foot of 26 mongrel dogs. All dogs were given daily whirlpool treatment and protective bandaging for 14 days following injury. In addition, certain dogs received a vasodilator, fasciotomy, or both vasodilator and fasciotomy following injury. Deep foot temperatures, foot volumes, tissue pressures, and 14 day tissue loss-salvage scores were compared. Significant differences between fasciotomy and nonfasciotomy dogs were seen in foot temperature, volume, and tissue pressure immediately following fasciotomy. Though there was no significant difference in 14 day tissue loss, there was clinically apparent prolongation of integrity of the local vascular system for 2 to 5 days following fasciotomy, and total foot salvage in several dogs receiving fasciotomy.