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.

NLRC3 expression in macrophage impairs glycolysis and host immune defense by modulating the NF-κB-NFAT5 complex during septic immunosuppression.

Impairment of innate immune cell function and metabolism underlies immunosuppression in sepsis; however, a promising therapy to orchestrate this impairment is currently lacking. In this study, high levels of NOD-like receptor family CARD domain containing-3 (NLRC3) correlated with the glycolytic defects of monocytes/macrophages from septic patients and mice that developed immunosuppression. Myeloid-specific NLRC3 deletion improved macrophage glycolysis and sepsis-induced immunosuppression. Mechanistically, NLRC3 inhibits nuclear factor (NF)-κB p65 binding to nuclear factor of activated T cells 5 (NFAT5), which further controls the expression of glycolytic genes and proinflammatory cytokines of immunosuppressive macrophages. This is achieved by decreasing NF-κB activation-co-induced by TNF-receptor-associated factor 6 (TRAF6) or mammalian target of rapamycin (mTOR)-and decreasing transcriptional co-activator p300 activity by inducing NLRC3 sequestration of mTOR and p300. Genetic inhibition of NLRC3 disrupted the NLRC3-mTOR-p300 complex and enhanced NF-κB binding to the NFAT5 promoter in concert with p300. Furthermore, intrapulmonary delivery of recombinant adeno-associated virus harboring a macrophage-specific NLRC3 deletion vector significantly improved the defense of septic mice that developed immunosuppression upon secondary intratracheal bacterial challenge. Collectively, these findings indicate that NLRC3 mediates critical aspects of innate immunity that contribute to an immunocompromised state during sepsis and identify potential therapeutic targets.

Mechanosensitive ion channel Piezo1 mediates mechanical ventilation-exacerbated ARDS-associated pulmonary fibrosis.

INTRODUCTION: Pulmonary fibrosis is a major cause of the poor prognosis of acute respiratory distress syndrome (ARDS). While mechanical ventilation (MV) is an indispensable life-saving intervention for ARDS, it may cause the remodeling process in lung epithelial cells to become disorganized and exacerbate ARDS-associated pulmonary fibrosis. Piezo1 is a mechanosensitive ion channel that is known to play a role in regulating diverse physiological processes, but whether Piezo1 is necessary for MV-exacerbated ARDS-associated pulmonary fibrosis remains unknown. OBJECTIVES: This study aimed to explore the role of Piezo1 in MV-exacerbated ARDS-associated pulmonary fibrosis. METHODS: Human lung epithelial cells were stimulated with hydrochloric acid (HCl) followed by mechanical stretch for 48 h. A two-hitmodel of MV afteracidaspiration-inducedlunginjuryin mice was used. Mice were sacrificed after 14 days of MV. Pharmacological inhibition and knockout of Piezo1 were used to delineate the role of Piezo1 in MV-exacerbated ARDS-associated pulmonary fibrosis. In some experiments, ATP or the ATP-hydrolyzing enzyme apyrase was administered. RESULTS: The stimulation of human lung epithelial cells to HCl resulted in phenotypes of epithelial-mesenchymal transition (EMT), which were enhanced by mechanical stretching. MV exacerbated pulmonary fibrosis in mice exposed to HCl. Pharmacologicalinhibitionorknockout of Piezo1 attenuated the MV-exacerbated EMT process and lung fibrosis in vivo and in vitro. Mechanistically, the observed effects were mediated by Piezo1-dependent Ca2+ influx and ATP release in lung epithelial cells. CONCLUSIONS: Our findings identify a key role for Piezo1 in MV-exacerbated ARDS-associated pulmonary fibrosis that is mediated by increased ATP release in lung epithelial cells. Inhibiting Piezo1 may constitute a novelstrategyfor the treatment of MV-exacerbated ARDS-associated pulmonary fibrosis.

Negative regulator NLRC3: Its potential role and regulatory mechanism in immune response and immune-related diseases.

NLRC3 is a member of the pattern recognition receptors nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) family, and plays a pivotal regulatory role in modulating the activation of immune cells. In macrophages, NLRC3 inhibits the activation of the NF-κB signaling pathway, the STING/TBK1 signaling pathway, and the formation of the inflammasome. In the context of T cells immune response, NLRC3 prevents the activation of T cells by regulating the function of dendritic cells and directly influencing the function of T cells. Different from other pattern recognition receptors, NLRC3 is more closely associated with regulatory activity than pathogens recognition, it influences the fates of cells, for example, prevents proliferation, promotes apoptosis and inhibits pyroptosis. These cellular functions regulated by NLRC3 are involved in the development processes of a variety of diseases, such as infectious disease, sterile inflammatory diseases, and cancer. However, its characteristics, function and regulatory mechanism in immune response and immune-related diseases have not been addressed fully. In this review, we elaborate the potential roles of NLRC3 from several different levels, include molecular mechanism, cellular functions in the immune-related diseases.

Mechanical stimulation improves rotator cuff tendon-bone healing via activating IL-4/JAK/STAT signaling pathway mediated macrophage M2 polarization.

Background: It is well known that appropriate mechanical stimulation benefits tendon-bone (T-B) healing, however, the mechanisms behind this are still uncovered completely. Here, we aimed to explore whether the IL-4/JAK/STAT signaling pathway mediated macrophage polarization was involved in mechanical stimulation induced T-B healing. Method: C57BL/6 mice rotator cuff (RC) repair model was established, and the mice were randomly allocated to the following group. 1. Mice were allowed for free cage activities after surgery (FC group); 2. Mice received treadmill running initiated on postoperative day 7 (TR group); 3. Mice only received a local injection of hydrogel containing IL-4 neutralizing antibody without postoperative intervention (FC â€‹+ â€‹AF-404-SP group); 4. Mice received a local injection of hydrogel containing IL-4 neutralizing antibody and postoperative treadmill running (TR â€‹+ â€‹AF-404-SP group). The expression of IL-4 within supraspinatus tendon (SST) enthesis was measured by Enzyme-linked immunosorbent assay (ELISA). In addition, the activation of JAK/STAT signaling pathway in macrophages and identification of macrophage phenotype at the RC insertion site was detected by Flow cytometry and qRT-PCR. T-B healing quality in this RC repair model was evaluated by histological staining, Micro-computed tomography (Micro-CT) scanning, and biomechanical testing. Result: In this study, using the RC repair model, we confirmed that generation of IL-4, activation of the JAK/STAT signaling pathway in macrophages, the ability of macrophages to polarize towards M2 subtype, and T-B healing quality were significantly enhanced in TR group compared to FC group. When comparing FC â€‹+ â€‹AF-404-SP group with TR â€‹+ â€‹AF-404-SP group, it was found that the mechanical stimulation induced this effect was depleted following the blockade of the IL-4/JAK/STAT signaling pathway. Conclusion: Our finding suggested that mechanical stimulation could accelerate T-B healing via activating the IL-4/JAK/STAT signaling pathway that modulates macrophages to polarize towards M2 subtype. The translational potential of this article: This is the first study to reveal a significant role of mechanical stimulation in the IL-4/JAK/STAT signaling pathway activation and macrophage polarization during RC T-B healing, which highlights the IL-4/JAK/STAT signaling pathway as a potential target to mediate macrophage M2 polarization and improves T-B healing for RC repair.

[Research advances of myeloid-derived suppressor cells in sepsis-induced immunosuppression].

Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. Most patients with sepsis underwent a state of immune suppression after surviving the acute inflammatory response, and were susceptible to secondary nosocomial infections, leading to a prolonged hospitalization and increased mortality rate. Myeloid-derived suppressor cells (MDSCs), a heterogeneous population with immunosuppressive activities, can contribute to the development of immunosuppression in patients with cancer and inhibit the host immune response, but the characteristics of MDSCs and their functional mechanism has not been fully addressed in the development of sepsis-induced immunosuppression. Thus, this review will summary the new findings on the mechanisms of MDSCs in septic immunosuppressionin order to provide ideas and directions for targeting MDSCs as treatment of septic immunosuppression.

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.

Efficacy and safety of a novel personalized navigation template in proximal femoral corrective osteotomy for the treatment of DDH.

BACKGROUND: This present study is aimed to retrospectively evaluate the efficacy and safety of a novel personalized navigation template in proximal femoral corrective osteotomy for the treatment of DDH. METHODS: Twenty-nine consecutive patients with DDH who underwent proximal femoral corrective osteotomy were evaluated between August 2013 and June 2017. Based on the different surgical methods, they were divided into the conventional group (n = 14) and navigation template group (n = 15). The osteotomy degrees, radiation exposure, and operation time were compared between the two groups. RESULTS: No major complications relating to osteotomy surgery such as redislocation or avascular necrosis occurred in the navigation template group, which had more accurate osteotomy degrees, less radiation exposure, and shorter operation time when compared with the conventional group (P < 0.05). Moreover, there was significant difference according to the McKay criteria between the two groups (P = 0.0362). CONCLUSIONS: The novel personalized navigation template in proximal femoral corrective osteotomy is effective and safe, which could improve the femoral osteotomy accuracy, reduce radiation exposure, and shorten operation time.

Curcumin ameliorates monosodium urate-induced gouty arthritis through Nod-like receptor 3 inflammasome mediation via inhibiting nuclear factor-kappa B signaling.

BACKGROUND: Monosodium urate (MSU) crystals-induced inflammation is a key initiator in gouty arthritis. Curcumin is an active ingredient possessing anti-inflammatory efficacy. But the underlying mechanism is not fully understood and its effect on gouty arthritis remains elusive. METHODS: We evaluated the effects of curcumin on cell viability in primary rat abdominal macrophages with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT). Then supernatants of MSU crystals-stimulated cells were collected and subjected to enzyme-linked immunosorbent assay for checking the modulation of curcumin on interleukin (IL)-1ß and tumor necrosis factor (TNF)-α. Meanwhile, cells were analyzed by using Western blot analysis and quantitative polymerase chain reaction (QPCR) to investigate the effects of curcumin on Nod-like receptor 3 (NLRP3) inflammasome/nuclear factor-kappa B (NF-κB) signaling. We also investigated the in vivo efficacy of curcumin with MSU-induced gouty arthritis rat models. RESULTS: Curcumin could reduce MSU crystals-induced IL-1ß and TNF-α in vitro. Western blot analysis and QPCR results revealed that curcumin regulated the production of these cytokines by suppressing the expression of inflammasome key components, including NLRP3, caspase-1. Further studies showed that the suppressive efficacy of curcumin on inflammasome was mediated by inhibiting MSU-induced NF-κB signaling activation. Intraperitoneal administration of curcumin could ameliorate symptoms of MSU-induced gouty arthritis, including the joint circumference, infiltration of neutrophils in knee joints, and production of IL-1ß, TNF-α, and elastase. Western blot analysis revealed that the levels of NLRP3, procaspase-1, caspase-1, pro-IL-1ß, and IL-1ß were downregulated by curcumin in vivo. CONCLUSIONS: These results indicated that curcumin could effectively ameliorate MSU crystal-induced gouty arthritis through NLRP3 inflammasome mediation via inhibiting NF-κB signaling both in vitro and in vivo, suggesting a promising active ingredient for the prevention and treatment of gouty arthritis.

Anatomic reconstruction of anterior talofibular ligament with tibial tuberosity-patellar tendon autograft for chronic lateral ankle instability.

INTRODUCTION: Anatomic repair of the anterior talofibular ligament (ATFL) is challenging when the local ligamentous tissue is severely attenuated. Anatomic reconstruction of the ATFL with tibial tuberosity-patellar tendon (TT-PT) autograft is a feasible choice that can avoid the complicated tendon-bone healing and restore ankle stability. MATERIALS AND METHODS: From 2009 to 2015, 31 chronic lateral ankle instability (CLAI) patients (31 ankles), who had a serious injury on the ATFL only, were treated with anatomic reconstruction of ATFL with TT-PT. American orthopedic foot and ankle society ankle-hindfoot score (AHS), visual analog scale for pain score (VAS), Karlsson-Peterson score, Tegner activity level, and objective examination comprehending range of motion were used to evaluate the clinical outcomes before and after operation. Radiographically, talar tilt angles and anterior drawer were assessed in pre- and postoperative ankle stress views. RESULTS: Among the 31 ankles, 17 ankles with single-bundle ATFL and 14 ankles with double-bundle ATFL were found at operation. At a mean follow-up of 42 months (24-82 months), all patients were satisfied with the procedure. Mean AHS significantly increased from 60.5 ± 8.2 to 93.5 ± 4.8. Mean Karlsson-Peterson score significantly increased from 55.2 ± 11.0 preoperatively to 91.2 ± 6.9 at final follow-up. Average VAS significantly decreased from 5.9 ± 1.6 preoperatively to 1.4 ± 1.0 at the latest follow-up. Mean Tegner activity level was 3.7 ± 0.9 before operation, compared with 7.0 ± 0.8 after operation. On stress radiographs, mean talar tilt angle was 17.0 ± 3.4° before operation and 3.8 ± 2.1° at the latest follow-up. In addition, mean anterior tibiotalar translation was 7.5 ± 2.2 mm before operation and 1.8 ± 1.1 mm at the latest follow-up. CONCLUSION: Anatomic reconstruction of the ATFL using a TT-PT autograft allows bone-bone healing in talus and tendon-tendon/periosteum healing in fibula rather than requiring tendon-bone healing, which is an alternative choice for treating CLAI caused by single ATFL insufficiency.