Methylene Blue Reduces Acute Cerebral Ischemic Injury via the Induction of Mitophagy

The treatment of stroke is limited by a short therapeutic window and a lack of effective clinical drugs. Methylene blue (MB) has been used in laboratories and clinics since the 1890s. Few studies have reported the neuroprotective role of MB in cerebral ischemia-reperfusion injury. However, whether and how MB protects against acute cerebral ischemia (ACI) injury was unclear. In this study, we investigated the effect of MB on this injury and revealed that MB protected against ACI injury by augmenting mitophagy. Using a rat middle cerebral artery occlusion (MCAO) model, we demonstrated that MB improved neurological function and reduced the infarct volume and necrosis after ACI injury. These improvements depended on the effect of MB on mitochondrial structure and function. ACI caused the disorder and disintegration of mitochondrial structure, while MB ameliorated the destruction of mitochondria. In addition, mitophagy was inhibited at 24 h after stroke and MB augmented mitophagy. In an oxygen-glucose deprivation (OGD) model in vitro, we further revealed that the elevation of mitochondrial membrane potential (MMP) by MB under OGD conditions mediated the augmented mitophagy. In contrast, exacerbating the decline of MMP during OGD abolished the MB-induced activation of mitophagy. Taken together, MB promotes mitophagy by maintaining the MMP at a relatively high level, which contributes to a decrease in necrosis and an improvement in neurological function, thereby protecting against ACI injury.     

Production of recombinant human lysozyme in the milk of transgenic pigs

In the swine industry pathogenic infections have a significant negative impact on neonatal survival. Piglets fed with human lysozyme, a natural antibiotic, might be more resistant to gastrointestinal infections. Here we describe the generation of transgenic swine expressing recombinant human lysozyme by somatic cell nuclear transfer. Three cloned female pigs were born, one of which expressed rhLZ at 0.32 ± 0.01 μg/ml in milk, 50-fold higher than that of the pig native lysozyme. Both the transgenic gilts and their progeny appear healthy. Introducing human lysozyme into pigs’ milk has a potential to benefit the piglets by enhancing immune function and defending against pathogenic bacteria, thereby increasing the new born survival rate. This advance could be of great value to commercial swine producers.     

Mechanisms of drug sensitization to TRA-8, an agonistic death receptor 5 antibody, involve modulation of the intrinsic apoptotic pathway in human breast cancer cells

TRA-8, a monoclonal antibody to death receptor 5 induces apoptosis in various cancer cells; however, the degree of sensitivity varies from highly sensitive to resistant. We have previously shown that resistance to TRA-8 can be reversed by using chemotherapeutic agents, but the mechanism underlying this sensitization was not fully understood. Here, we examined the combination of TRA-8 with doxorubicin or bortezomib in breast cancer cells. In TRA-8-resistant BT-474 and T47D cells, both chemotherapy agents synergistically sensitized cells to TRA-8 cytotoxicity with enhanced activation of apoptosis shown by cleavage of caspases and PARP, reduced Bid, increased proapoptotic Bcl-2 proteins, and increased mitochondrial membrane depolarization. Doxorubicin or bortezomib combined with TRA-8 also reduced Bcl-XL and X-linked inhibitors of apoptosis (XIAP) in treated cells. Furthermore, targeting these proteins with pharmacologic modulators, AT-101, BH3I-2' and AT-406, produced sensitization to TRA-8. TRA-8 combined with AT-101 or BH3I-2', inhibitors of antiapoptotic Bcl-2 proteins, produced synergistic cytotoxicity against ZR-75-1, BT-474, and T47D cells. The IAP-targeting compound, AT-406, was synergistic with TRA-8 in BT-474 cells, and to a lesser extent T47D cells. Activation of the intrinsic apoptotic pathway was a common mechanism associated with sensitization of TRA-8-resistant breast cancer cell lines. Collectively, these studies show that the Bcl-2 and IAP families of proteins are involved in TRA-8 and chemotherapy resistance via their modulation of the intrinsic apoptotic pathway. Targeting these proteins with novel agents sensitized TRA-8-resistant breast cancer cells, suggesting this approach may represent a potent therapeutic strategy in the treatment of breast cancer.     

Clinical Efficacy of Chemotherapy Combined with Verapamil in Metastatic Colorectal Patients

In order to determine the clinical efficacy and adverse reactions of chemotherapy and verapamil infusion through a target artery to treat colorectal cancer patients with metastasis after failure with previous conventional treatments. Patients with metastatic colon cancer (n = 36) received an infusion of verapamil, interleukin-2, oxaliplatin (or hydroxy camptothecin or irinotecan hydrochloride), fluorouracil and calcium folinate through target artery using the Seldinger puncture technique. From the second day of infusion, the patients were treated with fluorouracil and calcium folinate via systematic intravenous injection for 2–3 days. Efficacy was evaluated after at least two treatment courses. The objective response including complete or partial response was 58.3% in the 36 patients; clinical benefit rate, evaluated by Karnofsky Performance Status score was 91.7%; by weight was 83.3%; by the amount of painkiller consumed was 80.6%. No patient experienced side effects associated with heart function. Post-treatment, the P–R period, Q–T period, QRS, and heart rate were not significantly different than before treatment. Liver function was significantly improved. Side effects of chemotherapy were minor in comparison to those observed with intravenous chemotherapy. Infusion of verapamil and chemotherapy directly into pelvic tumor tissue can increase treatment efficacy and has been shown to be a relatively safe technique.     

Caveolin-1 plays a crucial role in inhibiting neuronal differentiation of neural stem/progenitor cells via VEGF signaling-dependent pathway

In the present study, we aim to elucidate the roles of caveolin-1(Cav-1), a 22 kDa protein in plasma membrane invaginations, in modulating neuronal differentiation of neural progenitor cells (NPCs). In the hippocampal dentate gyrus, we found that Cav-1 knockout mice revealed remarkably higher levels of vascular endothelial growth factor (VEGF) and the more abundant formation of newborn neurons than wild type mice. We then studied the potential mechanisms of Cav-1 in modulating VEGF signaling and neuronal differentiation in isolated cultured NPCs under normoxic and hypoxic conditions. Hypoxic embryonic rat NPCs were exposed to 1% O₂ for 24 h and then switched to 21% O₂ for 1, 3, 7 and 14 days whereas normoxic NPCs were continuously cultured with 21% O₂. Compared with normoxic NPCs, hypoxic NPCs had down-regulated expression of Cav-1 and up-regulated VEGF expression and p44/42MAPK phosphorylation, and enhanced neuronal differentiation. We further studied the roles of Cav-1 in inhibiting neuronal differentiation by using Cav-1 scaffolding domain peptide and Cav-1-specific small interfering RNA. In both normoxic and hypoxic NPCs, Cav-1 peptide markedly down-regulated the expressions of VEGF and flk1, decreased the phosphorylations of p44/42MAPK, Akt and Stat3, and inhibited neuronal differentiation, whereas the knockdown of Cav-1 promoted the expression of VEGF, phosphorylations of p44/42MAPK, Akt and Stat3, and stimulated neuronal differentiation. Moreover, the enhanced phosphorylations of p44/42MAPK, Akt and Stat3, and neuronal differentiation were abolished by co-treatment of VEGF inhibitor V1. These results provide strong evidence to prove that Cav-1 can inhibit neuronal differentiation via down-regulations of VEGF, p44/42MAPK, Akt and Stat3 signaling pathways, and that VEGF signaling is a crucial target of Cav-1. The hypoxia-induced down-regulation of Cav-1 contributes to enhanced neuronal differentiation in NPCs.     

Effects of elevated CO2 on the interspecific competition between two sympatric species of Aphis gossypii and Bemisia tabaci fed on transgenic Bt cotton

Abstract Effects of elevated CO₂ (twice ambient vs. ambient) and Bt Cry1Ac transgene (Bt cotton cv. 33^B vs. its nontransgenic parental line cv. DP5415) on the interspecific competition between two ecologically similar species of cotton aphid Aphis gossypii and whitefly biotype‐Q Bemisia tabaci were studied in open‐top chambers. The results indicated that elevated CO₂ and Bt cotton both affected the population abundances of A. gossypii and biotype‐Q B. tabaci when introduced solely (i.e., without interspecific competition) or two species coexisted (i.e., with interspecific competition). Compared with ambient CO₂, elevated CO₂ increased the population abundances of A. gossypii and biotype‐Q B. tabaci as fed on Bt and nontransgenic cotton on 45 (i.e., seedling stage) and 60 (i.e., flowering stage) days after planting (DAP), but only significantly enhanced aphid abundance without interspecific competition on the 45‐DAP nontransgenic cotton and 60‐DAP Bt cotton, and significantly increased whitefly abundance with interspecific competition on the 45‐DAP Bt cotton and 60‐DAP nontransgenic cotton. In addition, compared with nontransgenic cotton at elevated CO₂, Bt cotton significantly reduced biotype‐Q B. tabaci abundances without and with interspecific competition during seedling and flowering stage, while only significantly decreasing A. gossypii abundances without interspecific competition during the seedling stage. When the two insect species coexisted, the proportions of biotype‐Q B. tabaci were significantly higher than those of A. gossypii on Bt and nontransgenic cotton at the same CO₂ levels, and elevated CO₂ only significantly increased the percentages of biotype‐Q B. tabaci and significantly reduced the proportions of A. gossypii on seedling and flowering nontransgenic cotton. Therefore, the effects of elevated CO₂ were favorable for biotype‐Q B. tabaci to out‐compete A. gossypii under the predicted global climate change.     

Effects of selected insecticides on adults of two parasitoid species of Liriomyza trifolii: Ganaspidium nigrimanus (Figitidae) and Neochrysocharis formosa (Eulophidae)

Abstract Liriomyza trifolii is an important pest of vegetables and ornamental crops around the world. This pest is attacked by many parasitoid species. The principal management tactic used against L. trifolii is insecticide application. Insecticides vary in their effects on parasitoid species and insecticides that have less harmful effects should be preferred for the control of this pest. In this study, novaluron, abamectin, λ‐cyhalothrin and spinetoram were investigated for their lethal effects on adults of Neochrysocharis formosa and Ganaspidium nigrimanus, two important parasitoids of L. trifolii. Three different bioassays were used on adult parasitoids: direct insecticide application, insecticide intake and insecticide residue. Adult parasitoid response to novaluron exhibited the least lethal effects among the bioassays and insecticides tested. Abamectin had significant mortality to both parasitoid species in the direct application and insecticide intake bioassays and mortality were high for G. nigrimanus in the residue bioassay. Spinetoram was the most harmful insecticide to the adult parasitoids in all three bioassays. λ‐cyhalothrin effects varied between the two parasitoids. In the direct application, it was harmful to G. nigrimanus and had no effect on N. formosa. In the insecticide intake bioassay λ‐cyhalothrin had no effect in survival of either species, and in the residue bioassay it reduced parasitoid survival of both species. Potential tolerance of N. formosa to λ‐cyhalothrin is discussed.     

Develope monoclonal antibody against foot-and-mouth disease virus a type

In order to develop an anti-FMDV A Type monoclonal antibo by (mAb),BABL/c mice were immunized with FMDV A type.Monoclonal antibodies (mAbs) 7B11 and 8H4 against Foot-and-mouth disease virus (FMDV) serotype A were produced by fusing SP2/O myeloma cells with splenocyte from the mouse immunized with A/AV88.The microneutralization titer of the mAbs 7B11 and 8H4 were 1024 and 512,respectively.Both mAbs contain kappa light chains,the mAbs were IgG1.In order to define the mAbs binding epitopes,the reactivity of these mAbs against A Type FMDV,were examined using indirect ELISA,the result showed that both mAbs reacted with A Type FMDV.These mAbs may be used for further vaccine studies,diagnostic methods,prophylaxis,etiological and immunological research on FMDV.Characterization of these ncindicated that prepared anti-FMDV A mAbs had no cross-reactivity with Swine Vesicular Disease (SVD) or FMDV O,Asial and C Type antigens.Their titers in abdomen liquor were 1:5×106 and 1:2×106,respectively.7B11 was found to be of subtype IgG1,8H4 was classified as IgG2b subtype.The mAbs prepared in this study,are specific for detection of FMDV serotype A,and is potentially useful for pen-side diagnosis.