The effect of local nitric oxide synthase inhibition on the diameter of pulpal arteriole in dental bond material-induced vasodilation in rat

Local application of dental bond materials can cause pulpal vasodilation and hyperemia. Such local hemodynamic changes may be mediated by alterations in the levels of locally generated nitric oxide (NO). In different species systemic administration of NO synthase inhibitors leads to a decrease in pulpal blood flow. In contrast, the local administration of these inhibitors has not been tested yet. Therefore, the effect of locally blocked NO synthase on the internal diameter of rat pulpal arterioles under basal conditions and immediately after dental bond material application was studied by using vitalmicroscopic technique. The NO synthase blocker (L-NAME) was locally administered on a thinned dentine layer of the left lower incisor. L-NAME reduced the diameter of the pulpal arteriole both in basal and after bond material-induced hyperaemic conditions. These data suggest that the local formation of NO may have a significant role in the acute vasodilation induced by bond material application and also in maintenance of basal pulpal arteriolar tone.     

Influence of high-calorie (cafeteria) diets on the population of Paneth cells in the small intestine of the rat

A high-calorie (cafeteria) diet is known to cause changes in the intestinal morphology and functioning that seem to be related to calorie overfeeding. Among the cell lineages found in the small intestine epithelium, the Paneth cell (PC) population is known to be influenced by factors related mainly to the intestinal microbiota. The role of PCs in the intestinal cell concert remains unclear, because experimental evidence suggests PC involvement in local processes other than protection against pathogens. Participation of PC in digestive mechanisms has been proposed on this basis. We have analyzed the effect of high-carbohydrate (HC) and high-fat (HF) cafeteria diets on the PC population in the small intestine of the adult rat. For 8 weeks, both HC and HF diets caused a gain in body weight, but whereas the HC-fed rats showed reduced counts of intestinal crypts per 5-mum section, the HF-fed group showed the opposite. In control rats, the number of crypts per section showed a slight tendency to decrease along the duodenum - ileum axis, whereas the number of PCs per crypt was increased towards the ileum. As a result, the number of PCs per section (calculated from these data) remained constant along the three segments of the intestine. The hypercaloric diets did not modify the general tendencies seen in the crypt and PC counts, but reduced the number of PCs per section in the duodenum by 50%. HC-fed, but not HF-fed, rats showed a similar reduction in jejunum also. These changes do not correlate particularly with any of the predictable effects of diet composition, so that a multifactorial control of PC density is proposed.     

C3'-cis-Substituted carboxycyclopropyl glycines as metabotropic glutamate 2/3 receptor agonists: synthesis and SAR studies

The synthesis of a series of C3'-cis-substituted carboxycyclopropyl glycines bearing a wide variety of functional groups is described, and the structure-activity relationship for this series as agonists of group II metabotropic glutamate receptors is reported.     

HCV core protein localizes in the nuclei of nonparenchymal liver cells from chronically HCV-infected patients

Understanding the mechanism of hepatitis C virus (HCV) pathogenesis is an important part of HCV research. Recent experimental evidence suggests that the HCV core protein (HCcAg) has numerous functional activities. These properties suggest that HCcAg, in concert with cellular factors, may contribute to pathogenesis during persistent HCV infection. HCV is capable of infecting cells other than hepatocytes. Although the extrahepatic cellular tropism of HCV may play a role in the pathophysiology of this infection, the precise biological significance of the presence of HCV components in different liver cell types presently remains to be established. In this study, HCcAg was detected in nonparenchymal liver cells of six patients out of eight positive for serum HCV RNA. Immunostaining with anti-HCcAg mAbs revealed the presence of this protein in different liver cell types such as lymphocytes, Kupffer, polymorphonuclear, pit, endothelial, stellate, and fibroblast-like cells. Interestingly, HCcAg was immunolabeled not only in the cytoplasm but also in the nucleus of these cells. Remarkably, HCcAg co-localized with large lipid droplets present in stellate cells and with collagen fibers in the extracellular matrix. Moreover, HCcAg was immunolabeled in bile canaliculus suggesting the involvement of the biliary system in the pathobiology of HCV. Data suggest that nonparenchymal liver cells may constitute a reservoir for HCV replication. Besides, HCcAg may contribute to modulate immune function and fibrosis in the liver as well as steatosis.     

Functional characterization of the yeast Ppz1 phosphatase inhibitory subunit Hal3: a mutagenesis study

Saccharomyces cerevisiae Hal3 is a conserved protein that binds the carboxyl-terminal catalytic domain of the PP1c (protein phosphatase 1)-related phosphatase Ppz1 and potently inhibits its activity, thus modulating all of the characterized functions so far of the phosphatase. It is unknown how Hal3 binds to Ppz1 and inhibits its activity. Although it contains a putative protein phosphatase 1c binding-like sequence (263KLHVLF268), mutagenesis analysis suggests that this motif is not required for Ppz1 binding and inhibition. The mutation of the conserved His378 (possibly involved in dehydrogenase catalytic activity) did not impair Hal3 functions or Ppz1 binding. Random mutagenesis of the 228 residue-conserved central region of Hal3 followed by a loss-of-function screen allowed the identification of nine residues important for Ppz1-related Hal3 functions. Seven of these residues cluster in a relatively small region spanning from amino acid 446 to 480. Several mutations affected Ppz1 binding and inhibition in vitro, whereas changes in Glu460 and Val462 did not alter binding but resulted in Hal3 versions unable to inhibit the phosphatase. Therefore, there are independent Hal3 structural elements required for Ppz1 binding and inhibition. S. cerevisiae encodes a protein (Vhs3) structurally related to Hal3. Recent evidence suggests that both mutations are synthetically lethal. Surprisingly, versions of Hal3 carrying mutations that strongly affected Ppz1 binding or inhibitory capacity were able to complement lethality. In contrast, the mutation of His378 did not. This finding suggests that Hal3 may have both Ppz1-dependent and independent functions involving different structural elements.     

Direct and indirect interrelations between anthropometric and physiological variables in athletic and non-athletic adolescent girls: a path-analytic study

Our previous analysis of anthropometric and exercise test data of 62 athletic and 56 non-athletic girls (age range 10.5-15.5 years) showed that the intensity of habitual exercise failed to discriminate between the group means of the studied variables. However, the patterns of intervariable correlations differed between the subgroups categorized by physical activity. The present paper studies the problem of this difference further by using exploratory multivariate regression of aerobic power (VO2max) on 10 anthropometric variables and age. The VO2(max) regression was significant (Y(nonathletic) = 0.0194x(1) + 0.004x(2) - 0.371x(3) + 0.045x(4) - 0.177x(5) + 0.070x(6) - 0.271x(7) - 0.170x(8) + 0.015x(9) - 0.0005x(10) + 0.185x(11), SEE = 0.37, R2 = 0.71, F(11.44) = 9.63; Y(athletic) = 0.029x(1) + 0.063x(2) + 0.277x(3) - 0.030x(4) - 0.069x(5) + 0.151x(6) - 0.148x(7) + 0.001x(8) + 0.018x(9) - 0.019x(10) - 0.065x(11), SEE = 0.32, R2 = 0.71, F(11.50) = 11.30), but none or only one of the independent variables had a significant partial regression coefficient. The individual VO2(max) estimates were studied in both groups by using the other group's regression formula to rule out sample dependence. Both formulae gave good approximations of the observed values in spite of the dissimilar regression coefficients. The path analysis of the respective criterion-predictor correlation coefficients confirmed that the relationship of the predictor variables with VO2(max) involved quantitatively direct and indirect effects in the non-athletic and athletic groups.     

Mutations in TMEM76* cause mucopolysaccharidosis IIIC (Sanfilippo C syndrome)

Mucopolysaccharidosis IIIC (MPS IIIC, or Sanfilippo C syndrome) is a lysosomal storage disorder caused by the inherited deficiency of the lysosomal membrane enzyme acetyl-coenzyme A: alpha -glucosaminide N-acetyltransferase (N-acetyltransferase), which leads to impaired degradation of heparan sulfate. We report the narrowing of the candidate region to a 2.6-cM interval between D8S1051 and D8S1831 and the identification of the transmembrane protein 76 gene (TMEM76), which encodes a 73-kDa protein with predicted multiple transmembrane domains and glycosylation sites, as the gene that causes MPS IIIC when it is mutated. Four nonsense mutations, 3 frameshift mutations due to deletions or a duplication, 6 splice-site mutations, and 14 missense mutations were identified among 30 probands with MPS IIIC. Functional expression of human TMEM76 and the mouse ortholog demonstrates that it is the gene that encodes the lysosomal N-acetyltransferase and suggests that this enzyme belongs to a new structural class of proteins that transport the activated acetyl residues across the cell membrane.     

Segmental motions of rat thymidylate synthase leading to half‐the‐sites behavior

Thymidylate synthase (TS) is a homodimeric enzyme with two equivalent active sites composed of residues from both subunits. Despite the structural symmetry of the enzyme, certain experimental results are consistent with half‐the‐sites activity, suggesting negative cooperativity between the active sites. To gain insight into the mechanism behind this phenomenon, we explore segmental motions of rat TS in the absence of ligands, with normal mode analysis as a tool. Using solvent accessible surface area of the active site pocket as a monitor of the degree of opening of the active sites, we classified the first 25 nontrivial normal modes, obtained from the web server of the program ElNémo, according to the behavior of the active sites. We found seven modes that open and close both sites symmetrically and nine that do so in an anticorrelated fashion. We characterized the motions of these modes by visual inspection and through measurement of distances between selected atoms lining the active site pockets. The segments that regulate access to the active site correspond to the loop containing R44, helix K, and a long loop containing residues 103–125, in agreement with a large body of crystallographic studies. These elements can be activated together or in isolation. There are more asymmetric modes than symmetric ones in the set we analyzed, probably accounting for the half‐the‐sites behavior of the enzyme. Three of the asymmetric modes result in changes at the dimer interface and indicate the endpoints of possible communication pathways between the active sites. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 549–559, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com