Background Chikungunya virus (CHIKV), a serious health problem in several tropical countries, is the causative agent of chikungunya fever. Approved antiviral therapies or vaccines for the treatment or prevention of CHIKV infections are not available. As diverse natural phenolic compounds have been shown to possess antiviral activities, we explored the antiviral activity of α-Mangostin, a xanthanoid, against CHIKV infection. Methods The in vitro prophylactic and therapeutic effects of α-Mangostin on CHIKV replication in Vero E6 cells were investigated by administering it under pre, post and cotreatment conditions. The antiviral activity was determined by foci forming unit assay, quantitative RT-PCR and cell-based immune-fluorescence assay. The molecular mechanism of inhibitory action was further proposed using in silico molecular docking studies. Results In vitro studies revealed that 8 µM α-Mangostin completely inhibited CHIKV infectivity under the cotreatment condition. CHIKV replication was also inhibited in virus-infected mice. This is the first in vivo study which clearly showed that α-Mangostin is effective in vivo by significantly reducing virus replication in serum and muscles. Molecular docking indicated that α-Mangostin can efficiently interact with the E2–E1 heterodimeric glycoprotein and the ADP-ribose binding cavity of the nsP3 macrodomain. Conclusions The findings suggest that α-Mangostin can inhibit CHIKV infection and replication through possible interaction with multiple CHIKV target proteins and might act as a prophylactic/therapeutic agent against CHIKV.
Aortic dysfunctions (aneurysm, aortitis) lead to the most serious conditions related to aortic wall with life-threatening complications. The most common modality of management for such conditions is replacement (diseased part) of aorta by a larger diameter stent (reconstructive vascular surgery) which in itself is a big trial. The most natural way is to use a re-endothelized scaffold. Developing a scaffold with biomimetic properties is an experimental aim for most of the scientists and surgeons. We aim to structure a strategy to overcome the well-known problems associated with aorta. In this study, we plan to remold a larger diameter blood vessel such as aorta from xenogeneic origin using different protocols to decellularize and comparing them with normal aorta. The chemicals and enzymes used for bovine aorta decellularization are 1% SDS (group II), 70% ethanol + 0.25% trypsin (group III), 70% ethanol (group IV), and 0.25% trypsin (group V). Group I served as control (without decellularization). Histology and SEM study were conducted for cellular presence/absence in all scaffolds. Later, the scaffolds were coated with the fibrin glue (FG) and endothelial cells were proliferated over them. 3D images were taken showing the remolding of the endothelial cells on FG-coated surfaces. The re-endothelization was confirmed by lectin and vWF+/+ expression. Graft elasticity and burst pressure were confirmed by biomechanical tensile testing. Further, the absence of host tissue DNA and presence of cellular DNA after re-endothelialization were confirmed by PicoGreen assay. The acceptability for metabolically active cellular proliferation on scaffolds and its non-toxicity were proved by cell viability assay. Current findings accomplish that larger diameter aorta extracellular matrix scaffold (group II) can be fabricated and re-endothelialized to develop non-thrombotic surfaces with improved graft patency with promising results compared to other fabricated scaffold groups.
Background There is a significant pitfall in clinical translation of large-sized tissue-engineered grafts – a lack of vascularization. This study was carried out to find the answer in a plant leaf, as plants and animals share structural similarities. Methods and results We fabricated a scaffold using Brassica oleracea leaves (10%SDS) and expanded the endothelial cells onto them. The vascularity was demarcated by angiography. The thermal decomposition confirmed that the oxidation resistance of the scaffold is parallel to the natural leaf. The acellularity of the scaffold as well as the presence of cellular establishment after culture on the scaffold was confirmed by histology, scanning electron microscopy, periodic acid-Schiff, and DNA quantification. Further, we estimated various biochemical markers like MDA, catalase, total proteins, and total nitric oxide for confirming their metabolic activities. Cell-specific markers like vWF, lectin established their phenotype. Cytotoxicity and live-dead assay showed the viability of cells. Conclusion Our findings proved that the decellularized leaf scaffold preserves vascularity, exhibits non-toxicity, maintains the cell identity, and supports mammalian cells for their metabolic activities. The study gives a futuristic hope in combating the ever-growing issues of clinical applicability of large-sized grafts.
Various in vitro methods have been used for biological and synthetic scaffold fabrication. Some use polymers such as expanded polytetrafluoroethylene (ePTFE), polytetrafluoroethylene (PTFE), and polyethylene terephthalate (PET), while others use allogeneic or xenogeneic biological materials (e.g. blood vessels). While fabricating a biological scaffold, the first step is complete decellularization by enzymes (e.g. trypsin, collagenase, etc.) or chemicals (e.g. SDS, Triton-X, etc.), and the scaffolds should maintain its extracellular matrix (ECM). The second step involves re-endothelization so as to get fully biomimetic graft. In this study, we focused (concentrated) on the fabrication of a human saphenous vein scaffold by using various chemicals. We observed that cationic 1% SDS solution (Group B) performed excellent decellularization without altering the extracellular matrix as compared to the other chemicals like 0.25% trypsin and 70% ethanol (Groups C and D). Decellularization percentage and intactness of ECM (in all tunicae – intima, media, and adventitia) were confirmed based on histology. The PicoGreen assay showed that Group B (1% SDS decellularized scaffold, n = 3) had no detectable residual DNA. Re-endothelization on the complete decellularized scaffold (Group B) was done in both ways, without initial fibrin glue application (Group E) and with prior fibrin glue application (Group F). The vWF and lectin expressions suggested that endothelial cells did not alter their phenotype on human saphenous vein scaffolds. Uniaxial tensile testing revealed no significant differences in strain characteristics and modulus between native tissue and decellularized scaffolds. The live-dead (FDA/PI) and MTT assays confirmed the endothelial cell proliferation and viability, and the scanning electron microscope (SEM) data showed that the cells adhered to the scaffold matrix (Group F). We concluded that an allogeneic human saphenous vein scaffold with desirable properties can be fabricated and re-endothelialized to form a non-thrombogenic intimal surface in vitro using this protocol.
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