De novo transcriptome assemblies of red king crab (Paralithodes camtschaticus) and snow crab (Chionoecetes opilio) molting gland and eyestalk ganglia - Temperature effects on expression of molting and growth regulatory genes in adult red king crab.

Red king crab (Paralithodes camtschaticus) and snow crab (Chionoecetes opilio) are deep-sea crustaceans widely distributed in the North Pacific and Northwest Atlantic Oceans. These giant predators have invaded the Barents Sea over the past decades, and climate-driven temperature changes may influence their distribution and abundance in the sub-Arctic region. Molting and growth in crustaceans are strongly affected by temperature, but the underlying molecular mechanisms are little known, particularly in cold-water species. Here, we describe multiple regulatory factors in the two high-latitude crabs by developing de novo transcriptomes from the molting gland (Y-organ or YO) and eye stalk ganglia (ESG), in addition to the hepatopancreas and claw muscle of red king crab. The Halloween genes encoding the ecdysteroidogenic enzymes were expressed in YO, and the ESG contained multiple neuropeptides, including molt-inhibiting hormone (MIH), crustacean hyperglycemic hormone (CHH), and ion-transport peptide (ITP). Both crabs expressed a diversity of growth-related factors, such as mTOR, AKT, Rheb and AMPKα, and stress-responsive factors, including multiple heat shock proteins (HSPs). Temperature effects on the expression of key regulatory genes were quantified by qPCR in adult red king crab males kept at 4 °C or 10 °C for two weeks during intermolt. The Halloween genes tended to be upregulated in YO at high temperature, while the ecdysteroid receptor and several growth regulators showed tissue-specific responses to elevated temperature. Constitutive and heat-inducible HSPs were expressed in an inverse temperature-dependent manner, suggesting that adult red king crabs can acclimate to increased water temperatures.

Peptides from chicken processing by-product inhibit DPP-IV and promote cellular glucose uptake: potential ingredients for T2D management.

Inhibition of dipeptidyl peptidase IV (DPP-IV) and stimulation of muscle glucose uptake are two of the key strategies for management of type-2-diabetes (T2D). In the present study, four protein hydrolysates generated by enzymatic hydrolysis of chicken by-product, i.e., mechanical chicken deboning residue, were evaluated for their DPP-IV inhibitory activity as well as their effect on glucose uptake by skeletal muscle cells. The DPP-IV inhibitory assay was performed at two concentrations (1000 µg mL-1 and 10 µg mL-1) for the crude chicken protein hydrolysates. The hydrolysate with the highest DPP-IV inhibition was selected for preparative-scale fractionation using size-exclusion chromatography (SEC). The SEC fractions were tested for DPP-IV inhibitory activity as well as their effect on glucose uptake and metabolic activity of skeletal muscle cells. The muscle cells were treated with the SEC fractions and glucose uptake was measured based on luminescence detection of 2-deoxyglucose-6-phosphate (2DG6P). A fraction with peptides in the lower molecular weight range was shown to promote glucose uptake and to inhibit DPP-IV. Further chromatographic fractionation followed by inhibition assaying of the most potent SEC fraction led to isolation of five refined peptide fractions with more than 80% DPP-IV inhibition, which were subsequently analyzed with LC-HRMS/MS. This led to identification of 14 peptides as potential DPP-IV inhibitors from protein hydrolysates of mechanical chicken deboning residue.

Vitamin K2 improves proliferation and migration of bovine skeletal muscle cells in vitro.

Skeletal muscle function is highly dependent on the ability to regenerate, however, during ageing or disease, the proliferative capacity is reduced, leading to loss of muscle function. We have previously demonstrated the presence of vitamin K2 in bovine skeletal muscles, but whether vitamin K has a role in muscle regulation and function is unknown. In this study, we used primary bovine skeletal muscle cells, cultured in monolayers in vitro, to assess a potential effect of vitamin K2 (MK-4) during myogenesis of muscle cells. Cell viability experiments demonstrate that the amount of ATP produced by the cells was unchanged when MK-4 was added, indicating viable cells. Cytotoxicity analysis show that MK-4 reduced the lactate dehydrogenase (LDH) released into the media, suggesting that MK-4 was beneficial to the muscle cells. Cell migration, proliferation and differentiation was characterised after MK-4 incubation using wound scratch analysis, immunocytochemistry and real-time PCR analysis. Adding MK-4 to the cells led to an increased muscle proliferation, increased gene expression of the myogenic transcription factor myod as well as increased cell migration. In addition, we observed a reduction in the fusion index and relative gene expression of muscle differentiation markers, with fewer complex myotubes formed in MK-4 stimulated cells compared to control cells, indicating that the MK-4 plays a significant role during the early phases of muscle proliferation. Likewise, we see the same pattern for the relative gene expression of collagen 1A, showing increased gene expression in proliferating cells, and reduced expression in differentiating cells. Our results also suggest that MK-4 incubation affect low density lipoprotein receptor-related protein 1 (LRP1) and the low-density lipoprotein receptor (LDLR) with a peak in gene expression after 45 min of MK-4 incubation. Altogether, our experiments show that MK-4 has a positive effect on muscle cell migration and proliferation, which are two important steps during early myogenesis.