The effects of kefir,koumiss, yogurt and commercial probiotic formulations on PPARα and PPAR-β/δ expressions in mouse kidney

Commercial probiotic capsules that contain probiotic bacteria, kefir, koumiss and yogurt contain beneficial microorganisms that affect cholesterol levels and immune response, and are used for treatment of some diseases. We investigated using immunohistochemistry the effects of kefir, koumiss, yogurt and a commercial probiotic formulation on the expression levels of peroxisome proliferator-activated receptor-α (PPARα) and peroxisome proliferator-activated receptor-β/δ (PPAR-β/δ), which are members of the nuclear steroid hormone receptor superfamily in mouse kidney. Mice were assigned to five groups: group 1, commercial probiotic capsules; group 2, kefir; group 3, koumiss; group 4, yogurt; group 5, control. After oral administration for 15 days, body weights were recorded and kidney tissue samples were obtained. Hematoxylin & eosin staining and the streptavidin-biotin peroxidase complex (ABC) method were applied to tissue sections to examine histology and to determine the localization of PPARα and PPAR-β/δ in the kidneys. We found that the weights of the mice in the kefir, koumiss, yogurt and commercial probiotic capsules groups were increased compared to controls. No differences in kidney histology were observed in any of the experimental groups. Kefir, koumiss, yogurt and the commercial probiotic preparation increased PPARα and PPAR-β/δ expressions.     

DHA-fluorescent probe is sensitive to membrane order and reveals molecular adaptation of DHA in ordered lipid microdomains

Docosahexaenoic acid (DHA) disrupts the size and order of plasma membrane lipid microdomains in vitro and in vivo. However, it is unknown how the highly disordered structure of DHA mechanistically adapts to increase the order of tightly packed lipid microdomains. Therefore, we studied a novel DHA-Bodipy fluorescent probe to address this issue. We first determined if the DHA-Bodipy probe localized to the plasma membrane of primary B and immortal EL4 cells. Image analysis revealed that DHA-Bodipy localized into the plasma membrane of primary B cells more efficiently than EL4 cells. We then determined if the probe detected changes in plasma membrane order. Quantitative analysis of time-lapse movies established that DHA-Bodipy was sensitive to membrane molecular order. This allowed us to investigate how DHA-Bodipy physically adapted to ordered lipid microdomains. To accomplish this, we employed steady-state and time-resolved fluorescence anisotropy measurements in lipid vesicles of varying composition. Similar to cell culture studies, the probe was highly sensitive to membrane order in lipid vesicles. Moreover, these experiments revealed, relative to controls, that upon incorporation into highly ordered microdomains, DHA-Bodipy underwent an increase in its fluorescence lifetime and molecular order. In addition, the probe displayed a significant reduction in its rotational diffusion compared to controls. Altogether, DHA-Bodipy was highly sensitive to membrane order and revealed for the first time that DHA, despite its flexibility, could become ordered with less rotational motion inside ordered lipid microdomains. Mechanistically, this explains how DHA acyl chains can increase order upon formation of lipid microdomains in vivo.     

Single-cell paired-end genome sequencing reveals structural variation per cell cycle

The nature and pace of genome mutation is largely unknown. Because standard methods sequence DNA from populations of cells, the genetic composition of individual cells is lost, de novo mutations in cells are concealed within the bulk signal and per cell cycle mutation rates and mechanisms remain elusive. Although single-cell genome analyses could resolve these problems, such analyses are error-prone because of whole-genome amplification (WGA) artefacts and are limited in the types of DNA mutation that can be discerned. We developed methods for paired-end sequence analysis of single-cell WGA products that enable (i) detecting multiple classes of DNA mutation, (ii) distinguishing DNA copy number changes from allelic WGA-amplification artefacts by the discovery of matching aberrantly mapping read pairs among the surfeit of paired-end WGA and mapping artefacts and (iii) delineating the break points and architecture of structural variants. By applying the methods, we capture DNA copy number changes acquired over one cell cycle in breast cancer cells and in blastomeres derived from a human zygote after in vitro fertilization. Furthermore, we were able to discover and fine-map a heritable inter-chromosomal rearrangement t(1;16)(p36;p12) by sequencing a single blastomere. The methods will expedite applications in basic genome research and provide a stepping stone to novel approaches for clinical genetic diagnosis.     

Structure-driven HtL: design and synthesis of novel aminoindazole inhibitors of c-Jun N-terminal kinase activity

The design and synthesis of a new series of c-Jun N-terminal kinase inhibitors are reported. The novel series of substituted amino indazoles were designed based on a combination of hits from high-throughput screening and X-ray crystal structure information of the compounds crystallised into the JNK-1 ATP binding site.     

Postoperative death in sows as a result of gastric mucosal de-gloving at the pars oesophagea

It is well documented that pigs frequently die from postoperative acute gastric dilatation, and proximal gastric 'stress' ulceration. Three cases of gastric mucosal 'de-gloving' are reported. This was secondary to acute gastric dilatation and resulted in death from acute haemorrhage. All animals had undergone major abdominal surgery. Histology confirmed that the proximal gastric mucosa had been 'de-gloved', or torn from the gastro-oesophageal junction, leaving exposed muscle fibres. This syndrome has not been reported previously. The postmortem appearances of this mechanical injury could easily be mistaken for extensive oesophago-gastric peptic ulceration. This has major implications for prevention.     

Effect of additives on the thermostability ofBacillus stearothermophilus α-amylase

The effect of additives on the thermostability ofBacillus stearothermophilus -amylase was determined. Polyols, dimethyl formamide, and dimethyl sulfoxide all increased the half life of the enzyme approximately 2-fold when tested at a 10% (w/v) addition. These results suggest that the enzyme's structure is stabilized against thermal denaturation through ionic interactions. Addition of dextran or polyvinyl alcohol (hydrophilic polymers which increase the viscosity of the solution) had a slight positive effect on enzyme stability while addition of polyethylene glycol or polyvinylpyrrolidone (hydrophobic polymers which increase the viscosity of the solution) resulted in a 2-fold decrease in enzyme half life.     

A study of genetic linkage heterogeneity in 35 adult-onset polycystic kidney disease families

A genetic heterogeneity analysis of 35 kindreds with adult-onset polycystic kidney disease (ADPKD) was carried out using the D16S85, D16S84, D16S125 and D16S94 loci that are closely linked to the PKD1 locus on chromosome 16. The results show that the likelihood of two ADPKD loci is 2,514.9 times greater than for a single locus (P < 0.0001). The maximum likelihood lod score is 27.38 under heterogeneity with PKD1 lying 4.9 cM proximal to D16S85 (in males). At least 3% of kindreds are unlinked to PKD1, since the 95% confidence limits of alpha, the proportion of families linked to PKD1, are 0.54–0.97. Only 2 out of 35 kindreds (5.7%) show statistically significant evidence of non-linkage to PKD1, with conditional probabilities of 0.987 and 0.993 that the disease locus is unlinked. This confirms the existence of a small subgroup of ADPKD kindreds that are unlinked to PKD1 and provides a firm basis for genetic counselling of this population on the basis of DNA probes.     

Denaturation ofBacillus stearothermophilus α-amylase by urea and detergents

Denaturation ofBacillus stearothermophilus -amylase by urea and detergents was investigated for the purpose of understanding the mechanism of denaturation of this enzyme. The enzyme was extremely resistant to denaturation by detergents at 60° C, either in the presence or absence of added calcium. Addition of EDTA was necessary to obtain denaturation by detergents. The rate of denaturation of the -amylase by urea was strongly dependent on the incubation pH and presence or absence of calcium ions. Calcium-binding groups were shown to have pKa values of 5.5 for exogenous calcium and 4.7 to 4.8 for endogenous calcium. A mechanism is proposed for the denaturation ofBacillus stearothermophilus -amylase.