Jagged1 Expression Research Articles

Hydrogel contained valproic acid accelerates bone-defect repair via activating Notch signaling pathway in ovariectomized rats

The purpose was to observe whether valproic acid (VPA) has a positive effect on bone-defect repair via activating the Notch signaling pathway in an OVX rat model. The MC3T3-E1 cells were cocultured with VPA and induced to osteogenesis, and the osteogenic activity was observed by alkaline phosphatase (ALP) staining, Alizarin Red (RES) staining and Western blotting (WB). Then the hydrogel-containing VPA was implanted into the femoral epiphysis bone-defect model of ovariectomized (OVX) rats for 12 weeks. Micro-CT, biomechanical testing, histology, immunofluorescence, RT-qPCR, and WB analysis were used to observe the therapeutic effect and explore the possible mechanism. ALP and ARS staining and WB results show that the cell mineralization, osteogenic activity, and protein expression of ALP, OPN, RUNX-2, OC, Notch 1, HES1, HEY1, and JAG1 of VPA group is significantly higher than the control group. Micro-CT, biomechanical testing, histology, immunofluorescence, and RT-qPCR evaluation show that group VPA presented the stronger effect on bone strength, bone regeneration, bone mineralization, higher expression of VEGFA, BMP-2, ALP, OPN, RUNX-2, OC, Notch 1, HES1, HEY1, and JAG1 of VPA when compared with OVX group. Our current study demonstrated that local treatment with VPA could stimulate repair of femoral condyle defects, and these effects may be achieved by activating Notch signaling pathway and acceleration of blood vessel and bone formation.

Relationship of Notch Signal, Surfactant Protein A, and Indomethacin in Cervix During Preterm Birth: Mast Cell and Jagged-2 May Be Key in Understanding Infection-mediated Preterm Birth.

Although it is thought that there is a close relationship between Notch signal and preterm birth, the functioning of this mechanism in the cervix is unknown. The efficacy of surfactants and prostaglandin inhibitors in preterm labor is also still unclear. In this study, 48 female CD-1 mice were distributed to pregnant control (PC), Sham, PBS, indomethacin (2 mg/kg; intraperitoneally), lipopolysaccharides (LPS) (25 μg/100 μl; intrauterine), LPS + IND, and Surfactant Protein A Block (SP-A Block: SP-A B; the anti-SP-A antibody was applied 20 µg/100μl; intrauterine) groups. Tissues were examined by immunohistochemistry, immunofluorescence, and Western blot analysis. LPS administration increased the expression of N1 Dll-1 and Jagged-2 (Jag-2). Although Toll-like receptor (Tlr)-2 significantly increased in the LPS-treated and SP-A-blocked groups, Tlr-4 significantly increased only in the LPS-exposed groups. It was observed that Jag-2 is specifically expressed by mast cells. Overall, this experimental model shows that some protein responses increase throughout the uterus, starting at a specific point on the cervix epithelium. Surfactant Protein A, which we observed to be significantly reduced by LPS, may be associated with the regulation of the epithelial response, especially during preterm delivery due to infection. On the contrary, prostaglandin inhibitors can be considered an option to delay infection-related preterm labor with their dose-dependent effects. Finally, the link between mast cells and Jag-2 could potentially be a control switch for preterm birth.

Intrahepatic cholangiocyte regeneration from an Fgf-dependent extrahepatic progenitor niche in a zebrafishmodel of Alagille Syndrome.

Alagille Syndrome (ALGS) is a congenital disorder caused by mutations in the Notch ligand gene JAGGED1, leading to neonatal loss of intrahepatic duct (IHD) cells and cholestasis. Cholestasis can resolve in certain patients with ALGS, suggesting regeneration of IHD cells. However, the mechanisms driving IHD cell regeneration following Jagged loss remains unclear. Here, we show that cholestasis due to developmental loss of IHD cells can be consistently phenocopied in zebrafish with compound jagged1b and jagged2b mutations or knockdown. Leveraging the transience of jagged knockdown in juvenile zebrafish, we find that resumption of Jagged expression leads to robust regeneration of IHD cells through a Notch-dependent mechanism. Combining multiple lineage tracing strategies with whole-liver three-dimensional imaging, we demonstrate that the extrahepatic duct (EHD) is the primary source of multipotent progenitors that contribute to the regeneration, but not to the development, of IHD cells. Hepatocyte-to-IHD cell transdifferentiation is possible but rarely detected. Progenitors in the EHD proliferate and migrate into the liver with Notch signaling loss and differentiate into IHD cells if Notch signaling increases. Tissue-specific mosaic analysis with an inducible dominant-negative Fgf receptor suggests that Fgf signaling from the surrounding mesenchymal cells maintains this extrahepatic niche by directly preventing premature differentiation and allocation of EHD progenitors to the liver. Indeed, transcriptional profiling and functional analysis of adult mouse EHD organoids uncover their distinct differentiation and proliferative potential relative to IHD organoids. Our data show that IHD cells regenerate upon resumption of Jagged/Notch signaling, from multipotent progenitors originating from an Fgf-dependent extrahepatic stem cell niche. We posit that if Jagged/Notch signaling is augmented, through normal stochastic variation, gene therapy, or a Notch agonist, regeneration of IHD cells in patients with ALGS may be enhanced.

Down-regulation of miR-488 targeting to promote Jag1 expression inhibits hypoxia-reoxygenation myocardial H9c2 cell damage

To study the effect of down-regulating miR-488 targeting Jag1 on the injury of hypoxia-reoxygenation myocardial H9c2 cells. A hypoxic-reoxygenated myocardial H9c2 cell injury model was constructed. miR-488 inhibitor was used to transfect the cells. CCK-8 method and flow cytometry were used to detect cell proliferation and apoptosis in each group. Lactate dehydrogenase (LDH), superoxide dismutase (SOD), malonaldehyde (MDA), catalase (CAT) levels were detected. Western blotting was used to detect the expression of Bcl-2 associated X Protein (Bax) and B cell lymphoma/lewkmia-2 (Bcl-2). Target genes of miR-488 were predicted, and a luciferase reporter system was used to verify the targeting relationship between the two. Myocardial H9c2 cells were co-transfected with miR-488 inhibitor and Jag1 siRNA, and treated with hypoxia and reoxygenation, cell proliferation, apoptosis, LDH, SOD, MDA, CAT levels, and Bax, Bcl-2 protein expression were detected. The expression of miR-488 in the hypoxia-reoxygenated myocardial H9c2 cells was increased, along with reduced cell proliferation, increased apoptosis, increased Bax protein expression, decreased Bcl-2 protein expression, increased MDA, decreased CAT and SOD, and increased LDH level in the supernatant of cell culture. When myocardial H9c2 cells were transfected with miR-488 inhibitor and treated with hypoxia and reoxygenation, the expression of miR-488 was decreased, along with increased cell proliferation, decreased apoptosis, decreased Bax protein expression, increased Bcl-2 protein expression, decreased MDA, increased CAT and SOD, and decreased LDH level in the supernatant of cell culture. Down-regulation of miR-488 could target and down-regulate Jag1 expression. And Jag1 siRNA could reverse the effect of miR-488 inhibitor on the proliferation, apoptosis, LDH, SOD, MDA, CAT levels and the expression of Bax and Bcl-2 of hypoxic-reoxygenated myocardial H9c2 cells. Down-regulating miR-488 targeted Jag1 can attenuate hypoxia-reoxygenation induced myocardial H9c2 cell injury.

Dysregulated Notch Signaling in the Airway Epithelium of Children with Wheeze.

The airway epithelium of children with wheeze is characterized by defective repair that contributes to disease pathobiology. Dysregulation of developmental processes controlled by Notch has been identified in chronic asthma. However, its role in airway epithelial cells of young children with wheeze, particularly during repair, is yet to be determined. We hypothesized that Notch is dysregulated in primary airway epithelial cells (pAEC) of children with wheeze contributing to defective repair. This study investigated transcriptional and protein expression and function of Notch in pAEC isolated from children with and without wheeze. Primary AEC of children with and without wheeze were found to express all known Notch receptors and ligands, although pAEC from children with wheeze expressed significantly lower NOTCH2 (10-fold, p = 0.004) and higher JAG1 (3.5-fold, p = 0.002) mRNA levels. These dysregulations were maintained in vitro and cultures from children with wheeze displayed altered kinetics of both NOTCH2 and JAG1 expression during repair. Following Notch signaling inhibition, pAEC from children without wheeze failed to repair (wound closure rate of 76.9 ± 3.2%). Overexpression of NOTCH2 in pAEC from children with wheeze failed to rescue epithelial repair following wounding. This study illustrates the involvement of the Notch pathway in airway epithelial wound repair in health and disease, where its dysregulation may contribute to asthma development.

Cytochrome P450 2J2 inhibits the proliferation and angiogenesis of retinal vascular endothelial cells by regulating the Notch signaling pathway in a hypoxia-induced retinopathy model

ABSTRACT Retinopathy of Prematurity (ROP), a type of retinal neovascularization in premature infants, has become a serious problem that drastically affects the quality of life of premature infants. ROP is associated with angiogenesis and neovascularization. Here, we aimed to explain the function and latent roles of Cytochrome P450 2J2 (CYP2J2) in hypoxia-induced retinopathy in retinal vascular endothelial cells (HRVECs). HRVECs were stimulated with hypoxia for 24 h to establish an in vitro retinopathy model. Cell viability and migration were evaluated using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and Transwell assays, respectively. Protein and gene expression was determined by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and Western blot analysis. We observed that pcDNA3.1(+)-CYP2J2 promoted CYP2J2 and Jagged1 expression, while Dll4 was down-regulated in hypoxia-stimulated HRVECs. Additionally, pcDNA3.1(+)-CYP2J2 inhibited HRVEC viability, reduced PCNA expression, and inhibited the migration of HRVECs. Further, the Notch pathway was inhibited in the Hypoxia+pcDNA3.1(+)-CYP2J2 group. Opposite results were observed upon Terfenadone treatment in hypoxia induced HRVECs. Finally, our findings further verified that DAPT promotes the effects of CYP2J2 on cell viability, migration, and Notch signaling in hypoxia-induced HRVECs, while EDTA reversed the inhibitory effects of CYP2J2 on hypoxia-induced HRVECs. In conclusions, CYP2J2 was found to inhibit the viability and angiogenesis of HRVECs by inhibiting Notch signaling in a hypoxia-induced retinopathy model.

Hypoxia Upregulates NOTCH3 Signaling Pathway to Promote Endothelial-Mesenchymal Transition in Pulmonary Artery Endothelial Cells.

Background To investigate the effect of hypoxia on pulmonary artery endothelial cells and the role of NOTCH3 in endothelial-mesenchymal transition (EnMT) and to provide a research model for pulmonary disease and explain the pathogenesis of the pulmonary disease. Methods Pulmonary artery endothelial cells were divided into two groups and cultured in normoxic and hypoxic environments, respectively. QPCR, western blot, and immunofluorescence were used to detect endothelial cell-specific marker protein and mRNA expression in each group, and the ability of endothelial cells migration was evaluated by scratch and transwell experiment. Results The pulmonary artery endothelial cells in the normoxic group presented a typical pebble-like arrangement, and the endothelial cells in hypoxic culture showed a long spindle appearance. Hypoxia induced high expression of NOTCH3, Jagged-1, Hes1, c-Src, and CSL. Immunofluorescence showed that endothelial cells in hypoxic culture began to express the α-SMA, and the expression of vWF increased with hypoxia. Cell viability, scratch, and transwell results showed that endothelial cells in the hypoxic group were more capable of viability and migration than those in the normoxic group. The induction of EnMT by hypoxia can be inhibited by using notch3-specific inhibitor DAPT and Jagged-1. This study also found that miR-7-5p can regulate endothelial NOTCH3, indicating that miRNA is also involved in the process of endothelial-mesenchymal transformation. Conclusion Hypoxia promotes the transformation of endothelial cells into mesenchymal cells by opening the NOTCH3 pathway, which lays the foundation for disease progression or clinical prognosis, and is of great significance in the treatment of diseases.

MiR-571 affects the development and progression of liver fibrosis by regulating the Notch3 pathway

Exploring the expression of miR-571 in patients with liver fibrosis and its role in the progression of liver fibrosis. A total of 74 patients with liver fibrosis in our institution from September to December 2018 were collected for study, and the expression of miR-571, Notch3 and Jagged1 in patients with different progressions of liver fibrosis was determined by RT-PCR and Western blot analysis. Set up Notch3 up group and Notch3 down regulated group, RT-PCR and Western blot were used to determine the effect of Notch signaling on the expression of fibrogenic factors. CCK-8, cell scratch assays, Transwell assays, flow cytometry were used to determine the effect of miR-571 on LX-2 proliferation, migration, apoptosis in human stem stellate cells, and RT-PCR, Western blot assays were performed to determine the effect of miR-571 on the Notch3 signaling pathway and the expression of profibrogenic factors. miR-571, Notch3 and Jagged1 are up-regulated in patients with liver fibrosis and is associated with the progression of liver fibrosis. Notch3 signaling pathway can promote the expression of fibroblast in human hepatic stellate cells; miR-571 can inhibit the apoptosis of human hepatic stellate cells, promote cell proliferation and migration; up regulation of miR-571 can promote the expression of Notch3 and Jagged1, and up-regulation of miR-571 also promoted the expression of related fibroblasts. MiR-571 can promote the activation of human stem cell stellate cells and the expression of fibroblast related factors through Notch3 signaling pathway.

Total glycosides from Eucommia ulmoides seed promoted osteogenic differentiation of adipose-derived mesenchymal stem cells and bone formation in ovariectomized rats through regulating Notch signaling pathway

BackgroundOsteoporosis (OP) is a well-known chronic degenerative disease, with impaired mesenchymal stem cells (MSCs) function and suppressed osteogenic differentiation. Total glycosides from Eucommia ulmoides seed (TGEUS) was a Chinese medicine and have rich pharmacological effects. This study was designed to explore the mechanism of TGEUS in promoting osteogenic differentiation and bone formation in ovariectomized (OVX) rats.MethodsAdipose‐derived mesenchymal stem cells (ADSCs) were isolated and treated with different concentration of TGEUS. Cell viability was assessed using cell counting kit-8 (CCK-8) assay. Osteogenic capacity was identified by ALP staining and ARS staining. Moreover, RNA sequencing between control and TGEUS treated ADSCs were further performed to reveal the mechanism of TGEUS in promoting osteogenic differentiation. The expression of Jag1, Lfng and Hey1 were measured using quantitative real-time polymerase chain reaction (qRTPCR). Osteogenic markers were further assessed by western blot. DAPT and NICD were further used to identify whether Notch signaling pathway involved into TGEUS promoting osteogenic differentiation of ADSCs. Ovariectomy-induced bone loss rats model was established and divided into three groups: sham, OVX and OVX + TGEUS groups. HE staining and immunohistochemical staining were further performed to identify whether TGEUS could promote bone formation.ResultsTGEUS treatment significantly enhanced the cell viability and ALP activity than control group, the optimal dose of TGEUS was 5 μM. We selected 5 μM TGEUS for further study. TGEUS significantly enhanced ALP activity and calcium deposition than that of control group. Activation of Notch signaling fully blocked TGEUS-induced osteogenic differentiation of ADSCs. Following TGEUS treatment, the trabecular bone of the rats was significantly increased, thickened, and more connected compared to the OVX group. With the treatment of TGEUS, the expression of Osterix (Osx), Osteocalcin (OCN) and RUNX Family Transcription Factor 2 (RUNX2) increased than OVX group.ConclusionTGEUS enhanced osteogenic differentiation of ADSCs and promoted bone formation in ovariectomy-induced bone loss rats. Our study broadened the understanding of TGEUS as a therapeutic target against osteoporosis.

Cigarette smoke upregulates Notch-1 signaling pathway and promotes lung adenocarcinoma progression

Notch-1 pathway plays an important role in lung carcinoma, stem cell regulation, cellular communication, growth and differentiation. Cigarette smoke is involved in the regulation of Notch signaling. However, current data regarding the impact of cigarette smoke on the Notch pathway in lung cancer progression are limited.The present study aimed to explore whether cigarette smoke exposure altered Notch-1 pathway in ex-vivo (surgical samples of lung parenchyma from non-smoker and smoker patients with lung adenocarcinoma) and in vitro (adenocarcinoma A549 cell line) approaches.The expression of Notch-1, Jagged-1 and CD133 in surgical samples was evaluated by immunohistochemistry. A549 were exposed to cigarette smoke extracts (2.5 % and 5 % CSE for 6, 24 and 48 h) and the expression of Notch-1, Jagged-1 and Hes-1 was evaluated by Real-Time PCR and Western Blot (nuclear fractions). Expression and localization of Notch-1, Hes-1, CD133 and ABCG2 were assessed by immunofluorescence. The expression of survivin and Ki-67 was assessed by flow cytometry following CSE exposure and inhibition of Notch-1 signaling.Smokers lung parenchyma exhibited higher expression of Notch-1. CSE exposure increased Notch-1 and Hes-1 gene and nuclear protein expression in A549. Immunofluorescence confirmed higher expression of nuclear Hes-1 in CSE-stimulated A549 cells. CSE increased both survivin and Ki-67 expression and this effect was reverted by inhibition of the Notch-1 pathway.In conclusion, these data show that cigarette smoke may promote adenocarcinoma progression by activating the Notch-1 pathway thus supporting its role as hallmark of lung cancer progression and as a new target for lung cancer treatment.

Using Spatial Transcriptomics to Reveal Fetal Liver Hematopoietic Stem Cell-Niche Interactions

The hematopoietic stem cell (HSC) microenvironment, termed the niche, supports the proliferation, self-renewal, and differentiation abilities of HSCs. The definitive HSCs emerge from the hemogenic endothelium in the aorta-gonad-mesonephros (AGM) region after E11.5, and then migrate to the fetal liver after E12.5 for expansion. After E17.5, HSCs migrate to the bone marrow and reside in the bone marrow for the postnatal stage and adulthood. Because the fetal liver is thought to be a harbor for the rapid expansion of HSCs, numerous studies have focused on the fetal liver HSC niche in the search for novel niche factors and niche cells that support HSC expansion. However, to our knowledge, there are no successes in translating the niche factors to a clinical application for the expansion of HSCs ex vivo. In this study, we are using cutting-edge spatial transcriptomics to comprehensively study the transcriptomics and interactions between HSCs and the niche cells in the fetal liver, and in search of the niche cells and factors for HSC expansion.To understand the spatial distribution and interactions between HSCs and niche cells in the fetal liver, we introduced 2 spatial transcriptomic methods, slide-seq, and 10x Visium, in our study on E14.5 mouse fetal liver. By integrating with a parallel single-cell sequencing analysis, we revealed the spatial transcriptomics of HSCs and potential niche cells, including hepatoblasts, endothelium cells, macrophages, megakaryocytes, and hepatic stellate cells/perivascular mesenchymal cells (PMCs) in E14.5 mouse fetal liver. Interestingly, we found that the PMCs were characterized by enriched N-cadherin expression. Both slide-seq and 10x Visium showed that the N-cadherin-expressing PMCs are enriched in the portal vessel area. Importantly, the majority of fetal liver HSCs are in close proximity to N-cadherin-expressing PMCs, indicating a supportive role of N-cadherin-expressing PMCs in HSC maintenance. Subsequent CellPhoneDB (CPDB) analysis demonstrated that the N-cadherin-expressing PMCs are major niche-signaling senders with an enriched expression of niche factors, such as CXCL12 and KITL, and stemness pathway-related ligands, such as IGF1, IGF2, TGFβ2, TGFβ3, JAG2, and DLK1, indicating N-cadherin-expressing PMCs could be the major niche cells in supporting HSCs in the fetal liver. This finding was consistent with our previous finding that N-cadherin-expressing bone and marrow stromal progenitor cells can maintain reserve HSCs in the adult bone marrow. Moreover, CPDB analysis indicated that other potential niche cells, such as endothelium cells, macrophages, and megakaryocytes, may support HSCs in different signal transduction pathways. For example, endothelium cells have an enriched expression of KITL, IGF2, DLL1, TGFβ1, and TGFβ2; macrophages have enriched expression of KITL, IFNγ, and TGFβ1; megakaryocytes have enriched expression of PF4, JAG2 and TGFβ1. Intriguingly, our previous studies showed that megakaryocytes could promote HSC expansion under stress conditions in the bone marrow. To investigate the potential role of N-cadherin-expressing cells in supporting fetal liver HSCs, we generated an N-cad CreER;Cxcl12 and an N-cad CreER;Scf mouse model to conditionally knockout the well-studied niche factors, CXCL12 and SCF, in N-cadherin-expressing cells. Conditional knockout of either Cxcl12 or Scf in N-cadherin-expressing cells resulted in an increase in the number of HSCs. Moreover, conditional knockout of Cxcxl12 in N-cadherin-expressing cells also resulted in a myeloid-biased differentiation. We postulate that the knockout of Cxcl12 or Scf in N-cadherin-expressing cells leads to the migration of HSCs towards other potential niche cells, such as macrophages and megakaryocytes, which may induce HSC expansion and biased differentiation.In summary, by using cutting-edge spatial transcriptomics, we revealed a comprehensive spatial transcriptomics of HSCs and niche cells in E14.5 mouse fetal liver. The N-cadherin-expressing cells in the fetal liver is a major niche in maintaining HSCs, while other potential niches may be responsible for the expansion of HSCs. In the future, we will use multiple approaches, such as spatial transcriptomics and fluorescence in situ hybridization (FISH), to verify the distribution changes of HSCs in N-cad CreER;Cxcl12 mouse, and to reveal the niches in support of the expansion of HSCs. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Premature differentiation of nephron progenitor cell and dysregulation of gene pathways critical to kidney development in a model of preterm birth

Preterm birth is a leading cause of neonatal morbidity. Survivors have a greater risk for kidney dysfunction and hypertension. Little is known about the molecular changes that occur in the kidney of individuals born preterm. Here, we demonstrate that mice delivered two days prior to full term gestation undergo premature cessation of nephrogenesis, resulting in a lower glomerular density. Kidneys from preterm and term groups exhibited differences in gene expression profiles at 20- and 27-days post-conception, including significant differences in the expression of fat-soluble vitamin-related genes. Kidneys of the preterm mice exhibited decreased proportions of endothelial cells and a lower expression of genes promoting angiogenesis compared to the term group. Kidneys from the preterm mice also had altered nephron progenitor subpopulations, early Six2 depletion, and altered Jag1 expression in the nephrogenic zone, consistent with premature differentiation of nephron progenitor cells. In conclusion, preterm birth alone was sufficient to shorten the duration of nephrogenesis and cause premature differentiation of nephron progenitor cells. These candidate genes and pathways may provide targets to improve kidney health in preterm infants.

Preparation of Nano-Cerium Dioxide and Its Auxiliary Protection on Rats with Pulmonary Fibrosis via Notch/Jagged Signaling Pathway

The aim of this study was to explore the auxiliary protection of Notch/Jagged signaling pathway based on nano-cerium dioxide (nano-CeO2) on rats with pulmonary fibrosis. Nano-CeO2 was prepared by detonation method, and the nano-particles were detected in terms of the average particle size, Zeta potential, and purity. 30 Sprague-Dawley (SD) rats were selected as the research objects, and they were grouped randomly into a blank control group (1 mL/kg normal saline injection) and a pulmonary fibrosis model group (intraperitoneal injection of 1 mL/kg 2 g/L nano-CeO2 suspension). Rats were sacrificed at the 5th day, 10th day, 15th day, 20th day, 40th day, and 50th day in turn, and their lung tissues were collected. The differences in messenger ribonucleic acid (mRNA) and protein expression of Notch-1, Jagged-1, and transforming growth factor (TGF)-β1 genes were detected by Western blot and reverse transcription-polymerase chain reaction (RT-PCR). Lung tissue sections of rats were treated with hematoxylin-eosin (HE) staining for 50 days to observe the pathological changes of lung tissues. The results showed that the shape of nano-CeO2 was similar to a circle under the transmission electron microscope, the average particle size was 25.17±10.33 nm, and the average Zeta potential was −27.26±9.72 mV. The X-ray diffraction (XRD) map of nano-CeO2 presented 5 obvious characteristic peaks that were featured with good purity characteristics. After intraperitoneal injection of nano-CeO2 suspension into rats, the lung tissues of the pulmonary fibrosis model group showed marked congestion and fibrosis compared with the blank group. The mRNA and protein expressions of Notch-1, Jagged-1, and TGF-β1 in lung tissues of the pulmonary fibrosis model group increased gradually over time. Moreover, the expression levels of Notch-1, Jagged-1, and TGF-β1 of the pulmonary fibrosis model group were greater hugely than the levels of the black control group (P < 0.05). In conclusion, nano-CeO2 can induce pulmonary fibrosis by regulating the Notch/Jagged pathway.

Alveolar progenitor differentiation and lactation depends on paracrine inhibition of notch via ROBO1/CTNNB1/JAG1.

ABSTRACTIn the mammary gland, how alveolar progenitor cells are recruited to fuel tissue growth with each estrus cycle and pregnancy remains poorly understood. Here, we identify a regulatory pathway that controls alveolar progenitor differentiation and lactation by governing Notch activation in mouse. Loss of Robo1 in the mammary gland epithelium activates Notch signaling, which expands the alveolar progenitor cell population at the expense of alveolar differentiation, resulting in compromised lactation. ROBO1 is expressed in both luminal and basal cells, but loss of Robo1 in basal cells results in the luminal differentiation defect. In the basal compartment, ROBO1 inhibits the expression of Notch ligand Jag1 by regulating β-catenin (CTNNB1), which binds the Jag1 promoter. Together, our studies reveal how ROBO1/CTTNB1/JAG1 signaling in the basal compartment exerts paracrine control of Notch signaling in the luminal compartment to regulate alveolar differentiation during pregnancy.

Prediction of Key Candidate Genes for Platinum Resistance in Ovarian Cancer.

PurposeOvarian cancer is one of the common malignant tumors of female reproductive organs, which seriously threatens the life and health of women. Resistance to chemotherapeutic drugs for ovarian cancer is the root cause of recurrence in most patients. The purpose of this study is to determine the differentially expressed genes of platinum resistance in ovarian cancer, and to screen out molecular targets and diagnostic markers that could be used to treat ovarian cancer platinum resistance.MethodsWe downloaded 5 gene microarray datasets GSE58470, GSE45553, GSE41499, GSE33482, and GSE15372 from the Gene Expression Omnibus database, all of which are associated with ovarian cancer platinum resistance. Subsequently, the intersection of the statistically significant differentially expressed genes in 5 gene chips was taken, and relevant bioinformatics and clinical parameters were performed on the screened differential genes. qRT-PCR was utilized to examine the mRNA expression levels in ovarian cancer sensitive and cisplatin-resistant cells.ResultsThree differential genes, IFI27, JAG1, DNM3, may be closely related to platinum resistance of ovarian cancer, were screened by microarray datasets. According to the combined verification of bioinformatics, clinical case analyses and experiments, it was inferred that the increased expression of DNM3 was beneficial to patients with platinum resistance, but the high expression of IFI27 and JAG1 may lead to the risk of platinum resistance.ConclusionIFI27, JAG1 and DNM3 screened by relevant gene chips may serve as new biomarkers of platinum resistance in ovarian cancer.

Propranolol inhibits stemness of hemangioma through Jagged1.

BackgroundPropranolol is used clinically to treat infantile hemangioma (IH), although the exact mechanism that underlies its effectiveness is not fully understood. The Jagged1/Notch signaling pathway is downstream of the β2-adrenergic receptor (β2-AR). Propranolol is a non-selective β2-AR blocker that was shown to inhibit demethylation adrenaline-induced Jagged1 expression. A previous study has shown that propranolol dose-dependently inhibits the growth of IH. However, the effects of propranolol on stemness of IH are not known and are thus addressed in the current study.MethodsWe analyzed the expression of Jagged1 and Notch3 in IH specimens, using genetic tools to alter Notch signaling. The transduced IH cells were treated with different doses of propranolol, and the effects on IH cell proliferation, migration, and potential for tumor sphere formation were investigated. The effects of altered Notch signaling on tumor formation in vivo were also assessed.ResultsNotch3 and Jagged1 were significantly upregulated in IH. Augmented Notch signaling in IH cells increased cell proliferation, migration, the potential for tumor sphere formation and in vivo tumor formation. On the other hand, reduced Notch signaling in IH cells decreased cell proliferation, migration, the potential for tumor sphere formation and in vivo tumor formation.ConclusionsJagged1/Notch signaling regulated the stemness of IH, and propranolol inhibited it through suppression of Notch signaling.

Knockdown of lncRNA MEG8 inhibits cell proliferation and invasion, but promotes cell apoptosis in hemangioma, via miR‑203‑induced mediation of the Notch signaling pathway.

As a member of the long non-coding (lnc)RNA family, lncRNA maternally expressed 8, small nucleolar RNA host gene (MEG8), has been reported to serve an oncogenic role in several types of malignancies, including hepatocellular carcinoma, non-small cell lung cancer and pancreatic cancer. The current study aimed to investigate the effect of the knockdown of MEG8 on human hemangioma endothelial cell (HemEC) proliferation, apoptosis and invasion, in addition to determining the underlying molecular mechanism. The knockdown of lncRNA MEG8 was achieved by transfecting lncRNA MEG8 small interfering (si)RNA into HemECs, while the combined knockdown of lncRNA MEG8 knockdown and microRNA (miR)-203 was established by co-transfecting lncRNA MEG8 siRNA and a miR-203 inhibitor into HemECs. The cell proliferation, apoptosis and invasion and the expression levels of miR-34a, miR-200b, miR-200b and Notch signaling pathway-related factors were detected via CCK-8 Kit, flow cytometry, Transwell, reverse transcription-quantitative PCR and western blot assay, respectively. The knockdown of lncRNA MEG8 significantly inhibited proliferation (P<0.05) and invasion (P<0.05), but promoted apoptosis (P<0.01) in HemECs. Furthermore, lncRNA MEG8 knockdown upregulated miR-203 (P<0.01) expression, but did not alter miR-34a or miR-200b expression (both P>0.05). Subsequent experiments revealed that miR-203 silencing exerted no significant effect on the expression levels of lncRNA MEG8 (P>0.05) in HemECs. In addition, miR-203 silencing increased cell proliferation (P<0.05) and invasion (P<0.01), but suppressed apoptosis (P<0.05). miR-203 silencing also reversed the effect of lncRNA MEG8 knockdown on the proliferation (P<0.05), apoptosis (P<0.001) and invasion (P<0.01) of HemECs. Moreover, lncRNA MEG8 knockdown downregulated jagged canonical notch ligand 1 (JAG1; P<0.05) and Notch1 (P<0.05) expression levels, while miR-203 silencing upregulated JAG1 (P<0.01) and Notch1 (P<0.01) expression levels and reversed the effects of lncRNA MEG8 knockdown on JAG1 (P<0.01) and Notch1 (P<0.01) expression in HemECs. In conclusion, the findings of the present study suggested that lncRNA MEG8 knockdown may inhibit cell proliferation and invasion, but promote cell apoptosis in hemangioma via miR-203-induced mediation of the Notch signaling pathway.

SOX12 Promotes Stem Cell-Like Phenotypes and Osteosarcoma Tumor Growth by Upregulating JAGGED1.

SOX12 plays a role in promoting the growth of some tumors; however, its role in osteosarcoma remains unclear. From gene expression omnibus (GEO) and tumor alterations relevant for genomics-driven therapy (TARGET) databases, Kaplan–Meier analyses were conducted to establish relationships between SOX12 expression and osteosarcoma survival and recurrence in osteosarcoma patients. We also performed in vitro and in vivo assays to determine SOX12 function in osteosarcoma etiology. SOX12 expression was increased in osteosarcoma; high SOX12 expression levels were related to a poor prognosis and a high disease recurrence in patients. Moreover, SOX12 expression in osteosarcoma cell lines was increased, similar to osteosarcoma cancer stem cells. We also observed that SOX12 knockdown inhibited the spheroidization and expression of stemness markers in osteosarcoma cells and tumor formation in nude mice. In addition, SOX12 knockdown inhibited JAGGED1 and HES1 expression. Similarly, JAGGED1 knockdown also inhibited the formation of osteosarcoma cancer stem cells into pellets and reduced the expression of stemness markers and tumor formation capabilities in nude mice. Finally, during SOX12 knockdown, JAGGED1 overexpression rescued osteosarcoma cells from spheroidizing. SOX12 promotes stem cell-like phenotypes and osteosarcoma tumor growth by upregulating JAGGED1.

Schwann Cells Are Required for Efficient Corneal Wound Healing

BACKGROUND: Corneal nerves play a crucial role in maintaining corneal health, which includes regulation of activity of limbal stem cell (LSC). Their loss leads to neurotrophic keratopathy (NK), with corneal ulceration, scarring, and ultimately, blindness.1 Having identified nerve-ensheathing Schwann cells (SC) in the corneal limbus, we hypothesize that SCs, via paracrine interaction with LSC, play a key role in corneal epithelial maintenance and healing. In this study we wanted to (1) Define the role of SCs in corneal healing; (2) Determine the paracrine interaction between the limbal SCs and LSC. METHODS: (1) Local corneal ablation of SCs was induced in a genetically modified mouse where the topical application of tamoxifen induced SCs apoptosis.2 The corneal epithelium was then removed with an Amoils brush under anesthesia and fluorescein was used to assess healing over 4 days. (2) We performed single-cell RNA expression analysis of 10,000 cells derived from dissociated rat limbus with droplet-based high throughput 10× Genomics to identify ~3000 genes.3 We used the data to predict possible ligand-receptor interactions between the limbal SCs and LSC. RESULTS: (1) Ablation of SCs impaired corneal wound healing in mouse cornea, suggesting the involvement of SC in innervation-dependent corneal epithelial recovery. (2) Genomic analysis suggested the presence of paracrine crosstalk between SCs and LSCs, and relevant downstream intracellular signaling events in LSCs. The latter included activation of Notch signaling and VEGF-mediated cell migration and inhibition of apoptosis.4 Further expression analysis comparing the limbal region of healthy and wounded corneas indicated significant changes in the expression of jag1, Pdgfa, Tgfb1, and Ptn genes by SCs. All of these genes could potentially play a role in corneal recovery. CONCLUSIONS: Our findings (i) describe the presence of a high volume of SCs at the limbus, located in close spatial vicinity to LSCs, (ii) demonstrate the importance of the limbal SCs for corneal wound healing, and (iii) suggest the presence of paracrine SC-LSC interaction that may be responsible for the limbal nerve-mediated activation of LSCs during homeostasis or the epithelial wound healing after injury. These findings suggest new therapeutic targets for treating NK. REFERENCES: 1. Dua HS, Said DG, Messmer EM, et al. Neurotrophic keratopathy. Prog Retin Eye Res. 2018 Sep;66:107-131. 2. Johnston APW, Naska S, Jones K, et al. Sox2-mediated regulation of adult neural crest precursors and skin repair. Stem Cell Rep. 2013;1(1):38–45. doi:10.1016/j.stemcr.2013.04.004 3. Yuzwa SA, Borrett MJ, Innes BT, et al. Developmental emergence of adult neural stem cells as revealed by single-cell transcriptional profiling. Cell Rep. 2017;21(13):3970–3986. doi:10.1016/j.celrep.2017.12.017 4. Lindsell CE, Shawber CJ, Boulter J, et al. Jagged: A mammalian ligand that activates notch1. Cell. 1995;80(6):909–917. doi:10.1016/0092-8674(95)90294-5

Inhibition of the Notch1/Jagged1 pathway promotes homing of bone mesenchymal stem cells to improve asthma in rats

To explore the association of the Notch1/Jagged1 pathway with the homing of mesenchymal stem cells (BMSCs) to regulate Th1/Th2 drift in asthma. Twenty SD rats were randomly divided into normal control group, model group, BMSC transplantation group, and BMSC+Notch inhibitor group. Ovalbumin sensitization was used to establish rat models of asthma, and BMSCs were transplanted via the tail vein. The pathology of the lung tissue was examined with HE staining, and the contents of interleukin (IL)-5, IL-13, and interferon-γ (IFN-γ) in lung tissue homogenate were determined with enzyme-linked immunosorbent assay. The expressions of Notch1 and Jagged1 mRNA were detected with RT-PCR, and CXCR4 expression in the bronchial epithelial cells was examined using immunofluorescence staining; Western blotting was used to detect the protein expressions of T-bet, GATA-3, Notch1, and Jagged1 in the lung tissue. Compared with those in the control group, the expressions of IFN-γ and T-bet proteins decreased significantly and the pulmonary expressions of IL-5, IL-13, and GATA-3 proteins as well as Notch1 and Jagged1 mRNA and protein expressions all increased significantly in the model group (P < 0.05 or 0.01). Compared with those in the model group, CXCR4, IFN-γ, and T-bet protein expressions in BMSC group and BMSCs+Notch inhibitor group all increased significantly, and Notch1 and Jagged1 protein expressions in BMSCs group and IL-5, IL-13, Notch1, and Jagged1 mRNA and protein expressions in BMSCs + Notch inhibitor group all decreased significantly (P < 0.05 or 0.01). The expressions of CXCR4 and IFN-γ were significantly higher and the expressions of IL-13 and Notch1 mRNA were significantly lower in BMSCs+Notch inhibitor group than in BMSC group (P < 0.05). In asthmatic rats, the homing of the BMSCs to the lung tissue has a regulatory effect on Th1/Th2 drift, and the Notch1/Jagged1 pathway may participate in the homing of the BMSCs.