Molecular Mechanisms of Fetal Tendon Regeneration Versus Adult Fibrous Repair

Tendinopathies are painful, disabling conditions that afflict 25% of the adult human population. Filling an unmet need for realistic large-animal models, we here present an ovine model of tendon injury for the comparative study of adult scarring repair and fetal regeneration. Complete regeneration of the fetal tendon within 28 days is demonstrated, while adult tendon defects remained macroscopically and histologically evident five months post-injury. In addition to a comprehensive histological assessment, proteome analyses of secretomes were performed. Confirming histological data, a specific and pronounced inflammation accompanied by activation of neutrophils in adult tendon defects was observed, corroborated by the significant up-regulation of pro-inflammatory factors, neutrophil attracting chemokines, the release of potentially tissue-damaging antimicrobial and extracellular matrix-degrading enzymes, and a response to oxidative stress. In contrast, secreted proteins of injured fetal tendons included proteins initiating the resolution of inflammation or promoting functional extracellular matrix production. These results demonstrate the power and relevance of our novel ovine fetal tendon regeneration model, which thus promises to accelerate research in the field. First insights from the model already support our molecular understanding of successful fetal tendon healing processes and may guide improved therapeutic strategies.

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Abstract MP254: Decellularized Extracellular Matrix Hydrogel Lowers Fibrosis And Fibroblast Differentiation In Post-injury Mice Hearts Through Capg

Circulation Research ◽

10.1161/res.129.suppl_1.mp254 ◽

2021 ◽

Vol 129 (Suppl_1) ◽

Author(s):

Xinming Wang ◽

Samuel Senyo

Keyword(s):

Extracellular Matrix ◽

Molecular Mechanisms ◽

Smooth Muscle Actin ◽

Growth Factor Receptor ◽

Inhibitory Effect ◽

Extracellular Proteins ◽

Collagen Deposition ◽

Post Injury ◽

Capping Protein ◽

Decellularized Extracellular Matrix

Hypothesis and objective: We hypothesize that transplantation of decellularized cardiac extracellular matrix (dECM) lowers fibrosis and fibroblast differentiation. In this study we investigated collagen deposition and fibroblast differentiation in post-MI hearts and heart explants of various stiffness after dECM hydrogel treatments. The objectives are 1) determining if dECM derived from fetal and adult porcine hearts reduces fibrosis in injured hearts; and 2) identifying specific signaling pathways that regulate fibroblasts differentiation induced by extracellular proteins. Methods: Porcine dECM was injected immediately after ligating coronary artery in P1 mice. Histology was conducted on day 7 post-myocardial infarction (MI). A mice ventricle explant model was used to investigate the molecular mechanisms. Results: We observed that fetal dECM treatment lowered fibrosis and fibroblast differentiation in post-MI hearts (Fig.1). Fibroblast differentiation as indicated by α-smooth muscle actin expression in vimentin or platelet derived growth factor receptor α positive cells showed an inhibitory effect of fetal dECM on fibroblast differentiation. Using a heart explant model of modulated microenvironment stiffness, we demonstrated that increasing tissue stiffness stimulates fibroblast differentiation and collagen deposition. Fetal dECM treatment, however, inhibited fibroblast differentiation induced by increasing microenvironment stiffness. Transcriptome analysis revealed that two cytoskeleton-related genes, macrophage capping protein (CAPG) and leupaxin (LPXN), are modulated by dECM treatments. Using cytoskeleton polymerization modulators and siRNA, we demonstrated that fetal dECM lowers fibroblast differentiation through CAPG.

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R432 – Gene Expression of the Remodeling Rat Vocal Fold Wound

Otolaryngology ◽

10.1016/j.otohns.2008.05.587 ◽

2008 ◽

Vol 139 (2_suppl) ◽

pp. P189-P189

Author(s):

Tsunehisa Ohno ◽

Lesley C. French ◽

Bernard Rousseau

Keyword(s):

Gene Expression ◽

Extracellular Matrix ◽

Messenger Rna ◽

Vocal Fold ◽

Molecular Mechanisms ◽

Type I ◽

Post Injury ◽

Time Points ◽

Procollagen Type ◽

Procollagen Type I

Problem The authors investigated the expression of key extracellular matrix genes after vocal fold wounding in a rat model to better understand the reparative mechanisms of tissue repair during the remodeling phase of vocal fold injury. Methods Bilateral vocal fold wounds were created in 30 rats. Injured vocal fold specimens were harvested 1, 3, 7, 14, 28, and 56 days after wounding. 5 unwounded rats were used to establish baseline for polymerase chain reaction (PCR). The authors used real-time PCR to quantify messenger RNA expression of procollagen type I, III, interleukin-1 beta (IL-1 beta), decorin, and hyaluronan synthase (HAS) −1, −2, and −3. Analysis of variance was used to detect main effects for gene expression. Post-hoc tests were used to make comparisons between time points. Results Procollagen type I expression was decreased from baseline on post-injury day 1, 28, and 56. Procollagen type III was decreased on post-injury day 1 and 56, and increased from baseline on post-injury day 14. IL-1 beta expression was increased from baseline on post-injury day 1, 3, and 7. Decorin expression was decreased from baseline on post-injury day 1, 3, 7, and 56. HAS-1 expression was decreased from baseline at all post-injury time points. HAS-2 expression was increased from baseline on post-injury day 3, and decreased from baseline on post-injury day 14, 28, and 56. HAS-3 expression was decreased from baseline on post-injury day 1, 28, and 56. Conclusion Findings provide temporal changes in the expression of key extracellular matrix genes during a remodeling phase of vocal fold injury in a rat wound model. Significance Vocal fold wound models provide a means for investigating tissue reparative processes and molecular mechanisms controlling synthesis and degradation of the vocal fold extracellular matrix. Support Vanderbilt University Medical Center.

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Pifithrin-α, an inhibitor of p53 transactivation, alters the inflammatory process and delays tendon healing following acute injury

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00411.2005 ◽

2007 ◽

Vol 292 (1) ◽

pp. R321-R327 ◽

Cited By ~ 11

Author(s):

David Marsolais ◽

Claude H. Côté ◽

Jérôme Frenette

Keyword(s):

Extracellular Matrix ◽

Inflammatory Process ◽

Tendon Repair ◽

Tendon Healing ◽

Tendon Injury ◽

Acute Injury ◽

Inflammatory Phase ◽

Load To Failure ◽

Transcription Factor P53

Transcription factor p53, which was initially associated with cancer, has now emerged as an important regulator of inflammation and extracellular matrix homeostasis, two processes highly relevant to tendon repair. The goal of this study was to evaluate the effect of a p53 transactivation inhibitor, namely, pifithrin-α, on the pathophysiological sequence following collagenase-induced tendon injury. Administration of pifithrin-α during the inflammatory phase reduced the accumulation of neutrophils and macrophages by 30 and 40%, respectively, on day 3 postinjury. Pifithrin-α failed to reduce the percentage of apoptotic cells following collagenase injection but delayed functional recovery. In uninjured Achilles tendons, pifithrin-α increased metalloprotease activity 2.4-fold. Accordingly, pifithrin-α reduced the collagen content in intact tendons as well as in injured tendons 7 days posttrauma compared with placebo. The effect of pifithrin-α on load to failure and stiffness was also evaluated. The administration of pifithrin-α during the inflammatory phase did not significantly decrease the functional deficit 3 days posttrauma. More importantly, load to failure and stiffness were significantly decreased in the pifithrin-α group from day 7 to day 28 compared with placebo. Overall, our results suggest that administration of pifithrin-α alters the inflammatory process and delays tendon healing. The present findings also support the concept that p53 can regulate extracellular matrix homeostasis in vivo.

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Interplay of Forces and the Immune Response for Functional Tendon Regeneration

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.657621 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yuwei Yang ◽

Yicong Wu ◽

Ke Zhou ◽

Dongmei Wu ◽

Xudong Yao ◽

...

Keyword(s):

Immune Response ◽

Tensile Strength ◽

Tendon Healing ◽

Biomechanical Properties ◽

Tendon Injury ◽

Rapid Decline ◽

Sports Activity ◽

Tendon Regeneration ◽

Functional Regeneration ◽

External Forces

Tendon injury commonly occurs during sports activity, which may cause interruption or rapid decline in athletic career. Tensile strength, as one aspect of tendon biomechanical properties, is the main parameter of tendon function. Tendon injury will induce an immune response and cause the loss of tensile strength. Regulation of mechanical forces during tendon healing also changes immune response to improve regeneration. Here, the effects of internal/external forces and immune response on tendon regeneration are reviewed. The interaction between immune response and internal/external forces during tendon regeneration is critically examined and compared, in relation to other tissues. In conclusion, it is essential to maintain a fine balance between internal/external forces and immune response, to optimize tendon functional regeneration.

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CD146 Delineates an Interfascicular Cell Sub-Population in Tendon That Is Recruited during Injury through Its Ligand Laminin-α4

International Journal of Molecular Sciences ◽

10.3390/ijms22189729 ◽

2021 ◽

Vol 22 (18) ◽

pp. 9729

Author(s):

Neil Marr ◽

Richard Meeson ◽

Elizabeth F. Kelly ◽

Yongxiang Fang ◽

Mandy J. Peffers ◽

...

Keyword(s):

Expression Patterns ◽

Tendon Repair ◽

Tendon Healing ◽

Tendon Injury ◽

Focal Lesion ◽

Post Injury ◽

Surface Receptors ◽

Basement Membrane Protein ◽

Injury Model ◽

Cell Niche

The interfascicular matrix (IFM) binds tendon fascicles and contains a population of morphologically distinct cells. However, the role of IFM-localised cell populations in tendon repair remains to be determined. The basement membrane protein laminin-α4 also localises to the IFM. Laminin-α4 is a ligand for several cell surface receptors, including CD146, a marker of pericyte and progenitor cells. We used a needle injury model in the rat Achilles tendon to test the hypothesis that the IFM is a niche for CD146+ cells that are mobilised in response to tendon damage. We also aimed to establish how expression patterns of circulating non-coding RNAs alter with tendon injury and identify potential RNA-based markers of tendon disease. The results demonstrate the formation of a focal lesion at the injury site, which increased in size and cellularity for up to 21 days post injury. In healthy tendon, CD146+ cells localised to the IFM, compared with injury, where CD146+ cells migrated towards the lesion at days 4 and 7, and populated the lesion 21 days post injury. This was accompanied by increased laminin-α4, suggesting that laminin-α4 facilitates CD146+ cell recruitment at injury sites. We also identified a panel of circulating microRNAs that are dysregulated with tendon injury. We propose that the IFM cell niche mediates the intrinsic response to injury, whereby an injury stimulus induces CD146+ cell migration. Further work is required to fully characterise CD146+ subpopulations within the IFM and establish their precise roles during tendon healing.

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Tyrosine-Derived Polycarbonate Nerve Guidance Tubes Elicit Pro-Regenerative Extracellular Matrix Deposition When Used to Bridge Segmental Nerve Defects in Swine

10.1101/2020.01.11.902007 ◽

2020 ◽

Author(s):

JC Burrell ◽

D Bhatnagar ◽

DP Brown ◽

NS Murthy ◽

J Dutton ◽

...

Keyword(s):

Extracellular Matrix ◽

Schwann Cell ◽

Cell Infiltration ◽

Large Animal ◽

Matrix Deposition ◽

Regenerative Response ◽

Large Animal Models ◽

Segmental Nerve ◽

Collagen Iii ◽

Wide Range

AbstractPromising biomaterials should be tested in appropriate large animal models that recapitulate human inflammatory and regenerative responses. Previous studies have shown tyrosine-derived polycarbonates (TyrPC) are versatile biomaterials with a wide range of applications across multiple disciplines. The library of TyrPC has been well studied and consists of thousands of polymer compositions with tunable mechanical characteristics and degradation and resorption rates that are useful for nerve guidance tubes (NGTs). NGTs made of different TyrPCs have been used in segmental nerve defect models in small animals. The current study is an extension of this work and evaluates NGTs made using two different TyrPC compositions in a 1 cm porcine peripheral nerve repair model. We first evaluated a nondegradable TyrPC formulation, demonstrating proof-of-concept chronic regenerative efficacy up to 6 months with similar nerve/muscle electrophysiology and morphometry to the autograft repair control. Next, we characterized the acute regenerative response using a degradable TyrPC formulation. After 2 weeks in vivo, TyrPC NGT promoted greater deposition of pro-regenerative extracellular matrix (ECM) constituents (in particular collagen I, collagen III, collagen IV, laminin and fibronectin) compared to commercially available collagen-based NGTs. This corresponded with dense Schwann cell infiltration and axon extension across the lumen. These findings confirmed results reported previously in a mouse model and reveal that TyrPC NGTs were well tolerated in swine and facilitated host axon regeneration and Schwann cell infiltration in the acute phase across segmental defects - likely by eliciting a favorable neurotrophic ECM milieu. This regenerative response ultimately can contribute to functional recovery.

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Some Molecular Mechanisms of Cervical Ripening

International Journal of Biomedicine ◽

10.21103/article10(4)_ra3 ◽

2020 ◽

Vol 10 (4) ◽

pp. 324-329

Author(s):

Yu. Grigorieva ◽

G. Suvorova ◽

A. Chaulin ◽

S. Yukhimets ◽

S. Chemidronov ◽

...

Keyword(s):

Extracellular Matrix ◽

Preterm Labor ◽

Structural Changes ◽

Molecular Mechanisms ◽

Cervical Ripening ◽

Inflammatory Factors ◽

Cervical Insufficiency ◽

Cervical Remodeling ◽

Novel Approaches

Cervical remodeling is an active dynamic process that begins long before the onset of labor. The optimal course of the cervical ripening/remodeling processes is a prerequisite for successful vaginal delivery. Cervical remodeling is a slow progressive process that begins early in mammalian pregnancies, and can be loosely divided into four overlapping phases termed softening, ripening, dilation/labor, and postpartum repair. This review discusses some aspects of structural changes in the cervix at different stages of cervical ripening. In particular, the role of cervical epithelia, immune-inflammatory factors/cells, and components of the cervical extracellular matrix in cervical ripening is considered. A better understanding of the molecular-biochemical and histophysiological processes occurring during cervical remodeling is critical for the development of novel approaches to treat cervical insufficiency, preterm labor, and postpartum cervical disorders associated with its integrity.

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Evaluation of a Novel Degradable Synthetic Biomaterial Patch for Augmentation of Tendon Healing in a Large Animal Model

The Journal of Knee Surgery ◽

10.1055/s-0038-1646930 ◽

2018 ◽

Vol 32 (05) ◽

pp. 434-440 ◽

Cited By ~ 1

Author(s):

Wayne Gersoff ◽

Chantelle Bozynski ◽

Cristi Cook ◽

Ferris Pfeiffer ◽

Keiichi Kuroki ◽

...

Keyword(s):

Biomechanical Testing ◽

Tendon Repair ◽

Tendon Healing ◽

Standard Of Care ◽

Tendon Injury ◽

Large Animal Model ◽

Large Animal ◽

Range Of Movement ◽

Limb Function ◽

Group Outcomes

AbstractTendon injury is common in sports. The standard of care (SOC) for tendon repair is surgical treatment. However, restored tendons often lack complete strength and functionality, and surgical repair is often unsuccessful. This controlled laboratory study investigates the healing of an Artelon patch (AP)-augmented tendon versus tendon repair alone in a preclinical canine patellar tendon defect model. Full-thickness proximal and distal flap defects were created in the patella tendons of eight purpose-bred research mongrel dogs. Dogs were randomly allocated into either the AP-augmented repair group or the SOC group (N = 8; four knees per group). Outcomes measures included limb function and pain; range of motion (ROM) and ultrasound assessment at 2, 4, and 8 weeks; and measurements of elongation, biomechanical testing, and histology at 8 weeks. Data were compared for statistically significant differences to preoperative measures and between groups (p < 0.05). The AP group had higher limb function scores compared with the SOC group at 2, 4, and 8 weeks, with statistically significant differences observed at 2 weeks (AP: 7.1 ± 1.4, SOC: 5.5 ± 0.4, p < 0.05) and 8 weeks (AP: 9.5 ± 0.7, SOC: 7.0 ± 0.9, p < 0.05). The ROM was significantly higher for the AP group at 4 weeks (AP: 105 degrees ± 4, SOC: 89 degrees ± 5, p < 0.05). Pain scores were statistically significantly lower in the AP group at 4 (AP: 0.6 ± 0.5, SOC: 2.2 ± 0.5) and 8 weeks (p < 0.05 for both comparisons). All animals in the AP group displayed full bridging tissue at week 4, while most animals of the SOC group displayed full bridging by week 8. Minimal tendon elongation was observed in both groups. Significantly more force was required to elongate tendons in the AP group compared with the SOC group (p < 0.05). Animals with AP-augmented tendon repair show an earlier regain of function, earlier regain of range of movement, less postoperative pain, and improved tendon strength when compared with animals treated with tendon repair alone.

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Micro-RNA 92a as a Therapeutic Target for Cardiac Microvascular Dysfunction in Diabetes

Biomedicines ◽

10.3390/biomedicines10010058 ◽

2021 ◽

Vol 10 (1) ◽

pp. 58

Author(s):

Mostafa Samak ◽

Diana Kaltenborn ◽

Andreas Kues ◽

Ferdinand Le Noble ◽

Rabea Hinkel ◽

...

Keyword(s):

Molecular Mechanisms ◽

Microvascular Dysfunction ◽

Micro Rna ◽

Large Animal ◽

Cardiac Events ◽

Tissue Slices ◽

Large Animal Models ◽

Angiogenic Effect ◽

Impaired Angiogenesis

Microvascular dysfunction is a pathological hallmark of diabetes, and is central to the ethology of diabetes-associated cardiac events. Herein, previous studies have highlighted the role of the vasoactive micro-RNA 92a (miR-92a) in small, as well as large, animal models. In this study, we explore the effects of miR-92a on mouse and human cardiac microvascular endothelial cells (MCMEC, HCMEC), and its underlying molecular mechanisms. Diabetic HCMEC displayed impaired angiogenesis and a pronounced inflammatory phenotype. Quantitative PCR (qPCR) showed an upregulation of miR-92a in primary diabetic HCMEC. Downregulation of miR-92a by antagomir transfection in diabetic HCMEC rescued angiogenesis and ameliorated diabetic endothelial bed inflammation. Furthermore, additional analysis of potential in silico-identified miR-92a targets in diabetic HCMEC revealed the miR-92a dependent downregulation of an essential metalloprotease, ADAM10. Accordingly, downregulation of ADAM10 impaired angiogenesis and wound healing in MCMEC. In myocardial tissue slices from diabetic pigs, ADAM10 dysregulation in micro- and macro-vasculature could be shown. Altogether, our data demonstrate the role of miR-92a in cardiac microvascular dysfunction and inflammation in diabetes. Moreover, we describe for the first time the metalloprotease ADAM10 as a novel miR-92a target, mediating its anti-angiogenic effect.

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Abstract 16: Enhancing Myocardial Repair With CardioChimeras

Circulation Research ◽

10.1161/res.117.suppl_1.16 ◽

2015 ◽

Vol 117 (suppl_1) ◽

Author(s):

Pearl Quijada ◽

Hazel T Salunga ◽

Nirmala Hariharan ◽

Jonathan Cubillo ◽

Farid El-Sayed ◽

...

Keyword(s):

Infarct Size ◽

Troponin T ◽

Capillary Density ◽

Anterior Wall ◽

Left Ventricular ◽

Parent Cell ◽

Large Animal ◽

Myocardial Repair ◽

Post Injury ◽

Large Animal Models

Dual cell transplantation of cardiac progenitor cells (CPCs) and mesenchymal stem cells (MSCs) after infarction enhances myocardial repair and performance in large animal models relative to delivery of either cell population individually. However, a single stem cell to support both direct and indirect mechanisms of myocardial repair has yet to be identified. CardioChimeras (CCs), a progenitor cell formed by fusion between CPCs and MSCs were analysed for reparative potential after myocardial infarction (MI) relative to individual parents cell or combined parent cell delivery. Two representative CCs, CardioChimera 1 (CC1) and CardioChimera 2 (CC2) were used for this study. CC1 and CC2 improved left ventricular anterior wall thickness (AWT) at 4 weeks, but only CC1 treatment preserved AWT at 18 weeks relative to no cell treatment (PBS). Ejection fraction was enhanced at 6 weeks post injury in CC1 and CC2 groups, which was maintained in CC1, CC2 and CPC + MSC combined groups up to 18 weeks. Infarct size was decreased by 5% in CC1 and CC2 hearts, whereas CPC + MSC and CPC parent groups remained unchanged when comparing 4 to 12 week change in scar size. MSC and PBS groups displayed marked increases in infarct size (10-15%). CC1 and CC2 showed enhanced engraftment potential by 3-fold relative to CPC + MSC and CPC hearts. In contrast, MSCs were detected at low levels (0.04%). CC1 and CC2 discovered within the myocardium expressed early commitment marker cardiac troponin T relative to controls. CC1 and CC2 treatment increased capillary density within the infarct, indicating that cell persistence facilitates paracrine mediated vasculature stabilization and/or formation. CCs merge the application of distinct cells into a single entity for cellular therapeutic intervention in the progression of heart failure. CCs represent a tractable cellular system that improves upon combinatorial cell therapy approaches and supports myocardial regeneration.

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