Differentiation of Cultured Human Epidermal Keratinocytes at High Cell Densities is Mediated by Endogenous Activation of the Protein Kinase C Signaling Pathway

Normal human epidermal keratinocytes (NHEK) grown in serum-free medium on a plastic substrate spontaneously differentiate at high cell densities in vitro. Because protein kinase C (PKC) regulates murine keratinocyte differentiation triggered by a variety of stimuli, we examined the role of this signaling pathway in density-dependent activation of NHEK differentiation. Relative to subconfluent cultures, confluent NHEK expressed markedly higher levels of multiple differentiation markers assayed by immunoblotting, including keratin 1, loricrin, filaggrin, involucrin, TGK, and SPR-1. Expression of several of these markers continued to increase for several days after cells reached confluency. The total level of several PKC isoforms was not substantially altered in NHEK harvested at different cell densities, based on immunoblotting; however, subcellular fractionation revealed that PKCα underwent a redistribution to the particulate fraction in confluent and postconfluent NHEK cultures, suggesting that this isozyme was activated under these conditions and may be involved in triggering the terminal differentiation program. Supporting this concept, inhibition of PKC function using bryostatin 1 or GF 109203X blocked the induction of keratinocyte differentiation markers at high cell densities. These data suggest that endogenous activation of PKC is responsible for cell density-mediated stimulation of NHEK differentiation, establishing a critical role for this pathway in regulating human as well as murine keratinocyte differentiation

Radio Protective Effects of Ginseng Extract in Gamma-Rays Induced Chromosomal Damages of Human Lymphocyte

Ginsan, a polysaccharide extracted from Panax ginseng and subsequently referred as ginseng, posses various biological properties as an anticancer and antioxidant agent. Ginseng also approved effective against radiation effects through its immunomodulating actions in whole body irradiated mouse. But its protective effects on radiation induced DNA damage are not thoroughly investigated, mainly in human. This experiment aimed to assess the effects of ginseng at 2 working doses in suppressing radiation effects of human peripheral blood lymphocyte (PBL) i.e. chromosome aberration and micronuclei yields. The treatment times were 24 hours before, subsequently (0 hour) or 3 hours after and irradiation with gamma rays at doses of 0.5 - 2.0 Gy (dose rate of 3.16 Gy/min). Treated and untreated blood cultivation and metaphase spreading technique was done according to standard procedures. Results showed that without ginseng treatments, radiation significantly increased dicentrics and micronuclei frequencies. Different with the results in mouse study, however, our results indicated that none of the experimental concentrations of ginseng crude water extract tested had an effect on baseline chromosomal aberration and micronuclei (MN) yields in PBL. A protective effect was only seen in chromosome aberration yields of sample irradiated with 2.0 Gy and treated with ginseng 3 h post irradiation rather than 24 h pre-irradiation in one volunteer. Opposite results that ginseng suspected to be a weak radiosensitizer was found in some cases. This may be due to discrepancies exist in route of treatment and its fundamental mechanisms of protective action between both studies. Even though in general it was not effective, the possible mechanism involved in radioprotective influence of ginseng is discussed

Radiation Response Protein, Sialyltransferase (ST6Gal I)

Recently we identified β-galactoside α(2, 6)-sialyltransferase (ST6Gal I) as a candidate biomarker for ionizing radiation. The expression of ST6Gal I and the level of protein sialylation increased following radiation exposure in a dose-dependent manner. We also found that radiation induced ST6Gal I cleavage and the cleaved form of ST6Gal I was soluble and secreted. Sialylation of integrin β1, a glycosylated cell surface protein, was stimulated by irradiation and this increased its protein stability. Overexpression of ST6Gal I in SW480 colon cancer cells that initially showed a low enzyme activity of ST6Gal I increased the sialylation of integrin β1 and also increased the stability of the protein. Inhibition of sialylation by transfection with neuramidase or by treatment with short interfering (si) RNA targeting ST6Gal I (Si-ST6Gal I) reversed the effects of ST6Gal I expression. In addition, ST6Gal I overexpression increased clonogenic survival following radiation exposure and reduced radiation-induced cell death and caspase 3 activation. In conclusion, we suggest that exposure to ionizing radiation was found to increase sialylation of glycoproteins such as integrin β1 by inducing the expression of ST6 Gal I, and finally protein sialylation contributed to cellular radiation resistance

Synergistic Uric Acid-Lowering Effects of the Combination of Chrysanthemum indicum

Chrysanthemum indicum Linne flower (CF) and Cinnamomum cassia (L.) J. Persl bark (CB) extracts have served as the main ingredients in several prescriptions designed to treat hyperuricemia and gout in traditional Chinese and Korean medicine. However, little is known about the combination effects of a CF and CB (CC) mixture on hyperuricemia. In our study, we investigated the antihyperuricemic effects of CC mixture and the mechanisms underlying these effects in normal and potassium oxonate- (PO-) induced hyperuricemic rats. The CC mixture significantly decreased uric acid levels in normal and PO-induced hyperuricemic rats and showed the enhanced hypouricemic effect compared to CF or CB alone. Furthermore, the CC mixture increased renal uric acid excretion in PO-induced hyperuricemic rat. We found that CC mixture and its major components, chlorogenic acid, 3,4-dicaffeoylquinic acid (isochlorogenic acid), coumarin, cinnamaldehyde, trans-cinnamic acid, and o-methoxycinnamaldehyde, inhibit the activity of xanthine oxidase (XOD) in vitro. The CC mixture exerts antihyperuricemic effects accompanied partially by XOD activity inhibition. Therefore, the CC mixture may have potential as a treatment for hyperuricemia and gout

Growth differentiation factor 11 locally controls anterior-posterior patterning of the axial skeleton.

Growth and differentiation factor 11 (GDF11) is a transforming growth factor β family member that has been identified as the central player of anterior-posterior (A-P) axial skeletal patterning. Mice homozygous for Gdf11 deletion exhibit severe anterior homeotic transformations of the vertebrae and craniofacial defects. During early embryogenesis, Gdf11 is expressed predominantly in the primitive streak and tail bud regions, where new mesodermal cells arise. On the basis of this expression pattern of Gdf11 and the phenotype of Gdf11 mutant mice, it has been suggested that GDF11 acts to specify positional identity along the A-P axis either by local changes in levels of signaling as development proceeds or by acting as a morphogen. To further investigate the mechanism of action of GDF11 in the vertebral specification, we used a Cdx2-Cre transgene to generate mosaic mice in which Gdf11 expression is removed in posterior regions including the tail bud, but not in anterior regions. The skeletal analysis revealed that these mosaic mice display patterning defects limited to posterior regions where Gdf11 expression is deficient, whereas displaying normal skeletal phenotype in anterior regions where Gdf11 is normally expressed. Specifically, the mosaic mice exhibited seven true ribs, a pattern observed in wild-type (wt) mice (vs. 10 true ribs in Gdf11-/- mice), in the anterior axis and nine lumbar vertebrae, a pattern observed in Gdf11 null mice (vs. six lumbar vertebrae in wt mice), in the posterior axis. Our findings suggest that GDF11, rather than globally acting as a morphogen secreted from the tail bud, locally regulates axial vertebral patterning

Cognitive Enhancing and Neuroprotective Effect of the Embryo of the Nelumbo nucifera

The aim of the present study was to evaluate the effect of ENS on cognitive impairment induced by scopolamine and its potential neuroprotective effect against glutamate-induced cytotoxicity in HT22 cell and to investigate the underlying mechanisms. ENS (3, 10, 30, and 100 mg/kg), scopolamine (1 mg/kg), and donepezil (1 mg/kg) were administered to mice during a test period. Scopolamine impaired memory and learning in a water maze test and a passive avoidance test. The neuroprotective effect of ENS (10 and 100 μg/mL) was investigated on glutamate-induced cell death in HT22 cells by MTT assay. We investigated acetylcholinesterase inhibition in hippocampus and antioxidant activity, ROS levels, and Ca2+ influx in HT22 cells to elucidate the potential mechanisms of ENS. We found that ENS significantly ameliorated scopolamine-induced memory impairment and inhibited AChE activity in hippocampus. In vitro, ENS showed potent neuroprotective effects against glutamate-induced neurotoxicity in the HT22 cell. In addition, ENS induced a decrease in ROS production and intercellular Ca2+ accumulation and showed DPPH radical and H2O2 scavenging activity. In conclusion, ENS showed both a memory improving effect and a neuroprotective effect. Our results indicate that ENS may be of use in the treatment and prevention of neurodegenerative disorders

MMP9 Processing of HSPB1 Regulates Tumor Progression

Matrix metalloproteinases regulate pathophysiological events by processing matrix proteins and secreted proteins. Previously, we demonstrated that soluble heat shock protein B1 (HSPB1) is released primarily from endothelial cells (ECs) and regulates angiogenesis via direct interaction with vascular endothelial growth factor (VEGF). Here we report that MMP9 can cleave HSPB1 and release anti-angiogenic fragments, which play a key role in tumorprogression. We mapped the cleavage sites and explored their physiological relevance during these processing events. HSPB1 cleavage by MMP9 inhibited VEGF-induced ECs activation and the C-terminal HSPB1 fragment exhibited more interaction with VEGF than did full-length HSPB1. HSPB1 cleavage occurs during B16F10 lung progression in wild-type mice. Also, intact HSPB1 was more detected on tumor endothelium of MMP9 null mice than wild type mice. Finally, we confirmed that secretion of C-terminal HSPB1 fragment was significantly inhibited lung and liver tumor progression of B16F10 melanoma cells and lung tumor progression of CT26 colon carcinoma cells, compared to full-length HSPB1. These data suggest that in vivo MMP9-mediated processing of HSPB1 acts to regulate VEGF-induced ECs activation for tumor progression, releasing anti-angiogenic HSPB1 fragments. Moreover, these findings potentially explain an anti-target effect for the failure of MMP inhibitors in clinical trials, suggesting that MMP inhibitors may have pro-tumorigenic effects by reducing HSPB1 fragmentation