Alpha-actinin-4 is selectively required for insulin-induced GLUT4 translocation

Insulin induces GLUT4 translocation to the muscle cell surface. Using differential amino acid labeling and mass spectrometry, we observed insulin-dependent co-precipitation of actinin-4 (ACTN4) with GLUT4 (Foster, L. J., Rudich, A., Talior, I., Patel, N., Huang, X., Furtado, L. M., Bilan, P. J., Mann, M., and Klip, A. (2006) J. Proteome Res. 5, 64-75). ACTN4 links F-actin to membrane proteins, and actin dynamics are essential for GLUT4 translocation. We hypothesized that ACTN4 may contribute to insulin-regulated GLUT4 traffic. In L6 muscle cells insulin, but not platelet-derived growth factor, increased co-precipitation of ACTN4 with GLUT4. Small interfering RNA-mediated ACTN4 knockdown abolished the gain in surface-exposed GLUT4 elicited by insulin but not by platelet-derived growth factor, membrane depolarization, or mitochondrial uncoupling. In contrast, knockdown of alpha-actinin-1 (ACTN1) did not prevent GLUT4 translocation by insulin. GLUT4 colocalized with ACTN4 along the insulin-induced cortical actin mesh and ACTN4 knockdown prevented GLUT4-actin colocalization without impeding actin remodeling or Akt phosphorylation, maintaining GLUT4 in a tight perinuclear location. We propose that ACTN4 contributes to GLUT4 traffic, likely by tethering GLUT4 vesicles to the cortical actin cytoskeleton.     

Muscle-specific Pten deletion protects against insulin resistance and diabetes

Pten (phosphatase with tensin homology), a dual-specificity phosphatase, is a negative regulator of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway. Pten regulates a vast array of biological functions including growth, metabolism, and longevity. Although the PI3K/Akt pathway is a key determinant of the insulin-dependent increase in glucose uptake into muscle and adipose cells, the contribution of this pathway in muscle to whole-body glucose homeostasis is unclear. Here we show that muscle-specific deletion of Pten protected mice from insulin resistance and diabetes caused by high-fat feeding. Deletion of muscle Pten resulted in enhanced insulin-stimulated 2-deoxyglucose uptake and Akt phosphorylation in soleus but, surprisingly, not in extensor digitorum longus muscle compared to littermate controls upon high-fat feeding, and these mice were spared from developing hyperinsulinemia and islet hyperplasia. Muscle Pten may be a potential target for treatment or prevention of insulin resistance and diabetes.     

Spectroscopic identification of new ellagitannins and a trigalloyl-glucosylkaempferol from an extract of Euphorbia cotinifolia L. with antitumour and antioxidant activity

From an extract of leaves and small branches of Euphorbia cotinifolia L., 17 polyphenols were isolated including two new ellagitannins and a trigalloyl-glucosylkaempferol. Based on extensive spectral data (UV, ESI-MS, 1H NMR, DEPT and 1D/2D NMR) and chemical studies, their structures were characterized as 1-O-galloyl-3,6-hexahydroxydiphenoyl-D-B1,4-glucopyranose (5), 1-O-galloyl-3,6-valoneoyl-D-B1,4-glucopyranose (6), and kaempferol 3-O-(2",3",6"-tri-O-galloyl)-beta-D-glucopyranoside (13). Biological evaluation indicated that the 80% aqueous methanol extract (AME), chloroform extract (CE), and some pure compounds have potent scavenging activity in the DPPH assay with SC50 values lower than that of ascorbic acid, especially 5, 7-9, and a mixture of hyperin 6"-gallate (11) and isoquercitrin 6"-gallate (12). Moreover, using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) cell viability assay, 6 and 8 exhibited the highest inhibition of human hepatocellular carcinoma cells (Hep-G2), while AME, CE, 5, 7, 9, and the mixture of 11 and 12 were found to be moderate growth inhibitors according to their IC50 values. In addition, AME, 5, and 8 exhibited significant antiproliferative activity against colon carcinoma cells (HCT-116); however, CE and the other examined compounds displayed moderate to low antitumour activity against HCT-116 cells.     

Assessment of the ultrastructural and proliferative properties of human embryonic stem cell-derived cardiomyocytes

Assessment of early ultrastructural development and cell-cycle regulation in human cardiac tissue is significantly hampered by the lack of a suitable in vitro model. Here we describe the possible utilization of human embryonic stem cell (ES) lines for investigation of these processes. With the use of the embryoid body (EB) differentiation system, human ES cell-derived cardiomyocytes at different developmental stages were isolated and their histomorphometric, ultrastructural, and proliferative properties were characterized. Histomorphometric analysis revealed an increase in cell length, area, and length-to-width ratio in late-stage EBs (>35 days) compared with early (10-21 days) and intermediate (21-35 days) stages. This was coupled with a progressive ultrastructural development from an irregular myofibrillar distribution to an organized sarcomeric pattern. Cardiomyocyte proliferation, assessed by double labeling with cardiac-specific antibodies and either [3H]thymidine incorporation or Ki-67 immunolabeling, demonstrated a gradual withdrawal from cell cycle. Hence, the percentage of positively stained nuclei in early-stage cardiomyocytes ([3H]thymidine: 60 +/- 10%, Ki-67: 54 +/- 23%) decreased to 36 +/- 7% and 9 +/- 16% in intermediate-stage EBs and to <1% in late-stage cardiomyocytes. In conclusion, a reproducible temporal pattern of early cardiomyocyte proliferation, cell-cycle withdrawal, and ultrastructural maturation was noted in this model. Establishment of this unique in vitro surrogate system may allow to examine the molecular mechanisms underlying these processes and to assess interventions aiming to modify these properties. Moreover, the detailed characterization of the ES cell-derived cardiomyocyte may be crucial for the development of future cell replacement strategies aiming to regenerate functional myocardium.     

Impact of salinity stress on soil-borne fungi of sugarbeet

The sugarbeet cultivar Kaumera was found to be highly susceptible to infection by the root-rot pathogens Rhizoctonia solani and Sclerotium rolfsii in the absence of salinity stress. Under this environmental condition, R. solani was more efficient than S. rolfsii in producing cell wall-degrading enzymes in infected hypocotyls. Xylanase and galactanase were most effective. The rate of cell wall degradation by R. solani was nearly 2.5 times that of S. rolfsii when cells walls of healthy hypocotyls were used as sole carbon substrate for the in vitro produced crude enzymes.Under salinity stress the pathogenicity and the performance of cell wall-degrading enzymes of R. solani and S. rolfsii varied profoundly. Pathogenicity studies showed that R. solani appeared to be more tolerant than S. rolfsii of the salinity stresses applied, and relatively more virulent to cv Kaumera. The activities of cell wall enzymes of R. solani decreased and those of S. rolfsii increased with increased salt concentration when cell wall material was used as a sole carbon source. The metabolic products produced under salinity stress by R. solani and R. solani in the cell wall amended culture media shifted the initial pH towards neutrality or slight alkalinity for R. solani and to high acidity for S. rolfsii.When model substrates were used, xyland and galactan were the most responsive substrates for degradation by the cell wall enzymes of the two fungi studied. The rate of degradation was higher for S. rolfsii than for R. solani. The excessive acidity in salt stressed S. rolfsii culture media suggested reduced activities of the enzymes involved in cell wall degradation in vivo. This may explain the decreased virulence potentialities.     

MeSHer: identifying biological concepts in microarray assays based on PubMed references and MeSH terms

SUMMARY: MeSHer uses a simple statistical approach to identify biological concepts in the form of Medical Subject Headings (MeSH terms) obtained from the PubMed database that are significantly overrepresented within the identified gene set relative to those associated with the overall collection of genes on the underlying DNA microarray platform. As a demonstration, we apply this approach to gene lists acquired from a published study of the effects of angiotensin II (Ang II) treatment on cardiac gene expression and demonstrate that this approach can aid in the interpretation of the resulting 'significant' gene set. AVAILABILITY: The software is available at http://www.tm4.org. SUPPLEMENTARY INFORMATION: Results from the analysis of significant genes from the published Ang II study.     

Properties of monovalent nickel complexes with tetraaza-macrocyclic ligands in aqueous solutions

The effects of N-alkylation on the redox potential of the couples NiLⁱ²⁺/NiLⁱ⁺, L = tetraaza-14-membered-macrocyclic ligands, and on the properties of the monovalent nickel complexes in aqueous solutions are reported for 14 complexes. The spectra and lifetimes of the NiLⁱ⁺ complexes are reported. The self-exchange rates for the couples NiLⁱ²⁺/NiLⁱ⁺ were determined. Two of the ligands were synthesized for the first time for this study. Cyclic voltammetry and pulse radiolysis were used. The results point out that: (i) N-alkylation always shifts the redox potential to a less cathodic one; this effect stems to a large degree from the decrease in the solvation energy of the complex caused by the N-alkylation of the ligand. (ii). The lifetime of the monovalent complexes is not linearly related to the redox potential of the NiLⁱ²⁺/NiLⁱ⁺ couples. (iii) The NiLⁱ⁺ complexes exist in several isomeric forms; the rate of the isomerization depends on the structure of the ligand. (iv) Different isomers of NiLⁱ⁺ may be formed when the complex NiLⁱ²⁺ is reduced by different reagents; therefore, the pulse-radiolytically formed NiLⁱ⁺ complexes might have different properties than those formed electrochemically.     

Epidemiological Characteristics of Strongyloidiasis in Inhabitants of Indigenous Communities in Borneo Island,Malaysia

Epidemiological study on strongyloidiasis in humans is currently lacking in Malaysia. Thus, a cross-sectional study was carried out to determine the prevalence of Strongyloides stercoralis infection among the inhabitants of longhouse indigenous communities in Sarawak. A single stool and blood sample were collected from each participant and subjected to microscopy, serological and molecular techniques. Five species of intestinal parasites were identified by stool microscopy. None of the stool samples were positive for S. stercoralis. However, 11% of 236 serum samples were seropositive for strongyloidiasis. Further confirmation using molecular technique on stool samples of the seropositive individuals successfully amplified 5 samples, suggesting current active infections. The prevalence was significantly higher in adult males and tended to increase with age. S. stercoralis should no longer be neglected in any intestinal parasitic survey. Combination of more than 1 diagnostic technique is necessary to increase the likelihood of estimating the ‘true’ prevalence of S. stercoralis.