scholarly journals Notch Signaling Regulates MMP-13 Expression via Runx2 in Chondrocytes

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Di Xiao ◽  
Ruiye Bi ◽  
Xianwen Liu ◽  
Jie Mei ◽  
Nan Jiang ◽  
...  
Keyword(s):  
Notch Signaling ◽  
Chondrocyte Proliferation ◽  
Runx2 Expression ◽  
Over Expression ◽  
Proliferation And Differentiation ◽  
Important Regulator ◽  
Signaling Activation

Abstract Notch signaling is involved in the early onset of osteoarthritis. The aim of this study was to investigate the role of Notch signaling changes during proliferation and differentiation of chondrocyte, and to testify the mechanism of MMP-13 regulation by Notch and Runx2 expression changes during osteoarthritis. In this study, Chondrocytes were isolated from rat knee cartilages. Notch signaling was activated/inhibited by Jagged-1/DAPT. Proliferative capacity of Chondrocytes was analyzed by CCK-8 staining and EdU labeling. ColX, Runx2 and MMP-13 expressions were analyzed as cell differentiation makers. Then, Runx2 gene expression was interfered using lentivirus transfection (RNAi) and was over-expressed by plasmids transfected siRNA in chondrocytes, and MMP-13 expression was analyzed after Jagged-1/DAPT treatment. In vivo, an intra-articular injection of shRunx2 lentivirus followed with Jagged1/DAPT treatments was performed in rats. MMP-13 expression in articular cartilage was detected by immunohistochemistry. Finally, MMP-13 expression changes were analyzed in chondrocytes under IL-1β stimulation. Our findings showed that, CCK-8 staining and EdU labeling revealed suppression of cell proliferation by Notch signaling activation after Jagged-1 treatment in chondrocytes. Promoted differentiation was also observed, characterized by increased expressions of Col X, MMP-13 and Runx2. Meanwhile, Sox9, aggrecan and Col II expressions were down-regulated. The opposite results were observed in Notch signaling inhibited cells by DAPT treatment. In addition, Runx2 RNAi significantly attenuated the ‘regulatory sensitivity’ of Notch signaling on MMP-13 expression both in vitro and in vivo. However, we found there wasn’t significant changes of this ‘regulatory sensitivity’ of Notch signaling after Runx2 over-expression. Under IL-1β circumstance, MMP-13 expression could be reduced by both DAPT treatment and Runx2 RNAi, while Runx2 interference also attenuated the ‘regulatory sensitivity’ of Notch in MMP-13 under IL-1β stimulation. In conclusion, Notch signaling is an important regulator on rat chondrocyte proliferation and differentiation, and this regulatory effect was partially mediated by proper Runx2 expression under both normal and IL-1β circumstances. In the meanwhile, DAPT treatment could effectively suppress expression of MMP-13 stimulated by IL-1 β.

scholarly journals Downregulation of S100A4 Alleviates Cardiac Fibrosis via Wnt/β -Catenin Pathway in Mice

2018 ◽  
Vol 46 (6) ◽  
pp. 2551-2560 ◽  
Author(s):  
LiJun Qian ◽  
Jian Hong ◽  
YanMei Zhang ◽  
MengLin Zhu ◽  
XinChun Wang ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Group A ◽  
Ischemic Diseases ◽  
Fibrotic Diseases ◽  
Signaling Activation

Background/Aims: Cardiac fibrosis is a pathological change leading to cardiac remodeling during the progression of myocardial ischemic diseases, and its therapeutic strategy remains to be explored. S100A4, a calcium-binding protein, participates in fibrotic diseases with an unclear mechanism. This study aimed to investigate the role of S100A4 in cardiac fibrosis. Methods: Cardiac fibroblasts from neonatal C57BL/6 mouse hearts were isolated and cultured. Myocardial infarction was induced by ligating the left anterior descending coronary artery (LAD). The ligation was not performed in the sham group. A volume of 5×105pfu/g adenovirus or 5 µM/g ICG-001 was intramyocardially injected into five parts bordering the infarction zone or normal region. We used Western blotting, quantitative RT-PCR, immunofluorescence, immunohistochemistry and Masson’s trichrome staining to explore the function of S100A4. Results: We found significant increases of S100A4 level and cardiac fibrosis markers, and β-catenin signaling activation in vitro and in vivo. In addition, knockdown of S100A4 significantly reduced cardiac fibrosis and β-catenin levels. Moreover, the expression of S100A4 decreased after ICG-001 inhibited β-catenin signal pathway. Conclusion: Downregulation of S100A4 alleviates cardiac fibrosis via Wnt/β -catenin pathway in mice. S100A4 may be a therapeutic target of cardiac fibrosis.

scholarly journals Critical role of integrin CD11c in splenic dendritic cell capture of missing-self CD47 cells to induce adaptive immunity

2018 ◽  
Vol 115 (26) ◽  
pp. 6786-6791 ◽  
Author(s):  
Jiaxi Wu ◽  
Huaizhu Wu ◽  
Jinping An ◽  
Christie M. Ballantyne ◽  
Jason G. Cyster
Keyword(s):  
Critical Role ◽  
T Cell Proliferation ◽  
Cell Capture ◽  
Antigen Uptake ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation ◽  
Missing Self

CD11c, also known as integrin alpha X, is the most widely used defining marker for dendritic cells (DCs). CD11c can bind complement iC3b and mediate phagocytosis in vitro, for which it is also referred to as complement receptor 4. However, the functions of this prominent marker protein in DCs, especially in vivo, remain poorly defined. Here, in the process of studying DC activation and immune responses induced by cells lacking self-CD47, we found that DC capture of CD47-deficient cells and DC activation was dependent on the integrin-signaling adaptor Talin1. Specifically, CD11c and its partner Itgb2 were required for DC capture of CD47-deficient cells. CD11b was not necessary for this process but could partially compensate in the absence of CD11c. Mice with DCs lacking Talin1, Itgb2, or CD11c were defective in supporting T-cell proliferation and differentiation induced by CD47-deficient cell associated antigen. These findings establish a critical role for CD11c in DC antigen uptake and activation in vivo. They may also contribute to understanding the functional mechanism of CD47-blockade therapies.

scholarly journals STIL-a novel link in Shh and Wnt signaling, endowing oncogenic and stem like attributes to colorectal cancer

2020 ◽  
Author(s):  
Tapas Pradhan ◽  
Vikas Kumar ◽  
H Evangeline Surya ◽  
R Krishna ◽  
Samu John ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Clinical Importance ◽  
Disease Free Survival ◽  
Drug Resistant ◽  
Free Survival ◽  
Over Expression ◽  
Disease Free

AbstractDiscovery of potent gene regulating tumorigenesis and drug resistance is of high clinical importance. STIL is an oncogene, however its molecular insights and role in colorectal oncogenesis are unknown. In this study we have explored role of STIL in tumorigenesis and studied its molecular targets in colorectal cancer (CRC). STIL silencing reduced proliferation and tumor growth in CRC. Further, STIL was found to regulate stemness markers CD133 & CD44 and drug resistant markers Thymidylate synthase, ABCB1 & ABCG2 both in in-vitro and in-vivo CRC models. In addition, over expression of STIL mRNA was found to be associated with reduced disease free survival in CRC cases. To our surprise we observed an Shh independent regulation of stemness and drug resistant genes mediated by STIL. Interestingly, we found an Shh independent regulation of β-catenin mediated by STIL via p-AKT, which partially answers Shh independent regulatory mechanism of CSC markers by STIL. Our study suggest an instrumental role of STIL in molecular manifestation of CRC and progression.

Demyelination-remyelination in the Central Nervous System: Ligand-dependent Participation of the Notch Signaling Pathway

2019 ◽  
Vol 171 (1) ◽  
pp. 172-192 ◽  
Author(s):  
Patricia A Mathieu ◽  
María F Almeira Gubiani ◽  
Débora Rodríguez ◽  
Laura I Gómez Pinto ◽  
María de Luján Calcagno ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Central Nervous System ◽  
Nervous System ◽  
Notch Signaling ◽  
Progenitor Cells ◽  
Central Nervous System Disease ◽  
Notch Signaling Pathway ◽  
Signaling Activation

Abstract Multiple sclerosis (MS) is an immune-mediated central nervous system disease mostly affecting young people. Multiple sclerosis and other neurodegenerative and white matter disorders involve oligodendrocyte (OL) damage and demyelination. Therefore, elucidating the signaling pathways involved in the remyelination process through the maturation of OL progenitor cells (OPCs) may contribute to the development of new therapeutic approaches. In this context, this paper further characterizes toxic cuprizone (CPZ)-induced demyelination and spontaneous remyelination in rats and investigates the role of ligand-dependent Notch signaling activation along demyelination/remyelination both in vivo and in vitro. Toxic treatment generated an inflammatory response characterized by both microgliosis and astrogliosis. Interestingly, early demyelination revealed an increase in the proportion of Jagged1+/GFAP+ cells, which correlated with an increase in Jagged1 transcript and concomitant Jagged1-driven Notch signaling activation, particularly in NG2+ OPCs, in both the corpus callosum (CC) and subventricular zone (SVZ). The onset of remyelination then exhibited an increase in the proportion of F3/contactin+/NG2+ cells, which correlated with an increase in F3/contactin transcript during ongoing remyelination in the CC. Moreover, neurosphere cultures revealed that neural progenitor cells present in the brain SVZ of CPZ-treated rats recapitulate in vitro the mechanisms underlying the response to toxic injury observed in vivo, compensating for mature OL loss. Altogether, the present results offer strong evidence of cell-type and ligand-specific Notch signaling activation and its time- and area-dependent participation in toxic demyelination and spontaneous remyelination.

Notch Regulation by the Fbw7/hcdc4/Sel-10 Ubiquitin Ligase.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1420-1420
Author(s):  
Jonathan E. Grim ◽  
Olga Sala ◽  
Nack Gyun Chung ◽  
Jerald Radich ◽  
Barbara J. Varnum-Finney ◽  
...  
Keyword(s):  
T Cell ◽  
Ubiquitin Ligase ◽  
Heterozygous Mutation ◽  
Candidate Peptide ◽  
Human T Cell ◽  
Notch Protein ◽  
Important Regulator

Abstract T-cell neoplasms frequently sustain mutations in the Notch1 gene, leading to the expression of constitutively active Notch proteins. Such mutations often target the C-terminal PEST domain, which is known to be involved in protein stability. The ubiquitin ligase Fbw7/hdcd4/Sel-10 is a tumor suppressor that negatively regulates Notch function by targeting the Notch protein for ubiquitination and proteasomal degradation. Although the PEST domain is known to be important for Fbw7/Notch interactions, the specific residues that regulate binding of Notch to Fbw7 have not been defined. Based on the structural motifs (known as phosphodegrons) common to known substrates of Fbw7, we have identified two candidate peptide sequences within the Notch protein and have generated a series of mutants in these regions. Using co-immunoprecipitation assays, we show that one potential phosphodegron that is outside of the PEST domain does not appear to influence Notch binding to Fbw7. However, a second potential phosphodegron is present within the PEST domain and contains a conserved threonine residue (T2512) which is central to binding of Fbw7 to Notch. A mutant in which this residue is replaced by alanine (T2512A) shows a prolonged half life when compared to wild type Notch ICD, supporting its role in Notch stability. To evaluate the role of Fbw7 mediated Notch degradation in vitro and in vivo, we used lentiviral vectors to transfect hematopoietic cells with shRNA targeting Fbw7. These studies demonstrate that Fbw7 knockdown leads to phenotypes consistent with increased Notch activity. Because Notch is commonly mutated in human leukemias, we hypothesized that Fbw7 may also sustain mutations that lead to loss of Notch regulation. We evaluated primary human T cell leukemias for mutations in Fbw7 and found that 1 of 23 samples contains a heterozygous mutation in the Fbw7 common region (R505C). We show that this mutant is deficient in binding to Notch, suggesting that Fbw7 mutation may contribute to the deregulation of Notch that is commonly seen in T-cell neoplasms. Together, this work shows that Fbw7 is an important regulator of Notch function whose mutation may be an important step in leukemogenesis.

NOTCH1 Signaling Defines a Leukemia Initiating Cell Population in T-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1507-1507
Author(s):  
Wenxue Ma ◽  
Kristen M. Smith ◽  
Alejandro Gutierrez ◽  
Heather S. Leu ◽  
Qingfei Jiang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Notch Signaling ◽  
Lymphoblastic Leukemia ◽  
Therapeutic Resistance ◽  
Self Renewal ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Notch1 Receptor

Abstract Abstract 1507 Leukemia initiating cells (LIC) contribute to therapeutic resistance as a result of their capacity to accumulate mutations in pathways, such as the NOTCH1 receptor signaling pathway, that promote self-renewal and survival within specific niches. Activating mutations in NOTCH1 occur commonly in T cell acute lymphoblastic leukemia (T-ALL) and have been implicated in driving therapeutic resistance. However, the role of NOTCH1 activation in human T-ALL LIC propagation and LIC sensitivity to selective NOTCH1 receptor inhibition has not been examined. The difficulties in maintaining primary cultures of leukemia cells have hampered investigations into the biology of T-ALL LIC and underscore the need for a direct transplantation model to characterize human LIC in vivo and as a paradigm for screening candidate drugs that inhibit self-renewal pathways active in T-ALL LIC. Pediatric T-ALL serially transplantable LIC were found to be enriched in the CD34+CD4− and CD34+CD7− fractions of newly diagnosed patient samples. More recently, a CD7+CD1a− glucocorticoid resistant LIC population, capable of engrafting leukemia in NOD/SCID IL2Rƒn gamma null (NSG) mice, was identified in primary adult T-ALL without an in vitro expansion. In this study, we identified and molecularly characterized potential LIC populations in pediatric T-ALL without preceding in vitro culture and examined the role of NOTCH1 activation in LIC propagation. To further define the T-ALL LIC, CD34+CD2+CD7+ or CD34+CD2+CD7− cells were isolated from T-ALL primary patients' blood by FACS sorting and transplanted into neonatal RAG2−/− gamma chain−/− mice to determine their leukemic engraftment potential. Limiting dilution experiments were performed with cells from six T-ALL patient samples. Mice transplanted with CD34+CD2+CD7+ or CD34+CD2+CD7− cells developed a T-ALL-like disease characterized by pale bone marrow and enlarged spleen, thymus and liver. Hematopoietic organs were analyzed by flow cytometry and showed engraftment of bone marrow, spleen, thymus and liver. Furthermore, the disease could be serially transplanted. LIC were uniquely susceptible to targeted inhibition in vivo with a therapeutic human NOTCH1 negative regulatory region selective monoclonal antibody (mAb) while normal human hematopoietic progenitors were spared thereby highlighting the cell type and context specific effects of NOTCH signaling. Both the NOTCH1 mAb treatment and lentiviral shRNA knockdown of NOTCH1 reduced NOTCH1, HES1 and c-MYC transcript levels, underscoring the selectivity of NOTCH1 mAb inhibition of NOTCH signaling. These results demonstrate that CD34+CD2+CD7+ and CD34+CD2+CD7− subpopulations are enriched for LIC activity in pediatric T-ALL. Moreover, inhibition of NOTCH signaling by either mAb or shRNA-mediated Notch1 knockdown might be another strategy to target the LIC in T-ALL. Disclosures: No relevant conflicts of interest to declare.

In vivo and in vitro effect of 1,25-dihydroxyvitamin D3 and 1,25-dihydroxy-16-ene-23-yne-vitamin D3 on the proliferation and differentiation of avian chondrocytes: their role in tibial dyschondroplasia

1996 ◽  
Vol 148 (3) ◽  
pp. 465-474 ◽  
Author(s):  
C Farquharson ◽  
J S Rennie ◽  
N Loveridge ◽  
C C Whitehead
Keyword(s):  
Calcium Absorption ◽  
Biologically Active ◽  
Plasma Vitamin ◽  
Chondrocyte Differentiation ◽  
Chondrocyte Proliferation ◽  
Tibial Dyschondroplasia ◽  
Proliferation And Differentiation ◽  
Vitro Effect

Abstract 1,25-Dihydroxyvitamin D3 (1,25(OH)2D3) is regarded as the most biologically active metabolite of cholecalciferol. It prevents tibial dyschondroplasia (TD) in chicks where inhibition of chondrocyte differentiation within the growth plate occurs. However, it is unclear whether its mode of action is through direct interaction with its chondrocyte receptor and its known regulatory role in cell differentiation or is mediated by increased calcium absorption and mobilisation. Synthetic analogues of 1,25(OH)2D3 such as 1,25-dihydroxy-16-ene-23-yne cholecalciferol (RO 23–7553) with increased differentiation properties but reduced calcaemic activity have been synthesised. In this study, the in vitro and in vivo effects of 1,25(OH)2D3 and RO 23–7553 on chick chondrocyte growth and differentiation were examined. In addition, the in vivo effectiveness of these steroids in preventing TD in chicks was assessed. 1,25(OH)2D3 and RO 23–7553 (10−12-10−7 m) displayed biphasic concentration effects and had similar potencies in vitro in regulating chondrocyte proliferation and differentiation. However, while the incidence of TD in birds dosed with 1,25(OH)2D3 was lower (10%) than in control chicks (55%), RO 23–7553 was ineffective (50%). This may be the result of its reduced affinity (1000 times less) for the plasma vitamin D binding protein (DBP) and the chondrocyte receptor in comparison to that of 1,25(OH)2D3. A reduction in calcium supply to the chondrocyte may also result in decreased chondrocyte differentiation but blood ionised and plasma total calcium were normal in birds dosed with RO 23–7553. These data suggest that RO 23–7553 and 1,25(OH)2D3 regulate chondrocyte proliferation and differentiation similarly in vitro but not in vivo. This may be caused by differences in DBP binding and clearance rates of the two steroids in vivo. Journal of Endocrinology (1996) 148, 465–474

MiR-214 Attenuates the Osteogenic Effects of Mechanical Loading on Osteoblasts

2019 ◽  
Vol 40 (14) ◽  
pp. 931-940
Author(s):  
Yu Yuan ◽  
Jianmin Guo ◽  
Lingli Zhang ◽  
Xiaoyang Tong ◽  
Shihua Zhang ◽  
...  
Keyword(s):  
Mechanical Loading ◽  
Mirna Expression ◽  
Mechanical Strain ◽  
Mechanisms Of Action ◽  
Matrix Mineralization ◽  
Over Expression ◽  
Osteogenic Factors

AbstractExercise is an effective way to prevent osteoporosis, but its mechanism remains unclear. MicroRNAs (miRNAs) play an essential role in bone metabolism. Recently, mechanical loading was reported to induce changes in miRNA expression in osteoblasts. However, the role of miRNAs in bone under exercise and its underlining mechanisms of action still remain unknown. MiR-214 was reported to regulate the process of osteogenesis and is considered a biomarker of osteoporosis. In this study, we aimed to investigate whether exercise could induce changes in miRNA expression in bone and to study the effects of miR-214 on mechanical loading-induced osteogenesis in osteoblasts. The results showed that miR-214 was down-regulated in both tibia from C57BL/6 mice after exercise in vivo and in osteoblasts after mechanical strain in vitro. Mechanical strain could enhance the ALP activity, promote matrix mineralization, up-regulate the expression of osteogenic factors such as ATF4, Osterix, ALP and β-catenin, and down-regulate RANKL and RANK expression. Over-expression of miR-214 not only inhibited the expression of these osteogenic factors but also attenuated mechanical strain-enhanced osteogenesis in osteoblasts. Collectively, our results indicated that miR-214 could attenuate the osteogenic effects of mechanical loading on osteoblasts, suggesting that inhibition of miR-214 may be one of the ways in which exercise prevents osteoporosis.

scholarly journals Targeting cholesterol homeostasis in hematopoietic malignancies

Blood ◽  
2021 ◽  
Author(s):  
Andrea Brendolan ◽  
Vincenzo Russo
Keyword(s):  
Cholesterol Homeostasis ◽  
Cell Integrity ◽  
Cellular Functions ◽  
Hematopoietic Malignancies ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation ◽  
X Receptors

Cholesterol is a vital lipid for cellular functions. It is necessary for membrane biogenesis, cell proliferation and differentiation. In addition to maintaining cell integrity and permeability, increasing evidence indicates a strict link between cholesterol homeostasis, inflammation and haematological tumors. This makes cholesterol homeostasis an optimal therapeutic target for hematopoietic malignancies. Manipulating cholesterol homeostasis either interfering with its synthesis or activating the reverse cholesterol transport via the engagement of liver X receptors (LXRs), affects the integrity of tumor cells both in vitro and in vivo. Cholesterol homeostasis has also been manipulated to restore antitumor immune responses in preclinical models. These observations have prompted clinical trials in acute myeloid leukemia (AML) to test the combination of chemotherapy with drugs interfering with cholesterol synthesis, i.e. statins. We review the role of cholesterol homeostasis in hematopoietic malignancies, as well as in cells of the tumor microenvironment, and discuss the potential use of lipid modulators for therapeutic purposes.

scholarly journals RBP-J Deficiency Promoted The Proliferation and Differentiation of CD133-Positive Ependymal Cells In Vitro and In Vivo Studies

2021 ◽  
Author(s):  
Xin Ye ◽  
Mengyi Li ◽  
Wei Bian ◽  
Junwei Li ◽  
Ting Zhang ◽  
...  
Keyword(s):  
Notch Signaling ◽  
Signaling Pathway ◽  
Notch Signaling Pathway ◽  
In Vivo Studies ◽  
Ependymal Cells ◽  
Ependymal Layer ◽  
Proliferation And Differentiation ◽  
Mature Neurons

Abstract Although the ependymal cells were reported to have the characteristics of neural stem cells (NSCs), the properties of CD133-ependymal cells have not been uncovered, in particular, it is largely unknown about the effect of Notch signaling pathway on the neurogenesis of CD133-positive ependymal cells. By using the transgenic mouse and primarily cultured ependymal cells, we found that the immunoreactivity for prominin-1/CD133 was exclusively localized in the subventricular zone (SVZ) and ependymal layer of ventricles, moreover, most CD133-positive ependymal cells were co-labeled with Nestin. In addition, RBP-J, a key nuclear effector of Notch signaling pathway, was highly active in CD133-positive ependymal cells. Our results demonstrated that CD133-positive ependymal cells can differentiate into the immature and mature neurons, in particular, the number of CD133-positive ependymal cells differentiating into the immature and mature neurons was significantly increased following the deficiency or interference of RBP-J in vivo or in vitro. By using real-time qPCR and Western blot, we found that RBP-J and Hes1 were down-regulated while Notch1 was up-regulated in the expression levels of mRNAs and proteins following the deficiency or interference of RBP-J in vivo or in vitro. These results demonstrated RBP-J deficiency promoted the proliferation and differentiation of CD133-positive ependymal cells. Therefore, we speculated that RBP-J could maintain CD133-positive ependymal cells in the characteristics of NSCs possibly by regulating Notch1/RBP-J/Hes1 pathway.

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