Temporal effect of in vivo tendon fatigue loading on the apoptotic response explained in the context of number of fatigue loading cycles and initial damage parameters

Accumulation of damage is a leading factor in the development of tendinopathy. Apoptosis has been implicated in tendinopathy, but the biological mechanisms responsible for initiation and progression of these injuries are poorly understood. We assessed the relationship between initial induced damage and apoptotic activity 3 and 7 days after fatigue loading. We hypothesized that greater apoptotic activity (i) will be associated with greater induced damage and higher number of fatigue loading cycles, and (ii) will be higher at 7 than at 3 days after loading. Left patellar tendons were fatigue loaded for either 100 or 7,200 cycles. Diagnostic tests were applied before and after fatigue loading to determine the effect of fatigue loading on hysteresis, elongation, and loading and unloading stiffness (damage parameters). Cleaved Caspase‐3 staining was used to identify and calculate the percent apoptosis in the patellar tendon. While no difference in apoptotic activity occurred between the 100 and 7,200 cycle groups, greater apoptotic activity was associated with greater induced damage. Apoptotic activity was higher at 7 than 3 days after loading. We expect that the decreasing number of healthy cells that can repair the induced damage in the tendon predispose it to further injury. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1097–1103, 2014.     

The initial repair response of articular cartilage after mechanically induced damage

The regenerative potential of articular cartilage (AC) defects is limited and depends on defect size, biomechanical conditions, and age. Early events after overloading might be predictive for cartilage degeneration in the long term. Therefore, the present aim is to investigate the temporal response of cartilage to overloading at cell, matrix, and tissue level during the first period after mechanical overloading. In the present study, the effect of high loading (∼8 MPa) at a high rate (∼14 MPa/s) at day 0 during a 9 day period on collagen damage, gene expression, cell death, and biochemical composition in AC was investigated. A model system was developed which enabled culturing osteochondral explants after loading. Proteoglycan content was repeatedly monitored over time using μCT, whereas other evaluations required destructive measurements. Changes in matrix related gene expressions indicated a degenerative response during the first 6 h after loading. After 24 h, this was restored and data suggested an initial repair response. Cell death and microscopic damage increased after 24 h following loading. These degradative changes were not restored within the 9 day culture period, and were accompanied by a slight loss of proteoglycans at the articular surface that extended into the middle zones. The combined findings indicate that high magnitude loading of articular cartilage at a high rate induces an initial damage that later initiates a healing response that can probably not be retained due to loss of cell viability. Consequently, the matrix cannot be restored in the short term. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1265–1273, 2017.

     

Structural and mechanical effects of in vivo fatigue damage induction on murine tendon

The purpose of this study was to develop and validate an in vivo mouse model of tendon fatigue and use this model to investigate and quantify the physical manifestations of fatigue damage in mouse tendon. Patellar tendons of C57BL/6J mice were fatigue loaded at 2 Hz to three endpoints (4 N peak force per cycle for 1 h, 6 N for 1 h, and 4 N for 2 h), during which hysteresis, tangent stiffness, and peak strain of each cycle were measured. Damage accumulation was then quantified using in situ histology, and each tendon was loaded monotonically to failure. Histological damage increased significantly in all three groups (≥2-fold), and monotonic stiffness decreased significantly in the 6 N, 1 h and 4 N, 2-h groups (~25%), suggesting that damage initially manifests as changes to the collagen structure of the tendon and subsequently as changes to the function. For the fatigue loading protocols used in this study, none of the evaluated real-time parameters from fatigue loading correlated with damage area fraction measured structural damage or monotonic stiffness, suggesting that they are not suited to serve as proxies for damage accumulation. In future studies, this model will be used to compare the biological response of mouse tendon to fatigue damage across genetic strains.     

Distributing a fixed amount of cyclic loading to tendon explants over longer periods induces greater cellular and mechanical responses.

Tendon overuse injuries are a major source of clinical concern. Cyclic loading causes material damage and induces biochemical responses in tendon. The purpose of this study was to examine the biochemical and biomechanical tendon response after applying cyclical loading over varying durations. Avian flexor digitorum profundus tendons were loaded (3 or 12 MPa) to a fixed number of cycles across either 1 or 12 days in vitro. The tendon response evaluations included biomechanical data gathered during loading and subsequent failure testing. Evaluations also included cellular viability, cell death, and proteoglycan, collagen, collagenase, and prostaglandin E(2) (PGE(2)) content measurements obtained from tissue specimens and media samples. Significant strains (up to 2%) accumulated during loading. Loading to 12 MPa significantly reduced maximum stress (33% and 27%) and energy density (42% and 50%) when applied across 1 or 12 days, respectively. Loading to 3 MPa also caused a 40% reduction in energy density, but only when applied across 12 days. Cell death and collagenase activity increased significantly with increasing magnitude and duration. However, no differences occurred in cell viability or collagen content. Glycosaminoglycan content increased 50% with load magnitude, while PGE(2) production increased 2.5-fold with loading magnitude and 11-fold with increased duration. Mechanical fatigue-induced mechanical property changes were exhibited by the tendons in response to increased loading magnitude across just 1 day. However, when the same loading was applied over a longer period, most outcomes were magnified substantially, relative to the short duration regimens. This is presumably due to the increased response time for the complex cellular response to loading. A key contributor may be the inflammatory mediator, PGE(2), which exhibited large magnitude and duration dependent increases to cyclic loading.     

The role of the macrophage in tendinopathy and tendon healing

The role of the macrophage is an area of emerging interest in tendinopathy and tendon healing. The macrophage has been found to play a key role in regulating the healing process of the healing tendon. The specific function of the macrophage depends on its functional phenotype. While the M1 macrophage phenotype exhibits a phagocytic and proinflammatory function, the M2 macrophage phenotype is associated with the resolution of inflammation and tissue deposition. Several studies have been conducted on animal models looking at enhancing or suppressing macrophage function, targeting specific phenotypes. These studies include the use of exogenous biological and pharmacological substances and more recently the use of transgenic and genetically modified animals. The outcomes of these studies have been promising. In particular, enhancement of M2 macrophage activity in the healing tendon of animal models have shown decreased scar formation, accelerated healing, decreased inflammation and even enhanced biomechanical strength. Currently our understanding of the role of the macrophage in tendinopathy and tendon healing is limited. Furthermore, the roles of therapies targeting macrophages to enhance tendon healing is unclear. Clinical Significance: An increased understanding of the significance of the macrophage and its functional phenotypes in the healing tendon may be the key to enhancing tendon healing. This review will present the current literature on the function of macrophages in tendinopathy and tendon healing and the potential of therapies targeting macrophages to enhance tendon healing.     

Production of PGE2 increases in tendons subjected to repetitive mechanical loading and induces differentiation of tendon stem cells into non‐tenocytes

Whether tendon inflammation is involved in the development of tendinopathy or degenerative changes of the tendon remains a matter of debate. We explored this question by performing animal and cell culture experiments to determine the production and effects of PGE₂, a major inflammatory mediator in tendons. Mouse tendons were subjected to repetitive mechanical loading via treadmill running, and the effect of PGE₂ on proliferation and differentiation of tendon stem cells (TSCs) was assessed in vitro. Compared to levels in cage control mice, PGE₂ levels in mouse patellar and Achilles tendons were markedly increased in response to a bout of rigorous treadmill running. PGE₂ treatment of TSCs in culture decreased cell proliferation and induced both adipogenesis and osteogenesis of TSCs, as evidenced by accumulation of lipid droplets and calcium deposits, respectively. Effects of PGE₂ on both TSC proliferation and differentiation were apparently PGE₂–dose‐dependent. These findings suggest that high levels of PGE₂, which are present in tendons subjected to repetitive mechanical loading conditions in vivo as shown in this study, may result in degenerative changes of the tendon by decreasing proliferation of TSCs in tendons and also inducing differentiation of TSCs into adipocytes and osteocytes. The consequences of this PGE₂ effect on TSCs is the reduction of the pool of tenocytes for repair of tendons injured by mechanical loading, and production of fatty and calcified tissues within the tendon, often seen at the later stages of tendinopathy. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:198–203, 2010     

The role of interstitial fluid flow in the remodeling response to fatigue loading.

Load-induced fluid flow enhances molecular transport through bone tissue and relates to areas of bone resorption and apposition. Remodeling activity is highly coordinated and necessitates a means for cellular communication via intracellular and extracellular means. Osteocytes, osteoblasts, and osteoclasts, which reside in disparate locations within the tissue, communicate intracellularly via the cellular syncytium and extracellularly via the pericellular fluid space of the lacunocanalicular system. Both of these communications systems are physically disrupted by microdamage incurred during fatigue loading of bone. The purpose of this study was to develop an analytical model to understand the role of interstitial fluid flow in the remodeling response to fatigue loading. Adequate transport was assumed a prerequisite for maintenance of cell viability in bone. Diffusive and convective transport were simulated through the lacunocanalicular network in a healthy undamaged state as well as in a damaged state after fatigue loading. The model predicts that fatigue damage impedes transport from the blood supply, depleting the concentration of molecular entities in and downstream from areas of damage. Furthermore, the presence of microcracks alters the distribution of molecular entities between individual lacunae. These effects were confirmed by the results of an in vivo pilot study in which fluorescent, flow-visualizing agents pooled within microcracks and were absent from areas surrounding microcracks, corresponding to areas deprived of fluid flow. Loss of osteocyte viability is coupled to targeting and initiation of new remodeling activity. Taken as a whole, these data suggest a link between interstitial fluid flow, mass transport, maintenance of osteocyte viability, and modulation of remodeling activity.     

卵裂球受损对冻融胚胎移植周期临床结局的影响

目的探讨卵裂球受损对冻融胚胎移植周期临床结局的影响。方法回顾分析181个冻融胚胎移植周期,了解胚胎级别与冷冻受损的关系,分析受损程度(完整存活、受损率≤20%和>20%组)和移植平均≥6、<6细胞胚胎对妊娠率和种植率的影响。结果1、2、3级胚胎存活率差异显著(P<0.05),损伤率差异非常显著(P<0.001)。完整存活胚胎、受损率≤20%及>20%胚胎组的临床妊娠率、种植率均存在显著性差异(P<0.05,P<0.01),移植≥6细胞胚胎组的临床妊娠率、种植率显著高于<6细胞胚胎组(P<0.05)。结论胚胎卵裂球受损率评价胚胎受损程度优于胚胎存活率。在冻融胚胎移植周期,胚胎质量、移植时胚胎细胞期和受损程度是影响妊娠率和种植率的胚胎学因素。     

Simultaneous rupture of the quadriceps tendon and contralateral patellar tendon in a patient with chronic renal failure

Simultaneous quadriceps and patellar tendon rupture is rare. Mechanical factors and coexisting systemic and local factors are taken into consideration in the pathogenesis of these ruptures. In patients with some chronic systemic diseases, simultaneous rupture can occur spontaneously or with minor traumas. We present a case of simultaneous quadriceps and patellar tendon rupture in a 21-year-old man with chronic renal failure in this report. He was treated surgically by osseotendinous repair with suture anchors and supplemental cerclage wire fixation on both sides. He regained his normal knee joint functions 18 months after the operation.This study was conducted at Ankara Numune Education and Research Hospital, Third Orthopedics and Traumatology Clinic, Talatbaa Bulvar, Shhiye, Ankara, Turkey     

Cell kinetics of the initial response to orthodontically induced osteogenesis in rat molar periodontal ligament

Summary Initial cell kinetics (15 min-20 h) of mechanically induced osteogenesis was studied with³H-thymidine (³H-Tdr) autoradiography. Continuous orthodontic force elicited a three-stage cell proliferative reaction within rat molar periodontal ligament (PDL): (a) brief, generalized response, characterized by a burst of mitotic activity from 75 min to 2 h and a cyclic change (decrease, increase and return to control levels) in percent labeled cells from 15 min to 2 h; (b) transient, generalized response, involving an increased mitotic index (MI) from 2 to 13 h, associated with elevated labeling index (LI) from 6 to 12 h; and (c) sustained, localized osteogenic response (12 h to several days) in which cell proliferation was confined primarily to the immediate area of new bone formation. These results suggest that the initial effect of mechanical perturbation on relatively quiescent adult bone progenitor cells is a generalized, transient release of G₁ and G₂ blocked cells. This response may correlate clinically with the regional acceleration phenomenon (RAP), which follows bone trauma, fracture, or surgical injury.     

尿毒症并发一侧髌韧带断裂对侧股四头肌肌腱断裂1例

正患者,男,36岁,主因摔倒致双侧膝关节疼痛伴活动受限18 d于2016年1月22日入院。现病史:患者18 d前因摔倒致双侧膝关节疼痛,当即感疼痛难忍,不能独立行走,自诉听到双侧膝关节"咔"的一声,曾服止痛药物治疗,效果不佳,为进一步诊治收入我科。自发病以来睡眠、大小便、体重等基本无变化。查体:慢性肾病面容。双侧膝关节肿胀明显,皮温皮色正常,无窦道及瘢痕,左侧髌骨下方及右侧髌骨上方触之有明显的空虚感,双膝压痛(+),双膝浮髌     

The effect of interbody fusion cage design on the stability of the instrumented spine in response to cyclic loading: an experimental study

Background Context

In the lumbar spine, end plate preparation for the interbody fusion cages may critically affect the cage's long-term performance. This study investigated the effect of the interbody cage design on the compliance and cage subsidence of instrumented spines under cyclic compression.

In situ deflection of tendon cell‐cilia in response to tensile loading: an in vitro study

To determine if a correlation exists between tensile loading and the deflection of tendon cell‐cilia in situ, rat‐tail tendon fascicles were stained for tubulin and mounted in a loading device attached to the stage of a confocal microscope. Individual tendon cells (n = 13) were identified and sequential images taken at 0%, 2%, 4%, 6%, and 8% grip to grip strain. The change in ciliary deflection angle was then measured at each strain level. To determine the ability of cilia to return to their original orientation, additional fascicles were loaded to 6% strain and then unloaded to 0% and tendon cell‐ciliary (n = 10) deflection angle measured. There was a weak (r² = 0.40) but significant (p < 0.0001) correlation between the change in deflection angle and applied strain. Tensile loading produced a change in deflection angle from 0% to 3% (p = 0.039) and from 3% to 6% (p = 0.001) strain. There was no change (p = 1.000) in deflection angle from 6% to 8% strain. Reducing the strain from 6% to 0% resulted in a change (p = 0.048) in angle towards the pre‐load position. However, the angle did not return to the pre‐strain position (p = 0.025). These results demonstrate that tensile loading produces in situ deflection of tendon cell‐cilia and supports the concept that cilia are involved in the mechanotransduction response of tendon cells. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:925–930     

Cyclin E影响DNA损伤药物对乳腺癌细胞的治疗敏感性

目的探讨Cyclin E的表达水平对乳腺癌细胞化疗敏感性的影响。方法Westem blot检测不同乳腺癌细胞系中Cyclin E表达水平的差异;RNAi技术敲低Cyclin E在乳腺癌细胞中的表达水平后,用流式细胞术和细胞衰老相关β-gal染色检测Cyclin E敲低前后乳腺癌细胞对DNA损伤药物的反应敏感性。结果乳腺癌中Cyclin E的表达与ER状态没有直接关系,DNA损伤药物阿霉素可以明显导致Cyclin E表达敲低MDA-MB-435细胞的G_1期阻滞,进而减少进入S期的细胞数,Cyclin E表达较高的MDA-MB-435细胞处于G_1期的细胞百分比为56.21%,S期的细胞百分比为15.68%;而Cyclin E表达敲低的MDA-MB-435细胞处于G_1期的细胞百分比为65.85%,S期的细胞百分比为11.91%。同时,阿霉素导致Cyclin E表达敲低MDA-MB-435的衰老细胞增加,减少有增殖能力的细胞数,有效的控制细胞分裂和增殖。结论Cyclin E可能作为一种独立的预后因素评估乳腺癌的化疗敏感性。     

精液常规参数初检合格后不同时间点再分析结果及其与精子DNA损伤的关系

目的:探讨完全液化且常规参数初检合格的精液标本,于不同时间再分析的结果差异,及精子DNA碎片化指数(DFI)与精子活动力改变的相关性。方法:选取127份符合纳入标准的精液标本,分别于取样后15、30、60 min时采用计算机辅助精液分析(CASA)系统进行分析。精子形态分析采用Shorr染色法,吖啶橙试验(AOT)检测DFI。结果:3个时间点精子浓度、a级和b级精子百分率均无统计学差异(P>0.05)。取样15 min时a+b和a+b+c级精子百分率显著高于30和60 min时的结果(P<0.05),后两者间无统计学差异(P>0.05)。不同时间精子活动力各项指标中,至少有1项由"正常组"变为"异常组"的发生率为25.2%,两组间DFI和形态学无统计学差异(P>0.05)。取样后15到60 min变化的精子活动力指标中,a、a+b、a+b+c级下降值与DFI存在正相关性(P<0.05)。结论:取样后15 min内完全液化并初查参数合格的精液标本,30～60 min内复查时,a级和b级精子百分率波动并无显著差异,而a+b级及a+b+c级精子则可能显著下降,精子活动力指标可能出现异常。故应至少进行2次精液分析,综合评估生育力。如2次结果差异较大,尤其是a级精子百分率下降幅度较大,则可能与精子DNA损伤有关,需进一步行精子DNA损伤检测。     

Use of tactile stiffness to detect fatigue in the latissimus dorsi muscle

In clinical settings, no method has been established to examine the fatigue of a latissimus dorsi muscle (LDM) preconditioned for cardiomyoplasty. We examined the feasibility of measuring muscle stiffness (tactile stiffness) to evaluate muscle fatigue in situ using our tactile sensor. We stimulated canine LDM with burst pacing and monitored both stiffness and tension to determine their relationship. In both dissected LDM and LDM in situ, the decrements of these parameters during burst pacing were compared between preconditioned and unconditioned LDM. In measurement in situ, the sensor probe was placed on the LDM through a small incision. Strong statistical correlation was shown between stiffness and tension (r = 0.935). In decrements of stiffness in situ, there were statistically significant differences between preconditioned and unconditioned LDM. Our tactile sensor system can provide an efficient method for evaluating fatigue of muscles in situ without measuring muscle tension.