• Journal of Microbiology and Biotechnology
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• v.22 no.5
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• pp.585-591
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• 2012

The antagonistic effect of Trichoderma harzianum on a range of seed-borne fungal pathogens of wheat (viz. Fusarium graminearum, Bipolaris sorokiniana, Aspergillus spp., and Penicillium spp.) was assessed. The potential of T. harzianum as a biocontrol agent was tested in vitro and under field conditions. Coculture of the pathogens and Trichoderma under laboratory conditions clearly showed dominance of T. harzianum. Under natural conditions, biocontrol effects were also obtained against the test fungi. One month after sowing, field emergence (plant stand) was increased by 15.93% over that obtained with the control treatment, and seedling infection was reduced significantly. Leaf blight severity was decreased from 22 to 11 at the heading stage, 35 to 31 at the flowering stage, and 86 to 74 at the grain filling stage. At harvest, the number of tillers per plant was increased by 50%, the yield was increased by 31.58%, and the 1,000-seed weight was increased by 21%.

• Archives of Pharmacal Research
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• v.19 no.5
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• pp.368-373
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• 1996

Nitric oxide (NO) is synthesized by various cells involved in inflammatory reactions and may then act on mast cells. In the present work, we attempted to clarify the role of this molecule on the proliferation of mouse bone marrow derived-mast cells (BMMC). Swiss 3T3 fibroblastsproduced nitrite ($NO_{2}$) and nitrate ($NO_{3}$) upon treatment with interferon ${\gamma}$(IFN-${\gamma}$). This formation was dependent of L-arginine and could be inhibited by the -L-arginine analogue $N^{G}$-monomethyl-L-arginine ($N^{G}$MMA). The effect of IFN-g was drastically invreased by cotreatment with tumor necrosis factor g(IFN-g). BMMC were maintained in vitro for as long as 30 days when cocultured with Swiss 3T3 fibroblasts. coculture with $N^{G}$MMA, significantly increased the number of BMMC. These results indicate that NO involves the inhibition of proliferation of BMMC when cocultured with Swiss 3T3 fibroblasts.

• Molecules and Cells
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• v.42 no.7
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• pp.530-545
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• 2019

Tumor cells can vary epigenetically during ionizing irradiation (IR) treatment. These epigenetic variegations can influence IR response and shape tumor aggressiveness. However, epigenetic disturbance of histones after IR, implicating in IR responsiveness, has been elusive. Here, we investigate whether altered histone modification after IR can influence radiation responsiveness. The oncogenic CXCL12 mRNA and protein were more highly expressed in residual cancer cells from a hepatoma heterotopic murine tumor microenvironment and coculture of human hepatoma Huh7 and normal IMR90 cells after radiation. H3K4 methylation was also enriched and H3K9 methylation was decreased at its promoter region. Accordingly, invasiveness and the subpopulation of aggressive $CD133^+/CD24^-$ cells increased after IR. Histone demethylase inhibitor IOX1 attenuated CXCL12 expression and the malignant subpopulation, suggesting that responses to IR can be partially mediated via histone modifications. Taken together, radiation-induced histone alterations at the CXCL12 promoter in hepatoma cells are linked to CXCL12 upregulation and increased aggressiveness in the tumor microenvironment.

• Biomolecules & Therapeutics
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• v.27 no.2
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• pp.185-192
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• 2019

Coculture with adipose-derived stem cells (ADSCs) can stimulate proliferation and migration of melanocytes. To enhance outcomes of skin disorders caused by melanocyte loss or death, mixed transplantation with ADSCs has been suggested. However, role of cocultured ADSCs in proliferation and migration of melanocytes remains unclear. This study determined the effect of ADSCs on production of growth factors and expression levels of intergrins in primary culture of adult human melanocytes with or without ADSCs and in nude mice grafted with such melanocytes. Higher amounts of growth factors for melanocytes, such as bFGF and SCF were produced and released from ADSCs by coculturing with melanocytes. Relative levels of integrins ${\beta}1$, ${\alpha}5$, and ${\alpha}6$ as well as adhesion to fibronectin and laminin were increased in melanocytes cocultured with ADSCs. Such increases were inhibited by neutralization of bFGF or SCF. Relative levels of bFGF, SCF and integrins were increased in nude mice skin after grafting with melanocyte+ADSC cocultures. Collectively, these results indicate that ADSCs can stimulate proliferation and migration of melanocytes by increasing expression of integrins in melanocytes through upregulation of production/release of melanocyte growth factors such as bFGF and SCF.

• Journal of Microbiology and Biotechnology
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• v.32 no.4
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• pp.397-404
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• 2022

Natural killer (NK) cell activity is more attenuated in hepatocellular carcinoma (HCC) patients than normal. Hypoxic-inducible factor (HIF)-1α is highly expressed in tumors to maintain their metabolism in a hypoxic environment. The expression of HIF-1α in cancers can lead to cell growth, proliferation, invasion/metastasis and immune escape. Although apigenin, a flavonoid, is known to have various biological activities, it has not been demonstrated in NK cell immune activity in HCC cells. In this study, NK-92 cells were directly cocultured with HCC SK-Hep1 cells for 24 h to evaluate NK cell activity in HCC cells or HCC cells expressing HIF-1α by apigenin. NK cell cytotoxicity to HCC cells expressing HIF-1α was significantly increased, and NK cell-activating receptors, NKG2D, NKp30 and NKp44 were highly expressed. The activating effect of apigenin on NK cells substantially induced apoptosis in HCC cells expressing HIF-1α through high expression of CD95L on the surface of NK-92 cells. Moreover, apigenin excellently inhibited the level of TGF-β1 in a coculture of NK cells and HCC cells. In conclusion, apigenin seems to be a good compound that increases NK cell cytotoxicity to HCC cells by controlling HIF-1α expression.

• Journal of Microbiology and Biotechnology
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• v.32 no.9
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• pp.1209-1216
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• 2022

To better understand the effects of PEGylation and biotinylation on the delivery efficiency of proteins, the cationic protein lysozyme (LZ) and anionic protein bovine serum albumin (BSA) were chemically conjugated with poly(ethylene glycol) (PEG) and biotin-PEG to primary amine groups of proteins using N-hydroxysuccinimide reactions. Four types of protein conjugates were successfully prepared: PEGylated LZ (PEG-LZ), PEGylated BSA (PEG-BSA), biotin-PEG-conjugated LZ (Bio-PEG-LZ), and biotin-PEG-conjugated BSA (Bio-PEG-BSA). PEG-LZ and Bio-PEG-LZ exhibited a lower intracellular uptake than that of LZ in A549 human lung cancer cells (in a two-dimensional culture). However, Bio-PEG-BSA showed significantly improved intracellular delivery as compared to that of PEG-BSA and BSA, probably because of favorable interactions with cells via biotin receptors. For A549/fibroblast coculture spheroids, PEG-LZ and PEG-BSA exhibited significantly decreased tissue penetration as compared with that of unmodified proteins. However, Bio-PEG-BSA showed tissue penetration comparable to that of unmodified BSA. In addition, citraconlyated LZ (Cit-LZ) showed reduced spheroid penetration as compared to that of LZ, probably owing to a decrease in protein charge. Taken together, chemical conjugation of targeting ligands-PEG to anionic proteins could be a promising strategy to improve intracellular delivery and in vivo activity, whereas modifications of cationic proteins should be more delicately designed.

• Journal of Marine Bioscience and Biotechnology
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• v.16 no.1
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• pp.26-35
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• 2024

Chlorella has various biotechnological applications, including in the biomedical and pharmaceutical industries, because of its advantages, including rich nutrients, fast growth rate, easy cultivation, and high biomass. We used the Agrobacterium-mediated transformation method to express human GM-CSF and EGF proteins, which are widely used in regenerative medicine, cosmetics, and pharmaceutical materials in Chlorella. The codon-optimized hGM-CSF and hEGF genes were cloned into plant binary vectors and transformed into Chlorella vulgaris using the Agrobacterium-mediated coculture transformation method. After transformation, genomic DNA PCR was performed for each C. vulgaris line that was stably subcultured on an antibiotic-resistant solid medium to confirm the insertion of hGM-CSF and hEGF into the chromosome. Furthermore, PT-PCR and protein expression of hGM-CSF and hEGF in each transformed C. vulgaris were significantly increased compared to the untransformed Chlorella. This study suggests that high-value proteins, including hGM-CSF and hEGF, which are foreign genes of C. vulgaris, can be stably expressed through the Agrobacterium-mediated Chlorella transformation system.

• Journal of Plant Biotechnology
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• v.2 no.1
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• pp.43-49
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• 2000

An efficient and reliable Agrobacterium transformation procedure based on TDZ (thidiazuron)-induced organogenesis was established and applied to six Brazilian eggp1ant varieties. Optimum transgenic plants recovery was achieved upon the study of the following parameters affecting transformation efficiency, using F-100 variety as a model: i) explant source; ii) pre-culture period; iii) physical state of the pre-culture medium and iv) coculture conditions. The highest frequency of kanamycin-resistant calli derived from leaf explants (5%) was obtained without a pre-culture period and co-cultivation for 24 h in liquid medium followed by five days on solid RM (regeneration medium). For cotyledon explants, best results were achieved upon a pre-culture of 24 h in liquid RM and a co-cultivation period of 24 h in liquid RM followed by three days in solid RM, resulting in a transformation Sequency of 22.7%. Kanamycin-resistant organogenic calli were also obtained from cultivars Emb, Preta Comprida, Round nose Shaded, Campineira and Florida Market. The expression pattern of an epidermis-specific promoter was studied using transformants expressing a chimaeric construct comprised by the promoter Atgrp-5 transcriptionally fused to the coding region of the gus gene. The expression pattern was similar to that previously observed in tobacco and Arabidopsis thaliana, with preferential expression at the epidermis and the stem phloem. These results support the idea that the Atgrp-5 promoter can be used to drive defense genes in these tissues, which are sites of pathogen interaction and spread, in programs for the genetic improvement of eggplant.

• Clinical and Experimental Reproductive Medicine
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• v.43 no.3
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• pp.146-151
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• 2016

Objective: This study was conducted to compare the effects of static culture, dynamic culture, and the combination of dynamic culture with specialized surfaces involving co-culture on human embryonic development. Embryos cultured using conventional static culture (SC) techniques served as a control group. We compared dynamic culture using micro-vibration culture (MVC) and micro-vibration with co-culture (MCoC), in which autologous cumulus cells were used as a specialized surface. Methods: We conducted a chart review of patients who were treated between January 2011 and November 2014 in order to compare embryonic development rates and pregnancy rates among the groups. Zygotes were cultured in micro-droplets, and embryos were subsequently selected for transfer. Some surplus embryos were cryopreserved, and the others were cultured for blastocyst development. A micro-vibrator was set at the frequency of 42 Hz for duration of 5 seconds per 60 minutes to facilitate embryo development. Results: No significant differences among the groups were present in patient's characteristics. However, the clinical pregnancy rates were significantly higher in the MVC group and the MCoC group than in the SC group. No significant differences were found in the blastocyst development rate between the SC group and the MVC group, but the blastocyst development rate in the MCoC group was significantly higher than in the SC and MVC groups. Conclusion: The clinical pregnancy rate was significantly increased by the application of micro-vibration to the embryonic cultures of poor responders. The blastocyst development rate was significantly increased by the application of MCoC to surplus embryos.

• Biomedical Science Letters
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• v.21 no.2
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• pp.122-125
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• 2015

House dust mite (HDM) as a major allergen and damage-associated molecular pattern (DAMP) such as S100A8 and S100A9 trigger the pathogenesis and severity of allergic disease such as asthma. Regulation of neutrophil apoptosis is an important immune response and its dysregulation is involved in pathogenesis of allergic diseases. In this study, we examined the effects of HDM, S100A8 and S100A9 on spontaneous apoptosis of normal neutrophils. We considered the importance of the difference between in vitro and in vivo results and developed a new in vitro system consisting of a combination of immune cells and T helper (Th) cytokines. Extract of Dermatophagoides pteronyssinus (DP), S100A8, and S100A9 inhibited neutrophil apoptosis in culture of neutrophils alone without other leukocytes. DP and S100A8 more strongly suppressed neutrophil apoptosis in combinations of neutrophils, eosinophils, lymphocytes or monocytes than in a culture of neutrophils alone. Anti-apoptotic effect of S100A9 in the mixture of immune cells was similar to that in neutrophils. DP, S100A8, and S100A9 blocked neutrophil apoptosis, regardless of pretreatment with a T helper (Th) 1 cytokine (IFN-$\gamma$), Th2 cytokines (IL-4 and IL-10), a Th9 cytokine (IL-9), a Th17 cytokine (IL-17), a Treg-producing cytokine (TGF-$\beta$). These findings may enable elucidation of allergy pathogenesis due to HDM and DAMP.