摘要: Purposes: Gap junction intercellular communication (GJIC) is essential for articular cartilage to respond appropriately to physical or biological stimuli and maintain homeostasis. Connective tissue growth factor (CTGF), identified as an endochondral ossification genetic factor, plays a vital role in cell proliferation, migration and adhesion. However, how CTGF regulates GJIC in chondrocytes is still unknown. This study aims to explore the effects of CTGF on GJIC in chondrocytes and its potential biomechanism. Materials and methods: qPCR was performed to determine the expression of gene profile in the CCN family in chondrocytes. After CTGF treatment, CCK-8 assay and scratch assay were performed to explore cell proliferation and migration. A scrape loading/dye transfer assay was adopted to visualize GJIC in living chondrocytes. Western blot analysis was done to detect the expression of Cx43 and PI3K/Akt signalling. Immunofluorescence staining was used to show protein distribution. siRNA targeting CTGF was used to detect the influence on cell-cell communication. Results; The CTGF (CCN2) was shown to be the highest expressed member of the CCN family in chondrocytes. CTGF facilitated functional gap junction intercellular communication in chondrocytes through up-regulation of Cx43 expressions. CTGF activated PI3K/Akt signalling to promote Akt phosphorylation and translocation. Suppressing CTGF also reduced the expression of Cx43. The inhibition of PI3K/Akt signalling decreased the expressions of Cx43 and thus impaired gap junction intercellular communication enhanced by CTGF. Conclusions: For the first time, we provide evidence to show CTGF facilitates cell communication in chondrocytes via PI3K/Akt signalling pathway.

摘要: Objectives Successful pregnancy involves the homeostasis between maternal decidua and fetoplacental units, whose disruption contributes to compromised pregnancy outcomes, including recurrent spontaneous abortion (RSA). The role of cell heterogeneity of maternal decidua in RSA is yet to be illustrated. Materials and methods A total of 66,078 single cells from decidua samples isolated from patients with RSA and healthy controls were analysed by unbiased single-cell RNA sequencing (scRNA-seq). Results Our scRNA-seq results revealed that stromal cells are the most abundant cell type in decidua during early pregnancy. RSA samples are accompanied by aberrant decidualization and obviously obstructed communication between stromal cells and other cell types, such as abnormal activation of macrophages and NK cells. In addition, the over-activated TNF superfamily member 12 (TNFSF12, TWEAK) and FASLG in RSA are closely related to stromal cell demise and pregnancy failure. Conclusions Our research reveals that the cell composition and communications in normal and RSA decidua at early pregnancy and provides insightful information for the pathology of RSA and will pave the way for pregnancy loss prevention.

摘要: Objectives Diabetes is a risk factor for intervertebral disc degeneration (IVDD). Studies have demonstrated that diabetes may affect IVDD through transcriptional regulation; however, whether post-transcriptional regulation is involved in diabetic IVDD (DB-IVDD) is still unknown. This study was performed to illustrate the role of HuR, an RNA-binding protein, in DB-IVDD development and its mechanism. Materials and Methods The expression of HuR was evaluated in nucleus pulposus (NP) tissues from diabetic IVDD patients and in high glucose-treated NP cells. Senescence and autophagy were assessed in HuR over-expressing and downregulation NP cells. The mRNAs that were regulated by HuR were screened, and immunoprecipitation was applied to confirm the regulation of HuR on targeted mRNAs. Results The results showed that the expression of HuR was decreased in diabetic NP tissues and high glucose-treated NP cells. Downregulation of HuR may lead to increased senescence in high glucose-treated NP cells, while autophagy activation attenuates senescence in HuR deficient NP cells. Mechanistic study showed that HuR prompted Atg7 mRNA stability via binding to the AU-rich elements. Furthermore, overexpression of Atg7, but not HuR, may ameliorate DB-IVDD in rats in vivo. Conclusions In conclusion, HuR may suppress senescence through autophagy activation via stabilizing Atg7 in diabetic NP cells; while Atg7, but not HuR, may serve as a potential therapeutic target for DB-IVDD.

摘要: Objectives Pre-eclampsia is a leading cause of morbidity and mortality during pregnancy. Although the two forms of this disorder, early- (EOPE) and late-onset of pre-eclampsia (LOPE) are different, the underlying pathology remains elusive. We aim to unravel the difference and to identify novel biomarkers for EOPE and LOPE. Materials and Methods A complete comparison of both placental and peripheral blood transcriptomes was performed to investigate the pathology of pre-eclampsia. Single-cell transcriptomics of the maternal-fetal interface were integrated to identify novel biomarkers for EOPE and LOPE which were further verified at protein or mRNA level in patients. Results We found that the transcriptomes of placentae from EOPE, but not LOPE, were significantly different from their respective controls. Conversely, the transcriptomes of peripheral blood from LOPE were more different from their controls than EOPE. Importantly, we identified that several classical biomarkers of pre-eclampsia were expressed specifically in extravillous trophoblast and syncytiotrophoblast and only upregulated in EOPE, suggesting they should not be applied to all pre-eclampsia patients in general. We further identified novel biomarkers for EOPE and LOPE from differentially expressed genes (DEGs) of placental and peripheral blood, respectively. The new biomarkers EBI3, IGF2, ORMDL3, GATA2 and KIR2DL4 were experimentally verified with patient blood samples. Conclusion Our data demonstrate distinct pathology of EOPE and LOPE, and uncover new biomarkers that can be applied in diagnosis for pre-eclampsia.

摘要: Objectives: Cancer cell migration to secondary organs remains an essential cause of death among breast cancer (BrCa) patients. Cell motility mainly relies on actin dynamics. Our previous reports verified that dishevelled-associated activator of morphogenesis 1 (Daam1) regulates invadopodia extension and BrCa cell motility. However, how Daam1 is involved in actin filament assembly and promotes pseudopodia formation in BrCa cells remains unclear. Materials and methods: One hundred human BrCa samples were collected at Women's Hospital of Nanjing Medical University. Immunohistochemistry (IHC) was used to examine Daam1 and Fascin expression. Wound healing and Boyden chamber assays were used to explore cell migration and pseudopodia extension of BrCa cells. Co-IP/pull down and Western blotting were performed to study the physical interaction between Daam1 and Fascin. Immunofluorescence assays were performed to observe whether Daam1 and Fascin were colocalized and mediated actin filament assembly. Results: Fascin was upregulated in BrCa tissues compared with that in paracarcinoma tissues. The downregulation of Fascin caused a decline in pseudopodia formation and cell motility. Moreover, we found that Daam1 interacted with Fascin via formin homology (FH) domains, especially the FH2 domain. Immunofluorescence assays showed that Daam1 and Fascin partially colocalized to actin filaments, and the knockdown of Daam1 or Fascin failed to colocalize to short and curved actin filaments. Conclusions: Daam1 specifically binds to Fascin via FH domains and cooperatively facilitates pseudopodia formation and cell migration by promoting actin filament assembly in BrCa.