Modulation of chondrocyte migration and aggregation by insulin-like growth factor-1 in cultured cartilage

The effect of insulin-like growth factor-1 (IGF-1) on the behavior of rabbit chondrocytes in cultured collagen (CL) gels initially seeded with 2 × 10⁵ cells/ml was examined. On day 5, the frequency of migrating cells cultured in presence of 100 ng IGF-1/ml was 0.04, which was 54 % of the frequency in IGF-1-free culture. The presence of IGF-1 caused an increase in the frequency of dividing cells from 0.09 to 0.13. These results suggest that IGF-1 suppressed the migration of chondrocytes in the CL gels while stimulating cell division in the initial culture phase. The proteolytic migration of cells was thought to be suppressed by the down-regulation of membrane type 1 matrix metalloproteinase by IGF-1. This contributed to the formation of aggregates with spherical-shaped cells that produced collagen type II.     

Influence of high-pressure homogenization,ultrasonication, and supercritical fluid on free astaxanthin extraction from β-glucanase-treated Phaffia rhodozyma cells

In this study astaxanthin production by Phaffia rhodozyma was enhanced by chemical mutation using ethyl methane sulfonate. The mutant produces a higher amount of astaxanthin than the wild yeast strain. In comparison to supercritical fluid technique, high-pressure homogenization is better for extracting astaxanthin from yeast cells. Ultrasonication of dimethyl sulfoxide, hexane, and acetone-treated cells yielded less astaxanthin than β-glucanase enzyme-treated cells. The combination of ultrasonication with β-glucanase enzyme is found to be the most efficient method of extraction among all the tested physical and chemical extraction methods. It gives a maximum yield of 435.71 ± 6.55 µg free astaxanthin per gram of yeast cell mass.     

Identification of Alkaloid Compounds Arborinine and Graveoline from Ruta angustifolia (L.) Pers for their Antifungal Potential against Isocitrate lyase (ICL1) gene of Candida albicans

Mycopathologia - Candida albicans has been reported globally as the most widespread pathogenic species contributing candidiasis from superficial to systemic infections in immunocompromised...     

Identification of naphthalene metabolism by white rot fungus Pleurotus eryngii

The use of biomaterials or microorganisms in PAHs degradation had presented an eye-catching performance. Pleurotus eryngii is a white rot fungus, which is easily isolated from the decayed woods in the tropical rain forest, used to determine the capability to utilize naphthalene, a two-ring polycyclic aromatic hydrocarbon as source of carbon and energy. In the meantime, biotransformation of naphthalene to intermediates and other by-products during degradation was investigated in this study. Pleurotus eryngii had been incubated in liquid medium formulated with naphthalene for 14 days. The presence of metabolites of naphthalene suggests that Pleurotus eryngii begin the ring cleavage by dioxygenation on C1 and C4 position to give 1,4-naphthaquinone. 1,4-Naphthaquinone was further degraded to benzoic acid, where the proposed terepthalic acid is absent in the cultured extract. Further degradation of benzoic acid by Pleurotus eryngii shows the existence of catechol as a result of the combination of decarboxylation and hydroxylation process. Unfortunately, phthalic acid was not detected in this study. Several enzymes, including manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase and 2,3-dioxygenase are enzymes responsible for naphthalene degradation. Reduction of naphthalene and the presence of metabolites in liquid medium showed the ability of Pleurotus eryngii to utilize naphthalene as carbon source instead of a limited glucose amount.     

Metabolic alteration of <Emphasis Type="Italic">Catharanthus roseus</Emphasis> cell suspension cultures overexpressing <Emphasis Type="Italic">geraniol synthase</Emphasis> in the plastids or cytosol

Previous studies showed that geraniol could be an upstream limiting factor in the monoterpenoid pathway towards the production of terpenoid indole alkaloid (TIA) in Catharanthus roseus cells and hairy root cultures. This shortage in precursor availability could be due to (1) limited expression of the plastidial geraniol synthase resulted in a low activity of the enzyme to catalyze the conversion of geranyl diphosphate to geraniol; or (2) the limitation of geraniol transport from plastids to cytosol. Therefore, in this study, C. roseus’s geraniol synthase (CrGES) gene was overexpressed in either plastids or cytosol of a non-TIA producing C. roseus cell line. The expression of CrGES in the plastids or cytosol was confirmed and the constitutive transformation lines were successfully established. A targeted metabolite analysis using HPLC shows that the transformed cell lines did not produce TIA or iridoid precursors unless elicited with jasmonic acid, as their parent cell line. This indicates a requirement for expression of additional, inducible pathway genes to reach production of TIA in this cell line. Interestingly, further analysis using NMR-based metabolomics reveals that the overexpression of CrGES impacts primary metabolism differently if expressed in the plastids or cytosol. The levels of valine, leucine, and some metabolites derived from the shikimate pathway, i.e. phenylalanine and tyrosine were significantly higher in the plastidial- but lower in the cytosolic-CrGES overexpressing cell lines. This result shows that overexpression of CrGES in the plastids or cytosol caused alteration of primary metabolism that associated to the plant cell growth and development. A comprehensive omics analysis is necessary to reveal the full effect of metabolic engineering.     

Agonist Leukadherin-1 Increases CD11b/CD18-Dependent Adhesion Via Membrane Tethers

Integrin CD11b/CD18 is a key adhesion receptor that mediates leukocyte migration and immune functions. Leukadherin-1 (LA1) is a small molecule agonist that enhances CD11b/CD18-dependent cell adhesion to its ligand ICAM-1. Here, we used single-molecule force spectroscopy to investigate the biophysical mechanism by which LA1-activated CD11b/CD18 mediates leukocyte adhesion. Between the two distinct populations of CD11b/CD18:ICAM-1 complex that participate in cell adhesion, the cytoskeleton(CSK)-anchored elastic elements and the membrane tethers, we found that LA1 enhanced binding of CD11b/CD18 on K562 cells to ICAM-1 via the formation of long membrane tethers, whereas Mn²⁺ additionally increased ICAM-1 binding via CSK-anchored bonds. LA1 activated wild-type and LFA1^−/− neutrophils also showed longer detachment distances and time from ICAM-1-coated atomic force microscopy tips, but significantly lower detachment force, as compared to the Mn²⁺-activated cells, confirming that LA1 primarily increased membrane-tether bonds to enhance CD11b/CD18:ICAM-1 binding, whereas Mn²⁺ induced additional CSK-anchored bond formation. The results suggest that the two types of agonists differentially activate integrins and couple them to the cellular machinery, providing what we feel are new insights into signal mechanotransduction by such agents.     

Rice bran lipase catalyzed esterification of palm oil fatty acid distillate and glycerol in organic solvent

Rice bran lipase (RBL) was delipidated to enhance its stability in organic solvent and its esterification activity at elevated temperature. The esterification activity of delipidated RBL increased as temperature was increased from 45 to 65°C. The esterification activity of delipidated RBL at 65°C was about 14 times greater than that of the non-delipidated RBL. As temperature was further increased to 75°C, the non-delipidated RBL lost all esterification activity, whereas the delipidated RBL retained approximately 48% of its esterilication activity. The delipidated RBL maintained a relative esterification activity greater than 80% after 16 h of incubation in hexane, whereas the non-delipidated RBL maintained a relative esterification activity of only 50%. A method for production of acylglycerol using delipidated RBL to esterify palm oil fatty acid distillate (PFAD) with glycerol in hexane was successfully developed. The effects of reaction temperatures and type of water removal agents (silica gel and molecular sieve) on the degree of esterification were also examined. A 4 h reaction at 65°C, catalyzed by delipidated RBL and using silica gel as the water removal agent resulted in 53.8% esterification. Thin layer chromatography analysis suggested that the esterified product was primarily comprised of mono-and di-acylglycerols.     

Structural characterization of angiotensin I-converting enzyme in complex with a selenium analogue of captopril

Human somatic angiotensin I-converting enzyme (ACE), a zinc-dependent dipeptidyl carboxypeptidase, is central to the regulation of the renin-angiotensin aldosterone system. It is a well-known target for combating hypertension and related cardiovascular diseases. In a recent study by Bhuyan and Mugesh [Org. Biomol. Chem. (2011) 9, 1356-1365], it was shown that the selenium analogues of captopril (a well-known clinical inhibitor of ACE) not only inhibit ACE, but also protect against peroxynitrite-mediated nitration of peptides and proteins. Here, we report the crystal structures of human testis ACE (tACE) and a homologue of ACE, known as AnCE, from Drosophila melanogaster in complex with the most promising selenium analogue of captopril (SeCap) determined at 2.4 and 2.35 ? resolution, respectively. The inhibitor binds at the active site of tACE and AnCE in an analogous fashion to that observed for captopril and provide the first examples of a protein-selenolate interaction. These new structures of tACE-SeCap and AnCE-SeCap inhibitor complexes presented here provide important information for further exploration of zinc coordinating selenium-based ACE inhibitor pharmacophores with significant antioxidant activity.     

A Mutation in SLC24A1 Implicated in Autosomal-Recessive Congenital Stationary Night Blindness

Congenital stationary night blindness (CSNB) is a nonprogressive retinal disorder that can be associated with impaired night vision. The last decade has witnessed huge progress in ophthalmic genetics, including the identification of three genes implicated in the pathogenicity of autosomal-recessive CSNB. However, not all patients studied could be associated with mutations in these genes and thus other genes certainly underlie this disorder. Here, we report a large multigeneration family with five affected individuals manifesting symptoms of night blindness. A genome-wide scan localized the disease interval to chromosome 15q, and recombination events in affected individuals refined the critical interval to a 10.41 cM (6.53 Mb) region that harbors SLC24A1, a member of the solute carrier protein superfamily. Sequencing of all the coding exons identified a 2 bp deletion in exon 2: c.1613_1614del, which is predicted to result in a frame shift that leads to premature termination of SLC24A1 (p.F538CfsX23) and segregates with the disorder under an autosomal-recessive model. Expression analysis using mouse ocular tissues shows that Slc24a1 is expressed in the retina around postnatal day 7. In situ and immunohistological studies localized both SLC24A1 and Slc24a1 to the inner segment, outer and inner nuclear layers, and ganglion cells of the retina, respectively. Our data expand the genetic basis of CSNB and highlight the indispensible function of SLC24A1 in retinal function and/or maintenance in humans.