• Korean Journal of Microbiology
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• v.40 no.3
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• pp.183-188
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• 2004

Stenotrophomonas sp. OK-5 capable of degrading TNT has been found to have three nitroreductase fractions designated as NTR fractions I, II, and III. NTR in a previous study. This study was attempted to reveal physiological and molecular characteristics of NTR fractions I, II, and III in strain OK-5. Several chemicals (e.g., EDTA, NaCl, dithiothreitol, $\beta$-mercaptoethanol) were tested for their effect on enzyme activity of NTRs, demonstrating that enzyme activities of NTR fractions I, II, and III from OK-5 were inhibited in the presence of $\beta$-mercaptoethanol. Substrate specificity test showed that NTR fractions I, II, and III all have over 70% enzyme activities for nitrobenzene or RDX as a substrate. N-terminal amino acid sequence of NTR fraction I from Stenotrophomonas sp. OK-5 was $^1MSDLLNADAVVQLFRTARDS^20$ and exhibited 70% sequence homology with that of NTR from Xanthomonas campestris. NTR I gene from Stenotrophomonas sp. OK-5 (SmOK5nrI) shared extensive sequence homology in deduced amino acid sequence of PCR product with NTRs from Xanthomonas campestris (81 %), X. axonopodis (75%), Streptomyces avermitilis(30%), whereas they had low homology with that from P. putida KT2440 (pnrB) (16%).

• Journal of Korean Neurosurgical Society
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• v.62 no.6
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• pp.626-634
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• 2019

Objective : Nerve growth factor (NGF) is a member of the neurotrophic factor family and plays a vital role in the physiological processes of organisms, especially in the nervous system. Many recent studies have reported that NGF is also involved in the regulation of tumourigenesis by either promoting or suppressing tumor growth, which depends on the location and type of tumor. However, little is known regarding the effect of NGF on interspinal schwannoma (IS). In the present study, we aimed to explored whether mouse nerve growth factor (mNGF), which is widely used in the clinic, can influence the growth of interspinal schwannoma cells (ISCs) isolated from IS in vitro. Methods : ISCs were isolated, cultured and identified by S-100 with immunofluorescence analysis. S-100-positive cells were divided into five groups, and separately cultured with various concentrations of mNGF (0 [phosphate buffered saline, PBS], 40, 80, 160, and 320 ng/mL) for 24 hours. Western blot and quantantive real time polymerase chain reaction (PCR) were applied to detect tyrosine kinase A (TrkA) receptor and p75 neurotrophin receptor ($p75^{NTR}$) in each group. Crystal violet staining was selected to assess the effect of mNGF (160 ng/mL) on ISCs growth. Results : ISCs growth was enhanced by mNGF in a dose-dependent manner. The result of crystal violet staining revealed that it was significantly strengthened the cells growth kinetics when cultured with 160 ng/mL mNGF compared to PBS group. Western blot and quantantive real time PCR discovered that TrkA receptor and mRNA expression were both up-regualated under the condition of mNGF, expecially in 160 ng/mL, while the exoression of $p75^{NTR}$ demonstrated no difference among groups. Conclusion : From these data, we conclude that exogenous mNGF can facilitate ISC growth by activating both TrkA receptor and $p75^{NTR}$. In addition, patients who are suffering from IS should not be administered mNGF in the clinic.

• Journal of Life Science
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• v.21 no.8
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• pp.1134-1141
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• 2011

Transmembrane protein 21 (TMP21) is a member of the p24 cargo protein family and has been shown to modulate ${\alpha}$-secretase-mediated A${\beta}$ production which was specifically observed in the brains of subjects with Alzheimer's disease (AD). In order to investigate whether TMP21 could affect nerve growth factor (NGF) receptor signaling pathway, the alteration of NGF receptors and their downstream proteins were detected in TMP21 over-expressed cells. CMV/hTMP21 vector used in this study was successfully expressed into TMP21 proteins in B35 cells after lipofectamin transfection. Expressed TMP21 proteins induced the down-regulation of ${\gamma}$-secretase complex components including Presenlin-1 (PS-1), PS-2, Nicastrin (NST), Pen-2 and APH-1. Also, the expression level of NGF receptor $p75^{NTR}$ and RhoA were significantly higher in CMV/hTMP21 transfectants than vehicle transfectants, while their levels returned to vehicle levels after NGF treatment. However, the phosphorylation of NGF receptor TrkA was dramtically decreased in NGF No-treated CMV/hTMP21 transfectants compared with vehicle transfectants, and increased in NGF treated CMV/hTMP21 transfectants. In TrkA downstream signaling pathway, the phosphorylation level of ERK was also decreased in CMV/hTMP21 transfectants, while the phosphorylation of Akt was increased in the same transfectants. Furthermore, NGF treatment induced the increase of phosphorylation level of Akt and ERK in CMV/hTMP21 transfectants. Therefore, these results suggested that over-expression of TMP21may simultaneously induce the up-regulation of $p75^{NTR}$/RhoA expression and the down-regulation of TrkA/ERK phosphorylation through the inhibition of ${\gamma}$-secretase activity.

• Journal of Life Science
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• v.23 no.6
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• pp.812-824
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• 2013

Reserpine, an anti-hypertensive drug, is able to positively modulate several phenotypes associated with $A{\beta}$ toxicity in a Caenorhabditis elegans model of Alzheimer's disease (AD). We investigated into the therapeutic effects of reserpine on mammalian neurodegenerative disorders, and found that significant alteration of the key factors influencing AD was detected in Tg2576 mice after reserpine treatment for 30 days. The aggressive behavior of Tg2576 mice was significantly improved upon reserpine treatment, whereas their social contact was consistently maintained. Furthermore, the levels of $A{\beta}$-42 peptide in the hippocampus of the brain and blood serum were lower in the reserpine-treated group than in the vehicle-treated group. Among g-secretase components, the expression levels of PS-2, Pen-2, and APH-1 were slightly lower in reserpine-treated Tg2576 mice, although a significant change in nicastrin (NCT) expression was not detected. Furthermore, the serum level of nerve growth factor (NGF) increased in reserpine-treated Tg2576 mice compared with vehicle-treated mice. Among down-stream effectors of the NGF receptor TrkA signaling pathway, reserpine treatment induced elevation of TrkA phosphorylation and reduction of ERK phosphorylation. In addition, in the NGF receptor $p75^{NTR}$ signaling pathway, the expression levels of $p75^{NTR}$ and Bcl-2 were enhanced in reserpine-treated Tg2576 mice compared with vehicle-treated mice, whereas the expression level of RhoA declined. Overall, these results suggest that reserpine can help relieve AD pathogenesis in Tg2576 mice through downregulation of $A{\beta}$-42 deposition, alteration of ${\gamma}$-secretase components, and regulation of NGF metabolism.

• Asian Spine Journal
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• v.12 no.6
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• pp.998-1009
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• 2018

Study Design: Olfactory ensheathing cells (OECs) from rat olfactory mucosa were cultured, characterized, and transplanted into a rat model of spinal cord injury (SCI). Purpose: To evaluate different doses of OECs in a rat model of SCI. Overview of Literature: SCI causes permanent functional deficit because the central nervous system lacks the ability to perform spontaneous repair. Cell therapy strategies are being explored globally. The clinical use of human embryonic stem cell is hampered by ethical controversies. Alternatively, OECs are a promising cell source for neurotransplantation. This study aimed to evaluate the efficacy of different doses of allogenic OEC transplantation in a rat model of SCI. Methods: OECs were cultured from the olfactory mucosa of Albino Wistar rats; these cells were characterized using immunohistochemistry and flow cytometry. Rats were divided into five groups (n=6 rats each). In each group, different dosage ($2{\times}10^5$, $5{\times}10^5$, $10{\times}10^5$, and >$10{\times}10^5$) of cultured cells were transplanted into experimentally injured spinal cords of rat models. However, in the SCI group, only DMEM (Dulbecco's modified Eagle's medium) was injected. Rats were followed up upto 8 weeks post-transplantation. The outcome of transplantation was assessed using the Basso, Beattie, Bresnahan (BBB) scale; motor-evoked potential studies; and histological examination. Results: Cultured cells expressed 41% of p75NTR, a marker for OEC, and 35% of anti-fibronectin, a marker for olfactory nerve fibroblast. These cells also expressed $S100{\beta}$ and glial fibrillary acid protein of approximately 75% and 83%, respectively. All the transplanted groups showed promising BBB scores for hind-limb motor recovery compared with the SCI group (p<0.05). A motor-evoked potential study showed increased amplitude in all the treated groups compared with the SCI. Green fluorescent protein-labeled cells survived in the injured cord, suggesting their role in the transplantation-mediated repair. Transplantation of $5{\times}10^5$ cells showed the best motor outcomes among all the doses. Conclusions: OECs demonstrated a therapeutic effect in rat models with the potential for future clinical applications.

• Molecules and Cells
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• v.38 no.3
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• pp.221-228
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• 2015

Because Schwann cells perform the triple tasks of myelination, axon guidance and neurotrophin synthesis, they are candidates for cell transplantation that might cure some types of nervous-system degenerative diseases or injuries. However, Schwann cells are difficult to obtain. As another option, ectomesenchymal stem cells (EMSCs) can be easily harvested from the nasal respiratory mucosa. Whether fibrin, an important transplantation vehicle, can improve the differentiation of EMSCs into Schwann-like cells (SLCs) deserves further research. EMSCs were isolated from rat nasal respiratory mucosa and were purified using anti-CD133 magnetic cell sorting. The purified cells strongly expressed HNK-1, nestin, $p75^{NTR}$, S-100, and vimentin. Using nuclear staining, the MTT assay and Western blotting analysis of the expression of cell-cycle markers, the proliferation rate of EMSCs on a fibrin matrix was found to be significantly higher than that of cells grown on a plastic surface but insignificantly lower than that of cells grown on fibronectin. Additionally, the EMSCs grown on the fibrin matrix expressed myelination-related molecules, including myelin basic protein (MBP), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) and galactocerebrosides (GalCer), more strongly than did those grown on fibronectin or a plastic surface. Furthermore, the EMSCs grown on the fibrin matrix synthesized more neurotrophins compared with those grown on fibronectin or a plastic surface. The expression level of integrin in EMSCs grown on fibrin was similar to that of cells grown on fibronectin but was higher than that of cells grown on a plastic surface. These results demonstrated that fibrin not only promoted EMSC proliferation but also the differentiation of EMSCs into the SLCs. Our findings suggested that fibrin has great promise as a cell transplantation vehicle for the treatment of some types of nervous system diseases or injuries.

• Journal of Life Science
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• v.21 no.9
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• pp.1234-1243
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• 2011

Liriope platyphylla has been used in oriental medicine as an effective medical plant to improve symptoms of cough, sputum production, neurodegenerative disorders, obesity and diabetes for long time. In order to investigate the effects of novel extracts on nerve growth factors (NGF)-stimulated neuritic outgrowth, the alteration of NGF expression and NGF receptor signaling pathway were detected in neuroblastoma cells and C57BL/6 mice. Of a total of 13 novel extracts, 4 extracts (LP-E, LP-M, LP-M50, LP2E17PJ) showed high viability on MTT assay. Also, all of these extracts induced NGF secretion and NGF mRNA expression in neuroblastoma cells. However, the NGF-induced neuritic outgrowth from PC12 cells was only stimulated by LP-E, LP-M and LP-M50. Furthermore, we selected LP-M as a best candidate, based on method and amounts of extraction, in order to verify its effect in mice. C57BL/6 mice were treated with 50 mg/kg of LP-M for 2 weeks and the effects on NGF regulation were analyzed with various methods. The expression of NGF mRNA was significantly increased in LP-M treated mice compared to vehicle treated mice. Also, the signaling pathway of p75NTR was inhibited in the cortex by LP-M treatment, with no change in the hippocampus of brain. However, the signaling pathway of TrkA was dramatically activated in only hippocampus via LP-M treatment. Therefore, these results suggest that the novel four extracts of L. platyphylla may contribute to the regulation of NGF expression and secretion in neuronal cells. LP-M was especially considered to be an excellent candidate for a neurodegenerative disease-therapeutic drug.