• Journal of Korean Neurosurgical Society
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• v.64 no.5
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• pp.705-715
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• 2021

Objective : Through our previous clinical trials, the demonstrated therapeutic effects of MSC in chronic spinal cord injury (SCI) were found to be not sufficient. Therefore, the need to develop stem cell agent with enhanced efficacy is increased. We transplanted enhanced Wnt3-asecreting human mesenchymal stem cells (hMSC) into injured spines at 6 weeks after SCI to improve axonal regeneration in a rat model of chronic SCI. We hypothesized that enhanced Wnt3a protein expression could augment neuro-regeneration after SCI. Methods : Thirty-six Sprague-Dawley rats were injured using an Infinite Horizon (IH) impactor at the T9-10 vertebrae and separated into five groups : 1) phosphate-buffered saline injection (injury only group, n=7); 2) hMSC transplantation (MSC, n=7); 3) hMSC transfected with pLenti vector (without Wnt3a gene) transplantation (pLenti-MSC, n=7); 4) hMSC transfected with Wnt3a gene transplantation (Wnt3a-MSC, n=7); and 5) hMSC transfected with enhanced Wnt3a gene (1.7 fold Wnt3a mRNA expression) transplantation (1.7 Wnt3a-MSC, n=8). Six weeks after SCI, each 5×10⁵ cells/15 µL at 2 points were injected using stereotactic and microsyringe pump. To evaluate functional recovery from SCI, rats underwent Basso-Beattie-Bresnahan (BBB) locomotor test on the first, second, and third days post-injury and then weekly for 14 weeks. Axonal regeneration was assessed using growth-associated protein 43 (GAP43), microtubule-associated protein 2 (MAP2), and neurofilament (NF) immunostaining. Results : Fourteen weeks after injury (8 weeks after transplantation), BBB score of the 1.7 Wnt3a-MSC group (15.0±0.28) was significantly higher than that of the injury only (10.0±0.48), MSC (12.57±0.48), pLenti-MSC (12.42±0.48), and Wnt3a-MSC (13.71±0.61) groups (p<0.05). Immunostaining revealed increased expression of axonal regeneration markers GAP43, MAP2, and NF in the Wnt3a-MSC and 1.7 Wnt3a-MSC groups. Conclusion : Our results showed that enhanced gene expression of Wnt3a in hMSC can potentiate axonal regeneration and improve functional recovery in a rat model of chronic SCI.

• Journal of Microbiology and Biotechnology
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• v.19 no.9
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• pp.966-971
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• 2009

Peroxidase from Coprinus cinereus (CiP) has attracted attention for its high specific activity and broad substrate spectrum compared with other peroxidases. In this study, the functional expression of this peroxidase was successfully achieved in the methylotrophic yeast Pichia pastoris. The expression level of CiP was increased by varying the microbial hosts and the expression promoters. Since a signal sequence, such as the alpha mating factor of Saccharomyces cerevisiae, was placed preceding the cDNA of the CiP coding gene, expressed recombinant CiP (rCiP) was secreted into the culture broth. The Mut Pichia pastoris host showed a 3-fold higher peroxidase activity, as well as 2-fold higher growth rate, compared with the $Mut^s $ Pichia pastoris host. Furthermore, the AOX1 promoter facilitated a 5-fold higher expression of rCiP than did the GAP promoter.

• Journal of Sericultural and Entomological Science
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• v.28 no.2
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• pp.15-20
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• 1986

This study was carried out to investigate the elucidation of gene expression in embryo formation of the Kidney mutant, especially yolk proteins in unfertilized eggs and template activity of m-RNA extracted from them. The results were summarized as follow ; There was no recognized qualitative difference in yolk proteins of unferilized eggs between the Kidney mutant and normal. There was not any difference, between Kidney mutant and normal in the molecular species of m-RNA derived from maternal origin with the template activity in vitro.

• Development and Reproduction
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• v.17 no.1
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• pp.63-72
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• 2013

A specific inhibitor of RNA polymerase II, ${\alpha}$-amanitin is broadly used to block transcriptional activities in cells. Previous studies showed that ${\alpha}$-amanitin affects in vitro maturation of cumulus-oocyte-complex (COC). In this study, we evaluated the target of ${\alpha}$-amanitin, and whether it affects oocytes or cumulus cells (CCs), or both. We treated ${\alpha}$-amanitin with different time period during in vitro culture of denuded oocytes (DOs) or COCs in comparison, and observed the changes in morphology and maturation status. Although DOs did not show any change in morphology and maturation rates with ${\alpha}$-amanitin treatment, oocytes from COCs were arrested at metaphase I (MI) stage and CCs were more scattered than control groups. To discover causes of meiotic arrest and scattering of CCs, we focused on changes of cumulus expansion, gap junctions, and cellular metabolism which to be the important factors for the successful in vitro maturation of COCs. Expression of genes for cumulus expansion markers (Ptx3, Has2, and Tnfaip6) and gap junctional proteins (Gja1, Gja4, and Gjc1) decreased in ${\alpha}$-amanitin-treated CCs. However, these changes were not observed in oocytes. In addition, expression of genes related to metabolism (Prps1, Rpe, Rpia, Taldo1, and Tkt) decreased in ${\alpha}$-amanitin-treated CCs but not in oocytes. Therefore, we concluded that the transcriptional activities of CCs for supporting suitable transcripts, especially for its metabolic activities and formation of gap junctions among CCs as well as with oocytes, are important for oocytes maturation in COCs.

• The Journal of Korean Medicine
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• v.29 no.2
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• pp.133-150
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• 2008

Objective: This study was performed to examine Danggui (DG, Angelica gigas Nakai)'s potential activity for promoting axonal regeneration in the injured peripheral nerve. Methods: Using the sciatic nerve in the rats, DG extract 5 ${\mu}l$(10 mg/ml in 0.5% saline) was dripped into the injury site of the nerve. Results: DG treatment facilitated axonal elongation responses in the distal portion to the injury site. GAP-43 protein levels were upregulated by DG treatment in the injured nerve and also in the DRG, suggesting the induction of GAP-43 expression at gene expression level after nerve injury. Phospho-Erk1/2 protein levels were upregulated in the injured nerve area and also in the DRG, suggesting retrograde transport of phospho-Erk1/2 protein from the injury area to the cell body. Cdc2 protein levels were slightly upregulated by DG treatment. DG treatment increased the number of non-neuronal cells in the distal portion to the injury site. Conclusions: The present data suggest that DG is effective for enhanced axonal regrowth after sciatic nerve injury.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.2
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• pp.431-437
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• 2008

Sengmaek-san(Shengmai-san; SMS) is used in oriental medicine as one of the key herbal medicine for treating diverse symptoms including cardiovascular and neurological disorders. In the present study, the effects of SMS on axonal regeneration were investigated in the rat model given sciatic nerve injury. SMS treatment enhanced axonal regrowth into and the number of non-neuronal cells in the distal area after crush injury. GAP-43 protein levels were increased in the injured sciatic nerve compared to intact nerve and further upreguated by SMS treatment. GAP-43 protein was increased similarly in the dorsal root ganglion (DRG) at lumbar 4 - 6 by nerve injury and SMS treatment, suggesting GAP-43 induction at gene expression level. SMS-mediated increase in phospho-Erk1/2 protein was observed in the DRG as well as in the injured nerve implying its retrograde transport into the cell body as the process of lesion signal transmission. The present findings suggest that SMS may be involved in enhanced axonal regeneration via dynamic regulation of regeneration-associated proteins.

• Journal of Microbiology and Biotechnology
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• v.19 no.2
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• pp.208-214
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• 2009

Steroid hormones have long been known to have a profound influence on the immune system. Although the functions of the nuclear receptors in the development of T cells are fairly well studied, the differential expression of these receptors in T helper cells is poorly understood. Here, we investigated the differential expression of nuclear receptors and coregulators in Th1 and Th2 cells by genome-wide micro array analysis. The result showed that several nuclear receptors and coregulators are differentially expressed in these cells. The result was confirmed by RT-PCR. The result showed that $RXR{\alpha}$ is highly expressed in Th2 cells. Overexpression of $RXR{\alpha}$ in a Jurkat human T cell line induced IL4 but not IFN-${\gamma}$ gene expression, suggesting that $RXR{\alpha}$ plays a selective role in Th1 and Th2 differentiation. In summary, these results suggest that Th1/Th2 differentiation is influenced by differential regulation of nuclear receptors and coregulators.

• Applied Biological Chemistry
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• v.39 no.4
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• pp.260-264
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• 1996

Process of particle bombardment for efficient transformation of Cymbidium virescence rhizome microcross sections was investigated using Biolistic particle delivery system with pBI121 harboring the ${\beta}-glucuronidase$(GUS) and the neomycin phosphotransferaseII(nptII). The best result was obtained from the combination of $1.11{\;}{\mu}m$ tungsten particles coated with pBl121, $77.33kg/cm^2$ helium pressure, 6.35 mm gap distance, and 7.0 cm target distance. Transient expression of the reporter gene, GUS, bombarded into the rhizome microsections was observed by the histochemical assay. The marker gene, nptII, delivered by bombarding the tungsten particles coated with the plasmid DNA was identified in the transformed rhizome by polymerase chain reaction.

• Journal of Life Science
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• v.10 no.6
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• pp.630-639
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• 2000

The gastric pathogen Helicobacter pylori(H. pylori) establishes long-term chronic infection that can lead to atrophic gastritis, intestinal metaplasia, and gastric cancer. H. pylori, which express cytotoxic genes is now recohnized as a cause of peptic ulcer and is also a major risk factor for the development of gastric adenocarcinoma. We performed this study 1) to assess the detection rate of H. pylori according to direct investigation of bacteria of gastric biopsy specimen and two serologic tests of GAP test and Helico blot 2.0 system in the symptomatic and non-symptomatic group 2) to evaluate and compare the efficacy of two serologic tests of GAP test and Helico blot 2.0 system for the diagnosis of H. pylori infection. Forty-nine patients were positive for H pylori infection based on direct investigation of bacteria by histology. The detection rates of H. pylori infection based on direct investigation of bacteria by histology. The detection rates of H. phlori were significantly lower in gastric cancer than in other gastroduodenal disease(p<0.05). The concordance of two serologic tests of GAP test and Helico blot 2.0 system is poor. There was no statistically significant difference between the expression rate of CagA and VacA in the symptomatic and non-symptomatic group. Although Helico blot 2.0 system may not displace GAP test, it was a very sensitive serologic test for the diagnosis of H. pylori infection and it was used to detect IgG antibodies to H. pylori-specific antigens, including CagA, VacA and the various urease subunit. Our data suggest that further investigation is needed to determine whether or not the serologic expression of cytotoxic gene may be clinical usefulness of diagnostic methods in the gastroduodenal disease.

• Molecules and Cells
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• v.26 no.3
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• pp.243-249
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• 2008

Corticotropin releasing factor (CRF) mediates various responses to stress through CRF receptors 1 and 2. CRF receptor 2 has two forms, $2{\alpha}$ and $2{\beta}$ each of which appears to have distinct roles. Here we used dopaminergic neuron-derived MN9D cells to investigate the function of CRF receptor 2 in dopamine neurons. We found that n-butyrate, a histone deacetylase inhibitor, induced MN9D cell differentiation and increased gene expression of all CRF receptors. CRF receptor $2{\beta}$ was minimally expressed in MN9D cells; however, its expression dramatically increased during differentiation. CRF receptor $2{\beta}$ expression levels appeared to correlate with neurite outgrowth, suggesting CRF receptor $2{\beta}$ involvement in neuronal differentiation. To validate this statement, we made a CRF receptor $2{\beta}$-overexpressing $MN9D/CRFR2{\beta}$ stable cell line. This cell line showed robust neurite outgrowth and GAP43 overexpression, together with MEK and ERK activation, suggesting MN9D cell neuronal differentiation. From these results, we conclude that CRF receptor $2{\beta}$ plays an important role in MN9D cell differentiation by activating the MEK/ERK signaling pathway.