A novel defensin-like peptide C-13326 possesses protective effect against multidrug-resistant Aeromonas schubertii in hybrid snakehead (Channa maculate ♀ × Channa argus ♂).

The purpose of this study was to investigate whether a defensin-like antimicrobial peptide (C-13326 peptide) identified in Hermetia illucens could possess protective effect against multidrug-resistant Aeromonas schubertii in hybrid snakehead (Channa maculate ♀ × Channa argus ♂). The cDNA of C-13326 peptide comprised 243 nucleotides encoding 80 amino acids, with six conserved cysteine residues and the classical CSαß structure. The recombinant expression plasmid pPIC9K-C-13326 was constructed and transformed into GS115 Pichia pastoris, and the C-13326 peptide was expressed by induction with 1% methanol. The crude extract of C-13326 peptide was precipitated by ammonium sulfate, assayed by Braford method, detected by tricine-SDS-PAGE, evaluated by BandScan software and identified by liquid chromatography-mass spectrometry. The C-13326 peptide was shown to have inhibitory activity against the growth of multidrug-resistant A. schubertii DM210910 by using the minimum growth inhibitory concentration and Oxford cup method. In addition, scanning electron microscopy analysis suggested that C-13326 peptide inhibited the growth of A. schubertii DM210910 by damaging the bacterial cell membrane. To explore the role of peptide C-13326 in vivo, hybrid snakehead was fed with peptide C-13326 as feed additives for 7 days. The results revealed that C-13326 peptide could significantly down-regulate the expression levels of IL-1ß, IL-8, IL-12 and TNF-α (p < .05), and significantly improved the survival rate of hybrid snakehead after challenging with A. schubertii DM210910. Therefore, the C-13326 peptide is a promising antimicrobial agent for A. schubertii treatment in aquaculture.

"Star" miR-34a and CXCR4 antagonist based nanoplex for binary cooperative migration treatment against metastatic breast cancer.

Invasion and metastasis of tumor cells is one of the major obstacles in cancer therapy. The process of tumor metastasis and diffusion is coordinated by multiple pathways associated with chemokine signals and migration microenvironment. In our previous work, chemokine CXC receptor 4 (CXCR4) antagonists showed significant anti-metastatic effects by blocking the CXCR4/stromal cell-derived factor-1(SDF-1) axis in pancreatic cancer and breast cancer. Here, we proposed to achieve migration chain-treatment for metastatic tumors by introducing a cell adhesion molecules CD44 inhibitor (Star miR-34a) to deprive of cell migration capability on the basis of CXCR4 antagonism (cyclam monomer, CM). Dextrin modified 1.8 k PEI with CM-end was prepared to deliver therapeutic miR-34a (named DPC/miR-34a) for efficient anti-metastasis by downregulating adhesion protein CD44 and targeting the CXCR4/SDF-1 axis. Additionally, reduced expression of the anti-apoptotic protein Bcl2 caused by miR-34a could enhance the anti-tumor efficacy of DPC/miR-34a nanoplex administration. Compared with inhibition of the CXCR4/SDF-1 axis or CD44 expression, the multidimensional therapy (DPC/miR-34a) exhibited considerable suppression of cancer cell invasion as assessed by an in vitro cell invasion assay and in vivo anti-metastasis model. Moreover, DPC/miR-34a demonstrated a superior antitumor and anti-metastatic efficacy both in lung metastatic model and orthotopic MDA-MB-231 tumor models, thus providing an efficient approach for combating metastatic tumors.

H2O2-activated oxidative stress amplifier capable of GSH scavenging for enhancing tumor photodynamic therapy.

Photodynamic therapy (PDT) is a clinically approved cancer treatment approach that relies on the generation of excess reactive oxygen species (ROS) to eradicate tumor cells by inducing oxidative stress. Unfortunately, if the tumor's endogenous glutathione (GSH) is overexpressed, it will eliminate the ROS and restrict the therapeutic efficacy of PDT. Herein, we report a H2O2-activated oxidative stress amplifier (OSA) for enhancing the ROS generation for PDT via GSH scavenging. Cinnamaldehyde (Cin) and chlorin e6 (Ce6) were applied as the GSH scavenger and photosensitizer, respectively, which were assembled with the ROS-responsive amphipathic polymer (DPL) to form DPL@CC micelles as the OSA. In the circulation of blood, the OSA can effectively protect the Cin from albumin binding to retain its GSH depletion ability. Once the OSA reached the tumor site, the high level of H2O2 triggered the degradation of DPL and led to the release of Cin and Ce6. Subsequently, the released Cin reacted with the intracellular GSH by Michael Addition and downregulated the GSH level to about 18.9%, versus untreated cells, to weaken the anti-oxidation ability of tumor cells. Thus, it provided a suitable environment for PDT to obtain an amplifying effect on oxidative stress and superior anti-cancer efficacy of 94% growth inhibition. The preparation of the H2O2-activated oxidative stress amplifier is a convincing strategy for promoting intracellular ROS generation and enhancing the tumor PDT efficacy, which could also augment the clinical application of PDT.

SoNar, a Highly Responsive NAD+/NADH Sensor, Allows High-Throughput Metabolic Screening of Anti-tumor Agents.

The altered metabolism of tumor cells confers a selective advantage for survival and proliferation, and studies have shown that targeting such metabolic shifts may be a useful therapeutic strategy. We developed an intensely fluorescent, rapidly responsive, pH-resistant, genetically encoded sensor of wide dynamic range, denoted SoNar, for tracking cytosolic NAD(+) and NADH redox states in living cells and in vivo. SoNar responds to subtle perturbations of various pathways of energy metabolism in real time, and allowed high-throughput screening for new agents targeting tumor metabolism. Among > 5,500 unique compounds, we identified KP372-1 as a potent NQO1-mediated redox cycling agent that produced extreme oxidative stress, selectively induced cancer cell apoptosis, and effectively decreased tumor growth in vivo. This study demonstrates that genetically encoded sensor-based metabolic screening could serve as a valuable approach for drug discovery.

The biphasic redox sensing of SENP3 accounts for the HIF-1 transcriptional activity shift by oxidative stress.

AIM: To investigate the mechanisms underlying the biphasic redox regulation of hypoxia-inducible factor-1 (HIF-1) transcriptional activity under different levels of oxidative stress caused by reactive oxidative species (ROS). METHODS: HeLa cells were exposed to different concentrations of H(2)O(2) as a simple model for mild and severe oxidative stress. Luciferase reporter assay and/or quantitative real-time PCR were used to investigate the transcriptional activity. Immunoblot was used to detect protein expression. Chromatin immunoprecipitation assay was used to detect HIF-1/DNA binding. The interaction of p300 with HIF-1α or with SENP3, and the SUMO2/3 conjugation states of p300 were examined by coimmunoprecipitation. RESULTS: HIF-1 transcriptional activity in HeLa cells was enhanced by low doses (0.05-0.5 mmol/L) of H(2)O(2), but suppressed by high doses (0.75-8.0 mmol/L) of H(2)O(2). The amount of co-activator p300 bound to HIF-1α in HeLa cells was increased under mild oxidative stress, but decreased under severe oxidative stress. The ROS levels differentially modified cysteines 243 and 532 in the cysteine protease SENP3, regulating the interaction of SENP3 with p300 to cause different SUMOylation of p300, thus shifting HIF-1 transcriptional activity. CONCLUSION: The shift of HIF-1 transactivation by ROS is correlated with and dependent on the biphasic redox sensing of SENP3 that leads to the differential SENP3/p300 interaction and the consequent fluctuation in the p300 SUMOylation status.

Emodin enhances sensitivity of gallbladder cancer cells to platinum drugs via glutathion depletion and MRP1 downregulation.

Glutathione conjugation and transportation of glutathione conjugates of anticancer drugs out of cells are important for detoxification of many anticancer drugs. Inhibition of this detoxification system has recently been proposed as a strategy to treat drug-resistant solid tumors. Gallbladder carcinoma is resistant to many anticancer drugs, therefore, it is needed to develop a novel strategy for cancer therapy. In the present study, we tested the effect of emodin (1,3,8-trihydroxy-6-methylanthraquinone), a reactive oxygen species (ROS) generator reported by our group previously, in combination with cisplatin (CDDP), carboplatin (CBP) or oxaliplatin in treating the gallbladder carcinoma cell line SGC996. Our results showed that co-treatment with emodin could remarkably enhance chemosensitivity of SGC996 cells in comparison with cisplatin, carboplatin or oxaliplatin treatment alone. We found that the mechanisms may be attributed to reduction of glutathione level, and downregulation of multidrug resistance-related protein 1 (MRP1) expression in SGC996 cells. The experiments on tumor-bearing mice showed that emodin/cisplatin co-treatment inhibited the tumor growth in vivo via increasing tumor cell apoptosis and downregulating MRP1 expression. In conclusion, emodin can work as an adjunct to enhance the anticancer effect of platinum drugs in gallbladder cancer cells via ROS-related mechanisms.

[The influence of elective course of emergency treatment for medical students on the cultivation of first aid knowledge and skill of cardio-pulmonary resuscitation].

OBJECTIVE: To investigate the influence of elective course of emergency treatment for medical students on the cultivation of first aid knowledge and skills of cardio-pulmonary resuscitation. METHODS: Senior students major in medicine of our university were randomly divided into observation group and contrast group with 30 students in each group according to whether an elective course of emergency treatment was given or not. All of them then received a test of first aid knowledge and cardio-pulmonary resuscitation skills. RESULTS: The theoretical exam scores in observation group and contrast group were respectively 78.5+/-9.1 and 46.7+/-15.6. The scores in observation group were significantly higher than that in contrast group (P < 0.01). Cardio-pulmonary resuscitation skills scores in observation group and contrast group were respectively 7.32+/-0.83 and 6.63+/-0.91. The scores in observation group were significantly higher than that in contrast group (P < 0.01). The number of failure for closed cardiac massage in 60 times in observation group and contrast group was respectively 5.06+/-0.58 and 5.77+/-0.63. The number of mouth to mouth artificial respiration in 4 times in observation group and contrast group was 0.92+/-0.16 and 1.10+/-0.17, respectively. There were notable differences in the number of failure in resuscitation maneuvers between two groups (both P < 0.01), observation group being obviously poorer than contrast group. CONCLUSION: An elective course of emergency treatment given to medical students plays an important role in the cultivation of first aid knowledge and skills in cardio-pulmonary resuscitation. It is therefore necessary that emergency medicine is included as a required course in medical college.

[Effects of rosiglitazone on inflammatory reaction and insulin resistance in obese patients with newly diagnosed type 2 diabetes].

OBJECTIVE: To study the effect of rosiglitazone on serum high-sensitivity C-reactive protein (hs-CRP), interleukin-1beta (IL-1beta), IL-6, tumor necrosis factor-alpha (TNF-alpha) and insulin resistance in obese patients with newly diagnosed type 2 diabetes. METHODS: This study involved 118 patients with newly diagnosed type 2 diabetes and obesity, who were randomly assigned into two groups for a 12-week treatment with rosiglitazone (4 mg/day, group A) or sulfonylureas (group B). Serum hs-CRP, IL-1beta, IL-6, TNF-alpha, fasting plasma glucose (FPG) and fasting insulin (FINS) were measured before and after the treatment. Insulin resistance index was calculated according to the HOMA Model. RESULTS: In group A, rosiglitazone treatment resulted in significantly reduced serum hs-CRP, IL-1beta, IL-6, TNF-alpha, FPG and insulin resistance index (P<0.01). No difference in FPG was found between the two groups after the treatment (P>0.05), but serum hs-CRP, IL-1beta, IL-6, TNF-alpha and insulin resistance index were significantly lower in group A than in group B (P<0.05). CONCLUSION: Rosiglitazone can decrease FPG, reduce the inflammation reaction and improve insulin resistance in obese patients with type 2 diabetes.

Emodin enhances cytotoxicity of chemotherapeutic drugs in prostate cancer cells: the mechanisms involve ROS-mediated suppression of multidrug resistance and hypoxia inducible factor-1.

The intrinsic or acquired resistance to multiple drugs (MDR) of cancer cells remains one of the main obstacles for chemotherapy. Development of small molecule targeting to hypoxia inducible factor-1 (HIF-1) has been recently proposed as strategy for treatments of drug-resistant solid tumors. In the present study, emodin, proven as a reactive oxygen species (ROS) generator by our previous work, was applied in combination with cisplatin and other chemotherapeutic drugs in the multidrug resistant prostate carcinoma cell line DU-145 and normal human dermal fibroblasts. Results showed that emodin/cisplatin co-treatment remarkably elevated ROS level and enhanced chemosensitivity in DU-145 cells, compared with cisplatin-only treatment, but exerted little effect on non-tumor cells. The effect of co-treatment on MDR1 gene and its upstream regulator HIF-1 was then investigated in DU-145. Co-treatment downregulated MDR1 expression and promoted drug retention, and meanwhile suppressed transactivation of HIF-1 in response to hypoxia without changing expression of HIF-1 alpha. The experiments on tumor-bearing mice showed that co-treatment inhibited the tumor growth in vivo, owing to oxidative stress and MDR1 down-regulation within tumors. HIF-1 transactivation and clonegenesis were suppressed in cells isolated from the tumors. Finally, examinations for the body weight, the organ histology and the antioxidant capacity of serum suggested that no systemic toxicity related to co-treatment was discernable. In conclusions, emodin, as a novel small inhibitor of HIF-1, may be recognized an effective adjunctive to improve efficacy of cytotoxic drugs in prostate cancer cells with over-activated HIF-1 and potent MDR.

The endogenous reactive oxygen species promote NF-kappaB activation by targeting on activation of NF-kappaB-inducing kinase in oral squamous carcinoma cells.

Reactive oxygen species (ROS) could stimulate or inhibit NF-kappaB pathways. However, most results have been obtained on the basis of the exogenous ROS and the molecular target of ROS in NF-kappaB signalling pathways has remained unclear. Here, the oral squamous carcinoma (OSC) cells, with a mild difference in the endogenous ROS level, were used to investigate how slight fluctuation of the endogenous ROS regulates NF-kappaB activation. This study demonstrates that NF-kappaB-inducing kinase (NIK) is a critical target of the endogenous ROS in NF-kappaB pathways. The results indicate that ROS may function as a physiological signalling modulator on NF-kappaB signalling cascades through its ability to facilitate the activity of NIK and subsequent NF-kappaB transactivation. In addition, the data are useful to explain why the altered intracellular microenvironment related to redox state may influence biological behaviours of cancer cells.