Cisplatin resistance induced by decreased apoptotic activity in non-small-cell lung cancer cell lines

We have investigated defective steps in apoptosis that might account for the development of resistance. For this purpose, A549 and Calu1 NSCLC (non-small-cell lung cancer) cell lines were treated with cisplatin to obtain resistant sub-lines. Gene expression profiles and the phosphorylation status of the BAD (Bcl-2/Bcl-XL-antagonist, causing cell death) protein were determined for each cell line. Cell death and cytochrome c release were analysed after treating cell lines with their appropriate cisplatin doses. Gene expression of BAD, Bid, caspases 4 and 6 were clearly decreased in the resistant cell lines, and the differential phosphorylation status of BAD also seemed to play a role in the development of cisplatin resistance. Since this is a new cisplatin-resistant Calu1 cell line, it is noteworthy that DNA fragmentation, apoptotic cell ratio and cytochrome c levels were most decreased in the CR-Calu1 cell line.     

Genetic and pathogenic diversity of Neofusicoccum parvum in New Zealand vineyards

Genetic diversity of 50 isolates of Neofusicoccum parvum, the predominant species of the Botryosphaeriaceae recovered from grapevines displaying symptoms of dieback and decline in New Zealand, was compared to that of isolates from Australia, South Africa, and California. The eight universally primed polymerase chain reaction (UP-PCR) primers distinguished 56 genotypes, with only four clonal pairs found. Seven main groups were identified in a neighbour-joining (NJ) tree with isolates from different regions and vineyards of New Zealand, Australia, and California distributed in different groups, indicating a high level of intra and intervineyard genetic variation. All of the South African isolates were positioned in a separate UP-PCR group, indicating that these isolates were less related to the other N. parvum isolates. When compared to fungi that reproduce sexually the genetic diversity and Shannon diversity indices were low (0.076-0.249; 0.109-0.367, respectively), genetic identity levels were high (0.76-0.95), and genetic distance levels were low (0.04-0.27). The large number of genotypes and the low number of clones in the New Zealand N. parvum populations may be explained by parasexual recombination as anastomosis was observed between nonself pairings. Pathogenicity tests using isolates from different UP-PCR groups inoculated onto either green shoots or 1-y-old grapevines detected virulence diversity, indicating intra and intervineyard variation between isolates, however, no correlation was detected between UP-PCR group and virulence.     

Glucose-dependent insulinotropic polypeptide (GIP) stimulates transepithelial glucose transport

The purpose of this study was to characterize the effects of glucose-dependent insulinotropic peptide (GIP) on small intestinal glucose transport in vitro. Stripped proximal jejunum from fasted mice was mounted in Ussing chambers. The serosal side was bathed in Regular Ringer solution containing 5 mmol/l glucose, and the mucosal side, with solution containing 10 mmol/l 3-O-methyl glucose (3OMG). Intercellular cyclic adenosine monophosphate (cAMP), mucosa-to-serosa fluxes of 3OMG (J(ms)(3OMG)), and short-circuit current (I(SC)) were measured in the presence and absence of GIP. GIP increased cAMP by 2.5-fold in isolated enterocytes, consistent with a direct effect of GIP on these epithelial cells. GIP also increased I(SC) and J(ms)(3OMG) by 68 and 53%, respectively, indicating that the increase in J(ms)(3OMG) was primarily electrogenic, with a small electroneutral component. The stimulatory effect of GIP on J(ms)(3OMG) was concentration dependent. In addition, 1,000 nmol/l and 10 nmol/l GIP increased J(ms)(3OMG) by 70 and 30% over control, respectively, consistent with receptor activation. Phlorizin (20 mumol/l), an inhibitor of Na(+)-glucose cotransporter (SGLT-1), abolished the increase in I(SC) and decreased J(ms)(3OMG) by approximately 65%. These results indicate that stimulation of SGLT-1 activity by GIP partially accounts for the increase in J(ms)(30MG). These studies are the first to demonstrate direct stimulation of intestinal glucose transport by GIP independent of its insulinotropic properties. GIP stimulates cellular accumulation of cAMP and thereby upregulates glucose transport. The GIP-induced increase in glucose transport appears to be mediated, at least in part, by SGLT-1.     

Pollen morphology in theBoragineae (Boraginaceae) in relation to the taxonomy of the tribe

The results of a comparative survey on pollen morphology in theBoragineae by means of light, scanning and transmission electron microscopy are presented and discussed in relation to the taxonomy of the tribe. Multivariate analyses lead to the recognition of 15 pollen morphotypes for which a discriminant key is proposed. The discriminatory characters concern mainly the stereostructural and ultrastructural features of the grains, such as tectum sculpture and aperture morphology, while the number of apertures appears variable within genera and even single species. Seven out of the 12 investigated genera, as currently circumscribed, are matched by palynological data:Anchusella, Borago, Brunnera, Elizaldia, Lithodora, Symphytum andTrachystemon. On account of aperture shape,Lithodora is however closer to theLithospermeae than to theBoragineae. Other genera, and in particularNonea, show a wide variation in tectum ornamentation, shape of grains and number of apertures. Palynological data do not support a broad concept of the genusAnchusa, and point to the autonomy of the satellite generaHormuzakia, Gastrocotyle, Phyllocara, Pentaglottis andCynoglottis.     

The Phylogeography of Y-Chromosome Haplogroup H1a1a-M82 Reveals the Likely Indian Origin of the European Romani Populations

Linguistic and genetic studies on Roma populations inhabited in Europe have unequivocally traced these populations to the Indian subcontinent. However, the exact parental population group and time of the out-of-India dispersal have remained disputed. In the absence of archaeological records and with only scanty historical documentation of the Roma, comparative linguistic studies were the first to identify their Indian origin. Recently, molecular studies on the basis of disease-causing mutations and haploid DNA markers (i.e. mtDNA and Y-chromosome) supported the linguistic view. The presence of Indian-specific Y-chromosome haplogroup H1a1a-M82 and mtDNA haplogroups M5a1, M18 and M35b among Roma has corroborated that their South Asian origins and later admixture with Near Eastern and European populations. However, previous studies have left unanswered questions about the exact parental population groups in South Asia. Here we present a detailed phylogeographical study of Y-chromosomal haplogroup H1a1a-M82 in a data set of more than 10,000 global samples to discern a more precise ancestral source of European Romani populations. The phylogeographical patterns and diversity estimates indicate an early origin of this haplogroup in the Indian subcontinent and its further expansion to other regions. Tellingly, the short tandem repeat (STR) based network of H1a1a-M82 lineages displayed the closest connection of Romani haplotypes with the traditional scheduled caste and scheduled tribe population groups of northwestern India.     

MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus

Background

Dysregulation of microRNA (miRNA) expression in various tissues and body fluids has been demonstrated to be associated with several diseases, including Type 2 Diabetes mellitus (T2D). Here, we compare miRNA expression profiles in different tissues (pancreas, liver, adipose and skeletal muscle) as well as in blood samples from T2D rat model and highlight the potential of circulating miRNAs as biomarkers of T2D. In parallel, we have examined the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients.

Methodology/Principal Findings

Employing miRNA microarray and stem-loop real-time RT-PCR, we identify four novel miRNAs, miR-144, miR-146a, miR-150 and miR-182 in addition to four previously reported diabetes-related miRNAs, miR-192, miR-29a, miR-30d and miR-320a, as potential signature miRNAs that distinguished IFG and T2D. Of these microRNAs, miR-144 that promotes erythropoiesis has been found to be highly up-regulated. Increased circulating level of miR-144 has been found to correlate with down-regulation of its predicted target, insulin receptor substrate 1 (IRS1) at both mRNA and protein levels. We could also experimentally demonstrate that IRS1 is indeed the target of miR-144.

Conclusion

We demonstrate that peripheral blood microRNAs can be developed as unique biomarkers that are reflective and predictive of metabolic health and disorder. We have also identified signature miRNAs which could possibly explain the pathogenesis of T2D and the significance of miR-144 in insulin signaling.     

High level expression of a thermostable Bacillus xylose (glucose) isomerase in Escherichia coli

Summary A thermophilic Bacillus sp. producing xylose (glucose) isomerase has been isolated. Its xy/A gene when cloned in Escherichia coli and expressed gave 37.5 and 12.8 units/ mg protein respectively for xylose and glucose isomerase activities at 85°C. A single heat treatment of the crude extract purified the enzyme further yielding the highest ever recorded activities of 150 and 49.02 units /mg protein.     

In vitro lipofection with novel asymmetric series of 1,2-dialkoylamidopropane-based cytofectins containing single symmetric bis-(2-dimethylaminoethane) polar headgroups

Novel N,N'-diacyl-1,2-diaminopropyl-3-carbamoyl[bis-(2-dimethylaminoethane)] bivalent cationic lipids were synthesized and evaluated for in vitro transfection activity against a murine melanoma cell line. In the absence of the helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine), only the dioleoyl derivative 22 (1,2lb5) elicited transfection activity. The transfection activity of this lipid was reduced when formulated with DOPE. Contrary to that, the dimyristoyl derivative 19 (1,2lb2) mediated no activity when used alone but induced the highest levels of marker gene expression in the presence of DOPE. In an effort to correlate the transfection activity with cationic lipid structures, the physicochemical properties of cationic lipids in isolation and of lipoplexes were studied with surface tensiometry, photon correlation spectroscopy, gel electrophoresis mobility shift assay, and fluorescence techniques. In regard to the lipoplex properties, gel electrophoresis mobility shift assay and EtBr exclusion fluorescence assay revealed that the 1,2lb5 was the only lipid to associate and condense plasmid DNA, respectively. Photon correlation spectroscopy analysis found that 1,2lb5/DNA complexes were of relatively small size compared to all other lipoplexes. With respect to the properties of isolated lipids, Langmuir monolayer studies and fluorescence anisotropy on cationic lipid dispersions verified high two-plane elasticity and increased fluidity of the transfection competent dioleoyl derivative 1,2lb5, respectively. The results indicate that high transfection activity is mediated by cationic lipids characterized by an expanded mean molecular area, high molecular elasticity, and increased fluidity.     

Genetic heterogeneity among Vibrio alginolyticus isolated from shrimp farms by PCR fingerprinting

AIMS: To study the strain variability among Vibrio alginolyticus isolates from different sources by insertion sequence-targeted PCR fingerprinting and whole cell protein profile analysis. METHODS AND RESULTS: Eleven strains of V. alginolyticus were isolated from seven different sources including healthy, infected, farm-reared and wild shrimps. Following biochemical characterization, the isolates were analysed by PCR fingerprinting and whole cell protein analysis by SDS-PAGE. The strains were genetically different irrespective of the sources of isolation. CONCLUSIONS: Strain variation exists in V. alginolyticus isolates obtained even from the same source, and PCR fingerprinting is a simple and efficient method in identifying strain-specific variations among the different isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: Vibrio alginolyticus is implicated in severe vibriosis of marine aquaculture systems although many strains are avirulent and could be used as probiotic strains. As a wide variation exists among this species, differentiating the harmful and beneficial strains would help in finding ways of controlling the infections by eliminating harmful shrimp pathogenic vibrios.