A Systematic Review of Extracellular Vesicle-Derived Piwi-Interacting RNA in Human Body Fluid and Its Role in Disease Progression.

The state of host cells is reflected in the cargo carried by their extracellular vesicles (EVs). This makes EV a potential source of biomarkers for human diseases. Piwi-interacting RNA (piRNA) regulates gene expression through epigenetic regulation and post-transcriptional gene silencing. Thus, piRNA profiling in EVs derived from human clinical samples could identify markers that characterize disease stages and unveil their roles in disease pathology. This review aimed to report the expression profiles of EV-derived piRNA (EV-piRNA) in various human samples, as well as their role in each pathology. A systematic review was conducted to collate the findings of human EV-piRNA from original research articles published in indexed scientific journals up to February 16, 2022. Article searches were performed in PubMed, Web of Science, and Scopus databases, using a combination of keywords, including "EV" and "piRNA." A total of 775 nonredundant original articles were identified. After subjecting articles to inclusion and exclusion criteria, 34 articles were accepted for this review. The piRNA expression levels among the small RNA profiles of human-derived EVs range from 0.09% to 43.84%, with the lowest expression level reported in urine-derived EVs and the highest percentage in plasma-derived EVs. Differentially expressed EV-piRNAs have been identified in patients with specific disease conditions compared to their counterparts (healthy control), suggesting an association between piRNA and progression in various diseases. Seven articles identified piRNA putative target genes and/or the pathway enrichment of piRNA target genes, and one study demonstrated a direct role of piRNA candidates in disease pathology. In conclusion, EV-piRNA has been isolated successfully from various human body fluids. EV-piRNA is a new research niche in human disease pathology. The expression profiles of EV-piRNA in various tissue types and disease conditions remain largely unexplored. Furthermore, there is currently a lack of guidelines on piRNA bioinformatics analysis, which could lead to inconsistent results and thus hinder the progression of piRNA discoveries. Finally, the lack of published scientific evidence on the role of EV-piRNA supports the need for future research to focus on the functional analysis of EV-piRNA as part of the route in piRNA discoveries.

Patient-specific Instrumentation Versus Standard Surgical Instruments in Primary Reverse Total Shoulder Arthroplasty: A Retrospective Comparative Clinical Study.

Aims: Patient-specific instrumentation (PSI) in primary shoulder arthroplasty has been studied; results supported the positive impact of the PSI on the glenoid positioning. Nevertheless, no clinical outcomes have been reported. We compare the clinical outcomes of primary reverse total shoulder arthroplasty using PSI versus the standard methods. Methods: Fifty-three patients with full records and a minimum of 24-months follow-up were reviewed, 35 patients received primary standard RSTA, and 18 patients received primary PSI RSTA. All patients were operated on in a single center. The median follow-up was 46 months (53 months in the standard group vs 39 months in the PSI group). Results: There was an overall significant post-operative improvement in the whole cohort (P< 0.05). The standard group had more deformed glenoids (B2, B3, C&D) and significantly low preoperative constant score and forward flexion (P=0.02 & 0.034). Compared to the PSI group (all were A1, A2, B1 &one type D), there were no statistically significant differences in any clinical outcome postoperatively. PSI neither prolonged the waiting time to surgery (P=0.693) nor the intraoperative time (P=0.962). Radiologically, PSI secured a higher percentage of optimum baseplate position and screw anchorage; however, no statistical correlation was found. Conclusion: In this series, both groups achieved comparable good outcomes. PSI did not achieve significantly better clinical outcomes than Standard after primary RSTA. Yet comparison has some limitations. PSI did not negatively impact the waiting time or the surgical time.

Cartilage debris and osteoarthritis risk factors influence gene expression in the synovium in end stage osteoarthritis.

BACKGROUND: Gene expression in healthy synovium remains poorly characterised. Thus, synovial functional activity changes associated with osteoarthritis (OA) are difficult to define. This study sought to identify differentially expressed genes (DEG) of end-stage OA and assess the influence of OA risk factors on these DEG. METHODS: Anonymised patient clinical data and x-ray images were analysed. Osteoarthritic and non-osteoarthritic patients with soft tissue or traumatic knee injuries were matched for body mass index (BMI) and sex. Tissue samples were partitioned for immunocytochemistry (IHC) and microarray analysis. Multiple bioinformatics applications were utilised to determine changes in functional and canonical pathway activation. RESULTS: Age, disease-modifying injections and hypertension were confounding factors between patient groups. Inflammation was present in all tissues. Cartilage debris and inflammatory aggregates were noted in many osteoarthritic patient tissues. IHC and expression analyses revealed upregulation of synoviolin 1 (SYVN1) in osteoarthritic synovium. Significant differential expression was noted in 2084 genes. Osteoarthritic synovium displayed a significant upregulation of 95% of DEG coding for proteins, relative to non-osteoarthritic synovium tissues. Unfolded protein response (UPR)-related genes were upregulated in osteoarthritic synovium; gene expression of molecules within many canonical pathways including protein ubiquitination and UPR pathways was modified by BMI and sex. CONCLUSIONS: The synovium of all three pathologies exhibited elements of an inflammatory response. Cartilage debris, age, BMI and sex influence DEG of osteoarthritic synovium. UPR pathway is the top deregulated canonical pathway identified in osteoarthritic synovium regardless of BMI and sex, while typical OA-associated inflammatory and matrix gene responses were minimal.

Histopathological dataset and demographic details of synovial tissues from patients with end-stage osteoarthritis, soft tissue and traumatic injuries of the knee.

Degradation of articular cartilage is the defining feature of end-stage osteoarthritis (OA) with osteophytes, subchondral sclerosis, malalignment and joint space narrowing being additional indicators of advanced disease. Obesity, older age and female gender are OA risk factors. Differing degrees of synovitis are observed in OA, soft tissue and traumatic injuries of the knee. The synovium is also subject to systemic, enhanced lipids and inflammatory mediators characteristic of obesity. Synovial cellular composition changes specific to OA and associated with its handling of cartilage debris are unclear. Triangulation of data from three knee pathologies was used to highlight findings pertaining to OA compared to non-OA. OA patient data was compared to non-OA from knee ligament and tibial frature patients at surgery. Knee pathology, gender and BMI informed patient identification. Once consented, patient inclusion and characterisation utilised data from clinical assessments, blood tests, function scores, and radiological imaging, scores and intraoperative assessment. Intra-operative synovial tissues from the same site and processed identically underpins in-depth analyses and comparisons of histopathological images from these different knee pathologies. This supports the identification of distinct changes in the cellular composition of the knee synovium characteristic of OA. This data underpins a better understanding of OA pathogenesis and disease progression vital for the design of targeted therapeutics. The tissue and cell data include detailed results from the semi-quantitative synovitis score established by Krenn and observational data for morphological features such as cartilage debris inclusion, inflammatory cells aggregate and infiltration. This histopathological data is presented in the context of detailed clinical and functional information. This data and the holistic study design can be used as a foundation for the multifactorial collection and analysis of clinical data from OA patients, OA severity measures, tissue immuno-histology and synovial inflammation analysis to underpin the details and comparisons needed in further studies into OA and its treatment globally.

Naturally-derived endoplasmic reticulum stress inhibitors for osteoarthritis?

Osteoarthritis (OA) is a chronic degenerative disease that affects the whole synovial joint. OA causes severe pain and disability that significantly affects the livelihood of an individual and incurs a huge socioeconomic burden. Current management strategies are limited to supporting functional improvement with physiotherapy and pain reduction as there are no drugs available that can reverse the progression of OA with only joint replacement surgery for late stage OA. OA is associated with advancing age and obesity, both of which compromise the functions of key endoplasmic reticulum (ER) molecular chaperones leading to improper protein folding and ER stress. Failure to restore protein homeostasis leads to increased cellular stress and eventually apoptotic cell death. Cartilage is avascular and is dependent on its constituent cells, chondrocytes, for extracellular matrix maintenance. Chondrocytes have limited proliferative capacity and their apoptosis eventually leads to extracellular matrix loss and cartilage degeneration. Recent studies on attenuating ER stress and chondrocytes apoptosis offer a credible strategy for reducing OA progression. The established roles of ER stress responses in OA have paved the way for targeted drug discovery studies aiming to mitigate ER stress and OA progression. In this review, in vitro, pre-clinical and clinical evidence of naturally-derived ER stress inhibitors for OA, the prospect and challenges in bringing these compounds to clinics are discussed.

Modulation of the mechanical responses of synovial fibroblasts by osteoarthritis-associated inflammatory stressors.

OBJECTIVES: To compare mechanobiological response of synovial fibroblasts (SFb) from OA patient cohorts under mechanical load and inflammatory stressors for better understanding of SFb homeostatic functions. METHODS: Primary SFb isolated from knee synovium of OA obese (OA-ob:SFb), OA-pre-obese (OA-Pob:SFb), non-OA arthroscopic (scope:SFb), and non-OA arthroscopic with cartilage damage (scope-CD:SFb) were exposed to OA-conditioned media (OACM), derived from OA obese (OA-ob:CM), OA-pre-obese (OA-Pob:CM), and mechanical stretch at either 0 %, 6 % or 10 % for 24 h. Differences in the mRNA levels of genes involved in extracellular matrix production, inflammation and secretory activity were measured. RESULTS: Despite the significant BMI differences between the OA-ob and OA-Pob groups, OA-Pob has more patients with underlying dyslipidaemia, and low-grade synovitis with higher levels of secreted proteins, CXCL8, COL4A1, CCL4, SPARC and FGF2 in OA-Pob:CM. All primary SFb exhibited anti-proliferative activity with both OA-CM. Mechanical stretch stimulated lubricin production in scope:SFb, higher TGFß1 and COL1A1 expressions in scope-CD:SFb. OA-Pob:CM stimulated greater detrimental effects than the OA-ob:CM, with higher pro-inflammatory cytokines, IL1ß, IL6, COX2 and proteases such as aggrecanases, ADAMTS4 and ADAMTS5, and lower ECM matrix, COL1A1 expressions in all SFb. OA-ob:SFb were unresponsive but expressed higher pro-inflammatory cytokines under OA-Pob:CM treatment. CONCLUSION: Both mechanical and inflammatory stressors regulate SFb molecular functions with heterogeneity in responses that are dependent on their pathological tissue of origins. While mechanical stretch promotes a favorable effect with enhanced lubricin production in scope:SFb and TGFß1 and COL1A1 in scope-CD:SFb, the presence of excessively high OA-associated inflammatory mediators in OA-Pob:CM, predominantly SPARC, CXCL8 and FGF2 drive all SFb regardless of pathology, towards greater pro-inflammatory activities.

Development of a novel in vitro insulin resistance model in primary human tenocytes for diabetic tendinopathy research.

BACKGROUND: Type 2 diabetes mellitus (T2DM) had been reported to be associated with tendinopathy. However, the underlying mechanisms of diabetic tendinopathy still remain largely to be discovered. The purpose of this study was to develop insulin resistance (IR) model on primary human tenocytes (hTeno) culture with tumour necrosis factor-alpha (TNF-α) treatment to study tenocytes homeostasis as an implication for diabetic tendinopathy. METHODS: hTenowere isolated from human hamstring tendon. Presence of insulin receptor beta (INSR-ß) on normal tendon tissues and the hTeno monolayer culture were analyzed by immunofluorescence staining. The presence of Glucose Transporter Type 1 (GLUT1) and Glucose Transporter Type 4 (GLUT4) on the hTeno monolayer culture were also analyzed by immunofluorescence staining. Primary hTeno were treated with 0.008, 0.08, 0.8 and 8.0 µM of TNF-α, with and without insulin supplement. Outcome measures include 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-d-glucose (2-NBDG) assay to determine the glucose uptake activity; colourimetric total collagen assay to quantify the total collagen expression levels; COL-I ELISA assay to measure the COL-I expression levels and real-time qPCR to analyze the mRNA gene expressions levels of Scleraxis (SCX), Mohawk (MKX), type I collagen (COL1A1), type III collagen (COL3A1), matrix metalloproteinases (MMP)-9 and MMP-13 in hTeno when treated with TNF-α. Apoptosis assay for hTeno induced with TNF-α was conducted using Annexin-V FITC flow cytometry analysis. RESULTS: Immunofluorescence imaging showed the presence of INSR-ß on the hTeno in the human Achilles tendon tissues and in the hTeno in monolayer culture. GLUT1 and GLUT4 were both positively expressed in the hTeno. TNF-α significantly reduced the insulin-mediated 2-NBDG uptake in all the tested concentrations, especially at 0.008 µM. Total collagen expression levels and COL-I expression levels in hTeno were also significantly reduced in hTeno treated with 0.008 µM of TNF-α. The SCX, MKX and COL1A1 mRNA expression levels were significantly downregulated in all TNF-α treated hTeno, whereas the COL3A1, MMP-9 and MMP-13 were significantly upregulated in the TNF-α treated cells. TNF-α progressively increased the apoptotic cells at 48 and 72 h. CONCLUSION: At 0.008 µM of TNF-α, an IR condition was induced in hTeno, supported with the significant reduction in glucose uptake, as well as significantly reduced total collagen, specifically COL-I expression levels, downregulation of candidate tenogenic markers genes (SCX and MKX), and upregulation of ECM catabolic genes (MMP-9 and MMP-13). Development of novel IR model in hTeno provides an insight on how tendon homeostasis could be affected and can be used as a tool for further discovering the effects on downstream molecular pathways, as the implication for diabetic tendinopathy.

Subacromial impingement syndrome: An electromyographic study of shoulder girdle muscle fatigue.

Muscle fatigue affecting glenohumeral and/or scapular muscles is suggested as one of the contributing factors to the development of subacromial impingement syndrome (SAIS). Nonetheless, the fatigability of shoulder girdle muscles in association with the pathomechanics of SAIS has not been reported. This study aimed to measure and compare fatigue progression within the shoulder girdle musculature of patients and healthy controls. 75 participants including 39 patients (20 females; 19 males) and 36 healthy controls (15 females; 21 males) participated in the study. Study evaluated the progression of muscle fatigue in 15 shoulder girdle muscles by means of surface and fine-wire EMG during submaximal contraction of four distinct movements (abduction, flexion, internal and external rotation). Shoulder strength, subjective pain experience (McGill Pain Questionnaire), and psychological status (Hospital Anxiety and Depression Scale) were also assessed. The results were compared between patient and control groups according to the gender. Despite marked fatigue observed in the majority of muscles particularly during flexion and abduction at 90°, overall results indicated a lower tendency of fatigue progression in the impingement group across the tests (p < 0.05 - p < 0.001). Shoulder Strength, pain experience, and psychological status were significantly different between the two groups (P < .05). Lower tendency to fatigue progression in the impingement group can be attributed to the presence of fear avoidance and pain-related muscle inhibition, which in turn lead to adaptations in motor programme to reduce muscle recruitment and activation. The significantly higher levels of pain experience and anxiety/depression in the impingement group further support this proposition.

POSTURAL ALTERATIONS IN PATIENTS WITH SUBACROMIAL IMPINGEMENT SYNDROME.

BACKGROUND: An aberrant upper body posture has been proposed as one of the etiological factors contributing to the development of subacromial impingement syndrome (SAIS). Clinicians have translated this supposition into assessment and rehabilitation programs despite insufficient and conflicting evidence to support this approach. PURPOSE: The purpose of this study was to compare several postural variables between the SAIS patients and asymptomatic healthy controls. STUDY DESIGN: Case-Control Study. METHODS: A total of 75 participants including 39 patients (20 females; 19 males) and 36 healthy controls (15 females; 21 males) participated in the study. Study evaluated several postural variables including forward head posture (FHP), forward shoulder posture (FSP), thoracic kyphosis index (TKI), scapular index (SI), normalized scapular protraction (NSP), and the lateral scapular slide test (LSST). The variables were compared between patient and control groups according to sex. RESULTS: Significant differences were observed in the female patients compared to asymptomatic controls for the FHP (49.38 + 9.6o vs 55.5o+8.38, p=0.03), FSP (45.58 + 10.1o vs 53.68 + 7.08, p=0.02), and LSST in third position (10.2 + 2.1cm vs 11.5 + 0.7cm, p=0.01). Male patients showed a significant difference only in the FSP compared to controls (61.9o+9.4o vs 49.78 + 9.28, p<0.001). CONCLUSIONS: While inadequate data on the relationship between dysfunctional posture and SAIS has led to broad variations in current rehabilitation strategies, the results of the present study revealed different patterns of postural aberrations in female and male patients with SAIS. This clarifies the need to develop individualized or sex-specific approaches for assessing posture in men and women with SAIS and rehabilitation programs based on the assessment results. LEVEL OF EVIDENCE: 3b.

Shoulder muscle activation and coordination in patients with a massive rotator cuff tear: an electromyographic study.

Adaptive muscle activation strategies following a massive rotator cuff tear (MRCT) are inadequately understood, and the relationship among muscles during everyday activities has not been considered. Thirteen healthy subjects comprised the control group, and 11 subjects with a MRCT the patient group. Upper limb function was assessed using the Functional Impairment test-hand, neck, shoulder, and arm (FIT-HaNSA). Electromyography (EMG) was recorded from 13 shoulder muscles, comprising five muscle groups, during a shelf-lifting task. Mean FIT-HaNSA scores were significantly lower in MRCT patients (p≤0.001), reflecting a severe functional deficit. In MRCT patients, EMG signal amplitude was significantly higher for the biceps brachii-brachioradialis (p < 0.001), upper trapezius-serratus anterior (p= 0.025), muscle groups and for the latissimus dorsi (p = 0.010), and teres major (p=0.007) muscles. No significant differences in the correlation among muscle groups were identified, pointing to an unchanged neuromuscular strategy following a tear. In MRCT patients, a reorganization of muscle activation strategy along the upper limb kinetic chain is aimed at reducing demand on the glenohumeral joint. Increased activation of the latissimus dorsi and teres major muscles is an attempt to compensate for the deficient rotator cuff. Re-education towards an alternate neuromuscular control strategy appears necessary to restore function.

Normal shoulder muscular activation and co-ordination during a shoulder elevation task based on activities of daily living: an electromyographic study.

Studies of normal shoulder function have often failed to consider the inter-relationship between different muscle groups in activities relevant to daily life. Upper limb functional status was assessed in 12 healthy male volunteers using the Functional Impairment Test-Hand, Neck, Shoulder and Arm test (FIT-HaNSA). Electromyography was then used to study the activity and coordination of 13 muscles (10 by surface electrodes, 3 by fine-wire intramuscular electrodes) around the shoulder during a dynamic movement task based on the shelf-lifting task in FIT-HaNSA. Muscles were grouped for analysis into deltoid (anterior, middle, and posterior divisions), adductors (latissimus dorsi and teres major), rotator cuff (supraspinatus, infraspinatus, and subscapularis), and elbow flexor (brachioradialis, biceps brachii) groups. There were no significant inter-session effects. Using cross-correlation analysis to investigate the whole time-course of activation, there were highly significant positive correlations (p < 0.001) between the deltoid and rotator cuff, the deltoid and adductor and the adductor and rotator cuff groups, and a significant negative correlation between the deltoid and elbow flexor groups (p = 0.031). We conclude that the deltoid, adductor, and rotator cuff muscles all contribute to the muscular component of glenohumeral joint stability. Muscular stability can be adapted as required to meet task-specific demands.

Matrix metalloproteinase expression is related to angiogenesis and histologic grade in spindle cell soft tissue neoplasms of the extremities.

We defined the immunocytochemical expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in benign soft tissue neoplasms, fibromatoses, and sarcomas, together with the activity of gelatinase MMPs and TIMPs measured by zymography and reverse zymography in a subset of cases. The most strongly expressed MMP in all tumors was MMP-1, with weaker expression of MMP-10, MMP-11, and MMP-14 in most tumors. Nuclear expression of MMP-1, MMP-8, and MMP-13 was an unusual feature. TIMP-2 was expressed in all tumors, with stronger expression in fibromatoses than in sarcomas. Fibromatoses and high-grade sarcomas showed greater MMP-1 expression than other groups, and endothelial MMP-2 expression was more extensive in sarcomas. Differences in MMP and TIMP expression might be linked to the biologic behavior of soft tissue neoplasms. The activation of endothelial MMP-2 linked to widespread MMP-14 expression provides a mechanism for sarcomas to modulate their matrix and facilitate angiogenesis.