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.