Hypoxia Upregulates the Notch3 Signaling Pathway to Promote Epithelial-Mesenchymal Transition in Pulmonary Artery Cells.

Objective: To investigate the role and mechanism of Notch3 in a hypoxia-induced model of pulmonary hypertension, specifically pulmonary artery hypertension. Methods: A pulmonary artery hypertension rat model was induced using monocrotaline, and hepatic encephalopathy staining was used to observe the pathomorphological changes in pulmonary artery tissue. Primary isolation and extraction of rat pulmonary artery endothelial cells were performed, and a pulmonary artery hypertension cell model was established through hypoxia induction. Notch3 overexpression lentivirus (LV-Notch3) was used for intervention, and the expression of the Notch3 gene was detected using a real-time polymerase chain reaction. Western blotting was conducted to assess the expression of vascular endothelial growth factor, matrix metalloproteinase-2, and matrix metalloproteinase-9 proteins. Cell proliferation levels were measured using a medical training therapy assay. Results: Compared to the control group, the model group showed significant thickening of the pulmonary artery membrane, increased pulmonary angiogenesis, and endothelial cell damage. After Notch3 overexpression, the LV-Notch3 group showed further thickening of the pulmonary artery tunica media, increased pulmonary angiogenesis, and significantly improved endothelial cell injury. Compared to control cells, the model group showed a significant decrease in Notch3 expression (P < .05), while the expression levels of vascular endothelial growth factor, MMP-2, and MMP-9 proteins and cell proliferation ability increased significantly (P < .05). Following Notch3 overexpression, there was a significant increase in Notch3 expression (P < .05), and the expression levels of vascular endothelial growth factor, MMP-2, and MMP-9 proteins, as well as cell proliferation ability decreased significantly (P < .05). Conclusions: Notch3 can potentially reduce angiogenesis and proliferation in pulmonary artery endothelial cells and improve hypoxia-induced pulmonary artery hypertension in rats.

Omega-3 fatty acids impair miR-1-3p-dependent Notch3 down-regulation and alleviate sepsis-induced intestinal injury.

BACKGROUND: Sepsis is a troublesome syndrome that can cause intestinal injury and even high mortality rates. Omega-3 fatty acids (FAs) are known to protect against intestinal damage. Accordingly, the current study set out to explore if omega-3 FAs could affect sepsis-induced intestinal injury with the involvement of the microRNA (miR)-1-3p/Notch3-Smad axis. METHODS: First, cecal ligation and perforation (CLP) was performed to establish septic mouse models in C57BL/6J mice, and mouse intestinal epithelial MODE-K cells were induced by lipopolysaccharide (LPS) to establish sepsis cell models. The CLP-induced septic mice or LPS-exposed cells were subjected to treatment with Omega-3 FAs and activin (Smad signaling activator), miR-1-3p inhibitor and over-expressed/short hairpin RNA (oe-/sh)-Notch3 to explore their roles in inflammation, intestinal oxidative stress and cell apoptosis. A dual-luciferase reporter gene assay was further performed to verify the regulatory relationship between miR-1-3p and Notch3. RESULTS: Omega-3 FAs inhibited CLP-induced intestinal injury and ameliorated LPS-induced intestinal epithelial cell injury by down-regulating miR-1-3p, as evidenced by decreased levels of tumor necrosis factor-α, interleukin-1ß (IL-1ß) and IL-6, in addition to diminished levels of reactive oxygen species, malondialdehyde levels and superoxide dismutase activity. Furthermore, miR-1-3p could down-regulate Notch3, which inactivated the Smad pathway. CONCLUSION: Collectively, our findings indicated that omega-3 FAs elevate the expression of Notch3 by down-regulating miR-1-3p, and then blocking the Smad pathway to alleviate intestinal epithelial inflammation and oxidative stress injury caused by sepsis.

Vitamin C alleviates the senescence of periodontal ligament stem cells through inhibition of Notch3 during long-term culture.

Periodontal ligament stem cells (PDLSCs), as potential "seed cells" for periodontal tissue repair and regeneration, require to be expanded in vitro for a large scale. Senescence of PDLSCs occurred during long-term culture may compromise the therapeutic effects of PDLSCs. Medium supplements may be useful in antisenescence. However, the effects and mechanisms of vitamin C (Vc) treatment on PDLSCs during long-term culture are still unclear. In this study, we identified that Vc-treated PDLSCs cells maintained a slender morphology, higher growth rate and migration capacity, stemness, and osteogenic differentiation capability during a long-term culture. Moreover, we also identified that Notch3 was significantly upregulated during the cell senescence, and Vc treatment alleviated the senescence of PDLSCs through inhibition of Notch3 during long-term culture. In summary, Vc treatment suppressed PDLSCs senescence by reducing the expression of Notch3 and might be a simple and useful strategy to inhibit cellular senescence during the cell long-term culture.

Notch signaling pathway mediates the immunomodulatory mechanism of Yangfei Huoxue decoction alleviating bleomycin-induced pulmonary fibrosis in rats.

OBJECTIVE: To investigate the immunomodulatory mechanism by which Yangfei Huoxue decoction (YHD) alleviates bleomycin (BLM)-induced pulmonary fibrosis (PF) in rats. METHODS: Rats were randomly divided into two time-point groups (day 14 and 28), and each time-point group comprised the following six subgroups: control, BLM, dexamethasone (DXM), YHD high dose (YHD-H), YHD middle dose (YHD-M), and YHD low dose (YHD-L). Haematoxylin and eosin and Masson staining, flow cytometry, enzyme-linked immunosorbent assay, Western blotting and UPLC-QT of analyses were examined. RESULTS: The results showed that YHD reduced the degree of alveolar inflammation and fibrosis; downregulated the expression of CD28, CD80, CD86, Delta-like 1, Notch2, and Notch3; and upregulated the proportions of Th1/Th2 and Tc1/Tc2. The seven components of YHD were detected. CONCLUSION: The current study indicates that YHD mainly functions by regulating the immune system and that the molecular mechanism may be related to the regulation of the Notch signaling pathway.

Hdac1 Regulates Differentiation of Bipotent Liver Progenitor Cells During Regeneration via Sox9b and Cdk8.

BACKGROUND & AIMS: Upon liver injury in which hepatocyte proliferation is compromised, liver progenitor cells (LPCs), derived from biliary epithelial cells (BECs), differentiate into hepatocytes. Little is known about the mechanisms of LPC differentiation. We used zebrafish and mouse models of liver injury to study the mechanisms. METHODS: We used transgenic zebrafish, Tg(fabp10a:CFP-NTR), to study the effects of compounds that alter epigenetic factors on BEC-mediated liver regeneration. We analyzed zebrafish with disruptions of the histone deacetylase 1 gene (hdac1) or exposed to MS-275 (an inhibitor of Hdac1, Hdac2, and Hdac3). We also analyzed zebrafish with mutations in sox9b, fbxw7, kdm1a, and notch3. Zebrafish larvae were collected and analyzed by whole-mount immunostaining and in situ hybridization; their liver tissues were collected for quantitative reverse transcription polymerase chain reaction. We studied mice in which hepatocyte-specific deletion of ß-catenin (Ctnnb1flox/flox mice injected with Adeno-associated virus serotype 8 [AAV8]-TBG-Cre) induces differentiation of LPCs into hepatocytes after a choline-deficient, ethionine-supplemented (CDE) diet. Liver tissues were collected and analyzed by immunohistochemistry and immunoblots. We performed immunohistochemical analyses of liver tissues from patients with compensated or decompensated cirrhosis or acute on chronic liver failure (n = 15). RESULTS: Loss of Hdac1 activity in zebrafish blocked differentiation of LPCs into hepatocytes by increasing levels of sox9b mRNA and reduced differentiation of LPCs into BECs by increasing levels of cdk8 mRNA, which encodes a negative regulator gene of Notch signaling. We identified Notch3 as the receptor that regulates differentiation of LPCs into BECs. Loss of activity of Kdm1a, a lysine demethylase that forms repressive complexes with Hdac1, produced the same defects in differentiation of LPCs into hepatocytes and BECs as observed in zebrafish with loss of Hdac1 activity. Administration of MS-275 to mice with hepatocyte-specific loss of ß-catenin impaired differentiation of LPCs into hepatocytes after the CDE diet. HDAC1 was expressed in reactive ducts and hepatocyte buds of liver tissues from patients with cirrhosis. CONCLUSIONS: Hdac1 regulates differentiation of LPCs into hepatocytes via Sox9b and differentiation of LPCs into BECs via Cdk8, Fbxw7, and Notch3 in zebrafish with severe hepatocyte loss. HDAC1 activity was also required for differentiation of LPCs into hepatocytes in mice with liver injury after the CDE diet. These pathways might be manipulated to induce LPC differentiation for treatment of patients with advanced liver diseases.