cytochrome-c-t and genipin

Bone regeneration is a complicated process and includes a number of distinct and sequential stages of coordinated cellular actions under the regulation of multiple growth factors. Therefore, bone grafting materials in which growth factors can be incorporated and released in a programmed order in line with the bone tissue healing process may lead to desirable clinical outcomes. In the present study, a double-layered chitosan/gelatin/genipin (d-CSG/G) nanosphere coating is developed by using layer-by-layer electrophoretic deposition and genipin crosslinking. The surface morphology, physicochemical and mechanical properties of the coatings are explored. Cytochrome C is used as a therapeutic model protein and is successfully loaded on the inner and outer layers of the coating. The protein release can be controlled by the loading position, genipin concentration and thickness of the outer layer. Furthermore, the cell response to the coatings was evaluated. Real-time polymerase chain reactions, immunofluorescence staining and extracellular matrix mineralization assay confirmed that the functions of the loaded growth factor are fully preserved after fabrication. Overall, the d-CSG/G nanosphere coating could be a promising growth factor delivery system to promote bone tissue regeneration.

Topics: Animals; Biomimetics; Bone Morphogenetic Protein 2; Calcification, Physiologic; Cattle; Chitosan; Coated Materials, Biocompatible; Cross-Linking Reagents; Cytochromes c; Delayed-Action Preparations; Extracellular Matrix; Fluorescent Antibody Technique; Gelatin; Iridoids; Mesenchymal Stem Cells; Nanospheres; Osteocalcin; Rats; Real-Time Polymerase Chain Reaction; Recombinant Proteins; Solutions; Spectroscopy, Fourier Transform Infrared; Surface Properties; Transforming Growth Factor beta

Genipin, an active constituent of Gardenia fruit, has been reported to show an anti-tumor effect in several cancer cell systems. Here, we demonstrate how genipin exhibits a strong apoptotic cell death effect in human non-small-cell lung cancer H1299 cells. Genipin-mediated decrease in cell viability was observed through apoptosis as demonstrated by induction of a sub-G1 peak through flow cytometry, DNA fragmentation measured by TUNEL assay, and cleavage of poly ADP-ribose-polymerase. During genipin-induced apoptosis, the mitochondrial execution pathway was activated by caspase-9 and -3 activation as examined by a kinetic study, cytochrome c release, and a dose-dependent increase in Bax/Bcl-2 ratio. A search for the downstream pathway reveals that genipin-induced apoptosis was mediated by an increase in phosphorylated p38MAPK expression, which further activated downstream signaling by phosphorylating ATF-2. SB203580, a p38MAPK inhibitor, markedly blocked the formation of TUNEL-positive apoptotic cells in genipin-treated cells. Besides, the interference of p38MAPK inhibited Bax expression and cytochrome c release. Altogether, our observations imply that genipin causes increased levels of Bax in response to p38MAPK signaling, which results in the initiation of mitochondrial death cascade, and therefore it holds promise as a potential chemotherapeutic agent for the treatment of H1299 cells.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; bcl-2-Associated X Protein; Carcinoma, Non-Small-Cell Lung; Cell Survival; Cytochromes c; Dose-Response Relationship, Drug; Humans; Iridoids; Mitochondria; Molecular Targeted Therapy; p38 Mitogen-Activated Protein Kinases; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Tumor Cells, Cultured

This study examined the effects of genipin, isolated from Gardenia jasminoides Ellis, on d-galactosamine (GalN) and lipopolysaccharide (LPS)-induced hepatic apoptosis and liver failure. Mice were given an intraperitoneal injection of genipin (25, 50, 100 and 200mg/kg) 1h before GalN (700mg/kg)/LPS (10microg/kg) administration. The survival rate of the genipin group was significantly higher than that of the control. Genipin markedly reduced the increases in serum aminotransferase activities and lipid peroxidation. The glutathione content decreased in GalN/LPS group, and this decrease was attenuated by genipin. Increases in serum tumor necrosis factor-alpha (TNF-alpha), which were observed in GalN/LPS-treated mice, were significantly reduced by genipin. Genipin attenuated the GalN/LPS-induced apoptosis of hepatocytes, as estimated by the caspase-3 and -8 activity assay, TNF-R1 associated death domain (TRADD) protein measurement and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) method. Moreover, increased cytosolic cytochrome c protein was reduced by genipin. After 3h of GalN/LPS injection, nuclear phosphorylated c-Jun (p-c-Jun) level was significantly increased, whereas it was attenuated by genipin. Also, the increased nuclear level of nuclear factor-kappaB and the decreased cytosolic level of IkappaB-alpha protein were significantly attenuated by genipin. Our results suggest that genipin offers marked hepatoprotection against damage induced by GalN/LPS related with its antioxidative, anti-apoptotic activities, and inhibition of NF-kappaB nuclear translocation and nuclear p-c-Jun expression.

Topics: Animals; Apoptosis; Caspases; Cell Nucleus; Cytochromes c; Cytosol; Galactosamine; Gene Expression Regulation; Glutathione; I-kappa B Proteins; Iridoid Glycosides; Iridoids; Lipid Peroxidation; Lipopolysaccharides; Liver; Liver Failure, Acute; Male; Mice; Mice, Inbred ICR; NF-KappaB Inhibitor alpha; Oxidative Stress; Phosphorylation; Protein Transport; Proto-Oncogene Proteins c-jun; TNF Receptor-Associated Death Domain Protein; Transaminases; Transcription Factor RelA; Tumor Necrosis Factor-alpha