Uptake Of Retinoic Acid Research Articles

Electrophoretic mobility of neuron-like cells regenerated from iPSCs with induction of retinoic acid- and nerve growth factor-loaded solid lipid nanoparticles

Abstract The capillary electrophoresis (CE) technique has been widely used to improve our understanding of biochemistry, and has become one of the best choices to address critical issues in neurobiology. In this study, doubled-emulsified solid lipid nanoparticles (DESLNs) grafted with PPFLMLLKGSTR (PM12) were fabricated to carry retinoic acid (RnA) and nerve growth factor (NGF), and to differentiate induced pluripotent stem cells (iPSCs) into nerve cells. The membrane charge of differentiated neuron-like cells was investigated using a high-performance CE. We found that surface PM12 enhanced the uptake of RnA- and NGF-loaded DESLNs (RnA-NGF-DESLNs) via strong interaction with cellular integrin α3β1 and α5β1, as evidenced by immunofluorescence assay. Further, highly expressed βIII-tubulin, a typical neuronal marker, suggested the improved neurite outgrowth of differentiating iPSCs. The enhanced electrophoretic mobility also demonstrated the reproduction of neurons from iPSCs treated with various kinds of DESLNs. After longer induction, the major CE elution peak appeared in shorter time, evidencing an increase in electrostatic potential (more negative) and effective neuronal regeneration. Induction with PM12-RnA-NGF-DESLNs to differentiate iPSCs resulted in the largest electrophoretic mobility (more negative), zeta potential and membrane charge. Thus, this study showed the ability of CE to analyze the charge variation during guided differentiation of iPSCs with drug carriers, and the CE method can be promising in evaluating the formation of matured neurons from iPSCs.

Cytoglobin expression in the hepatic stellate cell line HSC-T6 is regulated by extracellular matrix proteins dependent on FAK-signalling.

BackgroundFibrosis is a physiological response to cellular injury in the liver and is mediated by the activation of hepatic stellate cells resulting in the replacement of hepatocytes with extracellular matrix comprised principally of collagen 1 to form a hepatic scar. Although the novel hexaco-ordinated globin cytoglobin was identified in activated hepatic stellate cells more than 10 years ago, its role in stellate cell biology and liver fibrosis remains enigmatic.ResultsIn the current study, we investigated the role of different extracellular matrix proteins in stellate cell proliferation, activation (alpha smooth muscle actin expression and retinoic acid uptake) and cytoglobin expression. Our results demonstrate that cytoglobin expression is correlated with a more quiescent phenotype of stellate cells in culture and that cytoglobin is regulated by the extracellular matrix through integrin signalling dependent on activation of focal adhesion kinase.ConclusionsAlthough further studies are required, we provide evidence that cytoglobin is a negative regulator of stellate cell activation and therefore may represent a novel target for anti-fibrotic treatments in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s13069-015-0032-y) contains supplementary material, which is available to authorized users.

Reverse transformation of hepatic myofibroblast-like cells by TGFβ1/LAP

The activation of hepatic stellate cells (HSCs) to myofibroblasts (MFBs) is a key process for initiation of hepatic fibrosis and accumulation of the extracellular matrix (ECM). In this process, transforming growth factor beta1 (TGFbeta1) plays an important role in activating HSCs. In this study, we determined whether the activation of HSC was suppressed by latency-associated peptide (LAP) that is a part of TGFbeta1 precursor peptide. An MFB-like cell line (MFBY2) established from a fibrotic rat liver was infected with a recombinant adenovirus expressing LAP (AxCALAP). As results, AxCALAP-infected MFBY2 arrested cell proliferation and significantly decreased in expression of TGFbeta1 and ECM components. Interestingly, the expression of glial fibrillary acidic protein and up-take of retinoic acid were enhanced by AxCALAP-infection, while expression of alpha-smooth muscle actin was inhibited. These results suggested that overexpression of LAP in MFBs induces the reverse transformation to HSC phenotype. The adenoviral vector used in this study may have possible therapeutic applications in liver fibrosis.

Factors modulating the effect of retinoids on cultured retinal pigment epithelial cell proliferation

The effect of several naturally-occurring retinoids and 13-cis-retinoic acid on the proliferation of cultured bovine retinal pigment epithelial (RPE) cells was investigated. None of the retinoids tested were toxic to the cultures and all, except retinylpalmitate, inhibited cell proliferation when given for more than 3 days. The relative potencies of the retinoids were; all-trans-retinoic acid greater than 13-cis-retinoic acid greater than all-trans-retinol approximately equal to all-trans-retinaldehyde. Uptake of retinoic acid by cultured RPE cells was 10-fold less than the uptake of retinol. Although retinoic acid-treated cultures showed strong density-dependent growth inhibition, cellular proliferation was inhibited more in sparse cultures than in dense ones. Retinoic acid did not significantly inhibit the proliferation of first passage bovine or rabbit RPE cells, but partially inhibited the proliferation of first passage human RPE cells. The sensitivity of all these cultures to growth inhibition by retinoic acid increased in subsequent subcultures, yet there was no effect of passage number on retinoic acid uptake. This study demonstrates that RPE cell proliferation can be inhibited by retinoic acid but the sensitivity of these cells to the retinoid's effects are modulated by incubation time, in vitro aging, and cell density.

Increase in membrane uptake of long-chain fatty acids early during preadipocyte differentiation.

An increase in early rates of oleate uptake, which reflected fatty acid (FA) entry into the cells, was apparent 2-3 days after confluence of differentiating BFC-1 preadipocytes. The increase was measured in cells kept without glucose and with arsenate, where greater than 95% of intracellular radioactivity was recovered as free unesterified oleate. Uptake of retinoic acid, a molecule structurally similar to long-chain FA, remained unaltered during cell differentiation. Increase in oleate transport was related to increase in transport Vmax (determined under arsenate treatment) from 0.2 to 2 nmol/min per 10(6) cells, whereas Km remained unchanged (2 x 10(-7) M). Oleate transport was maximal at about day 6 after cell confluence (day 0), as FA metabolism (incorporation into lipids) began to gradually increase. The increase in transport preceded induction of mRNAs for both cytosolic FA-binding protein, which appeared at day 6, and for the FA synthase, which appeared at day 10. Data indicated that increases in activities of FA transport and of lipoprotein lipase, early during cell differentiation, favored increased availability of exogenous FA at a stage when endogenous FA synthesis is limited. This result would promote FA esterification and lipid deposition by supplying a rate-limiting substrate. Furthermore, oleate addition to BFC-1 preadipocytes at confluence potentiated the effect of dexamethasone in inducing mRNA for cytosolic FA-binding protein. In adipocytes, FA from exogenous or endogenous sources was necessary to maintain levels of cytosolic FA-binding protein mRNA. Thus, the increase in FA availability might contribute to, or modulate, induction of proteins necessary for preadipocyte differentiation.