Polyamine-sensitive protein kinase from chick intestinal mucosa

Summary A cyclic nucleotide-independent protein kinase which phoshorylates preferentially acidic proteins such as casein or phosvitin was isolated from cytosol of chick duodenal mucosa. The enzyme was purified more than 633 fold to apparent homogeneity by ammonium sulfate fractionation, column chromatography on DEAE-cellulose, phosphocellulose, hydroxylapatite and by sucrose density gradient centrifugation. The native enzyme has a molecular weight of 131000 as measured by gel filtration. The enzyme is a complex protein containing three polypeptides of molecular weight of 39 000, 36 000 and 27 000. It behaves as a complex throughout its purification and gel filtration but its components are readily separated by electrophoresis in denaturing buffer. The 27 000 molecular weight band was selectively autophosphorylated when the enzyme was incubated in the presence of [-³²P]ATP.When casein was used as substrate, physiological concentrations of naturally occurring polyamines such as spermine and spermidine markedly stimulated enzyme activity. However with phosvitin as substrate polyamines were strong inhibitors of the enzyme activity. This contrasting effect on intestinal kinase activity was also apparent using cytoplasmic proteins as endogenous phosphate acceptors. A characterization of this differential effect is presented and some possible physiological implications are discussed.     

Human hydatidosis in Sardinia. Epidemiologic study of the cases operated on from 1974 to 1981

A retrospective survey of surgical cases to obtain baseline data on hydatid disease in Sardinia from 1974 to 1981 revealed an annual mean rate of 11.1/100.000 population. However, real prevalence is obviously higher, since infection is not always synonymous with disease and surgical incidence should be regarded as the figure that counts. Information on sex, age, residence, occupation, cyst location, number of re-operations was collected to assess the public health impact to hydatidosis within the island. The obtained results indicate that not all population is at equal risk, being hydatid disease most prevalent in rural areas and particularly in the districts where sheep-breeding is highly diffused (annual mean rate greater than 20/100.000 population). The highest rate was observed in farmers and shepherds (34.2/100.000) respect to retired (14.1/100.000), housewives (10.9/100.000), employed in services (8.9/100.000) and students (8.3/100.000). Over 55% of the cysts were found in the liver, about 30% in the lung and 15% in other sites. A correlation between age and cyst location and between profession and cyst location was shown. Pulmonary cysts were prevalent in children and young people, hepatic in grown-up people, whereas nearly the same hepatic and pulmonary frequency was observed in shepherds. Comparisons between previous surveyed periods were done and results were discussed, suggesting the need of a continuous and well-planned control programme.     

Polymorphism of human C2 detected by immunoblotting

Summary To allotype human complement component C2, thin layer agarose gel isoelectric focusing of human serum and/or EDTA-plasma was performed followed by direct immunofixation or by immunoblotting with a specific antihuman-C2 antibody. Using reference samples for C2 BC phenotypes and local samples from an HLA, C4, and Bf genotyped family, a differentiation of the C2^*B and C2^*C variants segregating with the respective HLA haplotype was achieved. The C2 BC phenotype is characterized by a double banding pattern similar to that observed in the haemolytic overlay assay usually used for the detection of C2 polymorphism.An homozygous C2^*Q0 reference sample determined by functional assays was shown to be biochemically deficient, as demonstrated by immunofixation and immunoblotting. The visual interpretation of C2 phenotypes was definitely easier after immunofixation and immunoblotting than after an haemolytic overlay assay. In addition, the method for C2 allotyping described here has several advantages, in particular it saves time and tolerates repeated thawing and freezing of the samples.     

Induction of autolysis of staphylococci by the basic peptide antibiotics Pep 5 and nisin and their influence on the activity of autolytic enzymes

Pep 5 and nisin are cationic bactericidal peptides which were shown to induce autolysis in Staphylococcus cohnii 22. In contrast to nisin, Pep 5 induced lysis could be stimulated in the presence of glucose. Addition of lipoteichoic acids (LTA) (d-alanine:phosphorus=0.475:1) inhibited all effects of Pep 5 on susceptible cells in a molar ratio LTA:Pep 5 of 10:1. Treatment of S. cohnii 22 with Pep 5 or nisin for 20 min and subsequent washing with 2.5 M NaCl released autolysin activity. Crude preparations of the hydrolyzing enzymes produced free amino groups as well as polysaccharide fragments from the murein backbone, suggesting the presence of a muramidase or glucosamidase, and endopeptidase or amidase. Both enzyme activities were inhibited by lipoteichoic acid; they could be fully reactivated by addition of Pep 5 in sufficient concentrations. The velocity of hydrolysis was not influenced by nisin, whereas it was doubled in presence of Pep 5. The results are discussed in view of a possible mechanism of induction of lysis by Pep 5 and nisin.Abbreviations A.U. arbitrary unit - CCCP carbonylcyanide-m-chlorophenyl hydrazone - DNase deoxyribonuclease - CYG casein yeast extract glucose - IT initial turbidity - LTA lipoteichoic acid - RNase ribonuclease - TSB Tryptone Soy Broth     

Chromatofocusing of apolipoproteins from human serum high density lipoprotein

Human HDL was delipidated and the apolipoproteins were fractionated by chromatofocusing. Chromatofocusing, which separates proteins due to their differing isoelectric points, resulted in 8 peaks with corresponding pI values of 7.40, 6.92, 6.64, 5.48, 5.30, 5.18, 4.92 and 4.63. By one single chromatofocusing run four apolipoproteins were obtained in pure form. Two additional polypeptides could be purified during the desalting step using phenyl-Sepharose.     

Immunological memory and lymphoblast-migration in mice infected with Hymenolepis nana

The presence of small cells carrying memory and lymphoblast migration in C57 Bl/6N inbred mice with the intestinal parasite Hymenolepis nana were investigated. Hymenolepis nana egg-infection stimulated an enhanced accumulation of mesenteric lymphoblasts at days 3, 6 and 9 after infection; lymphoblasts accumulated selectively in the mesenteric nodes (MLN) of mice suggesting a cell-trapping effect. The migration was studied using lymphoblasts from non-infected donors. Spleen cells and MLNC collected from donor mice 30 days after a primary infection and enriched for T cells were able to transfer an adoptive immunity, by contrast unseparated cells were uneffective. This result provides preliminary evidence for the existence of T memory cells in the spleen and in the mesenteric nodes.     

Modulation of cyclic nucleotide-independent protein kinase from chick intestine by naturally occurring polyamines and mucopolysaccharides

Summary Chick duodenal mucosa contains an endogenous factor which is capable to inhibit selectively a homologous polyamine-sensitive protein kinase. The inhibitor was partially purified and characterized, and it was found to contain typical mucopolysaccharidic components.Glycosidases digestion studies, selective degradation analysis and spectrophotometric titrations with metachromatic dyes indicated that the inhibitor preparation contained two major moieties identified as heparin-like and heparan sulfate-like structures. In chick intestine the inhibitor was specific for polyamine-sensitive protein kinase since selectively interacted with it and was inert towards other cAMP-independent and cAMP-dependent protein kinases. The inhibitory effect of the endogenous factor was counteracted by naturally occurring polyamines such as spermine. The order of potency of various polyamines was: spermine > thermine spermidine diamines. The release of inhibition by addition of physiological concentrations of spermine was also apparent when using cytosolic proteins as endogenous phosphate acceptors. These results suggest that a possible role of polyamine in the regulation of polyamine-sensitive protein kinase in the intestine is to protect the enzyme from the inhibitory action of endogenous heparinoids.     

The atokous-epitokous border is determined before the onset of heteronereid development in Platynereis dumerilii (Annelida,Polychaeta)

Summary In most nereids sexual maturation is accompanied by a dramatic reorganization of the body that enables swarming of the formerly benthic worms. However, a border exists between unchanged anterior (atokous) and metamorphosed posterior (epitokous) segments. The site of this atokous-epitokous border (a/e border) is different in sexually mature males and females of Platynereis dumerilii. There is no correlation between the total number of setigerous segments of a specimen and the location of the a/e border. The location of the a/e border and sexual development are affected neither by cutting off caudal segments of juveniles (including the prospective a/e border) nor by transecting the ventral nerve cord. When parapodia are transplanted from prospective epitokous regions to prospective atokous regions and vice versa, they maintain their original character during metamorphosis. The results presented here suggest that prospective atokous as well as epitokous characters are determined at or only very shortly after formation of the respective segments. Thus the a/e border is established well in advance of the onset of epitokous metamorphosis.     

Epidemiology of hydatid disease in Sardinia: a study of fertility of cysts in sheep

Hydatidosis, caused by Echinococcus granulosus, is a cyclozoonotic disease of economic significance in Sardinia. The life-cycle involves stray and sheep dogs as definitive hosts and sheep, pigs, goats and cattle as intermediate hosts. The most important intermediate host is sheep, due to home slaughtering with ready access of the viscera to dogs. This survey was undertaken in 1987 to ascertain the epidemiological significance of sheep in maintaining the life-cycle. A total of 700 (91.3%) of 767 sheep harboured hydatid cysts. The frequency distribution of the number of hydatid cysts was over-dispersed. Of 497 infected sheep, 7.6% had fertile cysts, 75.7% sterile cysts and 16.7% fertile + sterile cysts.