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Huang W.-C.,Taipei Veterans General Hospital | Huang W.-C.,Graduate Institute of Medical science | Huang W.-C.,National Yang Ming University | Kuo H.-S.,Taipei Veterans General Hospital | And 13 more authors.
Journal of Gene Medicine | Year: 2011

Background: Following spinal cord injury, the delivery of neurotrophic factors to the injured spinal cord has been shown to promote axonal regeneration and functional recovery. In previous studies, we showed that acidic fibroblast growth factor (aFGF) is a potent neurotrophic factor that promotes the regeneration of axotomized spinal cord or dorsal root ganglion neurones. Methods: We constructed a recombinant adeno-associated virus (AAV) vector to express human aFGF and evaluated aFGF expression and function in AAV-aFGF-infected PC12 cells. We analyzed AAV-green fluorescent protein (GFP) tropism and AAV-mediated aFGF expression in contused spinal cords. Animals received behavioural testing to evaluate the functional recovery. Results: Overexpression of aFGF was shown in AAV-aFGF-infected PC12 cells in a dose-dependent manner. Concurrently, neurite extension and cell number were significantly increased in AAV-aFGF infected cells. AAV-mediated GFP expression persisted for at least 5 weeks in contused spinal cords, and the most prominently transduced cells were neurones. Contusive injury reduced endogenous aFGF expression in spinal cords. Overexpression of aFGF was demonstrated in AAV-aFGF transduced spinal cords compared to AAV-GFP transduced spinal cords at 3 and 14 days post-injury. Evaluation of motor function revealed that the improvement of AAV-aFGF-treated rats was prominent. Both AAV-aFGF- and recombinant human aFGF-treated rats revealed significantly better recovery at 5 weeks post-injury, compared to vehicle- and AAV-GFP-treated rats. Conclusions: These data suggest that supplement of aFGF improve the functional recovery of spinal cord-contused rats and that AAV-aFGF-mediated gene transfer could be a clinically feasible therapeutic approach for patients after nervous system injuries. © 2011 John Wiley & Sons, Ltd.


Lin Y.-H.,National Taiwan University of Science and Technology | Fu K.-Y.,Tri Service General Hospital | Hong P.-D.,National Taiwan University of Science and Technology | Ma H.,Veteran General Hospital | And 11 more authors.
Annals of Plastic Surgery | Year: 2013

ABSTRACT: Embryonic stem cells (ESCs) are pluripotent cells that can differentiate into various cell types, including keratinocyte-like cells, within suitable microniches. In this study, we aimed to investigate the effects of culture media, cell coculture, and a tissue-engineering biocomposite on the differentiation of mouse ESCs (MESCs) into keratinocyte-like cells and applied these cells to a surgical skin wound model. MESCs from BALB/c mice (ESC26GJ), which were transfected using pCX-EGFP expressing green fluorescence, were used to track MESC-derived keratinocytes. Weak expression of the keratinocyte early marker Cytokeratin 14 (CK-14) was observed up to 12 days when MESCs were cultured in a keratinocyte culture medium on tissue culture plastic and on a gelatin/collagen/polycaprolactone (GCP) biocomposite. MESCs cocultured with human keratinocyte cells (HKCs) also expressed CK-14, but did not express CK-14 when cocultured with human fibroblast cells (HFCs). Furthermore, CK-14 expression was observed when MESCs were cocultured by seeding HKCs or HFCs on the same or opposite side of the GCP biocomposite. The highest CK-14 expression was observed by seeding MESCs and HKCs on the same side of the GCP composite and with HFCs on the opposite side. To verify the effectiveness of wound healing in vivo, adipose-derived stem cells were applied to treat surgical wounds in nude mice. An obvious epidermis multilayer and better collagen deposition during wound healing were observed, as assessed by Masson staining. This study demonstrated the potential of keratinocyte-like differentiation from mesenchymal stem cells for use in promoting wound closure and skin regeneration. Copyright © 2013 Lippincott Williams & Wilkins.


Lan H.-C.,Academia Sinica, Taiwan | Lan H.-C.,Institute of Biology and Anatomy | Wu C.-F.,Academia Sinica, Taiwan | Wu C.-F.,National Yang Ming University | And 3 more authors.
Journal of Biological Chemistry | Year: 2012

SF-1 is a key transcription factor for all steroidogenic genes. It up-regulates the expression of the steroidogenic Cyp11a1 gene in the adrenal in a pathway stimulated by cAMP through HIPK3-mediated JNK/c-Jun phosphorylation. In the present study, we have investigated the factors mediating cAMP-dependent HIPK3 action to potentiate the activity of SF-1 for Cyp11a1 transcription in mouse adrenocortical Y1 cells. We found Daxx, a HIPK kinase substrate in the apoptosis pathway, was phosphorylated by HIPK3 at Ser-669 in response to cAMP stimulation. Daxx participated in SF-1-dependent Cyp11a1 expression as shown by experiments involving both overexpression and down-regulation via a dominant negative Daxx mutant. The S669A mutant of Daxx, which could not be phosphorylated by HIPK3, lost the ability to potentiate SF-1 activity for Cyp11a1 expression. The enhancement of SF-1 activity by Daxx required JNK and c-Jun phosphorylation. Thus, Daxx functioned as a signal transducer linking cAMP-stimulated HIPK3 activity with JNK/c-Jun phosphorylation and SF-1-dependent Cyp11a1 transcription for steroid synthesis. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.


Huang K.-H.,Taipei Medical University | Pai M.-H.,Taipei Medical University | Wu C.-H.,Institute of Biology and Anatomy | Liu J.-J.,Taipei Medical University | Yeh S.-L.,Taipei Medical University
e-SPEN | Year: 2010

Background & aims: Arginine (Arg) was shown to have immunomodulatory effect and inhibits advanced glycation end product (AGE) formation in in vitro studies. This study investigated the effects of dietary Arg supplementation on renal receptor of AGE (RAGE) expressions and oxidative damage in diabetic rats. Methods: There were 1 normal control (NC) group and 2 diabetic groups in this study. Rats in the NC group were fed with a chow diet. One diabetic group (DM) was fed a common semipurified diet, while the other diabetic group received a diet in which part of the casein was replaced by Arg (DM-Arg) for 8 wk. Diabetes was induced by an intraperitoneal injection of nicotinamide followed by streptozotocin. Rats with blood glucose levels exceeding 180 mg/dL were considered diabetic. Blood samples were collected at the baseline and 8 wk. Kidneys of the animals were harvested at the end of the study for further analysis. Results: Plasma fructosamine contents were significantly higher in the diabetic groups than in the NC group. The DM group had higher fructosamine than the DM-Arg group. Kidney nitrotyrosine concentrations and nuclear factor-κB p65 protein expressions were significantly lower in the DM-Arg group than in the DM group. The result of immunohistochemical staining also showed that the expressions of RAGE in the kidneys were significantly lower in the DM-Arg group than in the DM group. Conclusions: These results suggest that dietary Arg supplementation may decrease renal RAGE expressions and oxidative damage in rats with type 2 diabetes. © 2010 European Society for Clinical Nutrition and Metabolism.


Lan H.-C.,Institute of Biology and Anatomy | Lin I.-W.,Institute of Biology and Anatomy | Yang Z.-J.,Institute of Biology and Anatomy | Lin J.-H.,Institute of Biology and Anatomy
Toxicological Sciences | Year: 2015

Certain commonly used compounds that interfere with the functions of the endocrine system are classified as endocrine-disrupting chemicals (EDCs). Bisphenol A (BPA) is an EDC that is widely used in food containers. BPA levels in human sera are commonly observed to be approximately 1-100 nM. Compared with the effects of BPA on the gonads, its effects on the adrenal gland are poorly understood. To investigate the influence of BPA on steroidogenesis, we examined the activity of the steroidogenic gene Cyp11a1 and its regulatory pathways in mouse Y1 adrenal cortex cells. Treatment with BPA at < 100 μM did not cause cell death. However, increased promoter activity and protein expression of Cyp11a1 were induced by low doses of BPA (10-1000 nM). Moreover, BPA induced c-Jun phosphorylation, and a specific inhibitor of c-Jun N-terminal kinase (JNK) significantly suppressed BPA-induced steroidogenesis. Thus, treatment of adrenal cells with low doses of BPA activated Cyp11a1 and increased corticosterone production through the JNK/c-Jun signaling pathway. Identical results were observed in rats after BPA injection. The abnormal induction of hormone synthesis by BPA in the adrenal gland might be linked to human metabolic defects and neuropsychiatric disorders. © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved.


Shyu H.-Y.,Armed Forces Taoyuan General Hospital | Shyu H.-Y.,Institute of Biology and Anatomy | Shieh J.-C.,Chung Shan Medical University | Lin J.-H.,Taipei General Hospital | And 2 more authors.
Journal of Atherosclerosis and Thrombosis | Year: 2012

Aim: Cigarette-smoking induced oxidative DNA damage to endothelial cells has been reported to play an etiological role in atherosclerosis development. Individual vulnerability to oxidative stress through smoking exposure and the ability to repair DNA damage, which plays a critical role in modifying the risk susceptibility of large artery atherosclerotic (LAA) stroke, is hypothesized. Thus, we examined the effect of genetic polymorphisms of DNA repair pathway genes and cigarette smoking in relation to risk susceptibility of LAA stroke. Methods: We enrolled 116 LAA stroke patients and 315 healthy controls from the Armed Forces Taoyuan General Hospital, Taoyuan, Taiwan. Genotyping of polymorphisms of the OGG1 (Ser326Cys), XRCC1 (Arg399Gln), ERCC2 (Lys751Gln), and ERCC5 (Asp1104His) genes was performed and used to evaluate LAA stroke susceptibility. Results: Of those non-synonymous polymorphisms, the ERCC2 Lys751Gln variant was found to be associated with LAA stroke risk (OR: 1.69, 95%CI: 1.02-2.86), and this association was more pronounced in smokers, manifesting a 2.73-fold increased risk of LAA stroke (p = 0.027). A joint effect on risk elevation of LAA stroke was seen in those patients with OGG1 and ERCC2 polymorphisms (OR: 2.75, 95%CI: 1.26-6.00). Moreover, among smokers carrying the OGG1 Ser326Cys polymorphism, there was a tendency toward an increased risk of LAA stroke in those patients who had a greater number of high-risk genotypes of XRCC1, ERCC2, and ERCC5 polymorphisms (p trend = 0.010). Conclusion: The susceptible polymorphisms of DNA repair pathway genes may have a modifying effect on the elevated risk of LAA stroke in smokers among ethnic Chinese in Taiwan.


Shyu H.-Y.,Armed Forces Taoyuan General Hospital | Shyu H.-Y.,Institute of Biology and Anatomy | Fong C.-S.,Buddhist Dalin Tzu Chi General Hospital | Fong C.-S.,Tzu Chi University | And 6 more authors.
Clinica Chimica Acta | Year: 2010

Background: γ-Glutamyl carboxylation, a reaction essential for the biosynthesis of vitamin K-dependent coagulation factors, requires the participation of the γ-glutamyl carboxylase (GGCX), vitamin K epoxide reductase (VKORC1), and NAD(P)H:quinone oxidoreductase (NQO1). We evaluated the role of these genotype polymorphisms in patients with large-artery atherosclerotic stroke. Methods: In this hospital-based case-control study, 117 patients who were categorized as having large-artery atherosclerotic stroke and 115 age- and gender-matched controls were recruited. Genotyping determination for the GGCX1 (Gln325Arg), NQO1 (Pro187Ser), and VKORC1 (rs9923231) polymorphisms was performed. The associations of genotype with ischemic stroke (IS) risk were examined. Results: A higher genotypic frequency of NQO1 C609T was found in the controls than in the patients, manifesting a 0.47-fold risk reduction in IS (95% CI=0.25-0.87). A tendency toward a reduced IS risk was statistically significant in those subjects who carried a greater number of the NQO1, GGCX, and VKORC1 polymorphisms (aOR=0.58, P trend=0.005). The synergistic effect of multiple genes on risk reduction was more significant in a subset of patients who were not alcoholics and who were non-smokers (P<0.05). Conclusions: Compartmentation of coagulation factor metabolism may account for the preferential role of NQO1, GGCX, and VKORC1 polymorphisms to lower the risk for large-artery atherosclerotic stroke. © 2010 Elsevier B.V.

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