Experimental DNA vaccine against filariasis

Filarial infections are major causes of morbidity in the tropics and sub-tropics, afflicting over 150 million people in about 80 countries, causing debilitating symptoms such as elephantiasis (lymphatic filariasis), dermatitis and blindness (onchocerciasis or river blindness). Current control of lymphatic filariais relies on mass drug treatment with diethylcarbamazine（DEC) and albendazole while ivermectin is used against onchocerciasis. Repeat treatment is frequently required and this highlights the possibility of development of drug resistance. In addition, risk of adverse reactions following treatment excludes some patients from control programmes. Such circumstances urgently call for the development of complementary control measures such as vaccination. DNA vaccines are novel type of subunit vaccine in which production of the immunizing antigen is induced in host cells after introduction of a plasmid or recombinant viral vector containing the gene that encodes the antigen. DNA vaccines are relatively simple and cheap to produce and their stability makes them particularly suitable for use in remote regions that lack the cold-chain storage facilities required of conventional vaccines. Filarial nematodes are tissue-dwelling parasites that survive for many years in immunocompetent hosts. It is proposed that this longevity is, in part, due to the capacity of the parasites to modulate potentially lethal Th2 responses of their hosts. Consequently, the efficiency of a filarial vaccine may depend on how well it circumvents filarial-driven immunosuppression. To test this hypothesis, a series of DNA vaccines were developed and tested in the Litomosoides sigmodontis-mouse model of filarial infections. The L. sigmodontis Abundant Larval Transcript-1 (Ls-ALT) and Cysteine Protease Inhibitor (Ls-CPI) genes were cloned and genetically engineered to ablate their immunomodulatory properties by deleting the acid domain and by site mutation, respectively. In addition, the L. sigmodontis Venom Allergen Homologue (Ls-VAH) and Thioredoxin Peroxidase (Ls-TPX) were used in their native forms as vaccine targets. To improve immunisation and antigen processing by the host, these parasite genes were fused to a DNA sequence encoding an antibody that specifically binds dendritic cell surface protein (αDEC205 single chain Fv). DNA plasmids carrying mutated forms and/or anti-DEC205 were then co-administered with plasmids encoding the Th2 promoting cytokine IL4, and/or antigen-presenting cell activating MIP1α and Flt3L. Mice immunized with mutated forms (ADDALT and CPImu) of parasite antigens produced more specific antibody post-challenge and showed strongly increased lymphocyte stimulation above controls immunized with the native form. The immune response was further enhanced when plasmids encoding IL4, MIP1α, Flt3L and anti-DEC-205 forms were co-administered, resulting in production of a Th2/IgG1 phenotype. Significant reduction of worm burden (82.3%) was achieved by a cocktail vaccination which combined the ADDALT and CPImu candidates. Mice immunized with Ls-VAH and Ls-TPX DNA carried by αDEC205 elicited Th2-biased responses with up-regulated IgG1 and IgE antibodies as well as enhanced IL5, IL4, and IL13 and diminished IFNγ production compared to controls. The immune responses were further driven towards the Th2/IgG1 phenotype when Ls-VAH and Ls-TPX were injected with plasmids encoding ADDALT and CPImu and with the adjuvants Flt3L, MIP1α and IL4. This resulted in reduction in worm burden of 55.7% (cocktail vaccine containing Ls-VAH) and 41.6% (cocktail vaccine containing Ls-TPX) respectively in vaccinated animals

Fabrication of a novel hierarchical fibrous scaffold for breast cancer cell culture

Supplementary data to this article can be found online at https://doi.org/10.1016/j.polymertesting.2019.106107.Scaffolds combining nano- and submicro-fibers closely mimicking extracellular matrix (ECM) have been poorly exploited for in vitro cancer cell culture. Herein, a combined electrospinning and modified in situ biosynthesis method has been developed to fabricate a novel scaffold consisting of bacterial cellulose (BC) nanofibers and electrospun cellulose acetate (CA) submicrofibers to mimic the fibrillar structure of natural ECM. The CA/BC nano/submicrofibrous scaffold was characterized by scanning electron microscopy (SEM), mechanical strength tests, porosity measurements, and cell studies using the MCF-7 breast cancer cells. In addition, the sensitivity of the cancer cells seeded in the CA/BC nano/submicrofibrous scaffold to an anticancer drug was assessed. It was found that the CA/BC scaffold exhibited an interconnected porous structure in which BC nanofibers penetrated into the submicrofibrous CA scaffold. Such sophisticated structure was responsible for the improved mechanical properties of CA/BC scaffold over the ones obtained using a single kind of fibers. More importantly, the CA/BC scaffold showed improved cell adhesion, migration, and proliferation over single BC or CA scaffold. Finally, cells grown on CA/BC scaffold exhibited a greater doxorubicin resistance than those on single CA or BC scaffold. The results suggest that the CA/BC nano/submicrofibrous scaffold has potential for application in in vitro tumor model for the study of cancer progression and drug screening.This work was supported by the Key Project of Natural Science Foundation of Jiangxi Province (Grant no. 20161ACB20018), the National Natural Science Foundation of China (Grant nos. 31870963, 31660264, and 51572187), the Youth Science Foundation of Jiangxi Province (Grant no. 20181BAB216010), and the Science and Technology Research Project of Jiangxi Education Department (Grant no. GJJ180348).info:eu-repo/semantics/publishedVersio

Ubiquitination Is Required for Effective Replication of Coxsackievirus B3

BACKGROUND: Protein ubiquitination and/or degradation by the ubiquitin/proteasome system (UPS) have been recognized as critical mechanisms in the regulation of numerous essential cellular functions. The importance of the UPS in viral pathogenesis has become increasingly apparent. Using murine cardiomyocytes, we have previously demonstrated that the UPS plays a key role in the replication of coxsackievirus B3 (CVB3), an important human pathogen associated with various diseases. To further elucidate the underlying mechanisms, we examined the interplay between the UPS and CVB3, focusing on the role of ubiquitination in viral lifecycle. METHODOLOGY/PRINCIPAL FINDINGS: As assessed by in situ hybridization, Western blot, and plaque assay, we showed that proteasome inhibition decreased CVB3 RNA replication, protein synthesis, and viral titers in HeLa cells. There were no apparent changes in 20S proteasome activities following CVB3 infection. However, we found viral infection led to an accumulation of protein-ubiquitin conjugates, accompanied by a decreased protein expression of free ubiquitin, implicating an important role of ubiquitination in the UPS-mediated viral replication. Using small-interfering RNA, we demonstrated that gene-silencing of ubiquitin significantly reduced viral titers, possibly through downregulation of protein ubiquitination and subsequent alteration of protein function and/or degradation. Inhibition of deubiquitinating enzymes apparently enhances the inhibitory effects of proteasome inhibitors on CVB3 replication. Finally, by immunoprecipitation, we showed that coxsackieviral polymerase 3D was post-translationally modified by ubiquitination and such modification might be a prerequisite for its function in transcriptional regulation of viral genome. CONCLUSION: Coxsackievirus infection promotes protein ubiquitination, contributing to effective viral replication, probably through ubiquitin modification of viral polymerase

Incorporating graphene oxide into biomimetic nano-microfibrous cellulose scaffolds for enhanced breast cancer cell behavior

Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10570-020-03078-w) contains supplementary material, which is available to authorized users.The impact of graphene oxide (GO) on normal cells has been widely investigated. However, much less is known on its effect on cancer cells. Herein, GO nanosheets were incorporated into electrospun cellulose acetate (CA) microfibers. The GO-incorporated CA (GO/CA) microfibers were combined with bacterial cellulose (BC) nanofibers via in situ biosynthesis to obtain the nano-microfibrous scaffolds. The GO/CA-BC scaffolds were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS). The GO/CA-BC scaffolds were used for breast cancer cell culture to evaluate the effect of GO on cancer cell behavior. Fluorescence images revealed large multicellular clusters on the surface of GO/CA-BC scaffolds. Compared to the bare CA-BC scaffold, the GO/CA-BC scaffolds not only showed enhanced mechanical properties but also improved cell proliferation. It is expected that the GO/CA-BC scaffolds would provide a suitable microenvironment for the culture of cancer cells which is necessary for drug screening and cell biology study.This work was supported by National Natural Science Foundation of China (Grant nos. 51572187, 51973058, 31660264, 31870963), the Key Research and Development Program of Jiangxi Province (No. 20192ACB80008), and the Youth Science Foundation of Jiangxi Province (No. 20181BAB216010), and Key Project of Natural Science Foundation of Jiangxi Province (No. 20161ACB20018).info:eu-repo/semantics/publishedVersio

Whole-genome transcription and DNA methylation analysis of peripheral blood mononuclear cells identified aberrant gene regulation pathways in systemic lupus erythematosus

Hypermethylated CpG sites in PBMC of SLE patients comparing normal controls by genome-wide DNA methylation analysis. Three group comparisons, SLE LN+ vs. NC (S3-1), SLE LN− vs. NC (S3-2), and SLE (LN− and LN+) vs. NC (S3-3), were performed to identify hypermethylated DNA CpG sites by fold change (FC) > 1.2 and q value < 5% within each group. The 1813 common hypermethylated sites identified in each of the three group comparisons were then selected and listed in S3-4. (XLSX 814 kb

Heparinization and hybridization of electrospun tubular graft for improved endothelialization and anticoagulation

Supplementary data to this article can be found online at https://doi.org/10.1016/j.msec.2020.111861.Constructing biomimetic structure and immobilizing antithrombus factors are two effective methods to ensure rapid endothelialization and long-term anticoagulation for small-diameter vascular grafts. However, few literatures are available regarding simultaneous implementation of these two strategies. Herein, a nano-micro-fibrous biomimetic graft with a heparin coating was prepared via a step-by-step in situ biosynthesis method to improve potential endothelialization and anticoagulation. The 4-mm-diameter tubular graft consists of electrospun cellulose acetate (CA) microfibers and entangled bacterial nanocellulose (BNC) nanofibers with heparin coating on dual fibers. The hybridized and heparinized graft possesses suitable pore structure that facilitates endothelia cells adhesion and proliferation but prevents infiltration of fibrous tissue and blood leakage. In addition, it shows higher mechanical properties than those of bare CA and hybridized CA/BNC grafts, which match well with native blood vessels. Moreover, this dually modified graft exhibits improved blood compatibility and endothelialization over the counterparts without hybridization or heparinization according to the testing results of platelet adhesion, cell morphology, and protein expression of von Willebrand Factor. This novel graft with dual modifications shows promising as a new small-diameter vascular graft. This study provides a guidance for promoting endothelialization and blood compatibility by dual modifications of biomimetic structure and immobilized bioactive molecules.This work was supported by the National Natural Science Foundation of China (grant nos. 51973058 and 31870963), the Key Research and Development Program of Jiangxi Province (No. 20192ACB80008), and the Key Project of Natural Science Foundation of Jiangxi Province (20202ACBL204013).info:eu-repo/semantics/publishedVersio

Controllable synthesis of biomimetic nano/submicro-fibrous tubes for potential small-diameter vascular grafts

Mimicking the morphological structure of native blood vessels is critical for the development of vascular grafts. Herein, small-diameter composite vascular grafts that integrate the nanofibrous bacterial cellulose (BC) and submicrofibrous cellulose acetate (CA) were fabricated via a combined electrospinning and step-by-step in situ biosynthesis. Scanning electron microscopy (SEM) observation shows the nano/submicro-fibrous morphology and well-interconnected porous structure of the BC/CA grafts. It is found that the BC/CA graft with a suitable BC content demonstrates lower potential of thrombus formation and enhanced endothelialization as compared to the BC and CA counterparts. Western blotting and RT-qPCR results suggest that the BC/CA-2 graft promotes endothelialization by improving expressions of genes vWF-1 and CD31 and protein CD31. The in vivo tests demonstrate much lower inflammatory response to the BC/CA graft. These results suggest that the BC/CA graft shows a great potential as an artificial graft for rapid formation of an endothelial cell monolayer.National Natural Science Foundation of China (Grant no. 51973058, 31870963, 31760265) and Key Research and Development Program of Jiangxi Province (No. 20192ACB80008)info:eu-repo/semantics/publishedVersio