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ABCD4 , a member of the ATP-binding cassette transporter superfamily, is associated with the transport of vitamin B 12 which is crucial for the development of red blood cells (RBCs) and may also be involved in its metabolism. However, the molecular function of ABCD4 during RBC development in zebrafish is mostly unknown. Using a morpholino-based knockdown approach, we found that abcd4 -knockdown resulted in abnormal RBCs of irregular shapes and various sizes. o-Dianisidine staining, as an indicator of hemoglobin in RBCs, further confirmed that abcd4 morphants possessed fewer hemoglobinized cells and impaired blood circulation. Multiple protein sequence alignment revealed that the amino acid sequence for residues 13–292, which is the domain of vitamin B 12 transport, of the zebrafish Abcd4 was highly conserved compared to that of other species. Accordingly, the abcd4 morphants can be rescued with human ABCD4 , demonstrating a conserved role of ABCD4 in vertebrates. Notably, the vitamin B 12 -deficient phenotype in abcd4 morphants, which causes anemia, was recapitulated in the newly-established abcd4 mutant, indicating the possibility that the abcd4 mutant could be used as a disease model of vitamin B 12 -deficiency anemia. Our study provides an insight that the analysis of the newly-established abcd4 mutant may contribute to understanding its roles in ABCD4-related vitamin B 12 -deficiency anemia and the associated pathogeneses in humans. ● Abcd4 knockdown in zebrafish results in anemic phenotype ● Zebrafish abcd4 mutant contains dysmorphic RBCs and induces juvenile death ● The role of ABCD4 in animal development may be conserved in vertebrates ● Zebrafish abcd4 mutant may serve as an animal model to study cobalamin metabolism [ABSTRACT FROM AUTHOR]

Transforming growth factor-beta (TGF-β) superfamily proteins play a critical role in proliferation, differentiation, and other functions of mesenchymal stem cells (MSCs). During chondrogenic differentiation of MSCs, TGF-β up-regulates chondrogenic gene expression by enhancing the expression of the transcription factor SRY (sex-determining region Y)-box9 (Sox9). In this study, we investigated the effect of continuous TGF-β1 overexpression in human synovium-derived MSCs (hSD-MSCs) on immunophenotype, differentiation potential, and proliferation rate. hSD-MSCs were transduced with recombinant retroviruses (rRV) encoding TGF-β1. The results revealed that continuous overexpression of TGF-β1 did not affect their phenotype as evidenced by flow cytometry and reverse transcriptase PCR (RT-PCR). In addition, continuous TGF-β1 overexpression strongly enhanced cell proliferation of hSD-MSCs compared to the control groups. Also, induction of chondrogenesis was more effective in rRV-TGFB-transduced hSD-MSCs as shown by RT-PCR for chondrogenic markers, toluidine blue staining and glycosaminoglycan (GAG)/DNA ratio. Our data suggest that overexpression of TGF-β1 positively enhances the proliferation and chondrogenic potential of hSD-MSCs. [ABSTRACT FROM AUTHOR]

Abstract: In this study, we attempted to elucidate the E3 ubiquitin ligase for topo IIα. When cullins and VHL were ectopically expressed in HT1080 and HEK293T cells, topo IIα was degraded most prominently in cullin 2- and VHL-expressing cells. Cullin 2 and the β domain (aa 114–123) of VHL, a subunit of the ECV (Elongin B/C-cullin 2-VHL protein) complex, specifically interact with the ATPase domain of topo IIα. We identified that topo IIα associated with endogenous Elongin C. In HT1080 cells co-transfected with deletion mutants of topo IIα GRDD (glucose-regulated destruction domain) and VHL, topo IIα was degraded by VHL expression. These results demonstrate that ECV acts as E3 ubiquitin ligase targeting GRDD-independent topo IIα to the ubiquitin–proteasome pathway. [Copyright &y& Elsevier]

In cell-based tissue engineering, fibrin hydrogel can be utilized to produce scaffolds to treat cartilage. However, the optimal fibrin formulation for cartilage regeneration has not yet been studied. This study aimed to find the optimal fibrin formulation and determine whether fibrin optimized with human adipose-derived stem cells (hASCs) increased the in-vivo chondrogenic potential. To find the optimal formulation, fibrin constructs were divided into twelve groups with different ratios of fibrinogen (10, 20, 30, and 50 mg/mL) to thrombin (10, 50, and 100 IU/mL), following which the physical and biological properties of cell-free and cell-embedded fibrin were investigated. The results from cell-free hydrogels showed that increases in the concentrations of fibrinogen and thrombin corresponded to increases in stiffness and initial weight. Moreover, hydrogel degradation was inhibited in high-concentration formulations. In cell-embedded fibrin constructs, the variation of gel formulation did not affect cell viability. However, cell behavior depended on the gel formulation. hASCs within high-concentration fibrinogen formulation maintained a round morphology similar to natural chondrocytes. Variations in thrombin concentration had a lesser effect on cell morphology. In terms of in-vivo cartilage formation, the formulation with 30 mg/mL fibrinogen and 100 IU/mL thrombin showed the highest cartilage formation, as evidenced through collagen type II alpha 1 chain (COL2) and safranin-O, 4 weeks after implantation. The results may lead to optimally designed 3D bio-scaffolds in which we can control both cell survival and chondrogenic potential for cartilage tissue engineering. Scaffolds made with the optimal fibrin formulation can be applied to develop cell therapies with mesenchymal stem cells to treat osteoarthritis. • The high fibrin formulation increased in stiffness and initial weight and also inhibited hydrogel degradation. • hASCs within high-concentration fibrinogen formulation maintained a round morphology similar to natural chondrocytes. • The formulation with 30 mg/mL fibrinogen and 100 IU/mL thrombin showed the highest cartilage formation. [ABSTRACT FROM AUTHOR]

Highlights: [•] Co-culture of hSDMSCs with SNP-stimulated chondrocytes improves anti-inflammation. [•] Co-culture system produces IGF-1. [•] Co-culture system suppresses inflammatory genes expression. [•] Co-culture system improves cell proliferation. [•] Exogenous IGF-1 inhibits inflammatory activity in SNP-stimulated chondrocytes. [ABSTRACT FROM AUTHOR]

Development of hepatocellular carcinoma (HCC) is a complex process involving multiple changes in gene expression and usually occurs in the presence of liver cirrhosis. In this research, we observed proteome alterations of three tissue types isolated from livers of HCC patients: normal, cirrhotic, and tumorous tissue. Proteome alterations were observed using two-dimensional polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Comparing the tissue types with each other, a significant change in expression level was found in 21 proteins. Of these proteins, sarcosine dehydrogenase, liver carboxylesterase, peptidyl-prolyl isomerase A, and lamin B1 are considered novel HCC marker candidates. In particular, lamin B1 may be considered as a marker for cirrhosis, because its expression level changes considerably in cirrhotic tissue compared with normal tissue. The proteins revealed in this experiment can be used in the future for studies pertaining to hepatocarcinogenesis, or as diagnostic markers and therapeutic targets for HCC. [Copyright &y& Elsevier]