Surface modification of chitosan films. Effects of hydrophobicity on protein adsorption

The surface of chitosan films was modified using acid chloride and acid anhydrides. Chemical composition at the film surface was analyzed by attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy (XPS). ATR-FTIR data verified that the substitution took place at the amino groups of chitosan, thus forming amide linkages, and the modification proceeded to the depth at least 1 microm. Choices of molecules substituted at the amino groups of the glucosamine units did affect the hydrophobicity of the film surface, as indicated by air-water contact angle analysis. The surface became more hydrophobic than that of non-modified film when a stearoyl group (C(17)H(35)CO-) was attached to the films. The reaction of chitosan films with succinic anhydride or phthalic anhydride, however, produced more hydrophilic films. Selected modified films were subjected to protein adsorption study. The amount of protein adsorbed, determined by bicinchoninic acid (BCA) assay, related to the types of attached molecules. The improved surface hydrophobicity affected by the stearoyl groups promoted protein adsorption. In contrast, selective adsorption behavior was observed in the case of the chitosan films modified with anhydride derivatives. Lysozyme adsorption was enhanced by H-bonding and charge attraction with the hydrophilic surface. While the amount of albumin adsorbed was decreased possibly due to negative charges that gave rise to repulsion between the modified surface and albumin. This study has demonstrated that it is conceivable to fine-tune surface properties which influence its response to bio-macromolecules by heterogeneous chemical modification.     

Comprehensive Evaluation of the Association of APOE Genetic Variation with Plasma Lipoprotein Traits in U.S. Whites and African Blacks

Although common APOE genetic variation has a major influence on plasma LDL-cholesterol, its role in affecting HDL-cholesterol and triglycerides is not well established. Recent genome-wide association studies suggest that APOE also affects plasma variation in HDL-cholesterol and triglycerides. It is thus important to resequence the APOE gene to identify both common and uncommon variants that affect plasma lipid profile. Here, we have sequenced the APOE gene in 190 subjects with extreme HDL-cholesterol levels selected from two well-defined epidemiological samples of U.S. non-Hispanic Whites (NHWs) and African Blacks followed by genotyping of identified variants in the entire datasets (623 NHWs, 788 African Blacks) and association analyses with major lipid traits. We identified a total of 40 sequence variants, of which 10 are novel. A total of 32 variants, including common tagSNPs (≥5% frequency) and all uncommon variants (<5% frequency) were successfully genotyped and considered for genotype-phenotype associations. Other than the established associations of APOE*2 and APOE*4 with LDL-cholesterol, we have identified additional independent associations with LDL-cholesterol. We have also identified multiple associations of uncommon and common APOE variants with HDL-cholesterol and triglycerides. Our comprehensive sequencing and genotype-phenotype analyses indicate that APOE genetic variation impacts HDL-cholesterol and triglycerides in addition to affecting LDL-cholesterol.     

Control of nanobiointerfaces generated from well-defined biomimetic polymer brushes for protein and cell manipulations

To better understand protein/material and cell/material interactions at the submolecular level, well-defined polymer brushes consisting of poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) on silicon wafers were prepared by atom transfer radical polymerization (ATRP). Silicon wafers were treated with 3-(2-bromoisobutyryl)propyl dimethylchlorosilane (BDCS) to form a monolayer that acts as initiators for ATRP. Silicon-supported BDCS monolayers were soaked in a methanol/water mixture solution containing Cu(I)Br, bipyridine, and a sacrificial initiator. After MPC was added to the solution, ATRP was carried out for 18 h. The molecular weight and thickness of the PMPC brush layer on the silicon surface increased with an increase in the polymerization time. The dense polymer brushes were obtained by the "grafting from" system. By selective decomposition of the BDCS monolayer by UV light-irradiation, the PMPC brush region and the sizes were well controlled, resulting in fabricating micropatterns of the PMPC brushes. When the thickness of the PMPC brush layer was greater than 5.5 +/- 1.0 nm (3 h polymerization), serum protein adsorption and fibroblast adhesion were effectively reduced, i.e., proteins and cells could recognize such thin polymer brushes on the surface. In addition, the density of the adherent cells on the patterned PMPC brush surface could be controlled by changing the size of the pattern.     

Lipoprotein lipase gene sequencing and plasma lipid profile

Lipoprotein lipase (LPL) plays a crucial role in lipid metabolism by hydrolyzing triglyceride (TG)-rich particles and affecting HDL cholesterol (HDL-C) levels. In this study, the entire LPL gene plus flanking regions were resequenced in individuals with extreme HDL-C/TG levels (n = 95), selected from a population-based sample of 623 US non-Hispanic White (NHW) individuals. A total of 176 sequencing variants were identified, including 28 novel variants. A subset of 64 variants [common tag single nucleotide polymorphisms (tagSNP) and selected rare variants] were genotyped in the total sample, followed by association analyses with major lipid traits. A gene-based association test including all genotyped variants revealed significant association with HDL-C (P = 0.024) and TG (P = 0.006). Our single-site analysis revealed seven independent signals (P < 0.05; r² < 0.40) with either HDL-C or TG. The most significant association was for the SNP rs295 exerting opposite effects on TG and HDL-C levels with P values of 7.5.10⁻⁴ and 0.002, respectively. Our work highlights some common variants and haplotypes in LPL with significant associations with lipid traits; however, the analysis of rare variants using burden tests and SKAT-O method revealed negligible effects on lipid traits. Comprehensive resequencing of LPL in larger samples is warranted to further test the role of rare variants in affecting plasma lipid levels.     

Oil palm (Elaeis guineensis Jacq.) linkage map,and quantitative trait locus analysis for sex ratio and related traits

Sex ratio and shell-thickness type are among the main components determining yield in oil palm. An integrated linkage map of oil palm was constructed based on 208 offspring derived from a cross between two tenera palms differing in inherited sex ratio. The map consisted of 210 genomic simple sequence repeats (SSRs), 28 expressed sequence tag SSRs, 185 amplified fragment length polymorphism markers, and the Sh locus, which controls shell-thickness phenotype, distributed across 16 linkage groups covering 1,931 cM, with an average marker distance of 4.6 cM. Quantitative trait locus (QTL) analysis identified eight QTLs across six linkage groups associated with sex ratio and related traits. These QTLs explained 8.1–13.1 % of the total phenotypic variance. The QTL for sex ratio on linkage group 8 overlapped with a QTL for number of male inflorescences. In most cases a specific QTL allele combination was responsible for genotype class mean differences, suggesting that most QTLs in heterozygous oil palm are likely to be segregating for multiple alleles with different degrees of dominance. In addition, two new SSRs were shown to flank the major Sh locus controlling the fruit variety type in oil palm.     

Schistosoma japonicum (Chinese): Changes of the tegument surface in cercariae, schistosomula and juvenile parasites during development

, , , and 1988. Schistosoma japonicum (Chinese): changes of the tegument surface in cercariae, schistosomula and juvenile parasites during development. International Journal for Parasitology18: 1093–1104. The surface of cercariae of S. japonicum (Chinese) and changes following their transformation into schistosomula were observed at 0.5,1, 3,6,12,24 and 48 hand 5,7,10 and 15 days by scanning electron microscopy. The cercarial surface shows two distinct regions: on the head and the body it is mildly undulating and covered throughout with numerous spines that are partially embedded in a thick layer of glycocalyx; on the tail it is highly folded into tall and thin circumferential ridges, spines are confined to the dorsal and ventral aspects and are prominent due to a thinner glycocalyx. Between 0.5 and 12 h following skin penetration, the surface of schistosomula is gradually cleared of glycocalyx, while small blebs and slender microvilli project out from the glycocalyx-free area and from spines. Subsequently, these structures are detached from the surface and thus responsible for the shedding of the original cercarial membrane and glycocalyx. Between 24 and 48 h, schistosomula show little change in the body size in comparison to earlier stages, and most of the surface is still covered with spines but devoid of blebs and microvilli. From day 2 onwards schistosomula are lengthened but with little net growth. Spines decrease in the middle region of the body; and the surface is invaginated by small pits and folded into thin ridges, which result in rapid increase of the surface area. Between days 5 and 7 schistosomula elongate further, and the disappearance of the spines in the middle region becomes even more pronounced, while those remaining are confined only to the anterior and posterior extremities. Ridges and pits are more developed, and the former begin to show regular foldings throughout all parts of the body. Between days 7 and 10 schistosomula grow rapidly and the body is quadrupled in size. Between days 10 and 15 sexes can be differentiated. The male has a flat ovoid shape body, whose ridges and pitting on the surface are highly developed almost to the same extent as in adult parasites, while spines remain mostly on the posterior end. Sensory papillae greatly increase in number, especially on the ventral and lateral aspects and around the suckers. The surface of the female is relatively smooth; ridges are developed only on the posterior part, while the remaining surface is flat and extensively pitted. Spines are present throughout but less concentrated in the middle region.     

Effects of ozone treatment on cell growth and ultrastructural changes in bacteria

Ozone appeared to inhibit growth and caused the death of gram negative and gram positive tested bacteria: Escherichia coli, Salmonella sp., Staphylococcus aureus and Bacillus subtilis. Bacterial cultures at 10(3), 10(4), 10(5), 10(6), and 10(7) cfu/ml dilution were exposed to 0.167/mg/min/L of ozone at different time intervals (0, 5, 10, 15, 30, 60, 90, 120, and 150 min). Cell viability was observed in all types of tested bacteria at 10(3), 10(4), 10(3) cfu/ml within 30 min after ozone exposure. However, cell inactivation was not significantly observed at concentrations of 10(6), 10(7) cfu/ml even after an exposure of 150 min. Ultrastructural changes of treated bacteria showed deformation, rough damage and surface destruction revealed by scanning electron microscopy. Some bacterial cells showed collapsed and shrunken patterns within 60 min and severe rupture and cellular lysis after 90 min of ozone treatment. This study supports the proposed mechanism of the bacteria inactivation by ozone that caused cell membrane destruction and finally lysis reaction. Thus, the precaution of using ozone as a biocide should be used to address appropriate concentrations of bacterial contamination in water.     

Paramphistomum cervi: Surface topography of the tegument of adult fluke

Adult Paramphistomum cervi or rumen fluke are pear-shaped, slightly concave ventrally and convex dorsally. The worm measures about 5-13 mm in length and 2-5 mm in width across the mid-section. As observed by scanning electron microscopy (SEM), the tegumental surface in all part of the body, appears highly corrugated with transverse folds alternating with grooves and is spineless. At high magnification, the surface of the fold is composed of microfolds or ridges separated by microgrooves or pits. Corrugations and invaginations of the ventral surface are also more extensive than on the dorsal surface of the body. Both anterior and posterior suckers have thick rims covered with transverse folds without spine. The genital pore is situated at the anterior third of the body. There are two types of sensory papillae on the surface: type 1 is bulbous in shape, measuring 10-15 μm in diameter at the base with nipple-like tips, and type 2 has a similar shape and size and also a short cilia on top. These sensory papillae usually occur in large clusters, each having between 5 and 20 units depending on the region of the body. Clusters of papillae on the ventral surface and around the anterior suckers tend to be more numerous and larger in size. The dorsal surface of the body has the least number of papillae.