Publications by authors named "Chia-Lung Wu"

21 Publications

  • Page 1 of 1

Single-cell RNA sequencing reveals the induction of novel myeloid and myeloid-associated cell populations in visceral fat with long-term obesity.

FASEB J 2021 03;35(3):e21417

Department of Orthopaedic Surgery, Washington University in St. Louis, St. Louis, MO, USA.

Macrophages and other immune cells are important contributors to obesity-associated inflammation; however, the cellular identities of these specific populations remain unknown. In this study, we identified individual populations of myeloid cells found in mouse epididymal/visceral adipose tissue by single-cell RNA sequencing, immunofluorescence, and flow cytometry. Multiple canonical correlation analysis identified 11 unique myeloid and myeloid-associate cell populations. In obese mice, we detected an increased percentage of monocyte-derived pro-inflammatory cells expressing Cd9 and Trem2, as well as significantly decreased percentages of multiple cell populations, including tissue-resident cells expressing Lyve1, Mafb, and Mrc1. We have identified and validated a novel myeloid/macrophage population defined by Ly6a expression, exhibiting both myeloid and mesenchymal characteristics, which increased with obesity and showed high pro-fibrotic characteristics in vitro. Our mouse adipose tissue myeloid cell atlas provides an important resource to investigate obesity-associated inflammation and fibrosis.
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http://dx.doi.org/10.1096/fj.202001970RDOI Listing
March 2021

Single Cell Omics for Musculoskeletal Research.

Curr Osteoporos Rep 2021 Apr 9;19(2):131-140. Epub 2021 Feb 9.

Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN, USA.

Purpose Of Review: The ability to analyze the molecular events occurring within individual cells as opposed to populations of cells is revolutionizing our understanding of musculoskeletal tissue development and disease. Single cell studies have the great potential of identifying cellular subpopulations that work in a synchronized fashion to regenerate and repair damaged tissues during normal homeostasis. In addition, such studies can elucidate how these processes break down in disease as well as identify cellular subpopulations that drive the disease. This review highlights three emerging technologies: single cell RNA sequencing (scRNA-seq), Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq), and Cytometry by Time-Of-Flight (CyTOF) mass cytometry.

Recent Findings: Technological and bioinformatic tools to analyze the transcriptome, epigenome, and proteome at the individual cell level have advanced rapidly making data collection relatively easy; however, understanding how to access and interpret the data remains a challenge for many scientists. It is, therefore, of paramount significance to educate the musculoskeletal community on how single cell technologies can be used to answer research questions and advance translation. This article summarizes talks given during a workshop on "Single Cell Omics" at the 2020 annual meeting of the Orthopedic Research Society. Studies that applied scRNA-seq, ATAC-seq, and CyTOF mass cytometry to cartilage development and osteoarthritis are reviewed. This body of work shows how these cutting-edge tools can advance our understanding of the cellular heterogeneity and trajectories of lineage specification during development and disease.
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http://dx.doi.org/10.1007/s11914-021-00662-2DOI Listing
April 2021

Single cell transcriptomic analysis of human pluripotent stem cell chondrogenesis.

Nat Commun 2021 01 13;12(1):362. Epub 2021 Jan 13.

Dept. of Orthopaedic Surgery, Washington University in Saint Louis, St. Louis, MO, 63110, USA.

The therapeutic application of human induced pluripotent stem cells (hiPSCs) for cartilage regeneration is largely hindered by the low yield of chondrocytes accompanied by unpredictable and heterogeneous off-target differentiation of cells during chondrogenesis. Here, we combine bulk RNA sequencing, single cell RNA sequencing, and bioinformatic analyses, including weighted gene co-expression analysis (WGCNA), to investigate the gene regulatory networks regulating hiPSC differentiation under chondrogenic conditions. We identify specific WNTs and MITF as hub genes governing the generation of off-target differentiation into neural cells and melanocytes during hiPSC chondrogenesis. With heterocellular signaling models, we further show that WNT signaling produced by off-target cells is responsible for inducing chondrocyte hypertrophy. By targeting WNTs and MITF, we eliminate these cell lineages, significantly enhancing the yield and homogeneity of hiPSC-derived chondrocytes. Collectively, our findings identify the trajectories and molecular mechanisms governing cell fate decision in hiPSC chondrogenesis, as well as dynamic transcriptome profiles orchestrating chondrocyte proliferation and differentiation.
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http://dx.doi.org/10.1038/s41467-020-20598-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806634PMC
January 2021

Is Obesity a Disease of Stem Cells?

Cell Stem Cell 2020 07;27(1):15-18

Department of Orthopaedic Surgery, Washington University in St. Louis, St. Louis, MO, USA; Shriners Hospitals for Children, St. Louis, MO, USA; Center of Regenerative Medicine, Washington University in St. Louis, St. Louis, MO, USA. Electronic address:

Obesity disrupts physiological homeostasis and alters both systemic and local microenvironments that impact stem cell plasticity and impair regenerative capacity. We present growing evidence that reveals the bidirectionality of obesity-induced stem cell dysfunction and how the molecular changes in stem cells residing in obese environments may accelerate disease severity.
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http://dx.doi.org/10.1016/j.stem.2020.04.019DOI Listing
July 2020

Gene therapy for follistatin mitigates systemic metabolic inflammation and post-traumatic arthritis in high-fat diet-induced obesity.

Sci Adv 2020 May 8;6(19):eaaz7492. Epub 2020 May 8.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA.

Obesity-associated inflammation and loss of muscle function play critical roles in the development of osteoarthritis (OA); thus, therapies that target muscle tissue may provide novel approaches to restoring metabolic and biomechanical dysfunction associated with obesity. Follistatin (FST), a protein that binds myostatin and activin, may have the potential to enhance muscle formation while inhibiting inflammation. Here, we hypothesized that adeno-associated virus 9 (AAV9) delivery of FST enhances muscle formation and mitigates metabolic inflammation and knee OA caused by a high-fat diet in mice. AAV-mediated FST delivery exhibited decreased obesity-induced inflammatory adipokines and cytokines systemically and in the joint synovial fluid. Regardless of diet, mice receiving FST gene therapy were protected from post-traumatic OA and bone remodeling induced by joint injury. Together, these findings suggest that FST gene therapy may provide a multifactorial therapeutic approach for injury-induced OA and metabolic inflammation in obesity.
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http://dx.doi.org/10.1126/sciadv.aaz7492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7209997PMC
May 2020

Transgenic conversion of ω-6 to ω-3 polyunsaturated fatty acids via fat-1 reduces the severity of post-traumatic osteoarthritis.

Arthritis Res Ther 2020 04 15;22(1):83. Epub 2020 Apr 15.

Department of Orthopaedic Surgery, Washington University in St. Louis, Campus Box 8233, Couch Biomedical Research Bldg, Room 3121, St. Louis, MO, 63110, USA.

Background: Dietary fatty acid (FA) content has been shown to influence the development of post-traumatic osteoarthritis (PTOA) in obesity. We used the fat-1 transgenic mouse to examine the hypothesis that endogenous reduction of ω-6 to ω-3 FA ratio, under the same dietary conditions, would mitigate metabolic inflammation and the pathogenesis of PTOA in obese male and female mice.

Methods: Male and female fat-1 and wild-type littermates were fed either a control diet or an ω-6 FA-rich high-fat diet and underwent destabilization of the medial meniscus (DMM) surgery to induce PTOA. OA severity, synovitis, and osteophyte formation were determined histologically, while biomarker and lipidomic analyses were performed to evaluate levels of adipokines, insulin, pro-/anti-inflammatory cytokines, and FAs in serum and joint synovial fluid. Multivariable models were performed to elucidate the associations of dietary, metabolic, and mechanical factors with PTOA.

Results: We found that elevated serum levels of ω-3 FAs in fat-1 mice as compared to wild-type controls fed the same diet resulted in reduced OA and synovitis in a sex- and diet-dependent manner, despite comparable body weights. The fat-1 mice showed trends toward decreased serum pro-inflammatory cytokines and increased anti-inflammatory cytokines. Multivariable analysis for variables predicting OA severity in mice resulted in correlations with serum FA levels, but not with body weight.

Conclusions: This study provides further evidence that circulating FA composition and systemic metabolic inflammation, rather than body weight, may be the major risk factor for obesity-associated OA. We also demonstrate the potential genetic use of ω-3 FA desaturase in mitigating PTOA in obese patients following injury.
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http://dx.doi.org/10.1186/s13075-020-02170-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7160898PMC
April 2020

Prospective isolation of chondroprogenitors from human iPSCs based on cell surface markers identified using a CRISPR-Cas9-generated reporter.

Stem Cell Res Ther 2020 02 18;11(1):66. Epub 2020 Feb 18.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, 63110, USA.

Background: Articular cartilage shows little or no capacity for intrinsic repair, generating a critical need of regenerative therapies for joint injuries and diseases such as osteoarthritis. Human-induced pluripotent stem cells (hiPSCs) offer a promising cell source for cartilage tissue engineering and in vitro human disease modeling; however, off-target differentiation remains a challenge during hiPSC chondrogenesis. Therefore, the objective of this study was to identify cell surface markers that define the true chondroprogenitor population and use these markers to purify iPSCs as a means of improving the homogeneity and efficiency of hiPSC chondrogenic differentiation.

Methods: We used a CRISPR-Cas9-edited COL2A1-GFP knock-in reporter hiPSC line, coupled with a surface marker screen, to identify a novel chondroprogenitor population. Single-cell RNA sequencing was then used to analyze the distinct clusters within the population. An unpaired t test with Welch's correction or an unpaired Kolmogorov-Smirnov test was performed with significance reported at a 95% confidence interval.

Results: Chondroprogenitors expressing CD146, CD166, and PDGFRβ, but not CD45, made up an average of 16.8% of the total population. Under chondrogenic culture conditions, these triple-positive chondroprogenitor cells demonstrated decreased heterogeneity as measured by single-cell RNA sequencing with fewer clusters (9 clusters in unsorted vs. 6 in sorted populations) closer together. Additionally, there was more robust and homogenous matrix production (unsorted: 1.5 ng/ng vs. sorted: 19.9 ng/ng sGAG/DNA; p < 0.001) with significantly higher chondrogenic gene expression (i.e., SOX9, COL2A1, ACAN; p < 0.05).

Conclusions: Overall, this study has identified a unique hiPSC-derived subpopulation of chondroprogenitors that are CD146/CD166/PDGFRβ/CD45 and exhibit high chondrogenic potential, providing a purified cell source for cartilage tissue engineering or disease modeling studies.
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http://dx.doi.org/10.1186/s13287-020-01597-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7026983PMC
February 2020

Intergenerational Transmission of Diet-Induced Obesity, Metabolic Imbalance, and Osteoarthritis in Mice.

Arthritis Rheumatol 2020 04 5;72(4):632-644. Epub 2020 Mar 5.

Washington University in St. Louis and Shriners Hospitals for Children, St. Louis, Missouri.

Objective: Obesity and osteoarthritis (OA) are 2 major public health issues affecting millions of people worldwide. Whereas parental obesity affects the predisposition to diseases such as cancer or diabetes in children, transgenerational influences on musculoskeletal conditions such as OA are poorly understood. This study was undertaken to assess the intergenerational effects of a parental/grandparental high-fat diet on the metabolic and skeletal phenotype, systemic inflammation, and predisposition to OA in 2 generations of offspring in mice.

Methods: Metabolic phenotype and predisposition to OA were investigated in the first and second (F1 and F2) generations of offspring (n = 10-16 mice per sex per diet) bred from mice fed a high-fat diet (HFD) or a low-fat control diet. OA was induced by destabilizing the medial meniscus. OA, synovitis, and adipose tissue inflammation were determined histologically, while bone changes were measured using micro-computed tomography. Serum and synovial cytokines were measured by multiplex assay.

Results: Parental high-fat feeding showed an intergenerational effect, with inheritance of increased weight gain (up to 19% in the F1 generation and 9% in F2), metabolic imbalance, and injury-induced OA in at least 2 generations of mice, despite the fact that the offspring were fed the low-fat diet. Strikingly, both F1 and F2 female mice showed an increased predisposition to injury-induced OA (48% higher predisposition in F1 and 19% in F2 female mice fed the HFD) and developed bone microarchitectural changes that were attributable to parental and grandparental high-fat feeding.

Conclusion: The results of this study reveal a detrimental effect of parental HFD and obesity on the musculoskeletal integrity of 2 generations of offspring, indicating the importance of further investigation of these effects. An improved understanding of the mechanisms involved in the transmissibility of diet-induced changes through multiple generations may help in the development of future therapies that would target the effects of obesity on OA and related conditions.
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http://dx.doi.org/10.1002/art.41147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7113102PMC
April 2020

Physiologic and pathologic effects of dietary free fatty acids on cells of the joint.

Ann N Y Acad Sci 2019 03 15;1440(1):36-53. Epub 2019 Jan 15.

Department of Orthopaedic Surgery, Washington University, St. Louis, Missouri.

Fatty acids (FAs) are potent organic compounds that not only can be used as an energy source during nutrient deprivation but are also involved in several essential signaling cascades in cells. Therefore, a balanced intake of different dietary FAs is critical for the maintenance of cellular functions and tissue homeostasis. A diet with an imbalanced fat composition creates a risk for developing metabolic syndrome and various musculoskeletal diseases, including osteoarthritis (OA). In this review, we summarize the current state of knowledge and mechanistic insights regarding the role of dietary FAs, such as saturated FAs, omega-6 polyunsaturated FAs (PUFAs), and omega-3 PUFAs on joint inflammation and OA pathogeneses. In particular, we review how different types of dietary FAs and their derivatives distinctly affect a variety of cells within the joint, including chondrocytes, osteoblasts, osteoclasts, and synoviocytes. Understanding the molecular mechanisms underlying the effects of FAs on metabolic behavior, anabolic, and catabolic processes, as well as the inflammatory response of joint cells, may help identify therapeutic targets for the prevention of metabolic joint diseases.
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http://dx.doi.org/10.1111/nyas.13999DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6420352PMC
March 2019

Effects of dietary fatty acid content on humeral cartilage and bone structure in a mouse model of diet-induced obesity.

J Orthop Res 2019 03 12;37(3):779-788. Epub 2019 Feb 12.

Department of Orthopaedic Surgery, Washington University, Couch Biomedical Research Bldg., Room 3121, Campus Box 8233, Saint Louis, Missouri 63110.

Obesity is a primary risk factor for osteoarthritis (OA), and previous studies have shown that dietary content may play an important role in the pathogenesis of cartilage and bone in knee OA. Several previous studies have shown that the ratio of ω-3 polyunsaturated fatty acids (PUFAs), ω-6 PUFAs, and saturated fatty acids can significantly influence bone structure and OA progression. However, the influence of obesity or dietary fatty acid content on shoulder OA is not well understood. The goal of this study was to investigate the role of dietary fatty acid content on bone and cartilage structure in the mouse shoulder in a model of diet-induced obesity. For 24 weeks, mice were fed control or high-fat diets supplemented with ω-3 PUFAs, ω-6 PUFAs, or saturated fatty acids. The humeral heads were analyzed for bone morphometry and mineral density by microCT. Cartilage structure and joint synovitis were determined by histological grading, and microscale mechanical properties of the cartilage extracellular and pericellular matrices were quantified using atomic force microscopy. Diet-induced obesity significantly altered bone morphology and mineral density in a manner that was dependent on dietary free fatty acid content. In general, high-fat diet groups showed decreased bone quality, with the ω-3 diet being partially protective. Cartilage mechanical properties and OA scores showed no changes with obesity or diet. These findings are consistent with clinical literature showing little if any relationship between obesity and shoulder OA (unlike knee OA), but suggest that diet-induced obesity may influence other joint tissues. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.
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http://dx.doi.org/10.1002/jor.24219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6662729PMC
March 2019

Step-Wise Chondrogenesis of Human Induced Pluripotent Stem Cells and Purification Via a Reporter Allele Generated by CRISPR-Cas9 Genome Editing.

Stem Cells 2019 01 31;37(1):65-76. Epub 2018 Oct 31.

Department of Orthopaedic Surgery, Washington University, St. Louis, Missouri, USA.

The differentiation of human induced pluripotent stem cells (hiPSCs) to prescribed cell fates enables the engineering of patient-specific tissue types, such as hyaline cartilage, for applications in regenerative medicine, disease modeling, and drug screening. In many cases, however, these differentiation approaches are poorly controlled and generate heterogeneous cell populations. Here, we demonstrate cartilaginous matrix production in three unique hiPSC lines using a robust and reproducible differentiation protocol. To purify chondroprogenitors (CPs) produced by this protocol, we engineered a COL2A1-GFP knock-in reporter hiPSC line by CRISPR-Cas9 genome editing. Purified CPs demonstrated an improved chondrogenic capacity compared with unselected populations. The ability to enrich for CPs and generate homogenous matrix without contaminating cell types will be essential for regenerative and disease modeling applications. Stem Cells 2019;37:65-76.
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http://dx.doi.org/10.1002/stem.2931DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6312762PMC
January 2019

Osteoarthritis as a disease of the cartilage pericellular matrix.

Matrix Biol 2018 10 22;71-72:40-50. Epub 2018 May 22.

Department of Molecular Epidemiology, Leiden University Medical Center, Leiden, the Netherlands.

Osteoarthritis is a painful joint disease characterized by progressive degeneration of the articular cartilage as well as associated changes to the subchondral bone, synovium, and surrounding joint tissues. While the effects of osteoarthritis on the cartilage extracellular matrix (ECM) have been well recognized, it is now becoming apparent that in many cases, the onset of the disease may be initially reflected in the matrix region immediately surrounding the chondrocytes, termed the pericellular matrix (PCM). Growing evidence suggests that the PCM - which along with the enclosed chondrocytes are termed the "chondron" - acts as a critical transducer or "filter" of biochemical and biomechanical signals for the chondrocyte, serving to help regulate the homeostatic balance of chondrocyte metabolic activity in response to environmental signals. Indeed, it appears that alterations in PCM properties and cell-matrix interactions, secondary to genetic, epigenetic, metabolic, or biomechanical stimuli, could in fact serve as initiating or progressive factors for osteoarthritis. Here, we discuss recent advances in the understanding of the role of the PCM, with an emphasis on the reciprocity of changes that occur in this matrix region with disease, as well as how alterations in PCM properties could serve as a driver of ECM-based diseases such as osteoarthritis. Further study of the structure, function, and composition of the PCM in normal and diseased conditions may provide new insights into the understanding of the pathogenesis of osteoarthritis, and presumably new therapeutic approaches for this disease.
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http://dx.doi.org/10.1016/j.matbio.2018.05.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6146061PMC
October 2018

Differentiation of human induced pluripotent stem cells into nucleus pulposus-like cells.

Stem Cell Res Ther 2018 03 9;9(1):61. Epub 2018 Mar 9.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA.

Background: Intervertebral disc (IVD) degeneration is characterized by an early decrease in cellularity of the nucleus pulposus (NP) region, and associated extracellular matrix changes, reduced hydration, and progressive degeneration. Cell-based IVD therapy has emerged as an area of great interest, with studies reporting regenerative potential for many cell sources, including autologous or allogeneic chondrocytes, primary IVD cells, and stem cells. Few approaches, however, have clear strategies to promote the NP phenotype, in part due to a limited knowledge of the defined markers and differentiation protocols for this lineage. Here, we developed a new protocol for the efficient differentiation of human induced pluripotent stem cells (hiPSCs) into NP-like cells in vitro. This differentiation strategy derives from our knowledge of the embryonic notochordal lineage of NP cells as well as strategies used to support healthy NP cell phenotypes for primary cells in vitro.

Methods: An NP-genic phenotype of hiPSCs was promoted in undifferentiated hiPSCs using a stepwise, directed differentiation toward mesodermal, and subsequently notochordal, lineages via chemically defined medium and growth factor supplementation. Fluorescent cell imaging was used to test for pluripotency markers in undifferentiated cells. RT-PCR was used to test for potential cell lineages at the early stage of differentiation. Cells were checked for NP differentiation using immunohistochemistry and histological staining at the end of differentiation. To enrich notochordal progenitor cells, hiPSCs were transduced using lentivirus containing reporter constructs for transcription factor brachyury (T) promoter and green fluorescent protein (GFP) fluorescence, and then sorted on T expression based on GFP intensity by flow cytometry.

Results: Periods of pellet culture following initial induction were shown to promote the vacuolated NP cell morphology and NP surface marker expression, including CD24, LMα5, and Basp1. Enrichment of brachyury (T) positive cells using fluorescence-activated cell sorting was shown to further enhance the differentiation efficiency of NP-like cells.

Conclusions: The ability to efficiently differentiate human iPSCs toward NP-like cells may provide insights into the processes of NP cell differentiation and provide a cell source for the development of new therapies for IVD diseases.
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http://dx.doi.org/10.1186/s13287-018-0797-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5845143PMC
March 2018

Genome Engineering for Personalized Arthritis Therapeutics.

Trends Mol Med 2017 10 5;23(10):917-931. Epub 2017 Sep 5.

Department of Orthopedic Surgery, Washington University, St. Louis, MO 63110, USA; Shriners Hospitals for Children - St. Louis, St. Louis, MO 63110, USA; Department of Cell Biology, Duke University Medical Center, Durham, NC 27710, USA; Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA; Cytex Therapeutics, Inc., Durham, NC 27705, USA. Electronic address:

Arthritis represents a family of complex joint pathologies responsible for the majority of musculoskeletal conditions. Nearly all diseases within this family, including osteoarthritis, rheumatoid arthritis, and juvenile idiopathic arthritis, are chronic conditions with few or no disease-modifying therapeutics available. Advances in genome engineering technology, most recently with CRISPR-Cas9, have revolutionized our ability to interrogate and validate genetic and epigenetic elements associated with chronic diseases such as arthritis. These technologies, together with cell reprogramming methods, including the use of induced pluripotent stem cells, provide a platform for human disease modeling. We summarize new evidence from genome-wide association studies and genomics that substantiates a genetic basis for arthritis pathogenesis. We also review the potential contributions of genome engineering in the development of new arthritis therapeutics.
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http://dx.doi.org/10.1016/j.molmed.2017.08.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5657581PMC
October 2017

Conditional Macrophage Depletion Increases Inflammation and Does Not Inhibit the Development of Osteoarthritis in Obese Macrophage Fas-Induced Apoptosis-Transgenic Mice.

Arthritis Rheumatol 2017 09 8;69(9):1772-1783. Epub 2017 Aug 8.

Washington University and Shriners Hospitals for Children, St. Louis, Missouri.

Objective: To investigate whether short-term, systemic depletion of macrophages can mitigate osteoarthritis (OA) following injury in the setting of obesity.

Methods: CSF-1R-GFP+ macrophage Fas-induced apoptosis (MaFIA)-transgenic mice that allow conditional depletion of macrophages were placed on a high-fat diet and underwent surgery to induce knee OA. A small molecule (AP20187) was administrated to deplete macrophages in MaFIA mice. The effects of macrophage depletion on acute joint inflammation, OA severity, and arthritic bone changes were evaluated using histology and micro-computed tomography. Immunohistochemical analysis was performed to identify various immune cells. The levels of serum and synovial fluid cytokines were also measured.

Results: Macrophage-depleted mice had significantly fewer M1 and M2 macrophages in the surgically operated joints relative to controls and exhibited decreased osteophyte formation immediately following depletion. Surprisingly, macrophage depletion did not attenuate the severity of OA in obese mice; instead, it induced systemic inflammation and led to a massive infiltration of CD3+ T cells and particularly neutrophils, but not B cells, into the injured joints. Macrophage-depleted mice also demonstrated a markedly increased number of proinflammatory cytokines including granulocyte colony-stimulating factor, interleukin-1β (IL-1β), IL-6, IL-8, and tumor necrosis factor in both serum and joint synovial fluid, although the mice showed a trend toward decreased levels of insulin and leptin in serum after macrophage depletion.

Conclusion: Our findings indicate that macrophages are vital for modulating homeostasis of immune cells in the setting of obesity and suggest that more targeted approaches of depleting specific macrophage subtypes may be necessary to mitigate inflammation and OA in the setting of obesity.
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http://dx.doi.org/10.1002/art.40161DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5611814PMC
September 2017

Serum and synovial fluid lipidomic profiles predict obesity-associated osteoarthritis, synovitis, and wound repair.

Sci Rep 2017 03 20;7:44315. Epub 2017 Mar 20.

Department of Orthopaedic Surgery, Washington University, St. Louis MO, 63110, USA.

High-fat diet-induced obesity is a major risk factor for osteoarthritis (OA) and diminished wound healing. The objective of this study was to determine the associations among serum and synovial fluid lipid levels with OA, synovitis, adipokine levels, and wound healing in a pre-clinical obese mouse model of OA. Male C57BL/6 J mice were fed either a low-fat (10% kcal) or one of three high-fat (HF, 60% kcal) diets rich in saturated fatty acids (SFAs), ω-6 or ω-3 polyunsaturated FAs (PUFAs). OA was induced by destabilization of the medial meniscus. Mice also received an ear punch for evaluating wound healing. Serum and synovial fluid were collected for lipidomic and adipokine analyses. We demonstrated that the serum levels of ω-3 PUFAs were negatively correlated with OA and wound size, but positively correlated with adiponectin levels. In contrast, most ω-6 PUFAs exhibited positive correlations with OA, impaired healing, and inflammatory adipokines. Interestingly, levels of pentadecylic acid (C15:0, an odd-chain SFA) and palmitoleic acid were inversely correlated with joint degradation. This study extends our understanding of the links of FAs with OA, synovitis and wound healing, and reports newly identified serum and synovial fluid FAs as predictive biomarkers of OA in obesity.
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http://dx.doi.org/10.1038/srep44315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5357837PMC
March 2017

Cartilage-Specific Knockout of the Mechanosensory Ion Channel TRPV4 Decreases Age-Related Osteoarthritis.

Sci Rep 2016 07 8;6:29053. Epub 2016 Jul 8.

Department of Orthopaedic Surgery, Washington University in St. Louis, Missouri, 63110, USA.

Osteoarthritis (OA) is a progressive degenerative disease of articular cartilage and surrounding tissues, and is associated with both advanced age and joint injury. Biomechanical factors play a critical role in the onset and progression of OA, yet the mechanisms through which physiologic or pathologic mechanical signals are transduced into a cellular response are not well understood. Defining the role of mechanosensory pathways in cartilage during OA pathogenesis may yield novel strategies or targets for the treatment of OA. The transient receptor potential vanilloid 4 (TRPV4) ion channel transduces mechanical loading of articular cartilage via the generation of intracellular calcium ion transients. Using tissue-specific, inducible Trpv4 gene-targeted mice, we demonstrate that loss of TRPV4-mediated cartilage mechanotransduction in adulthood reduces the severity of aging-associated OA. However, loss of chondrocyte TRPV4 did not prevent OA development following destabilization of the medial meniscus (DMM). These results highlight potentially distinct roles of TRPV4-mediated cartilage mechanotransduction in age-related and post-traumatic OA, and point to a novel disease-modifying strategy to therapeutically target the TRPV4-mediated mechanotransduction pathway for the treatment of aging-associated OA.
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http://dx.doi.org/10.1038/srep29053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4937413PMC
July 2016

Intravenous glutamine appears to reduce the severity of symptomatic platinum-induced neuropathy: a prospective randomized study.

J Chemother 2015 Aug 23;27(4):235-40. Epub 2015 Mar 23.

The purpose of this study is to evaluate the efficacy of intravenous (IV) glutamine or calcium/magnesium (Ca/Mg) infusion against platinum-induced neuropathy. Patients undergoing platinum-based (oxaliplatin or cisplatin) therapy were randomized to receive IV glutamine or Ca/Mg infusion during four cycles of chemotherapy, from the fifth cycle of therapy to the eighth cycle. The total neuropathy score (TNS) was evaluated at the end of the fourth course of chemotherapy (as baseline) and at the end of the eighth cycle (as end-of treatment). The intent-to-treat analysis of the end point included 29 patients in the glutamine arm and 26 patients in the Ca/Mg arm. The mean TNS of both cohorts increased significantly. The baseline and end-of-treatment TNSs between the two groups were not statistically different. Patients with symptoms at baseline (N = 29) had significantly lower scores at the end of the study in the glutamine group (P = 0.045). Besides, glutamine group patients who initially had only sensory symptoms (N = 23) also had significantly lower scores at the study's end (P = 0.035). Neither IV glutamine nor Ca/Mg infusion prevented further worsening of platinum-induced neuropathy. However, IV glutamine apparently reduced the severity of symptomatic platinum-induced neuropathy.
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http://dx.doi.org/10.1179/1973947815Y.0000000011DOI Listing
August 2015

Dietary fatty acid content regulates wound repair and the pathogenesis of osteoarthritis following joint injury.

Ann Rheum Dis 2015 Nov 10;74(11):2076-83. Epub 2014 Jul 10.

Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina, USA Department of Biomedical Engineering, Duke University Medical Center, Durham, North Carolina, USA Department of Cell Biology, Duke University Medical Center, Durham, North Carolina, USA.

Objective: The mechanisms linking obesity and osteoarthritis (OA) are not fully understood and have been generally attributed to increased weight, rather than metabolic or inflammatory factors. Here, we examined the influence of fatty acids, adipokines, and body weight on OA following joint injury in an obese mouse model.

Methods: Mice were fed high-fat diets rich in various fatty acids (FA) including saturated FAs (SFAs), ω-6 polyunsaturated FAs (PUFAs), and ω-3 PUFAs. OA was induced by destabilising the medial meniscus. Wound healing was evaluated using an ear punch. OA, synovitis and wound healing were determined histologically, while bone changes were measured using microCT. Activity levels and serum cytokines were measured at various time-points. Multivariate models were performed to elucidate the associations of dietary, metabolic and mechanical factors with OA and wound healing.

Results: Using weight-matched mice and multivariate models, we found that OA was significantly associated with dietary fatty acid content and serum adipokine levels, but not with body weight. Furthermore, spontaneous activity of the mice was independent of OA development. Small amounts of ω-3 PUFAs (8% by kcal) in a high-fat diet were sufficient to mitigate injury-induced OA, decreasing leptin and resistin levels. ω-3 PUFAs significantly enhanced wound repair, SFAs or ω-6 PUFAs independently increased OA severity, heterotopic ossification and scar tissue formation.

Conclusions: Our results indicate that with obesity, dietary FA content regulates wound healing and OA severity following joint injury, independent of body weight, supporting the need for further studies of dietary FA supplements as a potential therapeutic approach for OA.
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http://dx.doi.org/10.1136/annrheumdis-2014-205601DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4363043PMC
November 2015

Life-long caloric restriction does not alter the severity of age-related osteoarthritis.

Age (Dordr) 2014 1;36(4):9669. Epub 2014 Jul 1.

Department of Orthopaedic Surgery, Duke University Medical Center, 375 Medical Sciences Research Bldg, 3093, Durham, NC, 27710, USA.

Chronic adipose tissue inflammation and its associated adipokines have been linked to the development of osteoarthritis (OA). It has been shown that caloric restriction may decrease body mass index and adiposity. The objectives of this study were to investigate the effect of lifelong caloric restriction on bone morphology, joint inflammation, and spontaneously occurring OA development in aged mice. C57BL/NIA mice were fed either a calorie-restricted (CR) or ad libitum (AL) diet starting at 14 weeks of age. All mice were sacrificed at 24 months of age. Adipose tissue and knee joints were then harvested. Bone parameters of the joints were analyzed by micro-CT. OA and joint synovitis were determined using histology and semiquantitative analysis. Lifelong caloric restriction did not alter the severity of OA development in C57BL/NIA aged mice, and there was no difference in the total joint Mankin score between CR and AL groups (p = 0.99). Mice also exhibited similar levels of synovitis (p = 0.54). The bone mineral density of the femur and the tibia was comparable between the groups with a small increase in cancellous bone volume fraction in the lateral femoral condyle of the CR group compared with the AL group. Lifelong caloric restriction did not alter the incidence of OA or joint synovitis in C57BL/NIA mice, indicating that a reduction of caloric intake alone was not sufficient to prevent spontaneous age-related OA. Nonetheless, early initiation of CR continued throughout a life span did not negatively impact bone structural properties.
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http://dx.doi.org/10.1007/s11357-014-9669-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4150885PMC
March 2015

Intra-articular delivery of purified mesenchymal stem cells from C57BL/6 or MRL/MpJ superhealer mice prevents posttraumatic arthritis.

Cell Transplant 2013 10;22(8):1395-408. Epub 2012 Aug 10.

Department of Orthopaedic Surgery, Duke University Medical Center, Durham, NC 27710, USA.

Joint injury dramatically enhances the onset of osteoarthritis (OA) and is responsible for an estimated 12% of OA. Posttraumatic arthritis (PTA) is especially common after intra-articular fracture, and no disease-modifying therapies are currently available. We hypothesized that the delivery of mesenchymal stem cells (MSCs) would prevent PTA by altering the balance of inflammation and regeneration after fracture of the mouse knee. Additionally, we examined the hypothesis that MSCs from the MRL/MpJ (MRL) "superhealer" mouse strain would show increased multilineage and therapeutic potentials as compared to those from C57BL/6 (B6) mice, as MRL mice have shown exceptional in vivo regenerative abilities. A highly purified population of MSCs was prospectively isolated from bone marrow using cell surface markers (CD45-/TER119-/PDGFRα+/Sca-1+). B6 MSCs expanded greater than 100,000-fold in 3 weeks when cultured at 2% oxygen and displayed greater adipogenic, osteogenic, and chondrogenic differentiation as compared to MRL MSCs. Mice receiving only a control saline injection after fracture demonstrated PTA after 8 weeks, but the delivery of 10,000 B6 or MRL MSCs to the joint prevented the development of PTA. Cytokine levels in serum and synovial fluid were affected by treatment with stem cells, including elevated systemic interleukin-10 at several time points. The delivery of MSCs did not reduce the degree of synovial inflammation but did show increased bone volume during repair. This study provides evidence that intra-articular stem cell therapy can prevent the development of PTA after fracture and has implications for possible clinical interventions after joint injury before evidence of significant OA.
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http://dx.doi.org/10.3727/096368912X653264DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3891895PMC
April 2014