Macrophage migration inhibitory factor regulates joint capsule fibrosis by promoting TGF-β1 production in fibroblasts

Joint capsule fibrosis caused by excessive inflammation results in post-traumatic joint contracture (PTJC). Transforming growth factor (TGF)-β1 plays a key role in PTJC by regulating fibroblast functions, however, cytokine-induced TGF-β1 expression in specific cell types remains poorly characterized. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine involved in inflammation- and fibrosis-associated pathophysiology. In this study, we investigated whether MIF can facilitate TGF-β1 production from fibroblasts and regulate joint capsule fibrosis following PTJC. Our data demonstrated that MIF and TGF-β1 significantly increased in fibroblasts of injured rat posterior joint capsules. Treatment the lesion sites with MIF inhibitor 4-Iodo-6-phenylpyrimidine (4-IPP) reduced TGF-β1 production and relieved joint capsule inflammation and fibrosis. In vitro, MIF facilitated TGF-β1 expression in primary joint capsule fibroblasts by activating mitogen-activated protein kinase (MAPK) (P38, ERK) signaling through coupling with membrane surface receptor CD74, which in turn affected fibroblast functions and promoted MIF production. Our results reveal a novel function of trauma-induced MIF in the occurrence and development of joint capsule fibrosis. Further investigation of the underlying mechanism may provide potential therapeutic targets for PTJC.     

The herpesvirus accessory protein γ134.5 facilitates viral replication by disabling mitochondrial translocation of RIG-I

RIG-I and MDA5 are cytoplasmic RNA sensors that mediate cell intrinsic immunity against viral pathogens. While it has been well-established that RIG-I and MDA5 recognize RNA viruses, their interactive network with DNA viruses, including herpes simplex virus 1 (HSV-1), remains less clear. Using a combination of RNA-deep sequencing and genetic studies, we show that the γ₁34.5 gene product, a virus-encoded virulence factor, enables HSV growth by neutralization of RIG-I dependent restriction. When expressed in mammalian cells, HSV-1 γ₁34.5 targets RIG-I, which cripples cytosolic RNA sensing and subsequently suppresses antiviral gene expression. Rather than inhibition of RIG-I K63-linked ubiquitination, the γ₁34.5 protein precludes the assembly of RIG-I and cellular chaperone 14-3-3ε into an active complex for mitochondrial translocation. The γ₁34.5-mediated inhibition of RIG-I-14-3-3ε binding abrogates the access of RIG-I to mitochondrial antiviral-signaling protein (MAVS) and activation of interferon regulatory factor 3. As such, unlike wild type virus HSV-1, a recombinant HSV-1 in which γ₁34.5 is deleted elicits efficient cytokine induction and replicates poorly, while genetic ablation of RIG-I expression, but not of MDA5 expression, rescues viral growth. Collectively, these findings suggest that viral suppression of cytosolic RNA sensing is a key determinant in the evolutionary arms race of a large DNA virus and its host.     

Hypoxic bone marrow mesenchymal cell-extracellular vesicles containing miR-328-3p promote lung cancer progression via the NF2-mediated Hippo axis

Lung cancer is the most aggressive tumour afflicting patients on a global scale. Extracellular vesicle (EV)-delivered microRNAs (miRs) have been reported to play critical roles in cancer development. The current study aimed to investigate the role of hypoxic bone marrow mesenchymal cell (BMSC)-derived EVs containing miR-328-3p in lung cancer. miR-328-3p expression was determined in a set of lung cancer tissues by RT-qPCR. BMSCs were infected with lentivirus-mediated miR-328-3p knock-down and then cultured in normoxic or hypoxic conditions, followed by isolation of EVs. Following ectopic expression and depletion experiments in lung cancer cells, the biological functions of miR-328-3p were analysed using CCK-8 assay, flow cytometry and Transwell assay. Xenograft in nude mice was performed to test the in vivo effects of miR-328-3p delivered by hypoxic BMSC-derived EVs on tumour growth of lung cancer. Finally, the expression of circulating miR-328-3p was detected in the serum of lung cancer patients. miR-328-3p was highly expressed in EVs derived from hypoxic BMSCs. miR-328-3p was delivered to lung cancer cells by hypoxic BMSC-derived EVs, thereby promoting lung cancer cell proliferation, invasion, migration and epithelial-mesenchymal transition. miR-328-3p targeted NF2 to inactivate the Hippo pathway. Moreover, EV-delivered miR-328-3p increased tumour growth in vivo. Additionally, circulating miR-328-3p was bioactive in the serum of lung cancer patients. Taken together, our results demonstrated that hypoxic BMSC-derived EVs could deliver miR-328-3p to lung cancer cells and that miR-328-3p targets the NF2 gene, thereby inhibiting the Hippo pathway to ultimately promote the occurrence and progression of lung cancer.     

Sustained expression of MCP-1 induced low wall shear stress loading in conjunction with turbulent flow on endothelial cells of intracranial aneurysm

Shear stress was reported to regulate the expression of AC007362, but its underlying mechanisms remain to be explored. In this study, to isolate endothelial cells of blood vessels, unruptured and ruptured intracranial aneurysm (IA) tissues were collected from IA patients. Subsequently, quantitative real-time PCR (qRT-PCR), Western blot and luciferase assay were performed to investigate the relationships between AC007362, miRNAs-493 and monocyte chemoattractant protein-1 (MCP-1) in human umbilical vein endothelial cells (HUVECs) exposed to shear stress. Reduced representation bisulphite sequencing (RRBS) was performed to assess the level of DNA methylation in AC007362 promoter. Accordingly, AC007362 and MCP-1 were significantly up-regulated while miR-493 was significantly down-regulated in HUVECs exposed to shear stress. AC007362 could suppress the miR-493 expression and elevate the MCP-1 expression, and miR-493 was shown to respectively target AC007362 and MCP-1. Moreover, shear stress in HUVECs led to the down-regulated DNA methyltransferase 1 (DNMT1), as well as the decreased DNA methylation level of AC007362 promoter. Similar results were also observed in ruptured IA tissues when compared with unruptured IA tissues. In conclusion, this study presented a deep insight into the operation of the regulatory network of AC007362, miR-493 and MCP-1 upon shear stress. Under shear stress, the expression of AC007362 was enhanced by the inhibited promoter DNA methylation, while the expression of MCP-1 was enhanced by sponging the expression of miR-493.     

CX3CL1 promotes tumour cell by inducing tyrosine phosphorylation of cortactin in lung cancer

It has been reported that chemokine CX₃CL1 can regulate various tumours by binding to its unique receptor CX₃CR1. However, the effect of CX₃CL1-CX₃CR1 on the lung adenocarcinoma and lung squamous cell carcinoma is still unclear. Here, we showed that CX₃CL1 can further invasion and migration of lung adenocarcinoma A549 and lung squamous cell carcinoma H520. In addition, Western blot and immunofluorescence test indicated CX₃CL1 up-regulated the phosphorylation level of cortactin, which is a marker of cell pseudopodium. Meanwhile, the phosphorylation levels of c-Src and c-Abl, which are closely related to the regulation of cortactin phosphorylation, are elevated. Nevertheless, the src/abl inhibitor bosutinib and mutations of cortactin phosphorylation site could inhibit the promotion effect of CX₃CL1 on invasion and migration of A549 and H520. Moreover, these results of MTT, Hoechst staining and Western blot suggested that CX₃CL1 had no effect on the proliferation and apoptosis of A549 and H520 in vitro. The effects of CX₃CL1 were also verified by the subcutaneous tumour formation in nude mice, which showed that it could promote proliferation and invasion of A549 in vivo. In summary, our results indicated that CX₃CL1 furthered invasion and migration in lung cancer cells partly via activating cortactin, and CX₃CL1 may be a potential molecule in regulating the migration and invasion of lung 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.     

Hydrogen sulphide reduces hyperhomocysteinaemia-induced endothelial ER stress by sulfhydrating protein disulphide isomerase to attenuate atherosclerosis

Hyperhomocysteinaemia (HHcy)-impaired endothelial dysfunction including endoplasmic reticulum (ER) stress plays a crucial role in atherogenesis. Hydrogen sulphide (H₂S), a metabolic production of Hcy and gasotransmitter, exhibits preventing cardiovascular damages induced by HHcy by reducing ER stress, but the underlying mechanism is unclear. Here, we made an atherosclerosis with HHcy mice model by ApoE knockout mice and feeding Pagien diet and drinking L-methionine water. H₂S donors NaHS and GYY4137 treatment lowered plaque area and ER stress in this model. Protein disulphide isomerase (PDI), a modulation protein folding key enzyme, was up-regulated in plaque and reduced by H₂S treatment. In cultured human aortic endothelial cells, Hcy dose and time dependently elevated PDI expression, but inhibited its activity, and which were rescued by H₂S. H₂S and its endogenous generation key enzyme-cystathionine γ lyase induced a new post-translational modification-sulfhydration of PDI. Sulfhydrated PDI enhanced its activity, and two cysteine-terminal CXXC domain of PDI was identified by site mutation. HHcy lowered PDI sulfhydration association ER stress, and H₂S rescued it but this effect was blocked by cysteine site mutation. Conclusively, we demonstrated that H₂S sulfhydrated PDI and enhanced its activity, reducing HHcy-induced endothelial ER stress to attenuate atherosclerosis development.     

KDM5A silencing transcriptionally suppresses the FXYD3-PI3K/AKT axis to inhibit angiogenesis in hepatocellular cancer via miR-433 up-regulation

Hepatocellular cancer (HCC) has been reported to belong to one of the highly vascularized solid tumours accompanied with angiogenesis of human umbilical vein endothelial cells (HUVECs). KDM5A, an attractive drug target, plays a critical role in diverse physiological processes. Thus, this study aims to investigate its role in angiogenesis and underlying mechanisms in HCC. ChIP-qPCR was utilized to validate enrichment of H3K4me3 and KDM5A on the promotor region of miR-433, while dual luciferase assay was carried out to confirm the targeting relationship between miR-433 and FXYD3. Scratch assay, transwell assay, Edu assay, pseudo-tube formation assay and mice with xenografted tumours were conducted to investigate the physiological function of KDM5A-miR-433-FXYD3-PI3K-AKT axis in the progression of HCC after loss- and gain-function assays. KDM5A p-p85 and p-AKT were highly expressed but miR-433 was down-regulated in HCC tissues and cell lines. Depletion of KDM5A led to reduced migrative, invasive and proliferative capacities in HCC cells, including growth and a lowered HUVEC angiogenic capacity in vitro. Furthermore, KDM5A suppressed the expression of miR-433 by demethylating H3K4me3 on its promoterregion. miR-433 negatively targeted FXYD3. Depleting miR-433 or re-expressing FXYD3 restores the reduced migrative, invasive and proliferative capacities, and lowers the HUVEC angiogenic capacity caused by silencing KDM5A. Therefore, KDM5A silencing significantly suppresses HCC tumorigenesis in vivo, accompanied with down-regulated miR-433 and up-regulated FXYD3-PI3K-AKT axis in tumour tissues. Lastly, KDM5A activates the FXYD3-PI3K-AKT axis to enhance angiogenesis in HCC by suppressing miR-433.     

Implications of seed germination ecology for conservation of Camptotheca acuminata,a rare,endemic, and endangered species in China

Plant Ecology - Seed germination and seedling recruitment are among the most critical stages for plant population persistence and development, which may be influenced by habitat fragmentation and...