Base editing with high efficiency in allotetraploid oilseed rape by A3A‐PBE system

CRISPR/Cas‐base editing is an emerging technology that could convert a nucleotide to another type at the target site. In this study, A3A‐PBE system consisting of human A3A cytidine deaminase fused with a Cas9 nickase and uracil glycosylase inhibitor was established and developed in allotetraploid Brassica napus. We designed three sgRNAs to target ALS, RGA and IAA7 genes, respectively. Base‐editing efficiency was demonstrated to be more than 20% for all the three target genes. Target sequencing results revealed that the editing window ranged from C1 to C10 of the PAM sequence. Base‐edited plants of ALS conferred high herbicide resistance, while base‐edited plants of RGA or IAA7 exhibited decreased plant height. All the base editing could be genetically inherited from T₀ to T₁ generation. Several Indel mutations were confirmed at the target sites for all the three sgRNAs. Furthermore, though no C to T substitution was detected at the most potential off‐target sites, large‐scale SNP variations were determined through whole‐genome sequencing between some base‐edited and wild‐type plants. These results revealed that A3A‐PBE base‐editing system could effectively convert C to T substitution with high‐editing efficiency and broadened editing window in oilseed rape. Mutants for ALS, IAA7 and RGA genes could be potentially applied to confer herbicide resistance for weed control or with better plant architecture suitable for mechanic harvesting.     

IL-37 overexpression enhances the therapeutic effect of endometrial regenerative cells in concanavalin A-induced hepatitis

Background aimsMesenchymal stromal cells and immunosuppressive factor IL-37 can both suppress concanavalin A (Con A)-induced hepatitis in mice. Endometrial regenerative cells (ERCs), novel types of mesenchymal-like stromal cells, possess powerful immunomodulatory effects and are effective in treating various diseases. The aim of this study was to explore the effects of ERCs in suppressing Con A-induced hepatitis and determine whether IL-37 overexpression could enhance the therapeutic effect of ERCs in this process.MethodsERCs were extracted from the menstrual blood of healthy female volunteer donors. The IL-37 gene was transferred into ERCs, and the expression of IL-37 in cells was detected by western blot and enzyme-linked immunosorbent assay. Hepatitis was induced by Con A in C57BL/6 mice that were randomly divided into groups treated with phosphate-buffered saline, ERCs, IL-37 or ERCs transfected with the IL-37 gene (IL-37-ERCs). Cell tracking, liver function, histopathological and immunohistological changes, immune cell proportions and levels of cytokines were measured 24 h after Con A administration.ResultsCompared with ERC or IL-37 treatment, IL-37-ERCs further reduced levels of liver enzymes (alanine aminotransferase and aspartate aminotransferase) and improved histopathological changes in the liver. In addition, IL-37-ERC treatment further reduced the proportions of M1 macrophages and CD4⁺ T cells and increased the proportion of regulatory T cells. Moreover, IL-37-ERC treatment resulted in lower levels of IL-12 and interferon gamma, and higher level of transforming growth factor beta.ConclusionsThe results of this study suggest that ERCs can effectively alleviate Con A-induced hepatitis. Furthermore, IL-37 overexpression can significantly enhance the therapeutic efficacy of ERCs by augmenting the immunomodulatory and anti-inflammatory properties of ERCs. This study may provide a promising strategy for treatment of T-cell-dependent hepatitis.     

Sparstolonin B exerts beneficial effects on prostate cancer by acting on the reactive oxygen species-mediated PI3K/AKT pathway

Prostate cancer is a major health concern in males worldwide, owing to its high incidence. Sparstolonin B (SsnB), a component of the Chinese herbal medicine Sparganium stoloniferum, is used to treat many diseases. However, the effects and mechanisms of action of SsnB in prostate cancer have not yet been reported. In this study, we evaluated the effects of SsnB on cellular processes and tumour growth. In particular, we verified that SsnB could inhibit the proliferation, migration and invasion of prostate cancer cells and induce apoptosis by activating G2/M phase arrest in vitro based on a series of cytological experiments. In vivo, we found that SsnB could inhibit tumour growth in nude mouse xenograft models. We further confirmed that SsnB could repress the PI3K/AKT pathway by increasing reactive oxygen species (ROS) accumulation and oxidative stress. Collectively, SsnB inhibits tumour growth and induces apoptosis in prostate cancer via the suppression of the ROS-mediated PI3K/AKT pathway and may be a new alternative to adjuvant therapy for prostate cancer.     

Prostaglandin E3 attenuates macrophage-associated inflammation and prostate tumour growth by modulating polarization

Alternative polarization of macrophages regulates multiple biological processes. While M1-polarized macrophages generally mediate rapid immune responses, M2-polarized macrophages induce chronic and mild immune responses. In either case, polyunsaturated fatty acid (PUFA)-derived lipid mediators act as both products and regulators of macrophages. Prostaglandin E₃ (PGE₃) is an eicosanoid derived from eicosapentaenoic acid, which is converted by cyclooxygenase, followed by prostaglandin E synthase successively. We found that PGE₃ played an anti-inflammatory role by inhibiting LPS and interferon-γ-induced M1 polarization and promoting interleukin-4-mediated M2 polarization (M2a). Further, we found that although PGE₃ had no direct effect on the growth of prostate cancer cells in vitro, PGE₃ could inhibit prostate cancer in vivo in a nude mouse model of neoplasia. Notably, we found that PGE₃ significantly inhibited prostate cancer cell growth in a cancer cell-macrophage co-culture system. Experimental results showed that PGE₃ inhibited the polarization of tumour-associated M2 macrophages (TAM), consequently producing indirect anti-tumour activity. Mechanistically, we identified that PGE₃ regulated the expression and activation of protein kinase A, which is critical for macrophage polarization. In summary, this study indicates that PGE₃ can selectively promote M2a polarization, while inhibiting M1 and TAM polarization, thus exerting an anti-inflammatory effect and anti-tumour effect in prostate cancer.     

Baicalein alleviates osteoarthritis by protecting subchondral bone,inhibiting angiogenesis and synovial proliferation

Osteoarthritis (OA) is one of the most frequent chronic joint diseases with the increasing life expectancy. The main characteristics of the disease are loss of articular cartilage, subchondral bone sclerosis and synovium inflammation. Physical measures, drug therapy and surgery are the mainstay of treatments for OA, whereas drug therapies are mainly limited to analgesics, glucocorticoids, hyaluronic acids and some alternative therapies because of single therapeutic target of OA joints. Baicalein, a traditional Chinese medicine extracted from Scutellaria baicalensis Georgi, has been widely used in anti-inflammatory therapies. Previous studies revealed that baicalein could alleviate cartilage degeneration effectively by acting on articular chondrocytes. However, the mechanisms involved in baicalein-mediated protection of the OA are not completely understood in consideration of integrality of arthrosis. In this study, we found that intra-articular injection of baicalein ameliorated subchondral bone remodelling. Further studies showed that baicalein could decrease the number of differentiated osteoblasts by inhibiting pre-osteoblasts proliferation and promoting pre-osteoblasts apoptosis. In addition, baicalein impaired angiogenesis of endothelial cells and inhibited proliferation of synovial cells. Taken together, these results implicated that baicalein might be an effective medicine for treating OA by regulating multiple targets.     

TGF-β isoforms inhibit hepatitis C virus propagation in transforming growth factor beta/SMAD protein signalling pathway dependent and independent manners

Transforming growth factor beta (TGF-β) plays an important role in the viral liver disease progression via controlling viral propagation and mediating inflammation-associated responses. However, the antiviral activities and mechanisms of TGF-β isoforms, including TGF-β1, TGF-β2 and TGF-β3, remain unclear. Here, we demonstrated that all of the three TGF-β isoforms were increased in Huh7.5 cells infected by hepatitis C virus (HCV), but in turn, the elevated TGF-β isoforms could inhibit HCV propagation with different potency in infectious HCV cell culture system. TGF-β isoforms suppressed HCV propagation through interrupting several different stages in the whole HCV life cycle, including virus entry and intracellular replication, in TGF-β/SMAD signalling pathway–dependent and TGF-β/SMAD signalling pathway–independent manners. TGF-β isoforms showed additional anti-HCV activities when combined with each other. However, the elevated TGF-β1 and TGF-β2, not TGF-β3, could also induce liver fibrosis with a high expression of type I collagen alpha-1 and α-smooth muscle actin in LX-2 cells. Our results showed a new insight into TGF-β isoforms in the HCV-related liver disease progression.     

Prolonged humoral and cellular immunity in COVID-19-recovered patients

By the beginning of 2021, the battle against coronavirus disease 2019 (COVID-19) remains ongoing. Investigating the adaptive immune response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes COVID-19, in patients who have recovered from this disease could contribute to our understanding of the natural host immune response. We enrolled 38 participants in this study. 7 healthy participants and 31 COVID-19 patients who had recovered from COVID-19 and categorized them into 3 groups according to their previous clinical presentations: 10 moderate, 9 mild, and 12 asymptomatic. Flow cytometry analysis of peripheral lymphocyte counts in recovered patients showed significantly increased levels of CD4⁺ T cells in patients with a history of mild and moderate COVID-19 symptoms compared with those healthy individuals (p < 0.05 and p < 0.0001 respectively). whereas no significant difference was observed in the CD8⁺ T cell percentage in COVID-19-recovered patients compared with healthy individuals. Our study demonstrated that antibodies against the SARS-CoV-2 spike protein (anti-S) IgG antibody production could be observed in all recovered COVID-19 patients, regardless of whether they were asymptomatic (p < 0.05)or presented with mild (p < 0.0001) or moderate symptoms (p < 0.01). Anti-S IgG antibodies could be detected in participants up to 90 days post-infection. In conclusion, the lymphocyte levels in recovered patients were associated with the clinical presentation of the disease, and further analysis is required to investigate relationships between different clinical presentations and lymphocyte activation and function.     

Superior japonica rice variety YJ144 with improved rice blast resistance,yield, and quality achieved using molecular design and multiple breeding strategies

Molecular Breeding - The stem color of young mung bean is a very useful tool in germplasm identification. Flowering time and plant height (PH) are known to be strongly correlated with crop adaption...