Lori Setton | Duke University (original) (raw)

Papers by Lori Setton

Current Opinion in Orthopaedics, Jan 1, 2004

Wolters Kluwer Health may email you for journal alerts and information, but is committed to maint... more Wolters Kluwer Health may email you for journal alerts and information, but is committed to maintaining your privacy and will not share your personal information without your express consent. For more information, please refer to our Privacy Policy. ... Skip Navigation Links Home > October ...

Biophysical journal, Jan 1, 2008

Cells within fibrocartilaginous tissues, including chondrocytes and fibroblasts of the meniscus, ... more Cells within fibrocartilaginous tissues, including chondrocytes and fibroblasts of the meniscus, ligament, and tendon, regulate cell biosynthesis in response to local mechanical stimuli. The processes by which an applied mechanical load is transferred through the extracellular matrix to the environment of a cell are not fully understood. To better understand the role of mechanics in controlling cell phenotype and biosynthetic activity, this study was conducted to measure strain at different length scales in tissue of the fibrocartilaginous meniscus of the knee joint, and to define a quantitative parameter that describes the strain transferred from the far-field tissue to a microenvironment surrounding a cell. Experiments were performed to apply a controlled uniaxial tensile deformation to explants of porcine meniscus containing live cells. Using texture correlation analyses of confocal microscopy images, two-dimensional Lagrangian and principal strains were measured at length scales representative of the tissue (macroscale) and microenvironment in the region of a cell (microscale) to yield a strain transfer ratio as a measure of median microscale to macroscale strain. The data demonstrate that principal strains at the microscale are coupled to and amplified from macroscale principal strains for a majority of cell microenvironments located across diverse microstructural regions, with average strain transfer ratios of 1.6 and 2.9 for the maximum and minimum principal strains, respectively. Lagrangian strain components calculated along the experimental axes of applied deformations exhibited considerable spatial heterogeneity and intersample variability, and suggest the existence of both strain amplification and attenuation. This feature is consistent with an in-plane rotation of the principal strain axes relative to the experimental axes at the microscale that may result from fiber sliding, fiber twisting, and fiber-matrix interactions that are believed to be important for regulating deformation in other fibrocartilaginous tissues. The findings for consistent amplification of macroscale to microscale principal strains suggest a coordinated pattern of strain transfer from applied deformation to the microscale environment of a cell that is largely independent of these microstructural features in the fibrocartilaginous meniscus.

Journal of …, Jan 1, 2007

Mechanical stimuli are important factors that regulate cell proliferation, survival, metabolism a... more Mechanical stimuli are important factors that regulate cell proliferation, survival, metabolism and motility in a variety of cell types. The relationship between mechanical deformation of the extracellular matrix and intracellular deformation of cellular sub-regions and organelles has not been fully elucidated, but may provide new insight into the mechanisms involved in transducing mechanical stimuli to biological responses. In this study, a novel fluorescence microscopy and image analysis method was applied to examine the hypothesis that mechanical strains are fully transferred from a planar, deformable substrate to cytoplasmic and intranuclear regions within attached cells. Intracellular strains were measured in cells derived from the anulus fibrosus of the intervertebral disc when attached to an elastic silicone membrane that was subjected to tensile stretch. Measurements indicated cytoplasmic strains were similar to those of the underlying substrate, with a strain transfer ratio (STR) of 0.79. In contrast, nuclear strains were much smaller than those of the substrate, with an STR of 0.17. These findings are consistent with previous studies indicating nuclear stiffness is significantly greater than cytoplasmic stiffness, as measured using other methods. This study provides a novel method for the study of cellular mechanics, including a new technique for measuring intranuclear deformations, with evidence of differential magnitudes and patterns of strain transferred from the substrate to cell cytoplasm and nucleus.

Connective tissue …, Jan 1, 2009

Intervertebral disc (IVD) disorders are believed to be related to aging-related cell loss and phe... more Intervertebral disc (IVD) disorders are believed to be related to aging-related cell loss and phenotypic changes, as well as biochemical and structural changes in the extracellular matrix of the nucleus pulposus (NP) region. Previously, we found that the laminin γ1 chain was more highly expressed in immature NP porcine tissues, in parallel with the expression pattern for a laminin receptor, integrin α6 subunit, as compared to adjacent anulus fibrosus region; suggesting that cell-matrix interactions may be unique to the immature NP. However, the identity of laminin isoforms specific to immature or mature NP tissues, their associated receptors and functional significance are still poorly understood. In this study, we evaluated the zonal-specific expression of the laminin chains, receptors (i.e. integrins) and other binding proteins in immature tissue and isolated cells of rat, porcine and human intervertebral disc, towards the goal of revealing features of cellular environment and cellmatrix interactions in the immature NP. Results from both immuno-histochemical staining and flow cytometry analysis found that NP cells expressed higher levels of the laminin α5 chain, laminin receptors (integrin α3, α6, β4 subunit and CD239) and related binding proteins (CD151), as compared to cells from adjacent anulus fibrosus. These differences suggest that laminin interactions with NP cells are distinct from that of the anulus fibrosus, and that laminins may be important contributors to region-specific IVD biology. The revealed laminin isoforms, their receptors and related binding proteins may be used as distinguishing features of these immature NP cells in the intervertebral disc.

Journal of anatomy, Jan 1, 2007

Intervertebral disc cells are surrounded by a pericellular matrix that is biochemically and morph... more Intervertebral disc cells are surrounded by a pericellular matrix that is biochemically and morphologically distinct from other extracellular matrix regions. Although the function of the pericellular matrix is not fully understood, prior studies of pericellular matrix-chondrocyte regions in articular cartilage (termed 'chondrons') suggest that the size, shape, and mechanical properties of the pericellular matrix significantly influence the micromechanical environment of the contained cells. A first step in understanding the role of the pericellular matrix in the intervertebral disc is to quantify the three-dimensional morphology and zonal variations of these regions across the disc. In this study, three-dimensional reconstructions and morphometric measurements of pericellular matrixcell regions were obtained in situ using fluorescence confocal microscopy of en bloc sections of nucleus pulposus and anulus fibrosus of the rat disc immunolabeled for type VI collagen. The morphology of the pericellular matrix and cells varied significantly across regions, with distinct pericellular matrix aspect ratios (largest/smallest diameter) showing shapes that were generally large and rounded in the nucleus pulposus (average of 1.9), and ellipsoidal and discoidal in the inner (2.4) and outer anulus fibrosus (2.8). The average pericellular matrix volume per cell was found to be significantly larger in the nucleus (6424 µ m 3 ) than that of inner (1903 µ m 3 ) and outer (1433 µ m 3 ) anulus. Pericellular matrix regions containing 1 or 2 cells were the dominant subgroup in the rat intervertebral disc at both 1 and 12 months of age. Multicellular pericellular matrix regions were present more often in the younger nucleus pulposus and outer anulus fibrosus. The orientation of the pericellular matrix regions further varied significantly across the disc, reflecting local collagen matrix architecture. These studies provide new information on the organization and shape of intervertebral disc cells and their surrounding pericellular matrix, which may provide new insights into the mechanisms that regulate cell-matrix interactions.

Biomechanics and modeling …, Jan 1, 2006

The knee meniscus exhibits significant spatial variations in biochemical composition and cell mor... more The knee meniscus exhibits significant spatial variations in biochemical composition and cell morphology that reflect distinct phenotypes of cells located in the radial inner and outer regions. Associated with these cell phenotypes is a spatially heterogeneous microstructure and mechanical environment with the innermost regions experiencing higher fluid pressures and lower tensile strains than the outer regions. It is presently unknown, however, how meniscus tissue mechanics correlate with the local micromechanical environment of cells. In this study, theoretical models were developed to study mechanics of inner and outer meniscus cells with varying geometries. The results for an applied biaxial strain predict significant regional differences in the cellular mechanical environment with evidence of tensile strains along the collagen fiber direction of approximately 0.07 for the rounded inner cells, as compared to levels of 0.02-0.04 for the elongated outer meniscus cells. The results demonstrate an important mechanical role of extracellular matrix anisotropy and cell morphology in regulating the region-specific micromechanics of meniscus cells, that may further play a role in modulating cellular responses to mechanical stimuli.

Critical care …, Jan 1, 1995

To evaluate the possible role of tumor necrosis factor (TNF) in the development of lung injury af... more To evaluate the possible role of tumor necrosis factor (TNF) in the development of lung injury after bowel ischemia, and the ability of TNF-soluble receptors to negate TNF toxicity, using a rat small bowel ischemia and reperfusion model. Prospective, randomized, controlled laboratory study. Research laboratory. Forty adult male Sprague-Dawley rats weighing approximately 300 g. The rats were divided equally into four groups: a) ischemia and reperfusion alone; b) those animals receiving TNF antibodies (1 mL) before reperfusion; and c) those animals receiving 200 micrograms of human recombinant TNF soluble receptors. These 30 anesthetized rats underwent 60 mins of superior mesenteric artery occlusion per 4 hrs of reperfusion. The remaining ten animals were sham operated (laparotomy), serving as controls. Lung permeability was measured using bovine serum albumin labeled with 125I, and organ injury was assessed histologically. Thirty and 60 mins after declamping and reperfusion, plasma TNF concentrations increased to 830 +/- 66 and 173 +/- 56 pg/mL, respectively, compared with 10 pg/mL before ischemia (p < .001). In sham-operated control rats, TNF concentrations did not increase from baseline concentrations. Four hours after reperfusion, sequestration of neutrophils in the pulmonary microcirculation was noted (319 +/- 60 vs. 84 +/- 13 neutrophils/10 high-power fields in sham-operated rats [p < .04]). Pulmonary microvascular leak also occurred, as measured by translocation of radiolabeled albumin into the bronchoalveolar space and expressed as the ratio of bronchoalveolar lavage to blood concentrations. This ratio was 5.3 +/- 0.8 in ischemic control animals compared with 1.1 +/- 0.3 in sham animals (p < .03). Treatment with antibodies to TNF before reperfusion attenuated the pulmonary injury (75 +/- 6 neutrophils/10 high-power fields, permeability index 1.6 +/- 0.1) less than in ischemic controls (p < .005). A similar protection was achieved with soluble TNF receptors, which prevented bowel ischemia-induced lung neutrophil sequestration (117 +/- 35 neutrophils/10 high-power fields, pulmonary vascular leak ratio of 2.3 +/- 0.1, p < .05). The results of this study show that ischemia and subsequent reperfusion of the intestine in rats produce lung injury. This injury is mediated, at least in part, by TNF. Soluble TNF receptors are an effective tool in preventing lung TNF injury after intestinal ischemia.

The Journal of Bone and Joint Surgery (American), Jan 1, 2006

Mechanical loading of the intervertebral disc may contribute to disc degeneration by initiating d... more Mechanical loading of the intervertebral disc may contribute to disc degeneration by initiating degeneration or by regulating cell-mediated remodeling events that occur in response to the mechanical stimuli of daily activity. This article is a review of the current knowledge of the role of mechanical stimuli in regulating intervertebral disc cellular responses to loading and the cellular changes that occur with degeneration. Intervertebral disc cells exhibit diverse biologic responses to mechanical stimuli, depending on the loading type, magnitude, duration, and anatomic zone of cell origin. The innermost cells respond to low-to-moderate magnitudes of static compression, osmotic pressure, or hydrostatic pressure with increases in anabolic cell responses. Higher magnitudes of loading may give rise to catabolic responses marked by elevated protease gene or protein expression or activity. The key regulators of these mechanobiologic responses for intervertebral disc cells will be the micromechanical stimuli experienced at the cellular level, which are predicted to differ from that measured for the extracellular matrix. Large hydrostatic pressures, but little volume change, are predicted to occur for cells of the nucleus pulposus during compression, while the highly oriented cells of the anulus fibrosus may experience deformations in tension or compression during matrix deformations. In general, the pattern of biologic response to applied loads suggests that the cells of the nucleus pulposus and inner portion of the anulus fibrosus experience comparable micromechanical stimuli in situ and may respond more similarly than cells of the outer portion of the anulus fibrosus. Changes in these features with degeneration are critically understudied, particularly degeneration-associated changes in cell-level mechanical stimuli and the associated mechanobiology. Little is known of the mechanisms that regulate cellular responses to intervertebral mechanobiology, nor is much known with regard to the precise mechanical stimuli experienced by cells during loading. Mechanical factors appear to regulate responses of the intervertebral disc cells through mechanisms involving intracellular Ca(2+) transients and cytoskeletal remodeling that may regulate downstream effects such as gene expression and posttranslational biosynthesis. Future studies should address the broader biologic responses to mechanical stimuli in intervertebral disc mechanobiology, the involved signaling mechanisms, and the apparently important interactions among mechanical factors, genetic factors, cytokines, and inflammatory mediators that may be critical in the regulation of intervertebral disc degeneration.

Annals of biomedical …, Jan 1, 2005

Intervertebral disc (IVD) cells experience a broad range of physicochemical stimuli under physiol... more Intervertebral disc (IVD) cells experience a broad range of physicochemical stimuli under physiologic conditions, including alterations in their osmotic environment. Cellular responses to altered osmolarity have been documented at the transcriptional and post-translational level, but mainly for extracellular matrix proteins. In this study, the gene expression profile of human IVD cells was quantified with gene array technology following exposure to increased osmolarity in order to capture the biological responses for a broad set of targets. A total of 42 genes were identified in IVD cells as significantly changed following culture under hyper-osmotic conditions. Gene expression patterns were verified using RT-PCR. Genes identified in this study include those related to cytoskeleton remodeling and stabilization (ephrin-B2, muskelin), as well as membrane transport (ion transporter SLC21A12, osmolyte transporter SLC5A3, monocarboxylic acid SLC16A6). An unexpected finding was the differential regulation of the gene for the neurotrophin, brain-derived neurotrophic factor, by hyper-osmotic stimuli that suggests a capability of IVD cells to respond to physicochemical stimuli with factors that may regulate discogenic pain.

Biomaterials, Jan 1, 2006

We describe the fabrication of deformable microstructures by low-pressure-soft-microembossing (mS... more We describe the fabrication of deformable microstructures by low-pressure-soft-microembossing (mSEmb) that provides in vitro experimental ''test-beds'' to investigate the interplay of mechanical and chemical stimuli on cell behavior in a highly controlled environment. Soft microembossing exploits the softness and plasticity of parafilm to fabricate microstructures by pressing a silicon master or an elastomeric poly(dimethylsiloxane) stamp into the parafilm. We demonstrate that a protein-resistant comb polymer can be printed into the raised features of the embossed microstructures, which imparts protein, and hence cell resistance to those regions of the microstructures. These two features of our fabrication methodology-microembossing followed by spatially selective transfer of a nonfouling polymer-forms the core of our strategy to pattern cells within the parafilm microstructures, such that the cells are confined within bottoms of the microstructures. Cell culture experiments demonstrated the preferential cell attachment of NIH 3T3 fibroblasts to the fibronectin (FN) micropatterns by immunofluorescence microscopy. The actin cytoskeleton realigned along the axis of applied mechanical stress, and stretched cells showed altered gene expression of cytoskeletal and matrix proteins in response to mechanical deformation. The use of parafilm as a substrate and mSEmb as a fabrication method provides a simple and widely accessible methodology to investigate cellular behavior under well-defined conditions of plastic deformation and surface ligand density. r

Journal of Orthopaedic …

Interleukin-17 (IL-17) is a cytokine recently shown to be elevated, along with interferon-γ (IFNγ... more Interleukin-17 (IL-17) is a cytokine recently shown to be elevated, along with interferon-γ (IFNγ) and tumor necrosis factor (TNFα), in degenerated and herniated intervertebral disc (IVD) tissues, suggesting a role for these cytokines in intervertebral disc disease. The objective of our study was to investigate the involvement of IL-17 and costimulants IFNγ and TNFα in intervertebral disc pathology. Cells were isolated from anulus fibrosus and nucleus pulposus tissues of patients undergoing surgery for intervertebral disc degeneration or scoliosis. The production of inflammatory mediators, nitric oxide (NOx), prostaglandin E2 (PGE2) and interleukin-6 (IL-6), as well as intercellular adhesion molecule (ICAM-1) expression, were quantified for cultured cells following exposure to IL-17, IFNγ and TNFα. Intervertebral disc cells exposed to IL-17, IFNγ or TNFα showed a remarkable increase in inflammatory mediator release and ICAM-1 expression (GLM and ANOVA, p<0.05). Addition of IFNγ or TNFα to IL-17 demonstrated a synergistic increase in inflammatory mediator release, and a marked increase in ICAM-1 expression. These findings suggest that IVD cells not only respond with a catabolic phenotype to IL-17 and costimulants IFNγ and TNFα, but also express surface ligands with consequent potential to recruit additional lymphocytes and immune cells to the IVD microenvironment. IL-17 may be an important regulator of inflammation in the IVD pathologies.

Arthritis & …, Jan 1, 2009

Objective-Mice with Col9a1 gene inactivation (Col9a1 -/-) prematurely develop osteoarthritis and ... more Objective-Mice with Col9a1 gene inactivation (Col9a1 -/-) prematurely develop osteoarthritis and intervertebral disc degeneration. In this study, we investigate Col9a1 -/mice for functional and symptomatic changes that may associate with these pathologies.

Arthritis & …, Jan 1, 2008

Objective. Type IX collagen is an important component of the intervertebral disc extracellular ma... more Objective. Type IX collagen is an important component of the intervertebral disc extracellular matrix. Mutations in type IX collagen are associated with premature disc degeneration in mice and a predisposition to disc disorders in humans. The aim of this study was to assess the prevalence and timeline of intervertebral disc degeneration in mice homozygous for an inactivated Col9a1 gene.

Tissue Engineering Part B: …, Jan 1, 2009

Osteoarthritis (OA) is a degenerative joint disease that can result in joint pain, loss of joint ... more Osteoarthritis (OA) is a degenerative joint disease that can result in joint pain, loss of joint function, and deleterious effects on activity levels and lifestyle habits. Current therapies for OA are largely aimed at symptomatic relief and may have limited effects on the underlying cascade of joint degradation. Local drug delivery strategies may provide for the development of more successful OA treatment outcomes that have potential to reduce local joint inflammation, reduce joint destruction, offer pain relief, and restore patient activity levels and joint function. As increasing interest turns toward intra-articular drug delivery routes, parallel interest has emerged in evaluating drug biodistribution, safety, and efficacy in preclinical models. Rodent models provide major advantages for the development of drug delivery strategies, chiefly because of lower cost, successful replication of human OA-like characteristics, rapid disease development, and small joint volumes that enable use of lower total drug amounts during protocol development. These models, however, also offer the potential to investigate the therapeutic effects of local drug therapy on animal behavior, including pain sensitivity thresholds and locomotion characteristics. Herein, we describe a translational paradigm for the evaluation of an intraarticular drug delivery strategy in a rat OA model. This model, a rat interleukin-1b overexpression model, offers the ability to evaluate anti-interleukin-1 therapeutics for drug biodistribution, activity, and safety as well as the therapeutic relief of disease symptoms. Once the action against interleukin-1 is confirmed in vivo, the newly developed anti-inflammatory drug can be evaluated for evidence of disease-modifying effects in more complex preclinical models.

Journal of …, Jan 1, 2010

Objective: Interleukin-1 beta (IL1β) is a pro-inflammatory cytokine that mediates arthritic patho... more Objective: Interleukin-1 beta (IL1β) is a pro-inflammatory cytokine that mediates arthritic pathologies. Our objective was to evaluate pain and limb dysfunction resulting from IL1β overexpression in the rat knee and investigate the ability of local IL1 receptor antagonist (IL1Ra) delivery to reverse associated pathology.

Osteoarthritis and …, Jan 1, 2007

Journal of …, Jan 1, 2008

Object-Biochemical irritation of the dorsal root ganglion (DRG) after intervertebral disc herniat... more Object-Biochemical irritation of the dorsal root ganglion (DRG) after intervertebral disc herniation contributes to radiculopathy through tumor necrosis factor-α (TNFα)-mediated inflammation. Soluble TNF receptor Type II (sTNFRII) sequesters this cytokine, providing clinical benefit. Previous work involving conjugation of sTNFRII with thermally responsive elastin-like polypeptide (ELP) yielded a chimeric protein (ELP-sTNFRII) with in vitro anti-TNFα bioactivity. Furthermore, temperature-triggered ELP aggregation into a "depot" prolongs protein residence time following perineural injection. In this study the authors evaluated the inflammatory phenotype of DRG explants after TNFα stimulation, and assessed the abilities of sTNFRII or ELP-sTNFRII to attenuate these neuroinflammatory changes.

… Research Part B: …, Jan 1, 2009

Biodegradable polymeric scaffolds are of interest for delivering antibiotics to local sites of in... more Biodegradable polymeric scaffolds are of interest for delivering antibiotics to local sites of infection in orthopaedic applications, such as bone and diarthrodial joints. The objective of this study was to develop a biodegradable scaffold with ease of drug loading in aqueous solution, while providing for drug depot delivery via syringe injection. Elastin-like polypeptides (ELPs) were used for this application, biopolymers of repeating pentapeptide sequences that were thermally triggered to undergo in situ depot formation at body temperature. ELPs were modified to enable loading with the antibiotics, cefazolin and vancomycin, followed by induction of the phase transition in vitro. Cefazolin and vancomycin concentrations were monitored, as well as bioactivity of the released antibiotics, to test an ability of the ELP depot to provide for prolonged release of bioactive drugs. Further tests of formulation viscosity were conducted to test suitability as an injectable drug carrier. Results demonstrate sustained release of therapeutic concentrations of bioactive antibiotics by the ELP, with first-order time constants for drug release of ~ 25h for cefazolin and ~ 500h for vancomycin. These findings illustrate that an injectable, in situ forming ELP depot can provide for sustained release of antibiotics with an effect that varies across antibiotic formulation. ELPs have important advantages for drug delivery, as they are known to be biocompatible, biodegradable and elicit no known immune response. These benefits suggest distinct advantages over currently used carriers for antibiotic drug delivery in orthopaedic applications.

Protein …, Jan 1, 2009

We have previously developed a method to purify recombinant proteins, termed inverse transition c... more We have previously developed a method to purify recombinant proteins, termed inverse transition cycling (ITC) that eliminates the need for column chromatography. ITC exploits the inverse solubility phase transition of an elastin-like polypeptide (ELP) that is fused to a protein of interest. In ITC, a recombinant ELP fusion protein is cycled through its phase transition, resulting in separation of the ELP fusion protein from other Escherichia coli contaminants. Herein, we examine the role of the position of the ELP in the fusion protein on the expression levels and yields of purified protein for four recombinant ELP fusion proteins. Placing the ELP at the C-terminus of the target protein (protein-ELP) results in a higher expression level for the four ELP fusion proteins, which also translates to a greater yield of purified protein. The position of the fusion protein also has a significant impact on its specific activity, as ELP-protein constructs have a lower specific activity than protein-ELP constructs for three out of the four proteins. Our results show no difference in mRNA levels between protein-ELP and ELPprotein fusion constructs. Instead, we suggest two possible explanations for these results: first, the translational efficiency of mRNA may differ between the fusion protein in the two orientations and second, the lower level of protein expression and lower specific activity is consistent with a scenario that placement of the ELP at the N-terminus of the fusion protein increases the fraction of misfolded, and less active conformers, which are also preferentially degraded compared to fusion proteins in which the ELP is present at the C-terminal end of the protein.

Arthritis & …, Jan 1, 2010

Objective-Prior reports document macrophage and lymphocyte infiltration with proinflammatory cyto... more Objective-Prior reports document macrophage and lymphocyte infiltration with proinflammatory cytokine expression in pathologic intervertebral disc (IVD) tissues. Nevertheless, the role of the Th17 lymphocyte lineage in mediating disc disease remains uninvestigated. We undertook this study to evaluate the immunophenotype of pathologic IVD specimens, including interleukin-17 (IL-17) expression, from surgically obtained IVD tissue and from nondegenerated autopsy control tissue.