MicroRNA Profiling Reveals Distinct Signatures in Degenerative Rotator Cuff Pathologies

MicroRNAs (miRNAs) have emerged as key regulators orchestrating a wide range of inflammatory and fibrotic diseases. However, the role of miRNAs in degenerative shoulder joint disorders is poorly understood. The aim of this explorative case-control study was to identify pathology-related, circulating miRNAs in patients with chronic rotator cuff tendinopathy and degenerative rotator cuff tears (RCT). In 2017, 15 patients were prospectively enrolled and assigned to three groups based on the diagnosed pathology: (i) no shoulder pathology, (ii) chronic rotator cuff tendinopathy, and (iii) degenerative RCTs. In total, 14 patients were included. Venous blood samples (“liquid biopsies”) were collected from each patient and serum levels of 187 miRNAs were determined. Subsequently, the change in expression of nine candidate miRNAs was verified in tendon biopsy samples, collected from patients who underwent arthroscopic shoulder surgery between 2015 and 2018. Overall, we identified several miRNAs to be progressively deregulated in sera from patients with either chronic rotator cuff tendinopathy or degenerative RCTs. Importantly, for the several of these miRNAs candidates repression was also evident in tendon biopsies harvested from patients who were treated for a supraspinatus tendon tear. As similar expression profiles were determined for tendon samples, the newly identified systemic miRNA signature has potential as novel diagnostic or prognostic biomarkers for degenerative rotator cuff pathologies. © 2019 The Authors. Journal of Orthopaedic Research^® published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. Inc. J Orthop Res 38:202–211, 2020     

Implant surface alters compartmental-specific contributions to fixation strength in rats

Mechanical fixation of the implant to host bone is an important contributor to orthopedic implant survivorship. The relative importance of bone-implant contact, trabecular bone architecture, and cortical bone geometry to implant fixation strength has never been directly tested, especially in the settings of differential implant surface properties. Thus, using a rat model where titanium rods were placed into the intramedullary canal of the distal femur, we determined the relative contribution of bone-implant contact and peri-implant bone architecture to the fixation strength in implants with different surface roughness: highly polished and smooth (as-received) and dual acid-etched (DAE) implants. Using a training set that maximized variance in implant fixation strength, we initially examined correlation between implant fixation strength and outcome parameters from microcomputed tomography and found that osseointegration volume per total volume (OV/TV), trabecular bone volume per total volume (BV/TV), and cortical thickness (Ct.Th) were the single best compartment-specific predictors of fixation strength. We defined separate regression models to predict implant fixation strength for as-received and DAE implants. When the training set models were applied to independent validation sets, we found strong correlations between predicted and experimentally measured implant fixation strength, with r² = .843 in as received and r² = .825 in DAE implants. Interestingly, for as-received implants, OV/TV explained more of the total variance in implant fixation strength than the other variables, whereas in DAE implants, Ct.Th had the most explanatory power, suggesting that surface topography of implants affects which bone compartment is most important in providing implant fixation strength.     

Is Use of BMP-2 Associated with Tumor Growth and Osteoblastic Differentiation in Murine Models of Osteosarcoma?

BackgroundThe putative benefit of rhBMP-2 is in the setting of limb reconstruction using structural allografts, whether it be allograft-prosthetic composites, osteoarticular allografts, or intercalary segmental grafts. There are also potential advantages in augmenting osseointegration of uncemented endoprosthetics and in reducing infection. Recombinant human BMP-2 might mitigate nonunion in structural allograft augmented osteosarcoma limb salvage surgery; however, its use is limited because of concerns about the prooncogenic effects of the agent.Questions/purposes(1) To assess if BMP-2 signaling influences osteosarcoma cell line growth. (2) To characterize degree of osteosarcoma cell line osteoblastic differentiation in response to BMP-2. (3) To assess if BMP-2 signaling has a consistent effect on local or systemic tumor burden in various orthotopic murine models of osteosarcoma.MethodsIn this study, 143b, SaOS-2 and DLM8-M1 osteosarcoma cell lines were transfected with BMP-2 cDNA controlled by a constitutive promoter (experimental) or an empty vector (control) using a PiggyBac transposon system. Cellular proliferation was assessed using a quantitative MTT colorimetric assay. Osteoblastic differentiation was compared between control and experimental cell lines using quantitative real-time polymerase chain reaction of the osteoblastic markers connective tissue growth factor, Runx-2, Osterix, alkaline phosphatase and osteocalcin. Experimental and control cell lines were injected into the proximal tibia of either NOD-SCID (143b and SaOS-2 xenograft model), or C3H (DLM8-M1 syngeneic model) mice. Local tumor burden was quantitatively assessed using tumor volume caliper measurements and bioluminescence, and qualitatively assessed using post-mortem ex vivo microCT. Lung metastasis was qualitatively assessed by the presence of bioluminescence, and incidence was confirmed using histology. rhBMP-2 soaked absorbable collagen sponges (experimental) and sterile-H₂O soaked absorbable collagen sponges (control) were implanted adjacent to 143b proximal tibial cell line injections to compare the effects of exogenous BMP-2 application with endogenous upregulation.ResultsConstitutive expression of BMP-2 increased the in vitro proliferation of 143b cells (absorbance values 1.2 ± 0.1 versus 0.89 ± 0.1, mean difference 0.36 [95% CI 0.12 to 0.6]; p = 0.01), but had no effect on SaOS-2 and DLM8-M1 cell proliferation. In response to constitutive BMP-2 expression, 143b cells had no differences in osteoblastic differentiation, while DLM8-M1 cells downregulated the early marker connective tissue growth factor (mean ΔCt 0.2 ± 0.1 versus 0.6 ± 0.1; p = 0.002) and upregulated the early-mid range marker Runx-2 (mean ΔCt -0.8 ± 0.1 versus -1.1 ± 0.1; p = 0.002), and SaOS-2 cells upregulated the mid-range marker Osterix (mean ΔCt -2.1 ± 0.6 versus -3.9 ± 0.6; p = 0.002). Constitutive expression of BMP-2 resulted in greater 143b and DLM8-M1 local tumor volume (143b: 307.2 ± 106.8 mm³ versus 1316 ± 387.4 mm³, mean difference 1009 mm³ [95% CI 674.5 to 1343]; p < 0.001, DLM8-M1 week four: 0 mm³ versus 326.1 ± 72.8 mm³, mean difference 326.1 mm³ [95% CI 121.2 to 531]; p = 0.009), but modestly reduced local tumor growth in SaOS-2 (9.5 x 10⁸ ± 8.3x10⁸ photons/s versus 9.3 x 10⁷ ± 1.5 x 10⁸ photons/s, mean difference 8.6 x 10⁸ photons/s [95% CI 5.1 x 10⁸ to 1.2 x 10⁹]; p < 0.001). Application of exogenous rhBMP-2 also increased 143b local tumor volume (495 ± 91.9 mm³ versus 1335 ± 102.7 mm³, mean difference 840.3 mm³ [95% CI 671.7 to 1009]; p < 0.001). Incidence of lung metastases was not different between experimental or control groups for all experimental conditions.ConclusionsAs demonstrated by others, ectopic BMP-2 signaling has unpredictable effects on local tumor proliferation in murine models of osteosarcoma and does not consistently result in osteosarcoma cell line differentiation. Further investigations into other methods of safe bone and soft tissue healing augmentation and the use of differentiation therapies is warranted.Clinical RelevanceOur results indicate that BMP-2 has the potential to stimulate the growth of osteosarcoma cells that are poorly responsive to BMP-2 mediated osteoblastic differentiation. As this differentiation potential is unpredictable in the clinical setting, BMP-2 may promote the growth of microscopic residual tumor burden after resection. Our study provides further support for the recommendation to avoid the use of BMP-2 after limb-salvage surgery in patients with osteosarcoma.     

Rapid screening test for primary hyperaldosteronism: ratio of plasma aldosterone to renin concentration determined by fully automated chemiluminescence immunoassays

BACKGROUND: The ratio of plasma aldosterone concentration to plasma renin activity (PAC/PRA) is the most common screening test for primary hyperaldosteronism (PHA), but it is not standardized among laboratories. We evaluated new automated assays for the simultaneous measurement of PAC and plasma renin concentration (PRC). METHODS: We studied 76 healthy normotensive volunteers and 28 patients with confirmed PHA. PAC and PRC were measured immunochemically in EDTA plasma on the Nichols Advantage chemiluminescence analyzer, and PRA was determined by an activity assay. RESULTS: In volunteers, PAC varied from 33.3 to 1930 pmol/L, PRA from 1.13 to 19.7 ng.mL(-1).h(-1) (0.215 ng.mL(-1).h(-1) = 1 pmol.L(-1).s(-1)), and PRC from 5.70 to 116 mU/L. PAC/PRA ratios ranged from 4.35 to 494 (pmol/L)/(ng.mL(-1).h(-1)) and PAC/PRC ratios from 0.69 to 71.0 pmol/mU. In PHA patients, PAC ranged from 158 to 5012 pmol/L, PRA from 0.40 to 1.70 ng.mL(-1).h(-1), and PRC from 0.80 to 11.7 mU/L. PAC/PRA ratios were between 298 and 6756 (pmol/L)/(ng.mL(-1).h(-1)) and PAC/PRC ratios between 105 and 2328 pmol/mU. Whereas PAC or PRC showed broad overlap between PHA patients and volunteers, the PAC/PRC ratio indicated distinct discrimination of these two groups at a cutoff of 71 pmol/mU. CONCLUSION: The PAC/PRC ratio offers several practical advantages compared with the PAC/PRA screening method. The present study offers preliminary evidence that it may be a useful screening test for PHA. Further studies are required to validate these results, especially in hypertensive cohorts.     

国际医学杂志编辑委员会关于临床试验注册的声明

无私和信任是人体试验研究的核心。自愿参加研究的受试者是无私的,因为他们相信自己的参与将为他人健康的改善做出贡献,也相信研究者会尽最大努力减少受试者的风险。由于受试者的无私和信任,临床研究才成为可能,因此研究机构有义务使临床研究的实施符合伦理学要求,并诚实地报道研究结果。而诚实的报道首先需要知道所有现存的临床研究,即使其结果对赞助方不利。     

Distribution of Matrix Metalloproteinases in Human Atherosclerotic Carotid Plaques and Their Production by Smooth Muscle Cells and Macrophage Subsets

Purpose

In this study, the potential of matrix metalloproteinase (MMP) sense for detection of atherosclerotic plaque instability was explored. Secondly, expression of MMPs by macrophage subtypes and smooth muscle cells (SMCs) was investigated.

Procedures

Twenty-three consecutive plaques removed during carotid endarterectomy were incubated in MMPSense? 680 and imaged with IVIS® Spectrum. mRNA levels of MMPs, macrophage markers, and SMCs were determined in plaque specimens, and in in vitro differentiated M1 and M2 macrophages.

Results

There was a significant difference between autofluorescence signals and MMPSense signals, both on the intraluminal and extraluminal sides of plaques. MMP-9 and CD68 messenger RNA (mRNA) expression was higher in hot spots, whereas MMP-2 and αSMA expression was higher in cold spots. In vitro M2 macrophages had higher mRNA expression of MMP-1, MMP-9, MMP-12, and TIMP-1 compared to M1 macrophages.

Conclusion

MMP-9 is most dominantly MMP present in atherosclerotic plaques and is produced by M2 rather than M1 macrophages.

     

Decreased posterior cruciate and altered collateral ligament loading following ACL transection: A longitudinal study in the ovine model

Although ACL deficiency is shown to lead to joint degeneration, few quantitative data are reported on its effect on soft tissue structures surrounding the knee joint, specifically, the posterior cruciate and collateral ligaments. The kinematics of the stifle joint of sheep (N = 5) were measured during “normal” gait, as well as 4 and 20 weeks after ACL transection. These motions were reproduced using a unique robotic manipulator and the loads borne by PCL, MCL, and LCL during gait were determined. Our results demonstrated a significant decrease in mean PCL loads 20 weeks post‐ACL injury, at hoof‐strike (0% of gait, p = 0.034), hoof‐off (66% of gait, p = 0.006), peak‐swing (85% of gait, p = 0.026), and extension‐before‐hoof‐strike (95% of gait, p = 0.028). Mean MCL loads did not significantly increase following ACL transection, maybe due to large between‐animal variation. Finally, mean LCL loads indicated a significant decrease (p < 0.047) at 20 weeks across the entire gait cycle. From a clinical perspective, the load redistributions observed in cruciate and collateral ligaments following ACL injury indicate that these tissues can carry/adapt to the altered mechanical environment of the joint. The considerable variability in the magnitudes of change following ACL injury among animals also simulates clinical variability in humans after trauma. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:431–438, 2014.