Biology subjects

Ouyang, H.

Publications and source records attributed to Ouyang, H..

6 recordsLinked to original sources

bioRxiv2018

Development of whole-porcine monoclonal antibodies with potent neutralization activity against classical swine fever virus (CSFV) from single B cells

Classical swine fever (CSF) is a highly contagious swine disease found worldwide that has caused devastating economic losses. However, there are few efficacious mAbs against the CSF virus (CSFV) that can be used for treatment because most mAbs against CSFV are derived from mouse hybridoma cells and these murine mAbs have disadvantages of inefficient effector functions elicitations and high immunogenicity in vivo. Accordingly, we characterized whole-porcine anti-CSFV neutralizing mAbs (NAbs) isolated directly from single B cells sorted from a CSFV-vaccinated pig using the fluoresceinated conserved linear neutralizing epitope of the CSFV E2 protein and fluorophore conjugated goat anti-pig IgG. Immunoglobulin (Ig) genes were isolated via nested PCR, and two porcine mAbs termed HK24 and HK44 were produced. We determined that these mAbs can bind to E2 protein and recognize sites within this major antigenic epitope. In addition, we found that mAbs HK24 and HK44 exhibit potent neutralizing activity against CSFV, and they can protect PK-15 cells from infections in vitro with potent IC50 values of 9.3 g/ml and 0.62 g/ml, respectively. Notably, we demonstrated that these two mAbs can be used as novel reagents for detecting virus infection. These data suggest that our results not only provide a method for efficiently obtaining mAbs against CSFV but also offer promising mAb candidates for development of antibody-based diagnostic and antiviral agents.\n\nImportanceNeutralizing monoclonal antibodies (NAbs) can prevent and may slow the spread of virus infection. The discovery of NAbs that recognize classical swine fever virus (CSFV) necessitates new technologies because the NAbs produced by immunization and hybridoma technology could not be transferred to in vivo research. Multiple full-length human therapeutic antibodies have been produced via single-cell polymerase chain reactions but whole-porcine NAbs for CSFV have not been generated. In this study, two whole-porcine mAbs, named HK24 and HK44, were isolated from epitope-specific single B cells. We demonstrate that these two mAbs have potent neutralizing activity against CSFV and can protect cells against viral infection. Therefore, they may facilitate the development of vaccines or antiviral drugs that offer the advantages of stability and low immunogenicity.

molecular biology↗

bioRxiv2018

Genetically modified pigs are protected from classical swine fever virus

Classical swine fever (CSF) caused by classical swine fever virus (CSFV) is among the most detrimental diseases, and leads to significant economic losses in the swine industry. Despite efforts by many government authorities try to stamp out the disease from national pig populations, the disease remains widespread. Here, antiviral small hairpin RNAs (shRNAs) were selected and then inserted at the porcine ROSA26 (pROSA26) locus via a CRISPR/Cas9-mediated knock-in strategy. Finally, anti-CSFV transgenic (TG) pigs were produced by somatic nuclear transfer (SCNT). Importantly, in vitro and in vivo viral challenge assays demonstrated that these TG pigs could effectively limit the growth of CSFV and reduced CSFV-associated clinical signs and mortality, and the disease resistance was stably transmitted to F1-generation. The use of these TG pigs can improve the well-being of livestock and substantially reduce virus-related economic losses. Additionally, this antiviral approach may provide a reference for future antiviral research.\n\nAuthor summaryClassical swine fever (CSF), caused by classical swine fever virus (CSFV), and is a highly contagious, often fatal porcine disease with significant economic losses. Due to its economic importance to the pig industry, the biology and pathogenesis of CSFV have been investigated extensively. Despite efforts by many government authorities to stamp out the disease from national pig populations, the disease remains widespread in some regions and seems to be waiting for the reintroduction and the next round of disease outbreaks. These highlight the necessity and urgency of developing more effective approaches to eradicate the challenging CSFV. In this study, we successfully produced anti-CSFV transgenic pigs and confirmed that these transgenic pigs could effectively limit the growth of CSFV in vivo and in vitro and that the disease resistance traits in the TG founders can be stably transmitted to their F1-generation offspring. This study suggests that these TG pigs can improve the well-being of livestock and contribute to offer potential benefits over commercial vaccination. The use of these TG pigs can improve the well-being of livestock and substantially reduce CSFV-related economic losses.

genomics↗

bioRxiv2018

High resolution profile of body wide pathological changes induced by abnormal elastin metabolism in Loxl1 knockout mice

Abnormal ECM caused serious body wide diseases and elastin is one of the important ECM components. But its systemic function still has not yet been thoroughly illustrated due to limitations related to novel research technologies. To uncover the functions of elastin, a new method for body-wide organ transcriptome profiling, combined with single-cell mass cytometry of the blood, was developed. A body-wide organ transcriptomic (BOT) map was created by performing RNA-seq of 17 organs from both Loxl1 knockout (KO) and wide type (WT) mice. The BOT results showed a systematic up-regulation of genes related to immune response and proliferation process in multiple tissues of the KO mice; histological and immune staining also confirmed the hyperplasia and infiltration of local immune cells in the vagina, small intestine, and liver tissues of KO mice. Furthermore, using 32 markers, CYTOF mass cytometry analysis of the immune cell subpopulations from the peripheral blood revealed apparent systemic immune changes in the KO mice; data showed an activated NK cells and T cells with a higher expression of CD44 and CD38, and a suppressed B cells, macrophages and neutrophils with lower expressions of CD62L, CD44 and IL6. More interestingly, these findings also correlated well with the data obtained from cancer patient databases; tumor patients had higher mutation frequency of Loxl1, and the Loxl1-mutant tumor patients had up-regulated immune process, cell proliferation and decreased survival rate. Thus, this research provided a powerful strategy to screen body-wide organ functions of a particular gene; the findings also illustrated the important biological roles of elastin on multiple organ cells and systemic immunity. These strategy and discoveries are both of important value for the understanding of ECM biology and multi-organ cancer pathology.

cancer biology↗

bioRxiv2018

Identification and Structure-Activity Relationship of HDAC6 Zinc-finger Ubiquitin Binding Domain Inhibitors

HDAC6 plays a central role in the recruitment of protein aggregates for lysosomal degradation, and is a promising target for combination therapy with proteasome inhibitors in multiple myeloma. Pharmacologically displacing ubiquitin from the zinc-finger ubiquitin-binding domain (ZnF-UBD) of HDAC6 is an underexplored alternative to catalytic inhibition. Here, we present the discovery of a HDAC6 ZnF-UBD-focused chemical series and its progression from virtual screening hits to low micromolar inhibitors. A carboxylate mimicking the C-terminal extremity of ubiquitin, and an extended aromatic system stacking with W1182 and R1155 are necessary for activity. One of the compounds induced a conformational remodeling of the binding site where the primary binding pocket opens-up onto a ligand-able secondary pocket that may be exploited to increase potency. The preliminary structure-activity relationship accompanied by nine crystal structures should enable further optimization into a chemical probe to investigate the merit of targeting the ZnF-UBD of HDAC6 in multiple myeloma and other diseases.

biophysics↗

bioRxiv2018

Cell atlas of human uterus

The human uterus is a highly dynamic tissue that undergoes repeated damage repair and regeneration during the menstrual cycle, which make it ideal model to study tissue regeneration and pathological process. Stem/progenitors were speculated to be involved in the regeneration of endometrial epithelial and pathogenesis of endometriosis. But the identity, microenvironment and regulatory mechanisms of the uterus epithelial stem/progenitors in vivo remain unclear. Here, we dissected the cell heterogeneities of the full-thickness human uterus epithelial cells (11 clusters), stroma cells (6 clusters), endothelial cells (5 clusters), smooth muscle cells (2 clusters), myofibroblasts (2 clusters) and immune cells (6 clusters) from 2735 single cell by single cell RNA-seq. Further analysis identified a unique ciliated epithelial cell cluster showing characteristics of stem/progenitors with properties of epithelial-mesenchymal transition (EMT) that mainly localized in the upper functionalis of the endometrium. Ordering the cell subpopulations along the pseudo-space revealed cell clusters possess cellular states of stress, inflammation and apoptosis in the upper functionalis cellular ecosystem of the endometrium. Connectivity map between the human uterus subpopulations revealed potential inflammatory (cytokines and chemokines) and developmental (WNT, FGF, VEGF) signals within the upper functionalis cellular ecosystem of the endometrium, especially from other epithelial clusters, regulating cell plasticity of the EMT-epithelial clusters. This study reconstructed the heterogeneities, space-specific distribution and connectivity map of human uterus atlas, which would provide insight in the regeneration of uterus endometria and reference for the pathogenesis of uterus.

cell biology↗

bioRxiv2018

Local Delivery of Stromal Cell-Derived Factor-1α Improves the Pregnancy Rate of Injured Uterus through the Promotion of Endometrial and Vascular Regeneration

Severe infection and mechanical injury of the uterus may lead to infertility and miscarriage. Currently, there is a lack of effective treatment modality for functional repair of uterine injury. To address this clinical challenge, this study aimed to develop a chemotactic composite scaffold by incorporating recombinant human stromal cell-derived factor-1 (rhSDF-1) into a silk fibroin-bacterial cellulose (SF-BC) membrane carrier. A rat model of uterine injury was utilized for this study, which was composed of three groups: blank control, implantation with SF-BC only or SF-BC loaded with rhSDF-1. The tissue regeneration efficacy of the three groups were analyzed and compared. The results showed that SF-BC loaded with rhSDF-1 significantly enhanced endometrial regeneration and arteriogenesis of the injured rat uterus, which led to improved pregnancy outcomes, thus indicating much promise for functional uterine repair and regeneration.

bioengineering↗