Cell Morphology and Migration of Nucleus Pulposus Cells Depends on Substrate Stiffness and Ligand

2012 ◽  
Author(s):  
Priscilla Y. Hwang ◽  
Christopher L. Gilchrist ◽  
Aubrey T. Francisco ◽  
Jun Chen ◽  
Lori A. Setton
Keyword(s):  
Nucleus Pulposus ◽  
Cell Biology ◽  
Imaging Techniques ◽  
Matrix Proteins ◽  
Cell Phenotype ◽  
Nucleus Pulposus Cells ◽  
Cell Clustering ◽  
And Migration ◽  
And Function

Changes in nucleus pulposus (NP) cell phenotype and morphology are implicated in the progression of intervertebral disc (IVD) disorders. Understanding how changes in the NP cell microenvironment influence cell behavior and function is important for revealing how pathology-related changes in IVD extracellular matrix may affect NP cell biology. In this study, live-cell imaging techniques were utilized to study changes in cell migration and morphology when cultured upon substrates of different matrix proteins and stiffnesses. Results indicate that soft substrates containing matrix proteins promote cell clustering and cell-cell interactions which mimic in vivo conditions of healthy NP cells.

scholarly journals N-Cadherin Maintains the Healthy Biology of Nucleus Pulposus Cells under High-Magnitude Compression

2017 ◽  
Vol 43 (6) ◽  
pp. 2327-2337 ◽  
Author(s):  
Zhenyu Wang ◽  
Jiali Leng ◽  
Yuguang Zhao ◽  
Dehai Yu ◽  
Feng Xu ◽  
...  
Keyword(s):  
Cell Viability ◽  
Nucleus Pulposus ◽  
Cell Biology ◽  
Mechanical Load ◽  
Cell Phenotype ◽  
Nucleus Pulposus Cells ◽  
Keratin 19 ◽  
High Magnitude ◽  
And Control

Background/Aims: Mechanical load can regulate disc nucleus pulposus (NP) biology in terms of cell viability, matrix homeostasis and cell phenotype. N-cadherin (N-CDH) is a molecular marker of NP cells. This study investigated the role of N-CDH in maintaining NP cell phenotype, NP matrix synthesis and NP cell viability under high-magnitude compression. Methods: Rat NP cells seeded on scaffolds were perfusion-cultured using a self-developed perfusion bioreactor for 5 days. NP cell biology in terms of cell apoptosis, matrix biosynthesis and cell phenotype was studied after the cells were subjected to different compressive magnitudes (low- and high-magnitudes: 2% and 20% compressive deformation, respectively). Non-loaded NP cells were used as controls. Lentivirus-mediated N-CDH overexpression was used to further investigate the role of N-CDH under high-magnitude compression. Results: The 20% deformation compression condition significantly decreased N-CDH expression compared with the 2% deformation compression and control conditions. Meanwhile, 20% deformation compression increased the number of apoptotic NP cells, up-regulated the expression of Bax and cleaved-caspase-3 and down-regulated the expression of Bcl-2, matrix macromolecules (aggrecan and collagen II) and NP cell markers (glypican-3, CAXII and keratin-19) compared with 2% deformation compression. Additionally, N-CDH overexpression attenuated the effects of 20% deformation compression on NP cell biology in relation to the designated parameters. Conclusion: N-CDH helps to restore the cell viability, matrix biosynthesis and cellular phenotype of NP cells under high-magnitude compression.

LncRNA MALAT1 exhibits positive effects on nucleus pulposus cell biology in vivo and in vitro by sponging miR-503

2020 ◽  
Author(s):  
Hongyu Zheng ◽  
Tingting Wang ◽  
Xiangmin Li ◽  
Wei He ◽  
Zhiqiang Gong ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Cell Biology ◽  
Mapk Pathway ◽  
Critical Role ◽  
Mapk Signaling ◽  
Positive Effects ◽  
Lncrna Malat1

Abstract Background: Intervertebral disc degeneration (IDD) is characterized by the loss of nucleus pulposus cells (NPCs) and phenotypic abnormalities. Accumulating evidence suggests that long noncoding RNAs (lncRNAs) are involved in the pathogenesis of IDD. In this study, we aimed to investigate the functional effects of lncRNA MALAT1 on NPCs in IDD and the possible mechanism governing these effects. Results: We validated the decreased expression of MALAT1 in the IDD tissues, which was associated with decreased Collagen II and Aggrecan expression. In vitro, overexpressed MALAT1 could attenuate the effect of IL-1β on NPC proliferation, apoptosis, and Aggrecan degradation. In vivo, MALAT1 overexpression attenuated the severity of disc degeneration in IDD model rats. Our molecular study further demonstrated that MALAT1 could sponge miR-503, modulate the expression of miR-503, and activate downstream MAPK signaling pathways. The effects of MALAT1 on NPCs were partially reversed/aggregated by miR-503 mimics/inhibitor treatment. Conclusion: Our data suggested that the MALAT1-miR-503-MAPK pathway plays a critical role in NPCs, which may be a potential strategy for alleviating IDD.

scholarly journals The endothelial basement membrane acts as a checkpoint for entry of pathogenic T cells into the brain

2020 ◽  
Vol 217 (7) ◽  
Author(s):  
Xueli Zhang ◽  
Ying Wang ◽  
Jian Song ◽  
Hanna Gerwien ◽  
Omar Chuquisana ◽  
...  
Keyword(s):  
T Cells ◽  
Basement Membrane ◽  
Immune Cell ◽  
Cell Phenotype ◽  
T Cell Infiltration ◽  
And Migration ◽  
Endothelial Basement Membrane ◽  
Laminin 411

The endothelial cell basement membrane (BM) is a barrier to migrating leukocytes and a rich source of signaling molecules that can influence extravasating cells. Using mice lacking the major endothelial BM components, laminin 411 or 511, in murine experimental autoimmune encephalomyelitis (EAE), we show here that loss of endothelial laminin 511 results in enhanced disease severity due to increased T cell infiltration and altered polarization and pathogenicity of infiltrating T cells. In vitro adhesion and migration assays reveal higher binding to laminin 511 than laminin 411 but faster migration across laminin 411. In vivo and in vitro analyses suggest that integrin α6β1- and αvβ1-mediated binding to laminin 511–high sites not only holds T cells at such sites but also limits their differentiation to pathogenic Th17 cells. This highlights the importance of the interface between the endothelial monolayer and the underlying BM for modulation of immune cell phenotype.

Baicalein Inhibits the IL-1β-Induced Inflammatory Response in Nucleus Pulposus Cells and Attenuates Disc Degeneration In vivo

Inflammation ◽  
2019 ◽  
Vol 42 (3) ◽  
pp. 1032-1044 ◽  
Author(s):  
Haiming Jin ◽  
Qingqing Wang ◽  
Jianwei Wu ◽  
Xuyao Han ◽  
Tianchen Qian ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Nucleus Pulposus Cells

scholarly journals Knockdown of miR-660 protects nucleus pulposus cells from TNF-a-induced apoptosis by targeting serum amyloid A1

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Hao Jie Zhang ◽  
Xue Hai Ma ◽  
Song Lin Xie ◽  
Shu lian Qin ◽  
Cong Zhi Liu ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
General Characteristic ◽  
Luciferase Reporter ◽  
Healthy Controls ◽  
Nucleus Pulposus Cells ◽  
Apoptosis Rate ◽  
Tnf Α ◽  
Significant Difference ◽  
Induced Apoptosis

Abstract Background Intervertebral disc degeneration (IVDD) is a well-known cause of lower back pain, which is induced by multiple factors including increased apoptosis and decreased survival of nucleus pulposus cells. In this study, we evaluate the effect and potential mechanism of miR-660 on the nucleus pulposus cells apoptosis induced by TNF-α. Methods First, we collected tissue of nucleus pulposus from IVDD and healthy controls. General characteristic of the IVDD and healthy control was also collected. And, we also collected nucleus pulposus cells that stimulated by TNF-α or control. miRNA microarray was performed to identify the differentially expressed miRNAs. Apoptosis rate and miR-660 relative expression was measured after stimulated with different concentration of TNF-α to identify the optimal concentration of TNF-α. Second, we successfully constructed antigomiR-660 to block the miR-660 expression in nucleus pulposus cells and then stimulated with TNF-α (100 ng/ml, 12 h). The apoptosis rates and relative protein expression were then measured again. The target association between miR-660 and SAA1 was confirmed by dual-luciferase reporter. Results There was no significant difference between the age (IVDD: 39 ± 10 years, healthy controls: 36 ± 7 years), BMI and sex between IVDD and healthy controls. Microarray analysis found that miR-660 was significantly up-regulated in IVDD and TNF-α treated groups, which was further identified by PCR. We found that the rate of apoptosis and miR-660 expression increased with TNF-α concentration increased. Finally, TNF-a with 100 ng/ml was used for further experiment. Compared with TNF-α group, TNF-α + antigomiR-660 could significantly down-regulated the apoptosis rate and relative protein (c-Caspase3 and c-Caspase7). Dual-luciferase reporter revealed that miR-660 could directly binding to the SAA1 at 80–87 sites. Compared with TNF-α alone group, TNF-α + antigomiR-660 significantly up-regulated the SAA1 expression (P < 0.05). Conclusion These results indicated that knockdown of miR-660 protected the nucleus pulposus from apoptosis that induced TNF-α via up-regulation of SAA1. Further studies should focus on the role of miR-660 in protecting IVDD in vivo.

scholarly journals Metformin protects against apoptosis and senescence in nucleus pulposus cells and ameliorates disc degeneration in vivo

2016 ◽  
Vol 7 (10) ◽  
pp. e2441-e2441 ◽  
Author(s):  
Deheng Chen ◽  
Dongdong Xia ◽  
Zongyou Pan ◽  
Daoliang Xu ◽  
Yifei Zhou ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Nucleus Pulposus Cells

scholarly journals Non-viral reprogramming of human nucleus pulposus cells with FOXF1 via extracellular vesicle delivery: an in vitro and in vivo study

2021 ◽  
Vol 41 ◽  
pp. 90-107
Author(s):  
S Tang ◽  
◽  
A Salazar-Puerta ◽  
J Richards ◽  
S Khan ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Viral Vectors ◽  
Extracellular Vesicle ◽  
Proteoglycan Synthesis ◽  
Nucleus Pulposus Cells ◽  
Keratin 19 ◽  
Altered Gene ◽  
Ivd Degeneration

Intervertebral disc (IVD) degeneration is characterized by decreased cellularity and proteoglycan synthesis and increased inflammation, catabolism, and neural/vascular ingrowth. Regenerative methods for IVD degeneration are largely cell-therapy-based or involve viral vectors, which are associated with mutagenesis and undesired immune responses. The present study used bulk electroporation and engineered extracellular vesicles (EVs) to deliver forkhead-box F1 (FOXF1) mRNA to degenerate human nucleus pulposus (NP) cells as a minimally invasive therapeutic strategy for IVD regeneration. Bulk electroporation was used to investigate FOXF1 effects on human NP cells during a 4-week culture in 3D agarose constructs. Engineered EV delivery of FOXF1 into human IVD cells in monolayer was determined, with subsequent in vivo validation in a pilot mouse IVD puncture model. FOXF1 transfection significantly altered gene expression by upregulating healthy NP markers [FOXF1, keratin 19 (KRT19)], decreasing inflammatory cytokines [interleukin (IL)-1β, -6], catabolic enzymes [metalloproteinase 13 (MMP13)] and nerve growth factor (NGF), with significant increases in glycosaminoglycan accumulation in human NP cells. Engineered EVs loaded with FOXF1 demonstrated successful encapsulation of FOXF1 cargo and effective uptake by human NP cells cultured in monolayer. Injection of FOXF1-loaded EVs into the mouse IVD in vivo resulted in a significant upregulation of FOXF1 and Brachyury, compared to controls at 7 d post-injection, with no evidence of cytotoxicity. This is the first study to demonstrate non-viral delivery of FOXF1 and reprogramming of human NP cells in vitro and mouse IVD cells in vivo. This strategy represents a non-addictive approach for treating IVD degeneration and associated back pain.

scholarly journals Kruppel-Like Factors in Thrombosis and Hemostasis

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. SCI-45-SCI-45
Author(s):  
Mukesh Jain
Keyword(s):  
Cell Biology ◽  
Vascular System ◽  
Conflicts Of Interest ◽  
Human Subjects ◽  
Myeloid Cell ◽  
Nutrient Delivery ◽  
Inflammatory Processes ◽  
Endothelial Functions ◽  
And Function

Abstract Armed with the appreciation that the blood and vascular system share common origins and cooperate to ensure fundamental processes (e.g. blood flow/fluidity, oxygen/nutrient delivery, immunity) essential for organismal survival, we posited that shared molecular pathways may be operative in coordinating the function of both systems. Over the past 2 decades, studies from our group and others have identified a family of transcription factors termed Kruppel-like factors (KLFs) as essential for development, differentiation, and function of cellular constituents of both the hematopoietic and vascular systems. In this presentation, discussion will focus on the role KLFs in control of endothelium and myeloid cell biology in physiology and disease. Specifically, cellular and in vivo evidence will be discussed implicating KLFs as master regulators of all cardinal endothelial functions (permeability, vasoreactivity, blood fluidity, and inflammation). Further, studies demonstrating KLF-control of myeloid cell development, subset specification, and pro-inflammatory activation will be reviewed with particular emphasis on results of efforts altering myeloid KLFs in the context of acute (e.g. bacterial infection, sepsis) and chronic (e.g. atherosclerosis, arterial/venous thrombosis) inflammatory processes. Correlative studies in human subjects will be presented. And finally, insights into how targeting KLFs can be exploited for therapeutic gain will be discussed. Disclosures No relevant conflicts of interest to declare.

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