The Effect of Differential Modulation of the N-methyl-D-aspartate Receptor on Growth and intracellular Calcium Ion Concentration in Normal Human Oral Keratinocytes

The purpose of this study was to examine the interaction between the agents [N-methyl-D-aspartate (NMDA), arachidonic acid (AA), and Nitric Oxide Synthase Inhibitor (NOS-I)] and cultured normal human oral keratinocytes (NHOK) in order to elucidate the mechanisms by which epithelial growth and regeneration are regulated. We also examined whether AA and NOS-I could protect NHOK from glutamate cytotoxicity and the change of intracellular calcium ion concentration. NHOK were obtained from gingival tissue of 20 individuals aged 20 to 29, and third passage (P3) cells were used for this study. Cell viability was measured by the MTT assay and DNA synthesis by the BrdU assay. The microscopic features of NHOK were observed and changes in intracellular calcium ion concentration were measured. NMDA and NNA induced an initial increase in cell number. Low concentration of AA induced an increase in cell number while high concentrations of AA induced a decrease in cell number. The decrease in cell number induced by NMDA at the seventh day was abolished by the addition of low concentrations of AA or NOS inhibitors. Low concentrations of AA or NMDA with high concentrations of AA significantly increased the DNA synthesis rate at four hours. NMDA and AA both induced an increase in intracellular calcium ion concentration, and AA enhanced NMDA-induced intracellular calcium ion in a concentration-dependent manner. NMDA-induced NHOK death was associated with intracellular calcium ion change and the promotion of cell differentiation. Low concentrations of AA protected NHOK cells from NMDA-induced death.

Short-Term Effects of Combined Serial Casting and Botulinum Toxin Injection for Spastic Equinus in Ambulatory Children with Cerebral Palsy

PURPOSE: The purpose of this paper is to test the hypothesis that combination therapy of serial cast and botulinum toxin type A (BTX-A) injection can further enhance the effects of a BTX-A injection in ambulant children with cerebral palsy (CP) who have an equinus foot. MATERIALS AND METHODS: Children in group A (30 legs of 21 children) received a serial casting application after an injection of BTX-A, and children in group B (25 legs of 17 children) received only a BTX-A injection. Assessments were performed before the intervention and 1 month after the intervention. RESULTS: After the intervention, there were significant improvements in tone, dynamic spasticity, and passive range of motion (ROM) in both groups. However, the changes were greater in group A than in group B. Dimension D (standing) in Gross Motor Function Measure (GMFM)-66 was significantly improved in group A but not in group B. On the other hand, there were no significant changes in dimension E (walking, running, jumping) in GMFM-66 in either group. CONCLUSION: The results of our study suggest that a serial casting application after BTX-A injection can enhance the benefits of BTX-A injection in children with cerebral palsy.ope

The Effect of Calcipotriol on the Expression of Human β Defensin-2 and LL-37 in Cultured Human Keratinocytes

Background. Vitamin D has been reported to regulate innate immunity by controlling the expression of antimicrobial peptides (AMPs). Objective. We investigated the effect of calcipotriol on the expression of AMPs in human cultured keratinocytes. Methods. Keratinocytes were treated with lipopolysaccharide (LPS), TNF-α, Calcipotriol and irradiated with UVB, cultured, and harvested. To assess the expression of human beta defensin-2 and LL-37 in the control group, not exposed to any stimulants, the experimental group was treated with LPS, TNF-α, or UVB, and another group was treated again with calcipotriol; reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemical staining were performed. Results. In the experimental group treated with LPS, UVB irradiation, and TNF-α, the expression of β-defensin and LL-37 was increased more than in the control group and then decreased in the experimental group treated with calcipotriol. Conclusions. Calcipotriol suppressed HBD-2 and LL-37, which were stimulated by UVB, LPS, and TNF-α

A versatile approach to multiple gene RNA interference using microRNA-based short hairpin RNAs

Background: Effective and stable knockdown of multiple gene targets by RNA interference is often necessary to overcome isoform redundancy, but it remains a technical challenge when working with intractable cell systems. Results: We have developed a flexible platform using RNA polymerase II promoter-driven expression of microRNA-like short hairpin RNAs which permits robust depletion of multiple target genes from a single transcript. Recombination-based subcloning permits expression of multi-shRNA transcripts from a comprehensive range of plasmid or viral vectors. Retroviral delivery of transcripts targeting isoforms of cAMP-dependent protein kinase in the RAW264.7 murine macrophage cell line emphasizes the utility of this approach and provides insight to cAMP-dependent transcription. Conclusion: We demonstrate functional consequences of depleting multiple endogenous target genes using miR-shRNAs, and highlight the versatility of the described vector platform for multiple target gene knockdown in mammalian cells

The Neuroprotective Effect of Gugijihwang-Tang on Trimethyltin-Induced Memory Dysfunction in the Rat

Gugijihwang-Tang (the herbal formula PM012), a decoction consisting of several herbs including Rehmanniae Radix Preparata, has been widely used as herbal treatment for dementia. In order to investigate the neuroprotective action of this prescription, we examined the effect of Gugijihwang-Tang on learning and memory using the Morris water maze and [F-18]FDG micro PET neuroimaging technique. After injection of trimethyltin (TMT, 8.0 mg/kg, i.p.), which is a potent toxicant that selectively kills cells in the central nervous system, rats were administered Gugijihwang-Tang (100 mg/kg, p.o.) daily for two weeks, followed by the Morris water maze tasks and [F-18]FDG micro PET neuroimaging. In Gugijihwang-Tang administered TMT-treated rats, they showed improved learning and memory abilities in water maze tasks and glucose metabolism, suggesting that Gugijihwang-Tang plays effectively positive role in the improvement of brain function including learning and memory after TMT-induced neurodegeneration. Taken together, our results suggested that the Gugijihwang-Tang should be useful for developing strategies protecting nervous system and improving brain function

Potentiated therapeutic angiogenesis by primed human mesenchymal stem cells in a mouse model of hindlimb ischemia

Background: Human bone marrow-derived mesenchymal stem cells (hMSCs) are advantageous for cell-based therapy to treat ischemic diseases owing to their capacity to secrete various paracrine factors with potent angiogenic activity. Materials methods: In this study, we describe a method to increase secreted levels of VEGF and HGF from hMSCs without genetic modification. Results: We demonstrated that transplantation of primed hMSCs into ischemic limbs led to significantly greater improvements in tissue perfusion and limb salvage by increasing capillary formation compared with nonprimed hMSCs. The primed hMSCs also exhibited greater survival in vivo and secreted human VEGF and HGF in the ischemic tissue, supporting enhanced angiogenesis and cell survival. Conclusion: These findings indicate that priming hMSCs via methods described in this study enhances secretion of critical proangiogenic factors resulting in an enhanced therapeutic effect of cells for the treatment of ischemic diseases.This research was supported by grants from the Korea Health Technology R&D Project (M-S Chang, A100823) and the Innovative Research Institute for Cell Therapy, National University Hospital (A062260), both sponsored by the Ministry of Health, Welfare and Family, Seoul, Republic of Korea.OAIID:oai:osos.snu.ac.kr:snu2013-01/102/0000027724/1SEQ:1PERF_CD:SNU2013-01EVAL_ITEM_CD:102USER_ID:0000027724ADJUST_YN:YEMP_ID:A075930DEPT_CD:862CITE_RATE:3.718FILENAME:첨부된 내역이 없습니다.DEPT_NM:치의과학과EMAIL:[email protected]_YN:YCONFIRM:

Synthetic dsRNA Dicer substrates enhance RNAi potency and efficacy

RNA interference (RNAi) is the process of sequence-specific post-transcriptional gene silencing triggered by double-stranded RNAs. In attempts to identify RNAi triggers that effectively function at lower concentrations, we found that synthetic RNA duplexes 25–30 nucleotides in length can be up to 100-fold more potent than corresponding conventional 21-mer small interfering RNAs (siRNAs). Some sites that are refractory to silencing by 21-mer siRNAs can be effectively targeted by 27-mer duplexes, with silencing lasting up to 10 d. Notably, the 27-mers do not induce interferon or activate protein kinase R (PKR). The enhanced potency of the longer duplexes is attributed to the fact that they are substrates of the Dicer endonuclease, directly linking the production of siRNAs to incorporation in the RNA-induced silencing complex. These results provide an alternative strategy for eliciting RNAi-mediated target cleavage using low concentrations of synthetic RNA as substrates for cellular Dicer-mediated cleavage

The Alliance for Cellular Signaling Plasmid Collection: A Flexible Resource for Protein Localization Studies and Signaling Pathway Analysis

Cellular responses to inputs that vary both temporally and spatially are determined by complex relationships between the components of cell signaling networks. Analysis of these relationships requires access to a wide range of experimental reagents and techniques, including the ability to express the protein components of the model cells in a variety of contexts. As part of the Alliance for Cellular Signaling, we developed a robust method for cloning large numbers of signaling ORFs into Gateway® entry vectors, and we created a wide range of compatible expression platforms for proteomics applications. To date, we have generated over 3000 plasmids that are available to the scientific community via the American Type Culture Collection. We have established a website at www.signaling-gateway.org/data/plasmid/ that allows users to browse, search, and blast Alliance for Cellular Signaling plasmids. The collection primarily contains murine signaling ORFs with an emphasis on kinases and G protein signaling genes. Here we describe the cloning, databasing, and application of this proteomics resource for large scale subcellular localization screens in mammalian cell lines

Electrospun silk fibroin scaffolds with macropores for bone regeneration: An in vitro and in vivo study

We developed three-dimensional electrospun silk fibroin (ESF) scaffolds with controllable pore size. The purpose of this study was to evaluate ESF scaffolds with pores (P-ESF) for bone regeneration via in vitro and in vivo studies, with a comparison to a commercially available porous three-dimensional polylactic acid (PLA) scaffold. P-ESF supported significantly higher proliferation and alkaline phosphatase activity of osteoblasts than PLA in vitro (p<0.05). Moreover, higher expression levels of activated adhesion-related proteins, including focal adhesion kinase, were observed in the P-ESF than in PLA, as confirmed by western blot analyses. Microcomputed tomography revealed that 78.30% of the original bone volume was attained in the P-ESF implantation group at 7 weeks after critical bone defect formation in rat calvaria. Comparatively, the PLA implantation group showed only 49.31%. Histological evaluation also showed new bone tissue formation upon P-ESF implantation. Taken together, the P-ESF scaffold may be a good bone substitute for bone regeneration.The authors thank the Korea Science and Engineering Foundation (KOSEF) and Rural Development Administration, Republic of Korea, for sponsoring this research through the SRC=ERC Program of MOST=KOSEF (R11-2005-065), and BioGreen21 Program (200810FTH010102001)