Irving M Shapiro

Thomas Jefferson University, Filadelfia, Pennsylvania, United States

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Publications (291)1197.85 Total impact

  • Deborah J Gorth · Irving M Shapiro · Makarand V Risbud ·
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    ABSTRACT: The intervertebral disc is a unique avascular organ that supports axial skeleton flexion and rotation. The high proteoglycan content of the nucleus pulposus tissue, present at the center of the disc, is pivotal for its mechanical function, distribution of compressive loads. Chronic low back pain, a prevalent and costly condition, is strongly associated with disc degeneration. Degenerated discs exhibit high levels of inflammatory cytokines, matrix catabolizing enzymes, and an overall reduction in proteoglycan content. Although the cytokine profile of diseased discs has been widely studied, little is known of what initiates and drives inflammation and subsequent low back pain. Recent studies have shown that anaerobic bacteria are present in a high percentage of painful, herniated discs and long-term treatment with antibiotics resolves symptoms associated with chronic low back pain. It is thought that these anaerobic bacteria in the disc may stimulate inflammation through toll-like receptors to further exacerbate disc degeneration. Despite the promise and novelty of this theory, there are other possible inflammatory mediators that need careful consideration. The metabolic environment associated with diabetes and atypical matrix degradation products also have the ability to activate many of the same inflammatory pathways as seen during microbial infection. It is therefore imperative that the research community must investigate the contribution of all possible drivers of inflammation to address the wide spread problem of discogenic chronic low back pain.
    Discovery medicine 11/2015; 20(110):177-184. · 3.63 Impact Factor
  • Z I Johnson · Z R Schoepflin · H Choi · I M Shapiro · M V Risbud ·
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    ABSTRACT: The intervertebral disc is an important mechanical structure that allows range of motion of the spinal column. Degeneration of the intervertebral disc - incited by aging, traumatic insult, genetic predisposition, or other factors - is often defined by functional and structural changes in the tissue, including excessive breakdown of the extracellular matrix, increased disc cell senescence and death, as well as compromised biomechanical function of the tissue. Intervertebral disc degeneration is strongly correlated with low back pain, which is a highly prevalent and costly condition, significantly contributing to loss in productivity and health care costs. Disc degeneration is a chronic, progressive condition, and current therapies are limited and often focused on symptomatic pain relief rather than curtailing the progression of the disease. Inflammatory processes exacerbated by cytokines tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are believed to be key mediators of disc degeneration and low back pain. In this review, we describe the contributions of TNF-α and IL-1β to changes seen during disc degeneration at both cellular and tissue level, as well as new evidence suggesting a link between infection of the spine and low back pain, and the emerging therapeutic modalities aimed at combating these processes.
    European cells & materials 09/2015; 30:104-117. · 4.89 Impact Factor
  • Yanmei Yang · Harry C Blair · Irving M Shapiro · Bin Wang ·
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    ABSTRACT: Parathyroid hormone (PTH) induces osteoclast formation and activity by increasing the ratio of RANKL/OPG in osteoblasts. The proteasome inhibitor carfilzomib (CFZ) has been used as an effective therapy for multiple myeloma via the inhibition of pathologic bone destruction. But the effect of combination of PTH and CFZ on osteoclastogenesis is unknown. We now reported that CFZ inhibits PTH-induced RANKL expression and secretion without affecting PTH inhibition of OPG expression, and it does so by blocking HDAC4 proteasomal degradation in osteoblasts. Furthermore, we used different types of culture systems, including co-culture, indirect co-culture and transactivation, to assess the effect of CFZ on PTH action to induce osteoclastogenesis. Our results demonstrated that CFZ blocks PTH-induced osteoclast formation and bone resorption by its additional effect to inhibit RANKL-mediated I kappa B degradation and NF-kappa B activation in osteoclasts. This study showed for the first time that CFZ targets both osteoblasts and osteoclasts to suppress PTH-induced osteoclast differentiation and bone resorption. These findings warrant further investigation of this novel combination in animal models of osteoporosis and in patients. Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
    Journal of Biological Chemistry 05/2015; 290(27). DOI:10.1074/jbc.M115.663963 · 4.57 Impact Factor
  • Irving M Shapiro · William J Landis · Makarand V Risbud ·
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    ABSTRACT: Numerous studies have documented that matrix vesicles are unique extracellular membrane-bound microparticles that serve as initial sites for mineral formation in the growth plate and most other vertebrate mineralizing tissues. Microparticle generation is not confined to hard tissues, as cells in soft tissues generate similar structures; numerous studies have shown that a common type of extracellular particle, termed an exosome, a product of the endosomal pathway, shares many characteristics of matrix vesicles. Indeed, analyses of size, morphology and lipid and protein content indicate that matrix vesicles and exosomes are homologous structures. Such a possibility impacts our understanding of the biogenesis, processing and function of matrix vesicles (exosomes) in vertebrate hard tissues and explains in part how cells control the earliest stages of mineral deposition. Moreover, since exosomes influence a spectrum of functions, including cell-cell communication, it is suggested that this type of microparticle may provide a mechanism for the transfer of signaling molecules between cells within the growth plate and thereby regulate endochondral bone development and formation. Copyright © 2015. Published by Elsevier Inc.
    Bone 05/2015; 79. DOI:10.1016/j.bone.2015.05.013 · 3.97 Impact Factor
  • Sana S Dastgheyb · Cyrus B Toorkey · Irving M Shapiro · Noreen J Hickok ·
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    ABSTRACT: Allograft bone is commonly used to augment bone stock. Unfortunately, allograft is prone to bacterial contamination and current antimicrobial therapies are inadequate. Photoactivated porphyrins combat bacterial growth by production of reactive oxygen species (ROS); however, to our knowledge, they have not been tested in the setting of allograft bone. We asked: (1) Does 5,10,15,20-tetrakis-(4-aminophenyl)-porphyrin (TAPP) stably adsorb to morselized, mineralized allograft? (2) Does Staphylococcus aureus acquire TAPP from TAPP-allograft? (3) Is TAPP-allograft antibacterial to S. aureus? (4) Is ROS production critical for antimicrobial activity? (5) Does illuminated TAPP-allograft dislodge biofilm? (6) Could other photoactive dyes (TAPP, TMPyP, TSP, THP, and methylene blue) confer antimicrobial properties to allograft? TAPP adsorption to allograft (TAPP-allograft), its localization in S. aureus, and TAPP-allograft long-term stability were determined spectrophotometrically. Antimicrobial activity was measured while activated with light or in the dark during incubation with S. aureus or after allograft biofilm formation. Glutathione was added to illuminated TAPP-allograft to quench ROS and antimicrobial activity was determined. Light-dependent antimicrobial activity of other photoactive dyes (TMPyP, TSP, THP, and methylene blue) adsorbed to allograft was also tested. We found (1) porphyrins strongly adhere to bone allograft; and (2) the bacteria are not able to sequester TAPP from the TAPP-allograft; (3) when illuminated, TAPP-allograft is resistant to bacterial adherence; (4) the effects of TAPP are inhibited by the radical scavenger glutathione, indicating ROS-dependent antimicrobial activity; (5) illumination of TAPP-allograft disrupts biofilms; and, (6) other photoactive dyes impede biofilm formation on allograft bone in the presence of light. Porphyrins stably associate with allograft and are inactive until illuminated. Illuminated TAPP-allograft markedly reduces bacterial colonization, which is restored in the presence of radical scavengers. Finally, illuminated TAPP-allograft disrupts biofilms. The findings of this in vitro study suggest that loading bone allograft with biocompatible porphyrins before surgery might allow increased sterility of the allograft during implantation. Future testing in an animal model will determine if these in vitro activities can be used to prevent allograft-based infection in an establishing osteomyelitis.
    Clinical Orthopaedics and Related Research 04/2015; 473(9). DOI:10.1007/s11999-015-4299-5 · 2.77 Impact Factor
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    ABSTRACT: Periprosthetic joint infection (PJI) is one of the most challenging complications compromising the outcome of an otherwise successful operation. Considerable efforts have been invested in the recent years to address paradigm shifts in our understanding of the complex microbiological phenomena that contribute to the pathophysiology of PJI, such as microbial adherence, biofilm formation, and resistance to antibiotics. This article is an introduction to some of the recent advancements in the prevention, diagnosis, and treatment of PJI. It describes how industry, academic researchers, and government are increasing collaboration to address PJI through development of novel technologies, therapeutic strategies, and regulatory science that specifically target the unique biofilm-associated aspects of its pathogenesis. Copyright 2015 by the American Academy of Orthopaedic Surgeons.
    The Journal of the American Academy of Orthopaedic Surgeons 04/2015; 23 Suppl(suppl):S32-43. DOI:10.5435/JAAOS-D-14-00455 · 2.53 Impact Factor
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    Anthony V Florschutz · Brian S Parsley · Irving M Shapiro ·
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    ABSTRACT: Greater documentation of patient history and clinical course is crucial for identifying factors that can influence surgical outcomes. The Centers for Medicare and Medicaid Services have already begun public reporting of hospital data on readmission, complication, and infection rates and will soon launch a website to make physician-specific outcomes data public. The orthopaedic community has the opportunity to lead the way in ensuring that adequate and accurate data is collected to facilitate appropriate comparisons that are based on patients' true risk of complications and the complexity of treatment. Several studies have reported a link between oral pathogens and periprosthetic infection, although it remains unclear whether organisms unique to dental tissues are also present in osteoarthritic joints and tissues affected by periprosthetic joint infection. The American Academy of Orthopaedic Surgeons and the American Dental Association are aware of these concerns and have created guidelines for antibiotic prophylaxis in patients who have undergone total hip or knee arthroplasty and require high-risk dental procedures. Because these guidelines have received considerable criticism, recommendations that are based on scientific and case-controlled clinical studies and provide effective guidance on this important subject are needed. Copyright 2015 by the American Academy of Orthopaedic Surgeons.
    The Journal of the American Academy of Orthopaedic Surgeons 04/2015; 23 Suppl(suppl):S55-9. DOI:10.5435/JAAOS-D-14-00419 · 2.53 Impact Factor
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    ABSTRACT: Objectives of this study were to investigate whether AQP1 and AQP5 expression is altered during intervertebral disc degeneration and if hypoxia and HIF-1 regulate their expression in NP cells. AQP expression was measured in human tissues from different degenerative grades; regulation by hypoxia and HIF-1 was studied using promoter analysis and gain- and loss-of-function experiments. We show that both AQPs are expressed in the disc and that mRNA and protein levels decline with human disease severity. Bioinformatic analyses of AQP promoters showed multiple evolutionarily conserved HREs. Surprisingly, hypoxia failed to induce promoter activity or expression of either AQP. While genomic chromatin immunoprecipitation showed limited binding of HIF-1α to conserved HREs, their mutation did not suppress promoter activities. Stable HIF-1α suppression significantly decreased mRNA and protein levels of both AQPs, but HIF-1α failed to induce AQP levels following accumulation. Together, our results demonstrate that AQP1 and AQP5 expression is sensitive to human disc degeneration and that HIF-1α uniquely maintains basal expression of both AQPs in NP cells, independent of oxemic tension and HIF-1 binding to promoter HREs. Diminished HIF-1 activity during degeneration may suppress AQP levels in NP cells, compromising their ability to respond to extracellular osmolarity changes.
    Oncotarget 03/2015; 6(14). DOI:10.18632/oncotarget.3631 · 6.36 Impact Factor
  • Jun Li · Wen Yuan · Shuai Jiang · Wei Ye · Hao Yang · Irving M Shapiro · Makarand V Risbud ·
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    ABSTRACT: Prolyl-4-hydroxylase (PHD) proteins are key in sensing tissue hypoxia. In nucleus pulposus (NP) cells, our previous work has demonstrated that PHD isoforms have a differential contribution in controlling HIF-α degradation and activity. Recently we have shown that a regulatory relationship exists between PHD3 and inflammatory cytokines in NP cells. With respect to PHD2, the most abundant PHD isoform in NP cells, very little is known concerning its function and regulation under inflammatory conditions that characterize intervertebral disc degeneration. Here, we show that PHD2 is a potent regulator of the catabolic activities of TNF-α; silencing of PHD2 significantly decreased TNF-α-induced expression of catabolic markers including SDC4, MMP-3 and -13 and ADAMTS5 as well as several inflammatory cytokines and chemokines, while partially restoring aggrecan and collagen II expression. Use of NF-κB reporters with ShPHD2, SiHIF-1α, as well as p65-/-, PHD2-/- and PHD3-/- cells show that PHD2 serves as a co-activator of NF-κB/p65 signaling in HIF-1 independent fashion. Immunoprecipitation of endogenous and exogenously expressed tagged proteins as well as fluorescence microscopy indicates that following TNF-α treatment, PHD2 interacts with, and co-localizes with, p65. Conversely, loss-of-function experiments using lentivirally-delivered Sh-p65, Sh-IKKβ and NF-κB inhibitor confirmed that cytokine-dependent PHD2 expression in NP cells requires NF-κB signaling. These findings clearly demonstrate that PHD2 forms a regulatory circuit with TNF-α via NF-κB and thereby plays an important role in enhancing activity of this cytokine. We propose that during disc degeneration PHD2 may offer a therapeutic target to mitigate the deleterious actions of TNF-α, a key pro-inflammatory cytokine. Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
    Journal of Biological Chemistry 01/2015; 290(11). DOI:10.1074/jbc.M114.611483 · 4.57 Impact Factor
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    ABSTRACT: Antibiotic prophylaxis is standard for patients undergoing surgical procedures, yet, despite the wide use of antibiotics, breakthrough infections still occur. In the setting of total joint arthroplasty, such infections can be devastating. Recent findings have shown that synovial fluid causes marked staphylococcal aggregation, which can confer antibiotic insensitivity. We therefore ask whether clinical samples of synovial fluid that contain pre-operative prophylactic antibiotics can successfully eradicate a bacterial challenge by pertinent bacterial species. This study demonstrates that pre-operative prophylaxis with cefazolin results in high antibiotic levels. Furthermore, we show that even with antibiotic concentrations that far exceed expected bactericidal levels, Staphylococcus aureus added to the synovial fluid samples are not eradicated and are able to colonize model implant surfaces, i.e., titanium pins. Based on these studies, we suggest that current prophylactic antibiotic choices, despite high penetrance into synovial fluid, may need to be re-examined. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Antimicrobial Agents and Chemotherapy 01/2015; 59(4). DOI:10.1128/AAC.04579-14 · 4.48 Impact Factor
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    ABSTRACT: The objective of this study was to investigate whether expression of xylosyltransferase-1 (XT-1), a key enzyme in glycosaminoglycan biosynthesis, is responsive to disk degeneration and to inhibition by the inflammatory cytokines tumor necrosis factor α and IL-1β in nucleus pulposus (NP) cells. Analysis of human NP tissues showed that XT-1 expression is unaffected by degeneration severity; XT-1 and c-Jun, c-Fos, and Sp1 mRNA were positively correlated. Cytokines failed to inhibit XT-1 promoter activity and expression. However, cytokines decreased activity of XT-1 promoters containing deletion and mutation of the -730/-723 bp AP-1 motif, prompting us to investigate the role of AP-1 and Sp1/Sp3 in the regulation of XT-1 in healthy NP cells. Overexpression and suppression of AP-1 modulated XT-1 promoter activity. Likewise, treatment with the Sp1 inhibitors WP631 and mithramycin A or cotransfection with the plasmid DN-Sp1 decreased XT-1 promoter activity. Inhibitors of AP-1 and Sp1 and stable knockdown of Sp1 and Sp3 resulted in decreased XT-1 expression in NP cells. Genomic chromatin immunoprecipitation analysis showed AP-1 binding to motifs located at -730/-723 bp and -684/-677 bp and Sp1 binding to -227/-217 bp and -124/-114 bp in XT-1 promoter. These results suggest that XT-1 expression is refractory to the disease process and to inhibition by inflammatory cytokines and that signaling through AP-1, Sp1, and Sp3 is important in the maintenance of XT-1 levels in NP cells. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.
    American Journal Of Pathology 12/2014; 185(2). DOI:10.1016/j.ajpath.2014.09.021 · 4.59 Impact Factor
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    Zariel I. Johnson · Irving M. Shapiro · Makarand V. Risbud ·
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    ABSTRACT: Degeneration of the intervertebral disc is characterized by changes in proteoglycan status, loss of bound water molecules, decreased tissue osmotic pressure and a resulting mechanical failure of the disc. A similar spectrum of changes is evident in osteoarthritic articular cartilage. When healthy, resident cells in these skeletal tissues respond to applied mechanical loads by regulating their own osmotic state and the hydration of the extracellular matrix. The transcription factor Tonicity-Responsive Enhancer Binding Protein (TonEBP or NFAT5) is known to mediate the osmoadaptive response in these and other tissues. While the molecular basis of how osmotic loading controls matrix homeostasis is not completely understood, TonEBP regulates the expression of aggrecan and β1,3-glucoronosyltransferase in nucleus pulposus cells, in addition to targets that allow for survival under hypertonic stress. Moreover, in chondrocytes, TonEBP controls expression of several collagen subtypes and Sox9, a master regulator of aggrecan and collagen II expression. Thus, TonEBP-mediated regulation of the matrix composition allows disc cells and chondrocytes to modify the extracellular osmotic state itself. On the other hand, TonEBP in immune cells induces expression of TNF-α, ΙL-6 and MCP-1, pro-inflammatory molecules closely linked to matrix catabolism and pathogenesis of both disc degeneration and osteoarthritis, warranting investigations of this aspect of TonEBP function in skeletal cells. In summary, the TonEBP system, through its effects on extracellular matrix and osmoregulatory genes can be viewed primarily as a protective or homeostatic response to physiological loading.
    Matrix Biology 11/2014; 40. DOI:10.1016/j.matbio.2014.08.014 · 5.07 Impact Factor
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    ABSTRACT: As peri-prosthetic infection is one of the most devastating complications associated with implant placement, we have reasoned that such infection can be largely subverted by development of antibacterial implants. Our previous work demonstrated that covalent coupling of vancomycin to titanium alloy prevented colonization by the Gram-positive pathogens, Staphylococcus aureus and Staphylococcus epidermidis. Some orthopedic devices, including permanent prosthesis anchors, and most dental implants are transcutaneous or transmucosal and can be prone to colonization by Gram-negative pathogens. We report here the successful covalent coupling of the broad-spectrum antibiotic, tetracycline (TET), to titanium surfaces (Ti-TET) to retard Gram-negative colonization. Synthetic progress was followed by changes in water contact angle, while the presence of TET was confirmed by immunofluorescence. Ti-TET actively prevented colonization in the presence of bathing Escherichia coli, both by fluorescence microscopy and direct counting. Finally, the Ti-TET surface supported osteoblastic cell adhesion and proliferation over a 72-h period. Thus, this new surface offers a powerful means to protect transcutaneous implants from adhesion of Gram-negative pathogens, decreasing the need for replacement of this hardware. © 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2014.
    Journal of Biomedical Materials Research Part B Applied Biomaterials 11/2014; 103(7). DOI:10.1002/jbm.b.33310 · 2.76 Impact Factor
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    ABSTRACT: The intervertebral disc (IVD) is one of the largest avascular organs in vertebrates. The nucleus pulposus (NP), a highly hydrated and proteoglycan-enriched tissue, forms the inner portion of the IVD. The NP is surrounded by a multi-lamellar fibrocartilaginous structure, the annulus fibrosus (AF). This structure is covered superior and inferior side by cartilaginous endplates (CEP). The NP is a unique tissue within the IVD as it results from the differentiation of notochordal cells, whereas, AF and CEP derive from the sclerotome. The hypoxia inducible factor-1α (HIF-1α) is expressed in NP cells but its function in NP development and homeostasis is largely unknown. We thus conditionally deleted HIF-1α in notochordal cells and investigated how loss of this transcription factor impacts NP formation and homeostasis at E15.5, birth, 1 and 4 months of age, respectively. Histological analysis, cell lineage studies, and TUNEL assay were performed. Morphologic changes of the mutant NP cells were identified as early as E15.5, followed, postnatally, by the progressive disappearance and replacement of the NP with a novel tissue that resembles fibrocartilage. Notably, lineage studies and TUNEL assay unequivocally proved that NP cells did not transdifferentiate into chondrocyte-like cells but they rather underwent massive cell death, and were completely replaced by a cell population belonging to a lineage distinct from the notochordal one. Finally, to evaluate the functional consequences of HIF-1α deletion in the NP, biomechanical testing of mutant IVD was performed. Loss of the NP in mutant mice significantly reduced the IVD biomechanical properties by decreasing its ability to absorb mechanical stress. These findings are similar to the changes usually observed during human IVD degeneration. Our study thus demonstrates that HIF-1α is essential for NP development and homeostasis, and it raises the intriguing possibility that this transcription factor could be involved in IVD degeneration in humans.
    PLoS ONE 10/2014; 9(10):e110768. DOI:10.1371/journal.pone.0110768 · 3.23 Impact Factor
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    Sana Dastgheyb · Javad Parvizi · Irving M Shapiro · Noreen J Hickok · Michael Otto ·
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    ABSTRACT: The pathogenesis of joint infections is not well understood. In particular, we do not know why these infections respond poorly to antibiotic treatment. Here we show that methicillin-resistant Staphylococcus aureus, a major cause of joint infections, forms exceptionally strong biofilmlike aggregates in human synovial fluid (SF), to an extent significantly exceeding biofilm formation observed in growth medium or serum. Screening a transposon bank identified bacterial fibronectin- and fibrinogen-binding proteins as important for the formation of macroscopic clumps in SF, suggesting an important role of fibrin-containing clots in the formation of bacterial aggregates during joint infection. Pretreatment of SF with plasmin led to a strongly reduced formation of aggregates and increased susceptibility to antibiotics. These results give important insight into the pathogenesis of staphylococcal joint infection and the mechanisms underlying resistance to treatment. Furthermore, they point toward a potential novel approach for treating joint infections.
    The Journal of Infectious Diseases 09/2014; 211(4). DOI:10.1093/infdis/jiu514 · 6.00 Impact Factor
  • Xin Wang · Hua Wang · Hao Yang · Jun Li · Qiqing Cai · Irving M Shapiro · Makarand V Risbud ·
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    ABSTRACT: Matrix metalloproteinase-3 (MMP-3) plays an important role in intervertebral disc degeneration, a ubiquitous condition closely linked to low back pain and disability. Elevated expression of syndecan 4, a cell surface heparan sulfate proteoglycan, actively controls disc matrix catabolism. However, the relationship between MMP-3 expression and syndecan 4 in the context of inflammatory disc disease has not been clearly defined. We investigated the mechanisms by which cytokines control MMP-3 expression in rat and human nucleus pulposus cells. Cytokine treatment increased MMP-3 expression and promoter activity. Stable silencing of syndecan 4 blocked cytokine-mediated MMP-3 expression; more important, syndecan 4 did not mediate its effects through NF-κB or mitogen-activated protein kinase (MAPK) pathways. However, treatment with MAPK and NF-κB inhibitors resulted in partial blocking of the inductive effect of cytokines on MMP-3 expression. Loss-of-function studies confirmed that NF-κB, p38α/β2/γ/δ, and extracellular signal-regulated kinase (ERK) 2, but not ERK1, contributed to cytokine-dependent induction of MMP3 promoter activity. Similarly, inhibitor treatments, lentiviral short hairpin-p65, and short hairpin-I κ B kinase β significantly decreased cytokine-dependent up-regulation in MMP-3 expression. Finally, we show that transforming growth factor-β can block the up-regulation of MMP-3 induced by tumor necrosis factor (TNF)-α by counteracting the NF-κB pathway and syndecan 4 expression. Taken together, our results suggest that cooperative signaling through syndecan 4 and the TNF receptor 1-MAPK-NF-κB axis is required for TNF-α-dependent expression of MMP-3 in nucleus pulposus cells. Controlling these pathways may slow the progression of intervertebral disc degeneration and matrix catabolism.
    American Journal Of Pathology 07/2014; 184(9). DOI:10.1016/j.ajpath.2014.06.006 · 4.59 Impact Factor
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    ABSTRACT: The objective of this study was to determine the role of FIH-1 in regulating HIF-1 activity in the nucleus pulposus (NP) cells, and the control of this regulation by binding and sequestration of FIH-1 by Mint3. FIH-1 and Mint3 were both expressed in the NP, and were shown to strongly co-localize within the cell nucleus. While both mRNA and protein expression of FIH-1 decreased in hypoxia, only Mint3 protein levels were hypoxia sensitive. Overexpression of FIH-1 was able to reduce HIF-1 function as seen by changes in activities of HRE-luciferase reporter and HIF-1α-CTAD and HIF-2α-TAD. Moreover, co-transfection of either full-length Mint3 or the N-terminus of Mint3 abrogated FIH-1-dependent reduction in HIF-1 activity under both normoxia and hypoxia. Nuclear levels of FIH-1 and Mint3 decreased in hypoxia, and use of specific nuclear import and export inhibitors clearly showed that cellular compartmentalization of overexpressed FIH-1 was critical for its regulation of HIF-1 activity in NP cells. Interestingly, microarray results after stable silencing of FIH-1 showed no significant changes in transcripts of classical HIF-1 target genes. However, expression of several other transcripts, including those of Notch pathway changed in FIH-1 silenced cells. Moreover, co-transfection of Notch-ICD could restore suppression of HIF1-TAD activity by exogenous FIH-1. Taken together, these results suggest that possibly due to low endogenous levels and/or preferential association with substrates such as Notch, FIH-1 activity does not represent a major mechanism by which NP cells control HIF-1-dependent transcription, a testament to their adaptation to a unique hypoxic niche.
    Journal of Biological Chemistry 05/2014; 289(30). DOI:10.1074/jbc.M114.565101 · 4.57 Impact Factor
  • Y. Tian · P. Cao · W. Yuan · J. Li · I. Shapiro · M. Risbud ·

    Global Spine Journal 05/2014; 04(S 01). DOI:10.1055/s-0034-1376626
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    Global Spine Journal 05/2014; 04(S 01). DOI:10.1055/s-0034-1376558
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    ABSTRACT: Intervertebral disc degeneration is the leading cause of chronic back pain. Recent studies show that raised level of SDC4, a cell-surface heparan sulfate (HS) proteoglycan, plays a role in pathogenesis of disc degeneration. However, in nucleus pulposus (NP) cells of the healthy intervertebral disc, the mechanisms that control expression of SDC4 and its physiological function are unknown. Hypoxia induced SDC4 mRNA and protein expression by ∼2.4- and 4.4-fold (P<0.05), respectively, in NP cells. While the activity of the SDC4 promoter containing hypoxia response element (HRE) was induced 2-fold (P<0.05), the HRE mutation decreased the activity by 40% in hypoxia. Transfections with plasmids coding prolyl-4-hydroxylase domain protein 2 (PHD2) and ShPHD2 show that hypoxic expression of SDC4 mRNA and protein is regulated by PHD2 through controlling hypoxia-inducible factor 1α (HIF-1α) levels. Although overexpression of HIF-1α significantly increased SDC4 protein levels, stable suppression of HIF-1α and HIF-1β decreased SDC4 expression by 50% in human NP cells. Finally, suppression of SDC4 expression, as well as HS function, resulted in an ∼2-fold increase in sex-determining region Y (SRY)-box 9 (Sox9) mRNA, and protein (P<0.05) and simultaneous increase in Sox9 transcriptional activity and target gene expression. Taken together, our findings suggest that in healthy discs, SDC4, through its HS side chains, contributes to maintenance of the hypoxic tissue niche by controlling baseline expression of Sox9.-Fujita, N., Hirose, Y., Tran, C. M., Chiba, K., Miyamoto, T., Toyama, Y., Shapiro, I. M., Risbud, M. V. HIF-1-PHD2 axis controls expression of syndecan 4 in nucleus pulposus cells.
    The FASEB Journal 02/2014; 28(6). DOI:10.1096/fj.13-243741 · 5.04 Impact Factor

Publication Stats

12k Citations
1,197.85 Total Impact Points


  • 1988-2015
    • Thomas Jefferson University
      • • Department of Orthopaedic Surgery
      • • Department of Surgery
      • • Department of Medicine
      Filadelfia, Pennsylvania, United States
  • 2012
    • University of Michigan
      • Life Sciences Institute
      Ann Arbor, MI, United States
  • 2010-2012
    • Jefferson College
      Хиллсборо, Missouri, United States
  • 2003-2012
    • Thomas Jefferson University Hospitals
      Philadelphia, Pennsylvania, United States
  • 2009
    • Thomas Jefferson School of Law
      Filadelfia, Pennsylvania, United States
  • 1973-2009
    • University of Pennsylvania
      • • School of Dental Medicine
      • • Department of Bioengineering
      • • Department of Medicine
      • • Department of Pathology
      • • Department of Biochemistry
      • • School of Veterinary Medicine
      • • Department of Radiation Oncology
      • • Department of Orthodontics
      Philadelphia, Pennsylvania, United States
  • 1993-2007
    • The Children's Hospital of Philadelphia
      • • Department of Pathology and Laboratory Medicine
      • • Department of Radiology
      Filadelfia, Pennsylvania, United States
  • 2005
    • Drexel University
      Filadelfia, Pennsylvania, United States
  • 1978-1994
    • Philadelphia University
      • Department of Biochemistry
      Filadelfia, Pennsylvania, United States
  • 1983
    • Swarthmore College
      Swarthmore, Pennsylvania, United States
  • 1980-1981
    • University College London
      • Department of Cell and Developmental Biology
      Londinium, England, United Kingdom