Human glycated albumin affects glucose metabolism in L6 skeletal muscle cells by impairing insulin-induced insulin receptor substrate (IRS) signaling through a protein kinase C alpha-mediated mechanism

Nonenzymatic glycation is increased in diabetes and leads to increased levels of glycated proteins. Most studies have focused on the role of glycation products in vascular complications. Here, we have investigated the action of human glycated albumin (HGA) on insulin signaling in L6 skeletal muscle cells. Exposure of these cells to HGA inhibited insulin-stimulated glucose uptake and glycogen synthase activity by 95 and 80%, respectively. These effects were time- and dose-dependent, reaching a maximum after 12 h incubation with 0.1 mg/ml HGA. In contrast, exposure of the cells to HGA had no effect on thymidine incorporation. Further, HGA reduced insulin-stimulated serine phosphorylation of PKB and GSK3, but did not alter ERK1/2 activation. HGA did not affect either insulin receptor kinase activity or insulin-induced Shc phosphorylation on tyrosine. In contrast, insulin-dependent IRS-1 and IRS-2 tyrosine phosphorylation was severely reduced in cells preincubated with HGA for 24 h. Insulin-stimulated association of PI3K with IRS-1 and IRS-2, and PI3K activity were reduced by HGA in parallel with the changes in IRS tyrosine phosphorylation, while Grb2-IRS association was unchanged. In L6 myotubes, exposure to HGA increased PKC activity by 2-fold resulting in a similar increase in Ser/Thr phosphorylation of IRS-1 and IRS-2. These phosphorylations were blocked by the PKC inhibitor bisindolylmaleimide (BDM). BDM also blocked the action of HGA on insulin-stimulated PKB and GSK3 alpha. Simultaneously, BDM rescued insulin-stimulation of glucose uptake and glycogen synthase activity in cells exposed to HGA. The use of antibodies specific to PKC isoforms shows that this effect appears to be mediated by activated PKC alpha, independent of reactive oxygen species production. In summary, in L6 skeletal muscle cells, exposure to HGA leads to insulin resistance selectively in glucose metabolism with no effect on growth-related pathways regulated by the hormone

Synergistic effect of Aspergillus niger and Trichoderma reesei enzyme sets on the saccharification of wheat straw and sugarcane bagasse

Plant-degrading enzymes can be produced by fungi on abundantly available low-cost plant biomass. However, enzymes sets after growth on complex substrates need to be better understood, especially with emphasis on differences between fungal species and the influence of inhibitory compounds in plant substrates, such as monosaccharides. In this study, Aspergillus niger and Trichoderma reesei were evaluated for the production of enzyme sets after growth on two "second generation" substrates: wheat straw (WS) and sugarcane bagasse (SCB). A. niger and T. reesei produced different sets of (hemi-)cellulolytic enzymes after growth on WS and SCB. This was reflected in an overall strong synergistic effect in releasing sugars during saccharification using A. niger and T. reesei enzyme sets. T. reesei produced less hydrolytic enzymes after growth on non-washed SCB. The sensitivity to non-washed plant substrates was not reduced by using CreA/Cre1 mutants of T. reesei and A. niger with a defective carbon catabolite repression. The importance of removing monosaccharides for producing enzymes was further underlined by the decrease in hydrolytic activities with increased glucose concentrations in WS media. This study showed the importance of removing monosaccharides from the enzyme production media and combining T. reesei and A. niger enzyme sets to improve plant biomass saccharification

Disruption of bovine sperm functions in the presence of aplastic midpiece defect

Bulls are of great importance in the productive chain and for this reason they should have a good semen quality. There is no doubt that sperm morphology is very important to bull fertility, although little is known about how exactly the abnormal morphologies may affect sperm functions.To detail the morphological description of the aplastic midpiece defect (AMD), as well as to understand its consequences for male fertility based on membrane and acrosome status, mitochondrial membrane potential and DNA integrity parameters.The bulls were divided into two groups: control, consisting of satisfactory potential breeders (n = 3); and AMD, consisting of unsatisfactory potential breeders with a high percentage of AMD (n = 3). Bulls were evaluated by the breeding soundness evaluation; five ejaculates were collected from each animal and analyzed by flow cytometry. Spermatozoa from AMD group exhibited lower sperm motility and vigor (p < 0.05). In addition, it also exhibited lower mitochondrial membrane potential (p < 0.05), a higher percentage of spermatozoa with DNA fragmentation (p < 0.05), lower acrosome and plasma membrane integrity (p < 0.05), and higher lipid bilayer sperm membrane disorganization (p < 0.05) in comparison with control bulls. These findings may be due to oxidative stress and a reduction of the energy production capacity in addition to an alteration in the structural composition of the sperm cell. Moreover, semen with a high percentage of AMD may also be undergoing apoptosis.Bulls with a high percentage of AMD in their semen are not suitable for reproduction. Furthermore, it suggests there is a putative genetic basis for this sperm defect

Disruption of bovine sperm functions in the presence of aplastic midpiece defect

Bulls are of great importance in the productive chain and for this reason they should have a good semen quality. There is no doubt that sperm morphology is very important to bull fertility, although little is known about how exactly the abnormal morphologies may affect sperm functions.To detail the morphological description of the aplastic midpiece defect (AMD), as well as to understand its consequences for male fertility based on membrane and acrosome status, mitochondrial membrane potential and DNA integrity parameters.The bulls were divided into two groups: control, consisting of satisfactory potential breeders (n = 3); and AMD, consisting of unsatisfactory potential breeders with a high percentage of AMD (n = 3). Bulls were evaluated by the breeding soundness evaluation; five ejaculates were collected from each animal and analyzed by flow cytometry. Spermatozoa from AMD group exhibited lower sperm motility and vigor (p < 0.05). In addition, it also exhibited lower mitochondrial membrane potential (p < 0.05), a higher percentage of spermatozoa with DNA fragmentation (p < 0.05), lower acrosome and plasma membrane integrity (p < 0.05), and higher lipid bilayer sperm membrane disorganization (p < 0.05) in comparison with control bulls. These findings may be due to oxidative stress and a reduction of the energy production capacity in addition to an alteration in the structural composition of the sperm cell. Moreover, semen with a high percentage of AMD may also be undergoing apoptosis.Bulls with a high percentage of AMD in their semen are not suitable for reproduction. Furthermore, it suggests there is a putative genetic basis for this sperm defect