• KSBB Journal
• /
• 제31권4호
• /
• pp.228-236
• /
• 2016

The aim of this study was to characterize the hemolytic Aeromonas sp. MH-8 exposed to green tea catechin, epigallocatechin gallate (EGCG). Initially, the hemolytic Aeromonas sp. MH-8 was enriched and isolated from stale fish. Bactericidal effects of MH-8 exposed to EGCG ranging from 1 mg/mL to 4 mg/mL were monitored, and complete bactericidal effects were achieved within 3 h at 3 mg/mL and higher concentrations. SDS-PAGE with silver staining revealed that the amount of lipopolysaccharides increased or decreased in the strain MH-8 treated to different concentrations and exposing periods of EGCG in exponentially growing cultures. The stress shock proteins (70-kDa DnaK and 60-kDa GroEL), which might contribute to enhancing the cellular resistance to the cytotoxic effect of EGCG, were induced at different concentrations of EGCG exposed to cell culture of MH-8. Scanning electron microscopic analysis demonstrated the presence of irregular rod shapes with umbilicated surfaces for cells treated with EGCG. 2-DE of soluble protein fractions from MH-8 cultures showed 18 protein spots changed by EGCG exposure. These proteins involved in chaperons (e.g., DnaK, GroEL and trigger factor), enterotoxins (e.g., aerolysin and phospholipase C precursor), LPS synthesis (e.g., LPS biosynthesis protein and outer membrane protein A precursor), and various biosynthesis and energy metabolism were identified by peptide mass fingerprinting using MALDI-TOF. In consequence, EGCG was found to have substantial antibacterial effects against food-poisoning causing bacterium, hemolytic Aeromonas sp. MH-8. Also the results provide clues for understanding the mechanism of EGCG-induced stress and cytotoxicity on Aeromonas sp. MH-8.

• Biomolecules & Therapeutics
• /
• 제25권4호
• /
• pp.404-410
• /
• 2017

Benzylideneacetophenone derivative (1E)-1-(4-hydroxy-3-methoxyphenyl) hept-1-en-3-one (JC3) elicited cytotoxic effects on MDA-MB 231 human breast cancer cells-radiation resistant cells (MDA-MB 231-RR), in a dose-dependent manner, with an $IC_{50}$ value of $6{\mu}M$ JC3. JC3-mediated apoptosis was confirmed by increase in sub-G1 cell population. JC3 disrupted the mitochondrial membrane potential, and reduced expression of anti-apoptotic B cell lymphoma-2 protein, whereas it increased expression of pro-apoptotic Bcl-2-associated X protein, leading to the cleavage of caspase-9, caspase-3 and poly (ADP-ribose) polymerase. In addition, JC3 activated mitogen-activated protein kinases, and specific inhibitors of these kinases abrogated the JC3-induced increase in apoptotic bodies. JC3 increased the level of intracellular reactive oxygen species and enhanced oxidative macromolecular damage via lipid peroxidation, protein carbonylation, and DNA strand breakage. Considering these findings, JC3 is an effective therapy against radiation-resistant human breast cancer cells.

• BMB Reports
• /
• 제33권1호
• /
• pp.92-95
• /
• 2000

The HIV-1 p24(202-221) sequence ETINNEEEWDRVHPV HAGP contains a B-cell epitope with the earliest immune response and the highest antibody titer against anti-mouse sera obtained by immunization with p24 antigens. A novel mouse monoclonal antibody (mAb) was generated against the immunodominant B-cell epitope of the HIV-1 p24 capsid protein, p24(202-221). BALB/c mice were immunized with the four branched multiple antigenic peptide (MAP) containing the HIV-1p24(202-221) sequence, and antibody-secreting hybridoma were produced by fusion of mouse splenocytes with P3X63Ag8.653, mouse myeloma cells. One clone which produced the antigen-specific mAb named KI-24 (Isotype IgG1, light chain: ${\kappa}$) was identified. mAb KI-24 was highly specific for both the p24(202-221) and p24 proteins when analyzed by ELISA and Western blotting. Since p24(202-221) also contains a cytotoxic T-lymphocyte epitope, this specfic peptide epitope and the monoclonal antibody with specific reactivity against the p24 protein and p24(202-221) can be used in peptide vaccine development and p24 antigen detection from HIV patients.

• Asian Pacific Journal of Cancer Prevention
• /
• 제17권4호
• /
• pp.2277-2284
• /
• 2016

Ribosome-inactivating protein (RIP) from Mirabilis jalapa L. leaves has cytotoxic effects on breast cancer cell lines but is less toxic towards normal cells. However, it can easily be degraded after administration so it needs to be formulated into nanoparticles to increase its resistance to enzymatic degradation. The objectives of this study were to develop a protein extract of M. jalapa L. leaves (RIP-MJ) incorporated into nanoparticles conjugated with Anti-EpCAM antibodies, and to determine its cytotoxicity and selectivity in the T47D breast cancer cell line. RIP-MJ was extracted from red-flowered M. jalapa L. leaves. Nanoparticles were formulated based on polyelectrolyte complexation using low viscosity chitosan and alginate, then chemically conjugated with anti-EpCAM antibody using EDAC based on carbodiimide reaction. RIP-MJ nanoparticles were characterised for the particle size, polydispersity index, zeta potential, particle morphology, and entrapment efficiency. The cytotoxicity of RIP-MJ nanoparticles against T47D and Vero cells was then determined with MTT assay. The optimal formula of RIP-MJ nanoparticles was obtained at the concentration of RIP-MJ, low viscosity chitosan and alginate respectively 0.05%, 1%, and 0.4% (m/v). RIP-MJ nanoparticles are hexagonal with high entrapment efficiency of 98.6%, average size of 130.7 nm, polydispersity index of 0.380 and zeta potential +26.33 mV. The $IC_{50}$ values of both anti-EpCAM-conjugated and non-conjugated RIP-MJ nanoparticles for T47D cells (13.3 and $14.9{\mu}g/mL$) were lower than for Vero cells (27.8 and $33.6{\mu}g/mL$). The $IC_{50}$ values of conjugated and non-conjugated RIP-MJ for both cells were much lower than $IC_{50}$ values of non-formulated RIP-MJ (>$500{\mu}g/mL$).

• International Journal of Oral Biology
• /
• 제47권1호
• /
• pp.1-8
• /
• 2022

Inflammation is a protective mechanism against pathogens, but if maintained continuously, it destroys tissue structures. Aggregatibacter actinomycetemcomitans is a gram-negative, facultative anaerobic bacterium often found in severe periodontitis. A. actinomycetemcomitans invades epithelial cells and triggers inflammatory response in the immune cells. In this study, we investigated the effect of water-soluble rosehip extract on A. actinomycetemcomitans-induced inflammatory responses. A human monocytic cell line (THP-1) was differentiated to macrophages by phorbol 12-mystristate 13-acetate treatment. The cytotoxic effect of extract was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The effects of extract on bacterial growth were examined by measuring the optical densities using a spectrophotometer. THP-1-derived macrophages were infected A. actinomycetemcomitans after extract treatment, and culture supernatants were analyzed for cytokine production using enzyme-linked immunosorbent assay. Protein expression was measured by western blotting. Extract was not toxic to THP-1-derived macrophages. A. actinomycetemcomitans growth was inhibited by 1% extract. The extract suppressed A. actinomycetemcomitans-induced tumor necrosis factor-α, interleukin (IL)-1β, and IL-8 production. It also decreased mitogen-activated protein kinase (MAP kinase) and nuclear factor-κB (NF-κB) phosphorylation. Moreover, the extract inhibited the expression of inflammasome components, including nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3, Absent in Melanoma 2, and apoptosis associated speck-like protein containing a CARD. And cysteine-aspartic proteases-1 and IL-1β expression were decreased by the extract. In summary, extract suppressed A. actinomycetemcomitans growth and decreased inflammatory cytokine production by inhibiting activation of MAP kinase, NF-κB, and inflammasome signaling. Rosehip extract could be effective in the treatment of periodontal inflammation induced by A. actinomycetemcomitans infection.

• Oncology Letters
• /
• 제18권3호
• /
• pp.3256-3264
• /
• 2019

The induction of apoptosis is a useful strategy in anti-cancer research. Various Moon Hyung Yang (MHY) compounds have been developed as novel anti-cancer drug candidates; in the present study, the pro-apoptotic effects of (Z)-5-(3-ethoxy-4-hydroxybenzylidene)-2-thioxothiazolidin-4-one (MHY695) on HCT116 human colon cancer cells were assessed. MTT assays were performed to investigate the dose-dependent cytotoxic effects of MHY695 on HCT116 cells. Immunofluorescence staining and flow cytometry analyses were performed to identify apoptotic cell death, and western blot analysis was used to investigate the apoptotic-signaling pathways. A mouse xenograft model was also used to determine the effects of MHY695 in vivo. MHY695 decreased the viability of HCT116 cells and induced apoptotic cytotoxicity. The apoptotic mechanisms induced by MHY695 involved the dephosphorylation of Bcl-2-associated agonist of cell death protein following protein kinase B inactivation, induced myeloid leukaemia cell differentiation protein and BH3-interacting domain death agonist truncation, caspase-3 and -9 activation and poly (ADP-ribose) polymerase cleavage. In addition, MHY695 significantly suppressed tumor growth in the mouse xenograft model, compared with the vehicle control. Notably, MHY695 exhibited potent anti-cancer effects in four different types of human colon cancer cell line, including Caco-2, DLD-1, HT-29 and HCT116. Additionally, MHY695 showed reduced cytotoxicity in NCM460, normal colonic epithelial cells. Furthermore, MHY-induced cytotoxicity in colon cancer cells was independent of the tumor suppressor protein p53. Collectively, these observations suggested that MHY695 may be a novel drug for the treatment of colon cancer.

• 동의생리병리학회지
• /
• 제18권3호
• /
• pp.830-836
• /
• 2004

The unique biochemical pathways in melanocytes responsible for melanogenesis provide us with a rational mechanism-based means for developing both pharmacological regulators of pigmentation and cytotoxic chemotherapeutic drugs for melanoma, Myricetin is a polyphenolic antioxidant and a component from Biotae Semen, Biotae orientalis Folium. Therefore, the present study was conducted to evaluate the effects of myricetin on the melanogenesis in human melanoma (HM₃KO) cells. The cells were treated for 5 days with myricetin at several concentrations (10 - 100 μg/ml). Treatment with myricetin dose-dependently suppressed cell growth in HM₃KO cells, But melanin formation was markedly increased by myricetin as a dose dependent manner. Myricetin did not affect to tyrosinase activity, which is a key enzyme for melanogenesis, but significantly increased the level of tyrosinase protein expression, These results suggest that myricetin stimulate melanin synthesis of human melanoma cells through the modification of tyrosinase protein expression.

• 동의생리병리학회지
• /
• 제24권5호
• /
• pp.822-826
• /
• 2010

In order to validate the use of Faeces Trogopterori as an anti-inflammatory drug in the traditional Korean medicine, I have investigated the effect of water-soluble extract of F. Trogopterori (EFT) on the production of monocyte chemoattractant protein-1 (MCP-1), of which chemokine stimulates the migration of mononuclear cells, in human umbilical vein endothelial cells (HUVECs) stimulated with tumor necrosis factor-alpha. The extract inhibited dose-dependently MCP-1 production without its cytotoxic effect on HUVECs, as measured by enzyme-linked immunosorbent assay, and significantly decreased mRNA levels of MCP-1, as determined using reverse transcription polymerase chain reaction. These results suggest that F. Trogopterori may have therapeutic potential in the control of endothelial disorders caused by inflammation.

• BMB Reports
• /
• 제48권2호
• /
• pp.121-126
• /
• 2015

Here we report a new chemical inhibitor against HIV-1 with a novel structure and mode of action. The inhibitor, designated as A1836, inhibited HIV-1 replication and virus production with a 50% inhibitory concentration ($IC_{50}$) of $2.0{\mu}M$ in an MT-4 cell-based and cytopathic protection antiviral assay, while its 50% cytotoxic concentration ($CC_{50}$) was much higher than $50{\mu}M$. Examination of the effect of A1836 on in vitro HIV-1 reverse transcriptase (RT) and integrase showed that neither were molecular targets of A1836. The characterization and re-infection assay of the HIV-1 virions generated in the presence of A1836 showed that the synthesis of early RT products in the cells infected with the virions was inhibited dose-dependently, due in part to abnormal protein formation within the virions, thus resulting in an impaired infectivity. These results suggest that A1836 might be a novel candidate for the development of a new type of HIV-1 inhibitor.

• 대한본초학회지
• /
• 제22권1호
• /
• pp.111-117
• /
• 2007

Objectives : The purpose of this study was to investigate the anti-inflammatory effects of extract from Pulsatilla koreana $N_{AKAI}$ (PK) on the peritoneal macrophage. Methods : To evaluate of anti-inflammatory of PK, we examined cytokines and NO production in lipopolysacchride (LPS)-induced macrophages. Furthermore, we examined molecular mechanism using western blot. Results : 1.Extract from PK reduced LPS-induced NO, tumor necrosis factor-a ($TNF-{\alpha}$), interleukin (IL)-6 and IL-12 production in peritoneal macrophages. 2.Extract from PK itself does not have any cytotoxic effect. PK inhibited the activation of extracelluar signal-regulated kinase(ERK 1/2) but not another mitogen-activated protein kinases (MAPKs) such as p38, c-Jun NH2-terminal kinase (JNK) and the degradation of inhibitory kappa B a ($I_{k}B_{a}$) does not any effect in the LPS-stimulated peritoneal macrophages. Conclusion : PK down-regulated LPS-induced NO and cytokines production, which may be provide a clinical basis for anti-inflammatory properties of PK.