Transcriptome sequencing reveals inflammation and macrophage heterogeneity in subacromial bursa from degenerative shoulder disorders.

PURPOSE: We aimed to investigate the transcriptomic alterations that occur in the subacromial bursa (SAB) following degenerative or traumatic shoulder diseases. MATERIALS AND METHODS: RNA sequencing was employed to evaluate the transcriptomic alterations of the SAB in individuals afflicted with degenerative rotator cuff tear (RCT), traumatic RCT and proximal humerus fracture (PHF). To gain insights into the biological significance of differentially expressed genes (DEGs), we conducted an enrichment analysis utilizing Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. We further utilized single-cell RNA sequencing datasets of SAB from a recently published study to explore the associated cellular dynamics and alterations. RESULTS: We detected 1,790 up-regulated and 1,964 down-regulated DEGs between degenerative RCT and PHF, 2,085 up-regulated and 1,919 down-regulated DEGs between degenerative RCT and traumatic RCT, and 20 up-regulated and 12 down-regulated DEGs between traumatic RCT and PHF. Given the similar expression pattern between traumatic RCT and PHF, they were integrated as the traumatic group. In comparison with the traumatic group, 1,983 up-regulated and 2,205 down-regulated DEGs were detected in degenerative SAB. Enrichment analysis of up-regulated DEGs uncovered an elevated inflammatory and immunologic responses in degenerative SAB. Single-cell transcriptomic analysis revealed macrophage represented the immune cell with the most DEGs between the degenerative and traumatic RCT. CONCLUSION: Our results revealed that the SAB in degenerative RCT exhibited a different transcriptional signature compared to that in traumatic RCT, and enrichment analysis showed immunologic and inflammatory activations. Macrophages may play a fundamental role in this process.

Factors associated with genetic markers for rotator cuff disease in patients with atraumatic rotator cuff tears.

For atraumatic rotator cuff tears, genetics contributes to symptomatic tear risk and may influence rotator cuff healing after surgical repair. But little is known about how genetic factors influence rotator cuff tear patient characteristics at presentation. We collected saliva samples for genotyping from atraumatic rotator cuff tear patients. We examined nine single nucleotide polymorphisms (SNPs) associated with cuff tears in prior literature. We estimated associations of SNP dosage with (1) age at tear diagnosis, (2) bilateral atraumatic tear prevalence, and (3) tear size. Linear regression was used to estimate associations with diagnosis age adjusted for sex and principal components. Logistic regression and ordinal logistic regression were used to estimate associations with bilateral tear prevalence and tear size category, respectively, adjusting for age, sex, and principal components. Of 344 eligible patients, 336 provided sufficient samples for genotyping. Median age at tear diagnosis was 61, 22% (N = 74) had bilateral atraumatic tears, and 9% (N = 29) had massive tears. SNP rs13107325 in the SLC39A8 gene and rs11850957 in the STXBP6 gene were associated with younger diagnosis age even after accounting for multiple comparisons (rs13107325: -4 years, 95% CI = -6.5, -1.4; rs11850957: -2.7 years, 95% CI = -4.3, -1.1). No other significant associations were observed with diagnosis age, tear size, or bilateral tear prevalence. SLC39A8 encodes a Mn transporter. STXBP6 may play a role in inflammatory responses by altering phagocytosis and antigen presentation of monocytes and macrophages. Further research is needed to determine if genetic markers can be used alongside patient characteristics to aid in identifying optimal surgical repair candidates.

Gene Expression in Synovium of Rotator Cuff Tear Patients Determined by RNA Sequencing.

Rotator cuff tear (RCT) is a common shoulder disorder related to pain and dysfunction. However, the pathological mechanism of RCT remains unclear. Thus, this study aims to investigate the molecular events in RCT synovium and identify possible target genes and pathways as determined by RNA sequencing (RNA-Seq). The synovial tissue was biopsied from 3 patients with RCT (RCT group) and 3 patients with shoulder instability (Control group) during arthroscopic surgery. Then, differentially expressed (DE) mRNAs, long non-coding RNAs (lncRNAs) and micro RNAs (miRNAs) were comprehensively profiled by RNA-Seq. Gene ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and competing endogenous RNA (ceRNA) network analysis were performed to identify the potential functions of these DE genes. 447 mRNAs, 103 lncRNAs and 15 miRNAs were identified differentially expressed. The DE mRNAs were highlighted in inflammatory pathway including up-regulated T cell costimulation, positive regulation of T cell activation, and T cell receptor signaling. Down-regulated fatty acid degradation pathway and 5'-AMP-activated protein kinase (AMPK) signaling in RCT group are also enriched. Validation assay showed that the expression of pro-inflammatory molecules including IL21R, CCR5, TNFSF11, and MMP11 was significantly increased in RCT group compared with Control group. CeRNA analysis further revealed lncRNA-miRNA-mRNA regulatory networks involving IL21R and TNFSF11 in RCT. Activated synovial inflammation is the remarkable event of RCT. Importantly, increased T cell activation and disordered fatty acid metabolism signaling might play a significant role. ceRNA networks involving IL21R and TNFSF11 identified could potentially control the progression of RCT. In conclusion, our findings could provide new evidence for the molecular mechanisms of RCT and might identify new therapeutic targets.

Influence of Vitamin D Deficiency on the Expression of Genes and Proteins in Patients With Medium Rotator Cuff Tears.

BACKGROUND: Whether vitamin D deficiency is related to rotator cuff muscle and tendon physiology is controversial. PURPOSE: To assess the relationship between vitamin D deficiency and various gene expression patterns in patients with rotator cuff tears. STUDY DESIGN: Controlled laboratory study. METHODS: During arthroscopic surgery, samples from the supraspinatus muscle, deltoid muscle, and supraspinatus tendon were acquired from 12 patients with vitamin D deficiency (serum 25-hydroxyvitamin D concentration <20 ng/dL) and 12 patients with sufficient vitamin D levels (control group, serum 25-hydroxyvitamin D concentration ≥30 ng/dL) who were matched for age, sex, and tear size. Alterations in the expression of genes and proteins associated with myogenesis, muscle atrophy, adipogenesis, inflammation, and apoptosis, as well as in vitamin D receptor expression, were assessed using quantitative reverse transcription polymerase chain reaction, Western blotting, and immunohistochemistry and were compared between the 2 groups. RESULTS: Vitamin D receptor gene expression in the deltoid muscle was significantly lower in the vitamin D deficiency group than in the control group (P = .043). Additionally, in the deltoid muscle, myoDgene expression levels were lower and atrogin levels were higher in the vitamin D deficiency group than in the control group (P = .034 and P = .011, respectively). However, in the supraspinatus muscle, no differences were observed between groups in the expression of myogenesis- or atrophy-related genes (all P > .05). The expression of inflammation-related genes (interleukin (IL)-1ß and IL-6) was significantly higher in the vitamin D deficiency group, in both the deltoid and supraspinatus muscles (all P < .05). The supraspinatus tendon tissue did not show any significant differences in any gene expression evaluated (all P > .05). A correlation between gene and protein expression was observed for atrogin and IL-1ß in the deltoid muscle (P = .019 and P = .037, respectively) and for IL-6 in the supraspinatus muscle (P = .044). CONCLUSION: Vitamin D deficiency was not associated with the expression of myogenesis-related or muscle atrophy-related genes in the supraspinatus muscle of patients with rotator cuff tears, unlike in the deltoid muscle; rather, vitamin D deficiency was associated with increased proinflammatory cytokine expression. CLINICAL RELEVANCE: In patients with rotator cuff tears, vitamin D deficiency was observed to be associated with increased levels of proinflammatory cytokines in the rotator cuff muscles, without significant changes in gene expression related to myogenesis or muscle atrophy.

A Hyperglycemic Microenvironment Inhibits Tendon-to-Bone Healing through the let-7b-5p/CFTR Pathway.

BACKGROUND: Tendon-to-bone healing is a difficult process in treatment of rotator cuff tear (RCT). In addition, diabetes is an important risk factor for poor tendon-to-bone healing. Therefore, we investigated the specific mechanisms through which diabetes affects tendon-to-bone healing by regulating the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR). METHODS: Tendon-derived stem cells (TDSCs) were extracted from rats after which their proliferative capacities were evaluated by the MTT assay. The expression levels of CFTR and tendon-related markers were determined by qRT-PCR. Then, bioinformatics analyses and dual luciferase reporter gene assays were used to identify miRNAs with the ability to bind CFTR mRNA. Finally, CFTR was overexpressed in TDSCs to validate the specific mechanisms through which the high glucose microenvironment inhibits tendon-to-bone healing. RESULTS: The high glucose microenvironment downregulated mRNA expression levels of tendon-related markers and CFTR in TDSCs cultured with different glucose concentrations. Additionally, bioinformatics analyses revealed that let-7b-5p may be regulated by the high glucose microenvironment and can regulate CFTR levels. Moreover, a dual luciferase reporter gene assay was used to confirm that let-7b-5p targets and binds CFTR mRNA. Additional experiments also confirmed that overexpressed CFTR effectively reversed the negative effects of the hyperglycaemic microenvironment and upregulation of let-7b-5p on TDSC proliferation and differentiation. These findings imply that the hyperglycemic microenvironment inhibits CFTR transcription and, consequently, proliferation and differentiation of TDSCs in vitro by upregulating let-7b-5p. CONCLUSIONS: A hyperglycemic microenvironment inhibits TDSC proliferation in vitro via the let-7b-5p/CFTR pathway, and this is a potential mechanism in diabetes-induced poor tendon-to-bone healing.

Histological and molecular features of the subacromial bursa of rotator cuff tears compared to non-tendon defects: a pilot study.

BACKGROUND: The role of the subacromial bursa in the development or healing of shoulder pathologies is unclear. Due to this limited knowledge, we aimed to understand specific reactions of the subacromial bursa according to rotator cuff (RC) pathologies compared to non-tendon defects of the shoulder. We hypothesized that the tissue composition and inflammatory status of the bursa are likely to vary between shoulder pathologies depending on the presence and the extent of RC lesion. METHOD: Bursa samples from patients with either 1) shoulder instability with intact RC (healthy bursa, control), 2) osteochondral pathology with intact RC, 3) partial supraspinatus (SSP) tendon tear, or 4) full-thickness SSP tear were investigated histologically and on gene expression level. RESULT: Bursae from SSP tears differed from non-tendon pathologies by exhibiting increased chondral metaplasia and TGFß1 expression. MMP1 was not expressed in healthy bursa controls, but strongly increased with full-thickness SSP tears. Additionally, the expression of the inflammatory mediators IL1ß, IL6, and COX2 increased with the extent of SSP tear as shown by correlation analysis. In contrast, increased angiogenesis and nerve fibers as well as significantly upregulated IL6 and COX2 expression were features of bursae from patients with osteochondral pathology. Using immunohistochemistry, CD45+ leukocytes were observed in all examined groups, which were identified in particular as CD68+ monocytes/macrophages. CONCLUSION: In summary, besides the strong increase in MMP1 expression with SSP tear, molecular changes were minor between the investigated groups. However, expression of pro-inflammatory cytokines correlated with the severity of the SSP tear. Most pronounced tissue alterations occurred for the osteochondral pathology and full-thickness SSP tear group, which demonstrates that the bursal reaction is not exclusively dependent on the occurrence of an SSP tear rather than longstanding degenerative changes. The present bursa characterization contributes to the understanding of specific tissue alterations related to RC tears or non-tendon shoulder pathologies. This pilot study provides the basis for future studies elucidating the role of the subacromial bursa in the development or healing of shoulder pathologies.

Increasing transforming growth factor-beta concentrations with age decrease apelin in the rat rotator cuff.

BACKGROUND: The rotator cuff undergoes natural degeneration with age, leading to age-related rotator cuff tear; however, the precise mechanism remains unclear. Transforming growth factor-beta (TGF-ß) concentrations rise with age and TGF-ß contributes to the pathophysiology of skeletal muscle. TGF-ß has also been shown to suppress expression of the myokine, apelin, in skin fibroblasts. We hypothesized that TGF-ß expression in the rotator cuff changes with age and regulates apelin expression, thereby contributing to rotator cuff degeneration. METHODS: We used quantitative reverse-transcription polymerase chain reaction (Q-RT-PCR) to measure the expression of apelin and tendon-related genes (Tnmd, Col1a1, and Col3a1) in the rotator cuff of young (12 weeks), adult (24 weeks), and old (48 weeks) rats. Using Q-RT-PCR and enzyme-linked immunosorbent assay, we also measured Tgfb mRNA and TGF-ß protein levels, respectively. Furthermore, we used Q-RT-PCR to measure apelin mRNA levels in rotator cuff-derived cells after treatment with 0 (control) and 10 ng/mL recombinant TGF-ß. RESULTS: Apelin mRNA levels were significantly lower in old compared to young and adult rats. Similarly, tendon-related genes, Tnmd, Col1a1, and Col3a1, were significantly lower in adult and old rats than young rats. In contrast, Tgfb mRNA and TGF-ß protein were significantly higher in old compared to young rats. Stimulation with exogenous TGF-ß significantly decreased Apelin mRNA expression compared to control. CONCLUSIONS: TGF-ß regulates apelin expression in the rotator cuff and may play a key role in the degenerative pathology of the rotator cuff with age.

Distinct Gene Expression Profile in Patients With Poor Postoperative Outcomes After Rotator Cuff Repair: A Case-Control Study.

BACKGROUND: Impaired healing after rotator cuff repair is a major concern, with retear rates as high as 94%. A method to predict whether patients are likely to experience poor surgical outcomes would change clinical practice. While various patient factors, such as age and tear size, have been linked to poor functional outcomes, it is currently very challenging to predict outcomes before surgery. PURPOSE: To evaluate gene expression differences in tissue collected during surgery between patients who ultimately went on to have good outcomes and those who experienced a retear, in an effort to determine if surgical outcomes can be predicted. STUDY DESIGN: Case-control study; Level of evidence, 3. METHODS: Rotator cuff tissue was collected at the time of surgery from 140 patients. Patients were tracked for a minimum of 6 months to identify those with good or poor outcomes, using clinical functional scores and follow-up magnetic resonance imaging to confirm failure to heal or retear. Gene expression differences between 8 patients with poor outcomes and 28 patients with good outcomes were assessed using a multiplex gene expression analysis via NanoString and a custom-curated panel of 145 genes related to various stages of rotator cuff healing. RESULTS: Although significant differences in the expression of individual genes were not observed, gene set enrichment analysis highlighted major differences in gene sets. Patients who had poor healing outcomes showed greater expression of gene sets related to extracellular matrix production (P < .0001) and cellular biosynthetic pathways (P < .001), while patients who had good healing outcomes showed greater expression of genes associated with the proinflammatory (M1) macrophage phenotype (P < .05). CONCLUSION: These results suggest that a more proinflammatory, fibrotic environment before repair may play a role in poor healing outcome. With validation in a larger cohort, these results may ultimately lead to diagnostic methods to preoperatively predict those at risk for poor surgical outcomes.

Identification of a Novel Genetic Marker for Risk of Degenerative Rotator Cuff Disease Surgery in the UK Biobank.

BACKGROUND: While evidence indicates that familial predisposition influences the risk of developing degenerative rotator cuff disease (RCD), knowledge of specific genetic markers is limited. We conducted a genome-wide association study of RCD surgery using the UK Biobank, a prospective cohort of 500,000 people (40 to 69 years of age at enrollment) with genotype data. METHODS: Cases with surgery for degenerative RCD were identified using linked hospital records. The cases were defined as an International Classification of Diseases, Tenth Revision (ICD-10) code of M75.1 determined by a trauma/orthopaedic specialist and surgery consistent with RCD treatment. Cases were excluded if a diagnosis of traumatic injury had been made during the same hospital visit. For each case, up to 5 controls matched by age, sex, and follow-up time were chosen from the UK Biobank. Analyses were limited to European-ancestry individuals who were not third-degree or closer relations. We used logistic regression to test for genetic association of 674,405 typed and >10 million imputed markers, after adjusting for age, sex, population principal components, and follow-up. RESULTS: We identified 2,917 RCD surgery cases and 14,158 matched controls. We observed 1 genome-wide significant signal (p < 5 × 10-8) for a novel locus tagged by rs2237352 in the CREB5 gene on chromosome 7 (odds ratio [OR] = 1.17, 95% confidence interval [CI] = 1.11 to 1.24). The single-nucleotide polymorphism (SNP) rs2237352 was imputed with a high degree of confidence (info score = 0.9847) and is common, with a minor allele frequency of 47%. After expanding the control sample to include additional unmatched non-cases, rs2237352 and another SNP in the CREB5 gene, rs12700903, were genome-wide significant. We did not detect genome-wide significant signals at loci associated with RCD in previous studies. CONCLUSIONS: We identified a novel association between a variant in the CREB5 gene and RCD surgery. Validation of this finding in studies with imaging data to confirm diagnoses will be an important next step. CLINICAL RELEVANCE: Identification of genetic RCD susceptibility markers can guide understanding of biological processes in rotator cuff degeneration and help inform disease risk in the clinical setting. LEVEL OF EVIDENCE: Prognostic Level III. See Instructions for Authors for a complete description of levels of evidence.

A genome-wide association study for shoulder impingement and rotator cuff disease.

BACKGROUND: The purpose of the study was to identify genetic variants associated with rotator cuff disease by performing a genome-wide association study (GWAS) for shoulder impingement using the UK Biobank (UKB) cohort and then combining the GWAS data with a prior GWAS for rotator cuff tears. The loci identified by the GWAS and meta-analysis were examined for changes in expression following rotator cuff tearing using RNA sequencing. METHODS: A GWAS was performed using data from UKB with 3864 cases of shoulder impingement. The summary statistics from shoulder impingement and a prior study on rotator cuff tears were combined in a meta-analysis. Also, the previous association of 2 single-nucleotide polymorphisms (SNPs) with shoulder impingement from a published GWAS using the UKB was tested. Rotator cuff tendon biopsies were obtained from 24 patients with full-thickness rotator cuff tears who underwent arthroscopic rotator cuff repair (cases) and 9 patients who underwent open reduction internal fixation for a proximal humeral fracture (controls). Total RNA was extracted and differential gene expression was measured by RNA sequencing for genes with variants associated with rotator cuff tearing. RESULTS: The shoulder impingement GWAS identified 4 new loci: LOC100506457, LSP1P3, LOC100506207, and MIS18BP1/LINC00871. Combining data with a prior GWAS for rotator cuff tears in a meta-analysis resulted in the identification of an additional 7 loci: SLC39A8/UBE2D3, C5orf63, ASTN2, STK24, FRMPD4, ACOT9/SAT1, and LINC00890/ALG13. Many of the identified loci have known biologic functions or prior associations with diseases, suggesting possible biologic pathways leading to rotator cuff disease. RNA sequencing experiments show that expression of STK24 increases whereas expression of SAT1 and UBE2D3 decreases following rotator cuff tearing. Two SNPs previously reported to show an association with shoulder impingement from a prior UKB GWAS were not validated in our study. CONCLUSION: This is the first GWAS for shoulder impingement in which new data from UKB enabled the identification of 4 loci showing a genetic association. A meta-analysis with a prior GWAS for rotator cuff tearing identified an additional 7 loci. The known biologic roles of many of the 11 loci suggest plausible biologic mechanisms underlying the etiology of rotator cuff disease. The risk alleles from each of the genetic loci can be used to assess the risk for rotator cuff disease in individual patients, enabling preventative or restorative actions via personalized medicine.

Relationship between fatty infiltration and gene expression in patients with medium rotator cuff tear.

BACKGROUND: Fatty infiltration (FI) is a key prognostic factor that affects outcomes after rotator cuff repair and is radiologically evaluated using the Goutallier classification. The purpose of this study was to assess alterations in gene and protein expression according to the Goutallier classification in the supraspinatus muscle and any relationships among various gene expression profiles. METHODS: Twenty-four samples of the supraspinatus muscle from 12 patients with a high FI grade (grade 3 or 4) and 12 patients with a low FI grade (grade 1 or 2) with medium-sized tears were acquired during arthroscopic surgery. Alterations in the expression of genes and proteins associated with adipogenesis, fibrosis, inflammation, and muscle atrophy were compared between the high- and low-FI groups using reverse-transcription quantitative polymerase chain reaction, Western blotting, and immunohistochemistry. RESULTS: mRNA expression of not only the adipogenic genes (peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein α; P < .001 and P = .020) but also the fibrosis-related gene (α-smooth muscle actin; P < .001), inflammation-related genes (interleukin [IL]-1ß and tumor necrosis factor α; P = .041 and P = .039), and muscle atrophy-related genes (atrogin 1 and myostatin; P = .006 and P < .001) was higher in the high-FI group compared with that in the low-FI group. In addition, adipogenic gene expression was significantly correlated with the expression of other categories of genes (all P < .05, except atrogin 1). A correlation of gene and protein expression was observed for IL-1ß (P = .027) and myostatin (P = .029). CONCLUSIONS: The radiologic grading of FI was associated with the expression of various genes, including adipogenic, fibrotic, inflammatory, and atrophy-related genes, and these genes were closely correlated with each other in terms of expression. This information could be helpful in patient counseling.

Polymorphisms and alterations in gene expression associated with rotator cuff tear and healing following surgical repair: a systematic review.

BACKGROUND: Rotator cuff tears (RCTs) are a common cause of shoulder disability, yet both conservative and surgical treatment strategies can lead to poor results in some patient populations. Enhanced understanding of the genetic processes associated with RCTs can assist in the development of more effective management options and help predict individual responses to surgical treatment. This systematic review analyzes the current literature on the genetic footprint associated with RCTs and interprets these findings to enhance the current understanding of RCT pathogenesis, potential treatment regimens, and prognostic biomarkers of outcomes after surgical repair. METHODS: A systematic search of the Embase, PubMed, and Web of Science electronic databases was performed. Medical Subject Headings (MeSH) and Emtree index terms were formulated from the concept terms "rotator cuff tear," "genetics," and "human," and synonyms of these concepts were applied to the Web of Science search. Articles were screened against predefined inclusion and exclusion criteria. Eligible studies compared gene expression patterns and genetic polymorphisms between cases (with RCTs) and controls (without RCTs). Quality assessment was performed with studies being rated as high, moderate, or poor quality. A modified best-evidence synthesis was applied, and studies were determined to be of strong, moderate, or limited evidence. RESULTS: The search identified 259 articles. Of these studies, 26 were eligible for review. Two studies were considered poor quality; 15 studies, moderate quality; and 9 studies, high quality. Analysis of these articles found that RCTs were associated with alterations in genes that code for the extracellular matrix, cell apoptosis, immune and inflammatory responses, and growth factor pathways. In particular, there was strong evidence of a significant association between RCTs and the genes MMP3, TNC, and ESRRB. Strong evidence of an association between BMP5 upregulation and successful healing after surgical repair was also found. CONCLUSION: This review provides strong evidence of an genetic association with RCTs. The genotype and gene expression patterns detailed within this review can assist in deciphering the biological mechanisms resulting in RCTs, as well as predicting an individual's response to surgical repair. Future research could investigate whether manipulating these genes-or their associated signaling pathways-could assist in RCT healing and whether genetic biomarkers could be used clinically to predict patient outcomes after surgical repair of RCTs.

Genetic variants associated with rotator cuff tearing utilizing multiple population-based genetic resources.

BACKGROUND: The etiology of rotator cuff tearing is likely multifactorial, including a potential genetic predisposition. The purpose of the study was to identify genetic variants associated with rotator cuff tearing utilizing the UK Biobank (UKB) cohort, confirm variants using a separate genetic database, and evaluate tissue expression of genes with associated variants following rotator cuff tearing using RNA sequencing. METHODS: Genome-wide association study (GWAS): A GWAS was performed using data from UKB with 5701 cases of rotator cuff injury. RNA sequencing analyses: rotator cuff biopsies were obtained from 24 patients with full-thickness rotator cuff tears who underwent arthroscopic rotator cuff repair (cases) and 9 patients who underwent open reduction internal fixation for a proximal humerus fracture (controls). Total RNA was extracted and differential gene expression was measured by RNAseq for genes with variants associated with rotator cuff tearing. RESULTS: The results of the UKB GWAS identified 3 loci that reached genome-wide statistical significance: 2 loci on chromosome 7 in GLCCI1 (rs4725069; P = 5.0E-09) and THSD7A (rs575224171; P = 5.3E-09), and 1 locus on chromosome 2 in ZNF804A (rs775583810; P = 3.9E-09). The association with rotator cuff injury of the GLCCI1 single-nucleotide polymorphism (SNP; rs4725069) was confirmed in the Kaiser Permanente Research Bank cohort (P = .008). Twenty previously reported SNPs in 12 genes were evaluated using summary statistics from the UKB GWAS, which confirmed 3 SNPs in TNC with rotator cuff injury (rs1138545, rs72758637, and rs7021589; all P < .0024). Of 17 genes with variants associated with rotator cuff injury (14 previously from literature plus 3 new genes from current UKB GWAS), TIMP2, Col5A1, TGFBR1, and TNC were upregulated (P < .001 for all) and THSD7A was downregulated (P = .005) in tears vs. controls in the RNA sequencing data set. CONCLUSION: The UKB GWAS has identified 3 novel loci associated with rotator cuff tearing (ZNF804A, GLCCI1, THSD7A). Expression of the THSD7A gene was significantly downregulated in rotator cuff tears vs. controls supporting a potential functional role. Three previously reported SNPs in the TNC gene were validated in the UKB GWAS, supporting a role for this gene in rotator cuff tearing. Finally, TIMP2, Col5A1, TGFBR1, and TNC genes were found to have significantly upregulated tissue expression in cases vs. controls supporting a biologic role in tearing for these genes.

SAP30BP gene is associated with the susceptibility of rotator cuff tear: a case-control study based on Han Chinese population.

BACKGROUND: Multiple studies have indicated that genetic components contribute significantly to the risk of rotator cuff tears. Previous studies have suggested that the SAP30BP gene may play an essential role in the development of rotator cuff tears. The aim of this study was to evaluate the potential association of the SAP30BP gene with the susceptibility to rotator cuff tears in a Han Chinese population. METHODS: A total of 394 patients with rotator cuff tears and 998 healthy controls were included in the study. Twelve tag single nucleotide polymorphisms (SNPs) located in the region of the SAP30BP gene were selected for genotyping. Genetic association analyses were performed using χ2 tests for each SNP. Significant associations were searched in the GTEx database for their functional consequences. RESULTS: SNP rs820218 was significantly associated with rotator cuff tears (χ2 = 9.49, P = 0.0021, OR [95% CI] = 0.67 [0.52-0.87]). In addition, SNP rs820218 was found to be significantly associated with the gene expression level of SAP30BP in whole blood (NES = 0.12, P = 1.00 × 10-6). CONCLUSION: Our study has shown that the genetic polymorphism of SAP30BP contributes to the risk of rotator cuff tears in Chinese Han people. Individuals with the A allele for SNP rs820218 were less susceptible to developing rotator cuff tears.

The association between Interleukin-6 rs1800795/rs1800797 polymorphisms and risk of rotator cuff tear in a Chinese population.

Expression of proinflammatory cytokines, such as interleukin (IL)-6 (IL-6) and metalloproteases, are elevated in patients with rotator cuff tear (RCT). In order to investigate the role of IL-6 gene polymorphisms on RCT risk, we genotyped two SNPs on IL-6 gene (rs1800795 and rs1800797) in 138 RCT patients and 137 healthy controls using polymerase chain reaction (PCR) and Sanger sequencing. The IL-6 expression in shoulder joint synovial fluid was determined by using enzyme-linked immunosorbent assay (ELISA) method. The constant score and visual analog scale (VAS) were used to evaluate the clinical outcome of two s (surgicsal vs. conservative) for RCT patients. For rs1800795, individuals with the GG genotype or G allele had significantly higher risk of RCT. Elevated risk of tear size was associated with the GG genotype of the rs1800795 polymorphism. The IL-6 rs1800797 polymorphism was also associated with an increased risk of RCT, especially among female, drinkers, and individuals with B(MI) < 25 kg/m2. The elevated levels of IL-6 gene were observed among the mutant genotype of rs1800795/rs1800797 polymorphism. Surgical group is significantly better than conservative treatment from the perspective of constant score and VAS. Furthermore, CG genotype of rs1800795 polymorphism increased the constant score at 6 months in comparison with CC genotype. In conclusion, our study supports a role of IL-6 rs1800795/rs1800797 polymorphisms on increased RCT risk. The RCT patients with CG genotype of rs1800795 polymorphism have more obvious surgical treatment effects by influencing the IL-6 expression.

Genetic factors in rotator cuff pathology: potential influence of col 5A1 polymorphism in outcomes of rotator cuff repair.

BACKGROUND: Investigations in genetics have provided valuable information about the correlation between gene variants and tendinopathy. Single Nucleotide Polymorphisms of COL5A1 gene are reported to be involved in Achilles tendinopathy, chronic degenerative tendon changes at the elbow, and other tendinopathies. The influence of Single Nucleotide Polymorphisms of COL5A1 was previously analyzed in rotator cuff disease with confounding results. Moreover, the rs12722 polymorphism in COL5A1 gene has been implicated in the aetiology of musculoskeletal soft tissue injuries in several association studies. This study aims to analyse the possible influence of rs12722 polymorphism in COL5A1 in the outcomes of rotator cuff repair. METHODS: Seventy-nine patients were included in the study. DNA was extracted from 1.2 ml of venous blood and genotyped for COL5A1 SNPs rs12722. Rotator cuff muscle strength and range of motion (ROM) in anterior elevation, external and internal rotation of the shoulder were evaluated. RESULTS: Patients presenting COL5A1 SNP rs12722 CC showed a ROM of passive external rotation statistically significantly higher compared to patients with CT genotype and TT genotype. CONCLUSIONS: COL5A1 SNP rs12722 may influence the functional outcomes of RCRs, even though further studies are required to confirm these preliminary results.

Rotator Cuff Tear Susceptibility Is Associated With Variants in Genes Involved in Tendon Extracellular Matrix Homeostasis.

Rotator cuff tears (RCT) is a multifactorial disease with genetic factors contributing for the disease etiology. We hypothesized that genetic variants in genes involved in extracellular matrix (ECM) homeostasis may alter susceptibility to RCT. We evaluated 20 polymorphisms of genes involved in ECM homeostasis in 211 cases of full-thickness tears of the supraspinatus (Nfemales = 130; Nmales = 81) and 567 age-matched controls (Nfemales = 317; Nmales = 250). Multivariate logistic regressions were carried out with age, gender, genetic ancestry (based on the analysis of 61 biallelic short insertion/deletion polymorphisms), and common co-morbidities (diabetes, dyslipidemia, and smoking habits) as covariates. We observed that carriers of the rare allele of both studied variants of TGFB1, as well as their G/A (rs1800470/rs1800469) haplotype, were less susceptible to RCT (p < 0.05). In contrast, carriers of the G allele of MMP9 rs17576 (p = 0.014) or G/G haplotype (rs17576/rs17577; p < 0.001) had an increased risk for tendon tears. The presence of the T allele of MMP2 rs2285053 (p = 0.033), the T allele of MMP3 rs679620 (p = 0.024), and the TT-genotype of TIMP2 rs2277698 (p = 0.01) was associated with susceptibility to tears, especially in females. In males, the A allele of COL5A1 rs3196378 (p = 0.032) and the G allele of TGFBR1 rs1590 (p = 0.039) were independent risk factors for RCT. The C/T COL5A1 (rs3196378/rs11103544) haplotype was associated with a reduced risk of tears in males (p = 0.03). In conclusion, we identified the genetic variants associated with RCT susceptibility, thereby reinforcing the role of genes involved in the structure and homeostasis of the ECM of tendons in disease development. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:192-201, 2020.

Enhancement of tendon-bone healing after rotator cuff injuries using combined therapy with mesenchymal stem cells and platelet rich plasma.

OBJECTIVE: The injuries of rotator cuff will cause the shoulder dysfunctions. Due to limited self-regeneration abilities of the tendon-bone part, rotator cuff injuries remain a clinical challenge. Previous studies have proposed many strategies for treating this disease. In this work, we aimed to combine different strategies to achieve better beneficial effects on tendon-bone repair. MATERIALS AND METHODS: We isolated mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) and tested the effects of PRP on the gene expression, cell death resistance, and osteogenic differentiation of MSCs. Then, we utilized multiple strategies to treat rotator cuff injuries. We evaluated the expression of genes that related to tissue repair, bone formation, and tendon regeneration. We also tested the biomechanical property of repair tissues. RESULTS: We found that the in-vitro co-culture with PRP endowed MSCs with enhanced production of growth factors, better osteogenic differentiation ability, and stronger ability to resist cell death. Next, we applied MSCs, PRP, and MSCs-PRP combined therapies in rat rotator cuff injury model to compare their therapeutic effects in vivo. Through biomechanical testing, we found that the combined therapy was most efficient to promote tissue regeneration and enhance the biomechanical property of the newly generated bone. CONCLUSIONS: The combined treatment induced strongest signals related to angiogenesis, bone formation, and tendon generation in-situ. We demonstrated that the combination of MSCs and PRP synergistically promotes tendon-bone healing and holds great promise for the treatment of rotator cuff injuries.

Genetic basis of rotator cuff injury: a systematic review.

BACKGROUND: Rotator cuff disease is a widespread musculoskeletal pathology and a major cause of shoulder pain. Studies on familial predisposition suggest that genetic plays a role in the pathogenesis of rotator cuff disease. Several genes are responsible for rotator cuff disease. The aim of this study was to perform a systematic review on genetic association between rotator cuff disease and genes variations. METHODS: A systematic review of the literature was performed, in accordance with the PRISMA guidelines. PubMed, Medline, CINAHL, Cochrane, Embase and Google Scholar databases were searched comprehensively using the keywords: "Rotator cuff", "Gene", "Genetic", "Predisposition", "Single-nucleotide polymorphism" and "Genome-wide association". RESULTS: 8 studies investigating genes variations associated with rotator cuff tears were included in this review. 6 studies were case-control studies on candidate genes and 2 studies were GWASs. A significant association between SNPs and rotator cuff disease was found for DEFB1, FGFR1, FGFR3, ESRRB, FGF10, MMP-1, TNC, FCRL3, SASH1, SAP30BP, rs71404070 located next to cadherin8. Contradictory results were reported for MMP-3. CONCLUSION: Further investigations are warranted to identify complete genetic profiles of rotator cuff disease and to clarify the complex interaction between genes, encoded proteins and environment. This may lead to individualized strategies for prevention and treatment of rotator cuff disease. LEVEL OF EVIDENCE: Level IV, Systematic Review.