Families with recurrent cases of Waardenburg-Klein syndrome

Deaf children with the type I Waardenburg--Klein syndrome were studied. Secondary cases were found in 14 unrelated and 1 incest families. In 10 families probands and all their affected relatives had the type I Waardenburg--Klein syndrome. In 4 families patients were discovered with both type I and type II syndromes. In an incest family the proband seemed to have the type III, while his mother and father (sibs) had type II and their grandmother the type I syndrome. These results contradict the hypothesis claiming the origin of different types of Waardenburg--Klein syndrome to be due to the action of different genes. It is proposed that types I and II, or all types of the syndrome are caused by a single dominant gene. Potential mechanisms for clinical polymorphism of Waardenburg--Klein syndrome are related to incomplete penetrance and varying expression of this gene.     

Physical separation of aortic corticoid receptors with type I and type II specificities

Previous gel filtration binding assay studies indicated that rat vascular smooth muscle cells contained corticoid receptor I and corticoid receptor II sites which could be distinguished on the basis of their relative affinities for aldosterone and dexamethasone. Ion-exchange chromatography experiments were designed to separate the two sites for further studies on their physical characteristics and role in vascular smooth muscle cell physiology. Cultured aortic cells were incubated with 5-10 nM 3H steroid alone or in the presence of 10-fold non-radioactive steroid competitor for 30 min at 37 degrees C. Following cell lysis, total cellular protein-bound steroid was isolated using Sephadex G-25 and applied to a DEAE-cellulose ion-exchange column. Three peaks of radioactivity were eluted using a 1-200 mM sodium phosphate gradient: peak I (30-38 mM), peak II (52-64 mM), and peak III (92-102 mM). Peaks I and II contained 60% of the eluted radioactivity and exhibited the same steroid specificity as corticoid receptor II sites (dexamethasone greater than aldosterone). Peak III contained 40% of the eluted radioactivity and exhibited the same steroid specificity as corticoid receptor I sites (aldosterone greater than dexamethasone). These studies support the binding assay data on steroid specificity and relative proportion of type I and II sites. They also document the existence of type I and II corticoid receptors with different physicochemical characteristics in rat aortic smooth muscle cells.     

Isolation and Properties of Two New RNA Phages SP and FI

New RNA phages were isolated from feces of Siamang Gibbon and infants, and their characters were investigated. These phages, SP and FI were serologically unrelated to any of known RNA phages (group I, II and III) and were classified into groups IV and V. Several other characters, such as filtration and elution patterns through membrane filters, buoyant densities in CsCl, and UV sensitivities of SP and FI were similar to those of the group III phages, suggesting that a certain similarity between group III and IV or V might exist. Peculiar phage-host relations found in phages of FI group were also discussed. From these results, we propose new possible schemata for the grouping of RNA phages.     

Modified acetylenic steroids as potent mechanism-based inhibitors of cytochrome P-450SCC

Synthesized 20-(4-tetrahydropyranyl-1-butynyloxy)-5-pregnen-3 alpha,20 beta- diol [steroid I] and 20-(3-tetrahydropyranyl-1-propargyloxy)-5-pregnen- 3 alpha,20 beta-diol [steroid III] have been found to inactivate purified adrenocortical cytochrome P-450SCC. When incubated with the enzyme under turnover conditions, steroid I inactivated cytochrome P-450SCC by about 85% in 40 min. This is in contrast to the free triol analog, steroid II which inactivated the enzyme by only 45% within the same incubation period. A comparison of steroid III with its free triol analog, steroid IV, also showed that the diol is a more effective inactivator of the enzyme than the triol. The partition ratio was calculated by two different methods. Each of the steroids I-IV bound to the enzyme with spectrophotometric dissociation constant (Ks) in the micromolar range, producing Type II low spin spectra changes during titration of the enzyme. In addition, it was found that the binding of each of the compounds to the enzyme occurred without inactivation of the enzyme and that the inactivation under turnover condition, is not as a result of conversion to the denatured P-420 species. This demonstrated that steroids I and III could correctly be designated as mechanism-based (suicide) inhibitors. The kinetic studies demonstrated that steroids with the tetrahydropyranyl substituent are more potent inhibitors of cytochrome P-450SCC as shown by an initial turnover rate of 0.06 min-1, an inactivation rate constant of 0.05 min-1, and a partition ratio of about 1.0 for steroid I. Based on our finding, possible mechanisms of inactivation of cytochrome P-450SCC by these acetylenic steroids are proposed.     

CuZn-Superoxide Dismutases in Rice: Occurrence of an Active, Monomeric Enzyme and Two Types of Isozyme in Leaf and Non-Photosynthetic Tissues

Rice leaves and seed embryos contain four isozymes of CuZn-superoxidedismutase (SOD) and two isozymes of Mn-SOD. CuZn-SOD I is amajor enzyme in leaves, but not in embryos or etiolated seedlings.CuZn-SODs II,III and IV were found in the embryos but were alsofound as minor isozymes in leaves. CuZn-SODs I, II and IV were purified to homogeneity from riceleaves. CuZn-SODs I and II had similar properties with respectto molecular weight, dimeric structure, absorption spectrumand metal content, but their amino acid compositions differedfrom each other. The absorption spectrum of CuZn-SOD IV wassimilar to that of isozymes I and II, but this enzyme was amonomer with a molecular mass of 1.7 kDa. Antibody against CuZn-SODI from rice did not cross-react with isozymes II and IV. Antibodiesagainst CuZn-SOD from spinach leaves cross-reacted with isozymeI but not with isozymes II, III and IV. By contrast, the antibodiesagaist CuZn-SOD from spinach seeds cross-reacted with isozymesII, III and IV but not with isozyme I. Thus, the isozyme thatis expressed mainly in leaves (CuZn-SOD I) and the isozymesexpressed mainly in non-photosynthetic tissues (CuZn-SODs II,III, IV) are immunologically distinct. (Received October 7, 1988; Accepted January 27, 1989)     

Human apolipoprotein E isoprotein subclasses are genetically determined.

In a recent communication, we showed that human very low density lipoprotein (VLDL) apolipoprotein E (Apo E) from different individuals appears upon two-dimensional gel electrophoretic analysis in either one of two complex patterns. These have been designated class alpha and class beta. Mixing of VLDL from different subjects revealed that not all alpha or beta apo E patterns were the same. In this manner, we identified three subclasses of class alpha (alpha II, alpha III, and alpha IV) and three subclasses of class beta (beta II, beta III, and beta IV). We report here the results of family studies that reveal that the subclasses (alpha II, alph III, and alpha IV and beta II, beta III, and beta IV) of apo E are determined at a single genetic locus with three common alleles, epsilon II, epsilon III, and epsilon IV. The class beta phenotypes (beta II, beta III, and beta IV) represent homozygosity for two identical apo E alleles (epsilon). In contrast, class alpha phenotypes (alpha II, alpha III, and alpha IV) represent heterozygosity for two different apo E alleles. The apo E subclasses and their corresponding genotypes are as follows: beta II = epsilon II/epsilon II; beta III = epsilon III; beta IV = epsilon IV/epsilon IV; alpha II = epsilon II/epsilon III; alpha III = epsilon III/epsilon IV; and alpha IV = epsilon II/epsilon IV. To estimate the frequencies of the apo E alleles in the general population, apo E subclasses were then investigated in 61 unrelated volunteers and the results were: beta II = 1 (2%), beta III = 30 (49%), alpha II = 9 (15%, alpha III = 13 (31%), and alpha IV = 2 (3%). Utilizing the frequencies of these phenotypes, the gene frequencies were calculated to be epsilon II = 11%, epsilon III = 72%, and epsilon IV = 17%. In addition, apo E subclasses were studied in a clinic for individuals with plasma lipid disorders and the apo E subclass beta IV was found to be associated with type III hyperlipoproteinemia. There was no association of any apo E subclass with type II, type IV, or type VI hyperlipoproteinemia or plasma HDL cholesterol levels. This study explains the genetic basis for the common variation in a human plasma protein, apo E. Since the apo E subclass beta IV is associated with type III hyperlipoproteinemia, a disease characterized by xanthomatosis and premature atherosclerosis, understanding the genetic basis of the apo E subclasses should provide insight into the genetics of type III hyperlipoproteinemia.     

Iodine concentration in the blood and urine in children depending on the goiter size

Epidemiological investigations have shown that at the Bia?ystok Province about 30% of children and youth is afflicted with goitre. In this area drinking water is poor in iodine and iodine supply with food is quite unsatisfactory. The purpose of the present work has been to check the behaviour of the serum and urine iodine in children with thyroid goitre. The estimations of I in blood serum were made in 126 children with goitre (I, II, and III, according to the WHO classification) and in 100 healthy children. The method used included buthanol extraction according to Fisher and Morris. The concentration of iodine and its excretion rate in urine were assessed in 119 children with goitre of various advancement, an; in 170 children not affected with goitre, using the method of Fisher and Morris adopted to urine analysis. The authors found the following values for serum protein bound iodine: control group 4.9 microgram/100 ml, at average; 4.3 microgram/100 ml, 3.0 microgram/100 ml, and 2.6 microgram/100 ml in those affected with I, II, and III of the goitre, respectively. The 24 hr urine excretion rate of iodine in healthy children amounted to 66.4 microgram, and 64.2 microgram, 53.6 microgram, and 4o.2 microgram in those with I, II, and III goitre, respectively. The above results indicate a significant decrease of iodine concentration in blood serum and in urine in children with goitre; this decrease has been found to be dependent on the degree of the thyroid gland enlargement. The differences were statistically significant.     

Carboxylase Levels and Carbon Dioxide Fixation in Baker''s Yeast

Levels of pyruvate carboxylase (PC), phosphopyruvate carboxylase (PEPC), and malate dehydrogenase (decarboxylating) were compared in wild-type bakers' yeast (I), a cytoplasmic-respiratory mutant (II), a biotin-deficient wild-type yeast (III), and a biotin-deficient respiratory mutant (IV). PC activities were greatly reduced in III and IV, whereas PEPC was reduced in II and IV. Malate dehydrogenase (decarboxylating) could not be detected in any of the yeasts. With yeast I growing on glucose as the sole carbon source, PEPC decreased to negligible levels during the logarithmic phase of growth (glucose repression effect), whereas PC increased. Both enzymes reverted to their original levels during the stationary phase, when glucose in the medium was exhausted. In agreement with the leading role of PC for CO(2) assimilation, the rates of (14)CO(2) fixation in yeasts I and II were approximately equal and were much higher than that in yeast IV. With I and II, most of the (14)C was distributed similarly in oxalacetate derivatives; with yeast IV, most of (14)C appeared in a compound apparently unrelated to CO(2) fixation via C(4)-dicarboxylic acids.     

Comparison of wheat albumin inhibitors of α-amylase and trypsin

Wheat albumins were extracted from whole wheat flour with 150 mM sodium chloride solution and precipitated between 0·4 and 1·8 M ammonium sulphate. The albumin precipitate was separated by gel filtration on Sephadex G100 into five peaks. Three peaks (II, III, and IV), whose MWs were 60 000, 24 000 and 12 500 daltons respectively, were active toward several insect α-amylases, whereas only peak III inhibited human saliva and pancreatic α-amylases. Peaks III and IV also inhibited trypsin. In each active peak, we found several α-amylase inhibitors slightly different in their electrophoretic mobilities in a Tris—glycine buffer system (pH 8·5), whereas only one major trypsin inhibitor was present in peaks III and IV. In contrast to α-amylase inhibitors that were all anodic, trypsin inhibitors migrated to the cathode under our experimental conditions. From a quantitative standpoint, wheat albumins that inhibit trypsin are negligible, whereas about 2/3 of the total albumin inhibits amylases from different origins. All inhibitor components of peak III were active toward both insect and mammalian α-amylases. Moreover, they reversibly dissociated in the presence of 6 M guanidine hydrochloride giving two similar subunits.     

Effects of carvacrol on defects of ischemia-reperfusion in the rat liver

Many plants found in nature have been used to treat various illnesses. One such plant is oregano (Kekik in Turkish). Health beneficial effects of carvacrol obtained from oregano oil have been shown scientifically. We have investigated the comparative effects of carvacrol in the liver of rats subjected to ischemia-reperfusion defect, with silymarin. To test the effects we formed four groups using male Wistar albino rats. Group I was control. The other three groups of animals were administered 60 min prior to surgical operation single doses of physiological serum, carvacrol and silymarin, respectively. Group II, III and IV animal were subjected to 45 min long liver ischemia and 60 min reperfusion. Blood and tissue samples were collected for biochemical and histological analysis following the test.AST and ALT values obtained after biochemical analysis of the serums showed statistically significant difference in group II than the other three groups. A statistical evaluation of the serum AST levels among the groups II, III and IV showed that both groups III and IV which had no difference in between were significantly different in a positive way from group II (p<0.001). As to the serum ALT levels, difference between group II and group III (p<0.001) and group II and group IV (p<0.01) was found significant. No statistical difference was observed in groups I, III and IV for GSH, MDA and CAT levels of the liver. A statistical evaluation of the GSH level in group III and group IV was found to be significantly different from group II (p<0.001) without any difference between them. A similar evaluation for MDA and CAT levels among the revealed no difference between group III and group IV, however, group II showed difference with group II and group IV (p<0.05).Histological findings were in harmony with the biochemical results. We conclude that carvacrol protects the liver against defects caused by ischemia and reperfusion, and carvacrol is not hepatotoxic at the applied dosage.     

Low T3 syndrome in cancer patients in relation to weight loss, intravenous hyperalimentation therapy and age

In order to investigate the relation of weight loss and intravenous hyperalimentation therapy to low T3 syndrome, serum T3, T4. rT3 and TBG were determined by radioimmunoassay in 105 cancer patients. The cancer patients were classified into 3 groups, Group I, II and III depending on the grade of weight loss, ranging up to a 5% change in weight loss from a healthy condition, from 5 to 9%, and more than 10%, respectively. Cancer patients under age 59 showed no significant difference in serum T3, T4, rT3 and TBG among these 3 groups. However serum T3 and T3/T4 in cancer patients at age 60 and over were significantly reduced in group III, compared to groups I and II. Serum rT3 values were significantly elevated in group III of elderly cancer patients. The incidence of low T3 syndrome in group III of elderly cancer patients was also significantly higher than in groups I and II. In three out of 5 cancer patients with low T3 syndrome, serum T3 values increased after the intravenous hyperalimentation therapy, whereas no significant change in serum T3 values was observed in two patients who died within one day after the final examination. It is concluded that weight loss produced different effects on peripheral conversion of T4 to T3 between cancer patients under age 59 and over age 60 and glucose plays an important role in the pathogenesis of low T3 syndrome except cases with very poor prognosis.     

Identification of 10-methylsphinganine in cerebrosides of the guinea pig harderian gland

A large amount of branched long chain bases was detected in the cerebrosides of guinea pig Harderian gland. The long chain bases of cerebrosides were analyzed by GLC as trimethylsilyl derivatives. The branched long chain bases were separated into four peaks (I, II, III, IV) according to the number of carbon atoms and the position of branching. In the present work, the structures of long chain bases in the four peaks were analyzed by GLC and GC-MS after conversion of them to aldehydes, alcohols, and fatty acids. Furthermore the main component of long chain bases (Peak II) was isolated by HPLC as N-acetyl derivatives and analyzed by NMR. The structures of branched long chain bases in Peaks I, II, III, and IV are as follows. Branched long chain bases of Peak I are 2-amino-10- (main component), 2-amino-9-, and 2-amino-8-methylhexadecane-1,3-diol. Branched long chain bases of Peak II also consist of a mixture of 2-amino-10-, 2-amino-9-, and 2-amino-8-methyl-heptadecane-1,3-diol. The branched long chain base of Peak III is 2-amino-10-methyl-octadecane-1,3-diol, while that of Peak IV is 2-amino-16-methyloctadecane-1,3-diol. Among these branched long chain bases, 10-methylsphinganines are dominant though the chain lengths are different. These branched long chain bases, in which the substituted positions exist in the middle part of aliphatic chain (10-, 9-, or 8-methylsphinganine) are novel long chain bases in mammals.     

Biochemical and biophysical properties of flacherie virus of the silkworm

Flacherie virus of the silkworm (FVS) was extracted from diseased silkworms, both larvae and pupae, and purified by 15 to 30% sucrose density gradient centrifugation. FVS III and FVS IV, in addition to the FVS I and FVS II described in the previous paper (Himeno et al., 1974), were found. The FVS I, FVS III, and FVS IV showed the same mobility in 2.4% polyacrylamide gel electrophoresis and could not be distinguished from each other in the gel. However, the purified FVS II was separated into two bands, FVS IIa and FVS IIb, in 2.4% gel. FVS III was a spherical particle with a diameter of 28 ± 1 nm and showed a sedimentation coefficient of about 90 S. FVS III was easily decomposed into FVS IV which sedimented at about 30 S in sucrose gradient centrifugation. FVS I and FVS II each contained a single molecule of RNA which showed the same molecular weight. FVS I consisted of three polypeptides with molecular weights of 67,000, 50,000, and 33,000. FVS II consisted of 10 polypeptides; among them 2 polypeptides with molecular weights of 50,000 and 33,000 were also found. Labeling experiments with [³²P]orthophosphate revealed that FVS II was found at an early stage of infection and FVS I at a late stage. FVS II was also isolated at an early stage from silkworms infected with FVS II, and FVS I was found at a late stage in these silkworms. The correlation among FVS I, FVS II, FVS III, and FVS IV was discussed and it was suggested that they might be closely related to one another and that few particles in them were immature. It is possible that FVS II changes to FVS I via FVS III by cleavage of large polypeptides.     

Acid phosphatases from liver of Rana esculenta. Subcellular localization and partial characterization of multiple forms

1. Four acid phosphatases (AcPase I, II, III and IV) were found in the liver of the frog Rana esculenta. 2. AcPases I, II, III, and IV were associated with the microsomal, mitochondrial-lysosomal, nuclear and soluble fractions respectively and showed apparent molecular weights of about 240,000, 110,000, 38,000 and 17,000. 3. All the enzymes show acid pH optima, and similar Km values for p-nitrophenylphosphate. 4. AcPases I, II, and III hydrolyze most of the common phosphate esters whereas AcPase IV hydrolyzes effectively only p-nitrophenylphosphate, phenylphosphate and flavine mononucleotide. 5. AcPases I and II are inhibited by NaF and tartrate. AcPases III and IV are tartrate resistant. 6. Temperature inhibits AcPases I, II, IV, whereas it activates AcPase III.     

Blood and urine iodine levels in patients with gastric cancer

In this study, we aimed to investigate whether there is any relationship between gastric cancer and iodine concentrations in blood and urine in the northeast Anatolia region, where iodine deficiency is common. A total of 56 patients, diagnosed as gastric cancer and 25 healthy volunteers were included in the study. The methods used were based on the Sandell-Kolthoff reaction. The urine iodine concentration (UIC) and serum protein-bound iodine (PBI) levels were higher in patients with gastric cancer compared with healthy control subjects. The UIC in stage IV was higher than all other stages and the control group. The UIC was higher in stages III and IV compared with stages I and II. However, serum PBI levels in stage III were higher compared with stages I, and II and also control group. The serum PBI level in stage IV was higher than stage II and the control group. In the patient and control groups, there were no significant differences in serum PBI and UIC with regard to age or sex. Our results suggested that urinary and blood iodine concentration might be a useful marker for following the disease.     

Neonatal urinary steroids in Smith-Lemli-Opitz syndrome associated with 7-dehydrocholesterol reductase deficiency.

The biosynthetic abnormality in Smith-Lemli-Opitz syndrome (SLOS) is a deficiency of 7-dehydrocholesterol (7DHC) reductase, the enzyme responsible for catalyzing the final step in the Kandutsch-Russell pathway for cholesterol synthesis. Because the disposition of 7DHC and 8-dehydrocholesterol [8DHC; cholesta-5,8(9)-dien-3beta-ol] produced in this syndrome is little understood, we have analyzed urine from three young infants by gas chromatography/mass spectrometry to characterize its steroid metabolites. All steroid metabolites of adrenal origin found in normal infant urine were also found in urine from the patients with SLOS but in reduced amount. Quantitatively, the major steroids in these SLOS patients were identified by mass spectrometry as homologs of normal neonatal steroids possessing an additional double bond. Generally, two forms of each steroid were present in a similar amount. Because of the markedly increased levels of 7DHC and 8DHC in SLOS, these almost certainly represented the 5,7 and 5,8(9) unsaturated forms of each metabolite. The most abundant steroids were tentatively identified as 3beta,16alpha-dihydroxy-5,7-pregnadien-20-one and 3beta,16alpha-dihydroxy-5,8(9)-pregnadien-20-one, although similar 21-hydroxylated steroids and homologs of 16alpha-hydroxy-DHEA were also found. This study shows that all enzymatic steps used by cholesterol in the DHEA synthetic pathway are also functional for 7DHC and 8DHC.     

The effect of adjuvant and specific or non-specific vaccination on development of protective immunity of rabbits against Trichostrongylus colubriformis infection.

Adult New Zealand rabbits were vaccinated subcutaneously with one dose of 100 micrograms adult nematode phosphate buffered saline-soluble proteins (PBS-ASP, groups I and II), a detergent-soluble fraction of adult somatic proteins (DS-ASP, group III) or three doses of 1 mg normal rabbit serum proteins (group IV). Injections of the immunogens in groups II, III and IV were accompanied with beryllium hydroxide, Be(OH)2 as an adjuvant. Vaccinated rabbits and also those of group V (naive) were challenged orally with 10,000 infective larvae of T. colubriformis 14 days after antigen injection and necropsied 2 weeks later. A single dose of PBS-ASP induced 33.5% protection when the antigen was given alone (group I) and 69.4% when injected with Be(OH)2 (group II). A detergent-soluble fraction of ASP given with the adjuvant provided 87.2% protection (group III), whilst non-specific vaccination with serum proteins plus Be(OH)2 elicited 99% protection (group IV). Mesenteric lymph node leukocyte responses were measured using a leukocyte migration inhibition assay. A significant response was observed only in group IV. In ELISA tests IgA antibodies specific to PBS-ASP reached the highest level in the intestinal mucosa of groups I and II and in the bile of groups I and III. Antibody levels of IgG isotype were similar in the intestinal mucosa of all the immunized groups. Nematode antigen was detected using a 'sandwich' ELISA method in faecal protein extracts of rabbits of groups II and III on days 2-6 after challenge.     

Soluble adenosine diphosphate glucose–α-1,4-glucan α-4-glucosyltransferases from spinach leaves

Multiple forms of ADP-glucose-alpha-1,4-glucan alpha-4-glucosyltransferase were obtained from spinach leaves by gradient elution from a DEAE-cellulose column. In the presence of high concentrations of some salts and bovine serum albumin, unprimed activity was found in one (transglucosylase III) of the four fractions eluted from the column. In addition to having unprimed activity, transglucosylase III had a lower K(m) for ADP-glucose, a much higher K(m) for oyster glycogen, greater heat sensitivity and lower affinity for maltose, maltotriose and amylopectin beta-limit dextrin than fractions I, II and IV. In addition, the kinetics at low concentrations of amylose, amylopectin and rabbit liver glycogen were non-linear for transglucosylase III. The properties of transglucosylases I, II and IV were generally similar to each other. Rates of the unprimed reaction at physiological concentrations of ADP-glucose were greater than those found for the primed reaction of fraction III. The product formed by the unprimed reaction was a glucan containing principally alpha-1,4 linkages with some alpha-1,6 linkages. The primer, maltose, at a concentration of 0.5m inhibited the synthesis of the unprimed product.     

Maternal cocaine use and genitourinary tract malformations

A specific fetal disruption syndrome associated with maternal cocaine use has not as yet been documented. In the first 23 pregnancies evaluated in our program for chemically dependent women, one infant with prune belly syndrome and one infant with hypospadias were delivered to cocaine-using women. In the present study, all infants delivered in our program received a renal ultrasound at 2-3 days of life. Fifty infants born to cocaine-using women (group I) and 30 born to polydrug, noncocaine-using women (group II) were evaluated. The two groups of women were similar for age, gravidity, and race. Mean birth weight of the infants in the two groups was similar. Seven infants in group I demonstrated genitourinary tract malformations while there were no infants with such malformations in group II. It is hypothesized that the vasoconstrictive effects of cocaine could explain the abnormalities of the genitourinary tract found in the cocaine-exposed infants.