Human factor VIII has been isolated from a high purity factor VIII concentrate by immunoaffmity chromatography and HPLC on Mono Q gel. Two fractions of factor VIII were obtained with a specific activity of w7000 units/mg.The major fraction contained eight peptide chains of 200, 180, 160, 150, 135, 130, 115, and 105 kDa plus one doublet chain of 80 kDa. The minor fraction contained one peptide chain of 90 kDa plus the chain of 80 kDa. Both fractions were activated by thrombin to the same extent. Amino-terminal amino acid sequence analysis was performed on the 180-kDa, 130-kDa, and 90-kDa chains and showed an identical amino-terminal sequence in these chains. Each chain from 200 kDa to 90 kDa was linked to one 80-kDa chain by a metal-ion bridge(s).Studies on factor VIII in plasma and cryoprecipitate, prepared and gel ifitered in the presence of protease inhibitors, showed that one 200-kDa plus one 80-kDa chain were the only or dominating chains in the materials and may represent native factor VIII. The results indicated that all chains from 180 kDa to 90 kDa are fragments of the 200-kDa chain. All of these more or less fragmented chains form active factor VIII complexes with the 80-kDa chain.Factor VIII (antihemophilic factor) is the protein that is deficient or absent in individuals with classic hemophilia, an X-chromosome-linked bleeding disorder. It participates in the intrinsic pathway of blood coagulation as a cofactor in the activation of factor X by factor IXa, in the presence of phospholipid and calcium (1-7). In plasma, factor VIII is noncovalently bound to von Willebrand factor (vWF), a high molecular weight protein involved in primary hemostasis. Due to the low concentration of factor VIII in plasma and to the fact that factor VIII is highly susceptible to degradation by serine proteases, it is only recently that highly purified factor VIII, free of vWF, has been obtained. Bovine factor VIII, as isolated from plasma, is composed of three peptide chains of 93 kDa, 88 kDa, and 85 kDa (8). Factor VIII purified from porcine plasma has been shown to consist of subunits of 160 kDa, 130 kDa, 82 kDa, and 76 kDa (9). Purified human factor VIII has been prepared from commercial factor VIII concentrate and had a specific activity of 2294 units/mg (10). NaDodSO4/PAGE showed several peptide chains having molecular sizes of 188-79 kDa. The majority of the material was related to factor VIII as shown by immunoblot technique with use of monoclonal antibodies against factor VIII (11). A preparation of human factor VIII from cryoprecipitate, in the presence of seine protease inhibitors, has also been reported with the specific activity of 4740 units/mg (12). Unreduced NaDodSO4/PAGE showed predominant bands at 360 kDa, 210 kDa, and 90 kDa and an 80 kDa/79 kDa doublet band.Immunoblotting using monoclonal antibodies suggested that the 360 kDa component was a precursor to the other components. Thus different results are obtained when different starting materials and procedures for purification are used.The human fac...

Skeletal muscle development is a complex and highly orchestrated biological process mediated by a series of myogenesis regulatory factors. Numerous studies have demonstrated that circular RNAs (circRNAs) are involved in muscle differentiation, but the exact molecular mechanisms involved remain unclear. Here, we analyzed the expression of circRNAs at the adult and embryo development stages of cattle musculus longissimus . A stringent set of 1,318 circRNAs candidates were identified, and we found that 495 circRNAs were differentially expressed between embryonic and adult tissue libraries. We subsequently focused on one of the most downregulated circRNAs (using the adult stage expression as control), and this was named muscle differentiation-associated circular RNA (circMYBPC1). With RNA binding protein immunoprecipitation (RIP) and RNA pull-down assays, circMYBPC1 was identified to promote myoblast differentiation by directly binding miR-23a to relieve its inhibition on myosin heavy chain ( MyHC ). In addition, RIP assays demonstrated that circMYBPC1 could directly bind MyHC protein. In vivo observations also suggested that circMYBPC1 may stimulate skeletal muscle regeneration after muscle damage. These results revealed that the novel non-coding circRNA circMYBPC1 promotes differentiation of myoblasts and may promote skeletal muscle regeneration. Our results provided a basis for in-depth analysis of the role of circRNA in myogenesis and muscle diseases.

BackgroundRotavirus is the leading cause of severe dehydrating diarrhea in young children and the inner capsid protein VP6 is a potential vaccine candidate that can induce cross-protective immune responses against different Rotavirus strains. The use of ferritin nanoparticles as the scaffold of the antigen can improve the immunogenicity of the subunit vaccines and provide broader protection. We here present a non-live and self-assemble recombinant rotavirus VP6–ferritin (rVP6–ferritin) nanoparticle vaccine.ResultsThe rVP6–ferritin nanoparticles were expressed in E. coli and self-assembled to uniform spherical structure which similar to ferritin, and oral administration of them induced efficient humoral and mucosal immunogenicity in mice. The nanoparticles were further transgenically expressed in the milk of mice, and pup mice breastfed by transgenic rVP6–ferritin mothers had strongly induced immunogenicity and—compared to pups breastfed by wild type mothers—the proportion of rotavirus challenged pups with diarrhea symptoms, the duration and intensity of the diarrhea, and the deleterious effects on overall growth resulting from the diarrhea were all significantly reduced.ConclusionsThese results suggest that this recombinant VP6–ferritin nanoparticle vaccine can efficiently prevent the death and malnutrition induced by the rotavirus infection in infants and is a promising candidate vaccine for rotavirus.Electronic supplementary materialThe online version of this article (10.1186/s12951-019-0446-6) contains supplementary material, which is available to authorized users.

Beef is considered to be a good quality meat product because it contains linoleic acid and specific proteins, which can bring significant benefits to health. Circular RNAs (circRNAs) have been reported to regulate skeletal myogenesis. RNA-seq was used to investigate the circRNA molecular regulatory mechanisms with respect to differences in muscle quality between buffalo and cattle. A total of 10,449 circRNA candidates were detected, and 1128 of these were found to be differentially expressed between cattle and buffalo muscle tissue libraries. Differentially expressed 23 circRNAs were verified by qPCR. CircEch1, one of the most up-regulated circRNAs during muscle development, was subsequently characterized. CCK-8 (65.05 ± 2.33%, P < 0.0001), EdU (72.99 ± 0.04%, P < 0.001), and Western blotting assays showed that overexpression of circEch1 inhibited the proliferation of bovine myoblasts but promoted differentiation. In vivo studies suggested that circEch1 stimulates skeletal muscle regeneration. These results demonstrate that the novel regulator circEch1 induces myoblast differentiation and skeletal muscle regeneration. They also provide new insights into the mechanisms of circRNA regulation of beef quality.

Buffalo meat consist good qualitative characteristics as it contains "thined tender" which is favorable for cardavascular system. However, the regulatory mechanisms of long non-coding RNA (lncRNA), differences in meat quality are not well known. The chemical-physical parameters revealed the muscle quality of buffalo that can be equivalent of cattle, but there are significant differences in shearing force and muscle fiber structure. Then, we examined lncRNA expression profiles of buffalo and cattle skeletal muscle that provide first insights into their potential roles in buffalo myogenesis. Here, we profiled the expression of lncRNA in cattle and buffalo skeletal muscle tissues, and 16,236 lncRNA candidates were detected with 865 up-regulated lncRNAs and 1,296 down-regulated lncRNAs when comparing buffalo to cattle muscle tissue. We constructed coexpression and ceRNA networks, and found lncRNA MSTRG.48330.7, MSTRG.30030.4, and MSTRG.203788.46 could be as competitive endogenous RNA (ceRNA) containing potential binding sites for miR-1/206 and miR-133a. Tissue expression analysis showed that MSTRG.48330.7, MSTRG.30030.4, and MSTRG.203788.46 were highly and specifically expressed in muscle tissue. Present study may be used as a reference tool for starting point investigations into the roles played by several of those lncRNAs during buffalo myogenesis.