Browsing by Author "Richardson, William J"

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Open Access
A new non-enzymatic method for isolating human intervertebral disc cells preserves the phenotype of nucleus pulposus cells.
(Cytotechnology, 2014-12) Tang, Xinyan; Richardson, William J; Fitch, Robert D; Brown, Christopher R; Isaacs, Robert E; Chen, Jun
Cells isolated from intervertebral disc (IVD) tissues of human surgical samples are one of potential sources for the IVD cellular therapy. The purpose of this study was to develop a new non-enzymatic method, "tissue incubation", for isolating human IVD cells. The IVD tissues of annulus fibrosus (AF) and nucleus pulposus (NP) were incubated separately in tissue culture flasks with culture medium. After 7-10 days incubation, cells were able to migrate out of IVD tissues and proliferate in vitro. After 3-4 weeks culture, expanded cells were harvested by trypsinization, and the remaining tissues were transferred to a new flask for another round of incubation. The molecular phenotype of IVD cells from juvenile and adult human samples was evaluated by both flow cytometry analysis and immunocytochemical staining for the expression of protein markers of NP cells (CD24, CD54, CD239, integrin α6 and laminin α5). Flow cytometry confirmed that both AF and NP cells of all ages positively expressed CD54 and integrin α6, with higher expression levels in NP cells than in AF cells for the juvenile group sample. However, CD24 expression was only found in juvenile NP cells, and not in AF or older disc cells. Similar expression patterns for NP markers were also confirmed by immunocytochemistry. In summary, this new non-enzymatic tissue incubation method for cell isolation preserves molecular phenotypic markers of NP cells and may provide a valuable cell source for the study of NP regeneration strategies.
Open Access
Attenuation of inflammatory events in human intervertebral disc cells with a tumor necrosis factor antagonist.
(Spine, 2011-07) Sinclair, S Michael; Shamji, Mohammed F; Chen, Jun; Jing, Liufang; Richardson, William J; Brown, Christopher R; Fitch, Robert D; Setton, Lori A
Study design
The inflammatory responses of primary human intervertebral disc (IVD) cells to tumor necrosis factor α (TNF-α) and an antagonist were evaluated in vitro.
Objective
To investigate an ability for soluble TNF receptor type II (sTNFRII) to antagonize TNF-α-induced inflammatory events in primary human IVD cells in vitro.
Summary of background data
TNF-α is a known mediator of inflammation and pain associated with radiculopathy and IVD degeneration. sTNFRs and their analogues are of interest for the clinical treatment of these IVD pathologies, although information on the effects of sTNFR on human IVD cells remains unknown.
Methods
IVD cells were isolated from surgical tissues procured from 15 patients and cultured with or without 1.4 nmol/L TNF-α (25 ng/mL). Treatment groups were coincubated with varying doses of sTNFRII (12.5-100 nmol/L). Nitric oxide (NO), prostaglandin E₂ (PGE₂), and interleukin-6 (IL6) levels in media were quantified to characterize the inflammatory phenotype of the IVD cells.
Results
Across all patients, TNF-α induced large, statistically significant increases in NO, PGE₂, and IL6 secretion from IVD cells compared with controls (60-, 112-, and 4-fold increases, respectively; P < 0.0001). Coincubation of TNF-α with nanomolar doses of sTNFRII significantly attenuated the secretion of NO and PGE₂ in a dose-dependent manner, whereas IL6 levels were unchanged. Mean IC₅₀ values for NO and PGE₂ were found to be 35.1 and 20.5 nmol/L, respectively.
Conclusion
Nanomolar concentrations of sTNFRII were able to significantly attenuate the effects of TNF-α on primary human IVD cells in vitro. These results suggest this sTNFR to be a potent TNF antagonist with potential to attenuate inflammation in IVD pathology.
Open Access
Changes in midbrain pain receptor expression, gait and behavioral sensitivity in a rat model of radiculopathy.
(Open Orthop J, 2012) Hwang, Priscilla Y; Allen, Kyle D; Shamji, Mohammed F; Jing, Liufang; Mata, Brian A; Gabr, Mostafa A; Huebner, Janet L; Kraus, Virginia B; Richardson, William J; Setton, Lori A
Intervertebral disc herniation may contribute to inflammatory processes that associate with radicular pain and motor deficits. Molecular changes at the affected dorsal root ganglion (DRG), spinal cord, and even midbrain, have been documented in rat models of radiculopathy or nerve injury. The objective of this study was to evaluate gait and the expression of key pain receptors in the midbrain in a rodent model of radiculopathy. Radiculopathy was induced by harvesting tail nucleus pulposus (NP) and placing upon the right L5 DRG in rats (NP-treated, n=12). Tail NP was discarded in sham-operated animals (n=12). Mechanical allodynia, weight-bearing, and gait were evaluated in all animals over time. At 1 and 4 weeks after surgery, astrocyte and microglial activation was tested in DRG sections. Midbrain sections were similarly evaluated for immunoreactivity to serotonin (5HT(2B)), mu-opioid (µ-OR), and metabotropic glutamate (mGluR4 and 5) receptor antibodies. NP-treated animals placed less weight on the affected limb 1 week after surgery and experienced mechanical hypersensitivity over the duration of the study. Astroctye activation was observed at DRGs only at 4 weeks after surgery. Findings for pain receptors in the midbrain of NP-treated rats included an increased expression of 5HT(2B) at 1, but not 4 weeks; increased expression of µ-OR and mGluR5 at 1 and 4 weeks (periaqueductal gray region only); and no changes in expression of mGluR4 at any point in this study. These observations provide support for the hypothesis that the midbrain responds to DRG injury with a transient change in receptors regulating pain responses.
Open Access
Comparison of superior-level facet joint violations during open and percutaneous pedicle screw placement.
(Neurosurgery, 2012-11) Babu, Ranjith; Park, Jong G; Mehta, Ankit I; Shan, Tony; Grossi, Peter M; Brown, Christopher R; Richardson, William J; Isaacs, Robert E; Bagley, Carlos A; Kuchibhatla, Maragatha; Gottfried, Oren N
Background
Superior-level facet joint violation by pedicle screws may result in increased stress to the level above the instrumentation and may contribute to adjacent segment disease. Previous studies have evaluated facet joint violations in open or percutaneous screw cases, but there are no reports describing a direct institutional comparison.
Objective
To compare the incidence of superior-level facet violation for open vs percutaneous pedicle screws and to evaluate patient and surgical factors that affect this outcome.
Methods
We reviewed 279 consecutive patients who underwent an index instrumented lumbar fusion from 2007 to 2011 for degenerative spine disease with stenosis with or without spondylolisthesis. We used a computed tomography grading system that represents progressively increasing grades of facet joint violation. Patient and surgical factors were evaluated to determine their impact on facet violation.
Results
Our cohort consisted of 126 open and 153 percutaneous cases. Percutaneous procedures had a higher overall violation grade (P = .02) and a greater incidence of high-grade violations (P = .006) compared with open procedures. Bivariate analysis showed significantly greater violations in percutaneous cases for age < 65 years, obesity, pedicle screws at L4, and 1- and 2-level surgeries. Multivariate analysis showed the percutaneous approach and depth of the spine to be independent risk factors for high-grade violations.
Conclusion
This study demonstrates greater facet violations for percutaneously placed pedicle screws compared with open screws.
Open Access
Complications, revision fusions, readmissions, and utilization over a 1-year period after bone morphogenetic protein use during primary cervical spine fusions.
(The spine journal : official journal of the North American Spine Society, 2014-09) Goode, Adam P; Richardson, William J; Schectman, Robin M; Carey, Timothy S
Background context
Nationwide estimates examining bone morphogenetic protein (BMP) use with cervical spine fusions have been limited to perioperative outcomes.
Purpose
To determine the 1-year risk of complications, cervical revision fusions, hospital readmissions, and health care services utilization.
Study design
A retrospective cohort study from 2002 to 2009 using a nationwide claims database.
Patient sample
There were 61,937 primary cervical spine fusions of which 1,677 received BMP.
Outcome measures
Complications, revision fusions, 30-day hospital readmission, and health care utilization.
Methods
Data for these analyses come from the Thomson Reuters MarketScan Commercial Claims and Encounters Database 2010. Patients were aged 18 to 64 years, receiving and not receiving BMP with a primary (C2-C7) cervical spine fusion. All outcomes were defined by International Classification of Diseases, 9th edition Clinical Modification and Current Procedural and Terminology, 4th edition codes. Complications were analyzed as any complication and stratified by nervous system, wound, and dysphagia or hoarseness. Cervical revision fusions were determined in the 1-year follow-up. Hospital readmission discharge records defined 30-day hospital readmission and reason for the readmission. The utilization of at least one health care service of cervical spine imaging, epidural usage or rehabilitation service was examined. Poisson regression models were used to estimate the relative risk and 95% confidence interval (CI). Linear regression was used to determine the time to hospital readmission. Results were stratified by anterior or posterior and circumferential approaches.
Results
Patients receiving BMP were 29% more likely to have a complication (adjusted relative risk [aRR]=1.29 [95% CI, 1.14-1.46]) and a nervous system complication (aRR=1.42 [95% CI, 1.10-1.83]). Cervical revision fusions were more likely among patients receiving BMP (aRR=1.69 [95% CI, 1.35-2.13]). The risk of 30-day readmission was greater with BMP use (aRR=1.37 [95% CI, 1.07-1.73]) and readmission occurred 27.4% sooner on an average. Patients receiving BMP were more likely to receive computed tomography scans (aRR=1.34 [95% CI, 1.06-1.70]) and epidurals with anterior surgical approaches (aRR=1.29 [95% CI, 1.00-1.65]).
Conclusions
These findings question both the safety and effectiveness of off-label BMP use in primary cervical spine fusions.
Open Access
Determination of metarrestin (ML-246) in human plasma for a first-in-human clinical pharmacokinetic application by a simple and efficient uHPLC-MS/MS assay.
(Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2023-05) Richardson, William J; Zimmerman, Sara M; Reno, Annieka; Corvalan Cabanas, Natalia; Arisa, Oluwatobi; Rudloff, Udo; Figg, William D; Peer, Cody J
Metarrestin is a first-in-class small molecule inhibitor targeting the perinucleolar compartment, a subnuclear body associated with metastatic capacity. Promising preclinical results led to the clinical translation of the compound into a first-in-human phase I trial (NCT04222413). To characterize metarrestin's pharmacokinetic profile in humans, a uHPLC-MS/MS assay was developed and validated to determine the disposition of the drug in human plasma. Efficient sample preparation was accomplished through one-step protein precipitation paired with elution through a phospholipid filtration plate. Chromatographic separation was achieved with gradient elution through an Acuity UPLC® BEH C18 column (50 × 2.1 mm, 1.7 µm). Tandem mass spectrometry facilitated the detection of metarrestin and tolbutamide, the internal standard. The effective calibration range spanned 1-5000 ng/mL and was both accurate (range -5.9 % to 4.9 % deviation) and precise (≤9.0 %CV). Metarrestin proved stable (≤4.9 % degradation) under various assay-imposed conditions. Matrix effects, extraction efficiency, and process efficiency were assessed. Further, the assay was successfully able to determine the disposition of orally administered metarrestin in patients from the lowest dose cohort (1 mg) for 48 h post-administration. Thus, the validated analytical method detailed in this work is simple, sensitive, and clinically applicable.
Open Access
Differential expression of galectin-1 and its interactions with cells and laminins in the intervertebral disc.
(Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 2012-12) Jing, Liufang; So, Stephen; Lim, Shaun W; Richardson, William J; Fitch, Robert D; Setton, Lori A; Chen, Jun
Galectin-1 (Gal-1), an endogenous β-galactoside-binding protein, binds to laminins, which are highly expressed in the nucleus pulposus (NP) of the intervertebral disc (IVD). The objective of this study is to evaluate the expression of Gal-1 protein in IVD tissues during aging and the effect of Gal-1 on IVD cell adhesion to laminins. Tissues from rat, porcine, and human (scoliosis or disc degeneration) IVDs were used to evaluate Gal-1 expression via immunostaining, RT-PCR, and Western blot analysis. Attachment of isolated IVD cells (porcine and human) on select laminin isoforms (LM-111 and LM-511) was compared with/without pre-incubation with exogenous Gal-1. A biotinylated Gal-1(B-Gal-1) was used to evaluate for binding to IVD cells and to select for IVD cells by magnetic activated cell sorting (MACS). NP cells expressed high levels of Gal-1 protein as compared to anulus fibrosus (AF) cells in immature tissues, while exogenous Gal-1 increased both NP and AF cell attachment to laminins and exhibited a similar binding to both cell types in vitro. With aging, Gal-1 levels in NP tissue appeared to decrease. In addition, incubation with B-Gal-1 was able to promote the retention of more than 50% of IVD cells via MACS. Our results provide new findings for the presence and functional role of Gal-1 within IVDs. Similar staining patterns for Gal-1 and LM-511 in IVD tissue suggest that Gal-1 may serve as an adhesion molecule to interact with both cells and laminins. This MACS protocol may be useful for selecting pure IVD cells from mixed cells of pathological tissue.
Open Access
Distant Harrington rod migration 35 years after implantation.
(Journal of clinical neuroscience : official journal of the Neurosurgical Society of Australasia, 2013-10) Lark, Robert K; Caputo, Adam M; Brown, Christopher R; Michael, Keith W; Thacker, Julie K; Richardson, William J
Harrington rods have been successfully implanted in thousands of patients for the correction of scoliotic deformity since the 1950s. An exceedingly rare complication of Harrington rod placement is loosening with resultant migration. The authors present a 50-year-old woman who had a single Harrington rod placed when she was 15 years old. Thirty-five years later, she presented with acute sensory changes in her lower extremities. Imaging revealed rod failure and migration of the hardware distally, resulting in penetration of the wall of the rectum. Due to the unique anatomical position of the migrated hardware, sigmoidoscopy was used to directly visualize and remove the rod. The patient ultimately made a full recovery. Rod migration is an exceedingly rare complication that has been described only a few times since the introduction of Harrington rods over 60 years ago. The case herein is particularly unique given the extensive period of time that passed before migration (35 years) and the use of sigmoidoscopy for hardware removal.
Open Access
Experience with intrawound vancomycin powder for spinal deformity surgery.
(Spine, 2014-01) Martin, Joel R; Adogwa, Owoicho; Brown, Christopher R; Bagley, Carlos A; Richardson, William J; Lad, Shivanand P; Kuchibhatla, Maragatha; Gottfried, Oren N
Study design
Retrospective cohort study.
Objective
To evaluate the ability of local vancomycin powder to prevent deep wound infection after thoracolumbar and lumbar spinal fusion for open deformity cases.
Summary of background data
Recent studies report that local delivery of vancomycin powder is associated with a decrease in spinal surgical site infection (SSI). This study compares deformity fusion cases before and after the routine application of spinal vancomycin powder.
Methods
Posterior spinal deformity surgical procedures by a single institution were reviewed from January 2011 to April 2013. Routine application of vancomycin powder started in April 2012. Inclusion criteria included adult patients who underwent posterior fusion for deformity pathologies, including spondylolisthesis, kyphosis, sagittal imbalance, and scoliosis. Each cohort's baseline characteristics including infection risk factors, operative data, and rates of wound infection were compared. Associations between infection and vancomycin powder, with and without propensity score adjustment for risk factors were determined using logistic regression.
Results
A total of 306 patients were included in the study. All measured baseline and operative variables were statistically similar between untreated (n = 150) and those who received vancomycin powder (n = 156). No significant change in deep wound infection rate was seen between the control (5.3%) and intervention group (5.1%, P = 0.936). Logistic regression with and without propensity score adjusted for risk factors demonstrated that the use of vancomycin powder did not impact the development of SSI (odds ratio [95% confidence interval]: 1.01 [0.36-2.79], P = 0.9910) and (odds ratio [95% confidence interval]: 0.87 [0.31-2.42], P = 0.7876), respectively.
Conclusion
The local application of powdered vancomycin was not associated with a significant difference in the rate of deep SSI after spinal deformity surgery, and other treatment modalities are necessary to limit infection for this high-risk group. This study is in contrary to prior studies, which have reported a decrease in SSI with vancomycin powder.
Level of evidence
2.
Open Access
Hip Fracture in the Elderly Patients: A Sentinel Event.
(The American journal of hospice & palliative care, 2018-04) Koso, Riikka E; Sheets, Charles; Richardson, William J; Galanos, Anthony N
Importance
Hip fracture in the elderly patients is associated with increased morbidity and mortality. There is great need for advance care planning should a patient fail to rehabilitate or experience an adverse event during or after recovery. This study was performed to evaluate for palliative care consultation and changes in code status and/or advance directives in elderly patients with hip fracture.
Methods
We performed a retrospective review of 186 consecutive patients aged 65 years and older with a hip fracture due to a low-energy fall who underwent surgery at a large academic institution between August 1, 2013, and September 1, 2014. Risk factors assessed were patient demographics, home status, mobility, code status, comorbidities, medications, and hospitalizations prior to injury. Outcomes of interest included palliative care consultation, complications, mortality, and most recent code status, mobility, and home.
Results
About 186 patients with hip fractures were included. Three patients died, and 12 (6.5%) sustained major complications during admission. Nearly one-third (51 patients) died upon final follow-up approximately 1.5 years after surgery. Of the patients who died, palliative care consulted on 6 (11.8%) during initial admission. Eleven (21.6%) were full code at death. Three patients underwent cardiopulmonary resuscitation (CPR) and 1 underwent massive transfusion and extracorporeal membrane oxygenation prior to changing their code status to do not attempt resuscitation.
Conclusion
Hip fracture in elderly patients is an important opportunity to reassess the patient's personal health-care priorities. Advance directives, goals of care, and code status documentation should be updated in all elderly patients with hip fracture, should the patient's health decompensate.
Open Access
Identifying molecular phenotype of nucleus pulposus cells in human intervertebral disc with aging and degeneration.
(Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 2016-08) Tang, Xinyan; Jing, Liufang; Richardson, William J; Isaacs, Robert E; Fitch, Robert D; Brown, Christopher R; Erickson, Melissa M; Setton, Lori A; Chen, Jun
Previous study claimed that disc degeneration may be preceded by structure and matrix changes in the intervertebral disc (IVD) which coincide with the loss of distinct notochordally derived nucleus pulposus (NP) cells. However, the fate of notochordal cells and their molecular phenotype change during aging and degeneration in human are still unknown. In this study, a set of novel molecular phenotype markers of notochordal NP cells during aging and degeneration in human IVD tissue were revealed with immunostaining and flow cytometry. Furthermore, the potential of phenotype juvenilization and matrix regeneration of IVD cells in a laminin-rich pseudo-3D culture system were evaluated at day 28 by immunostaining, Safranin O, and type II collagen staining. Immunostaining and flow cytometry demonstrated that transcriptional factor Brachyury T, neuronal-related proteins (brain abundant membrane attached signal protein 1, Basp1; Neurochondrin, Ncdn; Neuropilin, Nrp-1), CD24, and CD221 were expressed only in juvenile human NP tissue, which suggested that these proteins may be served as the notochordal NP cell markers. However, the increased expression of CD54 and CD166 with aging indicated that they might be referenced as the potential biomarker for disc degeneration. In addition, 3D culture maintained most of markers in juvenile NP, and rescued the expression of Basp1, Ncdn, and Nrp 1 that disappeared in adult NP native tissue. These findings provided new insight into molecular profile that may be used to characterize the existence of a unique notochordal NP cells during aging and degeneration in human IVD cells, which will facilitate cell-based therapy for IVD regeneration. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1316-1326, 2016.
Open Access
In Silico Re-Optimization of Atezolizumab Dosing Using Population Pharmacokinetic Simulation and Exposure-Response Simulation.
(Journal of clinical pharmacology, 2023-06) Peer, Cody J; Schmidt, Keith T; Arisa, Oluwatobi; Richardson, William J; Paydary, Koosha; Goldstein, Daniel A; Gulley, James L; Figg, William D; Ratain, Mark J
Atezolizumab, a humanized monoclonal antibody against programmed cell death ligand 1 (PD-L1), was initially approved in 2016, around the same time that the sponsor published the minimum serum concentration to maintain the saturation of receptor occupancy (6 μg/mL). The initially approved dose regimen of 1200 mg every 3 weeks (q3w) was subsequently modified to 840 mg q2w or 1680 mg q4w through pharmacokinetic simulations. Yet, each standard regimen yields steady-state trough concentrations (C_MIN,SS ) far exceeding (≈ 40-fold) the stated target concentration. Additionally, the steady-state area under the plasma drug concentration-time curve (AUC_SS ) at 1200 mg q3w was significantly (P = .027) correlated with the probability of adverse events of special interest (AESIs) in patients with non-small cell lung cancer (NSCLC) and, coupled with excess exposure, this provides incentive to explore alternative dose regimens to lower the exposure burden while maintaining an effective C_MIN,SS . In this study, we first identified 840 mg q6w as an extended-interval regimen that could robustly maintain a serum concentration of 6 μg/mL (≥99% of virtual patients simulated, n = 1000), then applied this regimen to an approach that administers 2 "loading doses" of standard-interval regimens for a future clinical trial aiming to personalize dose regimens. Each standard dose was simulated for 2 loading doses, then 840 mg q6w thereafter; all yielded cycle-7 C_MIN,SS values of >6 μg/mL in >99% of virtual patients. Further, the AUC_SS from 840 mg q6w resulted in a flattening (P = .63) of the exposure-response relationship with adverse events of special interest (AESIs). We next aim to verify this in a clinical trial seeking to validate extended-interval dosing in a personalized approach using therapeutic drug monitoring.
Open Access
Injectable laminin-functionalized hydrogel for nucleus pulposus regeneration.
(Biomaterials, 2013-10) Francisco, Aubrey T; Mancino, Robert J; Bowles, Robby D; Brunger, Jonathan M; Tainter, David M; Chen, Yi-Te; Richardson, William J; Guilak, Farshid; Setton, Lori A
Cell delivery to the pathological intervertebral disc (IVD) has significant therapeutic potential for enhancing IVD regeneration. The development of injectable biomaterials that retain delivered cells, promote cell survival, and maintain or promote an NP cell phenotype in vivo remains a significant challenge. Previous studies have demonstrated NP cell - laminin interactions in the nucleus pulposus (NP) region of the IVD that promote cell attachment and biosynthesis. These findings suggest that incorporating laminin ligands into carriers for cell delivery may be beneficial for promoting NP cell survival and phenotype. Here, an injectable, laminin-111 functionalized poly(ethylene glycol) (PEG-LM111) hydrogel was developed as a biomaterial carrier for cell delivery to the IVD. We evaluated the mechanical properties of the PEG-LM111 hydrogel, and its ability to retain delivered cells in the IVD space. Gelation occurred in approximately 20 min without an initiator, with dynamic shear moduli in the range of 0.9-1.4 kPa. Primary NP cell retention in cultured IVD explants was significantly higher over 14 days when cells were delivered within a PEG-LM111 carrier, as compared to cells in liquid suspension. Together, these results suggest this injectable laminin-functionalized biomaterial may be an easy to use carrier for delivering cells to the IVD.
Open Access
Interleukin-17 synergizes with IFNγ or TNFα to promote inflammatory mediator release and intercellular adhesion molecule-1 (ICAM-1) expression in human intervertebral disc cells
(2011-01-01) Allen, Kyle D; Chen, Jun; Fitch, Robert; Gabr, Mostafa A; Helbling, Antonia R; Jing, Liufang; Richardson, William J; Setton, Lori A; Shamji, Mohammed F; Sinclair, S Michael
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.
Open Access
Interleukin-17 synergizes with IFNγ or TNFα to promote inflammatory mediator release and intercellular adhesion molecule-1 (ICAM-1) expression in human intervertebral disc cells.
(Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 2011-01) Gabr, Mostafa A; Jing, Liufang; Helbling, Antonia R; Sinclair, S Michael; Allen, Kyle D; Shamji, Mohammed F; Richardson, William J; Fitch, Robert D; Setton, Lori A; Chen, Jun
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.
Open Access
Kinematic and dynamic gait compensations in a rat model of lumbar radiculopathy and the effects of tumor necrosis factor-alpha antagonism.
(Arthritis research & therapy, 2011-08-26) Allen, Kyle D; Shamji, Mohammed F; Mata, Brian A; Gabr, Mostafa A; Sinclair, S Michael; Schmitt, Daniel O; Richardson, William J; Setton, Lori A
Tumor necrosis factor-α (TNFα) has received significant attention as a mediator of lumbar radiculopathy, with interest in TNF antagonism to treat radiculopathy. Prior studies have demonstrated that TNF antagonists can attenuate heightened nociception resulting from lumbar radiculopathy in the preclinical model. Less is known about the potential impact of TNF antagonism on gait compensations, despite being of clinical relevance. In this study, we expand on previous descriptions of gait compensations resulting from lumbar radiculopathy in the rat and describe the ability of local TNF antagonism to prevent the development of gait compensations, altered weight bearing, and heightened nociception.Eighteen male Sprague-Dawley rats were investigated for mechanical sensitivity, weight-bearing, and gait pre- and post-operatively. For surgery, tail nucleus pulposus (NP) tissue was collected and the right L5 dorsal root ganglion (DRG) was exposed (Day 0). In sham animals, NP tissue was discarded (n = 6); for experimental animals, autologous NP was placed on the DRG with or without 20 μg of soluble TNF receptor type II (sTNFRII, n = 6 per group). Spatiotemporal gait characteristics (open arena) and mechanical sensitivity (von Frey filaments) were assessed on post-operative Day 5; gait dynamics (force plate arena) and weight-bearing (incapacitance meter) were assessed on post-operative Day 6.High-speed gait characterization revealed animals with NP alone had a 5% decrease in stance time on their affected limbs on Day 5 (P ≤0.032). Ground reaction force analysis on Day 6 aligned with temporal changes observed on Day 5, with vertical impulse reduced in the affected limb of animals with NP alone (area under the vertical force-time curve, P <0.02). Concordant with gait, animals with NP alone also had some evidence of affected limb mechanical allodynia on Day 5 (P = 0.08) and reduced weight-bearing on the affected limb on Day 6 (P <0.05). Delivery of sTNFRII at the time of NP placement ameliorated signs of mechanical hypersensitivity, imbalanced weight distribution, and gait compensations (P <0.1).Our data indicate gait characterization has value for describing early limb dysfunctions in pre-clinical models of lumbar radiculopathy. Furthermore, TNF antagonism prevented the development of gait compensations subsequent to lumbar radiculopathy in our model.
Open Access
Molecular characterization of chordoma xenografts generated from a novel primary chordoma cell source and two chordoma cell lines.
(J Neurosurg Spine, 2014-09) Karikari, Isaac O; Gilchrist, Christopher L; Jing, Liufang; Alcorta, David A; Chen, Jun; Richardson, William J; Gabr, Mostafa A; Bell, Richard D; Kelley, Michael J; Bagley, Carlos A; Setton, Lori A
OBJECT: Chordoma cells can generate solid-like tumors in xenograft models that express some molecular characteristics of the parent tumor, including positivity for brachyury and cytokeratins. However, there is a dearth of molecular markers that relate to chordoma tumor growth, as well as the cell lines needed to advance treatment. The objective in this study was to isolate a novel primary chordoma cell source and analyze the characteristics of tumor growth in a mouse xenograft model for comparison with the established U-CH1 and U-CH2b cell lines. METHODS: Primary cells from a sacral chordoma, called "DVC-4," were cultured alongside U-CH1 and U-CH2b cells for more than 20 passages and characterized for expression of CD24 and brachyury. While brachyury is believed essential for driving tumor formation, CD24 is associated with healthy nucleus pulposus cells. Each cell type was subcutaneously implanted in NOD/SCID/IL2Rγ(null) mice. The percentage of solid tumors formed, time to maximum tumor size, and immunostaining scores for CD24 and brachyury (intensity scores of 0-3, heterogeneity scores of 0-1) were reported and evaluated to test differences across groups. RESULTS: The DVC-4 cells retained chordoma-like morphology in culture and exhibited CD24 and brachyury expression profiles in vitro that were similar to those for U-CH1 and U-CH2b. Both U-CH1 and DVC-4 cells grew tumors at rates that were faster than those for U-CH2b cells. Gross tumor developed at nearly every site (95%) injected with U-CH1 and at most sites (75%) injected with DVC-4. In contrast, U-CH2b cells produced grossly visible tumors in less than 50% of injected sites. Brachyury staining was similar among tumors derived from all 3 cell types and was intensely positive (scores of 2-3) in a majority of tissue sections. In contrast, differences in the pattern and intensity of staining for CD24 were noted among the 3 types of cell-derived tumors (p < 0.05, chi-square test), with evidence of intense and uniform staining in a majority of U-CH1 tumor sections (score of 3) and more than half of the DVC-4 tumor sections (scores of 2-3). In contrast, a majority of sections from U-CH2b cells stained modestly for CD24 (scores of 1-2) with a predominantly heterogeneous staining pattern. CONCLUSIONS: This is the first report on xenografts generated from U-CH2b cells in which a low tumorigenicity was discovered despite evidence of chordoma-like characteristics in vitro. For tumors derived from a primary chordoma cell and U-CH1 cell line, similarly intense staining for CD24 was observed, which may correspond to their similar potential to grow tumors. In contrast, U-CH2b tumors stained less intensely for CD24. These results emphasize that many markers, including CD24, may be useful in distinguishing among chordoma cell types and their tumorigenicity in vivo.
Open Access
Osteoimmunology: Interactions With the Immune System in Spinal Fusion.
(International journal of spine surgery, 2023-12) Bergin, Stephen M; Crutcher, Clifford L; Keeler, Carolyn; Rocos, Brett; Haglund, Michael M; Michael Guo, H; Gottfried, Oren N; Richardson, William J; Than, Khoi D
Spinal fusion is important for the clinical success of patients undergoing surgery, and the immune system plays an increasingly recognized role. Osteoimmunology is the study of the interactions between the immune system and bone. Inflammation impacts the osteogenic, osteoconductive, and osteoinductive properties of bone grafts and substitutes and ultimately influences the success of spinal fusion. Macrophages have emerged as important cells for coordinating the immune response following spinal fusion surgery, and macrophage-derived cytokines impact each phase of bone graft healing. This review explores the cellular and molecular immune processes that regulate bone homeostasis and healing during spinal fusion.
Open Access
Pharmacokinetics and tolerability of the dual TORC1/2 inhibitor sapanisertib in combination with the MEK inhibitor trametinib in dogs.
(Frontiers in veterinary science, 2022-01) Wei, Bih-Rong; Peer, Cody J; Richardson, William J; Hewitt, Stephen M; Figg, William D; Simpson, R Mark
Activation of one or both the Ras/MAPK and PI3K/Akt/mTOR signal transduction pathways are known to mediate oncogenicity of several canine and human cancers, including mucosal melanomas. Reciprocal cross activation between the two pathways can be a source of drug resistance. Consequently, oral dosing for plasma pharmacokinetic (PK) analysis and tolerability to a combination of sapanisertib, a dual TORC1/2 inhibitor, and trametinib, a MEK inhibitor, was evaluated in nontumor-bearing laboratory dogs for its potential application in parallel pathway targeting. Twelve dogs, divided into three equal cohorts, received either the combination or single agents. Animals were monitored for PK following single dose and 17-day repeat dosing, and by clinical observations, hematology, serum biochemistry, coagulation studies and urinalyses. A single trametinib dose (0.025 mg/kg), sulfated as dimethyl sulfoxide which enhanced its absorption, reached mean maximum concentration (C_max) 0.64 ng/mL [18% coefficient of variation (CV)] at a median time to maximum concentration (T_max) of 1.5 h (hr), and mean area under the concentration-time curve (AUC) 16.8 hr^*ng/mL (14%CV), which were similar when given alone or in combination with sapanisertib. A prolonged half-life afforded 3-4-fold plasma accumulation of trametinib with daily dosing, analogous to humans. Trametinib PK mirrored previous regulatory data in dogs, while exposure approximated some published human values but generally not all patients. Sapanisertib-alone in canine plasma following single 0.1 mg/kg dose [mean C_max 26.3 ng/mL (21%CV), median T_max 2.0 hr, and mean AUC 248 hr^*ng/mL (41%CV)] resembled levels in human therapeutic trials; whereas canine sapanisertib exposure was reduced when combined with trametinib, a known cytochrome P450 CYP3A4 inducer. Sex differences were not observed for either drug. Side effects upon repeat dosing with either or both drugs may include body weight loss, maldigestion, and cutaneous discoloration. The combination was tolerated without dose limiting toxicity, although clinical laboratory analyses revealed drug-induced acute-phase inflammation, proteinuria, and decreased blood reticulocytes, mild changes not necessitating intervention. Short-term results in dogs with this combination would appear to hold translational promise for clinical trial evaluation to target canine and possibly human melanoma, as well as other cancers having one or both signal transduction pathway activations.
Open Access
Proinflammatory cytokine expression profile in degenerated and herniated human intervertebral disc tissues.
(Arthritis and rheumatism, 2010-07) Shamji, Mohammed F; Setton, Lori A; Jarvis, Wingrove; So, Stephen; Chen, Jun; Jing, Liufang; Bullock, Robert; Isaacs, Robert E; Brown, Christopher; Richardson, William J
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.
Methods
Surgical IVD tissues were procured from patients with degenerative disc disease (n = 25) or herniated IVDs (n = 12); nondegenerated autopsy control tissue was also obtained (n = 8) from the anulus fibrosus and nucleus pulposus regions. Immunohistochemistry was performed for cell surface antigens (CD68 for macrophages, CD4 for lymphocytes) and various cytokines, with differences in cellularity and target immunoreactivity scores analyzed between surgical tissue groups and between autopsy control tissue regions.
Results
Immunoreactivity for IL-4, IL-6, IL-12, and interferon-gamma (IFNgamma) was modest in surgical IVD tissue, although expression was higher in herniated IVD samples and virtually nonexistent in control samples. The Th17 lymphocyte product IL-17 was present in >70% of surgical tissue fields, and among control samples was detected rarely in anulus fibrosus regions and modestly in nucleus pulposus regions. Macrophages were prevalent in surgical tissues, particularly herniated IVD samples, and lymphocytes were expectedly scarce. Control tissue revealed lesser infiltration by macrophages and a near absence of lymphocytes.
Conclusion
Greater IFNgamma positivity, macrophage presence, and cellularity in herniated IVDs suggests a pattern of Th1 lymphocyte activation in this pathology. Remarkable pathologic IVD tissue expression of IL-17 is a novel finding that contrasts markedly with low levels of IL-17 in autopsy control tissue. These findings suggest involvement of Th17 lymphocytes in the pathomechanism of disc degeneration.

Browsing by Author "Richardson, William J"

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