Influence factors of metagenomic next-generation sequencing negative results in diagnosed patients with spinal infection.

The aim of this study was to evaluate the influence factors of metagenomic next-generation sequencing (mNGS) negative results in the diagnosed patients with spinal infection. mNGS test was applied in a cohort of 114 patients with suspected spinal infection, among which 56 patients had a final diagnosis of spinal infection. mNGS achieved a sensitivity of 75.0% (95% CI, 61.6% to 85.6%) and a specificity of 84.5% (95% CI, 72.6% to 92.7%), using histopathology and culture results as reference. Diagnosed patients with a negative culture result had lower white blood cell account, percentage of neutrophilic granulocyte, C-reactive protein (all P<0.05) and relatively higher rate of prior antimicrobial treatment history (P=0.059). However, diagnosed patients with a negative mNGS result did not have such difference with mNGS-positive patients, suggesting that mNGS was not strictly limited by the above indicators, which presented the advantages of this technique from another point of view.

Anat-SFSeg: Anatomically-guided superficial fiber segmentation with point-cloud deep learning.

Diffusion magnetic resonance imaging (dMRI) tractography is a critical technique to map the brain's structural connectivity. Accurate segmentation of white matter, particularly the superficial white matter (SWM), is essential for neuroscience and clinical research. However, it is challenging to segment SWM due to the short adjacent gyri connection in a U-shaped pattern. In this work, we propose an Anatomically-guided Superficial Fiber Segmentation (Anat-SFSeg) framework to improve the performance on SWM segmentation. The framework consists of a unique fiber anatomical descriptor (named FiberAnatMap) and a deep learning network based on point-cloud data. The spatial coordinates of fibers represented as point clouds, as well as the anatomical features at both the individual and group levels, are fed into a neural network. The network is trained on Human Connectome Project (HCP) datasets and tested on the subjects with a range of cognitive impairment levels. One new metric named fiber anatomical region proportion (FARP), quantifies the ratio of fibers in the defined brain regions and enables the comparison with other methods. Another metric named anatomical region fiber count (ARFC), represents the average fiber number in each cluster for the assessment of inter-subject differences. The experimental results demonstrate that Anat-SFSeg achieves the highest accuracy on HCP datasets and exhibits great generalization on clinical datasets. Diffusion tensor metrics and ARFC show disorder severity associated alterations in patients with Alzheimer's disease (AD) and mild cognitive impairments (MCI). Correlations with cognitive grades show that these metrics are potential neuroimaging biomarkers for AD. Furthermore, Anat-SFSeg could be utilized to explore other neurodegenerative, neurodevelopmental or psychiatric disorders.

Exosomes Derived from Caerulein-Stimulated Pancreatic Acinar Cells Mediate Peritoneal Macrophage M1 Polarization and Pyroptosis via an miR-24-3p/MARCH3/NLRP3 Axis in Acute Pancreatitis.

OBJECTIVES: Acute pancreatitis (AP) has a high incidence of hospitalizations, morbidity, and mortality worldwide. A growing number of studies on AP pathogenesis are based on caerulein-induced experimental model, which simulates human AP in vivo. It has been demonstrated that both pancreatic acinar cells and peritoneal macrophages are involved in pancreatic inflammation and damage. However, their connection has not been well understood. METHODS: A caerulein-induced AP model was established on the pancreatic acinar cell line AR42J. Rat macrophages were isolated from the peritoneal cavity. The effects of caerulein-induced pancreatic exosomes on the peritoneal macrophage and pancreas in vivo and in vitro were examined. The underlying molecular mechanism was investigated by exploring the regulatory role of downstream molecules. RESULTS: We found that exosomes derived from caerulein-treated AR42J cells induced rat peritoneal macrophage M1 polarization and pyroptosis. miR-24-3p was upregulated in caerulein-stimulated exosomes, whereas the miR-24-3p inhibitor counteracted the effect of pancreatic exosomes on peritoneal macrophage M1 polarization and pyroptosis. Furthermore, miR-24-3p inhibited March3 expression, whereas MARCH3 mediated NLRP3 ubiquitination in rat peritoneal macrophages, which, in turn, contributed to the apoptosis, reactive oxygen species production, and inflammation in AR42J cells. CONCLUSIONS: Exosomes derived from caerulein-stimulated pancreatic acinar cells mediate peritoneal macrophage M1 polarization and pyroptosis via an miR-24-3p/MARCH3/NLRP3 axis in AP.

A Novel Serum Exosomal miRNA Signature in the Early Prediction of Persistent Organ Failure in Patients with Acute Pancreatitis.

OBJECTIVE: Current study aims to investigate whether serum exosomal microRNAs (miRNAs) could be potential biomarkers in predicting APs with POF at early phase. BACKGROUND: Novel biomarkers are sorely needed for early prediction of persistent organ failure (POF) in acute pancreatitis (AP) patients. METHODS: In the discovery stage, exosomal miRNAs were profiled in sera from APs with or without POF (5 vs. 5) using microarrays. POF-associated miRNA signatures then were assessed in training cohort (n=227) and further validated in three independent cohorts (n=516), including one nested case-control cohort. RESULTS: A total of 743 APs were recruited in this large-scale biomarker identification study with a nested case-control study. Data from the discovery cohort demonstrated that 90 exosomal miRNAs were significantly dysregulated in APs with POF compared with controls. One miRNA classifier (Cmi) comprising 3 miRNAs (miR-4265, 1208, 3127-5p) was identified in the training cohort, and was further evaluated in two validation cohorts for their predictive value for POF. AUCs for Cmi ranged from 0.88 to 0.90, which was statistically superior to AUCs of APACHE-II and BISAP, and outperformed BUN and creatinine in POF prediction across all cohorts (P<.05). Higher levels of Cmi indicated increased need for ICU admission, prolonged hospitalization, and elevated mortality rate, thus poor prognosis. In the nested case-control study, Cmi could help identify prediagnostic POF in post-ERCP pancreatitis cases within "golden hours" after ERCP with high efficacy. CONCLUSIONS: Serum exosomal Cmi may be an early predictor for POF in AP, even within "golden hours" after AP onset. TRIAL REGISTRATION: ClinicalTrials.gov (NCT02602808).

Benefit from Almonertinib after Osimertinib treat EGFR 19 exon deletion NSCLC induced Severe rash: a case report.

Osimertinib, a third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), has been recommended as a first-line treatment of EGFR-positive non-small cell lung cancer (NSCLC). Skin rash is one of the most common side effects of osimertinib, and can have an impact on patients' quality of life and follow-up. However, there are few reports on the safety and efficacy of switching therapy with osimertinib and the other three generations of TKIs. In this paper, we present a case of NSCLC with an EGFR exon 19 deletion (19del) and MET gene amplification who developed a severe rash after 2 months of treatment with osimertinib that did not recur after switching to replacement therapy with aumonertinib. Our findings indicate that aumonertinib is as effective as osimertinib in treating EGFR19del, while also exhibiting a lower occurrence of adverse skin reactions. This may result in an improved quality of life for patients.

Comprehensive expression analysis reveals several miRNAs against acute pancreatitis via modulating autophagy.

Acute pancreatitis (AP) had been one of the main reasons for hospitalization worldwide. However, the mechanisms related to AP remained to be unclear. This study identified 37 miRNAs and 189 mRNAs were differentially expressed in pancreatitis and normal samples. Bioinformatics analysis showed DEGs were significantly related to PI3K-Akt signaling, FoxO signaling, Oocyte meiosis, Focal adhesion, and Protein digestion and absorption. By constructing a signaling-DEGs regulation network, we found COL12A1, DPP4, COL5A1, COL5A2, and SLC1A5 were related to regulating Protein digestion and absorption, THBS2, BCL2, NGPT1, EREG, COL1A1 were related to regulating PI3K signaling, CCNB1, CDKN2B, IRS2, PLK2 were related to modulating FOXO signaling. Next, we constructed 1 miRNA-mRNA regulation network in AP, consisting of 34 miRNAs and 96 mRNAs. The protein-protein interaction networks and the miRNA-targets networks analysis show that hsa-miR-199a-5p, hsa-miR-150, hsa-miR-194, COL6A3 and CNN1 acted as hub regulators in AOf note, through comprehensive expression analysis, we found several miRNAs and mRNAs were significantly related to modulating autophagy signaling in AP, including hsa-miR-181c, hsa-miR-181d, hsa-miR-181b, hsa-miR-379 and hsa-miR-199a-5Overall, this study screening differently expressed miRNAs in AP and revealed miRNA- autophagy regulation may serve as a potential prognosis and Therapeutic marker for AP.

Comparison of postoperative recovery of primary pterygium excision combined with either limbal stem cell transplantation or amniotic membrane transplantation: a randomized controlled trial-based meta-analysis.

OBJECTIVE: To compare the postoperative recovery of primary pterygium excision combined with either limbal stem cell transplantation (LSCT) or amniotic membrane transplantation (AMT). METHODS: All relevant studies on the primary pterygium excision combined with either LSCT or AMT conducted before August 2022 were extracted from PubMed, EMBASE, Web of Science, and Cochrane Library databases. The main outcomes compared were tear film stability at 1, 3, and 6 months after surgery, postoperative corneal epithelial healing time, recurrence rate, and complications. RESULTS: Sixteen randomized controlled trials (RCTs) with 1390 eye cases were included in this meta-analysis. We found that patients of the AMT group improved significantly in the results of the tear break-up time (BUT) and Schirmer I test at 1 month after surgery (BUT: MD=-0.37, 95% CI: -0.62, -0.12, P<0.05; Schirmer I test: MD=-0.32, 95% CI: -0.57, -0.07, P<0.05) compared with those of the LSCT group, suggesting that the early stage of tear film stability after primary pterygium excision combined with AMT was superior to the LSCT combination. However, according to the Schirmer I test result, the patients in the LSCT group showed increased tear production compared to the AMT group at 3 and 6 months after surgery (3 months: MD=0.36, 95% CI: 0.08, 0.64, P<0.05; 6 months: MD=0.33, 95% CI: 0.07, 0.60, P<0.05), suggesting that the LSCT combination was superior to the AMT combination in long-term postoperative tear film stability. As for postoperative corneal epithelial healing time, the LSCT group exhibited shorter time than the AMT group (MD=-1.17, 95% CI: -2.15, -0.19, P<0.05). Furthermore, the recurrence rate was lower in the LSCT group than in the AMT group (RR=0.42, 95% CI: 0.30, 0.59, P<0.05). Lastly, there was no statistical difference in BUT and complication rate at 3 and 6 months after surgery between the LSCT and AMT groups. CONCLUSIONS: Our analysis suggests that primary pterygium excision combined with LSCT may be a better choice compared to the combination with AMT in postoperative recovery.

Radiation necrosis and therapeutic outcomes in patients treated with linear accelerator-based hypofractionated stereotactic radiosurgery for intact intracranial metastases.

INTRODUCTION: Balancing disease control and treatment-related toxicities can be challenging when treating higher-risk brain metastases (BMs) that are larger in size or eloquent anatomical locations. Hypofractionated stereotactic radiosurgery (hfSRS) is expected to offer superior or equal efficacy with lower toxicity profile compared with single-fraction SRS (sfSRS). We report the efficacy and toxicity profiles of hfSRS in a consecutive cohort of patients to support this predicted benefit from hfSRS for high-risk BMs. METHODS: We retrospectively analysed 185 consecutive individual lesions from 152 patients with intact BMs treated with hfSRS between 1 July 2016 and 31 October 2019 and followed up to 30 April 2022 with serial brain magnetic resonance imaging (MRI). The primary endpoint was the event of radiation necrosis (RN). Local control (LC) rate and distant brain failure (DBF) were reported as secondary outcomes. Kaplan-Meier method was used to report the cumulative incidence of RN and overall survival and the incidence of DBF. Potential risk factors for RN were assessed using univariable Cox regression analysis. RESULTS: The median follow-up was 38.0 months, and the median survival post-SRS was 9.5 months. The cumulative incidence rate of RN was 13.2% (95% CI: 7.0-24.7%), and 18.1% of patients with confirmed RN were symptomatic. Higher mean dose delivered to planning target volume (PTV) (HR 1.22, 95% CI: 1.05-1.42, P = 0.01), higher mean BED10 (biological equivalent dose assuming a tissue α / ß ratio of 10) (HR 1.12, 95% CI: 1.04-1.2, P < 0.001), and higher mean BED2 (HR 1.02, 95% CI: 1-1.04, P = 0.04) delivered to the lesion was associated with increased risk of RN. LC rate was 86% and the cumulative incidence of DBF was 36% with a median onset of 28.4 months. CONCLUSIONS: Our results support the predicted radiobiological benefit of the use of hfSRS in high-risk BMs to limit treatment-related toxicity with low risk for symptomatic RN comparable with lower risk population receiving sfSRS while achieving satisfactory local disease control.

Procyanidin B2 alleviates oxidative stress-induced nucleus pulposus cells apoptosis through upregulating Nrf2 via PI3K-Akt pathway.

Oxidative stress can lead to nucleus pulposus cell (NPC) apoptosis, which is considered to be one of the main contributors to intervertebral disc degeneration (IVDD). Procyanidin B2 is a natural antioxidant that protects against oxidative stress. However, whether procyanidin B2 protects NPCs from oxidative stress remains unknown. In this study, we demonstrated that procyanidin B2 could reduce tert-butyl hydroperoxide-induced reactive oxygen species in rat NPCs and attenuate rat NPC apoptosis. Further experiments revealed that procyanidin B2 upregulated the expression of both nuclear factor erythroid 2-related factor 2 (Nrf2) and phosphorylation of protein kinase B (Akt). We then used silencing of Nrf2 and LY294002 to silence Nrf2 expression and block the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, respectively, and found that the protective roles of procyanidin B2 in NPCs were inhibited. Therefore, we demonstrated that procyanidin B2 alleviated rat NPC apoptosis induced by oxidative stress by upregulating Nrf2 via activation of the PI3K/Akt signaling pathway. This study provides a potential therapeutic approach for procyanidin B2 in IVDD, which might help in the development of new drugs for IVDD treatment.

Scutellarin-mediated autophagy activates exosome release of rat nucleus pulposus cells by positively regulating Rab8a via the PI3K/PTEN/Akt pathway.

To provide a basis for promising exosome-based therapies against intervertebral disc degeneration (IDD), our present research aimed to identify a mechanism underlying the vesicle release from nucleus pulposus cells (NPCs). Scutellarin (SC) is a natural chemotherapeutic agent isolated from Erigeron breviscapus with a variety of biological activities. Here, we observed the significantly elevated autophagy levels in rat NPCs under the stimulation of SC, leading to a concomitant enhancement of intracellular vesicle release, which could be attributed to the inactivation of the phosphoinositide 3-kinase (PI3K)/phosphatase and tensin homolog (PTEN)/protein kinase B (Akt) pathway. To ensure that exosome release was driven by SC via the autophagic pathway, we implemented gain-of-function and loss-of-function studies by additionally using insulin-like growth factor-1 (IGF-1) and small-interfering RNA of autophagy-related gene 5 (ATG5), and the exosome secretion decreased in the case of attenuated autophagy. Evidently, the treatment with SC exerted the remarkable upregulation of Rab8a through the overexpression of ATG5. After the respective knockdown of ATG5 and Rab8a, the increased release of exosomes induced by SC was reversed, whereas the number of intracellular vesicles was restored. Overall, it can be concluded that SC contributes to the autophagy activation in NPCs by acting on the PI3K/PTEN/Akt pathway, which upregulates the expression of Rab8a and promotes the release of exosomes, inspiring novel therapeutic strategies in preventing IDD that might be fruitfully investigated.

Cross-Modal Interaction and Integration Through Stimulus-Specific Adaptation in the Thalamic Reticular Nucleus of Rats.

Stimulus-specific adaptation (SSA), defined as a decrease in responses to a common stimulus that only partially generalizes to other rare stimuli, is a widespread phenomenon in the brain that is believed to be related to novelty detection. Although cross-modal sensory processing is also a widespread phenomenon, the interaction between the two phenomena is not well understood. In this study, the thalamic reticular nucleus (TRN), which is regarded as a hub of the attentional system that contains multi-modal neurons, was investigated. The results showed that SSA existed in an interactive oddball stimulation, which mimics stimulation changes from one modality to another. In the bimodal integration, SSA to bimodal stimulation was stronger than to visual stimulation alone but similar to auditory stimulation alone, which indicated a limited integrative effect. Collectively, the present results provide evidence for independent cross-modal processing in bimodal TRN neurons.

Hesitate between confocal laser endomicroscopy and narrow-band imaging: how to choose a better method in the detection of focal precancerous state of gastric cancer.

BACKGROUND: With a high incidence globally, deaths form gastric cancer (GC) are not rare. Early diagnosis is crucial to ameliorate its prognosis. Confocal laser endomicroscopy (CLE) and narrow band imaging (NBI) have been extensively applied in gastroscopy, particularly when it comes to the detection and management of premalignant gastric lesion. Our meta-analysis intends to appraise the diagnostic capability and compare the efficacy of NBI and CLE for focal precancerous state of gastric cancer. METHODS: We performed a literature search up to November 5, 2020 in online databases and major conferences. Two investigators assessed the methodological bias by QUADAS-2, followed by sophisticated study selection and data exaction to make a comparison between sensitivity, specificity, positive and negative likelihood values, and diagnostic odds ratio. A symmetric summary receiver-operating curve (sROC) and its area under the curve (AUC) were used to estimate threshold effect. Additionally, we evaluated the publication bias by Deeks' asymmetry test. RESULTS AND CONCLUSIONS: Four studies involved 248 patients and 526 lesions. In analysis drawn from every lesion, the NBI's pooled sensitivity and specificity were 87% (95% CI: 0.80-0.92) and 85% (95% CI: 0.75-0.91), and those of CLE were 90% (95% CI: 0.85-0.91) and 87% (95% CI: 0.83-0.91). CLE illustrated that the pooled two were slightly higher than NBI when compared at the level of every lesion. The AUC for NBI and CLE was 0.92 (0.90-0.94) and 0.95 (0.92-0.96), and there might be a threshold effect, according to the shoulder-like distribution of scatter points in the sROC. We did not find obvious publication bias in our meta-analysis.

Long non-coding RNA FAM83A antisense RNA 1 (lncRNA FAM83A-AS1) targets microRNA-141-3p to regulate lung adenocarcinoma cell proliferation, migration, invasion, and epithelial-mesenchymal transition progression.

The current paper investigates how long non-coding RNA (lncRNA) FAM83A antisense RNA 1 (lncRNA FAM83A-AS1) affected the epithelial-mesenchymal transformation (EMT), growth, invasion and migration of lung adenocarcinoma (LUAD) via targeting miRNA-141-3p. The GEPIA and ENCORI databases were used to analyze differences in lncRNA FAM83A-AS1 levels within LUAD samples. FAM83A-AS1 and miR-141-3p levels were assessed using qRT-PCR among 30 LUAD samples and surrounding normal tissues. In addition, we analyzed how FAM83A-AS1 affected proliferation, invasion, migration, and EMT processes of LUAD cells by targeting miR-141-3p through EdU, CCK-8 assay, scratch assay, transwell migration and invasion assay, immunofluorescence (IF) staining and WB assay. MicroRNAs targeting FAM83A-AS1 were screened using AnnoLnc2 and identified by RT-qPCR. Dual-luciferase assays were utilized to evaluate the connection between FAM83A-AS1 and miR-141-3p. FAM83A-AS1 expression was remarkably raised in lung cancer cells and tissue samples; however, miR-141-3p level markedly reduced relative to healthy samples. FAM83A-AS1 silencing suppressed EMT, growth, invasion and migration of LUAD cells. MiR-141-3p was the possible FAM83A-AS1 binding target negatively associated with FAM83A-AS1. The miR-141-3p inhibitor partly abolished the FAM83A-AS1 knockdown-induced inhibition on EMT, cell growth, invasion and migration in LUAD cells. In addition, miR-141-3p down-regulation abolished the inhibition of E-box-bound zinc finger protein 1 and 2 protein production following FAM83A-AS1 knockdown. According to our results, FAM83A-AS1/miR-141-3p axis plays an important role in LUAD occurrence and development. FAM83A-AS1 sponged miR-141-3p to down-regulate the level of the latter within LUAD and thereby encouraging LUAD development and suggesting a possible novel therapeutic approach for LUAD.

Phase separation ability and phosphatase activity of the SHP1-R360E mutant.

SHP1 is a non-receptor protein tyrosine phosphatase that is widely expressed in hematopoietic cells such as white blood cells, neutrophils, and immune cells. SHP1 can regulate the occurrence and differentiation of immune cells and plays an important role as a tumor suppressor. Previous studies have suggested that SHP2, the homologous protein of phosphatase SHP1, can undergo liquid-liquid phase separation (LLPS). Therefore, in this study, we investigated if SHP1 is also capable of LLPS. To the best of our knowledge, our study is the first to reveal that SHP1 has the ability to undergo LLPS. In addition, we identified an important residue, SHP1-R360E, that can completely inhibit the LLPS ability of SHP1, but this mutation has no remarkable effect on SHP1's enzymatic activity. This allows us to explore the phosphatase activity and phase separation ability of SHP1 separately, providing a basis for future exploration of the phase separation mechanism of phosphatases.

Medical students' perceptions and expectations regarding digital health education and training: A qualitative study.

INTRODUCTION: Digital health - the convergence of digital technologies within health and health care to enhance the efficiency of health-care delivery - is fast becoming an integral part of routine medical practice. The integration of digital health into traditional practice brings significant changes. Logic dictates that for medical practitioners to operate in this new digitally enabled environment, they require specific knowledge, skills and competencies relating to digital health. However, very few medical programmes in Australia and globally include digital health within their regular curriculum. This pilot study aimed to explore medical students' perceptions and expectations of digital health education and training (ET). METHODS: An online survey and focus groups were used to collect information about medical students' perceptions and expectations relating to digital health and ET relating to this field within the medical programme at the University of Queensland. Sixty-three students took part in the survey, and 17 students were involved in four focus groups. RESULTS: Most participants had no formal ET in digital health. Most participants (n = 43; 68%) expressed a willingness to learn about digital health as part of their medical programme. DISCUSSION: Primarily, knowledge- and practice-related factors have motivated students to learn about digital health. The analysis of focus group data identified two superordinate themes: (a) drivers of digital health ET and (b) expectations relating to digital health ET. Students agreed that digital health is a relevant field for their future practice that should be taught as part of their regular curriculum.

A system dynamics model to simulate the water-energy-food nexus of resource-based regions: A case study in Daqing City, China.

Resource-based regions (RBRs) have made significant contributions to the social and economic development of nations. The long-term and high-intensity development of resources puts tremendous pressure on water, energy and food resources and the ecological environment. Exploring the water-energy-food nexus (WEF nexus, WEFN) of RBRs is key to making informed decisions about regional sustainable development. In this study, a feedback model for the WEFN of RBRs was developed using a system dynamics approach. The WEFN model not only describes the WEFN system from both the supply and demand sides, but also classifies WEF resources. Using Daqing, China, as a case study, five future scenarios were designed to explore the impacts of real policies designed by different government departments on the WEFN system. Comparing the predicted results of a scheme for business as usual, a scheme for developing bioenergy, a scheme for adjustment of the production structure, a scheme for strengthening the development of water and food resources and a scheme for saving WEF resources revealed that the schemes for adjustment of the production structure and for saving WEF resources will not only improve the security of WEF resources, but also reduce pollution of the water environment by human activities, which is conducive to improving the overall benefits of the WEFN system. Finally, some practical suggestions are put forward to promote the coordinated development of the WEFN system. The WEFN model is a multi-centric tool for integrated resources management, and can be expanded to other RBRs and provides scientific support for decision-makers.

Excessive mechanical strain accelerates intervertebral disc degeneration by disrupting intrinsic circadian rhythm.

Night shift workers with disordered rhythmic mechanical loading are more prone to intervertebral disc degeneration (IDD). Our results showed that circadian rhythm (CR) was dampened in degenerated and aged NP cells. Long-term environmental CR disruption promoted IDD in rats. Excessive mechanical strain disrupted the CR and inhibited the expression of core clock proteins. The inhibitory effect of mechanical loading on the expression of extracellular matrix genes could be reversed by BMAL1 overexpression in NP cells. The Rho/ROCK pathway was demonstrated to mediate the effect of mechanical stimulation on CR. Prolonged mechanical loading for 12 months affected intrinsic CR genes and induced IDD in a model of upright posture in a normal environment. Unexpectedly, mechanical loading further accelerated the IDD in an Light-Dark (LD) cycle-disrupted environment. These results indicated that intrinsic CR disruption might be a mechanism involved in overloading-induced IDD and a potential drug target for night shift workers.

TNF-α-stimulated nucleus pulposus cells induce cell apoptosis through the release of exosomal miR-16 targeting IGF-1 and IGF-1R in rats.

BACKGROUND: Exosomes may contain excess cellular components released by cells in response to harmful external stimuli to maintain cellular homeostasis. Inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α), can induce cell apoptosis, alter cellular component expression levels, and stimulate exosome release. In this study, we examined whether exosomes released from nucleus pulposus cells (NPCs) under inflammatory conditions could induce normal NP cell apoptosis in rats and its underlining mechanism. METHODS: Exosomes were isolated from TNF-α-treated NPCs and used to treat normal NPCs. The effects were assessed by flow cytometry and western blot analysis. Anti-apoptotic insulin-like growth factor-1 (IGF-1) expression in NPCs was assessed by western blot analysis. Given the exosomal miRNAs might be the key factors of exosomes, bioinformatics approaches and quantitative real-time polymerase chain reaction (qRT-PCR) were used to identify IGF-1-regulating micro RNAs (miRNAs), including miR-16. Luciferase reporter assay assessed miR-16 regulation of IGF-1 and IGF-1 receptor (IGF-1R). NPCs were transfected with miR-16 mimic, and exosomes were applied to normal NPCs. NPCs were pretreated with 10 ng/mL TNF-α, transfected with miR-16 inhibitors, and the exosomes were isolated. Cell and exosome miR-16 levels were detected by qRT-PCR. Western blot analysis determined IGF-1, IGF-1R, and apoptotic marker levels in exosome-treated NPCs. RESULTS: Exosomes from TNF-α-treated NPCs induced apoptosis in normal NPCs and repressed IGF-1 expression. Exosomal miR-16 regulated IGF-1 and induced NPC apoptosis. The dual-luciferase reporter assay revealed that miR-16 binds the 3' untranslated regions (3'-UTRs) of IGF-1 and IGF-1R. Exosomal miR-16 repressed IGF-1 and the IGF-1R/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway which therefore induced NPC apoptosis. Rescue experiments using miR-16 inhibitors further validated these findings. CONCLUSIONS: The inflammatory factor TNF-α stimulated exosome release from NPCs, which induced the apoptosis of normal NPCs through the actions of exosomal miR-16. Exosomal miR-16 directly repressed the anti-apoptotic IGF-1/IGF-1R pathway, increasing the apoptosis of NPCs.

Injectable Mussel-Inspired highly adhesive hydrogel with exosomes for endogenous cell recruitment and cartilage defect regeneration.

In the early stage of osteoarthritis (OA), cartilage degradation in the surface region leads to superficial cartilage defect. However, enhancing the regeneration of cartilage defect remains a great challenge for existing hydrogel technology because of the weak adhesion to wet tissue. In the present study, an injectable mussel-inspired highly adhesive hydrogel with exosomes was investigated for endogenous cell recruitment and cartilage defect regeneration. The hydrogel with high bonding strength to the wet surface was prepared using a crosslinked network of alginate-dopamine, chondroitin sulfate, and regenerated silk fibroin (AD/CS/RSF). Compared with commercial enbucrilate tissue adhesive, the AD/CS/RSF hydrogel provided a comparative lap shear strength of 120 kPa, with a similar gelation time and a higher capacity for maintaining adhesive strength. The AD/CS/RSF/EXO hydrogel with encapsulated exosomes recruited BMSCs migration and inflation, promoted BMSCs proliferation and differentiation. Most importantly, the AD/CS/RSF/EXO hydrogel accelerated cartilage defect regeneration in situ, and extracellular matrix remodeling after injection in rat patellar grooves. The exosomes released by the hydrogels could recruit BMSCs into the hydrogel and neo-cartilage via the chemokine signaling pathway. Our findings reveal an injectable and adhesive hydrogel for superficial cartilage regeneration, which is a promising approach for minimally treating cartilage defect with arthroscopic assistance.