• Journal of Microbiology and Biotechnology
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• 제2권1호
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• pp.26-34
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• 1992

The effect of induction temperature on fermentation parameters has been investigated extensively using Escherichia coli M5248[pNKM21], a producer of recombinant human interleukin-2 (rhIL-2). In this recombinant microorganism, the gene expression of rhIL-2 is regulated by the cI857 repressor and $P_L$ promoter system. The recombinant fermentation parameters studied in this work include the cell growth, protein synthesis, cell viability, plasmid stability, $\beta$-lactamase activity, and rhIL-2 productivity. Interrelationships of such fermentation parameters have been analyzed through a quantitative assessment of the experimental data set obtained at eight different culture conditions. While the expression of rhIL-2 gene was repressed at culture temperatures below $34^\circ{C}$ with little effect on other fermentation parameters, under the conditions of rhIL-2 production $>(36~44^\circ{C})$ the cell growth, plasmid stability, and $\beta$-lactamase activity were, as induction temperature was increased, more profoundly reduced. Although the rhIL-2 content in the insoluble protein fraction was maximum at $40^\circ{C}$, total rhIL-2 production in the culture volume was found to be highest at the induction temperature of $36^\circ{C}$. This was in contrast to the previously known optimum induction temperature of the P$_{L}$ promoter system $>(40~42^\circ{C})$.Explanations for such a discrepancy have been proposed based on a product formation kinetics, and their implications have been discussed in detail.l.

• International Journal of Oral Biology
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• 제42권4호
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• pp.203-211
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• 2017

The aim of this study was to evaluate the role of demineralized and particulate autogenous tooth, and interleukin-6 in bone regeneration. A demineralized and particulate autogenous tooth was prepared and human osteoblast-like cells (MG63) and human osteosarcoma cells were inoculated into the culture. The rate of cell adhesion, proliferation and mineralization were examined, and the appearance of cellular attachment was observed. An 8 mm critical size defect was created in the cranium of rabbits. Nine rabbits were divided into three groups including: An experimental group A (3 rabbits), in which a demineralised and particulate autogenous tooth was grafted; an experimental group B (3 rabbits), in which a demineralized, particulate autogenous tooth was grafted in addition to interleukin-6 (20 ng/mL); and a control group. The rabbits were sacrificed at 1, 2, 4 and 6 weeks for histopathological examination with H-E and Masson's Trichrome, and immunohistochemistry with osteocalcin. The cell-based assay showed a higher rate of cell adhesion, mineralization and cellular attachment in the experimental group A compared with the control group. The animal study revealed an increased number of osteoclasts, newly formed and mature bones in the experimental group A compared with the control group. Eventually, a higher number of osteoclasts were observed in the experimental group B. However, the emergence of newly formed and mature bone was lower than in the experimental group A. The current results suggest that treatment with demineralized and particulate autogenous tooth and interleukin-6 is not effective in stimulating bone regeneration during the bone grafting procedure.

• 대한치과교정학회지
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• 제28권1호
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• pp.165-174
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• 1998

교정력이 치아에 가해지면 치주인대의 재생과 치조골의 개조가 일어난다. 치주인대 섬유아세포는 collagenase와 TIMP-1을 분비하여 치주조직의 교원질의 분해와 합성을 담당한다. 본 연구에서는 치주인대 섬유아세포예 기계적 자극과 interleukin-$1{\beta}$를 가해 collagenase와 TIMP-1의 발현을 RT-PCR과 면역조직화학 염색을 사용하여 알아보았다. 4명의 10대 남자 교정환자에게서 아무런 병소가 없는 제1소구치를 발치후 치주인대 섬유아세포를 배양하여 4-6세대의 세포를 사용하였다. 대조군, $Petriperm dish^{\circledR}$ 바닥의 표면적을 $5\%$ 증가시킨 기계적 자극을 가한 군, interleukin-$1{\beta}$를 1.0 ng/ml를 가한 군과 기계적 자극과 interleukin-$1{\beta}$를 같이 가한 군으로 나누어 4명의 환자에서 얻은 세포군을 각 군별로 2, 4, 8시간 후 RT-PCR을 시행하여 그 산물을 반정량하여 대조군에 대한 각 실험군의 상대적인 증감을 나타내었고, 24시간후 면역조직화학 염색을 시행하여 다음과 같은 결과를 얻었다. 1. 광학 현미경으로 세포의 형태를 관찰한 결과 대조군에서는 전형적인 별모양과 길쭉한 모양을 함께 보였으나 기계적 자극과 interleukin-$1{\beta}$를 각각 혹은 동시에 준 군들에서는 별모양의 세포가 사라지고 모양이 더욱 길어졌다. 2. collagenase는 대조군에 비해 기계적 자극과 interleukin-$1{\beta}$를 각각 혹은 동시에 준 군들에서 증가하였고, 실험 8시간 후에서는 interleukin-$1{\beta}$를 준 군, 기계적 자극과 interleukin-$1{\beta}$를 동시에 준 군에서 뚜렷한 증가를 보였다. 3. TIMP-1은 세포 자극 2, 4시간 후에는 대조군에 비해 기계적 자극과 interleukin-$1{\beta}$를 각각 혹은 동시에 준 군들에서 감소하였지만, 실험 8시간 후에서는 증가를 보였다. 4. 면역조직화학 염색을 통해 collagenase와 TIMP-1이 대조군에 비해 기계적 자극과 interleukin-$1{\beta}$를 각각 혹은 동시에 준 군들에서 더욱 강한 염색상을 나타내었다. 본 실험의 결과 섬유아세포는 외부 자극이 가해지면 collagenase와 TIMP-1의 발현 조절을 통해 치주인대 재생과 치조골의 개조에 영향을 미쳐 항상성을 유지하려고 함을 알 수 있었다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제11권2호
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• pp.154-159
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• 2006

Interleukin-18 (IL-18), otherwise known as interferon-gamma-inducing factor (IGIF), is one of several well characterized and important cytokines that contribute to host defenses. The complementary DNA (cDNA) of mature human interleukin-18 gene (hIL-18) was fused with the signal peptide of the rice amylase 1A gene (Ramy1A) and introduced into the plant expression vector under the control of a duplicated CaMV 35S promoter. The recombinant plasmid was transformed into tobacco (Nicotiana tabacum L. cv Havana) using the Agrobacterium-mediated transformation method. The integration of the hlL-18 gene into the genome of transgenic tobacco plants was confirmed by polymerase chain reaction (PCR) amplification and its expression was observed in the suspension cells that were derived from the transgenic plant callus by using Northern blot analysis. The hlL-18 protein was detected in the extracts of the transgenic callus and in the medium of the transgenic tobacco suspension culture by using immunoblot analysis. Based upon enzyme-linked immunosorbant assay (ELISA) results, the expression level of the hlL-18 protein approximated $166{\mu}g/L$ in the suspension culture medium. Bioassay results from the induction of $interferon-{\gamma}$ from a KG-1 cell line indicated that the hlL-18 secreted into the suspension culture medium was bioactive.

• 대한치과교정학회지
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• 제43권6호
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• pp.294-301
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• 2013

Objective: To determine the interleukin (IL)-6 levels in gingival crevicular fluid (GCF) of patients with severe root resorption after orthodontic treatment and investigate the effects of different static compressive forces (CFs) on IL-6 production by human periodontal ligament (hPDL) cells and the influence of IL-6 on osteoclastic activation from human osteoclastic precursor (hOCP) cells in vitro. Methods: IL-6 levels in GCF samples collected from 20 patients (15 and 5 subjects without and with radiographic evidence of severe root resorption, respectively) who had undergone orthodontic treatment were measured by ELISA. The levels of IL-6 mRNA in hPDL cells and IL-6 protein in conditioned medium after the application of different uniform CFs (0, 1.0, 2.0, or 4.0 $g/cm^2$ for up to 72 h) were measured by real-time PCR and ELISA, respectively. Finally, the influence of IL-6 on mature osteoclasts was investigated by using hOCP cells on dentin slices in a pit-formation assay. Results: Clinically, the IL-6 levels were significantly higher in the resorption group than in the control group. In vitro, IL-6 mRNA expression significantly increased with increasing CF. IL-6 protein secretion also increased in a time- and magnitude-dependent manner. Resorbed areas on dentin slices were significantly greater in the recombinant human IL-6-treated group and group cultured in hPDL cell-conditioned medium with CF application (4.0 $g/cm^2$) than in the group cultured in hPDL cell-conditioned medium without CF application. Conclusions: IL-6 may play an important role in inducing or facilitating orthodontically induced inflammatory root resorption.

• International Journal of Industrial Entomology and Biomaterials
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• 제46권2호
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• pp.60-66
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• 2023

Osteoarthritis (OA) is one of the most prevalent degenerative joint diseases and is more common in older and obese individuals. Silkworm male pupae exerts tonic effects by increasing testosterone secretion and the forced swimming time and muscle ratio increased in mice consuming silkworm pupae, which may be beneficial to the older population. Therefore, it will be beneficial to investigate the effects of silkworm pupal extracts (SPE) on OA. To confirm this effect, we prepared SPE in different solvents, and their ability to attenuate matrix metalloproteinases (MMPs) and inflammatory mediators (interleukin-6 [IL-6], interleukin-8 [IL-8] and tumor necrosis factor-α [TNF-α]) were evaluated in an interleukin-1β (IL-1β)-induced SW1353 human chondrosarcoma cell line. 70% ethanolic SPE outperformed the other solvents, reducing MMP-1 and MMP-3 expression by up to 53% and 13%, respectively. Further experiments were performed using 70% ethanolic SPE from three distinct pupation stages in males and females. SPE treatment alleviated MMP-1 expression (43.9-47.4%) regardless of pupation stage and sex. Among the inflammatory mediators, 70% ethanolic SPE alleviated IL-6 and TNF-α levels, and the concentrations thereof were lowest in the early-stage male SPE-treated group (43.15% and 56.74%, respectively). In conclusion, 70% ethanolic SPE may prevent IL-1β-induced osteoarthritis by inhibiting MMPs and inflammatory cytokines. Therefore, SPE is a potential therapeutic agent for the treatment of OA.

• International Journal of Oral Biology
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• 제40권1호
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• pp.11-17
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• 2015

The gingival epithelium of the oral cavity is constantly exposed to exogenous stimuli such as bacterial toxins, allergens, and thermal changes. These exogenous stimuli are resisted by innate host defense in gingival epithelial cells. However, it is unclear exactly how the exogenous stimuli affect detrimentally on the human gingival epithelial cells. Here, we investigated whether the allergen, such as house dust mite (HDM) extract, is linked to $Ca^{2+}$ signaling and proinflammatory cytokine expression in primary cultured human gingival epithelial cells. HDM extract induced an increase in intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) in a dose-dependent manner. Extracellular $Ca^{2+}$ depletion did not affected on the HDM extract-induced increase in $[Ca^{2+}]_i$. The HDM extract-induced increase in $[Ca^{2+}]_i$ was abolished by the treatment with U73122 and 2-APB, which are inhibitors of phospholipase C (PLC) and inositol 1,4,5-trisphosphate ($IP_3$) receptor. Moreover, HDM extract induced the mRNA expression of pro-inflammatory cytokine, interleukin (IL)-8. These results suggest that HDM extract triggers $PLC/IP_3$-dependent $Ca^{2+}$ signaling and IL-8 mRNA expression in primary cultured human gingival epithelial cells.

• 한국임상수의학회지
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• 제17권2호
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• pp.305-314
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• 2000

Recombinant inteileukin-2 (IL-2) is a potent inductive stimulus for nitric oxide synthesis (NO.) and has been demonstrated as an antineoplastic agent in mice and human. But it is not let clear whether NO. can contribute to IL-2-induced therapeutic responses. Therefore, the current experiment was undertaken to clarify the effect of IL-2 on antitumor response against subcutaneous Meth-A tumor in mice. At the beginning of each experiment, normal BALB/c mice were injected subcuta-neously with $5{\times}10^6 Meth-A$ tumor cells. Some mice were implanted with osmotic minipumps con- taining 225 $\mu$l of 3.38 M $N^{\gamma}$ -monomethyl-L-arginine (MLA. an NOS inhibitor). Beginning on day 7, experimental groups were treated with a f-day course of IL-2 (50,000 lU,75,000 nJ,100,0007, 50,000 IU+MLA, 75,000 IU+MLA, 100,000 IU+MLA intraperitoneal injection every 12 hours for 5 days). The result of this experiment revealed that Meth-A tumor grew progressively in control mice. Intraperitoneal IL-2 treatment decreased tumor growth and prolonged survival. compared with con-trol mice. But no significant differences among 50.000 lU.75.000 lU and 100,000 lU of 7-2 treat-ment were observed. MLA administration prevented partially the decrease tumor growth and prolong survival of IL-2 treated mice compared with mice receiving IL-2 alone.

• The Korean Journal of Physiology and Pharmacology
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• 제1권1호
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• pp.71-78
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• 1997

In an attempt to investigate the role of phospholipase $A_2$($PLA_2$) in interleukin-l (IL-l) induced acute lung injury, mepacrine was tried to inhibit $PLA_2$ in IL-l induced ARDS rats. For confirmation of acute lung injury by IL-l, and to know the role of neutrophils in this injury, lung leak index, lung myeloperoxidase(MPO), number of neutrophils and protein content in the bronchoalveolar lavage (BAL) and wet lung weight were measured. At the same time lung $PLA_2$ was measured to know the effect of IL-l on $PLA_2$ activity. Pulmonary surfactant was also measured for an investigation of type II alveolar cell function. Neutrophil adhesion assay was performed to know the effect of $PLA_2$ inhibition in vitro with human umbilical vein endothelial cells (HUVEC). For precise location of injury by IL-l, morpholgical study was performed by electron microscopy. Five hours after instillation of IL-l (50 ng/rat), lung leak index, protein content, number of neutrophils, lung MPO and wet lung weight were increased significantly. Five hours after IL-l instillation lung $PLA_2$ activity was increased significantly, and increased surfactant release was observed in IL-l induced ARDS rats' BAL. In contrast, in rats given mepacrine and IL-l, there was decrease of acute lung injury i.e. decrease of lung leak index, wet lung weight, protein content, number of neutrophils in BAL and decreased lung MPO activity. Mepacrine decreased surfactant release also. Interestingly, inhibition of $PLA_2$ decreased adhesion of human neutrophils to HUVEC in vitro. Morphologically, IL-l caused diffuse necrosis of endothelial cells, type I and II epithelial cells and increased the infiltration of neutrophils in the interstitium of the lung but after mepacrine treatment these pathological findings were lessened. On the basis of these experimental results it is suggested that $PLA_2$ has a major role in the pathogenesis of acute lung injury mediated by neutrophil dependent manner in IL-l induced acute lung injury.

• 동의생리병리학회지
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• 제36권2호
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• pp.79-87
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• 2022

In this study, extracts of Boswellia serrata gum resin, Curcuma longa rhizome, and Terminalia chebula fruit were combined in different ratios, and their anti-osteoarthritis effects were compared to determine which combination had the best synergistic effect. B. serrata, C. longa, and T. chebula extracts in a 2:1:2 ratio exhibited higher antioxidative activity in scavenging DPPH radicals than did the individual extracts alone or the other extract combinations. Additionally, the 2:1:2 combination significantly improved the levels of enzymatic antioxidants and antioxidant-related proteins. Moreover, this same combination ratio decreased the protein levels of matrix metalloproteinase (MMP) 3 and MMP13 in interleukin-1β-stimulated human articular chondrocytes (HCHs) and increased those of aggrecan and collagen type II alpha 1 chain (COL2A1). Analysis of the underlying mechanisms revealed that the 2:1:2 combination significantly inhibited the phosphorylation of nuclear factor kappa B (NF-κB) p65, extracellular regulated protein kinase (ERK), and p38 mitogen-activated protein kinase (MAPK). Therefore, the 2:1:2 combination of these three plant extracts has the best potential for use as an effective dietary supplement for improving joint health compared with the individual extracts and their other combination ratios.