Collagenase Injection Research Articles

Quantitative analysis of tendon histopathology using digital pathology in rat models with Achilles tendon injury

ABSTRACT Although digital image analysis methods that quantify histopathologic features have emerged, no validated quantitative methods are available to evaluate tendon injury. This study aimed to propose and validate a quantitative analysis method to identify the histopathologic features of tendon injuries. The histopathologic features of two Achilles tendon injury models (a partial full-thickness defect model and a collagenase injection model) using Sprague-Dawley rats were evaluated by semiquantitative grading and a quantitative analysis method using a digital pathology software at weeks 1 and 4 after tendon injury (six tendons per group at each time point). The outcome variables between tendon injury models and between time points were compared, and the correlation between semiquantitative scores and the results of the quantitative analysis was investigated. The proposed analysis method quantified the severity of the histopathological features after tendon injury. Quantitative analysis differentiated the cell morphology between tendon injury models and time points better than semiquantitative scoring. The results from quantitative measurements correlated significantly with the semiquantitative scores. The proposed quantitative method can be effective in evaluating the histopathology of tendon injuries.

Additional Treatment after Collagenase Injections and Needle Fasciotomy for Dupuytren’s Disease: A Retrospective Cohort Study

Introduction The aim of this study was to assess the rate of additional treatment after collagenase injection and needle fasciotomy, and what factors are associated with additional procedures for recurrence.Materials and Methods We retrospectively identified 201 adult patients who underwent collagenase injection and 19 patients who underwent needle fasciotomy for Dupuytren’s disease from 2012 to 2014. Outcomes included additional treatment of the same ray for either recurrence or persistence of contracture. To evaluate associated factors, we performed a bivariate analysis.Results Additional treatment after collagenase injection for recurrence was performed in 24% of fingers at a median of 23 months (interquartile range [IQR]: 10.8–36.1) and was associated with bilateral disease (p = 0.008). Additional treatment for persistence was performed in 5.6% at a median of 1.9 months (IQR: 1.1–3.2). Additional treatment for recurrence after needle fasciotomy was performed in 13% of fingers at a median of 28.2 months (IQR: 27.5–28.2) and 4.2% for persistence at 1.1 months. Fingers treated with needle fasciotomy were more likely to undergo secondary open fasciectomy (13% vs. 5.1%, p = 0.022).Conclusion Additional treatment after collagenase injections was performed in 29% of fingers, mostly another collagenase injection, and was associated with bilateral disease. After needle fasciotomy, 17% of patients underwent additional treatment, primarily open partial fasciectomy.

Microglial replacement in the aged brain restricts neuroinflammation following intracerebral hemorrhage

Aged microglia display augmented inflammatory activity after neural injury. Although aging is a risk factor for poor outcome after brain insults, the precise impact of aging-related alterations in microglia on neural injury remains poorly understood. Microglia can be eliminated via pharmacological inhibition of the colony–stimulating factor 1 receptor (CSF1R). Upon withdrawal of CSF1R inhibitors, microglia rapidly repopulate the entire brain, leading to replacement of the microglial compartment. In this study, we investigated the impact of microglial replacement in the aged brain on neural injury using a mouse model of intracerebral hemorrhage (ICH) induced by collagenase injection. We found that replacement of microglia in the aged brain reduced neurological deficits and brain edema after ICH. Microglial replacement-induced attenuation of ICH injury was accompanied with alleviated blood-brain barrier disruption and leukocyte infiltration. Notably, newly repopulated microglia had reduced expression of IL-1β, TNF-α and CD86, and upregulation of CD206 in response to ICH. Our findings suggest that replacement of microglia in the aged brain restricts neuroinflammation and brain injury following ICH.

Heme as an inducer of cerebral damage in hemorrhagic stroke: potential therapeutic implications.

Heme as an inducer of cerebral damage in hemorrhagic stroke: potential therapeutic implications.

Effect of Oral Prednisolone after Collagenase Injection for Dupuytren’s Contracture: A Randomized, Controlled Trial with a 1-Year Follow-Up

Introduction This study verified the effectiveness of oral prednisolone after collagenase clostridium histolyticum (CCH) (10 mg/day for 2 weeks) for Dupuytren’s contracture with a 1-year follow-up.Materials and Methods This study included 31 patients with a contracture of the metacarpophalangeal joint of ≥ 30 degrees. A total of 16 patients were allocated randomly to treatment with prednisolone and 15 patients were treated without prednisolone (control group).Results At day 7, mean total active motion (TAM) was 235 degrees in the prednisolone group and 228 degrees in the control group. Mean Visual Analog Scale was 3.3 in the prednisolone group and 4.6 in the control group. There was significant difference between two groups. At day 30, mean TAM was 241 degrees in the prednisolone group and 233 degrees in the control group. There were significant difference between two groups The mean QuickDASH score was significantly higher in the control group (5.8 vs. 3.4). Recurrence was observed in 2/16 patients (13%) in the prednisolone group and 5/15 patients (33%) in the control group; there was no significant difference.Conclusion The administration of prednisolone decreased the likelihood of adverse effects, and also improved finger flexion range of motion and reduced pain after CCH.

Selective brain entry of lipid nanoparticles in haemorrhagic stroke is linked to biphasic blood-brain barrier disruption.

Haemorrhagic stroke represents a significant public health burden, yet our knowledge and ability to treat this type of stroke are lacking. Previously we showed that we can target ischaemic-stroke lesions by selective translocation of lipid nanoparticles through the site of blood-brain barrier (BBB) disruption. The data we presented in this study provide compelling evidence that haemorrhagic stroke in mice induces BBB injury that mimics key features of the human pathology and, more importantly, provides a gate for entry of lipid nanoparticles-based therapeutics selectively to the bleeding site.Methods: Haemorrhagic stroke was induced in mice by intra-striatal collagenase injection. lipid nanoparticles were injected intravenously at 3 h, 24 h & 48 h post-haemorrhagic stroke and accumulation in the brain studied using in-vivo optical imaging and histology. BBB integrity, brain water content and iron accumulation were characterised using dynamic contrast-enhanced MRI, quantitative T1 mapping, and gradient echo MRI.Results: Using in-vivo SPECT/CT imaging and optical imaging revealed biphasic lipid nanoparticles entry into the bleeding site, with an early phase of increased uptake at 3-24 h post-haemorrhagic stroke, followed by a second phase at 48-72 h. Lipid nanoparticles entry into the brain post-haemorrhage showed an identical entry pattern to the trans-BBB leakage rate (Ktrans [min-1]) of Gd-DOTA, a biomarker for BBB disruption, measured using dynamic contrast-enhanced MRI.Discussion: Our findings suggest that selective accumulation of liposomes into the lesion site is linked to a biphasic pattern of BBB hyper-permeability. This approach provides a unique opportunity to selectively and efficiently deliver therapeutic molecules across the BBB, an approach that has not been utilised for haemorrhagic stroke therapy and is not achievable using free small drug molecules.

Collagenase-Induced Patellar Tendinopathy with Neovascularization: First Results towards a Piglet Model of Musculoskeletal Embolization

Background: Therapeutic strategies targeting neovessels responsible for musculoskeletal chronic pain have emerged, including neovessels embolization. Our study aimed to develop a large animal model of patellar tendinopathy with neovascularization. Methods: Nine 3-month-old male piglets (18 patellar tendons) received percutaneous injections of increasing doses of collagenase (0 to 50 mg) at day 0 (D0). Tendinopathy was evaluated by ultrasound (D7 and D14). Neovascularization was evaluated visually and on angiographies. Bonar score was used for histological analysis (D14). Correlations were evaluated using Spearman’s rank (Rs) test. Results: Research protocol was well tolerated. All tendons were enlarged with a median increase of 31.58% [25–40.28] at D7 (p = 0.244) at D7 and 57.52% [48.41–91.45] at D14 (p = 0.065). Tendons with collagenase injection had more hypoechoic changes, with one tendon rupture (p = 0.012). Neovascularization was reported above 5 mg collagenase (p < 0.01) at D7 and D14 with dose-related neovessels induction (Rs = 0.8, p < 0.001). The Bonar score increased above 5 mg collagenase, correlated with the dose (Rs = 0.666, p = 0.003). Conclusions: The study shows the feasibility, safety and reproducibility of this new large animal model of patellar tendinopathy with neovascularization after collagenase injection. It will allow studying new treatments on direct embolization of neovessels by endovascular approach.

Cost-Effectiveness and Patient Outcomes of Injectable Collagenase to Treat Dupuytren's Contracture.

IntroductionDupuytren’s contracture is a disabling and progressive flexion contracture of the hand that is often treated by a surgical release. Collagenase clostridium histolyticum injection (CCH-I) was introduced to the UK in 2011 as an alternative and less invasive treatment for contracture. The purpose of this study was to evaluate the cost-effectiveness and patient-reported outcome measures (PROMs) of treating Dupuytren’s contracture with collagenase compared to surgery.MethodsA retrospective review identified 151 patients who underwent CCH-I (n=94), limited fasciectomy (LF; n=38) and percutaneous needle fasciotomy (PNF; n=19). Outcomes included PROMs (satisfaction, QuickDASH), complication rates (recurrence, reintervention) and direct costs.ResultsStandardised treatment costs for CCH-I, LF and PNF were £1,125.82, £3,438.28 and £1,143.32 respectively. Collagenase presented a cost-benefit of £88,205 had the LF/PNF group undergone CCH-I. At a mean six-year follow-up, there were no significant differences in complication rates (=0.621) or QuickDASH scores (p=0.157). Collagenase-treated patients reported the highest satisfaction and lowest recurrence rates.DiscussionCollagenase presents a significant cost reduction with superior PROMs relative to surgery for treating single-digit contracture.ConclusionOutpatient CCH-I is a cost-effective treatment with fewer clinical encounters, a similar risk profile to LF/PNF and high levels of patient satisfaction, which warrants serious consideration in light of overburdened waiting lists due to COVID-19.

Evaluation of Therapeutic Effects of Quercetin Against Achilles Tendinopathy in Rats via Oxidative Stress, Inflammation, Apoptosis, Autophagy, and Metalloproteinases

Background: Achilles tendinopathy, seen in athletes and manual labor workers, is an inflammatory condition characterized by chronic tendon pain. Owing to the toxicity that develops in various organs attributed to the use of anti–inflammatory drugs, there is a need for new therapeutic agents. Purpose: In the present study, the effects of quercetin (Que), the one that attracted the most attention of researchers studying this group of flavonoids, were investigated against collagenase–induced tendinopathy. Study Design: Controlled laboratory study. Methods: A total of 35 Sprague-Dawley rats were used in the study. Tendinopathy was created by injecting a single dose of collagenase (10 μL; 10 mg/mL) into the tendons of rats. Thirty minutes after the injection, Que was administered at doses of 25 or 50 mg/kg. Que administration was carried out for 7 days. Animals underwent a motility test at the end of the study. In addition, markers of oxidative stress, inflammation, apoptosis, and autophagy, as well as the expression levels of matrix metalloproteinases (MMPs 2, 3, 9, and 13), ICAM-1, and STAT3, were measured in tendon tissues with biochemical, molecular, and Western blot techniques. Results: The results showed that oxidative stress, inflammation, apoptosis, and autophagy were triggered by the injection of collagenase. In addition, MMPs, ICAM-1, and STAT3 were activated to participate in the development of tendinopathy. Que was found to reduce ICAM-1 levels in tendon tissue. Moreover, Que showed antioxidant, anti–inflammatory, antiapoptotic, and antiautophagic effects on tendons against tendinopathy. More important, Que suppressed the expression of MMPs in the tendon tissues. Conclusion: Que has protective properties against collagenase–induced tendon damage in rats. Clinical Relevance: We believe that with further study, Que may be shown to be an alternative treatment option for athletes or others who experience tendon injuries.

Collagenase Tenotomy for Dupuytren Boutonniere.

Treatment of boutonniere Dupuytren disease is rare and is resistant to treatment because of altered tendon dynamics. The authors used a small dose of collagenase clostridium histolyticum for an enzymatic tenotomy of the distal interphalangeal joint and showed that hyperextension at the distal interphalangeal joint improved significantly. Fifteen patients with boutonniere Dupuytren disease with severe proximal interphalangeal joint contractures averaging -69 degrees of extension were included in the study. Ten patients had at least one previous intervention, including surgical fasciectomy, Digit Widget treatment, and needle aponeurotomy. Collagenase clostridium histolyticum enzymatic tenotomy was performed in-office as a wide-awake procedure. All patients received varying doses of collagenase clostridium histolyticum for volar Dupuytren disease enzymatic fasciotomy and 0.1 mg of collagenase clostridium histolyticum into the distal extensor tendon for tenotomy to treat boutonniere deformity at the same time. Collagenase clostridium histolyticum enzymatic tenotomy significantly improved total active motion of the finger by 41.0 degrees (p = 0.001). Loss of extension at both the metacarpophalangeal joint and the proximal interphalangeal joint also improved with gains of 11.7 (p = 0.04) and 20.7 degrees (p = 0.0005) of extension, respectively. The average distal interphalangeal joint hyperextension was improved from 29.7 degrees to 14.0 degrees (p = 0.002). The authors show that collagenase injection led to significant average improvement in joint contracture at all finger joints and significantly increased the arc of motion at the proximal interphalangeal joint and metacarpophalangeal joint. Although collagenase has been previously used for flexion contractures in Dupuytren disease, we believe it has a role in treating the distal interphalangeal joint hyperextension deformity associated with boutonniere deformity in Dupuytren disease as well. Therapeutic, IV.

Inhibition of Aryl Hydrocarbon Receptor Attenuates Hyperglycemia-Induced Hematoma Expansion in an Intracerebral Hemorrhage Mouse Model.

BackgroundHyperglycemia is associated with greater hematoma expansion (HE) and worse clinical prognosis after intracerebral hemorrhage (ICH). However, the clinical benefits of intensive glucose normalization remain controversial, and there are no approved therapies for reducing HE. The aryl hydrocarbon receptor (AHR) has been shown to participate in hyperglycemia‐induced blood–brain barrier (BBB) dysfunction and brain injury after stroke. Herein, we investigated the role of AHR in hyperglycemia‐induced HE in a male mouse model of ICH.Methods and ResultsCD1 mice (n=387) were used in this study. Mice were subjected to ICH by collagenase injection. Fifty percent dextrose was injected intraperitoneally 3 hours after ICH. AHR knockout clustered regularly interspaced short palindromic repeat was administered intracerebroventricularly to evaluate the role of AHR after ICH. A selective AHR inhibitor, 6,2′,4′‐trimethoxyflavone, was administered intraperitoneally 2 hours or 6 hours after ICH for outcome study. To evaluate the effect of AHR on HE, 3‐methylcholanthrene, an AHR agonist, was injected intraperitoneally 2 hours after ICH. The results showed hyperglycemic ICH upregulated AHR accompanied by greater HE. AHR inhibition provided neurological benefits by restricting HE and preserving BBB function after hyperglycemic ICH. In vivo knockdown of AHR further limited HE and enhanced the BBB integrity. Hyperglycemia directly activated AHR as a physiological stimulus in vivo. The thrombospondin‐1/transforming growth factor‐β/vascular endothelial growth factor axis partly participated in AHR signaling after ICH, which inhibited the expressions of BBB‐related proteins, ZO‐1 and Claudin‐5.ConclusionsAHR may serve as a potential therapeutic target to attenuate hyperglycemia‐induced hematoma expansion and to preserve the BBB in patients with ICH.

Identification of motor patterns during bimanual tasks in children with unilateral cerebral palsy

We developed and validated an electronically administered patient-specific visual analogue survey (PVS) to evaluate changes in hand function after treatment with injectable collagenase clostridium histolyticum (CCH) in Dupuytren's contracture. The items in the PVS were authored and ranked in importance by the patients.In an open-label trial for patients with Dupuytren's contracture receiving CCH injection, 109 patients completed the PVS on the day of injection, day of manipulation and 30-day follow-up. For external validation, patients also completed standard patient-reported outcome measures, the Overall Treatment Effects Scale and QuickDASH, and underwent physician assessment of contracture via goniometry and the table top test.Responses were highly individualised with no single activity being chosen as important by more than 8% of patients. Sports-related activities were mentioned most often (23%). The PVS was highly responsive to changes in patients' conditions with CCH injection (effect size=1.49), much more so than the QuickDASH (effect size=0.50). Additionally, the PVS had no floor or ceiling effects, whereas the QuickDASH ceiling approached 20% post-injection. The PVS had excellent internal consistency (Cronbach's α=0.95) and correlated strongly with the QuickDASH post-injection (Spearman's=−0.67). PVS scores were significantly higher for patients reporting their condition had improved versus those reporting no change after injection. The test–retest reliability of the PVS was poor to fair, in part due to allowing patients to choose different activities at test and retest. However, test–retest reliability was good (intraclass correlation coefficient >0.7) and better than QuickDASH among patients who rated the same activities at test and retest.The PVS is simple to administer and enables individualised assessment in a large number of patients. It is also readily adaptable for use in other diseases, particularly within musculoskeletal medicine.Therapeutic II: Prospective cohort.

Dupuytren\u2019s interventions surgery versus collagenase (DISC) trial: study protocol for a pragmatic, two-arm parallel-group, non-inferiority randomised controlled trial

BackgroundDupuytren’s contracture is a fibro-proliferative disease of the hands affecting over 2 million UK adults, particularly the white, male population. Surgery is the traditional treatment; however, recent studies have indicated that an alternative to surgery—collagenase clostridium histolyticum (collagenase)—is better than a placebo in the treatment of Dupuytren’s contracture. There is however no robust randomised controlled trial that provides a definitive answer on the clinical effectiveness of collagenase compared with limited fasciectomy surgery. Dupuytren’s intervention surgery vs collagenase trial (DISC) trial was therefore designed to fill this evidence gap.Methods/designThe DISC trial is a multi-centre pragmatic two-arm parallel-group, randomised controlled trial. Participants will be assigned 1:1 to receive either collagenase injection or surgery (limited fasciectomy).We aim to recruit 710 adult participants with Dupuytren’s contracture. Potential participants will be identified in primary and secondary care, screened by a delegated clinician and if eligible and consenting, baseline data will be collected and randomisation completed.The primary outcome will be the self-reported patient evaluation measure assessed 1 year after treatment. Secondary outcome measures include the Unité Rhumatologique des Affections de la Main Scale, the Michigan Hand Questionnaire, EQ-5D-5L, resource use, further procedures, complications, recurrence, total active movement and extension deficit, and time to return to function. Given the limited evidence comparing recurrence rates following collagenase injection and limited fasciectomy, and the importance of a return to function as soon as possible for patients, the associated measures for each will be prioritised to allow treatment effectiveness in the context of these key elements to be assessed.An economic evaluation will assess the cost-effectiveness of treatments, and a qualitative sub-study will assess participants’ experiences and preferences of the treatments.DiscussionThe DISC trial is the first randomised controlled trial, to our knowledge, to investigate the clinical and cost-effectiveness of collagenase compared to limited fasciectomy surgery for patients with Dupuytren’s contracture.Trial registrationClinical.Trials.gov ISRCTN18254597. Registered on April 11, 2017.

Early-Stage Primary Anti-inflammatory Therapy Enhances the Regenerative Efficacy of Platelet-Rich Plasma in a Rabbit Achilles Tendinopathy Model

Background: Tendinopathy is a pervasive clinical problem that afflicts both athletes and the general public. Although the inflammatory changes in tendinopathy are well characterized, how the therapeutic effects of platelet-rich plasma (PRP) on tendinopathy are being modulated by the inflammatory environment is not well defined. Purpose/Hypothesis: In this study, we aimed to compare the therapeutic effects of PRP alone versus a combination of PRP with a primary glucocorticoid (GC) injection at the early stage of tendinopathy. We hypothesized that PRP treatment could promote better tendon regeneration through the suppression of inflammation with GC. Study Design: Controlled laboratory study. Methods: The gene expression profile of tendon stem/progenitor cells (TSPCs) cultured with PRP was analyzed with RNA sequencing. To evaluate the cell viability, senescence, and apoptosis of TSPCs under different conditions, TSPCs were treated with 0.1 mg/mL triamcinolone acetonide (TA) and/or 10% PRP in an IL1B–induced inflammatory environment. To further verify the effects of the sequential therapy of GCs and PRP, an early tendinopathy animal model was established through a local injection of collagenase in the rabbit Achilles tendon. The tendinopathy model was then treated with isopycnic normal saline (NS group), TA (TA group), PRP (PRP group), or TA and PRP successively (TA+PRP group). At 8 weeks after treatment, the tendons were assessed with magnetic resonance imaging (MRI), histological examination, transmission electron microscopy (TEM), and mechanical testing. Results: Gene Ontology enrichment analysis indicated that PRP treatment of TPSCs induced an inflammatory response, regulated cell migration, and remodeled the extracellular matrix. Compared with the sole use of PRP, successive treatment with TA followed by PRP yielded similar results in cell viability and senescence but less cell apoptosis in vitro. In vivo experiments demonstrated that the TA+PRP group achieved significantly better tendon regeneration, as confirmed by MRI, histological examination, TEM, and mechanical testing. Conclusion: This study showed that the primary use of GCs did not exert any obvious deleterious side effects on the treated tendon but instead enhanced the regenerative effects of PRP in early inflammatory tendinopathy. Clinical Relevance: The sequential therapy of GCs followed by PRP provides a promising treatment strategy for tendinopathy in clinical practice. PRP combined with the primary use of GCs appears to promote tendon regeneration in early inflammatory tendinopathy.

Report on the Evidence-Based Practice Committee’s Survey on Dupuytren Disease

PurposeThe Evidence-Based Practice Committee of the American Society for Surgery of the Hand (ASSH) set out to assess the membership’s practice patterns (PPs) and knowledge of evidence-based principles for Dupuytren disease (DD).MethodsA 21-item multiple-choice survey was distributed to all ASSH members via email in June 2020. Questions were divided into 2 types: evidence-based practice (EBP) and PPs. The survey addressed the following subtopics: nonsurgical, percutaneous, and open surgical management of DD.ResultsThe response rate was 18% (n = 419). Of 13 EBP questions, 5 were answered with the preferred response by >75% of surgeons. The remaining 8 EBP questions had greater frequencies of less preferred responses, which concerned the current evidence for percutaneous management, as well as nonsurgical and postoperative management of DD. Of the PP questions, there were differences in opinion on how to manage a painful nodule, the percutaneous technique (eg, collagenase injection vs percutaneous needle aponeurotomy), and the choice of surgical incision for open fasciectomy (eg, Bruner incision with Z-plasties, partial closure with an open transverse palmar component, or longitudinal incision with Z-plasties).ConclusionsHand surgeons continue to be well informed about current evidence-based practices for treating DD and can improve their knowledge by familiarizing themselves with current data on percutaneous and nonsurgical methods. There exist differences in PPs for DD in the ASSH membership, specifically with less invasive management; and knowledge of peer practices can help navigate differences, critically interpret the evidence, and optimize patient care.Type of study/level of evidenceEconomic/Decision Analyses V.

Dauricine alleviated secondary brain injury after intracerebral hemorrhage by upregulating GPX4 expression and inhibiting ferroptosis of nerve cells

Intracerebral hemorrhage (ICH) is a severe stroke subtype with high disability and mortality, and no effective treatment is available. Previous research on intracerebral hemorrhage secondary brain injury drugs mainly targeted at cell apoptosis, inflammation and oxidative stress, but did not achieve good effects. In recent years, ferroptosis has become a focus concern in neurological diseases. Ferroptosis is a new type of programmed cell death caused by iron-dependent accumulation of lipid peroxides, in which glutathione peroxidase 4 (GPX4) is a key protein affecting ferroptosis. In this study, we used the STRING protein database to predict the proteins that may be co-expressed with GPX4, and studied the ability of Dauricine(Dau) to up-regulate the expression of GPX4 against ferroptosis and neuroprotection after intracerebral hemorrhage in normal cells in vitro, glutathione peroxidase 4 (GPX4) knockdown cells and collagenase injection in vivo in mouse models of intracerebral hemorrhage. The results showed that glutathione reductase (GSR) was a possible co-expression protein with GPX4. Dau could up-regulate the expression of glutathione peroxidase 4 (GPX4) in intracerebral hemorrhage(ICH) model, normal cells and GPX4 knockdown cells in vitro, and simultaneously up-regulate the expression of GSR in ICH mice. Dau could also reduce the levels of iron and lipid peroxidation, and have a neuroprotective effect on intracerebral hemorrhage(ICH) mice. It was tesified that Dauricine(Dau) could inhibit ferroptosis of nerve cells and alleviate brain injury after intracerebral hemorrhage by upregulating glutathione peroxidase 4 (GPX4) and glutathione reductase (GSR) co-expression. Therefore, Dau may be an effective drug for inhibiting ferroptosis and treating intracerebral hemorrhage.

Profiling of Blood-Brain Barrier Disruption in Mouse Intracerebral Hemorrhage Models: Collagenase Injection vs. Autologous Arterial Whole Blood Infusion.

Disruption of the blood-brain barrier (BBB) and the subsequent formation of brain edema is the most severe consequence of intracerebral hemorrhage (ICH), leading to drastic neuroinflammatory responses and neuronal cell death. A better understanding of ICH pathophysiology to develop effective therapy relies on selecting appropriate animal models. The collagenase injection ICH model and the autologous arterial whole blood infusion ICH model have been developed to investigate the pathophysiology of ICH. However, it remains unclear whether the temporal progression and the underlying mechanism of BBB breakdown are similar between these two ICH models. In this study, we aimed to determine the progression and the mechanism of BBB disruption via the two commonly used murine ICH models: the collagenase-induced ICH model (c-ICH) and the double autologous whole blood ICH model (b-ICH). Intrastriatal injection of 0.05 U collagenase or 20 μL autologous blood was used for a comparable hematoma volume in these two ICH models. Then we analyzed BBB permeability using Evan’s blue and IgG extravasation, evaluated tight junction (TJ) damage by transmission electron microscope (TEM) and Western blotting, and assessed matrix metalloproteinase-9 (MMP-9) activity and aquaporin 4 (AQP4) mRNA expression by Gelatin gel zymography and RT-PCR, respectively. The results showed that the BBB leakage was associated with a decrease in TJ protein expression and an increase in MMP-9 activity and AQP4 expression on day 3 in the c-ICH model compared with that on day 5 in the b-ICH model. Additionally, using TEM, we found that the TJ was markedly damaged on day 3 in the c-ICH model compared with that on day 5 in the b-ICH model. In conclusion, the BBB was disrupted in the two ICH models; compared to the b-ICH model, the c-ICH model presented with a more pronounced disruption of BBB at earlier time points, suggesting that the c-ICH model might be a more suitable model for studying early BBB damage and protection after ICH.

Inhibition of P2X4R attenuates white matter injury in mice after intracerebral hemorrhage by regulating microglial phenotypes

BackgroundWhite matter injury (WMI) is a major neuropathological event associated with intracerebral hemorrhage (ICH). P2X purinoreceptor 4 (P2X4R) is a member of the P2X purine receptor family, which plays a crucial role in regulating WMI and neuroinflammation in central nervous system (CNS) diseases. Our study investigated the role of P2X4R in the WMI and the inflammatory response in mice, as well as the possible mechanism of action after ICH.MethodsICH was induced in mice via collagenase injection. Mice were treated with 5-BDBD and ANA-12 to inhibit P2X4R and tropomyosin-related kinase receptor B (TrkB), respectively. Immunostaining and quantitative polymerase chain reaction (qPCR) were performed to detect microglial phenotypes after the inhibition of P2X4R. Western blots (WB) and immunostaining were used to examine WMI and the underlying molecular mechanisms. Cylinder, corner turn, wire hanging, and forelimb placement tests were conducted to evaluate neurobehavioral function.ResultsAfter ICH, the protein levels of P2X4R were upregulated, especially on day 7 after ICH, and were mainly located in the microglia. The inhibition of P2X4R via 5-BDBD promoted neurofunctional recovery after ICH as well as the transformation of the pro-inflammatory microglia induced by ICH into an anti-inflammatory phenotype, and attenuated ICH-induced WMI. Furthermore, we found that TrkB blockage can reverse the protective effects of WMI as well as neuroprotection after 5-BDBD treatment. This result indicates that P2X4R plays a crucial role in regulating WMI and neuroinflammation and that P2X4R inhibition may benefit patients with ICH.ConclusionsOur results demonstrated that P2X4R contributes to WMI by polarizing microglia into a pro-inflammatory phenotype after ICH. Furthermore, the inhibition of P2X4R promoted pro-inflammatory microglia polarization into an anti-inflammatory phenotype, enhanced brain-derived neurotrophic factor (BDNF) production, and through the BDNF/TrkB pathway, attenuated WMI and improved neurological function. Therefore, the regulation of P2X4R activation may be beneficial for the reducing of ICH-induced brain injury.

Adipose micro-grafts enhance tendinopathy healing in ovine model: An in vivo experimental perspective study.

In Europe, approximatively 100 000 to 500 000 tendon repairs are performed every year. These procedures are associated with a considerable rate of postoperative complications (from 6% to 11%). Autologous micro‐grafts (AAMG) and stromal vascular fraction (SVF) have been shown to improve tendon healing in 60% to 70% of treated rodents. The purpose of this study was to evaluate the effects of AAMG in a sheep model with tendinopathy. We used sheep models because, as a large animal, they are more comparable to humans. The hypothesis was that SVF injection would improve tendon healing compared with the control group, reducing inflammatory and matrix degrading, while increasing anti‐inflammatory expression and collagen synthesis in the early stage of tendon injury. Sixteen Apennine sheep aged 2 to 5 years underwent 500 UI type I collagenase injection into both common calcaneal tendons (CCT) to induce tendinopathy. After 15 days (T0), one CCT in every ovine underwent randomly to 2.5 mL of AAMG obtained by mechanical disruption and the contralateral CCTs received no treatment. Clinical, ecographic, and sonographic evaluations were performed after 4 weeks (T1) and 8 weeks (T2). Histological, immunohistochemical, real‐time polymerase chain reaction (RT‐PCR), and biomechanical evaluations were performed at T2. At T2, the treated group showed a final tendon diameter (9.1 ± 1.4 mm) and a hardness expression (62%) that were similar to the original healthy tendon (8.1 ± 1.1 mm; 100%), with a significant recovery compared with the control group (9.5 ± 1.7 mm; 39%). Moreover, histological analysis of the treated group revealed an improvement in the fiber orientation score, fiber edema score, infiltrative‐inflammatory process, and necrosis score (4.3 ± 3.3) compared with control group (8.8 ± 2.9). Immunohistochemically, the treated group showed high expression of collagen 1, Factor VIII and significantly low expression of collagen 3. These data were confirmed by RT‐PCR analysis. The study findings suggested that AAMGs obtained through mechanical disruption present a safe, efficient, and reliable technique, enhancing tendon healing.

Disease-Modifying Potential of Metformin and Alendronate in an Experimental Mouse Model of Osteoarthritis.

Osteoarthritis (OA) is the most common degenerative joint disease causing progressive damages of the cartilage and subchondral bone, synovial inflammation, and severe pain. Despite the complex pathomorphological changes that occur in OA, the approach to different forms of OA is standardized. The global results from pharmacological treatment are not satisfactory. Hence, this study aimed to explore the effects of metformin, alendronate, and their combination on OA development and progression in mice with collagenase-induced osteoarthritis (CIOA). Female ICR (CD-2) mice were randomized to five groups: control group, CIOA untreated, CIOA + metformin, CIOA + alendronate, and CIOA + metformin + alendronate. OA was induced by the intra-articular (i.a.) injection of collagenase. OA phenotype was analyzed by flow cytometry (bone marrow cell differentiation), ELISA (serum levels of the adipokines leptin and resistin), and histology (pathological changes of the knee joint). Treatment with metformin, alendronate, or their combination inhibited the expression of RANK and RANKL on osteoblasts and osteoclasts obtained by ex vivo cultivation of bone marrow cells in mineralization or osteoclastogenic media. In addition, metformin treatment was effective for the attenuation of fibroblast differentiation, but not of mesenchymal stem cells (MSCs), while alendronate had an opposite effect. The combination of metformin and alendronate had a suppressive effect on both MSCs and fibroblasts differentiation. Treatment with metformin, alendronate, and their combination decreased serum concentrations of leptin and resistin in the chronic phase of arthritis. The histopathological examination showed that compared with the untreated CIOA group (OA score 9), the groups treated with metformin (OA score 4) or alendronate (OA score 6) had lower scores for cartilage changes. Metformin combined with alendronate significantly decreased the degree of cartilage degeneration (OA score 2), suggesting that this combination might be a useful approach for the treatment of OA patients.