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OBJECTIVE: Osteoarthritis (OA) is the most prevalent musculoskeletal disease affecting articulating joint tissues, resulting in local and systemic changes that contribute to increased pain and reduced function. Diverse technological advancements have culminated in the advent of high throughput "omic" technologies, enabling identification of comprehensive changes in molecular mediators associated with the disease. Amongst these technologies, genomics and epigenomics - including methylomics and miRNomics, have emerged as important tools to aid our biological understanding of disease. DESIGN: In this narrative review, we selected articles discussing advancements and applications of these technologies to OA biology and pathology. We discuss how genomics, deoxyribonucleic acid (DNA) methylomics, and miRNomics have uncovered disease-related molecular markers in the local and systemic tissues or fluids of OA patients. RESULTS: Genomics investigations into the genetic links of OA, including using genome-wide association studies, have evolved to identify 100+ genetic susceptibility markers of OA. Epigenomic investigations of gene methylation status have identified the importance of methylation to OA-related catabolic gene expression. Furthermore, miRNomic studies have identified key microRNA signatures in various tissues and fluids related to OA disease. CONCLUSIONS: Sharing of standardized, well-annotated omic datasets in curated repositories will be key to enhancing statistical power to detect smaller and targetable changes in the biological signatures underlying OA pathogenesis. Additionally, continued technological developments and analysis methods, including using computational molecular and regulatory networks, are likely to facilitate improved detection of disease-relevant targets, in-turn, supporting precision medicine approaches and new treatment strategies for OA.
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Metilación de ADN , Epigenómica , Genómica , Osteoartritis , Humanos , Osteoartritis/genética , Estudio de Asociación del Genoma Completo , MicroARNs/genética , Predisposición Genética a la EnfermedadRESUMEN
This study analyzed knee synovial fluid after anterior cruciate ligament (ACL) tear and in osteoarthritis (OA) to test the hypotheses that concentrations of cytokines, chemokines, and growth factors differ (a) by diagnosis and (b) after ACL tear by time from injury and presence/absence of concomitant meniscus tear. Synovial fluid samples were collected from two groups, ACL tears (with or without meniscus tear) (N = 13) and Kellgren-Lawrence grade 3 and 4 OA (N = 16), undergoing clinically indicated aspiration of the knee joint. Multiple cytokines, chemokines, and growth factors were assessed using a multiplexed 45-protein panel. Comparisons were made for the concentrations of all molecules between ACL tear and OA patients, isolated versus combined ACL and meniscus tears, and categorized by time from injury: acute or early subacute (<15 days, N = 8) versus late subacute or chronic (>15 days and <3 months, N = 5). ACL tear patients have higher levels of six molecules (IL-4, IL-5, IL-13, PlGF-1, bNGF, TNF-α) in knee synovial fluid compared to OA patients. Isolated ACL tears express higher levels of IL-4, IL-13 and IFN-γ and lower levels of IL-7 than ACL tears with a concomitant meniscus tear. SDF-1α, PlGF-1, IL-1RA, HGF, bNGF, and BDNF levels are elevated immediately after injury and drop off significantly in the late subacute phase (after 15 days). Synovial fluid from knees with ACL tears have elevated metabolic activity compared to knees with OA. The cytokine profiles after ACL tears are influenced by the time from injury and the presence of meniscus tears. These findings offer valuable insights into the levels of cytokines, chemokines, and growth factors in the knee after ACL injury, information which may have important implications for the diagnosis, prognosis and treatment of this common pathology.
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Lesiones del Ligamento Cruzado Anterior , Citocinas , Péptidos y Proteínas de Señalización Intercelular , Osteoartritis de la Rodilla , Líquido Sinovial , Humanos , Líquido Sinovial/metabolismo , Líquido Sinovial/química , Lesiones del Ligamento Cruzado Anterior/metabolismo , Lesiones del Ligamento Cruzado Anterior/complicaciones , Femenino , Masculino , Persona de Mediana Edad , Adulto , Citocinas/metabolismo , Citocinas/análisis , Osteoartritis de la Rodilla/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Péptidos y Proteínas de Señalización Intercelular/análisis , Adulto Joven , Quimiocinas/metabolismo , Quimiocinas/análisis , Anciano , Lesiones de Menisco Tibial/metabolismoRESUMEN
OBJECTIVE: Osteoarthritis (OA) is a complex disease involving contributions from both local joint tissues and systemic sources. Patient characteristics, encompassing sociodemographic and clinical variables, are intricately linked with OA rendering its understanding challenging. Technological advancements have allowed for a comprehensive analysis of transcripts, proteomes and metabolomes in OA tissues/fluids through omic analyses. The objective of this review is to highlight the advancements achieved by omic studies in enhancing our understanding of OA pathogenesis over the last three decades. DESIGN: We conducted an extensive literature search focusing on transcriptomics, proteomics and metabolomics within the context of OA. Specifically, we explore how these technologies have identified individual transcripts, proteins, and metabolites, as well as distinctive endotype signatures from various body tissues or fluids of OA patients, including insights at the single-cell level, to advance our understanding of this highly complex disease. RESULTS: Omic studies reveal the description of numerous individual molecules and molecular patterns within OA-associated tissues and fluids. This includes the identification of specific cell (sub)types and associated pathways that contribute to disease mechanisms. However, there remains a necessity to further advance these technologies to delineate the spatial organization of cellular subtypes and molecular patterns within OA-afflicted tissues. CONCLUSIONS: Leveraging a multi-omics approach that integrates datasets from diverse molecular detection technologies, combined with patients' clinical and sociodemographic features, and molecular and regulatory networks, holds promise for identifying unique patient endophenotypes. This holistic approach can illuminate the heterogeneity among OA patients and, in turn, facilitate the development of tailored therapeutic interventions.
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Osteoartritis , Proteómica , Humanos , Metabolómica , Perfilación de la Expresión Génica , Proteoma , Osteoartritis/genética , Osteoartritis/metabolismoRESUMEN
OBJECTIVE: To elucidate the scientific advances made in the last 12 months within the realm of osteoarthritis genetics, genomics, and epigenetics. This review paper highlights major research publications that enhance our current understanding of the role of genetics, genomics, and epigenetics in osteoarthritis. METHODS: A systematic literature search was conducted on pubmed.ncbi.nlm.nih.gov on "March 17, 2023", using the following keywords: "osteoarthritis" in combination with any of these terms: "genetic(s)", "mutation(s)", "genomic(s)", "epigenetic(s)", "DNA methylation", "noncoding RNA", "lncRNA", "circular RNA", "microRNA", "transcriptomic(s)", "RNA sequencing", "single cell RNA sequencing", or "single nucleus RNA sequencing". The selection comprised original research articles published in the English language between the OARSI congresses of 2022 and 2023. RESULTS: A total of 2178 research articles were identified, which subsequently reduced to 67 unique articles relevant to the field. Current trends in osteoarthritis genetics research involve meta-analyses of various cohorts to explore the impact of gene variants on osteoarthritis-related outcomes, such as pain. Early developmental changes within the joint were also found to influence osteoarthritis through genetic variations. Researchers also prioritize testing the mechanisms and functions of miRNAs, circRNAs, and lncRNAs. Potential drug targets began to emerge; however, independent validation studies are lacking. Single cell RNA sequencing studies revealed unique immune cell populations in the knee; however, no study reported single nucleus RNA sequencing analysis. CONCLUSIONS: This review focused on recent advances in the above-mentioned themes within the field of osteoarthritis. These advances improve our understanding of the disease's complexity and guide us toward functional assessments of genetic/epigenetic outcomes and toward their translational and clinical applications.
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MicroARNs , Osteoartritis , Humanos , Epigénesis Genética , Genómica , Epigenómica , MicroARNs/genética , Osteoartritis/genética , ARN CircularRESUMEN
BACKGROUND: Anterior cruciate ligament (ACL) tears often result in knee effusion and an increased risk for developing knee osteoarthritis (OA) in the long run. The molecular profile of these effusions could be informative regarding initial steps in the development of posttraumatic OA after an ACL tear. HYPOTHESIS: The proteomics of knee synovial fluid changes over time after ACL injury. STUDY DESIGN: Descriptive laboratory study. METHODS: Synovial fluid was collected from patients with an acute traumatic ACL tear presenting to the office for evaluation (18.31 ± 19.07 days from injury) (aspiration 1) and again at the time of surgery (35.41 ± 58.15 days after aspiration 1 (aspiration 2). High-resolution liquid chromatography mass spectrometry was used to assess the quantitative protein profile of synovial fluid, and differences in protein profile between the 2 aspirations were determined computationally. RESULTS: A total of 58 synovial fluid samples collected from 29 patients (12 male, 17 female; 12 isolated ACL tear, 17 combined ACL and meniscal tear) with a mean age and body mass index of 27.01 ± 12.78 years and 26.30 ± 4.93, respectively, underwent unbiased proteomics analysis. The levels of 130 proteins in the synovial fluid changed over time (87 high, 43 low). Proteins of interest that were significantly higher in aspiration 2 included CRIP1, S100A11, PLS3, POSTN, and VIM, which represent catabolic/inflammatory activities in the joint. Proteins with a known role in chondroprotection and joint homeostasis such as CHI3L2 (YKL-39), TNFAIP6/TSG6, DEFA1, SPP1, and CILP were lower in aspiration 2. CONCLUSION: Synovial fluid from knees with ACL tears exhibits an increased burden of inflammatory (catabolic) proteins relevant to OA with reduced levels of chondroprotective (anabolic) proteins. CLINICAL RELEVANCE: This study identified a set of novel proteins that provide new biological insights into the aftermath of ACL tears. Elevated inflammation and decreased chondroprotection could represent initial disruption of homeostasis, potentially initiating the development of OA. Longitudinal follow-up and mechanistic studies are necessary to assess the functional role of these proteins in the joint. Ultimately, these investigations could lead to better approaches to predict and possibly improve patient outcomes.
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Lesiones del Ligamento Cruzado Anterior , Quitinasas , Humanos , Masculino , Femenino , Lesiones del Ligamento Cruzado Anterior/cirugía , Líquido Sinovial/metabolismo , Ligamento Cruzado Anterior/cirugía , Proteómica , Articulación de la Rodilla/cirugía , Biomarcadores/metabolismo , Quitinasas/metabolismoRESUMEN
BACKGROUND: Shoulder instability is a common pathology associated with an elevated risk of osteoarthritis (OA). Little is known about gene expression in the cartilage of the glenohumeral joint after dislocation events, particularly as it relates to the risk of posttraumatic OA. This study tested the hypothesis that gene expression in glenoid cartilage varies among acute instability (<3 dislocations), chronic instability (≥3 dislocations), and OA. METHODS: Articular cartilage was collected from the anteroinferior glenoid of consenting patients undergoing shoulder stabilization surgery (n = 17) or total shoulder arthroplasty (n = 16). Digital quantitative polymerase chain reaction was used to assess the relative expression of 57 genes (36 genes from OA risk allele studies, 21 genes from differential expression studies), comparing (1) OA versus instability (acute and chronic combined), (2) acute versus chronic instability, (3) OA versus acute instability, and (4) OA versus chronic instability. RESULTS: The expression of 11 genes from OA risk allele studies and 9 genes from differential expression studies was significantly different between cartilage from patients with instability and those with OA. Pro-inflammatory genes from differential expression studies and genes from OA risk allele studies were more highly expressed in cartilage in the OA group compared with the instability group, which expressed higher levels of extracellular matrix and pro-anabolic genes. The expression of 14 genes from OA risk allele studies and 4 genes from differential expression studies, including pro-inflammatory genes, anti-anabolic genes, and multiple genes from OA risk allele studies, was higher in the acute instability group compared with the chronic instability group. Cartilage in the OA group displayed higher expression of CCL3, CHST11, GPR22, PRKAR2B, and PTGS2 than cartilage in the group with acute or chronic instability. Whereas cartilage in both the acute and chronic instability groups had higher expression of collagen genes, cartilage in the OA group had expression of a subset of genes from OA risk allele studies or from differential expression studies that was lower than in the acute group and higher than in the chronic group. CONCLUSIONS: Glenoid cartilage has an inflammatory and catabolic phenotype in shoulders with OA but an anabolic phenotype in shoulders with instability. Cartilage from shoulders with acute instability displayed greater (cellular) metabolic activity compared with shoulders with chronic instability. CLINICAL RELEVANCE: This exploratory study identified genes of interest, such as CCL3, CHST11, GPR22, PRKAR2B, and PTGS2, that have elevated expression in osteoarthritic glenoid cartilage. These findings provide new biological insight into the relationship between shoulder instability and OA, which could lead to strategies to predict and potentially modify patients' risk of degenerative arthritis due to shoulder instability.
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Cartílago Articular , Inestabilidad de la Articulación , Osteoartritis , Luxación del Hombro , Articulación del Hombro , Humanos , Articulación del Hombro/cirugía , Articulación del Hombro/patología , Cartílago Articular/patología , Inestabilidad de la Articulación/complicaciones , Ciclooxigenasa 2 , Osteoartritis/genética , Osteoartritis/cirugía , Osteoartritis/complicaciones , Expresión Génica , Luxación del Hombro/genética , Luxación del Hombro/cirugía , Luxación del Hombro/complicacionesRESUMEN
BACKGROUND: Anterior cruciate ligament (ACL) tears are associated with posttraumatic osteoarthritis, but the early biological changes that initiate joint degeneration after this injury are not well characterized. ACL tears typically result in effusion in the knee, which may provide insight into the initial response of the joint to injuries. HYPOTHESIS: Patient- and injury-specific factors are associated with the proteomics of synovial fluid in knees with ACL tears. STUDY DESIGN: Descriptive laboratory study. METHODS: Synovial fluid was collected from 105 patients (38 male, 67 female) with an acute traumatic ACL tear. Patient- and injury-specific factors such as age, sex, body mass index, time from injury, presence/absence of concomitant meniscal tears, and location of concomitant bone bruises (if present) were recorded. The protein concentration of synovial fluid was measured, followed by benchmarking of samples for multi-affinity high-abundance protein depletion. An isotropically labeled high-resolution nano-liquid chromatography with tandem mass spectrometry-based proteomic approach was used to determine the synovial fluid protein profile. Data were processed, quality controlled, and analyzed computationally for each patient and injury factor. RESULTS: The proteomics of synovial fluid from ACL tears was associated with patient sex, injury pattern, and location of bone bruises but not with patient age, body mass index, or time from injury. Knees with an isolated ACL tear had higher glutathione peroxidase 1 (GPX1) and plastin 3 levels than knees with an ACL tear and meniscal tear. A bone bruise on the lateral femoral condyle was associated with elevated leptin and glucose-6-phosphate dehydrogenase (G6PD) levels. A bone bruise on the lateral tibial plateau was associated with decreased GPX1 levels. Male patients had higher matrix metalloproteinase 9 and lower G6PD levels than female patients. CONCLUSION: Patient sex, injury pattern, and bone bruise location were important determinants of the proteomic profile of effusion resulting from ACL tears. CLINICAL RELEVANCE: Longitudinal follow-ups to see if and how proteomic differences relate to clinical outcomes and mechanistic studies to assess the role that specific proteins play in the joint are warranted. Ultimately, these investigations could lead to better approaches to predict clinical outcomes and identify possible interventions to optimize outcomes in these patients.
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Lesiones del Ligamento Cruzado Anterior , Enfermedades de los Cartílagos , Contusiones , Traumatismos de la Rodilla , Ligamento Cruzado Anterior/metabolismo , Lesiones del Ligamento Cruzado Anterior/complicaciones , Contusiones/complicaciones , Femenino , Humanos , Traumatismos de la Rodilla/complicaciones , Traumatismos de la Rodilla/metabolismo , Articulación de la Rodilla , Masculino , Proteómica , Líquido Sinovial/metabolismoRESUMEN
Osteoarthritis is the most common joint disease in the world with significant societal consequences but lacks effective disease-modifying interventions. The pathophysiology consists of a prominent inflammatory component that can be targeted to prevent cartilage degradation and structural defects. Intracellular metabolism has emerged as a culprit of the inflammatory response in chondrocytes, with both processes co-regulating each other. The role of glutamine metabolism in chondrocytes, especially in the context of inflammation, lacks a thorough understanding and is the focus of this work. We display that mouse chondrocytes utilize glutamine for energy production and anabolic processes. Furthermore, we show that glutamine deprivation itself causes metabolic reprogramming and decreases the inflammatory response of chondrocytes through inhibition of NF-κB activity. Finally, we display that glutamine deprivation promotes autophagy and that ammonia is an inhibitor of autophagy. Overall, we identify a relationship between glutamine metabolism and inflammatory signaling and display the need for increased study of chondrocyte metabolic systems.
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Condrocitos , Osteoartritis , Animales , Cartílago , Condrocitos/metabolismo , Glutamina/metabolismo , Ratones , FN-kappa B/metabolismo , Osteoartritis/metabolismoRESUMEN
Rotator cuff tendinopathy, a major cause of shoulder disability, occurs due to trauma or degeneration. Our molecular understanding of traumatic and degenerative tears remains elusive. Here, we probed transcript level differences between traumatic and degenerative tears. Subacromial bursa tissues were collected from patients with traumatic or degenerative tears during arthroscopy (N = 32). Transcripts differentially expressed by tear etiology were detected by RNA-seq. RNA-seq results were validated by real-time quantitative polymerase chain reaction. We identified 334 protein-coding transcripts differentially expressed between traumatic and degenerative tears in females and 167 in males at a fold-change greater than 2. In females, XIRP2, MYL1, MYBPC1, TNNT1, and LMOD2, were highly expressed in traumatic tears whereas TPSD1, CDSN, RCVRN, LTBP4, and PTGS1 were elevated in degen tears. Transcripts elevated in traumatic tears represented muscle cell differentiation and development, and muscle contraction whereas those elevated in degenerative tears represented cell activation and immune response. In males, AZGP1, CNTFR, COL9A1, ZNF98, and EREG were highly elevated in traumatic tears whereas MYL2, HOXD11, SLC6A7, CADM1, and MMP17 were highly expressed in degenerative tears. Transcripts elevated in traumatic tears represented metabolic/catabolic processes, and transmembrane protein transport while processes related to cell cycle were mainly enriched in degenerative tears. Numerous long noncoding RNAs were differentially expressed between traumatic and degenerative tears in both sexes. In summary, this study provides insights into molecular biology of bursa in patients with rotator cuff tendon disease based on tear acuity and novel sex-based transcript differences that could inform clinical decision making in treating patients with traumatic or degenerative shoulder injuries.
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Lesiones del Manguito de los Rotadores , Transcriptoma , Femenino , Humanos , Masculino , RNA-Seq , Manguito de los Rotadores , Lesiones del Manguito de los Rotadores/genética , Rotura , HombroRESUMEN
Rotator cuff tears are a common pathology in the shoulder and generally have two underlying etiologies: traumatic and degenerative. Little is known about the molecular underpinning of these etiologies. Here we queried transcript level differences in tear etiology stratified by sex in 31 patients with rotator cuff tears. Tendon tissues were isolated from females (N = 16) and males (N = 15) with traumatic (N = 16) or degenerative (N = 15) tears during arthroscopy. Differentially expressed transcripts were identified by RNA-seq and biological processes were probed computationally. Expression of some transcripts was validated by real-time quantitative polymerase chain reaction (qPCR). We identified 339 and 336 transcripts differentially expressed by tear etiology in females and males, respectively, at a fold-change greater than |2|. In females, GSTM1, MT1G, S1008A, ACSM3, DSC, FAM110C, and VNN2 were elevated in traumatic tears representing metabolic/catabolic processes, and immune response whereas CHAD, CLEC3A, IBSP, TNMD, APLNR, and CPA3 were elevated in degenerative tears representing tissue morphogenesis and developmental processes, angiogenesis, and extracellular matrix organization. In males, ELOA3B, CXCL8, ADM, TNS4, and SPOCK1 were elevated in traumatic tears representing localization of endoplasmic reticulum, chromosome organization, leukocyte/neutrophil degranulation, and protein transport whereas MYL2, TNNC1, MB, CPA3, APLNR, and CA3 were highly upregulated in degenerative tears representing muscle cell differentiation and development and angiogenesis. Numerous novel lncRNAs were identified to be differentially expressed by tear etiology in both sexes. Real-time qPCR confirmed RNA-seq data. This study improves our understanding of tendon biology based on underlying etiology (trauma or degeneration) in a sex-specific manner. These findings may help drive clinical decision-making in females and males with traumatic and degenerative shoulder injuries.
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Lesiones del Manguito de los Rotadores , Manguito de los Rotadores , Transcriptoma , Femenino , Humanos , Masculino , RNA-Seq , Manguito de los Rotadores/patología , Lesiones del Manguito de los Rotadores/genética , Lesiones del Manguito de los Rotadores/patología , Rotura , Tendones/patologíaRESUMEN
Emerging evidence delineates that obesity, a complex metabolic disorder, impairs the structure and function of stromal cells residing in various tissues. The exuberant adipose tissue mass observed in obesity is, in part, associated with hyperplasia of adipocytes resulting from recruitment of multipotent stromal cells within the stromal vascular fraction of adipose tissues. However, a clear understanding of the causal role of stromal cells and biological factors in obesity is lacking. In our quest to understanding the role of kinesin family member 26B (KIF26B), we found that KIF26B regulates osteogenic and chondrogenic differentiation of stromal/progenitor cells. In this study, we sought to examine the effects of Kif26b loss-of-function on adipogenic differentiation of murine C3H10T1/2 multipotent stromal cells. In vitro loss-of-function studies demonstrated that Kif26b knockdown by lentivirus mediated shRNA markedly dampened the differentiation potential of C3H10T1/2â¯cells to adipocytes and suppressed the expression of adipogenesis-related genes e.g., Pparg, C/ebpα, Fabp4 and Adipoq. Analysis of cell cycle revealed that Kif26b knockdown resulted in elevated expression of cyclins (Ccnd1, Ccnb1, Ccna2) along with rapid cell cycle progression from G0/G1 to S and G2 phases. Mechanistically, reduced adipogenic differentiation of Kif26b-deficient cells was partly dependent on PPARγ, a key transcription factor implicated in adipogenesis. This observation was experimentally supported as loss of adipogenesis was partially rescued by the addition of PPARγ agonist, rosiglitazone in Kif26b-deficient cells. We further found that silencing of Kif26b lessened the protein levels of phospho-AKT(Ser473), phospho-S6(Ser235/236), and phospho-mTOR(Ser2448), the major component of AKT/mTOR complex 1 (mTORC1) signaling at the basal level. Together, these data define a novel role of Kif26b in regulating the commitment of C3H10T1/2 multipotent stromal cells to the adipocyte lineage and provide a practical framework for further experiments to establish its therapeutic potential for the treatment of problems associated with adipogenesis such as obesity at the cellular and molecular level.
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Adipogénesis , Cinesinas/metabolismo , Células Madre Multipotentes/metabolismo , Adipocitos/citología , Adipocitos/metabolismo , Animales , Ciclo Celular , Línea Celular , Técnicas de Silenciamiento del Gen , Ratones , PPAR gamma/metabolismo , Transducción de Señal , Células del Estroma/metabolismoRESUMEN
Ectopic calcification is an osteogenic process that leads to the formation of inappropriate bone within intra-articular soft tissues, often in response to injury or surgery. The molecular mechanisms governing this phenotype have yet to be determined. Using a population genetics approach, we identified an association of the kinesin superfamily member 26b (Kif26b) with injury-induced ectopic calcification through quantitative trait locus analysis of recombinant inbred mouse strains, consistent with a genomewide association study that identified KIF26B as a severity locus for ectopic calcification in patients with hip osteoarthritis. Despite these associations of KIF26B with ectopic calcification, its mechanistic role and functional implications have not yet been fully elucidated. Here, we aim to decipher the functional role of KIF26B in osseous and chondrogenic transdifferentiation of human and murine progenitor/stem cells and in a murine model of non-invasive injury-induced intra-articular ectopic calcification. We found that KIF26B ablation via lentivirus-mediated shRNA significantly arrested osteogenesis of progenitor/stem cells and suppressed the expression of typical osteogenic marker genes. Conversely, KIF26B loss-of-function increased chondrogenesis as demonstrated by enhanced Safranin-O staining and by the elevated expression of chondrogenic marker genes. Furthermore, cell function analysis revealed that KIF26B knockdown significantly decreased cell viability and proliferation and induced cellular apoptosis. Mechanistically, loss of osteogenesis was reverted by the addition of a Wnt agonist, SKL2001, demonstrating a role of KIF26B in canonical Wnt/ß-catenin signaling. Finally, intra-articular delivery of Kif26b shRNA in B6-129SF2/J mice significantly hampered the development of intra-articular ectopic calcification at 8 weeks after injury compared with mice treated with non-target scrambled shRNA. In summary, these observations highlight that KIF26B plays a crucial role in ectopic bone formation by repressing osteogenesis, but not chondrogenesis, potentially via modulating Wnt/ß-catenin signaling. These findings establish KIF26B as a critical determinant of the osteogenic process in pathologic endochondral bone formation and an actionable target for pharmacotherapy to mitigate ectopic calcification (and heterotopic ossification). © 2021 American Society for Bone and Mineral Research (ASBMR).
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Calcinosis/genética , Transdiferenciación Celular , Cinesinas , Osteogénesis , Células Madre/citología , Animales , Modelos Animales de Enfermedad , Silenciador del Gen , Cinesinas/genética , Ratones , Vía de Señalización WntRESUMEN
OBJECTIVE: Recent evidence delineates an emerging role of periostin in osteoarthritis (OA), since its expression after knee injury is detrimental to the articular cartilage. We undertook this study to examine whether intraarticular (IA) knockdown of periostin would ameliorate posttraumatic OA in a murine model. METHODS: Posttraumatic OA was induced in 10-week-old male C57BL/6J mice (n = 24) by destabilization of the medial meniscus (DMM), and mice were analyzed 8 weeks after surgery. Periostin expression was inhibited by small interfering RNA (siRNA) delivered IA using a novel peptide-nucleotide polyplex. Following histologic assessment of the mouse knee cartilage, the extent of cartilage degeneration was determined using Osteoarthritis Research Society International (OARSI) cartilage damage score, and severity of synovitis was also assessed. Bone changes were measured using micro-computed tomography. The effect and mechanism of periostin silencing were investigated in human chondrocytes that had been stimulated with interleukin-1ß (IL-1ß) with or without the IκB kinase 2 inhibitor SC-514. RESULTS: Periostin expression in mice with posttraumatic OA was significantly abolished using IA delivery of a peptide-siRNA nanoplatform. OARSI cartilage damage scores were significantly lower in mice receiving periostin siRNA (mean ± SEM 10.94 ± 0.66) compared to untreated mice (22.38 ± 1.30) and mice treated with scrambled siRNA (22.69 ± 0.87) (each P = 0.002). No differences in the severity of synovitis were observed. Subchondral bone sclerosis, bone volume/total volume, volumetric bone mineral density, and heterotopic ossification were significantly lower in mice that had received periostin siRNA treatment. Immunostaining of cartilage revealed that periostin knockdown reduced the intensity of DMM-induced matrix metalloproteinase 13 (MMP-13) expression and also diminished the phosphorylation of p65 and immunoreactivity of the aggrecan neoepitope DIPEN. Periostin knockdown also suppressed IL-1ß-induced MMP-13 and ADAMTS-4 expression in chondrocytes. Mechanistically, periostin-induced MMP-13 expression was abrogated by SC-514, demonstrating a link between periostin and NF-κB. CONCLUSION: IA delivery of the periostin-siRNA nanocomplex represents a promising clinical approach to mitigate the severity of joint degeneration in OA. Our findings may thus provide an unequivocal scientific rationale for longitudinal studies of this approach. Utilizing a cartilage-specific gene-knockout strategy will further illuminate the functional role of periostin in OA.
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Cartílago Articular/metabolismo , Moléculas de Adhesión Celular/genética , Articulación de la Rodilla/metabolismo , Osteoartritis/metabolismo , Proteína ADAMTS4/metabolismo , Animales , Densidad Ósea/fisiología , Cartílago Articular/diagnóstico por imagen , Moléculas de Adhesión Celular/metabolismo , Condrocitos/metabolismo , Silenciador del Gen , Articulación de la Rodilla/diagnóstico por imagen , Metaloproteinasa 13 de la Matriz/metabolismo , Ratones , FN-kappa B/metabolismo , Nanopartículas , Osteoartritis/diagnóstico por imagen , Osteoartritis/genética , ARN Interferente Pequeño , Sinovitis/diagnóstico por imagen , Sinovitis/genética , Sinovitis/metabolismoRESUMEN
BACKGROUND: Elevated levels of periostin (Postn) in the cartilage and bone are associated with osteoarthritis (OA). However, it remains unknown whether Postn loss-of-function can delay or prevent the development of OA. In this study, we sought to better understand the role of Postn in OA development and assessed the functional impact of Postn deficiency on post-traumatic and age-related OA in mice. METHODS: The effects of Postn deficiency were studied in two murine experimental OA models using Postn-/- (n = 32) and littermate wild-type (wt) mice (n = 36). Post-traumatic OA was induced by destabilization of the medial meniscus (DMM) in 10-week-old mice (n = 20); age-related OA was analyzed in 24-month-old mice (n = 13). Cartilage degeneration was assessed histologically using the OARSI scoring system, and synovitis was evaluated by measuring the synovial lining cell layer and the cells density in the synovial stroma. Bone changes were measured by µCT analysis. Serum levels of Postn were determined by ELISA. Expression of Postn and collagenase-3 (MMP-13) was measured by immunostaining. RNA-seq was performed on chondrocytes isolated from 21-day old Postn-/- (n = 3) and wt mice (n = 3) to discover genes and pathways altered by Postn knockout. RESULTS: Postn-/- mice exhibited significantly reduced cartilage degeneration and OARSI score relative to wt mice in post-traumatic OA after 8 weeks (maximum: 2.37 ± 0.74 vs. 4.00 ± 1.20, P = 0.011; summed: 9.31 ± 2.52 vs. 21.44 ± 6.01, P = 0.0002) and spontaneous OA (maximum: 1.93 ± 0.45 vs. 3.58 ± 1.16, P = 0.014; summed: 6.14 ± 1.57 vs. 11.50 ± 3.02, P = 0.003). Synovitis was significantly lower in Postn-/- mice than wt only in the DMM model (1.88 ± 1.01 vs. 3.17 ± 0.63; P = 0.039). Postn-/- mice also showed lower trabecular bone parameters such as BV/TV, vBMD, Tb.Th, and Tb.N and high Tb. Sp in both models. Postn-/- mice had negligible levels of serum Postn compared with wt. Immunofluorescent studies of cartilage indicated that Postn-/- mice expressed lower MMP-13 levels than wt mice. RNA-seq revealed that cell-cell-adhesion and cell-differentiation processes were enriched in Postn-/- mice, while those related to cell-cycle and DNA-repair were enriched in wt mice. CONCLUSIONS: Postn deficiency protects against DMM-induced post-traumatic and age-related spontaneous OA. RNA-seq findings warrant further investigations to better understand the mechanistic role of Postn and its potential as a therapeutic target in OA.
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Enfermedades de los Cartílagos , Cartílago Articular , Osteoartritis , Animales , Condrocitos , Modelos Animales de Enfermedad , Meniscos Tibiales , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoartritis/genéticaRESUMEN
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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Desarrollo Musculoesquelético/fisiología , Enfermedades Musculoesqueléticas/fisiopatología , Sistema Musculoesquelético/citología , Análisis de la Célula Individual/métodos , Secuenciación de Inmunoprecipitación de Cromatina , Citometría de Flujo , Homeostasis/fisiología , Humanos , RNA-SeqRESUMEN
OBJECTIVE: The purpose of this study is to assess if incident radiographic osteoarthritis (OA) is associated with clinical OA symptoms at midterm follow-up after arthroscopic partial medial meniscectomy (APMM). DESIGN: A total of 44 patients (43% females, mean age 50.1 ± 2.8 years, minimum 5.6-year follow-up) with isolated medial meniscal tears and no-to-mild preoperative radiographic OA underwent APMM. Incident radiographic OA was assessed using the modified Kellgren-Lawrence (K-L) classification. Patients completed the Knee Injury and Osteoarthritis Outcomes Score (KOOS), and subscale thresholds for assessment of a symptomatic knee (KOOS OA criteria) and for Patient non-Acceptable Symptom State (PASS-N) following anterior cruciate ligament reconstruction (ACL-R) were calculated. RESULTS: Incident medial compartment OA occurred in 50% of patients. Morbidly obese patients (body mass index ≥35 kg/m2) were more likely to demonstrate incident radiographic OA (100% vs. 41%, P = 0.002). Forty-three percent of patients met KOOS OA criteria, while 77% were PASS-N. Females were more likely to meet KOOS OA criteria (73% vs. 21%, P = 0.009). Patients with incident radiographic OA in any compartment were more likely than those without radiographic OA to meet KOOS OA criteria (71% vs. 17%, P = 0.008). Patients with preoperative K-L grade 2 changes in any compartment were more likely to meet KOOS OA criteria than those without K-L grade 2 changes in any compartment (83% vs. 35%, P = 0.037). CONCLUSIONS: Roughly half of APMM patients will have incident radiographic OA within 6 years of APMM, and this risk increases with obesity. Females and patients with incident radiographic OA are more likely to meet clinical thresholds for OA.
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Reconstrucción del Ligamento Cruzado Anterior , Obesidad Mórbida , Osteoartritis , Femenino , Estudios de Seguimiento , Humanos , Masculino , Meniscectomía/efectos adversos , Persona de Mediana Edad , Osteoartritis/diagnóstico por imagen , Osteoartritis/cirugíaRESUMEN
BACKGROUND: While knees with meniscal tears are associated with a heightened risk of developing osteoarthritis (OA), it is difficult to predict which patients are at the greatest risk for OA. Gene signatures in menisci that are resected during arthroscopic partial meniscectomy (APM) may provide insight into the risk of OA progression. HYPOTHESIS: Meniscal gene signatures at the time of APM will predict radiographic OA progression. STUDY DESIGN: Case series; Level of evidence, 4. METHODS: Meniscal fragments were collected from 38 patients without OA during clinically indicated APM of the medial meniscus. The expression of 28 candidate genes with known roles in cartilage homeostasis, OA, extracellular matrix degradation, and obesity was assessed by quantitative real-time polymerase chain reaction. Weightbearing radiographs obtained before surgery and at final follow-up were graded by a musculoskeletal radiologist using the Kellgren-Lawrence classification of OA. The association of meniscal gene expression at baseline with the progression of radiographic OA was determined. RESULTS: Gene expression and baseline and follow-up radiographic data were available from 31 patients (81.6%) at a mean follow-up of 6.2 ± 1.3 years. Patients without OA progression had significantly higher expression of 7 genes: MMP9 (5.1-fold; P = .002), IL8 (2.9-fold; P = .016), CCL3 (3.7-fold; P = .032), CCL3L1 (4.5-fold; P = .008), CXCL6 (6.2-fold; P = .010), LEP (5.2-fold; P = .004), and RETN (46-fold; P = .008). CONCLUSION: Gene expression in the meniscus at the time of APM may be associated with the risk for progression of OA after surgery. Elevated expression of the aforementioned genes may reflect a chondroprotective response. Stratifying the risk for OA progression after APM could facilitate targeted interventions to delay or prevent the development of OA. Further studies in a larger cohort with an extended follow-up, and inclusion of additional genes, are warranted to better characterize this association.
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Platelet-rich plasma (PRP) is an autologous blood product that contains a variety of growth factors (GFs) that are released upon platelet activation. Despite some therapeutic potential of PRP in vitro, in vivo data are not convincing. Bolus injection of PRP is cleared rapidly from the body diminishing its therapeutic efficacy. This highlights a need for a delivery vehicle for a sustained release of PRP to improve its therapeutic effect. In this study, we used microfluidics to fabricate biodegradable PRP-loaded polyethylene glycol (PEG) microspheres. PRP was incorporated into the microspheres as a lyophilized PRP powder either as is (powder PRP) or first solubilized and pre-clotted to remove clots (liquid PRP). A high PRP loading of 10% w/v was achieved for both PRP preparations. We characterized the properties of the resulting PRP-loaded PEG microspheres including swelling, modulus, degradation, and protein release as a function of PRP loading and preparation. Overall, loading powder PRP into the PEG microspheres significantly affected the properties of microspheres, with the most pronounced effect noted in degradation. We further determined that microsphere degradation in the presence of powder PRP was affected by platelet aggregation and clotting. Platelet aggregation did not prevent but prolonged sustained PRP release from the microspheres. The delivery system developed and characterized herein could be useful for the loading and releasing of PRP to promote tissue regeneration and wound healing or to suppress tissue degeneration in osteoarthritis, and intervertebral disc degeneration.