Mercury (Hg) pollutes marine ecosystems and accumulates in benthic species. This ecological case ... more Mercury (Hg) pollutes marine ecosystems and accumulates in benthic species. This ecological case study investigated the temporal accumulation of Hg in American lobster (Homarus americanus; H. Milne Edwards, 1837) from coastal Maine (Casco Bay, ME, USA). We analyzed total Hg levels in legal-sized lobsters (carapace length: 8.255–12.5 cm; n = 34) collected during the early (May–July 1) or late (July 15–October) recreational harvest seasons. Morphometric data show that body size correlates with body weight (R2 = 0.76; p < 0.001), and average body sizes were similar in early and late seasons. The average chelipod size was ~7% larger in male lobsters (p < 0.02), reflecting sexual dimorphism. Hg levels in select tissues from boiled lobsters were analyzed using atomic absorption spectroscopy. Hg in ambient water was undetectable, indicating that Hg in tissues reflects bioaccumulation. Hg content correlated with the lengths (cm) and weights (g) of cephalothorax, carapace, chelipod, and hepatopancreas in both male and female lobsters. Total Hg levels in most tissues were within safe and acceptable limits for human consumption (<0.2 ppm). Compared to late-season lobsters, early-season lobsters had significantly higher Hg levels in tail (~55% increase; 0.130 ppm vs. 0.084 ppm; p < 0.05) and hepatopancreas tissues (~29% increase; 0.099 ppm vs. 0.077 ppm; p < 0.05), suggesting that seasonal factors influence Hg content (e.g., spring river runoff, lobster migration, inert biological cycles). Observed seasonal fluctuations in lobster Hg levels may inform future strategies for mitigating pollution in coastal marine ecosystems.
Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control o... more Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control of their Antarctic prey species. However, population trends and genetic diversity of leopard seals remain understudied, limiting our understanding of their ecological role. We investigated the genetic diversity, effective population size and demographic history of leopard seals to provide fundamental data that contextualizes their predatory influence on Antarctic ecosystems. Ninety leopard seals were sampled from the northern Antarctic Peninsula during the austral summers of 2008-2019 and a 405bp segment of the mitochondrial control region was sequenced for each individual. We uncovered moderate levels of nucleotide (π = 0.013) and haplotype (Hd = 0.96) diversity, and the effective population size was estimated at around 24,000 individuals (NE = 24,376; 95% CI: 16,876-33,126). Consistent with findings from other ice-breeding pinnipeds, Bayesian skyline analysis also revealed evidence for population expansion during the last glacial maximum, suggesting that historical population growth may have been boosted by an increase in the abundance of sea ice. Although leopard seals can be found in warmer, sub-Antarctic locations, the species' core habitat is centered on the Antarctic, making it inherently vulnerable to the loss of sea ice habitat due to climate change. Therefore, detailed assessments of past and present leopard seal population trends are needed to inform policies for Antarctic ecosystems.
The biological interface between an orthopedic implant and the surrounding host tissue may have a... more The biological interface between an orthopedic implant and the surrounding host tissue may have a dramatic effect upon clinical outcome. Desired effects include bony ingrowth (osseointegration), stimulation of osteogenesis (osteoinduction), increased vascularization, and improved mechanical stability. Implant loosening, fibrous encapsulation, corrosion, infection, and inflammation, as well as physical mismatch may have deleterious clinical effects. This is particularly true of implants used in the reconstruction of load-bearing synovial joints such as the knee, hip, and the shoulder. The surfaces of orthopedic implants have evolved from solid-smooth to roughened-coarse and most recently, to porous in an effort to create a three-dimensional architecture for bone apposition and osseointegration. Total joint surgeries are increasingly performed in younger individuals with a longer life expectancy, and therefore, the postimplantation lifespan of devices must increase commensurately. This review discusses advancements in biomaterials science and cell-based therapies that may further improve orthopedic success rates. We focus on material and biological properties of orthopedic implants fabricated from porous metal and highlight some relevant developments in stem-cell research. We posit that the ideal primary and revision orthopedic load-bearing metal implants are highly porous and may be chemically modified to induce stem cell growth and osteogenic differentiation, while minimizing inflammation and infection. We conclude that integration of new biological, chemical, and mechanical methods is likely to yield more effective strategies to control and modify the implant-bone interface and thereby improve long-term clinical outcomes.
Total knee arthroplasty (TKA) is a durable and reliable procedure to alleviate pain and improve j... more Total knee arthroplasty (TKA) is a durable and reliable procedure to alleviate pain and improve joint function. However, failures related to flexion instability sometimes occur. The goal of this study was to define biological differences between tissues from patients with and without flexion instability of the knee after TKA. Human knee joint capsule tissues were collected at the time of primary or revision TKAs and analyzed by RT-qPCR and RNA-seq, revealing novel patterns of differential gene expression between the two groups. Interestingly, genes related to collagen production and extracellular matrix (ECM) degradation were higher in samples from patients with flexion instability. Partitioned clustering analyses further emphasized differential gene expression patterns between sample types that may help guide clinical interpretations of this complication. Future efforts to disentangle the effects of physical and biological (e.g., transcriptomic modifications) risk factors will aid in further characterizing and avoiding flexion instability after TKA.
Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. A... more Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. Adjunct pharmacologic modalities may permit clinical prevention and treatment of recalcitrant joint contractures. We investigated the therapeutic potential of rosiglitazone by intra-articular delivery via oligo[poly(ethylene glycol)fumarate] (OPF) hydrogels in an established rabbit model of arthrofibrosis. OPF hydrogels loaded with rosiglitazone were characterized for drug elution properties upon soaking in minimum essential media (MEM) with 10% fetal bovine serum and measurements of drug concentrations via High Performance Liquid Chromatography (HPLC). Drug-loaded scaffolds were surgically implanted into 24 skeletally mature female New Zealand White rabbits that were divided into equal groups receiving OPF hydrogels loaded with rosiglitazone (1.67 mg), or vehicle control (10 ml DMSO). After 8 weeks of joint immobilization, rabbits were allowed unrestricted cage activity for 16 weeks. Contracture angles of rabbit limbs treated with rosiglitazone showed statistically significant improvements in flexion compared to control animals (mean angles, respectively, 64.4˚vs. 53.3˚, p < 0.03). At time of sacrifice (week 24), animals in the rosiglitazone group continued to exhibit less joint contracture than controls (119.0˚vs. 99.5˚, p ¼ 0.014). The intra-articular delivery of rosiglitazone using implanted OPF hydrogels decreases flexion contractures in a rabbit model of arthrofibrosis without causing adverse effects (e.g., gross inflammation or arthritis). Statement of Clinical Significance: Post-traumatic joint contractures are common and debilitating, with limited available treatment options. Pharmacologic interventions can potentially prevent and treat such contractures. This study is translational in that a commercially approved medication has been repurposed through a novel delivery device.
Epigenetic changes in articular chondrocytes are associated with osteoarthritis (OA) disease prog... more Epigenetic changes in articular chondrocytes are associated with osteoarthritis (OA) disease progression. Numerous studies have identified differentially methylated cytosines in OA tissues; however, the consequences of altered CpG methylation at single nucleotides on gene expression and phenotypes are difficult to predict. With the objective of detecting novel genes relevant to OA, we conducted a genome-wide assessment of differentially methylated sites (DMSs) and differentially methylated regions (DMRs). DNA was extracted from visually damaged and normal appearing, non-damaged human knee articular cartilage from the same joint and then subjected to reduced representation bisulfite sequencing. DMRs were identified using a genome-wide systematic bioinformatics approach. A sliding-window of 500bp was used for screening the genome for regions with clusters of DMSs. Gene expression levels were assessed and cell culture demethylation experiments were performed to further examine top candidate genes associated with damaged articular cartilage. More than 1,000 DMRs were detected in damaged osteoarthritic cartilage. Nineteen of these contained five or more DMSs and were located in gene promoters or first introns and exons. Gene expression assessment revealed that hypermethylated DMRs in damaged samples were more consistently associated with gene repression than hypomethylated DMRs were with gene activation. Accordingly, a demethylation agent induced expression of most hypermethylated genes in chondrocytes. Our study revealed the utility of a systematic DMR search as an alternative to focusing on single nucleotide data. In particular, this approach uncovered promising candidates for functional studies such as the hypermethylated protein-coding genes FOXP4 and SHROOM1, which appear to be linked to OA pathology in humans and warrant further investigation.
Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pa... more Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pathogenesis of arthrofibrosis, a debilitating condition after joint replacement and other orthopedic procedures. Cyclooxygenase 2 (COX-2) inhibitors may mitigate the inflammatory response and formation of arthrofibrosis, but oral delivery is associated with risk of systemic side effects in many patients. The nonsteroidal anti-inflammatory drug, celecoxib, may have therapeutic benefits for arthrofibrosis, but current methods for its local delivery (e.g., biologically derived microspheres) are not translatable to immediate clinical use. Therefore, we investigated the use of a drug scaffold for sustainable intra-articular delivery of therapeutic doses of celecoxib. Materials and Methods: Celecoxib was eluted from clinically approved biodegradable collagen membranes over 7 days as measured by UV spectroscopy and high-performance liquid chromatography/mass spectroscopy. Eluted concentrations of celecoxib were examined for toxicity (live/dead staining) and profibrotic gene expression (real-time-quantitative polymerase chain reaction) in rabbit knee capsular fibroblasts in vitro. Results: Sustained concentrations of celecoxib eluted from the membrane over 7 days from both a wet and dry scaffold, with a burst release (30-45%) of celecoxib in the first 2 h. Rabbit cells treated with eluted concentrations experienced a toxic response to the burst release doses, and inhibitory effects on profibrotic genes were seen in response to the sustained doses eluted from the scaffold. Conclusions: This study characterized the novel use of collagen scaffolds for intra-articular drug delivery to treat arthrofibrosis. Scaffolds exhibit celecoxib release through an initial burst release followed by sustained release of antifibrotic doses over 7 days. Thus, collagen scaffolds are promising for clinician-directed treatment of arthrofibrosis.
Substance P (SP), a neurotransmitter released after injury, has been linked to deregulated tissue... more Substance P (SP), a neurotransmitter released after injury, has been linked to deregulated tissue repair and fibrosis in musculoskeletal tissues and other organs. Although SP inhibition is an effective treatment for nausea, it has not been previously considered as an anti-fibrotic therapy. Although there are extensive medical records of individuals who have used SP antagonists, our analysis of human registry data revealed that patients receiving these antagonists and arthroplasty are exceedingly rare, thus precluding a clinical evaluation of their potential effects in the context of arthrofibrosis. Therefore, we pursued in vivo studies to assess the effect of SP inhibition early after injury on pro-fibrotic gene expression and contractures in an animal model of post-traumatic joint stiffening. Skeletally mature rabbits (n=24) underwent surgically induced severe joint contracture, while injected with either fosaprepitant (a selective SP antagonist) or saline (control) early after surgery (3, 6, 12, and 24 hours). Biomechanical testing revealed that differences in mean contracture angles between the groups were not statistically significant (p=0.27), suggesting that the drug neither mitigates nor exacerbates joint contracture. However, microarray gene expression analysis revealed that mRNA levels for proteins related to cell signaling, proangiogenic, pro-inflammatory and collagen matrix production were significantly different between control and fosaprepitant treated rabbits (p<0.05). Hence, our study demonstrates that inhibition of SP alters expression of pro-fibrotic genes in vivo. This finding will motivate future studies to optimize interventions that target SP to reduce the formation of post-traumatic joint contractures.
Total hip and knee arthroplasty are effective interventions for management of end-stage arthritis... more Total hip and knee arthroplasty are effective interventions for management of end-stage arthritis. Indeed, about seven million Americans are currently living with artificial hip and knee joints. The majority of these individuals, however, will outlive their implants and require revision surgeries, mostly due to poor implant osseointegration and aseptic loosening. Revisions are potentially avoidable with better management of patient-related risk factors that affect the osseointegration of orthopedic implants. In this review, we summarize the published clinical literature on the role of demographics, biologic factors, comorbidities, medications, and aseptic loosening risk. We focus on several systemic and local factors that are particularly relevant to implant osseointegration. Examples include physiological and molecular processes that are linked to hyperglycemia, oxidative stress, metabolic syndrome, and dyslipidemia. We discuss how orthopedic implant osseointegration can be affected by a number of molecular therapies that are antiresorptive or bone anabolic (i.e., calcium, vitamin D, bisphosphonates, calcitonin, strontium, hormone replacement therapy, selective estrogen receptor modulators).
Background: Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are cha... more Background: Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are challenging to manage and often require surgical reconstructions with limited long-term success. Thus, characterizations of these tissues are urgently needed to better understand cellular mechanisms that regulate tissue homeostasis and healing. Explant culturing systems allow for ex vivo analysis of tissues in an environment that mimics the native microenvironment in vivo. Methods: Collaborative efforts within our institution facilitated the establishment of a novel explant culturing system. Tissue specimens cultured in single wells, with individual applied loading and/or biological environment, allowed characterization of tissue cultured under a variety of biological loading conditions. Quantitative PCR analysis for selected gene markers was our primary outcome. Results: Data were stratified for analysis by either culture environment or loading condition. Our gene expression results show that specimens clustered by culture condition may differ in molecular markers related to ECM production (e.g., Col1a1, Adamts4) and/or organization (e.g., Tnc, Dnc). In contrast, loading condition did significantly alter the median gene expression levels of tissues in comparison to unloaded control samples, although gene expression values related to ECM degradation (e.g., Mmp1, Mmp10) were altered in tendons cultured under tension in the device. Conclusion: Our study demonstrates promising utility of a novel explant culturing system for further characterization of musculoskeletal tissues such as native tendons and ligaments, as well as pathologic fibrotic tissues resulting from arthrofibrosis or Dupuytren's disease.
Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control o... more Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control of their Antarctic prey species. However, population trends and genetic diversity of leopard seals remain understudied, limiting our understanding of their ecological role. We investigated the genetic diversity, effective population size and demographic history of leopard seals to provide fundamental data that contextualizes their predatory influence on Antarctic ecosystems. Ninety leopard seals were sampled in the northern Antarctic Peninsula, during the austral summers of 2008–2019 and a 405bp region of the mitochondrial control region was sequenced for each individual. We uncovered moderate levels of nucleotide (π = 0.013) and haplotype (Hd = 0.96) diversity, and the effective population size was estimated at around 24,000 individuals (NE= 24,376; range 16,876 – 33,126). Consistent with findings from other ice-breeding pinnipeds, Bayesian skyline analysis also revealed evidence for pop...
Metal orthopedic implants are largely biocompatible and generally achieve long-term structural fi... more Metal orthopedic implants are largely biocompatible and generally achieve long-term structural fixation. However, some orthopedic implants may loosen over time even in the absence of infection. In vivo fixation failure is multifactorial, but the fundamental biological defect is cellular dysfunction at the host-implant interface. Strategies to reduce the risk of short- and long-term loosening include surface modifications, implant metal alloy type, and adjuvant substances such as polymethylmethacrylate cement. Surface modifications (e.g., increased surface rugosity) can increase osseointegration and biological ingrowth of orthopedic implants. However, the localized responses of cells to implant surface modifications need to be better characterized. As an in vitro model for investigating cellular responses to metallic orthopedic implants, we cultured mesenchymal stromal/stem cells (MSCs) on clinical-grade titanium disks (Ti6Al4V) that differed in surface roughness as high (porous-structured), medium (grit-blasted), and low (bead-blasted). Topological characterization of clinically relevant Ti materials combined with differential mRNA expression analyses (RNA-seq and RT-qPCR) revealed alterations to the biological phenotype of cells cultured on titanium structures that favor early ECM production and observable responses to oxidative stress and heavy metal stress. These results provide a descriptive model for the interpretation of cellular responses at the interface between native host tissues and 3-D printed modular orthopedic implants, and will guide future studies aimed at increasing the long-term retention of such materials after TJA.
This is a PDF file of an article that has undergone enhancements after acceptance, such as the ad... more This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Purpose of Review MicroRNAs (miRNAs) are single-stranded, short (~22 nt) non-coding RNAs that con... more Purpose of Review MicroRNAs (miRNAs) are single-stranded, short (~22 nt) non-coding RNAs that control gene expression in most metazoan taxa. These vital post-transcriptional regulators are emerging as a novel class of relatively well-conserved biomarkers useful to molecular ecologists working on non-model marine organisms. The purpose of this review is to provide researchers with a brief background on miRNAs and to explore recent applications in marine biology. Recent Findings MiRNA datasets have been broadly employed in studies concerning commercially important species (oysters and crustaceans), phylogenetics (particularly deep evolutionary splits), and environmental stressor responses (temperature and salinity). Most progress has been made in the characterization of cnidarian miRNAs and bivalve and crustacean immune-related miRNAs. The use of miRNAs in phylogenetics is still under debate due to the secondary loss of miRNAs in some lineages, but they have been successfully applied in the resolution of deep evolutionary splits. Finally, miRNAs have been investigated in abiotic stress responses, but data interpretation is limited by the high number of species-specific miRNAs detected in these studies. Improvements in miRNA database curation and functional annotation should provide more confidence in their use. Summary Due to their evolutionary conservation, resilience to degradation, and amenable bioinformatics workflows, miRNAs are a powerful molecular tool in marine genomics. MiRNA investigations regarding environmental stress response will be particularly useful due to their potential to reveal physiological alterations and disease. Thus, they may be ultimately utilized as bio-indicators of environmental health.
Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pa... more Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pathogenesis of arthrofibrosis, a debilitating condition after joint replacement and other orthopedic procedures. Cyclooxygenase 2 (COX-2) inhibitors may mitigate the inflammatory response and formation of arthrofibrosis, but oral delivery is associated with risk of systemic side effects in many patients. The nonsteroidal anti-inflammatory drug, celecoxib, may have therapeutic benefits for arthrofibrosis, but current methods for its local delivery (e.g., biologically derived microspheres) are not translatable to immediate clinical use. Therefore, we investigated the use of a drug scaffold for sustainable intra-articular delivery of therapeutic doses of celecoxib. Materials and Methods: Celecoxib was eluted from clinically approved biodegradable collagen membranes over 7 days as measured by UV spectroscopy and high-performance liquid chromatography/mass spectroscopy. Eluted concentrations of celecoxib were examined for toxicity (live/dead staining) and profibrotic gene expression (real-time-quantitative polymerase chain reaction) in rabbit knee capsular fibroblasts in vitro. Results: Sustained concentrations of celecoxib eluted from the membrane over 7 days from both a wet and dry scaffold, with a burst release (30-45%) of celecoxib in the first 2 h. Rabbit cells treated with eluted concentrations experienced a toxic response to the burst release doses, and inhibitory effects on profibrotic genes were seen in response to the sustained doses eluted from the scaffold. Conclusions: This study characterized the novel use of collagen scaffolds for intra-articular drug delivery to treat arthrofibrosis. Scaffolds exhibit celecoxib release through an initial burst release followed by sustained release of antifibrotic doses over 7 days. Thus, collagen scaffolds are promising for clinician-directed treatment of arthrofibrosis.
Journal of Plastic, Reconstructive & Aesthetic Surgery, 2019
Mesenchymal stromal cells (MSCs) secrete many soluble growth factors and have previously been sho... more Mesenchymal stromal cells (MSCs) secrete many soluble growth factors and have previously been shown to stimulate nerve regeneration. MSC-seeded processed nerve allografts could potentially be a promising method for large segmental motor nerve injuries. Further progress in our understanding of how the functions of MSCs can be leveraged for peripheral nerve repair is required before making clinical translation. The present study, therefore, investigated whether interactions of adipose-derived MSCs with decellularized nerve allografts can improve gene and protein expression of growth factors that may support nerve regeneration. Human nerve allografts (n = 30) were decellularized and seeded with undifferentiated human adipose-derived MSCs. Subsequently, the MSCs and MSC-seeded grafts were isolated on days 3, 7, 14, and 21 in culture for RNA expression analysis by qRT-PCR. Evaluated genes Parts of this article have been presented at the following meeting/conference: 1.
Journal of orthopaedic research : official publication of the Orthopaedic Research Society, Jan 14, 2018
Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. A... more Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. Adjunct pharmacologic modalities may permit clinical prevention and treatment of recalcitrant joint contractures. We investigated the therapeutic potential of rosiglitazone by intra-articular delivery via oligo[poly(ethylene glycol)fumarate] (OPF) hydrogels in an established rabbit model of arthrofibrosis. OPF hydrogels loaded with rosiglitazone were characterized for drug elution properties upon soaking in minimum essential media (MEM) with 10% fetal bovine serum and measurements of drug concentrations via High Performance Liquid Chromatography (HPLC). Drug-loaded scaffolds were surgically implanted into 24 skeletally mature female New Zealand White rabbits that were divided into equal groups receiving OPF hydrogels loaded with rosiglitazone (1.67 mg), or vehicle control (10 µL DMSO). After 8 weeks of joint immobilization, rabbits were allowed unrestricted cage activity for 16 weeks. Con...
Mercury (Hg) pollutes marine ecosystems and accumulates in benthic species. This ecological case ... more Mercury (Hg) pollutes marine ecosystems and accumulates in benthic species. This ecological case study investigated the temporal accumulation of Hg in American lobster (Homarus americanus; H. Milne Edwards, 1837) from coastal Maine (Casco Bay, ME, USA). We analyzed total Hg levels in legal-sized lobsters (carapace length: 8.255–12.5 cm; n = 34) collected during the early (May–July 1) or late (July 15–October) recreational harvest seasons. Morphometric data show that body size correlates with body weight (R2 = 0.76; p < 0.001), and average body sizes were similar in early and late seasons. The average chelipod size was ~7% larger in male lobsters (p < 0.02), reflecting sexual dimorphism. Hg levels in select tissues from boiled lobsters were analyzed using atomic absorption spectroscopy. Hg in ambient water was undetectable, indicating that Hg in tissues reflects bioaccumulation. Hg content correlated with the lengths (cm) and weights (g) of cephalothorax, carapace, chelipod, and hepatopancreas in both male and female lobsters. Total Hg levels in most tissues were within safe and acceptable limits for human consumption (<0.2 ppm). Compared to late-season lobsters, early-season lobsters had significantly higher Hg levels in tail (~55% increase; 0.130 ppm vs. 0.084 ppm; p < 0.05) and hepatopancreas tissues (~29% increase; 0.099 ppm vs. 0.077 ppm; p < 0.05), suggesting that seasonal factors influence Hg content (e.g., spring river runoff, lobster migration, inert biological cycles). Observed seasonal fluctuations in lobster Hg levels may inform future strategies for mitigating pollution in coastal marine ecosystems.
Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control o... more Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control of their Antarctic prey species. However, population trends and genetic diversity of leopard seals remain understudied, limiting our understanding of their ecological role. We investigated the genetic diversity, effective population size and demographic history of leopard seals to provide fundamental data that contextualizes their predatory influence on Antarctic ecosystems. Ninety leopard seals were sampled from the northern Antarctic Peninsula during the austral summers of 2008-2019 and a 405bp segment of the mitochondrial control region was sequenced for each individual. We uncovered moderate levels of nucleotide (π = 0.013) and haplotype (Hd = 0.96) diversity, and the effective population size was estimated at around 24,000 individuals (NE = 24,376; 95% CI: 16,876-33,126). Consistent with findings from other ice-breeding pinnipeds, Bayesian skyline analysis also revealed evidence for population expansion during the last glacial maximum, suggesting that historical population growth may have been boosted by an increase in the abundance of sea ice. Although leopard seals can be found in warmer, sub-Antarctic locations, the species' core habitat is centered on the Antarctic, making it inherently vulnerable to the loss of sea ice habitat due to climate change. Therefore, detailed assessments of past and present leopard seal population trends are needed to inform policies for Antarctic ecosystems.
The biological interface between an orthopedic implant and the surrounding host tissue may have a... more The biological interface between an orthopedic implant and the surrounding host tissue may have a dramatic effect upon clinical outcome. Desired effects include bony ingrowth (osseointegration), stimulation of osteogenesis (osteoinduction), increased vascularization, and improved mechanical stability. Implant loosening, fibrous encapsulation, corrosion, infection, and inflammation, as well as physical mismatch may have deleterious clinical effects. This is particularly true of implants used in the reconstruction of load-bearing synovial joints such as the knee, hip, and the shoulder. The surfaces of orthopedic implants have evolved from solid-smooth to roughened-coarse and most recently, to porous in an effort to create a three-dimensional architecture for bone apposition and osseointegration. Total joint surgeries are increasingly performed in younger individuals with a longer life expectancy, and therefore, the postimplantation lifespan of devices must increase commensurately. This review discusses advancements in biomaterials science and cell-based therapies that may further improve orthopedic success rates. We focus on material and biological properties of orthopedic implants fabricated from porous metal and highlight some relevant developments in stem-cell research. We posit that the ideal primary and revision orthopedic load-bearing metal implants are highly porous and may be chemically modified to induce stem cell growth and osteogenic differentiation, while minimizing inflammation and infection. We conclude that integration of new biological, chemical, and mechanical methods is likely to yield more effective strategies to control and modify the implant-bone interface and thereby improve long-term clinical outcomes.
Total knee arthroplasty (TKA) is a durable and reliable procedure to alleviate pain and improve j... more Total knee arthroplasty (TKA) is a durable and reliable procedure to alleviate pain and improve joint function. However, failures related to flexion instability sometimes occur. The goal of this study was to define biological differences between tissues from patients with and without flexion instability of the knee after TKA. Human knee joint capsule tissues were collected at the time of primary or revision TKAs and analyzed by RT-qPCR and RNA-seq, revealing novel patterns of differential gene expression between the two groups. Interestingly, genes related to collagen production and extracellular matrix (ECM) degradation were higher in samples from patients with flexion instability. Partitioned clustering analyses further emphasized differential gene expression patterns between sample types that may help guide clinical interpretations of this complication. Future efforts to disentangle the effects of physical and biological (e.g., transcriptomic modifications) risk factors will aid in further characterizing and avoiding flexion instability after TKA.
Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. A... more Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. Adjunct pharmacologic modalities may permit clinical prevention and treatment of recalcitrant joint contractures. We investigated the therapeutic potential of rosiglitazone by intra-articular delivery via oligo[poly(ethylene glycol)fumarate] (OPF) hydrogels in an established rabbit model of arthrofibrosis. OPF hydrogels loaded with rosiglitazone were characterized for drug elution properties upon soaking in minimum essential media (MEM) with 10% fetal bovine serum and measurements of drug concentrations via High Performance Liquid Chromatography (HPLC). Drug-loaded scaffolds were surgically implanted into 24 skeletally mature female New Zealand White rabbits that were divided into equal groups receiving OPF hydrogels loaded with rosiglitazone (1.67 mg), or vehicle control (10 ml DMSO). After 8 weeks of joint immobilization, rabbits were allowed unrestricted cage activity for 16 weeks. Contracture angles of rabbit limbs treated with rosiglitazone showed statistically significant improvements in flexion compared to control animals (mean angles, respectively, 64.4˚vs. 53.3˚, p < 0.03). At time of sacrifice (week 24), animals in the rosiglitazone group continued to exhibit less joint contracture than controls (119.0˚vs. 99.5˚, p ¼ 0.014). The intra-articular delivery of rosiglitazone using implanted OPF hydrogels decreases flexion contractures in a rabbit model of arthrofibrosis without causing adverse effects (e.g., gross inflammation or arthritis). Statement of Clinical Significance: Post-traumatic joint contractures are common and debilitating, with limited available treatment options. Pharmacologic interventions can potentially prevent and treat such contractures. This study is translational in that a commercially approved medication has been repurposed through a novel delivery device.
Epigenetic changes in articular chondrocytes are associated with osteoarthritis (OA) disease prog... more Epigenetic changes in articular chondrocytes are associated with osteoarthritis (OA) disease progression. Numerous studies have identified differentially methylated cytosines in OA tissues; however, the consequences of altered CpG methylation at single nucleotides on gene expression and phenotypes are difficult to predict. With the objective of detecting novel genes relevant to OA, we conducted a genome-wide assessment of differentially methylated sites (DMSs) and differentially methylated regions (DMRs). DNA was extracted from visually damaged and normal appearing, non-damaged human knee articular cartilage from the same joint and then subjected to reduced representation bisulfite sequencing. DMRs were identified using a genome-wide systematic bioinformatics approach. A sliding-window of 500bp was used for screening the genome for regions with clusters of DMSs. Gene expression levels were assessed and cell culture demethylation experiments were performed to further examine top candidate genes associated with damaged articular cartilage. More than 1,000 DMRs were detected in damaged osteoarthritic cartilage. Nineteen of these contained five or more DMSs and were located in gene promoters or first introns and exons. Gene expression assessment revealed that hypermethylated DMRs in damaged samples were more consistently associated with gene repression than hypomethylated DMRs were with gene activation. Accordingly, a demethylation agent induced expression of most hypermethylated genes in chondrocytes. Our study revealed the utility of a systematic DMR search as an alternative to focusing on single nucleotide data. In particular, this approach uncovered promising candidates for functional studies such as the hypermethylated protein-coding genes FOXP4 and SHROOM1, which appear to be linked to OA pathology in humans and warrant further investigation.
Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pa... more Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pathogenesis of arthrofibrosis, a debilitating condition after joint replacement and other orthopedic procedures. Cyclooxygenase 2 (COX-2) inhibitors may mitigate the inflammatory response and formation of arthrofibrosis, but oral delivery is associated with risk of systemic side effects in many patients. The nonsteroidal anti-inflammatory drug, celecoxib, may have therapeutic benefits for arthrofibrosis, but current methods for its local delivery (e.g., biologically derived microspheres) are not translatable to immediate clinical use. Therefore, we investigated the use of a drug scaffold for sustainable intra-articular delivery of therapeutic doses of celecoxib. Materials and Methods: Celecoxib was eluted from clinically approved biodegradable collagen membranes over 7 days as measured by UV spectroscopy and high-performance liquid chromatography/mass spectroscopy. Eluted concentrations of celecoxib were examined for toxicity (live/dead staining) and profibrotic gene expression (real-time-quantitative polymerase chain reaction) in rabbit knee capsular fibroblasts in vitro. Results: Sustained concentrations of celecoxib eluted from the membrane over 7 days from both a wet and dry scaffold, with a burst release (30-45%) of celecoxib in the first 2 h. Rabbit cells treated with eluted concentrations experienced a toxic response to the burst release doses, and inhibitory effects on profibrotic genes were seen in response to the sustained doses eluted from the scaffold. Conclusions: This study characterized the novel use of collagen scaffolds for intra-articular drug delivery to treat arthrofibrosis. Scaffolds exhibit celecoxib release through an initial burst release followed by sustained release of antifibrotic doses over 7 days. Thus, collagen scaffolds are promising for clinician-directed treatment of arthrofibrosis.
Substance P (SP), a neurotransmitter released after injury, has been linked to deregulated tissue... more Substance P (SP), a neurotransmitter released after injury, has been linked to deregulated tissue repair and fibrosis in musculoskeletal tissues and other organs. Although SP inhibition is an effective treatment for nausea, it has not been previously considered as an anti-fibrotic therapy. Although there are extensive medical records of individuals who have used SP antagonists, our analysis of human registry data revealed that patients receiving these antagonists and arthroplasty are exceedingly rare, thus precluding a clinical evaluation of their potential effects in the context of arthrofibrosis. Therefore, we pursued in vivo studies to assess the effect of SP inhibition early after injury on pro-fibrotic gene expression and contractures in an animal model of post-traumatic joint stiffening. Skeletally mature rabbits (n=24) underwent surgically induced severe joint contracture, while injected with either fosaprepitant (a selective SP antagonist) or saline (control) early after surgery (3, 6, 12, and 24 hours). Biomechanical testing revealed that differences in mean contracture angles between the groups were not statistically significant (p=0.27), suggesting that the drug neither mitigates nor exacerbates joint contracture. However, microarray gene expression analysis revealed that mRNA levels for proteins related to cell signaling, proangiogenic, pro-inflammatory and collagen matrix production were significantly different between control and fosaprepitant treated rabbits (p<0.05). Hence, our study demonstrates that inhibition of SP alters expression of pro-fibrotic genes in vivo. This finding will motivate future studies to optimize interventions that target SP to reduce the formation of post-traumatic joint contractures.
Total hip and knee arthroplasty are effective interventions for management of end-stage arthritis... more Total hip and knee arthroplasty are effective interventions for management of end-stage arthritis. Indeed, about seven million Americans are currently living with artificial hip and knee joints. The majority of these individuals, however, will outlive their implants and require revision surgeries, mostly due to poor implant osseointegration and aseptic loosening. Revisions are potentially avoidable with better management of patient-related risk factors that affect the osseointegration of orthopedic implants. In this review, we summarize the published clinical literature on the role of demographics, biologic factors, comorbidities, medications, and aseptic loosening risk. We focus on several systemic and local factors that are particularly relevant to implant osseointegration. Examples include physiological and molecular processes that are linked to hyperglycemia, oxidative stress, metabolic syndrome, and dyslipidemia. We discuss how orthopedic implant osseointegration can be affected by a number of molecular therapies that are antiresorptive or bone anabolic (i.e., calcium, vitamin D, bisphosphonates, calcitonin, strontium, hormone replacement therapy, selective estrogen receptor modulators).
Background: Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are cha... more Background: Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are challenging to manage and often require surgical reconstructions with limited long-term success. Thus, characterizations of these tissues are urgently needed to better understand cellular mechanisms that regulate tissue homeostasis and healing. Explant culturing systems allow for ex vivo analysis of tissues in an environment that mimics the native microenvironment in vivo. Methods: Collaborative efforts within our institution facilitated the establishment of a novel explant culturing system. Tissue specimens cultured in single wells, with individual applied loading and/or biological environment, allowed characterization of tissue cultured under a variety of biological loading conditions. Quantitative PCR analysis for selected gene markers was our primary outcome. Results: Data were stratified for analysis by either culture environment or loading condition. Our gene expression results show that specimens clustered by culture condition may differ in molecular markers related to ECM production (e.g., Col1a1, Adamts4) and/or organization (e.g., Tnc, Dnc). In contrast, loading condition did significantly alter the median gene expression levels of tissues in comparison to unloaded control samples, although gene expression values related to ECM degradation (e.g., Mmp1, Mmp10) were altered in tendons cultured under tension in the device. Conclusion: Our study demonstrates promising utility of a novel explant culturing system for further characterization of musculoskeletal tissues such as native tendons and ligaments, as well as pathologic fibrotic tissues resulting from arthrofibrosis or Dupuytren's disease.
Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control o... more Leopard seals (Hydrurga leptonyx) are top predators that can exert substantial top-down control of their Antarctic prey species. However, population trends and genetic diversity of leopard seals remain understudied, limiting our understanding of their ecological role. We investigated the genetic diversity, effective population size and demographic history of leopard seals to provide fundamental data that contextualizes their predatory influence on Antarctic ecosystems. Ninety leopard seals were sampled in the northern Antarctic Peninsula, during the austral summers of 2008–2019 and a 405bp region of the mitochondrial control region was sequenced for each individual. We uncovered moderate levels of nucleotide (π = 0.013) and haplotype (Hd = 0.96) diversity, and the effective population size was estimated at around 24,000 individuals (NE= 24,376; range 16,876 – 33,126). Consistent with findings from other ice-breeding pinnipeds, Bayesian skyline analysis also revealed evidence for pop...
Metal orthopedic implants are largely biocompatible and generally achieve long-term structural fi... more Metal orthopedic implants are largely biocompatible and generally achieve long-term structural fixation. However, some orthopedic implants may loosen over time even in the absence of infection. In vivo fixation failure is multifactorial, but the fundamental biological defect is cellular dysfunction at the host-implant interface. Strategies to reduce the risk of short- and long-term loosening include surface modifications, implant metal alloy type, and adjuvant substances such as polymethylmethacrylate cement. Surface modifications (e.g., increased surface rugosity) can increase osseointegration and biological ingrowth of orthopedic implants. However, the localized responses of cells to implant surface modifications need to be better characterized. As an in vitro model for investigating cellular responses to metallic orthopedic implants, we cultured mesenchymal stromal/stem cells (MSCs) on clinical-grade titanium disks (Ti6Al4V) that differed in surface roughness as high (porous-structured), medium (grit-blasted), and low (bead-blasted). Topological characterization of clinically relevant Ti materials combined with differential mRNA expression analyses (RNA-seq and RT-qPCR) revealed alterations to the biological phenotype of cells cultured on titanium structures that favor early ECM production and observable responses to oxidative stress and heavy metal stress. These results provide a descriptive model for the interpretation of cellular responses at the interface between native host tissues and 3-D printed modular orthopedic implants, and will guide future studies aimed at increasing the long-term retention of such materials after TJA.
This is a PDF file of an article that has undergone enhancements after acceptance, such as the ad... more This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Purpose of Review MicroRNAs (miRNAs) are single-stranded, short (~22 nt) non-coding RNAs that con... more Purpose of Review MicroRNAs (miRNAs) are single-stranded, short (~22 nt) non-coding RNAs that control gene expression in most metazoan taxa. These vital post-transcriptional regulators are emerging as a novel class of relatively well-conserved biomarkers useful to molecular ecologists working on non-model marine organisms. The purpose of this review is to provide researchers with a brief background on miRNAs and to explore recent applications in marine biology. Recent Findings MiRNA datasets have been broadly employed in studies concerning commercially important species (oysters and crustaceans), phylogenetics (particularly deep evolutionary splits), and environmental stressor responses (temperature and salinity). Most progress has been made in the characterization of cnidarian miRNAs and bivalve and crustacean immune-related miRNAs. The use of miRNAs in phylogenetics is still under debate due to the secondary loss of miRNAs in some lineages, but they have been successfully applied in the resolution of deep evolutionary splits. Finally, miRNAs have been investigated in abiotic stress responses, but data interpretation is limited by the high number of species-specific miRNAs detected in these studies. Improvements in miRNA database curation and functional annotation should provide more confidence in their use. Summary Due to their evolutionary conservation, resilience to degradation, and amenable bioinformatics workflows, miRNAs are a powerful molecular tool in marine genomics. MiRNA investigations regarding environmental stress response will be particularly useful due to their potential to reveal physiological alterations and disease. Thus, they may be ultimately utilized as bio-indicators of environmental health.
Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pa... more Introduction: The inflammatory cascade and production of prostaglandins may play a role in the pathogenesis of arthrofibrosis, a debilitating condition after joint replacement and other orthopedic procedures. Cyclooxygenase 2 (COX-2) inhibitors may mitigate the inflammatory response and formation of arthrofibrosis, but oral delivery is associated with risk of systemic side effects in many patients. The nonsteroidal anti-inflammatory drug, celecoxib, may have therapeutic benefits for arthrofibrosis, but current methods for its local delivery (e.g., biologically derived microspheres) are not translatable to immediate clinical use. Therefore, we investigated the use of a drug scaffold for sustainable intra-articular delivery of therapeutic doses of celecoxib. Materials and Methods: Celecoxib was eluted from clinically approved biodegradable collagen membranes over 7 days as measured by UV spectroscopy and high-performance liquid chromatography/mass spectroscopy. Eluted concentrations of celecoxib were examined for toxicity (live/dead staining) and profibrotic gene expression (real-time-quantitative polymerase chain reaction) in rabbit knee capsular fibroblasts in vitro. Results: Sustained concentrations of celecoxib eluted from the membrane over 7 days from both a wet and dry scaffold, with a burst release (30-45%) of celecoxib in the first 2 h. Rabbit cells treated with eluted concentrations experienced a toxic response to the burst release doses, and inhibitory effects on profibrotic genes were seen in response to the sustained doses eluted from the scaffold. Conclusions: This study characterized the novel use of collagen scaffolds for intra-articular drug delivery to treat arthrofibrosis. Scaffolds exhibit celecoxib release through an initial burst release followed by sustained release of antifibrotic doses over 7 days. Thus, collagen scaffolds are promising for clinician-directed treatment of arthrofibrosis.
Journal of Plastic, Reconstructive & Aesthetic Surgery, 2019
Mesenchymal stromal cells (MSCs) secrete many soluble growth factors and have previously been sho... more Mesenchymal stromal cells (MSCs) secrete many soluble growth factors and have previously been shown to stimulate nerve regeneration. MSC-seeded processed nerve allografts could potentially be a promising method for large segmental motor nerve injuries. Further progress in our understanding of how the functions of MSCs can be leveraged for peripheral nerve repair is required before making clinical translation. The present study, therefore, investigated whether interactions of adipose-derived MSCs with decellularized nerve allografts can improve gene and protein expression of growth factors that may support nerve regeneration. Human nerve allografts (n = 30) were decellularized and seeded with undifferentiated human adipose-derived MSCs. Subsequently, the MSCs and MSC-seeded grafts were isolated on days 3, 7, 14, and 21 in culture for RNA expression analysis by qRT-PCR. Evaluated genes Parts of this article have been presented at the following meeting/conference: 1.
Journal of orthopaedic research : official publication of the Orthopaedic Research Society, Jan 14, 2018
Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. A... more Trauma, surgery, and other inflammatory conditions can lead to debilitating joint contractures. Adjunct pharmacologic modalities may permit clinical prevention and treatment of recalcitrant joint contractures. We investigated the therapeutic potential of rosiglitazone by intra-articular delivery via oligo[poly(ethylene glycol)fumarate] (OPF) hydrogels in an established rabbit model of arthrofibrosis. OPF hydrogels loaded with rosiglitazone were characterized for drug elution properties upon soaking in minimum essential media (MEM) with 10% fetal bovine serum and measurements of drug concentrations via High Performance Liquid Chromatography (HPLC). Drug-loaded scaffolds were surgically implanted into 24 skeletally mature female New Zealand White rabbits that were divided into equal groups receiving OPF hydrogels loaded with rosiglitazone (1.67 mg), or vehicle control (10 µL DMSO). After 8 weeks of joint immobilization, rabbits were allowed unrestricted cage activity for 16 weeks. Con...
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Papers by Eric Lewallen