Blood flow restriction training improves the efficacy of routine intervention in patients with chronic ankle instability.

As a new means of rehabilitation, blood flow restriction training (BFRT) is widely used in the field of musculoskeletal rehabilitation. To observe whether BFRT can improve the efficacy of routine rehabilitation intervention in patients with chronic ankle instability (CAI). Twenty-three patients with CAI were randomly divided into a routine rehabilitation group (RR Group) and a routine rehabilitation â€‹+ â€‹blood flow restriction training group (RR â€‹+ â€‹BFRT Group) according to the Cumberland Ankle Instability Tool (CAIT) score. The RR Group was treated with routine rehabilitation means for intervention, and the RR â€‹+ â€‹BFRT Group was treated with a tourniquet to restrict lower limb blood flow for rehabilitation training based on routine training. Before and after the intervention, the CAIT score on the affected side, standing time on one leg with eyes closed, comprehensive scores of the Y-balance test, and surface electromyography data of tibialis anterior (TA) and peroneus longus (PL) were collected to evaluate the recovery of the subjects. Patients were followed up 1 year after the intervention. After 4 weeks of intervention, the RR â€‹+ â€‹BFRT Group CAIT score was significantly higher than the RR Group (19.33 VS 16.73, p â€‹< â€‹0.05), the time of standing on one leg with eyes closed and the comprehensive score of Y-balance were improved, but there was no statistical difference between groups (p â€‹> â€‹0.05). RR â€‹+ â€‹BFRT Group increased the muscle activation of the TA with maximum exertion of the ankle dorsal extensor (p â€‹< â€‹0.05) and had no significant change in the muscle activation of the PL with maximum exertion of the ankle valgus (p â€‹> â€‹0.05). There was no significant difference in the incidence of resprains within 1 year between the groups (36.36% VS 16.67%, p â€‹> â€‹0.05). The incidence of ankle pain in the RR â€‹+ â€‹BFRT Group was lower than that in the RR Group (63.64% VS 9.09%, p â€‹< â€‹0.01). Therefore, four-weeks BFRT improves the effect of the routine intervention, and BFRT-related interventions are recommended for CAI patients with severe ankle muscle mass impairment or severe pain.

A Combined Treatment of BMP2 and Soluble VEGFR1 for the Enhancement of Tendon-Bone Healing by Regulating Injury-Activated Skeletal Stem Cell Lineage.

BACKGROUND: Bone morphogenetic protein 2 (BMP2) is an appealing osteogenic and chondrogenic growth factor for promoting tendon-bone healing. Recently, it has been reported that soluble vascular endothelial growth factor (VEGF) receptor 1 (sVEGFR1) (a VEGF receptor antagonist) could enhance BMP2-induced bone repair and cartilage regeneration; thus, their combined application may represent a promising treatment to improve tendon-bone healing. Moreover, BMP2 could stimulate skeletal stem cell (SSC) expansion and formation, which is responsible for wounded tendon-bone interface repair. However, whether the codelivery of BMP2 and sVEGFR1 increases tendon enthesis injury-activated SSCs better than does BMP2 alone needs further research. PURPOSE: To study the effect of BMP2 combined with sVEGFR1 on tendon-bone healing and injury-activated SSC lineage. STUDY DESIGN: Controlled laboratory study. METHODS: A total of 128 C57BL/6 mice that underwent unilateral supraspinatus tendon detachment and repair were randomly assigned to 4 groups: (1) untreated control group; (2) hydrogel group, which received a local injection of the blank hydrogel at the injured site; (3) BMP2 group, which received an injection of hydrogel with BMP2; and (4) BMP2 with sVEGFR1 group, which received an injection of hydrogel with BMP2 and sVEGFR1. Histology, micro-computed tomography, and biomechanical tests were conducted to evaluate tendon-bone healing at 4 and 8 weeks after surgery. In addition, flow cytometry was performed to detect the proportion of SSCs and their downstream differentiated subtypes, including bone, cartilage, and stromal progenitors; osteoprogenitors; and pro-chondrogenic progenitors within supraspinatus tendon enthesis at 1 week postoperatively. RESULTS: The repaired interface in BMP2 with sVEGFR1 group showed a significantly improved collagen fiber continuity, increased fibrocartilage, greater newly formed bone, and elevated mechanical properties compared with the other 3 groups. There were more SSCs; bone, cartilage, and stromal progenitors; osteoprogenitors; and pro-chondrogenic progenitors in the BMP2 with sVEGFR1 group than that in the other groups. CONCLUSION: Our study suggests that the combined delivery of BMP2 and sVEGFR1 could promote tendon-bone healing and stimulate the expansion of SSCs and their downstream progeny within the injured tendon-bone interface. CLINICAL RELEVANCE: Combining BMP2 with sVEGFR1 may be a good clinical treatment for wounded tendon enthesis healing.

Argonaute3-SF3B3 complex controls pre-mRNA splicing to restrain type 2 immunity.

Argonaute (AGO) proteins execute microRNA (miRNA)-mediated gene silencing. However, it is unclear whether all 4 mammalian AGO proteins (AGO1, AGO2, AGO3, and AGO4) are required for miRNA activity. We generate Ago1, Ago3, and Ago4-deficient mice (Ago134Δ) and find AGO1/3/4 to be redundant for miRNA biogenesis, homeostasis, or function, a role that is carried out by AGO2. Instead, AGO1/3/4 regulate the expansion of type 2 immunity via precursor mRNA splicing in CD4+ T helper (Th) lymphocytes. Gain- and loss-of-function experiments demonstrate that nuclear AGO3 interacts directly with SF3B3, a component of the U2 spliceosome complex, to aid global mRNA splicing, and in particular the isoforms of the gene Nisch, resulting in a dysregulated Nisch isoform ratio. This work uncouples AGO1, AGO3, and AGO4 from miRNA-mediated RNA interference, identifies an AGO3:SF3B3 complex in the nucleus, and reveals a mechanism by which AGO proteins regulate inflammatory diseases.

TPL2 kinase activity regulates microglial inflammatory responses and promotes neurodegeneration in tauopathy mice.

Tumor progression locus 2 (TPL2) (MAP3K8) is a central signaling node in the inflammatory response of peripheral immune cells. We find that TPL2 kinase activity modulates microglial cytokine release and is required for microglia-mediated neuron death in vitro. In acute in vivo neuroinflammation settings, TPL2 kinase activity regulates microglia activation states and brain cytokine levels. In a tauopathy model of chronic neurodegeneration, loss of TPL2 kinase activity reduces neuroinflammation and rescues synapse loss, brain volume loss, and behavioral deficits. Single-cell RNA sequencing analysis indicates that protection in the tauopathy model was associated with reductions in activated microglia subpopulations as well as infiltrating peripheral immune cells. Overall, using various models, we find that TPL2 kinase activity can promote multiple harmful consequences of microglial activation in the brain including cytokine release, iNOS (inducible nitric oxide synthase) induction, astrocyte activation, and immune cell infiltration. Consequently, inhibiting TPL2 kinase activity could represent a potential therapeutic strategy in neurodegenerative conditions.

TGF-ß1 derived from macrophages contributes to load-induced tendon-bone healing in the murine rotator cuff repair model by promoting chondrogenesis.

It has been established that mechanical stimulation benefits tendon-bone (T-B) healing, and macrophage phenotype can be regulated by mechanical cues; moreover, the interaction between macrophages and mesenchymal stem cells (MSCs) plays a fundamental role in tissue repair. This study aimed to investigate the role of macrophage-mediated MSC chondrogenesis in load-induced T-B healing in depth. C57BL/6 mice rotator cuff (RC) repair model was established to explore the effects of mechanical stimulation on macrophage polarization, transforming growth factor (TGF)-ß1 generation, and MSC chondrogenesis within T-B enthesis by immunofluorescence and enzyme-linked immunosorbent assay (ELISA). Macrophage depletion was performed by clodronate liposomes, and T-B healing quality was evaluated by histology and biomechanics. In vitro, bone marrow-derived macrophages (BMDMs) were stretched with CELLOAD-300 load system and macrophage polarization was identified by flow cytometry and quantitative real-time polymerase chain reaction (qRT-PCR). MSC chondrogenic differentiation was measured by histochemical analysis and qRT-PCR. ELISA and qRT-PCR were performed to screen the candidate molecules that mediated the pro-chondrogenic function of mechanical stimulated BMDMs. Mechanical stimulation promoted macrophage M2 polarization in vivo and in vitro. The conditioned media from mechanically stimulated BMDMs (MS-CM) enhanced MSC chondrogenic differentiation, and mechanically stimulated BMDMs generated more TGF-ß1. Further, neutralizing TGF-ß1 in MS-CM can attenuate its pro-chondrogenic effect. In vivo, mechanical stimulation promoted TGF-ß1 generation, MSC chondrogenesis, and T-B healing, which were abolished following macrophage depletion. Macrophages subjected to appropriate mechanical stimulation could polarize toward the M2 phenotype and secrete TGF-ß1 to promote MSC chondrogenesis, which subsequently augments T-B healing.

Exosomes from CD133+ human urine-derived stem cells combined adhesive hydrogel facilitate rotator cuff healing by mediating bone marrow mesenchymal stem cells.

Background: The inadequate regeneration of natural tissue (mainly fibrocartilage) between tendon and bone during rotator cuff (RC) repair results in an unsatisfactory quality of RC healing. Cell-free therapy based on stem cell exosomes is a safer and more promising approach for tissue regeneration. Here, we investigated the effect of exosomes from human urine-derived stem cells (USCs) and their subpopulations (CD133+USCs) on RC healing. Methods: USCs were isolated from urine and sorted by flow cytometry to obtain CD133+ urine-derived stem cells (CD133+ USCs). Urine-derived stem cell exosomes (USC-Exos) and CD133+ urine-derived stem cell exosomes (CD133+ USC-Exos) were subsequently isolated from the cell supernatant and identified by transmission electron microscopy (TEM), particle size analysis, and Western blot. We performed in vitro functional assays to evaluate the effects of USC-Exos and CD133+ USC-Exos on human bone marrow mesenchymal stem cells (BMSCs) proliferation, migration, osteogenic differentiation, and chondrogenic differentiation. In vivo experiments were performed by local injection of exosome-hydrogel complexes for the treatment of RC injury. The effects of CD133+ USC-Exos and USC-Exos on RC healing were assessed from imaging, histological, and biomechanical tests. Results: CD133+ USCs were positive for CD29, CD44, CD73, CD90, CD133, but negative for CD34 and CD45. Differentiation ability test results showed that both USCs and CD133+ USCs had the potential for osteogenic, chondrogenic, and adipogenic differentiation, but CD133+ USCs had stronger chondrogenic differentiation ability. CD133+ USC-Exos and USC-Exos could be efficiently taken up by BMSCs and promote their migration, osteogenic and chondrogenic differentiation. However, CD133+ USC-Exos could promote the chondrogenic differentiation of BMSCs more than USC-Exos. Compared with USC-Exos, CD133+ USC-Exos could promote the healing of bone-tendon interface (BTI) more effectively, which might be related to its ability to promote the differentiation of BMSCs into chondroblasts. Although the two exosomes exhibited the same effect in promoting subchondral bone repair in BTI, the CD133+ USC-Exos group had higher histological scores and stronger biomechanical properties. Conclusion: CD133+ USC-Exos hydrogel complex may become a promising therapeutic approach for RC healing based on stem cell exosomes. The translational potential of this article: This is the first study to assess the specific role of CD133+ USC-Exos in RC healing which may be related to the activation of BMSCs by CD133+ USC-Exos towards chondrogenic differentiation. Further, our study provides a reference for possible future treatment of BTI by applying CD133+ USC-Exos hydrogel complex.

Exosomes secreted by hypoxia-stimulated bone-marrow mesenchymal stem cells promote grafted tendon-bone tunnel healing in rat anterior cruciate ligament reconstruction model.

Background: After anterior cruciate ligament (ACL) reconstruction in clinic, firm and rapid integration of the grafted tendon into the bone tunnel remains a challenge. Exosomes from hypoxia-treated stem cells are beneficial for promoting angiogenesis and then coupling with osteogenesis. Therefore, exosomes from hypoxia-cultured bone-marrow mesenchymal stem cells (Hypo-Exos) may be a cell-free therapy for enhancing graft-bone incorporation after ACL reconstruction. Methods: Exosomes from normoxia-cultured bone-marrow mesenchymal stem cells (Norm-Exos) or Hypo-Exos were respectively cultured with human umbilical vein endothelial cells (HUVECs) for in-vitro evaluating their functions in HUVECs proliferation, migration, and tube formation. A total of 87 rats with single-bundle ACL reconstructions in the right knee were randomly allocated into 3 different treatments: phosphate-buffered saline (PBS) with the adhesive hydrogel injection as control (Ctrl), Norm-Exos with the adhesive hydrogel injection (Norm-Exos), and Hypo-Exos with the adhesive hydrogel injection (Hypo-Exos). At postoperative weeks 2, 4, or 8, the ACL graft-bone integrations were evaluated. Results: Hypo-Exos was a better stimulator for in-vitro HUVECs proliferation, migration, and tube formation compared to PBS or Norm-Exos. Hypo-Exos within the adhesive hydrogel could be sustained-released at least 14 days around the peri-graft site. Radiologically, at week 4 or 8, femoral or tibial bone tunnel areas (BTA), as well as bone volume/total volume ratio (BV/TV) of the femoral or tibial peri-graft bone in the Hypo-Exos group, improved significantly better than these parameters of the Ctrl and Norm-Exos groups (P<0.05 for all). Histologically, the grafted tendon-bone interface in the Hypo-Exos group showed significantly higher histologic scores at week 4 or 8 as compared with the other groups (P<0.05 for all). Immunofluorescent staining verified that type H vessels were more abundant in the Hypo-Exos group when compared to the Ctrl or Norm-Exos group at week 2. Biomechanically, the Hypo-Exos group exhibited a significantly heightened failure load compared with the Ctrl and Norm-Exos groups (P<0.05 for all) at 8 weeks. Meanwhile, the stiffness in the Hypo-Exos group was the greatest among the three groups. Conclusion: Peri-graft Hypo-Exos injection accelerates grafted tendon-bone tunnel integration after ACL reconstruction by improving peri-graft bone microarchitecture.

Intermittent fasting promotes repair of rotator cuff injury in the early postoperative period by regulating the gut microbiota.

Background: The repair of rotator cuff injury is affected by lifestyle and metabolic factors. Intermittent fasting (IF) can promote repair of damaged tissue by regulating intestinal flora, which provides an idea of therapy for rotator cuff injury. The aim of this study was to investigate the effects of fasting on rotator cuff repair after injury, and the role of intestinal flora or a single strain in this process. Methods: Mice underwent rotator cuff injury were treated with intermittent fasting or fed ad libitum. Fasting began one month before surgery and continued until euthanasia. Fresh feces were collected at 2 weeks before surgery, on the day of surgery, and 2, 4, 8 weeks postoperatively for 16S rRNA microbiome sequencing. Supraspinatus tendon-humerus â€‹(SSTH) complex was collected at 2, 4 and 8 weeks after surgery. Live parabacteroides distasonis (Parabacteroides distasonis) was used for repair of rotator cuff injury, with equal amount of pasteurized P. distasonis (KPD) or sterile anaerobic phosphate buffer saline (PBS) as control. Biomechanical, radiological, histological analysis were used to assess the effect of rotator cuff repair. Results: Biomechanical, radiological and histological analysis indicated that intermittent fasting significantly promoted the repair of rotator cuff injury in the early postoperative period (P ＜ 0.05), but significantly inhibited the repair of rotator cuff injury at 4 weeks postoperatively (P ＜ 0.05). 16S rRNA Microbiome sequencing result showed that P. distasonis was the species with the most obvious changes in intestinal flora of mice after fasting. The results of tensile test, X-ray analysis and histological analysis indicated that the live P. distasonis (LPD) significantly impaired the biomechanical properties, bone regeneration and fibrocartilage regeneration of enthesis postoperatively (P ＜ 0.05). Conclusion: Intermittent fasting promoted repair of rotator cuff injury in the early postoperative period by regulating the gut microbiota, in which P. distasonis played an important role. The translational potential of this article: Intermittent fasting (IF) may be a beneficial lifestyle for the repair of rotator cuff injury in the early postoperative period in clinical, and the influence of a certain strain on the repair of rotator cuff injury may also provide an idea for the treatment of rotator cuff injury in the future.

A Slotted Decellularized Osteochondral Scaffold With Layer-Specific Release of Stem Cell Differentiation Stimulators Enhances Cartilage and Bone Regeneration in Osteochondral Defects in a Rabbit Model.

BACKGROUND: Owing to the disappointing regenerative ability of osteochondral tissue, without treatment an osteochondral defect would progress to osteoarthritis. This situation motivates the need for new strategies to enhance the regeneration of osteochondral defects. PURPOSE: To develop a tissue-engineering scaffold by tethering bone morphogenetic protein 2 (BMP2) and transforming growth factor beta 3 (TGFß3) in a layer-specific manner on a slotted decellularized osteochondral matrix (SDOM) and to evaluate the efficacy of this scaffold for osteochondral regeneration. STUDY DESIGN: Controlled laboratory study. METHODS: Normal osteochondral tissue from the rabbit patellofemoral groove was sectioned into a slot shape and decellularized for fabricating an SDOM. The collagen-binding domain (CBD) was fused into the N-terminus of BMP2 or TGFß3 to synthesize 2 recombinant growth factors (GFs) (CBD-BMP2 or CBD-TGFß3), which were tethered to the bone layer and cartilage layer, respectively, of the SDOM to prepare a tissue-engineering scaffold (namely, CBD-GFs/SDOM). After examining the influence of the CBD-GFs/SDOM on the viability and layer-specific differentiation of bone marrow mesenchymal stem cells in vitro, we determined the regeneration potential of the CBD-GFs/SDOM on osteochondral regeneration in a rabbit model. A total of 72 New Zealand White rabbits with a cylindrical osteochondral defect in the patellofemoral groove were randomly assigned to 3 groups: defect only (control [CTL] group), defect patched with an SDOM (SDOM group), and defect patched with the CBD-GFs/SDOM (CBD-GFs/SDOM group). At 6 or 12 weeks postoperatively, the rabbits were euthanized to harvest the knee joint, which was then evaluated via gross observation, micro-computed tomography, histological staining, and mechanical testing. RESULTS: In vitro, the CBD-GFs/SDOM was noncytotoxic, showed high biomimetics with normal osteochondral tissue, was suitable for cell adhesion and growth, and had good layer-specific ability in inducing stem cell differentiation. Macroscopic images showed that the CBD-GFs/SDOM group had significantly better osteochondral regeneration than the CTL and SDOM groups had. Micro-computed tomography demonstrated that much more bony tissue was formed at the defect sites in the CBD-GFs/SDOM group compared with the defect sites in the CTL or SDOM group. Histological analysis showed that the CBD-GFs/SDOM group had a significant enhancement in osteochondral regeneration at 6 and 12 weeks postoperatively in comparison with the CTL or SDOM group. At 12 weeks postoperatively, the mechanical properties of reparative tissue were significantly better in the CBD-GFs/SDOM group than in the other groups. CONCLUSION: The CBD-GFs/SDOM is a promising scaffold for osteochondral regeneration. CLINICAL RELEVANCE: The findings of this study indicated that the CBD-GFs/SDOM is an excellent candidate for reconstructing osteochondral defects, which may be translated for clinical use in the future.

Activation-Induced Cytidine Deaminase Impacts the Primary Antibody Repertoire in Naive Mice.

Genetic and environmental cues shape the evolution of the B cell Ig repertoire. Activation-induced cytidine deaminase (AID) is essential to generating Ig diversity through isotype class switching and somatic mutations, which then directly influence clonal selection. Impaired B cell development in AID-knockout mice has made it difficult to study Ig diversification in an aging repertoire. Therefore, in this report, we used a novel inducible AID-knockout mouse model and discovered that deleting AID in adult mice caused spontaneous germinal center formation. Deep sequencing of the IgH repertoire revealed that Ab diversification begins early in life and evolves over time. Our data suggest that activated B cells form germinal centers at steady state and facilitate continuous diversification of the B cell repertoire. In support, we identified shared B cell lineages that were class switched and showed age-dependent rates of mutation. Our data provide novel context to the genesis of the B cell repertoire that may benefit the understanding of autoimmunity and the strength of an immune response to infection.

Local Administration of Metformin Improves Bone Microarchitecture and Biomechanical Properties During Ruptured Canine Achilles Tendon-Calcaneus Interface Healing.

BACKGROUND: Tendon-bone interface (TBI) healing is a clinical dilemma that is closely relevant to new bone formation and remodeling at the repair site. Previous studies showed that metformin is an osteogenic inducer for stem cells in vitro and capable of stimulating bone regeneration in vivo. HYPOTHESIS: Metformin would be effective for promoting TBI healing by enhancing new bone formation and remodeling. STUDY DESIGN: Controlled laboratory study. METHODS: Canine bone marrow stem cells (BMSCs) were cultured with various concentrations of metformin (0, 10, 50, 100, 200 µM). The effect of metformin on the osteogenic differentiation of canine BMSCs was evaluated via alizarin red staining and osteogenic gene expression. Eighteen mature beagles were included in a bilateral Achilles tendon-calcaneus (ATC) interface injury model. The right interface was reattached via surgical repair only, while the left was surgically reattached after implanting a fibrin glue containing metformin. At postoperative week 4 or 8, the healing quality of the wounded ATC interfaces was evaluated. RESULTS: In vitro experiments determined that metformin was an osteogenic inducer for canine BMSCs. In vivo experiments showed that the metformin-treated ATC interfaces were repaired with significantly greater failure load and stiffness than was the no-metformin control site (P < .05 for all). Micro-computed tomography analysis showed that the metformin-treated specimens presented significantly higher bone volume/total volume and trabecular thickness than did the no-metformin control specimens (P < .05 for all), as confirmed via hematoxylin and eosin staining. Immunohistochemical staining showed that significantly more osteocalcin-positive cells were located at the newly formed bones treated with metformin than at the no-metformin control site at week 4 (P < .05). Masson trichrome staining showed that significantly more oriented collagen fibers anchored into the newly formed bone of the metformin-treated site than the no-metformin control site (P < .05). CONCLUSION: Metformin induced the osteogenesis of canine BMSCs in vitro, and local administration of metformin provided an improvement of bone microarchitecture at the calcaneus as well as an increase in the tensile properties of the repaired ATC interfaces in canines. CLINICAL RELEVANCE: Findings of the study indicate that local administration of metformin may be an effective strategy for TBI healing in clinic.

Steroid-induced fibroblast growth factors drive an epithelial-mesenchymal inflammatory axis in severe asthma.

Asthma and inflammatory airway diseases restrict airflow in the lung, compromising gas exchange and lung function. Inhaled corticosteroids (ICSs) can reduce inflammation, control symptoms, and improve lung function; however, a growing number of patients with severe asthma do not benefit from ICS. Using bronchial airway epithelial brushings from patients with severe asthma or primary human cells, we delineated a corticosteroid-driven fibroblast growth factor (FGF)-dependent inflammatory axis, with FGF-responsive fibroblasts promoting downstream granulocyte colony-stimulating factor (G-CSF) production, hyaluronan secretion, and neutrophilic inflammation. Allergen challenge studies in mice demonstrate that the ICS, fluticasone propionate, inhibited type 2-driven eosinophilia but induced a concomitant increase in FGFs, G-CSF, hyaluronan, and neutrophil infiltration. We developed a model of steroid-induced neutrophilic inflammation mediated, in part, by induction of an FGF-dependent epithelial-mesenchymal axis, which may explain why some individuals do not benefit from ICS. In further proof-of-concept experiments, we found that combination therapy with pan-FGF receptor inhibitors and corticosteroids prevented both eosinophilic and steroid-induced neutrophilic inflammation. Together, these results establish FGFs as therapeutic targets for severe asthma patients who do not benefit from ICS.

Oncostatin M expression induced by bacterial triggers drives airway inflammatory and mucus secretion in severe asthma.

Exacerbations of symptoms represent an unmet need for people with asthma. Bacterial dysbiosis and opportunistic bacterial infections have been observed in, and may contribute to, more severe asthma. However, the molecular mechanisms driving these exacerbations remain unclear. We show here that bacterial lipopolysaccharide (LPS) induces oncostatin M (OSM) and that airway biopsies from patients with severe asthma present with an OSM-driven transcriptional profile. This profile correlates with activation of inflammatory and mucus-producing pathways. Using primary human lung tissue or human epithelial and mesenchymal cells, we demonstrate that OSM is necessary and sufficient to drive pathophysiological features observed in severe asthma after exposure to LPS or Klebsiella pneumoniae. These findings were further supported through blockade of OSM with an OSM-specific antibody. Single-cell RNA sequencing from human lung biopsies identified macrophages as a source of OSM. Additional studies using Osm-deficient murine macrophages demonstrated that macrophage-derived OSM translates LPS signals into asthma-associated pathologies. Together, these data provide rationale for inhibiting OSM to prevent bacterial-associated progression and exacerbation of severe asthma.

Characterize the microstructure change after tendon enthesis injury using synchrotron radiation µCT.

The microstructure of the bone-tendon interface (BTI) deserves in-depth investigation. In this study, we first aimed to extend the application of synchrotron radiation µCT to characterize the gradient structure of supraspinatus tendon (SST) enthesis, from both tissue morphology to cell distribution. Second, to acquire detailed morphological information of SST enthesis when after injury. Our results showed that in normal enthesis, the phenotype of chondrocyte in BTI was dependent on its distance to subchondral bone. After injury, the fibrocartilage cells were disrupted, as evidenced by reduced lacunae size. Our observation may partly explain the loss of BTI mechanical properties after injury, and we believe the application of synchrotron radiation microcomputed tomography will have promising potential for characterizing the morphology changes in enthesis and for evaluating the therapeutic effects of interventions in preclinical studies.

Early treadmill running delays rotator cuff healing via Neuropeptide Y mediated inactivation of the Wnt/ß-catenin signaling.

BACKGROUND: Defining the optimal rehabilitation programs for rotator cuff healing remains a challenge. Early treadmill running may have negative effects on tendon-bone interface (TBI) healing with increased expression of Neuropeptide Y (NPY). However, the underlying mechanism is still unknown. METHODS: The mice were randomly assigned to four groups: control group, treadmill group, treadmill â€‹+ â€‹BIBO3304 group and BIBO3304 group alone. Specifically, the control group was allowed free cage activity without any treatment after surgery. The treadmill group received early treadmill running initiated from postoperative day 2. The treadmill â€‹+ â€‹BIBO3304 group received treadmill running combined with intra-articular injection of BIBO3304 postoperatively. The BIBO3304 group only received type 1 NPY receptor (Y1 receptor, Y1R) antagonist BIBO3304 postoperatively. Healing outcomes of the rotator cuff were evaluated by histological analysis, synchrotron radiation micro-computed tomography (SR-µCT) scanning, and biomechanical testing at 4 and 8 weeks after surgery. The expression of NPY and its Y1 receptor during the treadmill running were tested by immunofluorescence. In addition, the related signaling pathway of Neuropeptide Y among all groups was detected by immunohistochemistry and western-blot. RESULTS: Immunofluorescence results show that early treadmill training could lead to a significant increase in the expression of NPY at the healing site, and Y1R was widely expressed in both normal or injured rotator cuff without statistical difference. At the same time, early treadmill running delayed the healing of rotator cuff, as indicated with unsatisfactory outcomes, including a significantly lower histological score, decreased bone formation and inferior biomechanical properties at postoperative week 4 and 8. Moreover, the use of BIBO3304 could partly alleviate the negative effects of early treadmill running on the healing of rotator cuff and promote the natural healing process of rotator cuff, as evidenced by significant differences observed between the treadmill and treadmill â€‹+ â€‹BIBO3304 groups, as well as observed between the control and BIBO3304 groups. On the other hand, the expressions of Wnt3a and ß-catenin in the treadmill group were significantly lower compared with the other groups, while the expression in the BIBO3304 group was the highest, as evaluated by immunohistochemistry and western-blot. CONCLUSIONS: Early treadmill running increased the expression of NPY at the RC healing site, which might burden the expression of Wnt3a/ß-catenin and delay the healing process, inhibition of Y1 receptor with BIBO3304 could promote bone-tendon healing through the Wnt/ß-catenin signaling.The translational potential of this article: This is the first study to evaluate the specific role of the NPY-Y1R axis and its underlying mechanism by which early treadmill running delays bone-tendon healing. Further, our study may provide references of precise and individualized exercise-based rehabilitation strategies for TBI healing in clinic. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: This is the first study to evaluate the specific role of the NPY-Y1R axis and its underlying mechanism by which early treadmill running delays bone-tendon healing. Further, our study may provide references of precise and individualized exercise-based rehabilitation strategies for TBI healing in clinic.

Tpl2 kinase regulates inflammation but not tumorigenesis in mice.

Tumor progression locus 2 (Tpl2, gene name MAP3K8), a mitogen-activated protein kinase, is widely expressed in immune and non-immune cells to integrate tumor necrosis factor (TNF), toll-like receptors (TLRs), and interleukin-1 (IL1) receptor signaling to regulate inflammatory response. Given its central role in inflammatory response, Tpl2 is an attractive small molecule drug target. However, the role of Tpl2 as an oncogene or tumor suppressor gene remains controversial, and its function outside immune cells is not understood. We therefore utilized a Tpl2 kinase dead (Tpl2-KD) mouse model in an 18-month aging study to further elucidate Tpl2 effects on lifespan and chronic disease. Histopathological studies revealed the incidence and severity of spontaneous tumors and non-neoplastic lesions were comparable between wild type and Tpl2-KD mice. The only finding was that male Tpl2-KD mice had higher bodyweight and an increased incidence of liver steatosis, suggesting a sex-specific role for Tpl2 in hepatic lipid metabolism. In conclusion, loss of Tpl2 kinase activity did not lead to increased tumorigenesis over aging in mice but affected likely alterations in lipid metabolism in male animals.

Treatment of chronic lateral ankle instability by double-band anatomical reconstruction of the anterior talofibular ligament's fibular enthesis. / è·è “å‰éŸ§å¸¦è “éª¨æ­¢ç‚¹åŒæŸè§£å‰–é‡å»ºæ²»ç–—æ ¢æ€§è¸å ³èŠ‚å¤–ä¾§ä¸ç¨³.

OBJECTIVES: Anterior talofibular ligament (ATFL) injury is one of the most common injuries in sports medicine, resulting in chronic lateral ankle instability (CLAI). The patients' daily life may be seriously affected by ankle osteoarthritis and other irreversible damages, if the ATFL injury is not treated in time and drags on. Patients with ATFL injury who show no significant recovery after 3-6 months of conservative treatment should consider surgical treatment as soon as possible to restore ankle stability and function. This study aims to investigate the effect of double-bands anatomical reconstruction of the ATFL's fibular enthesis for the treatment of CLAI. METHODS: A retrospective review was conducted on 67 patients diagnosed with CLAI in the Department of Sports Medicine, Xiangya Hospital, Central South University from January 2015 to January 2018, including 42 males and 25 females, aged from 17 to 41 years old, with disease course of (12.6±3.2) months. Of the 67 patients, 29 left ankles and 38 right ankles were included in this study. Patients suffered from repeated sprains which leaded to pain, swelling and obvious ankle relaxation. There were obvious tenderness at the ATFL insertion and the calcaneal fibular ligament insertion. Both the anterior ankle drawer test and the varus stress test were positive. Other ankle disorders were excluded by X-ray. Preoperative color Doppler ultrasonography and magnetic resonance examination were performed to observe ATFL injury. All the patients had surgical indications and no obvious contraindications, and they were treated with arthroscopic debridement and double-bundle anatomical reconstruction of the AFTL's fibular enthesis under anesthesia. Postoperative routine nursing and standardized rehabilitation exercise were recommended. Outpatient follow-up was conducted at 3, 6, 12, and 24 months postoperatively. American Orthopaedic Foot and Ankle Society (AOFAS) scores, Karlsson Ankle Functional (KAF) score, and the Japanese Society for Surgery of the Foot (JSSF) scale were used to evaluate the clinical outcomes. RESULTS: Intraoperative arthroscopic examination of 67 patients showed inflammatory synovial hyperplasia in 52 cases (77.6%), obvious osteophyte hyperplasia in 12 cases (17.9%), talus osteochondral injury of grade II-III in 23 cases (34.3%), and cartilage injury of grade IV in 5 cases (7.5%). All operations were carried out successfully, and both the anterior ankle drawer test and the varus stress test were negative under anesthesia after surgery. The anchors were in good position. Among them, 3 patients (4.5%) got temporary superficial peroneal nerve palsy and skin numbness at ankle joint after surgery, which gradually recovered within 2 weeks. There were no serious perioperative complications such as infection and suppurative arthritis. Postoperative follow-up was conducted for 12-24 (15.64±3.17) months. At the last follow-up, all patients were walking normally. Most patients had no pain or occasionally mild pain. Ankle function and motion were restored without re-instability. Sixty-four patients (95.5%) worked and exercised as before the surgery. Standing X-ray examination indicated normal joint space without stenosis, and the internal fixation was in good position. Postoperative AOFAS scores (94.78±6.37) were significantly better than the preoperative scores (64.17±12.43, P<0.01). Besides, the KAF scores and the JSSF ankle/hindfoot scale before surgery were significantly increased (KAF: 91.04±11.36 vs 59.74±13.63, P<0.01; JSSF: 95.32±10.21 vs 66.92±14.38, P<0.01). CONCLUSIONS: Arthroscopic debridement and double-bands anatomical reconstruction of the ATFL's fibular enthesis for the treatment of CLAI gains beneficial short-term effects for its minimal invasion and quick recovery.

Characterization of the Subchondral Bone and Pain Behavior Changes in a Novel Bipedal Standing Mouse Model of Facet Joint Osteoarthritis.

BACKGROUND: The subchondral bone parallels with the progression of osteoarthritis (OA). However, the biomechanical properties and histopathological changes of subchondral bone changes in the lumbar facet joint (LFJ) after long-term axial loading on the spine have not been explored. In this study, we aimed to investigate the subchondral bone histopathological changes that occur in the LFJ and pain behaviors in a novel bipedal standing mouse model. METHODS: Sixteen 8-week-old male C57BL/6 mice were randomly assigned into bipedal standing and control groups. A finite element stimulate model based on the micro-CT data was generated to simulate the von Mises stress distribution on the LFJ during different positions. The spine pain behaviors tests were analysis. In addition, the change in the subchondral bone of the LFJ was assessed by histological and immunohistochemistry staining. RESULTS: The computerized simulation of the von Mises stress distribution in the superior articular process of LFJ at the spine level 5 in the lying position increased and reached a maximum value at the bipedal standing posture. The spine pain behavior test revealed that the threshold of pressure tolerance decreased significantly in bipedal groups relative to control groups, whereas the mechanical hyperalgesia of the hind paw increased significantly in bipedal groups relative to control groups. The axial load accelerates LFJ degeneration with increased histological scores in bipedal groups. The expression of type II collagen and aggrecan (ACAN) was significantly decreased in the bipedal groups compared with the control groups, whereas the expression of MMP13 was increased. Compared with the control groups, the osteoclast activity was activated with higher TRAP-positive staining and associated with increased CD-31-positive vessels and GCRP-positive nerve ending expression in the subchondral bone of LFJ. CONCLUSION: Collectively, long-term axial loading induces the development of spine hyperalgesia in mice associate with increased osteoclast activity and aberrant angiogenesis and nerve invasion into the subchondral bone of LFJ that stimulates the natural pathological change in human LFJ OA. These results indicate that aberrant bone remodeling associate with aberrant nerve innervation in the subchondral bone has a potential as a therapeutic target in LFJ OA pain.

Mucosal CD8 T Cell Responses Are Shaped by Batf3-DC After Foodborne Listeria monocytogenes Infection.

While immune responses have been rigorously examined after intravenous Listeria monocytogenes (Lm) infection, less is understood about its dissemination from the intestines or the induction of adaptive immunity after more physiologic models of foodborne infection. Consequently, this study focused on early events in the intestinal mucosa and draining mesenteric lymph nodes (MLN) using foodborne infection of mice with Lm modified to invade murine intestinal epithelium (InlAMLm). InlAMLm trafficked intracellularly from the intestines to the MLN and were associated with Batf3-independent dendritic cells (DC) in the lymphatics. Consistent with this, InlAMLm initially disseminated from the gut to the MLN normally in Batf3-/- mice. Activated migratory DC accumulated in the MLN by 3 days post-infection and surrounded foci of InlAMLm. At this time Batf3-/- mice displayed reduced InlAMLm burdens, implicating cDC1 in maximal bacterial accumulation in the MLN. Batf3-/- mice also exhibited profound defects in the induction and gut-homing of InlAMLm-specific effector CD8 T cells. Restoration of pathogen burden did not rescue antigen-specific CD8 T cell responses in Batf3-/- mice, indicating a critical role for Batf3 in generating anti-InlAMLm immunity following foodborne infection. Collectively, these data suggest that DC play diverse, dynamic roles in the early events following foodborne InlAMLm infection and in driving the establishment of intestinal Lm-specific effector T cells.

The kinase IRAK4 promotes endosomal TLR and immune complex signaling in B cells and plasmacytoid dendritic cells.

The dysregulation of multiple signaling pathways, including those through endosomal Toll-like receptors (TLRs), Fc gamma receptors (FcγR), and antigen receptors in B cells (BCR), promote an autoinflammatory loop in systemic lupus erythematosus (SLE). Here, we used selective small-molecule inhibitors to assess the regulatory roles of interleukin-1 receptor (IL-1R)-associated kinase 4 (IRAK4) and Bruton's tyrosine kinase (BTK) in these pathways. The inhibition of IRAK4 repressed SLE immune complex- and TLR7-mediated activation of human plasmacytoid dendritic cells (pDCs). Correspondingly, the expression of interferon (IFN)-responsive genes (IRGs) in cells and in mice was positively regulated by the kinase activity of IRAK4. Both IRAK4 and BTK inhibition reduced the TLR7-mediated differentiation of human memory B cells into plasmablasts. TLR7-dependent inflammatory responses were differentially regulated by IRAK4 and BTK by cell type: In pDCs, IRAK4 positively regulated NF-κB and MAPK signaling, whereas in B cells, NF-κB and MAPK pathways were regulated by both BTK and IRAK4. In the pristane-induced lupus mouse model, inhibition of IRAK4 reduced the expression of IRGs during disease onset. Mice engineered to express kinase-deficient IRAK4 were protected from both chemical (pristane-induced) and genetic (NZB/W_F1 hybrid) models of lupus development. Our findings suggest that kinase inhibitors of IRAK4 might be a therapeutic in patients with SLE.