Apoptosis ; Caspase 3 ; Caspases ; physiology ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Isotretinoin ; pharmacology ; Melanoma ; pathology ; Receptors, Retinoic Acid ; physiology ; Retinoid X Receptors ; physiology ; Tretinoin ; pharmacology

All-trans retinoic acid (ATRA) is a vitamin A derivative and plays an important role in the regulation of cell aggregation, differentiation, apoptosis, proliferation, and inflammatory response. In recent years, some progress has been made in the role of ATRA in renal diseases, especially its protective effect on podocytes. This article reviews the research advances in podocyte injury, characteristics of ATRA, podocyte differentiation and regeneration induced by ATRA, and the protective effect of ATRA against proliferation, deposition of fibers, and apoptosis.
Apoptosis ; drug effects ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cytoprotection ; Humans ; Podocytes ; drug effects ; physiology ; Tretinoin ; pharmacology

The present study was designed to analyze the metabolites of all-trans-retinal (atRal) and compare the cytotoxicity of atRal versus its derivative all-trans-retinoic acid (atRA) in human retinal pigment epithelial (RPE) cells. We confirmed that atRA was produced in normal pig neural retina and RPE. The amount of all-trans-retinol (atROL) converted from atRal was about 2.7 times that of atRal-derived atRA after incubating RPE cells with 10 μmol/L atRal for 24 h, whereas atRA in medium supernatant is more plentiful (91 vs. 29 pmol/mL), suggesting that atRA conversion facilitates elimination of excess atRal in the retina. Moreover, we found that mRNA expression of retinoic acid-specific hydroxylase CYP26b1 was dose-dependently up-regulated by atRal exposure in RPE cells, indicating that atRA inactivation may be also initiated in atRal-accumulated RPE cells. Our data show that atRA-caused viability inhibition was evidently reduced compared with the equal concentration of its precursor atRal. Excess accumulation of atRal provoked intracellular reactive oxygen species (ROS) overproduction, heme oxygenase-1 (HO-1) expression, and increased cleaved poly(ADP-ribose) polymerase 1 (PARP1) expression in RPE cells. In contrast, comparable dosage of atRA-induced oxidative stress was much weaker, and it could not activate apoptosis in RPE cells. These results suggest that atRA generation is an antidotal metabolism pathway for atRal in the retina. Moreover, we found that in the eyes of ABCA4^-/-RDH8^-/- mice, a mouse model with atRal accumulation in the retina, the atRA content was almost the same as that in the wild type. It is possible that atRal accumulation simultaneously and equally promotes atRA synthesis and clearance in eyes of ABCA4^-/-RDH8^-/- mice, thus inhibiting the further increase of atRA in the retina. Our present study provides further insights into atRal clearance in the retina.
ATP-Binding Cassette Transporters/physiology* ; Alcohol Oxidoreductases/physiology* ; Animals ; Cell Survival/drug effects* ; Cells, Cultured ; Humans ; Inactivation, Metabolic ; Mice ; Retina/metabolism* ; Retinal Pigment Epithelium/metabolism* ; Swine ; Tretinoin/pharmacology*

OBJECTIVETo study the effect of RAR-beta transfection plus treatment with the corresponding ligand ATRA on the proliferation and phenotype of platelet-derived growth factor (PDGF)-activated hepatic stellate cells (HSC).

METHODSPDGF-activated hepatic stellate cells of rats were transfected with eukaryotic expression vector pCMV-script-RAR-beta, which was verified by western blot. The proliferation of transfected HSC was assayed by BrdU incorporation as well as MTT methods. Their phenotype (alpha-SMA and desmin) was observed by immunocytochemistry assay with image analysis and RAR-beta protein expression was detected by western blot.

RESULTSTransfection of RAR-beta gene and treatment with ligand ATRA could increase the expression of RAR-beta protein for at least 144h and inhibit the proliferation and the expression of alpha-SMA and desmin in PDGF-activated HSC. Significant statistical differences were perceived comparing with sham-transfected, only-PDGF treated, non-ligand treated and irrelevant ligand-treated HSC.

CONCLUSIONSTransfected with RAR-beta gene as well as using related ligand ATRA could suppress the proliferation and reverse the activation phenotype of activated HSC.

Animals ; Blotting, Western ; Cell Division ; Liver ; cytology ; Phenotype ; Platelet-Derived Growth Factor ; pharmacology ; Rats ; Receptors, Retinoic Acid ; physiology ; Transfection ; Tretinoin ; pharmacology

OBJECTIVETo study the effects of TrkB-BDNF signal pathway on the synthesis and secretion of vascular endothelial growth factor (VEGF) in human neuroblastoma cells (NB).

METHODSTrkB protein expression in SY5Y cells before and after all-trans-retinoicacid (ATRA) treatment was detected by Western blot. P-TrkB protein expression in SY5Y cells before and after the treatment of ATRA along with BDNF was also detected by Western blot. VEGF concentrations in the SY5Y cell culture supernatants were measured using ELISA after the treatment with ATRA, BDNF, tyrosine kinase inhibitor K252a and PI3k inhibitor LY294002.

RESULTSTrkB protein was undetectable in SY5Y cells before ATRA treatment. After the treatment of 1, 10 and 100 nM/L ATRA for five days, TrkB protein was expressed in SY5Y cells and the TrkB protein level increased with the increasing ATRA concentration. P-TrkB protein was not expressed in SY5Y cells treated only with 10 nM/L ATRA, but it was detectable after the treatment of ATRA along with BDNF. VEGF concentrations in the group treated with ATRA+BDNF were significantly higher than those in the untreated control and the ATRA alone treatment groups (P<0.01). VEGF concentrations in the K252a pretreated ATRA+BDNF group were significantly lower than those in the group treated with ATRA+BDNF (P<0.05). VEGF concentrations in the LY294002 treatment group (ATRA+LY294002+BDNF group) were also significantly lower than those in the group treated with ATRA+BDNF (P<0.01).

CONCLUSIONSActivation of TrkB-BDNF signal pathway may increase the synthesis and secretion of VEGF in human NB cells. The synthesis and secretion of VEGF can be inhibited by blocking TrkB-BDNF signal pathway with K252a or blocking the TrkB-BDNF downstream signal pathway PI3K/Akt with LY294002.

Brain-Derived Neurotrophic Factor ; physiology ; Cell Line, Tumor ; Enzyme-Linked Immunosorbent Assay ; Humans ; Neuroblastoma ; metabolism ; pathology ; Phosphatidylinositol 3-Kinases ; physiology ; Proto-Oncogene Proteins c-akt ; physiology ; Receptor, trkB ; physiology ; Signal Transduction ; physiology ; Tretinoin ; pharmacology ; Vascular Endothelial Growth Factor A ; analysis ; biosynthesis

This study was directed at assessing the effects of embryonic bodies on differentiation of the embryonic stem cells. The embryonic bodies of different days gained by means of hanging, suspending and hung-suspended were processed by all-trans retinotic acids for 4 days. Then they were detected by immunocytochemistry methods and were measured to detect the fluorescence intensity changes of calcium after Glu stimulation. During this period, we observed the differentiation process and compared the differentiation ratio of embryonic stem cells processed by means of hanging, suspending for several days. It was found that the differentiation ratio of EBs obtained by suspending for 3 days, or by hanging for 3 days and suspending for 1 day, is relatively higher, which can help us find out the intrinsic differentiation mechanism of embryonic stem cells.
Animals ; Cell Differentiation ; physiology ; Cells, Cultured ; Embryo, Mammalian ; cytology ; Embryonic Induction ; Mice ; Mice, Inbred ICR ; Neurons ; cytology ; Stem Cells ; cytology ; Tretinoin ; pharmacology

OBJECTIVETo study the role of extracellular matrix (ECM) in neural differentiation of mouse embryonic stem cells (ESCs).

METHODSMouse ESCs were incubated in the ESC conditioned medium, and the formation of embryonic bodies (EBs) were induced in bacteriological dishes using high-concentration all-trans retinoic acid (RA). The EBs were seeded on different matrixes (gelatin, fibronectin, and laminin/poly-L-ornithine) to test their impact on neural differentiation of the ESCs using immunofluorescence assay. The effect of laminin/poly-L-ornithine on the growth of neurites was evaluated with fluorescence microscopy.

RESULTSHigh-concentration RA activated and accelerated the differentiation of ESCs toward nestin-positive neural progenitor cells. Fibronectin supplement in the matrix dose-dependently promoted ESC differentiation into neural progenitor cells, while laminin/poly-L-ornithine increased the growth of the neurites and induced the maturation of the differentiated neural cells.

CONCLUSIONECM plays an important role in neural differentiation of mouse ESCs, and application of FN produces the most conspicuous effect during the differentiation of the ESCs into the neural progenitor cells;laminin/poly-L-ornithine is the most effective during their differentiation into neural cells.

Animals ; Cell Differentiation ; drug effects ; Cells, Cultured ; Culture Media ; Embryonic Stem Cells ; cytology ; drug effects ; Extracellular Matrix ; physiology ; Fibronectins ; pharmacology ; Laminin ; pharmacology ; Mice ; Neurons ; cytology ; drug effects ; Peptides ; pharmacology ; Tretinoin ; pharmacology

Neurogenesis can be induced by pathological conditions such as cerebral ischemia. However the molecular mechanisms or modulating reagents of the reactive neurogenesis after the cerebral ischemia are poorly characterized. Retinoic acid (RA) has been shown to increase neurogenesis by enhancing the proliferation and neuronal differentiation of forebrain neuroblasts. Here, we examined whether RA can modulate the reactive neurogenesis after the cerebral ischemia. In contrast to our expectation, RA treatment decreased the reactive neurogenesis in subventricular zone (SVZ), subgranular zone (SGZ) and penumbral region. Furthermore, RA treatment also decreased the angiogenesis and gliosis in penumbral region.
Animals ; Brain/blood supply ; Cell Differentiation ; Cell Proliferation ; Ischemic Attack, Transient/metabolism/*pathology ; Male ; Neovascularization, Pathologic ; Neuroglia/pathology/physiology ; Neurons/pathology/*physiology ; Rats ; Rats, Sprague-Dawley ; Tretinoin/*pharmacology/physiology

Cell motility is essential for a wide range of cellular activities including anigogenesis as well as metastasis of tumor cells. Ras has been implicated in cell migration and invasion, and functions at upstream of mitogen-activated protein kinase (MAPK) families, which include extracellular-signal regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 MAPK. In the present study, we examined the role of JNK in endothelial cell motility using stable transfectant (DAR-ECV) of ECV304 endothelial cells expressing previously established oncogenic H-Ras (leu 61). DAR-ECV cells showed an enhanced angiogenic potential and motility (approximately 2-fold) compared to ECV304 cells. Western blot analysis revealed constitutive activation of JNK in DAR-ECV cells. Pretreatment of JNK specific inhibitors, curcumin and all trans-retinoic acid, decreased the basal motility of DAR-ECV cells in a dose-dependent manner. These inhibitors also suppressed the motility stimulated by known JNK agonists such as TNFalpha and anisomycin. To further confirm the role of JNK, ECV304 cells expressing dominant active SEK1 (DAS-ECV) were generated. Basal non-stimulated levels of the cellular migration were greater in DAS-ECV clones than those in control ECV304 cells. These results suggest that Ras-SEK1-JNK pathway regulates motility of endothelial cells during angiogenesis.
Anisomycin/pharmacology ; Cell Line ; *Cell Movement ; Curcumin/pharmacology ; Endothelium, Vascular/cytology/*physiology ; Enzyme Activation ; Enzyme Inhibitors/pharmacology ; Extracellular Matrix/metabolism ; Genes, ras/genetics ; Human ; Matrix Metalloproteinases/physiology ; Mitogen-Activated Protein Kinases/*metabolism ; Neovascularization, Physiologic ; Support, Non-U.S. Gov't ; Tretinoin/pharmacology ; Tumor Necrosis Factor/pharmacology ; Umbilical Veins/cytology ; Urinary Plasminogen Activator/physiology

Cell motility is essential for a wide range of cellular activities including anigogenesis as well as metastasis of tumor cells. Ras has been implicated in cell migration and invasion, and functions at upstream of mitogen-activated protein kinase (MAPK) families, which include extracellular-signal regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 MAPK. In the present study, we examined the role of JNK in endothelial cell motility using stable transfectant (DAR-ECV) of ECV304 endothelial cells expressing previously established oncogenic H-Ras (leu 61). DAR-ECV cells showed an enhanced angiogenic potential and motility (approximately 2-fold) compared to ECV304 cells. Western blot analysis revealed constitutive activation of JNK in DAR-ECV cells. Pretreatment of JNK specific inhibitors, curcumin and all trans-retinoic acid, decreased the basal motility of DAR-ECV cells in a dose-dependent manner. These inhibitors also suppressed the motility stimulated by known JNK agonists such as TNFalpha and anisomycin. To further confirm the role of JNK, ECV304 cells expressing dominant active SEK1 (DAS-ECV) were generated. Basal non-stimulated levels of the cellular migration were greater in DAS-ECV clones than those in control ECV304 cells. These results suggest that Ras-SEK1-JNK pathway regulates motility of endothelial cells during angiogenesis.
Anisomycin/pharmacology ; Cell Line ; *Cell Movement ; Curcumin/pharmacology ; Endothelium, Vascular/cytology/*physiology ; Enzyme Activation ; Enzyme Inhibitors/pharmacology ; Extracellular Matrix/metabolism ; Genes, ras/genetics ; Human ; Matrix Metalloproteinases/physiology ; Mitogen-Activated Protein Kinases/*metabolism ; Neovascularization, Physiologic ; Support, Non-U.S. Gov't ; Tretinoin/pharmacology ; Tumor Necrosis Factor/pharmacology ; Umbilical Veins/cytology ; Urinary Plasminogen Activator/physiology