• Journal of Microbiology and Biotechnology
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• v.31 no.8
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• pp.1175-1182
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• 2021

We investigated the effect of bovine lactoferricin (Lfcin-B), a peptide derived from bovine lactoferrin, on activation of intestinal epithelial cells in IEC-6 intestinal cell, and protection against in vivo rotavirus (RV) infection. Treatment with Lfcin-B significantly enhanced the growth of IEC-6 cells and increased their capacity for attachment and spreading in culture plates. Also, Lfcin-B synergistically augmented the binding of IEC-6 cells to laminin, a component of the extracellular matrix (ECM). In the analysis of the intracellular mechanism related to Lfcin-B-induced activation of IEC-6 cells, this peptide upregulated tyrosine-dependent phosphorylation of focal adhesion kinase (FAK) and paxillin, which are intracellular proteins associated with cell adhesion, spreading, and signal transduction during cell activation. An experiment using synthetic peptides with various sequences of amino acids revealed that a sequence of 9 amino acids (FKCRRWQWR) corresponding to 17-25 of the N-terminus of Lfcin-B is responsible for the epithelial cell activation. In an in vivo experiment, treatment with Lfcin-B one day before RV infection effectively prevented RV-induced diarrhea and significantly reduced RV titers in the bowels of infected mice. These results suggest that Lfcin-B plays meaningful roles in the maintenance and repair of intestinal mucosal tissues, as well as in protecting against intestinal infection by RV. Collectively, Lfcin-B is a promising candidate with potential applications in drugs or functional foods beneficial for intestinal health and mucosal immunity.

• Journal of Periodontal and Implant Science
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• v.23 no.1
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• pp.37-47
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• 1993

The native connective tissue attachment of the periodontium is known to be a complex consisting of gingival fibroblasts, periodontal ligament cells, gingival epithelial cells, cementum, alveolar bone and extensive extracellular matrix (collagen, glycoprotein and proteoglycans). The purpose of this study was to evaluate the effects of natural extracts on DNA, collagen and protein synthesis and inhibition of cytokine production in the gingival and periodontal ligament fibroblasts and gingival epithelial cells. Healthy gingival tissue was obtained from orthodontic treatment patients, and gingival epithelial cells, gingival fibroblasts and periodontal ligament cells were isolated and cultured from the samples. After treated with Ginseng protein, Pluronic F-68, Scutellariae Radix, centella asiatica, PDGF, IGF, DNA synthesis, total protein and collagen synthesis, and cytokine production of gingival epithelial cell, gingival fibroblast and periodontal ligamentcells were measured. MTT method for DNA synthesis, Peterkofsky and Dingerman method for total protein and collagen synthesis, and IL-1 ELISA kit for cytokine production were used. The proliferation of epithelial cells was enhanced in Centella asiatica, Ginseng protein, Pluronic F-68 and Scutellariae Radix. The activities of PDL cells were increased in PDGF, IGF, and Pluronic F-68. Higher collagen synthesis was observed in Scutellariae Radix and total protein synthesis was increased in Scutellariae Radix and PDGF. The inhibitory effects on IL-1, IL-6, $TNF-{\alpha}$ were observed in all exrracts.

• BMB Reports
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• v.55 no.8
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• pp.370-379
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• 2022

Human pregnancy is a delicate and complex process where multiorgan interactions between two independent systems, the mother, and her fetus, maintain pregnancy. Intercellular interactions that can define homeostasis at the various cellular level between the two systems allow uninterrupted fetal growth and development until delivery. Interactions are needed for tissue remodeling during pregnancy at both fetal and maternal tissue layers. One of the mechanisms that help tissue remodeling is via cellular transitions where epithelial cells undergo a cyclic transition from epithelial to mesenchymal (EMT) and back from mesenchymal to epithelial (MET). Two major pregnancy-associated tissue systems that use EMT, and MET are the fetal membrane (amniochorion) amnion epithelial layer and cervical epithelial cells and will be reviewed here. EMT is often associated with localized inflammation, and it is a well-balanced process to facilitate tissue remodeling. Cyclic transition processes are important because a terminal state or the static state of EMT can cause accumulation of proinflammatory mesenchymal cells in the matrix regions of these tissues and increase localized inflammation that can cause tissue damage. Interactions that determine homeostasis are often controlled by both endocrine and paracrine mediators. Pregnancy maintenance hormone progesterone and its receptors are critical for maintaining the balance between EMT and MET. Increased intrauterine oxidative stress at term can force a static (terminal) EMT and increase inflammation that are physiologic processes that destabilize homeostasis that maintain pregnancy to promote labor and delivery of the fetus. However, conditions that can produce an untimely increase in EMT and inflammation can be pathologic. These tissue damages are often associated with adverse pregnancy complications such as preterm prelabor rupture of the membranes (pPROM) and spontaneous preterm birth (PTB). Therefore, an understanding of the biomolecular processes that maintain cyclic EMT-MET is critical to reducing the risk of pPROM and PTB. Extracellular vesicles (exosomes of 40-160 nm) that can carry various cargo are involved in cellular transitions as paracrine mediators. Exosomes can carry a variety of biomolecules as cargo. Studies specifically using exosomes from cells undergone EMT can carry a pro-inflammatory cargo and in a paracrine fashion can modify the neighboring tissue environment to cause enhancement of uterine inflammation.

• Archives of Pharmacal Research
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• v.25 no.6
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• pp.903-909
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• 2002

Research in the cytokine field has grown exponentially in recent years, and the validity of such studies relies heavily on the appropriate measurement of levels of cytokines in various biological samples. Transforming growth factor (TGF)-$\beta$, a hormonally active polypeptide found in normal and transformed tissue, is a potent regulator of cell growth and differentiation. The most widely used bioassay for TGF-$\beta$ is the inhibition of the proliferation of mink lung epithelial cells. Though detection of [$^3$H]thymidine incorporation is more sensitive than the MTT assay, it presents some disadvantages due to the safety and disposal problems associated with radioisotopes. In this study, we attempted to ascertain the experimental conditions which could be used for measuring the in vitro biological activity of TGF-$\beta$ in a safer and more sensitive way compared with the currently available methods. We compared the commonly used method, the MTT assay, to the XTT assay using different parameters including cell number, incubation time and the wave length used for detecting the product. We examined the anti-proliferative activities of TGF-$\beta$ in three different cell lines: Mv-1-Lu mink lung epithelial cells, MCF10A human breast epithelial cells and H-ras-transformed MCF10A cells. Herein, we present an experimental protocol which provides the most sensitive method of quantifying the biological activity of TGF-$\beta$, with a detection limit of as low as 10 pg/ml: Mv-1-Lu or H-ras MCF10A cells ($1{\times}10^5/well$) were incubated with TGF-$\beta$ at $37^{\circ}C$ in a humidified $CO_2$ incubator for 24 hr followed by XTT treatment and determination of absorbance at 450 or 490 nm. Our results may contribute to the establishment of an in vitro bioassay system, which could be used for the satisfactory quantitation of TGF-$\beta$.

• Journal of Periodontal and Implant Science
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• v.30 no.3
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• pp.527-538
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• 2000

Several extraction cases with advanced bone loss as a result of periodontal disease, root or labial bone fracture, extensive caries, and periapical lesions occur esthetic, functional problems and severe bone loss. Therefore, to treat these cases used several surgical methods and socket preservation among this therapies have been evaluated simple, effective and good prognosis in the implant placement. Socket preservation therapy have been used with barrier membranes or/and graft materials. Deproteinized bovine bone mineral have been evaluated ideal grafting materials. Recently, calcium-phosphate thin film coated bovine bone powders were developed in our country, but the study for these material wasn't reported. When two types of xenograft materials were implanted in extraction sockets of Beagle dogs, the effects of these were analyzed after 4 weeks and 8 weeks histological views. The results of this study were as follows. 1. In control groups, 4 weeks after implantation, the extraction sockets were filled with connective tissue which has dilated vessels and epithelial growth. And after 8 weeks, irregular connective bundles were observed. But new bone formation was not seen. 2. In Bio-Oss groups, epithelial growth was not seen and bone powder was covered with connective tissue fiber. New bone formation was found around the interproximal bone. There was no special change seen after 8 weeks, connective tissue fibers became more regular, and bone growth near bone powder was not made well. 3. In Ca-P BBP groups, epithelial cells didn't grow in the extraction sockets, there was a lot of new bone made around the bone powder after 8 weeks, new bone around bone powder was replaced with mature bone. It is thought that bone powder grafting into the extraction sockets is very useful for conservation of ridge, and Ca-P BBP is more effective in bone formation than Bio-Oss.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.3
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• pp.781-784
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• 2012

The aim of this study was to evaluate the effect of the adenovirus-mediated double suicide gene (CD/TK) for selective killing of gastric cancer cells. Gastric cancer cells SCG7901 and normal gastric epithelial cell lines were infected by adenoviruses Ad-survivin/GFP and Ad-survivin/CD/TK. GFP expression and CD-TK were detected by fluorescence microscopy and reverse transcriptase polymerase chain reaction (RT-PCR), respectively. After treatment of the infected cells with the pro-drugs ganciclovir (GCV) and/or 5-FC, the cell growth status was evaluated by methyl thiazolyl tetrazolium assay. Cell cycle changes were detected using flow cytometry. In nude mice bearing human gastric cancer, the recombinant adenovirus vector was injected directly into the tumor followed by an intraperitoneal injection of GCV and/or 5-FC. The subsequent tumor growth was then observed. The GFP gene driven by survivin could be expressed within the gastric cancer line SCG7901, but not in normal gastric epithelial cells. RT-PCR demonstrated the presence of the CD/TK gene product in the infected SCG7901 cells, but not in the infected normal gastric epithelial cells. The infected gastric cancer SCG7901, but not the gastric cells, was highly sensitive to the pro-drugs. The CD/TK fusion gene system showed significantly greater efficiency than either of the single suicide genes in killing the target cells (P<0.01). Treatment of the infected cells with the pro-drugs resulted in increased cell percentage in G0-Gl phase and decreased percentage in S phase. In nude mice bearing SCG7901 cells, treatment with the double suicide gene system significantly inhibited tumor growth, showing much stronger effects than either of the single suicide genes (P<0.01). The adenovirus-mediated CD/TK double suicide gene driven by survivin promoter combined with GCV an 5-FC treatment could be an effective therapy against experimental gastric cancer with much greater efficacy than the single suicide gene CD/TK combined with GCV or 5-FC.

• Tuberculosis and Respiratory Diseases
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• v.82 no.2
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• pp.133-142
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• 2019

Background: Idiopathic pulmonary fibrosis involves irreversible alveolar destruction. Although alveolar epithelial type II cells are key functional participants within the lung parenchyma, how epithelial cells are affected upon bleomycin (BLM) exposure remains unknown. In this study, we determined whether BLM could induce cell cycle arrest via regulation of Schlafen (SLFN) family genes, a group of cell cycle regulators known to mediate growth-inhibitory responses and apoptosis in alveolar epithelial type II cells. Methods: Mouse AE II cell line MLE-12 were exposed to $1-10{\mu}g/mL$ BLM and $0.01-100{\mu}M$ baicalein (Bai), a G1/G2 cell cycle inhibitor, for 24 hours. Cell viability and levels of pro-inflammatory cytokines were analyzed by MTT and enzyme-linked immunosorbent assay, respectively. Apoptosis-related gene expression was evaluated by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Cellular morphology was determined after DAPI and Hoechst 33258 staining. To verify cell cycle arrest, propidium iodide (PI) staining was performed for MLE-12 after exposure to BLM. Results: BLM decreased the proliferation of MLE-12 cells. However, it significantly increased expression levels of interleukin 6, tumor necrosis factor ${\alpha}$, and transforming growth factor ${\beta}1$. Based on Hoechst 33258 staining, BLM induced condensation of nuclear and fragmentation. Based on DAPI and PI staining, BLM significantly increased the size of nuclei and induced G2/M phase cell cycle arrest. Results of qRT-PCR analysis revealed that BLM increased mRNA levels of BAX but decreased those of Bcl2. In addition, BLM/Bai increased mRNA levels of p53, p21, SLFN1, 2, 4 of Schlafen family. Conclusion: BLM exposure affects pulmonary epithelial type II cells, resulting in decreased proliferation possibly through apoptotic and cell cycle arrest associated signaling.

• Animal cells and systems
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• v.2 no.1
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• pp.1-8
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• 1998

Hepatocyte growth factor (HGF), originally discovered and cloned as a powerful mitogen for hepatocytes, is a four kringle-containing growth factor which specifically binds to membrane-spanning tyrosine kinase, c-Met/HGF receptor. HGF has mitogenic, motogenic (enhancement of cell movement), morphogenic (e.g., induction of branching tubulogenesis), and anti-apoptotic activities for a wide variety of cells. During embryogenesis, HGF supports organogenesis and morphogenesis of various tissues, including liver, kidney, lung, gut, mammary gland, and tooth. In adult tissues HGF elicits an organotrophic function which supports regeneration of organs such as liver, kidney, lung, and vascular tissues. HGF is also a novel member of neurotrophic factor in nervous systems. Together with the preferential expression of HGF in mesenchymal or stromal cells, and c-Met/HGF receptor In epithelial or endothelial cells, the HGF-Met coupling seems to orchestrate dynamic morphogenic processes through epithelial-mesenchymal (or-stromal) interactions for organogenesis and organ regeneration. HGF or HGF gene may well become unique therapeutic tools for treatment of patients with various organ failure, through its actions to reconstruct organized tissue architectures. This review focuses on recently characterized biological and physiological functions integrated by HGF-Met coupling during organogenesis and organ regeneration.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.17 no.4
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• pp.331-336
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• 1995

To use cultured mucosa as a graft of full thickness, our laboratory has been involved in the development of techniques to grow epidermis together with connective tissue. Human oral mucosa was obtained at dental surgery. Under sterile conditions the tissues were cut into explants of 0.1 $cm^2$ which were placed in the center of 24 well tissue culture dishes and incubated in a growth medium. The growth medium used for epithelial was MEM(Minimum Essential Medium) supplemented with 10% fetal calf serum, 0.5% dimethyl sulfoxide, glutamine (0.292 g/l), epidermal growth factor (40 ug/ml), cholera toxin (30 ng/ml), hydrocortisone (2 ug/ml), insulin (40 ug/ml) and transferin (5 ug/ml). The medium for stratification of epithelial cells was MEM supplemented with 10% fetal calf serum, 0.5% dimethyl sulfoxide and glutamine (0.292 g/l). The medium used for fibroblasts was MEM supplemented with 10% fetal calf serum. With the three types of media used alternatively, a mucosa composed of epidermis and connective tissue was obtained after 3 weeks of culture.

• Applied Microscopy
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• v.45 no.2
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• pp.80-88
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• 2015

The structural change and distribution of mitochondrial enzyme (ATPase, cytochrome-c-oxidase), cell proliferation (proliferating cell nuclear antigen, PCNA), cell death (caspase-3) and cell growth factor (fibroblast growth factor 8, FGF-8) in the Sprague-Dawley rat bile duct during Clonorchis sinensis infection was investigated. Experimental groups were divided into C. sinensis infection, superinfection and reinfection of C. sinensis after 'praziquantel' treatment group. As a result, C. sinensis infected rat bile ducts showed the features of chronic clonorchiasis, i.e., connective tissue thickening, ductal fibrosis and epithelial tissue dilatation. PCNA for cell proliferation increased in the infection group, and decreased after praziquantel treatment. Caspase-3 was distributed in reinfection group only. FGF-8 was distributed in the rat bile duct after praziquantel treatment but not distributed in infection and reinfection group. Overall, C. sinensis infection causes physical and chemical irritations and then brings on the abnormalities of intracellular energy metabolism and cellular growth factors, which hinders bile duct tissue from functioning properly, and resultingly, fibrosis occurs and epithelial cells dilated abnormally. More intense infection makes tissue fibrosis chronical and activates apoptosis factors.