Purification, characterization and immunolocalization of fimbrial protein from Porphyromonas (bacteroides) gingivalis.

Rapid and reproducible method is described here for the purification of the 43 kDa fimbrial protein from P. gingivalis by preferential fractionation in the presence of 1% SDS and 0.2M of a bivalent cation at pH 6.5. Homogeneity of the purified 43 kDa was confirmed by SDS-PAGE and Western blot analysis using monoclonal and polyclonal antibodies raised against this protein. Amino acid composition and the amino acid sequence of the first 30 amino acid residues of the purified fimbriae are consistent with the composition and sequence predicted from the cloned gene of the fimbrial subunit. Circular dichroism spectra shows high levels of beta-sheet structure. The purified 43 kDa polymer shows fimbriae-like morphology under the electron microscope. Ultrastructural localization of the 43 kDa protein by the immunogold technique revealed specific labeling of the fimbriae with a diameter of approximately 3.5 to 5.0 nm. Localization of this protein suggest that the 43 kDa component is a fimbrial subunit.     

Design,synthesis, and evaluation of curcumin analogues as potential inhibitors of bacterial sialidase

Sialidases are key virulence factors that remove sialic acid from the host cell surface glycan, unmasking receptors that facilitate bacterial adherence and colonisation. In this study, we developed potential agents for treating bacterial infections caused by Streptococcus pneumoniae Nan A that inhibit bacterial sialidase using Turmeric and curcumin analogues. Design, synthesis, and structure analysis relationship (SAR) studies have been also described. Evaluation of the synthesised derivatives demonstrated that compound 5e was the most potent inhibitor of S. pneumoniae sialidase (IC₅₀?=?0.2?±?0.1?µM). This compound exhibited a 3.0-fold improvement in inhibitory activity over that of curcumin and displayed competitive inhibition. These results warrant further studies confirming the antipneumococcal activity 5e and indicated that curcumin derivatives could be potentially used to treat sepsis by bacterial infections.     

Effectiveness of Losartan-Loaded Hyaluronic Acid (HA) Micelles for the Reduction of Advanced Hepatic Fibrosis in C3H/HeN Mice Model

Advanced hepatic fibrosis therapy using drug-delivering nanoparticles is a relatively unexplored area. Angiotensin type 1 (AT1) receptor blockers such as losartan can be delivered to hepatic stellate cells (HSC), blocking their activation and thereby reducing fibrosis progression in the liver. In our study, we analyzed the possibility of utilizing drug-loaded vehicles such as hyaluronic acid (HA) micelles carrying losartan to attenuate HSC activation. Losartan, which exhibits inherent lipophilicity, was loaded into the hydrophobic core of HA micelles with a 19.5% drug loading efficiency. An advanced liver fibrosis model was developed using C3H/HeN mice subjected to 20 weeks of prolonged TAA/ethanol weight-adapted treatment. The cytocompatibility and cell uptake profile of losartan-HA micelles were studied in murine fibroblast cells (NIH3T3), human hepatic stellate cells (hHSC) and FL83B cells (hepatocyte cell line). The ability of these nanoparticles to attenuate HSC activation was studied in activated HSC cells based on alpha smooth muscle actin (α-sma) expression. Mice treated with oral losartan or losartan-HA micelles were analyzed for serum enzyme levels (ALT/AST, CK and LDH) and collagen deposition (hydroxyproline levels) in the liver. The accumulation of HA micelles was observed in fibrotic livers, which suggests increased delivery of losartan compared to normal livers and specific uptake by HSC. Active reduction of α-sma was observed in hHSC and the liver sections of losartan-HA micelle-treated mice. The serum enzyme levels and collagen deposition of losartan-HA micelle-treated mice was reduced significantly compared to the oral losartan group. Losartan-HA micelles demonstrated significant attenuation of hepatic fibrosis via an HSC-targeting mechanism in our in vitro and in vivo studies. These nanoparticles can be considered as an alternative therapy for liver fibrosis.     

Modulation of Intracellular Calcium Levels by Calcium Lactate Affects Colon Cancer Cell Motility through Calcium-Dependent Calpain

Cancer cell motility is a key phenomenon regulating invasion and metastasis. Focal adhesion kinase (FAK) plays a major role in cellular adhesion and metastasis of various cancers. The relationship between dietary supplementation of calcium and colon cancer has been extensively investigated. However, the effect of calcium (Ca²⁺) supplementation on calpain-FAK-motility is not clearly understood. We sought to identify the mechanism of FAK cleavage through Ca²⁺ bound lactate (CaLa), its downstream signaling and role in the motility of human colon cancer cells. We found that treating HCT116 and HT-29 cells with CaLa immediately increased the intracellular Ca²⁺ (iCa²⁺) levels for a prolonged period of time. Ca²⁺ influx induced cleavage of FAK into an N-terminal FAK (FERM domain) in a dose-dependent manner. Phosphorylated FAK (p-FAK) was also cleaved in to its p-N-terminal FAK. CaLa increased colon cancer cells motility. Calpeptin, a calpain inhibitor, reversed the effects of CaLa on FAK and pFAK cleavage in both cancer cell lines. The cleaved FAK translocates into the nucleus and modulates p53 stability through MDM2-associated ubiquitination. CaLa-induced Ca²⁺ influx increased the motility of colon cancer cells was mediated by calpain activity through FAK and pFAK protein destabilization. In conclusion, these results suggest that careful consideration may be given in deciding dietary Ca²⁺ supplementation to patient undergoing treatment for metastatic cancer.     

Salmonella serotypes isolated from human enteric processes in Recife, Pernambuco, during 1978-1980

From 13,196 faecal cultures made in Recife-Pernambuco during the period from 1978 to 1980, 1,720 strains of Salmonella were isolated. Serological typing on 1,387 of the isolates recognized 63 serotypes, 73.18% of which belonged to group B. The prevalent serotypes adding up to 1,231 strains (88.75% of the total of the isolates) were: S. typhimurium, S. saint-paul, S. poona, S. derby, S. agona, S. newport, S. oranienburg, S. infantis, S. tshiongwe and S. ndolo.     

The distribution of zinc and copper in plasma, erythrocytes and erythrocyte membranes of rainbow trout (Salmo gairdneri)

1. The zinc and copper concentration of plasma was determined in rainbow trout, lake trout, walleye and whitefish. 2. These fish had mean plasma zinc concentrations ranging from 9.3 to 15.1 ppm and copper concentrations from 0.6 to 1.3 ppm. 3. In rainbow trout, the concentration of zinc and copper is greater in the erythrocyte membrane than in the total erythrocyte. 4. Ultrafilterable plasma zinc and copper concentration in rainbow trout was determined to be 0.03 and 0.019 ppm, respectively. 5. Dialysis of rainbow trout plasma against 20 mM EDTA results in removal of 99% of the zinc and 88% of the copper from plasma proteins.