Functional Analysis of Membrane-Associated Scaffolding Tight Junction (TJ) Proteins in Tumorigenic Characteristics of B16-F10 Mouse Melanoma Cells.

Tight junction (TJ) proteins (Tjps), Tjp1 and Tjp2, are tight junction-associated scaffold proteins that bind to the transmembrane proteins of tight junctions and the underlying cytoskeleton. In this study, we first analyzed the tumorigenic characteristics of B16-F10 melanoma cells, including cell proliferation, migration, invasion, metastatic potential, and the expression patterns of related proteins, after the CRISPR-Cas9-mediated knockout (KO) of Tjp genes. The proliferation of Tjp1 and Tjp2 KO cells significantly increased in vitro. Other tumorigenic characteristics, including migration and invasion, were significantly enhanced in Tjp1 and Tjp2 KO cells. Zonula occludens (ZO)-associated protein Claudin-1 (CLDN-1), which is a major component of tight junctions and functions in controlling cell-to-cell adhesion, was decreased in Tjp KO cells. Additionally, Tjp KO significantly stimulated tumor growth and metastasis in an in vivo mouse model. We performed a transcriptome analysis using next-generation sequencing (NGS) to elucidate the key genes involved in the mechanisms of action of Tjp1 and Tjp2. Among the various genes affected by Tjp KO-, cell cycle-, cell migration-, angiogenesis-, and cell-cell adhesion-related genes were significantly altered. In particular, we found that the Ninjurin-1 (Ninj1) and Catenin alpha-1 (Ctnna1) genes, which are known to play fundamental roles in Tjps, were significantly downregulated in Tjp KO cells. In summary, tumorigenic characteristics, including cell proliferation, migration, invasion, tumor growth, and metastatic potential, were significantly increased in Tjp1 and Tjp2 KO cells, and the knockout of Tjp genes significantly affected the expression of related proteins.

Molecular mechanisms underlying the vulnerability of Pacific abalone (Haliotis discus hannai) to Vibrio harveyi infection at higher water temperature.

Climate change is one of the most important threats to farmed abalone worldwide. Although abalone is more susceptible to vibriosis at higher water temperatures, the molecular mode of action underlying this has not been fully elucidated. Therefore, this study aimed to address the high susceptibility of Halitotis discus hannai to V. harveyi infection using abalone hemocytes exposed to low and high temperatures. Abalone hemocytes were divided into four groups, 20C, 20 V, 25C, and 25 V, depending on co-culture with (V)/without (C) V. harveyi (MOI = 12.8) and incubation temperature (20 °C or 25 °C). After 3 h of incubation, hemocyte viability and phagocytic activity were measured, and RNA sequencing was performed using Illumina Novaseq. The expression of several virulence-related genes in V. harveyi was analyzed using real-time PCR. The viability of hemocytes was significantly decreased in the 25 V group compared to cells in the other groups, whereas phagocytic activity at 25 °C was significantly higher than at 20 °C. Although a number of immune-associated genes were commonly upregulated in abalone hemocyte exposed to V. harveyi, regardless of temperature, pathways and genes regarding pro-inflammatory responses (interleukin-17 and tumor necrosis factor) and apoptosis were significantly overexpressed in the 25 V group compared to the 25C group. Notably, in the apoptosis pathway, genes encoding executor caspases (casp3 and casp7) and pro-apoptotic factor, bax were significantly up-regulated only in the 25 V group, while the apoptosis inhibitor, bcl2L1 was significantly up-regulated only in the 20 V group compared to the control group at the respective temperatures. The co-culture of V. harveyi with abalone hemocytes at 25 °C up-regulated several virulence-related genes involved in quorum sensing (luxS), antioxidant activity (katA, katB, and sodC), motility (flgI), and adherence/invasion (ompU) compared to those at 20 °C. Therefore, our results showed that H. discus hannai hemocytes exposed to V. harveyi at 25 °C were highly stressed by vigorously activated inflammatory responses and that the bacterial pathogen overexpressed several virulence-related genes at the high temperature tested. The transcriptomic profile of both abalone hemocytes and V. harveyi in the present study provide insight into differential host-pathogen interactions depending on the temperature conditions and the molecular backgrounds related to increased abalone vulnerability upon global warming.

The Role of Human Endogenous Retrovirus (HERV)-K119 env in THP-1 Monocytic Cell Differentiation.

Human endogenous retrovirus (HERV)-K was reportedly inserted into the human genome millions of years ago and is closely related to various diseases, including cancer and immune regulation. In our previous studies, CRISPR-Cas9-enabled knockout (KO) of the HERV-K env gene was found to potentially reduce cell proliferation, cell migration, and invasion in colorectal and ovarian cancer cell lines. The immune response involves the migration and invasion of cells and is similar to cancer; however, in certain ways, it is completely unlike cancer. Therefore, we induced HERV-K119 env gene KO in THP-1, a monocytic cell that can be differentiated into a macrophage, to investigate the role of HERV-K119 env in immune regulation. Cell migration and invasion were noted to be significantly increased in HERV-K119 env KO THP-1 cells than in MOCK, and these results were contrary to those of cancer cells. To identify the underlying mechanism of HERV-K119 env KO in THP-1 cells, transcriptome analysis and cytokine array analysis were conducted. Semaphorin7A (SEMA7A), which induces the production of cytokines in macrophages and monocytic cells and plays an important role in immune effector cell activation during an inflammatory immune response, was significantly increased in HERV-K119 env KO THP-1 cells. We also found that HERV-K119 env KO THP-1 cells expressed various macrophage-specific surface markers, suggesting that KO of HERV-K119 env triggers the differentiation of THP-1 cells from monocytic cells into macrophages. In addition, analysis of the expression of M1 and M2 macrophage markers showed that M1 macrophage marker cluster of differentiation 32 (CD32) was significantly increased in HERV-K119 env KO cells. These results suggest that HERV-K119 env is implicated in the differentiation of monocytic cells into M1 macrophages and plays important roles in the immune response.

Inhibition of Urban Particulate Matter-Induced Airway Inflammation by RIPK3 through the Regulation of Tight Junction Protein Production.

Urban particulate matter (UPM) is a high-hazard cause of various diseases in humans, including in the respiratory tract, skin, heart, and even brain. Unfortunately, there is no established treatment for the damage caused by UPM in the respiratory epithelium. In addition, although RIPK3 is known to induce necroptosis, its intracellular role as a negative regulator in human lungs and bronchial epithelia remains unclear. Here, the endogenous expression of RIPK3 was significantly decreased 6 h after exposure to UPM. In RIPK3-ovexpressed cells, RIPK3 was not moved to the cytoplasm from the nucleus. Interestingly, the overexpression of RIPK3 dramatically decreased TEER and F-actin formation. Its overexpression also decreased the expression of genes for pro-inflammatory cytokines (IL-6 and IL-8) and tight junctions (ZO-1, -2, -3, E-cadherin, and claudin) during UPM-induced airway inflammation. Importantly, overexpression of RIPK3 inhibited the UPM-induced ROS production by inhibiting the activation of iNOS and eNOS and by regulating mitochondrial fission processing. In addition, UPM-induced activation of the iκB and NF-κB signaling pathways was dramatically decreased by RIPK3, and the expression of pro-inflammatory cytokines was decreased by inhibiting the iκB signaling pathway. Our data indicated that RIPK3 is essential for the UPM-induced inflammatory microenvironment to maintain homeostasis. Therefore, we suggest that RIPK3 is a potential therapeutic candidate for UPM-induced pulmonary inflammation.

Phloroglucinol Inhibits Oxidative-Stress-Induced Cytotoxicity in C2C12 Murine Myoblasts through Nrf-2-Mediated Activation of HO-1.

Phloroglucinol is a class of polyphenolic compounds containing aromatic phenyl rings and is known to have various pharmacological activities. Recently, we reported that this compound isolated from Ecklonia cava, a brown alga belonging to the family Laminariaceae, has potent antioxidant activity in human dermal keratinocytes. In this study, we evaluated whether phloroglucinol could protect against hydrogen peroxide (H2O2)-induced oxidative damage in murine-derived C2C12 myoblasts. Our results revealed that phloroglucinol suppressed H2O2-induced cytotoxicity and DNA damage while blocking the production of reactive oxygen species. We also found that phloroglucinol protected cells from the induction of apoptosis associated with mitochondrial impairment caused by H2O2 treatment. Furthermore, phloroglucinol enhanced the phosphorylation of nuclear factor-erythroid-2 related factor 2 (Nrf2) as well as the expression and activity of heme oxygenase-1 (HO-1). However, such anti-apoptotic and cytoprotective effects of phloroglucinol were greatly abolished by the HO-1 inhibitor, suggesting that phloroglucinol could increase the Nrf2-mediated activity of HO-1 to protect C2C12 myoblasts from oxidative stress. Taken together, our results indicate that phloroglucinol has a strong antioxidant activity as an Nrf2 activator and may have therapeutic benefits for oxidative-stress-mediated muscle disease.

Downregulated pol-miR-140-3p induces the expression of the kinesin family member 5A against Streptococcus parauberis infection in olive flounder.

MicroRNAs (miRNAs) are small non-coding RNAs that participate in various biological and cellular processes by regulating target gene expression. miRNAs are also known to play vital roles in the pathogenesis of various diseases, including infections, as well as the disease progression and defense responses. In this study, we examined the expression levels of pol-miR-140-3p and its target gene, kinesin family member 5A (KIF5A), in association with the Streptococcus parauberis (S. parauberis) infection, a major bacterial pathogen that causes streptococcosis in olive flounder (Paralichthys olivaceus). KIF5A is a heavy chain isoform of kinesin-1, which is known to be brain-specific, and this study is the first examination of KIF5A expression related to the regulation of miRNA in olive flounder (named PoKIF5A). There were significant differences in expression levels between infected and healthy olive flounder as the expression of pol-miR-140-3p in the infected fish was lower than that in the control, while the expression of PoKIF5A was higher in the infected fish than in the healthy controls. These contradictory results suggest that downregulated pol-miR-140-3p induces the expression of PoKIF5A against S. parauberis infection in olive flounder.

Induction of apoptosis through inactivation of ROS-dependent PI3K/Akt signaling pathway by platycodin D in human bladder urothelial carcinoma cells.

Platycodin D (PD) is a triterpenoid saponin, a major bioactive constituent of the roots of Platycodon grandiflorum, which is well known for possessing various pharmacological properties. However, the anti-cancer mechanism of PD in bladder cancer cells remains poorly understood. In the current study, we investigated the effect of PD on the growth of human bladder urothelial carcinoma cells. PD treatment significantly reduced the cell survival of bladder cancer cells associated with induction of apoptosis and DNA damage. PD inhibited the expression of inhibitor of apoptosis family members, activated caspases, and induced cleavage of poly (ADP-ribose) polymerase. PD also increased the release of cytochrome c into the cytoplasm by disrupting the mitochondrial membrane potential while upregulating the expression ratio of Bax to Bcl-2. The PD-mediated anti-proliferative effect was significantly inhibited by pre-treatment with a pancaspase inhibitor, but not by an inhibitor of necroptosis. Moreover, PD suppressed the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway, and the apoptosis-inducing effect of PD was further enhanced by a PI3K inhibitor. In addition, PD increased the accumulation of reactive oxygen species (ROS), whereas N-acetyl cysteine (NAC), an ROS inhibitor, significantly attenuated the growth inhibition and inactivation of the PI3K/Akt/mTOR signaling caused by PD. Furthermore, NAC significantly suppressed apoptosis, DNA damage, and decreased cell viability induced by PD treatment. Collectively, our findings indicated that PD blocked the growth of bladder urothelial carcinoma cells by inducing ROS-mediated inactivation of the PI3K/Akt/mTOR signaling.

The regulation of the TLR4/NF-κB and Nrf2/HO-1 signaling pathways is involved in the inhibition of lipopolysaccharide-induced inflammation and oxidative reactions by morroniside in RAW 264.7 macrophages.

Morroniside, a major iridoid glycoside isolated from Cornus officinalis, has a variety of beneficial pharmacological properties. Although morroniside has recently been reported to exhibit anti-inflammatory and antioxidant effects, the detailed mechanism has not yet been fully elucidated. In this study, we investigated the inhibitory effect of morroniside on inflammatory and oxidative stress activated by lipopolysaccharide (LPS) in RAW 264.7 macrophages. Our results indicated that morroniside pretreatment significantly inhibited the LPS-induced phagocytic activity and release of pro-inflammatory factors, which was associated with blocking the expression of their regulatory genes. Morroniside also markedly suppressed the expression of myeloid differentiation factor 88 as well as Toll-like receptor 4 (TLR4), and attenuated the translocation of nuclear factor-κB (NF-κB) to the nucleus in LPS-treated RAW 264.7 macrophages. Furthermore, morroniside prevented the binding of LPS to the TLR4 on the cell surface. In addition, morroniside abolished reactive oxygen species (ROS) generation, and enhanced the expression of heme oxygenase-1 (HO-1) following activation of nuclear factor-E2-related factor 2 (Nrf2) in LPS-stimulated RAW 264.7 macrophages. However, zinc protoporphyrin, a specific inhibitor of HO-1, reversed the morroniside-mediated inhibition of inflammatory response in LPS-treated RAW 264.7 macrophages. In conclusion, our findings suggest that morroniside exerts LPS-induced anti-inflammatory and antioxidant effects by targeting the TLR4/NF-κB and Nrf2/HO-1 signaling pathways in RAW 264.7 macrophages. Taken together, our findings suggest that morroniside interacted structurally and electrochemically with TLR4/MD2 complex, consequently can be a potential functional agent to prevent inflammatory and oxidative damage.

Human Endogenous Retrovirus (HERV)-K env Gene Knockout Affects Tumorigenic Characteristics of nupr1 Gene in DLD-1 Colorectal Cancer Cells.

Human endogenous retroviruses (HERVs) are suggested to be involved in the development of certain diseases, especially cancers. To elucidate the function of HERV-K Env protein in cancers, an HERV-K env gene knockout (KO) in DLD-1 colorectal cancer cell lines was generated using the CRISPR-Cas9 system. Transcriptome analysis of HERV-K env KO cells using next-generation sequencing (NGS) was performed to identify the key genes associated with the function of HERV-K Env protein. The proliferation of HERV-K env KO cells was significantly reduced in in vitro culture as well as in in vivo nude mouse model. Tumorigenic characteristics, including migration, invasion, and tumor colonization, were also significantly reduced in HERV-K env KO cells. Whereas, they were enhanced in HERV-K env over-expressing DLD-1 cells. The expression of nuclear protein-1 (NUPR1), an ER-stress response factor that plays an important role in cell proliferation, migration, and reactive oxygen species (ROS) generation in cancer cells, significantly reduced in HERV-K env KO cells. ROS levels and ROS-related gene expression was also significantly reduced in HERV-K env KO cells. Cells transfected with NUPR1 siRNA (small interfering RNA) exhibited the same phenotype as HERV-K env KO cells. These results suggest that the HERV-K env gene affects tumorigenic characteristics, including cell proliferation, migration, and tumor colonization through NUPR1 related pathway.

Apoptotic Effects of Anthocyanins from Vitis coignetiae Pulliat Are Enhanced by Augmented Enhancer of the Rudimentary Homolog (ERH) in Human Gastric Carcinoma MKN28 Cells.

Evidence suggests that augmented expression of a certain gene can influence the efficacy of targeted and conventional chemotherapies. Here, we tested whether the high expression of enhancer of the rudimentary homolog (ERH), which serves as a prognostic factor in some cancers, can influence the efficacy of anthocyanins isolated from fruits of Vitis coignetiae Pulliat, Meoru in Korea (AIMs) on human gastric cancer cells. The anticancer efficacy of AIMs was augmented in ERH-transfected MKN28 cells (E-MKN28 cells). Molecularly, ERH augmented AIM-induced caspase-dependent apoptosis by activating caspase-3 and -9. The ERH-augmented apoptotic effect was related to mitochondrial depolarization and inhibition of antiapoptotic proteins, XIAP, and Bcl-2. In addition, reactive oxygen species (ROS) generation was augmented in AIMs-treated E-MKN28 cells compared to AIMs-treated naïve MKN28 cells. In conclusion, ERH augmented AIM-induced caspase-dependent mitochondrial-related apoptosis in MKN28 cells. A decrease in expression of Bcl-2 and subsequent excessive ROS generation would be the mechanism for ERH-augmented mitochondrial-related apoptosis in AIMs-treated MKN28 cells. A decrease in expression of XIAP would be another mechanism for ERH-augmented caspase-dependent apoptosis in AIMs-treated MKN28 cells.

Spermidine Attenuates Oxidative Stress-Induced Apoptosis via Blocking Ca2+ Overload in Retinal Pigment Epithelial Cells Independently of ROS.

Retinal pigment epithelial (RPE) cells occupy the outer layer of the retina and perform various biological functions. Oxidative damage to RPE cells is a major risk factor for retinal degeneration that ultimately leads to vision loss. In this study, we investigated the role of spermidine in a hydrogen peroxide (H2O2)-induced oxidative stress model using human RPE cells. Our findings showed that 300 µM H2O2 increased cytotoxicity, apoptosis, and cell cycle arrest in the G2/M phase, whereas these effects were markedly suppressed by 10 µM spermidine. Furthermore, spermidine significantly reduced H2O2-induced mitochondrial dysfunction including mitochondrial membrane potential and mitochondrial activity. Although spermidine displays antioxidant properties, the generation of intracellular reactive oxygen species (ROS) upon H2O2 insult was not regulated by spermidine. Spermidine did suppress the increase in cytosolic Ca2+ levels resulting from endoplasmic reticulum stress in H2O2-stimulated human RPE cells. Treatment with a cytosolic Ca2+ chelator markedly reversed H2O2-induced cellular dysfunction. Overall, spermidine protected against H2O2-induced cellular damage by blocking the increase of intracellular Ca2+ independently of ROS. These results suggest that spermidine protects RPE cells from oxidative stress, which could be a useful treatment for retinal diseases.

Exercise training reduces the risk of opportunistic infections after acute exercise and improves cytokine antigen recognition.

In general, acute exercise is thought to inhibit immune function and increase the risk of opportunistic infections, but there is some opposition to this due to a lack of quantitative evaluation. Therefore, we quantified the effect of exercise on immune function and observed the interaction between antigens and cytokines using an intramuscular infection with Trichinella spiralis (T. spiralis), a common parasitic infection model. C57BL/6 mice were used for a non-infection experiment and an infection (Inf) experiment. Each experiment was divided further into three groups: one control (CON) group, and an exercise pre-infection (PIE)-only group and exercise-sustained (ES) group, each of which was subjected to exercise for 7 weeks. All animals in the infection experiment were infected with T. spiralis 30 min after acute exercise. After infection, the ES and Inf-ES groups continued exercise for 7 additional weeks. The number of T. spiralis nurse cells remaining in skeletal muscles was fewer in the infected exercise groups compared with the infected control. Expression of interleukin-6 (IL-6) and interleukin-10 (IL-10) was higher in the Inf-CON group and transforming growth factor beta (TGF-ß) expression was lower in the Inf-CON group than in the CON group, as measured by RT-PCR. In the infection experiment, only IL-10 had significant differences between the groups. Immunofluorescence revealed that most cytokines were specifically expressed around the antigenic nurse cells following exercise. In conclusion, exercise training does not increase the risk of opportunistic infections even after acute exercise, but rather reduces it. These results may be due to antigen-specific immune responses.

Co-Expression Network Analysis of Spleen Transcriptome in Rock Bream (Oplegnathus fasciatus) Naturally Infected with Rock Bream Iridovirus (RBIV).

Rock bream iridovirus (RBIV) is a notorious agent that causes high mortality in aquaculture of rock bream (Oplegnathus fasciatus). Despite severity of this virus, no transcriptomic studies on RBIV-infected rock bream that can provide fundamental information on protective mechanism against the virus have been reported so far. This study aimed to investigate physiological mechanisms between host and RBIV through transcriptomic changes in the spleen based on RNA-seq. Depending on infection intensity and sampling time point, fish were divided into five groups: uninfected healthy fish at week 0 as control (0C), heavy infected fish at week 0 (0H), heavy mixed RBIV and bacterial infected fish at week 0 (0MH), uninfected healthy fish at week 3 (3C), and light infected fish at week 3 (3L). We explored clusters from 35,861 genes with Fragments Per Kilo-base of exon per Million mapped fragments (FPKM) values of 0.01 or more through signed co-expression network analysis using WGCNA package. Nine of 22 modules were highly correlated with viral infection (|gene significance (GS) vs. module membership (MM) |> 0.5, p-value < 0.05). Expression patterns in selected modules were divided into two: heavy infected (0H and 0MH) and control and light-infected groups (0C, 3C, and 3L). In functional analysis, genes in two positive modules (5448 unigenes) were enriched in cell cycle, DNA replication, transcription, and translation, and increased glycolysis activity. Seven negative modules (3517 unigenes) built in this study showed significant decreases in the expression of genes in lymphocyte-mediated immune system, antigen presentation, and platelet activation, whereas there was significant increased expression of endogenous apoptosis-related genes. These changes lead to RBIV proliferation and failure of host defense, and suggests the importance of blood cells such as thrombocytes and B cells in rock bream in RBIV infection. Interestingly, a hub gene, pre-mRNA processing factor 19 (PRPF19) showing high connectivity (kME), and expression of this gene using qRT-PCR was increased in rock bream blood cells shortly after RBIV was added. It might be a potential biomarker for diagnosis and vaccine studies in rock bream against RBIV. This transcriptome approach and our findings provide new insight into the understanding of global rock bream-RBIV interactions including immune and pathogenesis mechanisms.

Activation of the Nrf2/HO-1 signaling pathway contributes to the protective effects of baicalein against oxidative stress-induced DNA damage and apoptosis in HEI193 Schwann cells.

Baicalein, a flavonoid extracted from the roots of Scutellaria baicalensis Georgi., has various pharmacological effects due to its high antioxidant activity. However, no study has yet been conducted on the protective efficacy of baicalein against oxidative stress in Schwann cells. In this study, we evaluated the protective effect of baicalein on DNA damage and apoptosis induced by hydrogen peroxide (H2O2) in HEI193 Schwann cells. For this purpose, HEI193 cells exposed to H2O2 in the presence or absence of baicalein were applied to cell viability assay, immunoblotting, Nrf2-specific small interfering RNA (siRNA) transfection, comet assay, and flow cytometry analyses. Our results showed that baicalein effectively inhibited H2O2-induced cytotoxicity and DNA damage associated with the inhibition of reactive oxygen species (ROS) accumulation. Baicalein also weakened H2O2-induced mitochondrial dysfunction, increased the Bax/Bcl-2 ratio, activated caspase-9 and -3, and degraded poly(ADP-ribose) polymerase. In addition, baicalein increased not only the expression but also the phosphorylation of nuclear factor-erythroid 2 related factor 2 (Nrf2) and promoted the expression of heme oxygenase-1 (HO-1), a critical target enzyme of Nrf2, although the expression of kelch-like ECH-associated protein-1 was decreased. However, the inhibition of Nrf2 expression by transfection with Nrf2-siRNA transfection abolished the expression of HO-1 and antioxidant potential of baicalein. These results demonstrate that baicalein attenuated H2O2-induced apoptosis through the conservation of mitochondrial function while eliminating ROS in HEI193 Schwann cells, and the antioxidant efficacy of baicalein implies at least a Nrf2/HO-1 signaling pathway-dependent mechanism. Therefore, it is suggested that baicalein may have a beneficial effect on the prevention and treatment of peripheral neuropathy induced by oxidative stress.

Protective Effect of Phloroglucinol on Oxidative Stress-Induced DNA Damage and Apoptosis through Activation of the Nrf2/HO-1 Signaling Pathway in HaCaT Human Keratinocytes.

Phloroglucinol (PG) is a component of phlorotannins, which are abundant in marine brown alga species. Recent studies have shown that PG is beneficial in protecting cells from oxidative stress. In this study, we evaluated the protective efficacy of PG in HaCaT human skin keratinocytes stimulated with oxidative stress (hydrogen peroxide, H2O2). The results showed that PG significantly inhibited the H2O2-induced growth inhibition in HaCaT cells, which was associated with increased expression of heme oxygenase-1 (HO-1) by the activation of nuclear factor erythroid 2-related factor-2 (Nrf2). PG remarkably reversed H2O2-induced excessive ROS production, DNA damage, and apoptosis. Additionally, H2O2-induced mitochondrial dysfunction was related to a decrease in ATP levels, and in the presence of PG, these changes were significantly impaired. Furthermore, the increases of cytosolic release of cytochrome c and ratio of Bax to Bcl-2, and the activation of caspase-9 and caspase-3 by the H2O2 were markedly abolished under the condition of PG pretreatment. However, the inhibition of HO-1 function using zinc protoporphyrin, a HO-1 inhibitor, markedly attenuated these protective effects of PG against H2O2. Overall, our results suggest that PG is able to protect HaCaT keratinocytes against oxidative stress-induced DNA damage and apoptosis through activating the Nrf2/HO-1 signaling pathway.

Anti-Proliferative and Pro-Apoptotic Effects of Licochalcone A through ROS-Mediated Cell Cycle Arrest and Apoptosis in Human Bladder Cancer Cells.

Licochalcone A (LCA) is a chalcone that is predominantly found in the root of Glycyrrhiza species, which is widely used as an herbal medicine. Although previous studies have reported that LCA has a wide range of pharmacological effects, evidence for the underlying molecular mechanism of its anti-cancer efficacy is still lacking. In this study, we investigated the anti-proliferative effect of LCA on human bladder cancer cells, and found that LCA induced cell cycle arrest at G2/M phase and apoptotic cell death. Our data showed that LCA inhibited the expression of cyclin A, cyclin B1, and Wee1, but increased the expression of cyclin-dependent kinase (Cdk) inhibitor p21WAF1/CIP1, and increased p21 was bound to Cdc2 and Cdk2. LCA activated caspase-8 and -9, which are involved in the initiation of extrinsic and intrinsic apoptosis pathways, respectively, and also increased caspase-3 activity, a typical effect caspase, subsequently leading to poly (ADP-ribose) polymerase cleavage. Additionally, LCA increased the Bax/Bcl-2 ratio, and reduced the integrity of mitochondria, which contributed to the discharge of cytochrome c from the mitochondria to the cytoplasm. Moreover, LCA enhanced the intracellular levels of reactive oxygen species (ROS); however, the interruption of ROS generation using ROS scavenger led to escape from LCA-mediated G2/M arrest and apoptosis. Collectively, the present data indicate that LCA can inhibit the proliferation of human bladder cancer cells by inducing ROS-dependent G2/M phase arrest and apoptosis.

Thymoquinone-Induced Tristetraprolin Inhibits Tumor Growth and Metastasis through Destabilization of MUC4 mRNA.

Tristetraprolin (TTP), a well-characterized AU-rich element (ARE) binding protein, functions as a tumor suppressor gene. The purpose of this study was to investigate whether a bioactive substance derived from a natural medicinal plant affects the induction of TTP and to elucidate its mechanism. We examined the effects of natural bioactive materials including Resveratrol (RSV), thymoquinone (TQ) and curcumin on the expression of TTP in cancer cell. TQ derived from a natural plant Nigella sativa increased the expression levels of TTP mRNA and proteins in a dose-dependent manner in gastric and breast cancer cells. TQ-induced TTP increased the instability of MUC4 mRNA by direct binding of TTP to ARE in the 3'UTR of MUC4 mRNA. The induction of TTP by TQ also reduced the proliferation, migration and invasion of cancer cells. The expression of the epithelial-mesenchymal (EMT)-related genes, which were target genes of TTP, was also decreased by the TQ treatment. In the in vivo experiments using mouse melanoma cells, TQ-induced TTP inhibited metastasis of tumor cells. We have found that TQ-induced TTP might inhibit metastasis by reducing tumor cell migration and invasion through destabilization of MUC4 mRNA, which suggest the MUC4 as a novel target to TTP.

Ten Cases of Taenia saginata Infection Confirmed by Analysis of the Internal Transcribed Spacer 1 rDNA Region in the Republic of Korea.

From October 2015 to August 2018, tapeworm proglottids were obtained from 10 patients who were residents of Daegu and Gyeongbuk provinces and had a history of raw beef consumption. Most of them had no overseas travel experience. The gravid proglottids obtained from the 10 cases had 15-20 lateral uterine branches. A part of internal transcribed spacer 1 (ITS1) DNA of the 10 cases, amplified by polymerase chain reaction (PCR) and digested with AleI restriction enzyme, produced the same band pattern of Taenia saginata, which differentiated from T. asiatica and T. solium. Sequences of ITS1 and cytochrome c oxidase subunit 1 (cox1) showed higher homology to T. saginata than to T. asiatica and T. solium. Collectively, these 10 cases were identified as T. saginata human infections. As taeniasis is one of the important parasitic diseases in humans, it is necessary to maintain hygienic conditions during livestock farming to avoid public health concerns.

Induction of Angiogenesis by Malarial Infection through Hypoxia Dependent Manner.

Malarial infection induces tissue hypoxia in the host through destruction of red blood cells. Tissue hypoxia in malarial infection may increase the activity of HIF1α through an intracellular oxygen-sensing pathway. Activation of HIF1α may also induce vascular endothelial growth factor (VEGF) to trigger angiogenesis. To investigate whether malarial infection actually generates hypoxia-induced angiogenesis, we analyzed severity of hypoxia, the expression of hypoxia-related angiogenic factors, and numbers of blood vessels in various tissues infected with Plasmodium berghei. Infection in mice was performed by intraperitoneal injection of 2×106 parasitized red blood cells. After infection, we studied parasitemia and survival. We analyzed hypoxia, numbers of blood vessels, and expression of hypoxia-related angiogenic factors including VEGF and HIF1α. We used Western blot, immunofluorescence, and immunohistochemistry to analyze various tissues from Plasmodium berghei-infected mice. In malaria-infected mice, parasitemia was increased over the duration of infection and directly associated with mortality rate. Expression of VEGF and HIF1α increased with the parasitemia in various tissues. Additionally, numbers of blood vessels significantly increased in each tissue type of the malaria-infected group compared to the uninfected control group. These results suggest that malarial infection in mice activates hypoxia-induced angiogenesis by stimulation of HIF1α and VEGF in various tissues.

Blood-Stage Plasmodium Berghei ANKA Infection Promotes Hepatic Fibrosis by Enhancing Hedgehog Signaling in Mice.

BACKGROUND/AIMS: Malaria is the most deadly parasitic infection in the world, resulting in damage to various organs, including the liver, of the infected organism; however, the mechanism causing this damage in the liver remains unclear. Liver fibrosis, a major characteristic of liver diseases, occurs in response to liver injury and is regulated by a complex network of signaling pathways. Hedgehog (Hh) signaling orchestrates a number of hepatic responses including hepatic fibrogenesis. Therefore, we investigated whether Hh signaling influenced the liver's response to malarial infection. METHODS: Eight-week-old male C57BL/6 mice inoculated with blood containing Plasmodium berghei ANKA (PbA)-infected erythrocytes were sacrificed when the level of parasitemia in the blood reached 10% or 30%, and the livers were collected for biochemical analysis. Liver responses to PbA infection were examined by hematoxylin and eosin staining, real-time polymerase chain reaction, immunohistochemistry and western blot. RESULTS: Severe hepatic injury, such as ballooned hepatocytes, sinusoidal dilatation, and infiltrated leukocytes, was evident in the livers of the malaria-infected mice. Hypoxia was also induced in 30% parasitemia group. With the accumulation of Kupffer cells, inflammation markers, TNF-α, interleukin-1ß, and chemokine (C-X-C motif) ligand 1, were significantly upregulated in the infected group compared with the control group. Expression of fibrotic markers, including transforming growth factor-ß, α-smooth muscle actin (α-SMA), collagen 1a1, thymosin ß4, and vimentin, were significantly higher in the infected groups than in the control group. With increased collagen deposition, hepatic stellate cells expressing α-SMA accumulated in the liver of the PbA-infected mice, whereas those cells were rarely detected in the livers of the control mice. The levels of Hh signaling and Yes-associated protein (YAP), two key regulators for hepatic fibrogenesis, were significantly elevated in the infected groups compared with the control group. Treatment of mice with Hh inhibitor, GDC-0449, reduced hepatic inflammation and fibrogenesis with Hh suppression in PbA-infected mice. CONCLUSION: Our results demonstrate that HSCs are activated in and Hh and YAP signaling are associated with this process, contributing to increased hepatic fibrosis in malaria-infected livers.