Epstein-Barr Virus_Encoded LMP1 Upregulates MicroRNA-21 to Promote the Resistance of Nasopharyngeal Carcinoma Cells to Cisplatin-Induced Apoptosis by Suppressing PDCD4 and Fas-L

Approximately 30% of patients with Epstein-Barr virus (EBV)-positive advanced nasopharyngeal carcinoma (NPC) display chemoresistance to cisplatin-based regimens, but the underlying mechanisms are unclear. The Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1), a functional homologue of the tumor necrosis factor receptor family, contributes substantially to the oncogenic potential of EBV through the activation of multiple signaling pathways, and it is closely associated with a poorer prognosis for NPC. Recent studies show that EBV infection can induce the expression of many cellular miRNAs, including microRNA-21, a biomarker for chemoresistance. However, neither a link between LMP1 expression and miR-21 upregulation nor their cross talk in affecting chemoresistance to cisplatin have been reported. Here, we observed that stable LMP1-transformed NPC cells were less sensitive to cisplatin treatment based on their proliferation, colony formation, the IC₅₀ value of cisplatin and the apoptosis index. Higher levels of miR-21 were found in EBV-carrying and LMP1-positive cell lines, suggesting that LMP1 may be linked to miR-21 upregulation. These data were confirmed by our results that exogenous LMP1 increased miR-21 in both transiently and stably LMP1-transfected cells, and the knock down of miR-21 substantially reversed the resistance of the NPC cells to cisplatin treatment. Moreover, the proapoptotic factors programmed cell death 4 (PDCD4) and Fas ligand (Fas-L), which were negatively regulated by miR-21, were found to play an important role in the program of LMP1-dependent cisplatin resistance. Finally, we demonstrated that LMP1 induced miR-21 expression primarily by modulating the PI3K/AKT/FOXO3a signaling pathway. Taken together, we revealed for the first time that viral LMP1 triggers the PI3K/Akt/FOXO3a pathway to induce human miR-21 expression, which subsequently decreases the expression of PDCD4 and Fas-L, and results in chemoresistance in NPC cells.     

Gene expression profiles during differentiation and transdifferentiation of bovine myogenic satellite cells

The molecular mechanisms underlying myogenic satellite cells (MSCs) differentiation into myotube-formed cells (MFCs) and transdifferentiation into adipocyte-like cells (ALCs) are unclear. As a step towards understanding the molecular mechanisms underlying MSC differentiation and transdifferentiation, we attempted to identify the genes differentially expressed during differentiation and transdifferentiation using gene microarray analysis (GMA). Thirty oligonucleotide arrays were used with two technical replicates and nine and six biological replicates for MFCs vs. MSCs and ALCs vs. MSCs, respectively, to contrast expression profile differences. GMA identified 1,224 differentially expressed genes by at least 2-fold during differentiation and transdifferentiation of MSCs. To select the highly expressed genes for future functional study, genes with a 4-fold expression difference were selected for validation by real time RT-PCR and approximately 96.9% of the genes were validated. The up-regulation of marker genes for myogenesis (MYL2, MYH3) and adipogenesis (PPAR??, and FABP4) was observed during the differentiation and transdifferentiation of MSCs into MFCs and ALCs, respectively. KOG analysis revealed that the most of the genes up-regulated during differentiation and transdifferentiation of MSCs were related to signal transduction. Again the exact location of 109 differentially expressed genes by 4-fold were analyzed by chromosome mapping. Among those, co-localization of 29 genes up-regulated during transdifferentiation with QTL for marbling score and intramuscular fat percentage supports the involvement of these genes in cellular transdifferentiation. Interestingly, some genes with unknown function were also identified during the process. Functional studies on these genes may unfold the molecular mechanisms controlling MSC differentiation and transdifferentiation.     

Gualou Xiebai Banxia decoction ameliorates Poloxamer 407-induced hyperlipidemia

Ethnopharmacological relevance: Gualou Xiebai Banxia (GLXBBX) decoction is a well-known traditional Chinese herbal formula that was first discussed in the Synopsis of the Golden Chamber by Zhang Zhongjing in the Eastern Han Dynasty. In traditional Chinese medicine, GLXBBX is commonly prescribed to treat cardiovascular diseases, such as coronary heart disease and atherosclerosis.Objective: The present study aimed to examine GLXBBX’s preventative capacity and elucidate the potential molecular mechanism of Poloxamer 407 (P407)-induced hyperlipidemia in rats.Materials and methods: Both the control and model groups received pure water, and the test group also received a GLXBBX decoction. For each administration, 3 ml of the solution was administered orally. To establish hyperlipidemia, a solution mixed with 0.25 g/kg P407 dissolved in 0.9% normal saline was injected slowly into the abdominal cavity. At the end of the study, the rats’ plasma lipid levels were calculated using an automatic biochemical analyzer to evaluate the preventative capability of the GLXBBX decoction, and the serum and liver of the rats were collected.Results: The GLXBBX decoction significantly improved P407-induced hyperlipidemia, including increased plasma triglycerides (TGs), aspartate aminotransferase (AST) elevation, and lipid accumulation. Moreover, GLXBBX decoction treatment increased lipoprotein lipase (LPL) activity and mRNA expression of LPL. Furthermore, GLXBBX significantly suppressed the mRNA expression of stearoyl-CoA desaturase (SCD1).Conclusion: GLXBBX significantly improved P407-induced hyperlipidemia, which may have been related to enhanced LPL activity, increased LPL mRNA expression, and decreased mRNA expression of SCD1.     

Microtubule stability, Golgi organization, and transport flux require dystonin-a2-MAP1B interaction

Loss of function of dystonin cytoskeletal linker proteins causes neurodegeneration in dystonia musculorum (dt) mutant mice. Although much investigation has focused on understanding dt pathology, the diverse cellular functions of dystonin isoforms remain poorly characterized. In this paper, we highlight novel functions of the dystonin-a2 isoform in mediating microtubule (MT) stability, Golgi organization, and flux through the secretory pathway. Using dystonin mutant mice combined with isoform-specific loss-of-function analysis, we found dystonin-a2 bound to MT-associated protein 1B (MAP1B) in the centrosomal region, where it maintained MT acetylation. In dt neurons, absence of the MAP1B-dystonin-a2 interaction resulted in altered MAP1B perikaryal localization, leading to MT deacetylation and instability. Deacetylated MT accumulation resulted in Golgi fragmentation and prevented anterograde trafficking via motor proteins. Maintenance of MT acetylation through trichostatin A administration or MAP1B overexpression mitigated the observed defect. These cellular aberrations are apparent in prephenotype dorsal root ganglia and primary sensory neurons from dt mice, suggesting they are causal in the disorder.     

CHIP modulates APP‐induced autophagy‐dependent pathological symptoms in Drosophila

Dysregulation of autophagy is associated with the neurodegenerative processes in Alzheimer's disease (AD), yet it remains controversial whether autophagy is a cause or consequence of AD. We have previously expressed the full‐length human APP in Drosophila and established a fly AD model that exhibits multiple AD‐like symptoms. Here we report that depletion of CHIP effectively palliated APP‐induced pathological symptoms, including morphological, behavioral, and cognitive defects. Mechanistically, CHIP is required for APP‐induced autophagy dysfunction, which promotes Aβ production via increased expression of BACE and Psn. Our findings suggest that aberrant autophagy is not only a consequence of abnormal APP activity, but also contributes to dysregulated APP metabolism and subsequent AD pathogenesis.     

Improved method for production of antibodies against T-2 toxin and diacetoxyscirpenol in rabbits.

A new, improved approach for the production of antibodies against T-2 toxin and diacetoxyscirpenol (DAS) was developed. The method involves the use of immunogens which were prepared by conjugating O-carboxymethoxyl oxime (CMO) derivatives of both toxins to bovine serum albumin (BSA). Isomers a and b of CMO-T-2 toxin and isomer b of CMO-DAS were tested. Antibodies against both toxins were demonstrated as early as 4 weeks after immunization. a-CMO-T-2-BSA conjugate was a better immunogen than the b isomer, and the highest titers (6,000) were reached 14 weeks after immunization and one booster injection. Antibody titers for rabbits immunized with the b isomer of CMO-T-2 never reached more than 2,000. The specificity of antibodies obtained from rabbits after immunization with CMO-T-2-BSA was similar to that of hemisuccinate-T-2-BSA. Anti-b-T-2 antibodies had slightly higher cross-reactivity with H-T-2 toxin than did the antibody obtained from rabbits immunized with the conjugate of the a isomer. The relative cross-reactivities of anti-a-CMO-T-2 antibody with T-2, acetyl-T-2, H-T-2, T-2-triol, 3'-OH-T-2, and T-2 tetraol were 1, 4.5, 5.7, 250, 500, and 3,000, respectively. The relative cross-reactivities of anti-b-T-2 antibody with T-2, acetyl-T-2, H-T-2, and T-2 triol were 1, 2, 3, and 488, respectively. Antibodies against b-CMO-DAS showed a high degree of cross-reactivity with monoacetoxyscirpenols (MAS). The relative cross-reactivities of anti-B-DAS antibody with DAS, 4-MAS, 15-MAS, acetyl-deoxynivalenol, T-2-toxin, acetyl-T-2, and neosolaniol were 1, 4, 5, 76, 107, 147, and 266, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

The anti-tumor effects of the combination of microwave hyperthermia and lobaplatin against breast cancer cells in vitro and in vivo

Background: Breast cancer is the main lethal disease among females. The combination of lobaplatin and microwave hyperthermia plays a crucial role in several kinds of cancer in the clinic, but its possible mechanism in breast cancer has remained indistinct.Methods: Mouse models were used to detect breast cancer progression. Cell growth was explored with MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphonyl)-2H-tetrazolium) and colony formation assays. Cell migration and invasion were investigated with a transwell assay. Cell apoptosis was probed with flow cytometry. The expression of apoptosis-associated proteins was examined with Western blots.Result: Combination treatment decreased breast cancer cell viability, colony formation, cell invasion and metastasis. In addition, the treatment-induced breast cancer cell apoptosis and autophagy, activated the c-Jun N-terminal kinase (JNK) signaling pathway, suppressed the protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway, and down-regulated IAP and Bcl-2 family protein expression.Conclusion: These results indicate that lobaplatin is an effective breast cancer anti-tumor agent. Microwave hyperthermia was a useful adjunctive treatment. Combination treatment was more efficient than any single therapy. The possible mechanism for this effect was mainly associated with activation of the JNK signaling pathway, inactivation of the AKT/mTOR signaling pathway and down-regulation of the Bcl-2 and IAP families.     

pH heterogeneity at intracellular and extracellular plasma membrane sites in HT29-C1 cell monolayers

In the colonic mucosa, short-chain fatty acids changeintracellular pH (pH_i) and extracellular pH(pH_e). In this report, confocal microscopy anddual-emission ratio imaging of carboxyseminaphthorhodofluor-1 were usedfor direct evaluation of pH_i and pH_e in asimple model epithelium, HT29-C1 cells. Live cell imaging along theapical-to-basal axis of filter-grown cells allowed simultaneousmeasurement of pH in the aqueous environment near the apical membrane,the lateral membrane, and the basal membrane. Subapical cytoplasmreported the largest changes in pH_i after isosmoticaddition of 130 mM propionate or 30 mM NH₄Cl. In restingcells and cells with an imposed acid load, lateral membranes hadpH_i values intermediate between the relatively acidicsubapical region (pH 6.3-6.9) and the relatively alkaline basalpole of the cells (pH 7.4-7.1). Transcellular pH_igradients were diminished or eliminated during an induced alkalineload. Propionate differentially altered pH_e near the apicalmembrane, in lateral intracellular spaces between adjacent cells, andnear the basal membrane. Luminal or serosal propionate causedalkalinization of the cis compartment (where propionate wasadded) but acidification of the trans compartment only inresponse to luminal propionate. Addition of NH₄Cl produced qualitatively opposite pH_e excursions. The microscopicvalues of pH_i and pH_e can explain a portion ofthe selective activation of polarized Na/H exchangers observed inHT29-C1 cells in the presence of transepithelial propionate gradients.

     

Concurrent expression of heme oxygenase-1 and p53 in human retinal pigment epithelial cell line

Heme oxygenase-1 (HO-1) is a stress-responsive protein that is known to regulate cellular functions such as cell proliferation, inflammation, and apoptosis. Here, we investigated the effects of HO activity on the expression of p53 in the human retinal pigment epithelium (RPE) cell line ARPE-19. Cobalt protoporphyrin (CoPP) induced the expression of both HO-1 and p53 without significant toxicity to the cells. In addition, the blockage of HO activity with the iron chelator DFO or with HO-1 siRNA inhibited the CoPP-induced expression of p53. Similarly, zinc protoporphyrin (ZnPP), an inhibitor of HO, suppressed p53 expression in ARPE-19 cells, although ZnPP increased the level of HO-1 protein while inhibiting HO activity. Also, CoPP-induced p53 expression was not affected by the formation of reactive oxygen species (ROS). Based on these results, we conclude that HO activity is involved in the regulation of p53 expression in a ROS-independent mechanism, and also suggest that the expression of p53 in ARPE-19 cells is associated with heme metabolites such as biliverdin/bilirubin, carbon monoxide, and iron produced by the activity of HO.