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Objectives: Frozen shoulder is a common shoulder disease that significantly affects the patient's life and work. Ferroptosis is a new type of programmed cell death, which is involved in many diseases. However, there have been no studies reporting the relationship between frozen shoulders and ferroptosis. This study identified potential molecular markers of ferroptosis in frozen shoulders to provide more effective strategies for the treatment of frozen shoulders., Methods: GSE238053 was downloaded from the Gene Expression Omnibus (GEO) dataset and intersected with ferroptosis genes to obtain differentially expressed genes (DEGs). The signaling pathways and biological functions of DEGs were performed by WebGestalt and Metascape. The interactions related to these DEGs and the key genes between frozen shoulders and ferroptosis was performed by STRING and Cytoscape. A frozen shoulders rat model was used to validate our predicted genes, Western Blot and qRT-PCR was used to assess the expression levels of our genes of interest., Results: A total of 34 DEGs between GSE238053 and Ferroptosis Database were obtained, most of which were involved in the HIF-1 signaling pathway and inflammatory response. A protein-protein interaction network was obtained by Cytoscape and the key genes (IL-6, HMOX1 and TLR4) were screened by MCODE. Our results of Western Blot showed that the protein expression level of TLR4 and HMOX1 were elevated, and the protein level of IL-6 decreased in frozen shoulders rat model. The mRNA level after frozen shoulders showed that IL-6 was upregulated, whereas TLR4 and HMOX1were downregulated., Conclusions: The results demonstrated that ferroptosis may affect the pathological process of frozen shoulders through these signaling pathways and genes. The identification of IL-6, HMOX1 and TLR4 genes can provide new therapeutic targets for frozen shoulders., (© 2024. The Author(s).)

The study aimed to explore the pathogenesis of secondary frozen shoulder and its influence on synovium tissue and angiogenesis by constructing a rat secondary frozen shoulder model along with transforming growth factor. 40 healthy male rats aged 8 weeks were divided into Sham group (n=10, no modeling treatment), Control group (n=10, modeling treatment), Low group (n=10, modeling treatment, and 10 mL/d transforming growth factor), and High group (n=10, modeling treatment, and 20 mL/d transforming growth factor). Hematoxylin and Eosin (HE) method was used for histological detection, and Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and immunohistochemical staining method were adopted to detect the expression of Matrix metalloproteinase-14 (MMP-14), mitogen-activated protein kinase (p38MAPK), and Vascular endothelial growth factor (VEGF). Compared with Sham group, the range of abduction and external rotation of rat glenohumeral joint in Control group, Low group, and High group was significantly reduced, and High group had the smallest range. Compared with the Sham group, the synovium in the Control group, the Low group, and the High group had obvious hyperplasia, and the blood vessels were significantly increased. Immunohistochemical staining and RT-PCR results showed that compared with Sham group, MMP-14, p38 MAPK, and VEGF in Control group, Low group, and High group all increased significantly, among which High group increased most. The secondary frozen shoulder is mainly manifested as synovial hyperplasia and increased blood vessels, which are related to the induction of MMP-14, p38 MAPK, and VEGF by transforming growth factor, which reveals the pathogenesis of secondary frozen shoulder to a certain extent, and lays a foundation for subsequent clinical treatment of secondary frozen shoulder.

As a common musculoskeletal disorder, frozen shoulder is characterized by thickened joint capsule and limited range of motion, affecting 2-5% of the general population and more than 20% of patients with diabetes mellitus. Pathologically, joint capsule fibrosis resulting from fibroblast activation is the key event. The activated fibroblasts are proliferative and contractive, producing excessive collagen. Albeit high prevalence, effective anti-fibrosis modalities, especially fibroblast-targeting therapies, are still lacking. In this study, microRNA-122 was first identified from sequencing data as a potential therapeutic agent to antagonize fibroblast activation. Then, Agomir-122, an analog of microRNA-122, was loaded into poly(lactic-co-glycolic acid) (PLGA) nanoparticles (Agomir-122@NP), a carrier with excellent biocompatibility for the agent delivery. Moreover, relying on the homologous targeting effect, we coated Agomir-122@NP with the cell membrane derived from activated fibroblasts (Agomir-122@MNP), with an attempt to inhibit the proliferation, contraction, and collagen production of abnormally activated fibroblasts. After confirming the targeting effect of Agomir-122@MNP on activated fibroblasts in vitro, we proved that Agomir-122@MNP effectively curtailed fibroblasts activation, ameliorated joint capsule fibrosis, and restored range of motion in mouse models both prophylactically and therapeutically. Overall, an effective targeted delivery method was developed with promising translational value against frozen shoulder., (© 2024. The Author(s).)

Frozen shoulder (FS) is a common disorder characterized by spontaneous onset of shoulder pain accompanied by progressive loss of range-of-motions. The cause of FS is still unclear, and radical therapy has not been established. With the final aim of preventing or curing FS at an earlier stage, we reviewed the pathological and biological features of this disease. Many studies indicate that the main pathology of FS is inflammation initially and fibrosis later. There are inflammatory cytokines, immune cells, fibrotic growth factors, and type-III collagen in the synovium and the joint capsule. The immune cell landscape switches from the macrophages to T cells. Activated fibroblasts seem to regulate the inflammatory and fibrotic processes. The imbalance between matrix metalloproteinases and tissue inhibitors of metalloproteases might promote fibrosis. Additionally, advanced glycation end-products are noted in the FS synovium. Diabetes mellitus and hypothyroidism are closely related to the development of FS. In terms of nonsurgical treatment, oral or intra-articular glucocorticoids are the only drugs that provide early benefit. Some other anti-inflammatory or antifibrotic drugs may potentially control the FS, but have not been proven effective in the clinical setting. Future studies should be targeted to develop steroid-sparing agents that inhibit biological events in FS., (© Japan College of Rheumatology 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Adipokines represent a spectrum of bioactive molecules that could modulate fibroblastic and inflammatory processes. The role of adipokines in the pathogenesis of frozen shoulder (FS), a common musculoskeletal disorder characterized by chronic inflammation, remains obscure., Purpose: To evaluate whether adipokines contribute to the pathogenic mechanisms of FS and to evaluate any potential correlation of adipokines with patients' symptoms., Study Design: Controlled laboratory study., Methods: Shoulder capsule specimens were obtained from 10 patients with FS and 10 patients with shoulder instability (control group). The specimens were dyed using hematoxylin and eosin and immunohistochemically assessed with antibodies targeting adipokines, collagen I, collagen III, and tumor necrosis factor α. Immunoreactivity was graded from "no" to "strong" in a blinded manner. Reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) analysis was conducted. Before the surgery, patient-reported frequency of pain, severity of pain, stiffness, and shoulder range of motion were documented., Results: In comparison with the control group, patients with FS had significantly greater pain frequency, pain severity, and stiffness and more limited shoulder range of motion ( P < .001). Hematoxylin and eosin- and Masson trichrome-stained samples from the FS group displayed hypercellularity and increased collagen fibers. Immunohistochemistry and RT-qPCR analyses indicated that expression of adipokines was significantly increased in FS capsules compared with the control group. The expression of collagen I, collagen III, and tumor necrosis factor α was also increased in FS capsules. No significant correlation was noted between adipokine expression and patient-reported outcomes in the control group, whereas in patients with FS, adiponectin expression was correlated with pain frequency ( r = 0.78; P = .01) and stiffness ( r = 0.73; P = .02). Visfatin was also correlated with pain frequency ( r = 0.70; P = .02)., Conclusion/clinical Relevance: This study indicated a potential role for adipokines in the pathogenesis of FS and demonstrated a correlation between adipokine expression and patients' pain and stiffness.

Objective: This study aims to explore the mechanism of miR-211-5p in extracellular vesicles (EVs) derived from bone marrow mesenchymal stem cells (BMSCs) in improving frozen shoulder (FS) in rat models., Methods: Rat BMSCs and EVs derived from rat BMSCs were isolated, identified, and then injected into rats to assess the expression of TGF-β, MMP1, MMP3, MMP12, GAP43, and PGP9.5 in shoulder capsule tissues. The range of motion of bilateral glenohumeral joints was assessed and pathological changes of shoulder capsule tissues were observed after hematoxylin-eosin staining. The binding sites of miR-211-5p to KDM2B and LACC1 to H3K4me3 were measured. FS rat models with LACC1 highly expressed were established to assess the motion of bilateral glenohumeral joints and expression of arthritis related factors in rats., Results: EVs were successfully extracted from BMSCs. Injection of BMSCs-EVs could improve the activity of bilateral glenohumeral joints and the pathological condition of joint capsule in rats. Elevated expression of miR-211-5p was found in rats injected with BMSCs-EVs. Dual luciferase assay showed that miR-211-5p had a binding site with KDM2B. ChIP, qRT-PCR, and western blot experiments showed BMSCs-EVs injection resulted in elevated enrichment of LACC1 promoter in shoulder capsule tissues of FS rats, and decreased mRNA and protein expression of KDM2B and increased H3K4me3 methylation. Overexpression of LACC1 could also improve the pathological condition of joint capsule tissue., Conclusion: miR-211-5p in EVs derived from BMSCs increased H3K4me3 methylation in shoulder capsule tissue of rats by binding KDM2B, resulting in up-regulated transcription level of LACC1 and improving FS., Availability of Data and Materials: The datasets used or analyzed during the current study are available from the corresponding author on reasonable request., Competing Interests: Conflict of interests The authors declare there is no conflict of interests., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Fibrosis is the main cause of limited range of motion (ROM) of shoulder in patients with frozen shoulder (FS). Overexpression of Interleukin 6 (IL-6) has been correlated with pathogenesis of FS. However, the underlying mechanism remains largely unexplored. In the current study, we focused on isolating synovial fibroblasts of FS and determining the influence of IL-6 as well as PI3K-Akt signaling pathway on the fibrotic process of synovial fibroblasts in FS by using RNA Sequencing (RNA-seq) and other molecular biology techniques. Synovial fibroblasts of FS express more extra cellular matrix (ECM) than that of control. RNA-seq results and bioinformatic analysis indicate that PI3K-Akt signaling pathway play an important role in the fibrotic process of FS, and IL-6 is the most related gene among those related to this process. The expression levels of IL-6 / IL-6R in FS synovial fibroblasts and IL-6 in culture supernatant were both significantly increased. siRNA interference with the expression of IL-6 attenuates the fibrosis level of FS as well as phosphorylation level of Akt. The findings suggest that synovial fibroblasts are key effector cells of fibrosis of FS. Activation of PI3K-Akt pathway can promote fibrosis of synovial fibroblasts in FS. IL-6 is up-regulated in synovial fibroblasts of FS and promoted the FS fibrosis through PI3K-Akt signaling pathway., Competing Interests: Declaration of Competing Interest The authors of the current study declare no conflicts of interest. Conflicts of interest The authors of the current study declare no conflicts of interest., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Objectives: Primary frozen shoulder (pFS) has three phases that differ in clinical presentation. It is characterized by contracture of the joint capsule. We hypothesized that there is a general upregulation of collagens in pFS, and that this is highest in the first phase of the disease. The aims of this study were to investigate the expression of various collagens and degradation of collagens in patients with primary pFS and relate this to the three phases of the condition., Methods: From twenty-six patients with pFS and eight control patients with subacromial impingement, biopsies were obtained during shoulder arthroscopy from the middle glenohumeral ligament and the anterior capsule, and mRNA levels for collagens, MMP-2 and -14 and TGF-β1, - β2 and -β3 in the tissue were analysed using real-time PCR., Results: Genes for collagens type I, III, IV, V, VI and XIV, were activated in pFS, and the total mRNA for all collagens was increased (P < 0.05). This upregulation was independent of disease phases in pFS. In addition, MMP-2, MMP-14, TGF-β1 and TGF-β3 were upregulated in all phases of the disease., Conclusion: There is a general upregulation and an increased degradation of collagens in pFS in all three phases of the disease. This indicates a constantly increased turnover of the fibrotic tissue in the capsule from pFS. The difference in clinical presentation of pFS observed in the three phases of the disease is not primarily a result of variations in collagen production., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Frozen shoulders (FS) is a major clinical concern, where chronic synovial inflammation, abnormal angiogenesis, and fibrosis represent the critical pathologies in the glenohumeral capsule. However, no pharmacotherapy has been introduced to treat this pathology. Tetrandrine (TET) has been proposed as a treatment for many diseases due to its strong anti-inflammatory, anti-angiogenic, and anti-fibrotic effects., Purpose: To study the anti-inflammatory, anti-angiogenic, and anti-fibrotic effects of TET on FS, and identify whether TET can prevent the development of FS in rats., Study Design: A controlled laboratory study., Methods: Forty-eight male Sprague-Dawley (SD) rats were randomly divided into control, TET, and FS groups. The TET group was intraperitoneally injected with TET every 2 days. TET and saline treatment were started on the day of FS surgery. After 8 weeks, the animals were sacrificed, and samples were collected for X-ray examination, glenohumeral range of motion (ROM) evaluation, histology and immunohistochemistry analysis, transmission electron microscopy (TEM) observation, and profibrogenic factors as well as proinflammatory cytokines measurements., Results: No significant difference in shoulder ROM was observed between the TET and control groups, but a significant difference was noted between these groups and the FS group (P < 0.01). Immunohistochemical staining showed no abnormal angiogenesis or fibrosis in the TET group or the control group. However, significant angiogenesis, collagen remodeling, and fibrosis were observed in the FS group, and the expression and proportion of type I and type III collagen in the FS group were significantly higher than those in the TET group or the control group (P < 0.01). TEM observation showed that TET protected the ultrastructure of collagen fibrous reticular arrangement of the articular capsule and prevented the formation of scar-like fibrotic structures, which are unique to FS. The significantly increased expression of Smad7 and the suppressed expression of Smad 2 in the TET group compared with that of the FS group indicated that TET also significantly inhibited the TGF-β1 intracellular signal pathway. The expression of profibrogenic factors and proinflammatory cytokines in the TET group and the control group was significantly lower than that in the TET group (P < 0.01)., Conclusion: The results demonstrated that TET protected the normal reticular structure of the capsule during the freezing period and prevented the development of FS by inhibiting inflammation, angiogenesis, and fibrosis in a rat FS model., Clinical Relevance: TET may be a safe and effective clinical medication for preventing and treating FS., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Adhesive capsulitis (AC) is a disabling condition of the shoulder joint affecting 2 to 5% of the general population. Our understanding of the molecular mechanisms is limited. The present study aimed to determine potential biomarkers of AC through transcriptomic analysis. This multi-centre study investigated patients undergoing arthroscopic capsulotomy surgery for resistant AC compared to those undergoing arthroscopic stabilization surgery for glenohumeral instability (control). Tissue samples were harvested from the anterior capsule during surgery. Total RNA was extracted and RNA-sequencing-based transcriptomics were performed. A number of genes deemed differentially expressed in RNA-sequencing analysis were validated using real-time reverse transcription polymerase chain reaction (RT-PCR). Baseline characteristics of the AC group (n = 22) were; mean age 52.7 years (SD: 10.2), 73% female, and Oxford Shoulder Score 19.6 (SD: 8.0), compared with the control group (n = 26), average age 23.9 years (SD: 5.2), 15% female, and Oxford Shoulder Score 39.0 (SD: 7.4). Transcriptomic analysis with false discovery rate correction and log 2 fold change cut-off of ±1.5 revealed 545 differentially expressed genes in AC relative to control. Bioinformatic analyses were carried out to identify biological processes and pathways enriched in this dataset. Real-time RT-PCR using two different normalization processes confirmed increased expression of matrix metallopeptidase 13 (MMP13) and platelet-derived growth factor subunit B (PDGFB), in patients with AC, while tumor necrosis factor α (TNFA) expression was reduced. These findings provide a comprehensive assessment of transcriptional changes associated with AC that give insights into the aetiology of the disease and provides a resource for molecular targets to better diagnose and treat this condition., (© 2020 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Background: The etiology of frozen shoulder (FS) remains uncertain. Advanced glycation end-products (AGEs) cause the cross-linking and stabilization of collagen and are increased in FS. The purpose of this study was to elucidate the pathogenesis of FS by evaluating the receptor of AGE (RAGE)-dependent pathways., Methods: Tissue samples of the coracohumeral ligament (CHL) and anterior inferior glenohumeral ligament (IGHL) were collected from 33 patients with FS, with severe stiffness, and 25 with rotator cuff tears (RCTs) as controls. Gene expression levels of RAGE, high-mobility group box 1 (HMGB1), Toll-like receptor 2 (TLR2), TLR4, nuclear factor-kappa B (NF-kB), and cytokines were evaluated using a quantitative real-time polymerase chain reaction. The immunoreactivities of carboxymethyllysine (CML), pentosidine, and RAGE were also evaluated. CML and pentosidine were further evaluated using high-performance liquid chromatography., Results: Gene expression levels of RAGE, HMGB1, TLR2, TLR4, and NF-kB were significantly greater in the CHLs and IGHLs from the FS group than in those from the RCT group. Immunoreactivities of RAGE and CML were stronger in the CHLs and IGHLs from the FS group than in those from the RCT group. Pentosidine was weakly immunostained in the CHLs and IGHLs from the FS group. CML using high-performance liquid chromatography was significantly greater in the CHLs and IGHLs from the FS group than in those from the RCT group., Conclusions: AGEs and HMGB1 might play important roles in the pathogenesis of FS by binding to RAGE and activating NF-kB signaling pathways. Suppression of these pathways could be a treatment option for FS., (Copyright © 2020 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

Rotator cuff lesion with shoulder stiffness is a major cause of shoulder pain and motionlessness. Subacromial bursa fibrosis is a prominent pathological feature of the shoulder disorder. MicroRNA-29a (miR-29a) regulates fibrosis in various tissues; however, the miR-29a action to subacromial bursa fibrosis remains elusive. Here, we reveal that subacromial synovium in patients with rotator cuff tear with shoulder stiffness showed severe fibrosis, hypertrophy, and hyperangiogenesis histopathology along with significant increases in fibrotic matrices collagen (COL) 1A1, 3A1, and 4A1 and inflammatory cytokines, whereas miR-29a expression was downregulated. Supraspinatus and infraspinatus tenotomy-injured shoulders in transgenic mice overexpressing miR-29a showed mild swelling, vascularization, fibrosis, and regular gait profiles as compared to severe rotator cuff damage in wild-type mice. Treatment with miR-29a precursor compromised COL3A1 production and hypervascularization in injured shoulders. In vitro, gain of miR-29a function attenuated COL3A1 expression through binding to the 3'-untranslated region (3'-UTR) of COL3A1 in inflamed tenocytes, whereas silencing miR-29a increased the matrix expression. Taken together, miR-29a loss is correlated with subacromial bursa inflammation and fibrosis in rotator cuff tear with shoulder stiffness. miR-29a repressed subacromial bursa fibrosis through directly targeting COL3A1 mRNA, improving rotator cuff integrity and shoulder function. Collective analysis offers a new insight into the molecular mechanism underlying rotator cuff tear with shoulder stiffness. This study also highlights the remedial potential of miR-29a precursor for alleviating the shoulder disorder.

Background: Although frozen shoulder (FS) is a common shoulder disorder, its pathogenesis is not yet determined. The function of matrix metalloproteinases (MMPs) is related to extracellular matrix remodeling. The purposes of this study were to investigate the pattern of sequential expression of MMPs in a rat model of shoulder contracture and to compare the expression of MMPs in the joint capsule between patients with FS and a control group., Methods: We obtained joint capsules from rats immobilized by molding plaster (a shoulder contracture model) at baseline, 3 days, 1 week, and 3 weeks (4 rats per time point; 16 rats in total). The expression of the inflammatory cytokine interleukin 6 (IL-6), MMP-2, and MMP-9 was examined by immunohistochemistry. We also obtained joint capsules from 21 patients with FS and 13 control patients with instability to quantify the expression levels of MMP-2 and MMP-9 by immunohistochemistry., Results: In the rat model, IL-6 and MMP-9 tended to be overexpressed in the joint capsule at 3 days and 1 week and MMP-2 at 3 days, 1 week, and 3 weeks. MMP-2 and MMP-9 were significantly overexpressed in the joint capsules of the patients with FS compared with those of control patients., Conclusion: The results from both human and animal studies suggest the involvement of MMP-2 and MMP-9 in the development of FS. Animal study showed that the sequential expression of IL-6 and MMPs may be associated with fibrosis of the joint capsule., (Copyright © 2018 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

Background: The etiology of frozen shoulder (FS) is unclear. Accordingly, this study used a label-free quantitative shotgun proteomic approach to elucidate the pathogenesis of FS based on protein expression levels., Methods: Tissue samples from the rotator interval (RI), middle glenohumeral ligament (MGHL), and anterior-inferior glenohumeral ligament (IGHL) were collected from 12 FSs with severe stiffness and 7 shoulders with a rotator cuff tear (RCT) as controls. Protein mixtures were digested and analyzed by nano-liquid chromatography/electrospray ionization-tandem mass spectrometry. Relative protein expression levels were calculated by the signal intensity of identified peptide ions on mass spectra. Differentially expressed proteins between FS and RCT samples were evaluated by a gene enrichment analysis using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes., Results: We identified 1594 proteins, 1358 of which were expressed in all 6 tissue groups. We detected more upregulated proteins in the upper (RI and MGHL) FS groups and the lower (IGHL) RCT group than in the comparative groups, respectively. Various proteins with functions in tissue repair, collagen metabolism and fibrillation, cell-cell and cell-matrix adhesion, blood coagulation, and the immune response were expressed more highly in the RI and MGHL FS groups than in the RCT group. Proteins with functions in phagocytosis, glutathione metabolism, retinoid metabolism, and cholesterol metabolism were expressed more highly in the IGHL RCT group than in the FS group., Conclusions: The pathophysiology of FS differs between the upper and lower parts of the joint capsule. Different treatment strategies for FS may be appropriate, depending on the location., (Copyright © 2018 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

It is fairly well understood that frozen shoulder involves several stages, which reflect the series of process from capsular inflammation and fibrosis to spontaneous resolution of this fibrosis. However, the underlying pathophysiologic process remains poorly determined. For this reason, management of frozen shoulder remains controversial. Determining the pathophysiological processes of frozen shoulder is a pivotal milestone in the development of novel treatment for patients with frozen shoulder. This article reviews what is known to date about the biological pathophysiology of frozen shoulder. Although articles for the pathophysiology of frozen shoulder provide inconsistent and inconclusive results, they have suggested both inflammation and fibrosis mediated by cytokines, growth factors, matrix metalloproteinases, and immune cells. Proinflammatory cytokines and growth factors released from immune cells control the action of fibroblast and matrix remodeling is regulated by the matrix metalloproteinases and their inhibitors. To improve our understanding of the disease continuum, better characterizing the biology of these processes at clearly defined stages will be needed. Further basic studies that use standardized protocols are required to more narrowly identify the role of cytokines, growth factors, matrix metalloproteinases, and immune cells. The results of these studies will provide needed clarity into the control mechanism of the pathogenesis of frozen shoulder and help identify new therapeutic targets for its treatment.

Background: The pathophysiological mechanisms behind proliferation of fibroblasts and deposition of dense collagen matrix in idiopathic frozen shoulder remain unclear. Alarmins (also known as danger signals) are endogenous molecules that are released into the extracellular milieu after infection or tissue injury and that signal cell and tissue damage., Purpose: To investigate whether the presence of alarmins is higher in patients with idiopathic frozen shoulder than in control subjects., Study Design: Controlled laboratory study., Methods: Shoulder capsule samples were collected from 10 patients with idiopathic frozen shoulder and 10 patients with unstable shoulders (control). The samples were stained with hematoxylin and eosin (H&E) and analyzed by immunohistochemistry using antibodies against alarmin molecules including high-mobility group protein B1 (HMGB1), interleukin 33, S100A8, S100A9, and the peripheral nerve marker PGP9.5. Immunoreactivities were rated in a blinded fashion from "none" to "strong." Immunohistochemical distribution within the capsule was noted. Before surgery, patient-ranked pain frequency, severity, stiffness, and the range of passive shoulder motion were recorded and statistically analyzed., Results: Compared with control patients, patients with frozen shoulder had greater frequency and severity of self-reported pain ( P = .02) and more restricted range of motion in all planes ( P < .05). H&E-stained capsular tissue from frozen shoulder showed fibroblastic hypercellularity and increased subsynovial vascularity. Immunoreactivity of alarmins was significantly stronger in frozen shoulder capsules compared with control capsules ( P < .05). Furthermore, the expression of the alarmin molecule HMGB1 significantly correlated ( r > 0.9, P < .05) with the severity of patient-reported pain., Conclusion: This study demonstrates a potential role for key molecular danger signals in frozen shoulder and suggests an association between the expression of danger molecules and the pain experienced by patients.

Knee pain is commonly seen in orthopedic and rehabilitation outpatient clinical settings. Patients with knee osteoarthritis (OA) are often complicated with joint soreness, swelling, weakness, and pain. These complaints are often caused by the excessive amount of synovial fluid (SF) accumulated in the bursae around the knee joint. This study was aimed to evaluate the effectiveness of platelet rich plasma (PRP) in treating patients with minor to moderate knee osteoarthritis (OA) combined with supra-patellar bursitis using a proteomic approach and clinical evaluation tool. In this study, 24 elderly patients with minor to moderate knee OA combined with supra-patellar bursitis were recruited. Musculoskeletal ultrasound was used for accurate needle placement for the aspiration of SF followed by subsequent PRP injections. Three monthly PRP injections were performed to the affected knees for a total of 3months. Approximately after the 2nd PRP injection, significant decreases in SF total protein concentrations, volumes, and Lequesne index values were observed. SF proteins associated with chelation and anti-aging physiological functions such as matrilin, transthyretin, and complement 5 increased at least 2-fold in concentrations. Proteins associated with inflammation, such as apolipoprotein A-I, haptoglobin, immunoglobulin kappa chain, transferrin, and matrix metalloproteinase decreased at least 2-fold in concentrations. Therefore, at least two monthly PRP injections may be beneficial for treating patients with minor to moderate knee OA combined with supra-patellar bursitis., (Copyright © 2017. Published by Elsevier Inc.)

Objectives: We evaluated mRNA expression levels of genes that encode TGF-β1; the TGF-β1 receptor; the collagen-modifying enzymes LOX, PLOD1, and PLOD2; and the extracellular matrix proteins COMP, FN1, TNC and TNXB in synovial/capsule specimens from patients with idiopathic adhesive capsulitis. Possible associations between the measured mRNA levels and clinical parameters were also investigated., Methods: We obtained glenohumeral joint synovium/capsule specimens from 9 patients with idiopathic adhesive capsulitis who had not shown improvement in symptoms after 5 months of physiotherapy. Adhesive capsulitis was confirmed in all patients by magnetic resonance imaging. We also obtained specimens from 8 control patients who had underwent surgery for acute acromioclavicular joint dislocation and who had radiological indication of glenohumeral capsule alteration based on arthroscopic evaluation. mRNA expression in the synovium/capsule specimens was analyzed by quantitative reverse transcription PCR. The B2M and HPRT1 genes were used as references to normalize target gene expression in the shoulder tissue samples., Results: The synovium/capsule samples from the patients with adhesive capsulitis had significantly higher TNC and FN1 expression than those from the controls. Additionally, symptom duration directly correlated with expression of TGFβ1 receptor I., Conclusion: Elevated levels of TNC and FN1 expression may be a marker of capsule injury. Upregulation of TGFβ1 receptor I seems to be dependent on symptom duration; therefore, TGFβ signaling may be involved in adhesive capsulitis. As such, TNC, FN1 and TGFβ1 receptor I may also play roles in adhesive capsulitis by contributing to capsule inflammation and fibrosis.

Background: The pathophysiologic mechanisms behind proliferation of fibroblasts and deposition of dense collagen matrix in idiopathic frozen shoulder remain unclear. Accumulation of advanced glycation end products (AGEs) with cross-linking and stabilization of collagen has been hypothesized to contribute to this pathophysiologic process. This study investigated whether the immunoreactivity of AGEs is higher in patients with idiopathic frozen shoulder than in the control groups., Methods: Shoulder capsule samples were collected from 8 patients with idiopathic frozen shoulder, 6 with unstable shoulders (control 1), and 8 with rotator cuff tears (control 2). The samples were hematoxylin and eosin stained and analyzed by immunohistochemistry using antibodies against AGEs. Immunoreactivities were rated in a blinded fashion from none (0) to strong (3). Immunohistochemical distribution within the capsule was noted., Results: Frozen shoulder patients had greater frequency and severity of self-reported pain (P = .02) than rotator cuff tear patients and more restricted range of motion in all planes (P < .05) than patients of the instability and rotator cuff tear groups. Hematoxylin and eosin-stained capsular tissue from frozen shoulder showed fibroblastic proliferation, increased numbers of adipocytes, and increased subsynovial vascularity. Immunoreactivity of AGEs was stronger in frozen shoulder capsules (2.8) than in instability (0.3; P = .0001) and rotator cuff tear (1.1; P = .016) capsules., Conclusion: This study highlights a potential role for AGEs in the pathogenesis of frozen shoulder. The overexpression of AGEs may explain the fibroblastic proliferation and deposition of collagen matrix in idiopathic frozen shoulder., Level of Evidence: Basic Science Study; Histology., (Copyright © 2016 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

Frozen shoulder is a relatively common disorder that leads to severe pain and stiffness in the shoulder joint. Although this disorder is self-limiting in nature, the symptoms often persist for years, resulting in severe disability. Recent studies using human specimens and animal models have shown distinct changes in the gene expression patterns in frozen shoulder tissue, indicating that novel therapeutic intervention could be achieved by controlling the genes that are potentially involved in the development of frozen shoulder. To achieve this goal, it is imperative to develop a reliable animal joint contracture model in which gene expression can be manipulated by gene targeting and transgenic technologies. Here, we describe a novel shoulder contracture mouse model. We found that this model mimics the clinical presentation of human frozen shoulder and recapitulates the changes in the gene expression pattern and the histology of frozen shoulder and joint contracture in humans and other larger animal models. The model is highly reproducible, without any major complications. Therefore, the present model may serve as a useful tool for investigating frozen shoulder etiology and for identifying its potential target genes., (© 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

The purpose of this study was to evaluate the expression of acid-sensing ion channels (ASICs) in the capsule and synovial fluid of patients with frozen shoulder. Capsular tissue and synovial fluid were obtained from 18 patients with idiopathic frozen shoulder (FS group) and 18 patients with instability of the shoulder (control group). The expressions of ASIC1, ASIC2, and ASIC3 in the capsule were determined using the reverse transcriptase-polymerase chain reaction, immunoblot analysis, and immunohistochemistry (IHC). The concentrations in synovial fluid were evaluated using an enzyme-linked immunosorbent assay. The mRNA expression of ASIC1, ASIC2 and ASIC3 in the capsule were significantly increased in the FS group compared with the control group. The protein levels of these three ASICs were also increased. The increased expressions were confirmed by IHC. Of the ASICs, ASIC3 showed the greatest increase in both mRNA and levels of expression compared with the control group. The levels of ASIC1 and ASIC3 in synovial fluid were significantly increased in the FS group. This study suggests that ASICs may play a role as mediators of inflammatory pain and be involved in the pathogenesis of frozen shoulder., (©2015 The British Editorial Society of Bone & Joint Surgery.)

Purpose: Frozen Shoulder Syndrome is a fibrosis of the shoulder joint capsule that is clinically associated with Dupuytren's disease, a fibrosis of the palmar fascia. Little is known about any commonalities in the pathophysiology of these connective tissue fibroses. β-catenin, a protein that transactivates gene expression, and levels of IGF2 mRNA, encoding insulin-like growth factor-II, are elevated in Dupuytren's disease. The aim of this study was to determine if correlating changes in β-catenin levels and IGF2 expression are evident in Frozen Shoulder Syndrome., Methods: Tissue from patients with Frozen Shoulder Syndrome and rotator cuff tear were obtained during shoulder arthroscopies. Total protein extracts were prepared from tissue aliquots and β-catenin immunoreactivity was assessed by Western immunoblotting. In parallel, primary fibroblasts were derived from these tissues and assessed for IGF2 expression by quantitative PCR., Results: β-catenin levels were significantly increased in Frozen Shoulder Syndrome relative to rotator cuff tear when assessed by Western immunoblotting analyses. IGF2 mRNA levels were significantly increased in primary fibroblasts derived from frozen shoulder syndrome tissues relative to fibroblasts derived from rotator cuff tissues., Conclusions: As in Dupuytren's disease, β-catenin levels and IGF2 expression are elevated in Frozen Shoulder Syndrome. These findings support the hypothesis that these connective tissue fibroses share a common pathophysiology.

Background: Nocturnal pain is commonly observed in patients with shoulder disorders such as a rotator cuff tear or frozen shoulder. This study was conducted to explore the possibility that melatonin plays a role as a mediator of nocturnal pain in patients with a rotator cuff tear or frozen shoulder., Methods: Subacromial bursa and joint capsule samples were collected from sixty-three patients: twenty-one patients with a rotator cuff tear, twenty-two with frozen shoulder, and twenty with shoulder instability (control group). The expression of melatonin receptor 1A (MTNR1A) and 1B (MTNR1B) and of acid-sensing ion channel 3 (ASIC3) in the subacromial bursa and the joint capsule were determined by real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis. The protein level of ASIC3 was measured by immunoblot analysis. To determine the effect of melatonin as a pain mediator, an in vitro study with use of primary cultured fibroblast-like synoviocytes was performed by semiquantitative RT-PCR analysis, immunoblot analysis, and enzyme-linked immunosorbent assay (ELISA)., Results: MTNR1A, MTNR1B, and ASIC3 expression was significantly increased in both the rotator cuff tear and frozen shoulder groups compared with the control group of patients with shoulder instability. Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) significantly stimulated the expression of MTNR1A and MTNR1B in primary cultured fibroblast-like synoviocytes treated with proinflammatory cytokines. Melatonin treatment at a physiological concentration (10 nM) induced ASIC3 expression and IL-6 production. Treatment with luzindole, a melatonin-receptor antagonist, reversed melatonin-stimulated ASIC3 expression and IL-6 production., Conclusions: Our study suggests that melatonin may play a role as a mediator of nocturnal pain with a rotator cuff tear or frozen shoulder, and this effect may be mediated via melatonin receptors., Clinical Relevance: Melatonin may be a therapeutic target of chronotherapy., (Copyright © 2014 by The Journal of Bone and Joint Surgery, Incorporated.)

Knee pain is commonly seen in orthopedic and rehabilitation outpatient clinical settings, and in the aging population. Bursitis of the knee joint, especially when the volume of the synovial fluid is large enough, can compress and distend the nearby soft tissues, causing pain in the knee joint. Out of all the bursae surrounding the knee joint, supra-patellar bursitis is most often associated with knee pain. Treatment strategies in managing supra-patellar bursitis include the aspiration of joint synovial fluid and then followed by steroid injection into the bursa. When supra-patellar bursitis is caused by degenerative disorders, the concept of viscosupplementation treatment may be effective by injecting hyaluronic acid into the bursa. However, the rheology or the changes in the concentrations of proteins (biomarkers) that are related to the development of bursitis in the synovial fluid is virtually unexplored. Therefore, this study aimed to identify the concentration changes in the synovial fluid total protein amount and individual proteins associated with supra-patellar bursitis using the Bradford protein assay and western immunoglobulin methods. A total of 20 patients were divided into two groups with 10 patients in each group. One group received the high molecular weight hyaluronic acid product of Synvisc Hylan G-F 20 and the other group received the low molecular weight hyaluronic acid product of Hya-Joint Synovial Fluid Supplement once per week injection into the bursa for a total of 3 weeks. Significant decreases in the synovial fluid total protein concentrations were observed after the second dosage of high molecular weight hyaluronic acid injections. Apolipoprotein A-I, interleukin 1 beta, alpha 1 antitrypsin, and matrix metalloproteinase 1 proteins revealed a trend of decreasing western immunoblotting band densities after hyaluronic acid injections. The decreases in apolipoprotein A-I and interleukin 1 beta protein band densities were significant in the high molecular weight hyaluronic acid injection group. Transthyretin, complement 5, and matrilin 3 proteins revealed a trend of increasing western immunoblotting band densities after hyaluronic acid injections. Transthyretin revealed significant increases in protein band densities in both the high and low molecular weight hyaluronic acid injection groups. This study may provide the rationale for targeting several biomarkers associated with lipid transport, inflammation, and anti-aging as possible disease modifying therapies for the treatment of supra-patellar bursitis and even degenerative joint disorders., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Frozen shoulder is a debilitating condition characterized by gradual loss of glenohumeral motion with chronic inflammation and capsular fibrosis. Yet its pathogenesis remains largely unknown. We hypothesized that the subacromial bursa may be responsible for the pathogenesis of frozen shoulder by producing inflammatory cytokines., Materials and Methods: We obtained joint capsules and subacromial bursae from 14 patients with idiopathic frozen shoulder and from 7 control subjects to determine the expression levels of interleukin (IL) 1α, IL-1β, IL-6, tumor necrosis factor α (TNF-α), cyclooxygenase (COX) 1, and COX-2 by real-time reverse transcriptase-polymerase chain reaction, immunohistochemistry, and enzyme-linked immunosorbent assay., Results: IL-1α, IL-1β, TNF-α, COX-1, and COX-2 were expressed at significantly high levels in the joint capsules of the frozen shoulder group compared with those of the control group. Intriguingly, IL-1α, TNF-α, and COX-2 were also expressed at significantly high levels in the subacromial bursae of the frozen shoulder group compared with those of the control group. Immunohistochemical analysis showed increased expression of COX-2 in both the joint capsules and subacromial bursae of the frozen shoulder group., Conclusions: These findings imply that elevated levels of inflammatory cytokines in the subacromial bursa may be associated with the pathogenesis of inflammation evolving into fibrosis., (Copyright © 2013 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Mosby, Inc. All rights reserved.)

Background: The purpose of this study was to investigate the presence of intercellular adhesion molecule-1 (ICAM-1) in shoulders with adhesive capsulitis ("frozen shoulder")., Methods: Glenohumeral capsular tissue was obtained from twenty-six patients (seventeen with adhesive capsulitis and nine controls), and ICAM-1 was evaluated with use of oligonucleotide arrays, real-time reverse transcription-polymerase chain reaction (RT-PCR), and immunohistochemistry. ICAM-1 was also evaluated in synovial fluid with use of western blotting (six patients with adhesive capsulitis and two controls) and in peripheral blood with use of an enzyme-linked immunosorbent assay (ELISA) (thirty-two patients with adhesive capsulitis, twenty with diabetes mellitus, and fourteen controls). The effect of ICAM-1 treatment on gene expression of cytokines related to inflammation and fibrosis was evaluated in cultured normal human synovial cells., Results: The level of ICAM-1 was significantly greater in capsular tissue from the glenohumeral joint of patients with adhesive capsulitis compared with controls as measured by oligonucleotide array analysis (0.12 ± 0.01 compared with 0.09 ± 0.00 arbitrary units) (p = 0.001), real-time RT-PCR (1.70 ± 0.19 compared with 0.67 ± 0.24 arbitrary units) (p < 0.05), and immunohistochemical staining. ICAM-1 was also significantly increased in the synovial fluid of patients with adhesive capsulitis (1.70 ± 0.18 arbitrary units) compared with normal controls (0.48 ± 0.17) (p < 0.05) and in serum of patients with adhesive capsulitis (633.22 ± 59.14 ng/mL) and patients with diabetes mellitus (671.25 ± 27.08 ng/mL) compared with controls (359.86 ± 44.29 ng/mL) (p < 0.05). Gene expression of cytokines related to inflammation and fibrosis in synoviocytes cultured in vitro was greater after three days of treatment with ICAM-1 and with ICAM-1 with glucose compared with untreated cells., Conclusions: ICAM-1 was increased in patients with adhesive capsulitis, similar to the increase that has been reported in patients with diabetes mellitus.

Background: Our understanding of the pathogenesis of frozen shoulder and why it is so painful is undetermined. This study investigated the expression of neuronal proteins in the capsular tissue of frozen shoulder., Methods: Shoulder capsular samples were collected from 8 patients with idiopathic adhesive capsulitis and 10 patients with a rotator cuff tear but no stiffness (controls). Samples were analyzed by immunohistochemistry using antibodies against protein gene product 9.5 (PGP9.5), a general nerve marker; growth associated protein 43 (GAP43), a nerve growth marker; nerve growth factor receptor p75; and CD34, an endothelial cell marker., Results: Samples from frozen shoulders showed subsynovial hypercellularity and fibroblastic proliferation, with increased expression of nerve growth factor receptor p75 and CD34 compared with controls. Nerves positive for PGP9.5 and GAP43 were more abundant in samples of frozen shoulder (2.8 ± 0.2 and 2.4 ± 0.4 per field; P < .01) compared with controls (1.6 ± 0.3 and 1.3 ± 0.3 per field; P < .05). Expression of neuronal proteins followed that of CD34., Conclusion: Increased expression of nerve growth factor receptor and new nerve fibers were found in the shoulder capsular tissue of patients with frozen shoulder compared with those without a frozen shoulder. These data suggest that neoinnervation and neoangiogenesis in the shoulder capsule are important events in the pathogenesis of frozen shoulder and may help explain the often-severe pain of patients with frozen shoulder., (Copyright © 2012 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Mosby, Inc. All rights reserved.)

Objective: To analyze changes in the capsule from idiopathic frozen shoulders and clarify their etiology., Materials and Methods: Samples (the rotator interval capsule, middle glenohumeral ligament (MGHL), and inferior glenohumeral ligament (IGHL)) were collected from 12 idiopathic frozen shoulders with severe stiffness and 18 shoulders with rotator cuff tears as a control. The number of cells was counted and the tissue elasticity of the samples was calculated by scanning acoustic microscopy (SAM). The amount of glycosaminoglycan content was assessed by alcian blue staining. Gene and protein expressions related to fibrosis, inflammation, and chondrogenesis were analyzed by quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC). Furthermore, the total genes of the two groups were compared by DNA microarray analysis., Results: The number of cells was significantly higher and the capsular tissue was significantly stiffer in idiopathic frozen shoulders compared with shoulders with rotator cuff tears. Staining intensity of alcian blue was significantly stronger in idiopathic frozen shoulders. Gene expressions related to fibrosis, inflammation, and chondrogenesis were significantly higher in idiopathic frozen shoulders compared with shoulders with rotator cuff tears assessed by both qPCR and DNA microarray analysis., Conclusion: In addition to fibrosis and inflammation, which used to be considered the main pathology of frozen shoulders, chondrogenesis is likely to have a critical role in pathogenesis of idiopathic frozen shoulders., (Copyright © 2011 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.)

There is a growing body of evidence supporting the use of hyaluronan (HA) in patients with adhesive capsulitis of the shoulder, although the mechanisms of the effect have not yet been clarified. This in vitro study examined the effects of HA on glenohumeral synovial/capsular fibroblasts (GSCFs) from patients with adhesive capsulitis of the shoulder. The study subjects were seven patients with primary or secondary adhesive capsulitis of the shoulder (average age: 55 years; range: 42-65). Synovial/capsular specimens were obtained from the rotator interval of each patient during arthroscopy. Part of the tissue specimen was used for histological analysis. The remainder of the tissue was prepared for cell culture. Various concentrations of HA (0.0-4.0 mg/mL) were added to the monolayer-cultured GSCFs from these patients. Histological analysis consistently demonstrated chronic nonspecific inflammation with synovial hyperplasia, proliferation of vessels and fibroblasts, and increased amount of extracellular matrix. Treatment with HA at various concentrations significantly and dose-dependently inhibited cell proliferation and decreased the expression levels of mRNA for adhesion-related procollagens and cytokines. Pretreatment with OS/37 did not reverse the inhibitory effect of HA. These results suggest that HA modulates cell proliferation and expression of the mRNA of adhesion-related procollagens and cytokines in GSCFs, preventing the progression of adhesion formation in patients with adhesive capsulitis of the shoulder., ((c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Background: Mitogen-activated protein (MAP) kinases are well-known molecules that play key roles in mechanical stress signals during skeletal development. To test our hypothesis that the synovium in frozen shoulders is induced by MAP kinases, immunohistochemical analyses for detecting expression and signal transduction of MAP kinases were performed in synovial tissue obtained from the rotator interval (RI) in frozen shoulders., Methods: Synovial tissues were examined from 10 frozen shoulder patients with a mean age of 55.4 years (46-62 years). Synovial tissues between the long head of the biceps tendon (LHB) and the RI in frozen shoulders were stained with hematoxylin and eosin (H&E) and then examined with immunohistochemical staining. Extracellular signal-regulated (ERK), the Jun N-terminal (JNK), and p38 mitogen-activated protein (MAP) kinases, nuclear factor kappaB (NF-kappaB), p50, CD29 (beta(1)-integrin), matrix metalloproteinase (MMP)-3, interleukin-6 (IL-6), CD56, CD68, S-100, and vascular endothelial growth factor (VEGF) were analyzed to detect expression patterns., Results: H&E showed vascular proliferation with fibrin and fibrous tissue in the synovium of frozen shoulders. ERK was expressed in the epithelial cells of vascular tissue, and JNK was expressed strongly in the interstitial cells around vascular tissue; p38 MAPK was not expressed. NF-kappaB was expressed in vascular tissue, and IL-6 was expressed around vascular tissue. CD29 (beta1-integrin) was expressed in vascular tissue and in superficial cells of synovial tissue. MMP-3 and VEGF were expressed on the surface layer of synovial tissue and vascular tissue, and CD68 was expressed on the surface layer. Nerve-related proteins, CD56 and S-100, were expressed weakly., Conclusions: Mechanical stress on the LHB and RI in the shoulder may induce ERK and JNK to express NF-kappaB by CD29 to develop capsule contracture, producing MMP-3, IL-6, and VEGF.

Several studies have demonstrated that inflammation in the subacromial bursa is an important component in the pathogenesis of impingement syndrome. We have demonstrated in a previous study that many inflammatory cytokines, including stromal cell-derived factor 1 (SDF-1, CXCL12), are increased in the subacromial bursa [Blaine et al. 2005. J Shoulder Elbow Surg 14(Suppl 1):84S-89S]. SDF-1 is a potent chemotactic and angiogenic factor that stimulates recruitment of inflammatory cells. In the current study, we proposed that the resident cells in subacromial bursal tissue produce SDF-1, which can play a role in the inflammatory reponse of bursal tissue, and that this chemokine can be regulated by steroid (dexamethasone) and nonsteroidal anti-inflammatory medications (NSAIDs). Twenty-two subacromial bursa tissues (18 bursitis and 4 normal bursa) were obtained intraoperatively from patients during shoulder surgery and analyzed using the cDNA Array technique in accordance with an IRB approved protocol. cDNA array results were confirmed with real-time reverse transcription-polymerase chain reaction (RT-PCR). Bursal cells (from 4 normal bursa, 3 bursitis) and two normal bone marrow with whole tissue explants were cultured for one passage. Cell culture supernatants were collected and SDF-1 protein was detected with enzyme-linked immunosorbent assay (ELISA). Cultured bursal cells were treated with a COX-2 inhibitor and dexamethasone, and cells was harvested at 1-day and 4-day intervals. SDF-1 expression was evaluated by real-time RT-PCR and ELISA. cDNA Array analysis demonstrated that the gene expression of SDF-1 was increased in patients with subacromial bursitis compared to controls (p < 0.05). Real-time RT-PCR also revealed that the mRNA expression of SDF-1 in bursitis tissue is increased 10-fold over control tissue. While the normal bursal cells produced negligible amounts of SDF-1 protein, cultured cells derived from bursitis lesion released as much SDF-1 protein (235 pg/100,000 cells) as normal bone marrow stromal cells (283 pg/100,000 cells) as measured by ELISA. The addition of a COX-2 inhibitor and dexamethasone to bursitis cell lines led to decreased SDF-1 expression levels compared to untreated bursitis cell lines. These studies demonstrate that there is a significant elevation of SDF-1 expression in the subacromial bursa of patients with rotator cuff disease. Furthermore, this chemokine can be downregulated by COX-2 inhibitors and steroids. These results provide biologic evidence for the use of steroid and NSAIDs in the treatment of subacromial bursitis. In the future, targeted inhibition of molecules such as SDF-1 in the subacromial bursa may present a therapeutic strategy that may avoid the side effects of these other (steroid and NSAID) medications.

Purpose: The pathophysiology of subacromial impingement syndrome is poorly understood. We investigated the expression of inflammatory cytokines, metalloproteases, and the cyclooxygenases in the subacromial bursa in control patients and in patients with rotator cuff tear., Type of Study: Basic science evaluation., Methods: Eighteen patients undergoing shoulder surgery had a subacromial bursa biopsy examination. Patients were divided into 2 groups. Group I (study group) had 10 patients with a full-thickness rotator cuff tear (RCT). Group II (control group) had 8 patients. Seven of 8 underwent shoulder arthroscopy with anterior capsular reconstruction for instability; 1 of 8 underwent open reduction internal fixation for acute proximal humerus fracture. None of the patients in group II had any history of symptoms or signs consistent with subacromial impingement. H&E and immunohistochemical antibody (MMP-1, MMP-9, IL-1, IL-6, TNF-alpha, COX-1, and COX-2) stained specimens were examined by 2 blinded observers using a histologic scale (grade 0 = no staining to grade 4 = intense staining)., Results: Histologic evidence of inflammation was present in all patients with RCT (group I). No or mild inflammation was noted in group II. The average staining grade for inflammatory cytokines (IL-1, IL-6, TNF-alpha) and proteinases (MMP-1 and MMP-9) was significantly more pronounced in the RCT group (P < .001). Cyclooxygenase enzymes (COX-1 and COX-2) were also increased in group II (P < .001)., Conclusions: A high level of expression of inflammatory cytokines, proteinases, and cyclooxygenase enzymes, known to produce a catabolic environment, is present in the subacromial bursa of patients with rotator cuff tear., Clinical Relevance: These findings support the role of nonsteroidal anti-inflammatory drugs and corticosteroids in RCT treatment, and emphasize the importance of subacromial bursectomy to reduce inflammation in RCT surgery.

Background: To compare the in vitro effects of selective COX-2 inhibitors (L-745,337, NS-398 and DFU) and of COX-unspecific diclofenac on release of PGE(2 )and 6-keto-PGF(1alpha) from inflamed bursa subacromialis tissue (IBST) obtained from a total of 35 patients with shoulder impingement syndrome (SIS)., Patients and Methods: Bursal specimens were incubated in the presence of drugs (0.01-1000 microM) for 20 min and 16 h., Results: After 20 min 10 microM diclofenac significantly inhibited formation of PGE(2) and 6-keto-PGF(1alpha), whereas L-745,337 and NS-398 (10-1000 microM) induced significant inhibition only at concentrations > or =100 microM. In contrast to equimolar diclofenac, DFU (0.01-10 microM) induced no inhibition of bursal PGE(2) release but a dose-dependent, although statistically not significant inhibition after 16 h. The inhibitory potency of diclofenac (0.01-10 microM) was even more increased during long-term incubation showing greater inhibition than DFU at all concentrations studied., Conclusion: The data suggest that in IBST in SIS in vitro the majority of PG is generated via the COX-1 pathway.

Little information exists on the molecular and biochemical pathophysiology of subacromial bursitis and rotator cuff disease. We investigated the pattern of expression of cytokines (interleukin [IL]-1beta, IL-1, IL-6, tumor necrosis factor [TNF] alpha, small inducible cytokines), metalloproteases, and cyclooxygenases in the subacromial bursa in patients with rotator cuff disease. Subacromial bursa specimens were prepared for molecular and biochemical analysis in patients undergoing shoulder surgery following an institutional review board-approved protocol. Specimens were analyzed for the presence of cytokines, metalloproteases, and cyclooxygenases by use of microarray for gene expression and immunohistocytochemistry. Microarray analysis for gene expression and immunohistochemistry demonstrated that the expression of several cytokine genes (TNF, IL-1alpha, IL-1beta, and IL-6) was increased in patients with subacromial bursitis compared with control specimens. Furthermore, the expression of metalloproteases (MMP-1 and MMP-9) and cyclooxygenases (COX-1 and COX-2) in the bursitis group was found to be increased as compared with controls. Although further investigation is required, these studies suggest that inflammation of the subacromial bursa does occur in patients with rotator cuff disease. These findings support the role of anti-inflammatory agents in the treatment of subacromial impingement and emphasize the importance of subacromial bursectomy to reduce inflammation in rotator cuff disease.

Adhesive capsulitis (AC) is a disorder that is characterized by shoulder pain and progressive limitation of both active and passive shoulder motion. Although the underlying pathological mechanisms of the disease are not well understood, the inflammatory reactions depending on the stage have been demonstrated histologically. The purpose of the study is to investigate the inflammatory changes that can be demonstrated with Tc-99m HIG in AC, and to determine the presence of correlations between scintigraphic findings and the clinical assessment. Twenty-one patients (12 females and 9 males) with a mean age of 50.57+/-8.49 were included in the study. AC was diagnosed according to recognized criteria. The planar X-ray images of the affected shoulders of all patients were normal. The patients were evaluated with the Constant Scoring System, and the functional and pain assessment parts of the American Shoulder and Elbow Surgeons' Form (ASES). Three phase bone scans and Tc-99m HIG scintigraphy were performed at least two days apart. Bone scan and Tc-99m HIG scintigraphy were evaluated visually and HIG uptake was evaluated in comparison with the contralateral normal shoulder. Bone scan demonstrated hypervascularity in 9 of the 21 patients (43%), whereas increased osteoblastic activity was detected in 19 (90%) in the affected shoulder. Tc-99m HIG uptake was positive in 12 (57%), and negative in 9 (43%) patients. All patients with increased Tc-99m HIG accumulation in the affected shoulder, also had increased osteoblastic activity on Tc-99m bone scintigraphy. A significant correlation was found between HIG uptake and constant, functional and pain scores. The difference between these scores was also statistically significant in patients with HIG positive and negative uptake. This study indicates that there is a good correlation between Tc-99m HIG scan findings and clinical scores. Tc-99m HIG accumulation in the affected shoulder may be related to continuing inflammatory reaction to AC. Tc-99m HIG scan may be a noninvasive, complementary method for demonstrating continuing inflammatory changes and may help in staging the disease.

Inflammation in the subacromial bursa causes pain in patients suffering from rotator cuff tear, with this long-lasting inflammation leading to fibrosis and thickening of the subacromial bursa. Both inflammatory cytokines and mechanical stress, and impingement in the subacromial space, might induce and worsen this inflammation. However, little is known of the mechanism of this inflammation. In this study, we used immunohistological staining to demonstrate the expression of Interleukin-1 beta (IL-1 beta), Tumor necrosis factor alpha (TNF-alpha), transforming growth factor beta (TGF-beta), and basic fibroblast growth factor (bFGF) in subacromial bursa derived from the patients suffering from rotator cuff tear. On the other hand the expression of these inflammatory cytokines and growth factors were little detected only to a small degree in patients with anterior shoulder instability who did not have severe shoulder pain and impingement in the subacromial space. Our findings suggest that those inflammatory cytokines and growth factors may play an important role in inflammation of the subacromial bursa. Controlling the expression of these cytokines and growth factors might be important for treating patients suffering from shoulder pain due to rotator cuff tear.

The subacromial bursa is recognized as a site associated with the shoulder pain caused by rotator cuff disease in the middle-aged and elderly. Substance P is contained in primary afferent nerves, and its quantity increases during chronic pain. The amount of substance P in the subacromial bursa of patients with rotator cuff disease was examined. Radioimmunoassay and immunohistochemistry were employed to quantify and localize substance P. The preoperative pain level was measured with a visual analogue scale with 0 as no pain, 5 as moderate, and 10 as severe. Thirty-seven patients that had undergone operation were divided into two groups: one composed of 19 patients with subacromial bursitis and a partial-thickness tear of the rotator cuff (nonperforated cuff) and the other composed of 18 patients with a full-thickness tear (perforated cuff). Subacromial bursae obtained from seven fresh cadavers with no shoulder pain before death were used as controls. The visual analogue scale showed significantly greater pain in the group with the nonperforated rotator cuff than in the group with the perforated cuff. Consistent with these results, the amount of substance P in the subacromial bursa was significantly greater in the former group than in the latter. Nerve fibers immunoreactive to substance P were localized around the vessels, with a larger number of fibers in the group with the nonperforated rotator cuff. Therefore, an increased amount of substance P in the subacromial bursa appears to correlate with the pain caused by rotator cuff disease.

Objective: To analyze the clinical features of acute gouty synovitis associated with thick, milky white, "chalky," urate laden synovial effusions, and to investigate the effects on synovial white blood cell (WBC) counts when leukocyte-rich rheumatoid effusions are incubated with a urate packed milky synovial fluid., Methods: Five patients (all men, mean age 70.8 years) with acute gouty synovitis (acute arthritis in 3, acute bursitis in 2) associated with "urate milk" were studied between 1993 and 1996., Results: Synovial effusions were thick, "chalky," and appeared "milky" white. The fluids were packed with monosodium urate (MSU) crystals, which sedimented upon standing, leaving a clear supernatant containing a few MSU crystals. The presence of massive amounts of MSU crystals and crystal clumps interfered with accurate determination of synovial WBC counts. Four fluids showed "a few leukocytes," and one a WBC count of 6750/mm3 with 91% neutrophils and several intraleukocytic crystals. Four patients had subcutaneous tophi. Of the risk factors associated with development of gout, the most frequent was ethanol abuse, in 4 and possibly all 5 patients. Incubation of leukocyte-rich rheumatoid synovial effusions with urate laden knee fluid from Patient 5 produced a greater reduction in synovial WBC counts compared to controls., Conclusion: Milky white synovial effusions containing massive quantities of urate crystals (referred to as "urate milk") may rarely occur in the setting of acute gouty arthritis or bursitis. Ethanol abuse appears to be a risk factor associated with the development of hyperuricemia and gout in these patients.

The purpose of this study was to test the hypothesis that specific cytokines are involved in the initiation and evolution of the fibrotic process in adhesive capsulitis of the shoulder. After approval from the Institutional Review Board, biopsies of shoulder capsule and synovium were collected during shoulder arthroscopy from 19 patients with adhesive capsulitis, 14 patients with nonspecific synovitis and no fibrosis or clinical evidence of adhesive capsulitis, and seven patients undergoing surgery for another pathology who had a normal capsule and synovium. Immunohistochemical localization with monoclonal antibodies to transforming growth factor-beta and its receptor, platelet-derived growth factor and its receptor, basic fibroblast growth factor, interleukin-1 beta, tumor necrosis factor-alpha, and hepatocyte growth factor was performed using standard immunoperoxidase techniques. The frequency of cytokine staining was correlated with the clinical diagnosis. Synovial cells, fibroblasts, T-cells, and B-cells were identified with specific antibodies, and newly synthesized matrix was examined for type-I and type-III collagen by immunohistochemical staining. The predominant cell types present were synovial cells and fibroblasts. Staining for type-III collagen in adhesive capsulitis tissues indicated new deposition of collagen in the capsule. There was staining for transforming growth factor-beta and its receptor, platelet-derived growth factor and its receptor, interleukin-1 beta, and tumor necrosis factor-alpha in adhesive capsulitis and nonspecific synovitis tissues, compared with minimal staining in normal capsule. Staining was more frequent in synovial cells than in capsular cells. The frequency of cell and matrix staining for transforming growth factor-beta, platelet-derived growth factor, and hepatocyte growth factor was greater in adhesive capsulitis tissues than in those from patients with nonspecific synovitis. No difference in the frequency of staining between primary (idiopathic) and secondary adhesive capsulitis was found. The results of this study indicate that adhesive capsulitis involves both synovial hyperplasia and capsular fibrosis. Cytokines such as transforming growth factor-beta and platelet-derived growth factor may be involved in the inflammatory and fibrotic processes in adhesive capsulitis. Matrix-bound transforming growth factor-beta may act as a persistent stimulus, resulting in capsular fibrosis. Understanding the basic pathophysiology of adhesive capsulitis is an important step in the development of clinically useful antifibrotic agents that may serve as novel treatments for patients with this conditions.

Cholesterol crystals were found in two patients with classic rheumatoid arthritis (RA). In one patient, cholesterol crystals were found in synovial fluid from both shoulder joints, and in the second they were in an olecranon bursa. To examine the possible systemic etiology of cholesterol crystals in synovial and bursal fluid, lipid concentrations and the presence of serum antilipoprotein antibodies were measured. Antilipoprotein antibodies were not found. The concentration of lipid and lipoproteins, as well as the normal pattern of lipoprotein on agarose gel, eliminates the possibility of hyperlipoproteinemia. Results seemed to exclude a systemic etiology for the formation of cholesterol crystals in synovial and bursal fluid in the RA patients. It appears that several local factors such as defective drainage, local destruction, increased permeability of synovial membrane, and intraarticular (bursal) bleeding are possible etiologies.

Basic calcium phosphate (BCP) crystals refer to a family of crystals including partially carbonate substituted hydroxyapatite, octacalcium phosphate, and tricalcium phosphate. These crystals have been found in and around joints and have been associated with several forms of arthritis and periarthritis. Identification of BCP crystals remains problematic because of the lack of a simple, reliable analytic procedure. Methods currently in use include alizarin red S staining, labelled diphosphonate binding, scanning and transmission electron microscopy with energy dispersive X-ray microanalysis, X-ray diffraction, and atomic force microscopy. Periarthropathies associated with BCP crystals include calcific tendinitis and bursitis. Intra-articular BCP crystal deposition is common in osteoarthritis, often found together with calcium pyrophosphate dihydrate crystals. Uncommon conditions in which BCP crystals are found include destructive shoulder arthropathies, acute inflammatory attacks of arthritis, and erosive arthritis. Secondary deposition of BCP crystals has been observed in chronic renal failure, in patients with "collagen vascular" diseases, following neurologic injury and after local corticosteroid injection.

Fifteen cases of acute gouty bursitis were seen among 136 crystal-proved cases of gout. Bursal aspirate yielded yellow or pink fluid in 10, chalky white fluid in 1, and a small amount of bloody fluid in 4. Monosodium urate crystals were present in all. Bursal fluid leucocyte counts averaged 2.9 X 10(9)/1 compared with synovial fluid leucocyte counts that averaged 25.5 X 10(9)/1 in cases of articular gout (P less than 0.05). Gouty, septic, and idiopathic (traumatic) bursitis share clinical features, and detailed bursal fluid analysis is crucial for diagnosis.