Uncoupling protein 1 (UCP1) is a unique mitochondrial membranous protein expressed in brown adipose tissue (BAT) in mammals. While its expression in response to cold temperatures and adipogenic inducers is well-characterized in mammals and human infants, the molecular characterization and expression of UCP1 in fish remain unexplored. To address this gap, we analyzed UCP1 expression in response to adipogenic inducers in a fish cell line, rainbow trout gonadal cells (RTG-2), and compared it with UCP1 expression in three mammalian preadipocytes, 3T3-L1, T37i, and WT1 exposed to the Peroxisome proliferator-activated receptor gamma (PPARγ) agonists, rosiglitazone (Rosi). In mammalian preadipocytes, UCP1 protein was highly expressed by Rosi, with an induction of adipogenesis observed in a time-dependent manner. This suggests that UCP1 plays a significant role in adipogenesis in mammals. However, RTG-2 cells showed no response to adipogenic inducers and exhibited only marginal expressions of UCP1. These results imply that RTG-2 cells may lack crucial responsive mechanisms to adipogenic signals or that the adipogenic response is regulated by other mechanisms. Further studies are needed to confirm these phenomena in fish preadipocytes when an appropriate cell line is established in future research.

We investigated the effect of a synthetic complement peptide C3a on the outcome of Brucella abortus 544 infection in a murine macrophage cell line RAW264.7 cell. First, we determined the highest non-cytotoxic concentration of the peptide in the cell line. We also found that the peptide significantly increased the growth of the bacteria at 8 and 24 h. Although the number of bacterial CFU was also elevated at 48 and 72 h, the increases were not significant as compared to controls. We further investigated the effect of C3a peptide on the growth of Brucella by pre-incubating the peptide at various temperatures and found that the effect was reversed at 24 h post-incubation suggesting that incubation of peptide at high temperatures including 65°C or 95°C could inactivate its action. This also could indicate the beneficial effect of high temperature during infection. Although several studies reported the inhibitory effect of different antimicrobial peptides including C3a, the present study preliminarily revealed that it had no positive contribution on the control of B. abortus 544 infection in vitro and indirectly to its receptor, CD88, which belongs to GPCR. Moreover, the encouraged further exploration of the effect of other similar peptides would be performed for the purpose of finding Brucella-host cell interaction for the control of disease progression.

Spirodelae Herba (SH) and Perilla Frutescens (PF) extracts have been widely used in clinical practice with various disorders for thousands of years. There are some reports regarding the anticancer effects of SH and PF each by each, but their mixture have not been investigated and their mechanisms also have not been clear. The purpose of the present study was to investigate the anticancer mechanisms and their effects of the mixture of SH and PF extracts on head and neck cancer cell line. Head and neck carcinoma KB cells were treated with SH, PF and their mixture. Anticancer effects were investigated by searching cancer cell death pathway; apoptosis and autophagy, which have been regarded to be effective and safe methods. Apoptosis, which is termed a programmed cell death, was observed by TUNNEL assay. Autophagy, which is termed a type II programmed cell death, was observed by acridine orange red staining. Additionally, the protein expressions associated with apoptosis and autophagy were detected for their mechanism by western blots. The mixture of SH and PF extracts induced autophagic and apoptotic cell death simultaneously in cancer cells. And 0.4 mg/ml of the mixture with SH and PF extracts down-regulated the expression of mTOR, however, the expressions of ATG5 and LC3-II, which induced autophagy, up-regulated. The mixture of SH and PF extracts also down-regulated the expressions of Bcl-2, but up-regulate the expressions of PARP-1 cleavage, Caspase-9 cleavage, Caspase-3 cleavage and BAX, which induced apoptosis. Taken together, these results suggested that the mixture of SH and PF extracts induce autophagic and apoptotic cell death simultaneously in head and neck cancer cells and it could be used as an alternative for anti-cancer drugs.

Salivary gland adenocarcinoma(AdCa NOS) is one of the major causes of mortality among malignant salivary gland tumors. New therapeutic measure are needed to improve the outcome for patients with AdCa NOS because current therapy does not significantly improve survival rates. Transglutaminase 2(TGase 2) was implicated in forming cross-linked protein polymer, apoptosis and matrix interaction. And also TGase 2 expression is up-regulated in proliferation, migration, invasion, and metastasis of cancer cells. shRNA which has emerged as an effective method to target specific genes for silencing has provided new opportunities for cancer therapy. But there has been rarely reported using shRNA-TGase 2 transfection in AdCa NOS. The purpose of this study were to examine the specific inhibition of TGase 2 mRNA and protein expression by siRNA transfection of TGase 2 through RT-PCR and immunoslot blotting, and to study proliferation, migration and invasion assay of SGT cell line from AdCa NOS. Cell cycle analysis showed that the downregulation of shRNA-TGase 2 caused the accumulation of cells in the sub-G0/G1 phase. In migration assay, suppressing shRNA-TGase 2 inhibited the capacity of the cells to migrate compared to parental cells. In invasion assay, cells transfected with shRNA-TGase 2 decreased in invasion when compared to SGT and vector transfected cells. shRNA-TGase 2 expressing plasmids efficiently downregulated TGase 2 mRNA and TGase 2 protein expression. It suggested that the shRNA-TGase 2 targeting system against TGase 2 could have a therapeutic potentiality for malignant salivary gland tumors, especially in inhibiting and/or preventing cancer cell proliferation, migration and invasion.

Despite existing chemotherapy and surgical resection strategies, salivary gland adenocarcinoma(AdCa NOS) is one of the major causes of mortality among malignant salivary gland tumors. New therapeutic measure are needed to improve the outcome for patients with AdCa. Overexpression of urokinase-type plasminogen activator receptor/urokinase-type plasminogen activator(uPAR-uPA) has been implicated in progression and metastasis of oral cancer. RNA interference(RNAi) which has emerged as an effective method to target specific genes for silencing has provided new opportunities for cancer therapy. But there has been rarely reported using RNAi-uPAR/uPA transfection in salivary gland AdCa. The purpose of this study were to examine the specific inhibition of uPAR/uPA mRNA and protein expression by RNAi transfection of uPAR/uPA through RT-PCR and Immunoslot blot, and to study tumor cell proliferation activity, adhesion, invasion and migration of SGT cell line in vitro compared to the controls. In adhesion assay, cells transfected with RNAi-uPAR/uPA inhibited markedly adhesion to vitronectin compared to parental cells. Angiogenic assays revealed a significant decrease in the angiogenic potential of SGT cells downregulated by both uPAR and uPA. In migration assay, suppressing uPAR and uPA inhibited the capacity of the cells to migrate compared to parental cells. In invasion assay, cells transfected with RNAi-uPAR/uPA showed the maximum decrease in invasion when compared to all other treatment conditions. RNAi expressing plasmids efficiently downregulated mRNA and protein expression of uPAR and uPA. Cell cycle analysis showed that the simultaneous downregulation of uPAR and uPA caused the accumulation of cells in the sub-G0/G1 phase in SGT cells. Immunoslot blot analysis revealed that downregulation of uPAR and uPA caused the prominent activation of caspase 8. It suggested that the RNAi targeting of the uPAR/uPA system could have a therapeutic potentiality for malignant salivary gland tumors.

Autophagy is recently receiving the spotlight as the development strategy for promising anticancer drugs. In particular, the majority of anticancer drugs originating from natural products are known to induce autophagy. Saururus chinensis has been used for treating various inflammatory diseases. Recent research has revealed that the extract of Saururus chinensis possess cytotoxicity for various types of human cancer cells. However, the exact action mechanism of Saururus chinensis extract for oral squamous cell carcinoma (OSCC) has not been studied yet. Therefore, the authors of this research aim to study the effect of methanol extract of S. chinensis (MESC) on OSCC cells. To observe the cell proliferation inhibitory effect of MESC on HSC3 cells, the authors conducted the trypan blue exclusion assay. Also, the action mechanism of MESC was studied by conducting the cell cycle analysis, acidic vesicular organelle (AVO) staining and flow cytometry analysis, monodansylcadaverine (MDC) staining, propidium iodide staining, and Western blotting on MESC-treated HSC3 cells. When HSC3 cells were treated in MESC, the cell proliferation was suppressed in time-dependent and dose-dependent manners. Also, the number of sub-G1 arrested cells increased in a dose-dependent manner. MDC punctate and AVO puncta significantly increased respectively. Western blot analysis demonstrated the expression of autophagy-related proteins increased, but apoptotic proteins were not observed. Also, the pAkt protein was reduced, while the p-p38 protein and pERK protein increased. According to our results, MESC induced autophagy and accompanied changes in the cell cycle in HSC3 cells. Also, the alteration in Akt, ERK, and p38 pathways were confirmed. This result suggested the possibility of MESC as the new promising adjuvant for treating OSCC patients.

국내에서 오래 전부터 오미자와 칠해목은 약용식물로 이용해져 왔다. 오미자의 경우 생리활성 물질인 lignan을 통하여 여러가지 효능들이 연구를 통하여 알려졌으나, 칠해목의 경우 항염증과 관련된 연구가 진행되었을 뿐, 산화 스트레스와 관련된 연구는 미비한 실정이었다. 이에 본 연구에서는 오미자․칠해목의 추출혼합물을 이용하여, 과산화수소로 산화 스트레스가 유도된 SH-SY5Y 신경세포에서의 보호 효과를 알아보고자 하였다. 과산화수소의 처리 농도의 경우, 신경세포 독성 실험을 통하여 100 μM의 농도를 본 연구에 사용하였다. 또한, 오미자와 칠해목 추출물은 SH-SY5Y 신경세포에서 세포 독성이 없음을 실험을 통해 확인하였으며, 과산화수소를 이용한 신경세포 보호효과를 확인한 결과, 30% 에탄올 추출물 50 μg/mL의 농도에서 각각의 추출물에서 가장 높은 보호 효과를 확인할 수 있었다. 이에 오미자․칠해목 추출혼합물의 최적 효능 비율을 알아보기 위해서 각각의 추출물을 다양한 수율로써 6:4, 7:3, 8:2의 비율로 신경세포 보호 활성을 확인한 결과, 오미자와 칠해목 7:3의 비율과 50 μg/mL의 농도에서 가장 높은 신경세포 보호효과가 나왔으며, 이에 본 연구에서 오미자․칠해목 추출혼합물의 최적의 비율과 처리 농도로 사용하였다. Annexin V와 PI를 이용한 SH-SY5Y 신경 세포의 세포사멸을 확인해 본 결과에서도 오지마․칠해목 추출혼합물은 SH-SY5Y 신경세포의 세포사멸를 억제시키는 것을 확인할 수 있었다. 이에 본 연구에서는 오미자․칠해목 추출혼합물이 과산화수소로 유도된 산화적 스트레스에서 SH- SY5Y 신경세포에서 보호 효과가 있다는 것을 확인할 수 있었지만, 그에 대한 메카니즘에 대한 연구는 미비한 실정이다. 이에 차후 연구에서는 오미자․칠해목 추출혼합물의 산화 스트레스에 대한 정확한 기전을 파악하기 위하여 In vitro와 In-vivo에서의 후속적 연구가 필요하다고 사료된다.

Natural products are vastly utilized as a source of chemotherapeutic agents for human cancers. Kochia scopraia is traditionally used for the cure of urological and dermatological diseases. Recently, methanol extract of Kochia scoparia (MEKS) has been shown to have anti-cancer activity to various human cancers. However, there is no report demonstrating the anti-cancer activity of MEKS in human mucoepidermoid carcinoma (MEC) cells. In this study, the authors studied the effects of MEKS on the cell proliferation and underlying mechanism in YD15 human MEC cells. MEKS decreased YD15 cell proliferation proven by trypan blue exclusion assay and induced apoptosis, evidenced by cell cycle analysis and western blotting. Autophagy induction by MEKS was verified by western blotting. In addition, MEKS regulated the expression of phosphorylated Akt, phosphorylated p38 and Nrf2 protein. This results can imply that MEKS might be a potential candidate for the treatment of human MEC cells.

This study was carried out to identify medicinal mushrooms with protective effects against oxidative stress in PC12 neuronal cell line, followed by evaluation of their antioxidant property. Extracts of medicinal mushrooms, including Ganoderma lucidum extract (GLE), antler-shaped Ganoderma lingzhi extract (AGLE), Hericium erinaceus extract (HEE), and Sanghuangporus baumii extract (SBE), were screened for cytotoxicity using MTT assay. None of the extracts up to 10 μg/ml concentration affected cell viability. These extracts were further checked for their protective effect against oxidative stress-induced reactive oxygen species (ROS) production. Exposure to 50 μM H₂O₂ induced ROS generation in PC12 cells, which was inhibited only by treatment with AGLE. In addition, inhibition of H₂O₂-induced ROS generation by AGLE was found to be in a dose-dependent manner (2.5, 5, and 10 μg/ml). Microscopic examination of DCF fluorescence for detection of ROS showed a similar pattern. Further, antioxidant activity of AGLE was determined by ABTS radical cation assay, and its IC50 was found to be 46.90±0.31 μg/ml. Taken together, these results suggest that AGLE may help to alleviate oxidative stress in PC12 neuronal cells.

5-fluorouracil (5-FU) is a pyrimidine analog which can work as antineoplastic antimetabolite by blocking thymidylate synthetase conversion of deoxyuridylic acid to thymidylic acid in DNA synthesis. This study is aimed to know the anticancer effect of 5-FU on the expressions of important signaling proteins in KB cells through immunoprecipitation high performance liquid chromatography (IP-HPLC). KB cells were treated with 5 μM 5-FU and cultured for 12, 24, 48, 72, and 96 hours, and followed by IP-HPLC analysis using 32 antisera. 5-FU suppressed the proliferation of KB cells by decreases in the expressions of proliferation-related proteins, Ki-67, PCNA, CDK4, and MPM2 to 82.6%, 92.4%, 95.2%, and 95.9%, respectively, but increases of antiproliferation-related proteins, p16 and p21 to 106.7% and 125.5%, respectively, during 96 hours of experiment. This proliferation reduction was also negatively regulated by cMyc/MAX/MAD network signaling. The cellular protection and survival were consistently arrested by 5-FU treatment in KB cells. The expressions of NFkB, MDR, p-mTOR, and TNFα were decreased to 95.1%, 92.8%, 93.4%, and 90.3% in 48-72 hours, respectively, while cellular stress was increased by upregulation of p38 to 111.3% in 48 hours. And the expressions of pAKT1/2/3, hTERT, and AMPK were also decreased to 93.3%, 97.4%, and 89.3% in 24-48 hours, respectively, while the cellular transformation might be undergone by upregulation of TGF-β1 to 117% until 96 hours. Particularly, 5-FU treatment greatly induced the cellular apoptosis in KB cells by increased expressions of PARP, cPARP, caspase 9, c-caspase 9, caspase 8, and caspase 3 in the lack of p53/BAX and FASL/FAS signaling. The expressions of PARP and c-PARP were increased maximum to 119.2% in 24 hours, and followed by increases of caspase 9, c-caspase 9, caspase 8, and caspase 3 to 111.2%, 125.9%, 108.6%, and 116.3% in 72-96 hours. Therefore, it is presumed that 5-FU induced cellular apoptosis in KB cells may be derived from the overexpression of PARP due to the increased DNA defect caused by 5-FU, which can lead to ATP depletion and subsequent cellular apoptosis.

To compare functional Chinese cabbage(‘Amtak’ baechu; F1 hybrid cultivar between Brassica rapa and B. perkinensis, AB) with general Chinese cabbage (‘Chunkwang’ baechu; general spring cultivar, CB), two kinds of kimchi(ABK and CBK) prepared with AB and CB cultivar were fermented at 10°C for 10 days. Their fermentative characteristics and anti-proliferative activities against mouse carcinoma cell lines were investigated. General kimchi(CBK) showed mature pH on the 6th day of fermentation, whereas functional kimchi(ABK) reached pH on the 9th day. CBK also exhibited acidity of mature stage on the 6th day, but ABK reached mature acidity on the 9th day. Although ABK and CBK were salted in the same condition, ABK had lower salinity than CBK, throughout the fermentation time. The highest total bacterial and lactic bacterial counts of CBK showed on the 8th day of fermentation, but ABK showed the highest total bacterial and lactic bacterial counts on the 10th day. The texture of ABK was harder than CBK for fermentation time. This seems to be corrleated with the slower fermentation rate of ABK. ABK showed significantly higher anti-proliferative activity (54.6% cell viability of control) in B16BL6 at 1,000 μg/mL. ABK was also higher in anti-proliferative activity than CBK throughout the fermentation time. However, there was no significant difference in the anti-proliferative activity of ABK between the fermentation times. In conclusion, fermentation of ABK showed a better texture, due to the slow fermentation rate and more anti-proliferative activity against mouse carcinoma cell line than those of CBK.

Wolbachia is intra-cellular bacteria and frustrate host biology, such as parthenogenesis, feminization, male killing andsperm-egg incompatibility. It has been reported that > 90% of Aedes albopictus are infected with Wolbachia in the Republicof Korea (ROK). However any roles of the Wolbachia strains have not been reported in ROK. In this study, we infectedWolbachia to Vero cell line to investigate the cell migration which is related with the development of multi-cellular organisms,immune response and inflammation. Changes of wound healing and viability in vero cell after Wolbachia infection wereassessed. Cell migration was induced by Wolbachia after 24hr. Cell viability was not affected by Wolbachia after 24hr.This study will be helpful to understand the role of Wolbachia strain and support various information in Dengue andZika vector management programs.