Alternative 3'-untranslated regions regulate high-salt tolerance of Spartina alterniflora.

High-salt stress continues to challenge the growth and survival of many plants. Alternative polyadenylation (APA) produces mRNAs with different 3'-untranslated regions (3' UTRs) to regulate gene expression at the post-transcriptional level. However, the roles of alternative 3' UTRs in response to salt stress remain elusive. Here, we report the function of alternative 3' UTRs in response to high-salt stress in S. alterniflora (Spartina alterniflora), a monocotyledonous halophyte tolerant of high-salt environments. We found that high-salt stress induced global APA dynamics, and ∼42% of APA genes responded to salt stress. High-salt stress led to 3' UTR lengthening of 207 transcripts through increasing the usage of distal poly(A) sites. Transcripts with alternative 3' UTRs were mainly enriched in salt stress-related ion transporters. Alternative 3' UTRs of HIGH-AFFINITY K+ TRANSPORTER 1 (SaHKT1) increased RNA stability and protein synthesis in vivo. Regulatory AU-rich elements were identified in alternative 3' UTRs, boosting the protein level of SaHKT1. RNAi-knock-down experiments revealed that the biogenesis of 3' UTR lengthening in SaHKT1 was controlled by the poly(A) factor CLEAVAGE AND POLYADENYLATION SPECIFICITY FACTOR 30 (SaCPSF30). Over-expression of SaHKT1 with an alternative 3' UTR in rice (Oryza sativa) protoplasts increased mRNA accumulation of salt-tolerance genes in an AU-rich element-dependent manner. These results suggest that mRNA 3' UTR lengthening is a potential mechanism in response to high-salt stress. These results also reveal complex regulatory roles of alternative 3' UTRs coupling APA and regulatory elements at the post-transcriptional level in plants.

Emodin ameliorates matrix degradation and apoptosis in nucleus pulposus cells and attenuates intervertebral disc degeneration through LRP1 in vitro and in vivo.

Low back pain (LBP) is the leading cause of disability worldwide, with a strong correlation to intervertebral disc degeneration (IDD). Inflammation-induced extracellular matrix (ECM) degradation plays a major role in IDD's progression. Emodin, known for its anti-inflammatory effects and ability to inhibit ECM degradation in osteoarthritis, but its role in IDD is unclear. Our study aimed to explore emodin's role and mechanisms on IDD both in vivo and in vitro. We discovered that emodin positively regulated anabolic markers (COL2A1, aggrecan) and negatively impacted catabolic markers (MMP3, MMP13) in nucleus pulposus cells, while also inhibiting cell apoptosis under inflammation environment. We revealed that emodin inhibits inflammation-induced NF-ĸB activation by suppressing the degradation of LRP1 via the proteasome pathway. Additionally, LRP1 was validated as essential to emodin's regulation of ECM metabolism and apoptosis, both in vitro and in vivo. Ultimately, we demonstrated that emodin effectively alleviates IDD in a rat model. Our findings uncover the novel pathway of emodin inhibiting ECM degradation and apoptosis through the inhibition of NF-κB via LRP1, thus alleviating IDD. This study not only broadens our understanding of emodin's role and mechanism in IDD treatment but also guides future therapeutic interventions.

Incomplete histologic healing and diminished biomechanical strength of meniscus-bone interface after medial meniscus posterior root transosseous repair in a goat model.

PURPOSE: To enhance the understanding of histological healing after repairing medial meniscal posterior root tear (MMPRT) at an early stage, utilizing a goat model. METHODS: Eighteen adult goats, totaling thirty-six knee joints, were allocated into three groups (n = 12): Sham group (Sham), Root Tear group (RT), and Root Tear with Transosseous Suture group (RTS). At 12- and 24-week intervals post-surgery, all the knees were harvested for imaging, macroscopic, histological, and biomechanical assessments. RESULTS: The intact root served as a meniscus-bone interface which connected the tibial and the circular fibers of the meniscus, with a bony insertion and a root-meniscus transition. A direct-fibrous-connection displayed at the bony insertion proximal to the synovium in the RTS group, while the remaining regions of the root displayed indirect-fibrous healing. The healing in the RT group was disjointed and reminiscent of scar tissue. The RTS group exhibited a more pronounced coronal extrusion compared to the Sham group (0.42 ± 0.09 vs. 0.19 ± 0.02, P = 0.0012) but was improved relative to that of the RT group (0.49 ± 0.02, P = 0.0028). The failure load and stiffness of the RTS group were notably higher than those of the RT group, with a strength of 42.67% and a stiffness of 83.75% of the intact root. All the samples ruptured at the root-meniscus transitions. CONCLUSION: The incomplete healing may be attributed to the histological factors underlying the low healing rate and persistent MME. Notably, the region attached to the posterior-cruciate-ligament exhibited superior healing compared to other regions of the bony insertion in the repaired group. Conversely, the root-meniscus transition displayed discontinuity, representing a mechanical weakness in the healing process. CLINICAL RELEVANCE: Modifications of bone tunnel positioning and suture placement could be undertaken in subsequent studies to particularly enhance the healing of the root-meniscus transition.

Correlation between posterior paraspinal muscle atrophy and lumbar intervertebral disc degeneration in patients with chronic low back pain.

BACKGROUND: Although enormous studies have been devoted to solving the problem of intervertebral disc degeneration/herniation, little attention is paid to the effect of paraspinal muscles on it. We aimed to investigate the correlation between paraspinal muscle atrophy and lumbar disc degeneration to recognize paraspinal muscle atrophy and its importance to the spine. PATIENTS AND METHODS: A total of 107 patients were enrolled in the study (65 females, 42 males; age 50.87 ± 15.391 years old). Cross-sectional area, functional cross-sectional area, and fatty infiltration of the posterior paraspinal muscles were measured at the level of L4/5, and the degree of facet joint degeneration was evaluated at the levels of L3/4, L4/5, and L5/S1 by MRI. After controlling the confounding factors by multiple linear regression, the correlations among paraspinal muscle atrophy, disc degeneration, and facet joint degeneration were analyzed. Meanwhile, Pearson/Spearson rank analysis was used to analyze the correlation between clinical symptoms (VAS and ODI) and paraspinal muscle atrophy. RESULTS: There was a strong correlation between paraspinal muscle atrophy and disc degeneration after controlling the confounding factors (p < 0.05, R > 0.5). There was a weak correlation between paraspinal muscle atrophy and facet joint degeneration (p < 0.05, R < 0.5). There was a significant correlation between facet joint degeneration and intervertebral disc degeneration (p < 0.05, R > 0.7). The fatty infiltration of paraspinal muscle was weakly correlated with ODI (p < 0.05, R < 0.3), but VAS was not. CONCLUSIONS: The degree of paraspinal muscle atrophy increased with lumbar disc degeneration and facet joint degeneration and fatty infiltration of multifidus was more susceptible to weight.

Radiologically anatomic measurement analysis for the third sacral foramen and an efficient implantation protocol for sacral neuromodulation.

AIMS: To define radiologically anatomic measurements for the S3 foramen and develop an efficient and straightforward implantation protocol for sacral neuromodulation (SNM) in a southern Chinese population. METHODS: We retrospectively reviewed 793 adults without sacrococcygeal abnormalities or bony injury. We generated CT-derived three-dimensional volumetric models and collected radiologically anatomic measurements of S3 foramen. A multiple generalized linear regression model was applied to evaluate implantation protocol for SNM. RESULTS: Height, weight, and sacral height were significantly different between enrolled patients based on sex (p < 0.001 for all), while age and sacral width were similar. The measurements L1, L2, A1, and A2 on both sides were similar, while based on sex, those of L1, A1, A2 showed significant differences [7.02 ± 0.90 (M) vs. 6.10 ± 0.99 cm (F) (p < 0.001); 83.05 ± 6.23 (M) vs. 82.08 ± 7.53° (F) (p = 0.005); and 13.31 ± 5.69 (M) vs. 11.97 ± 4.91° (F) (p < 0.001), respectively]. The multiple generalized linear regression model demonstrated that sex was a common independent factor for estimating L1, L2, A1, A2, while the consistency rate between the estimated model and actual measurements was poor. The measurements L1, L2, A1, and A2 were approximately 7.0 cm (M) versus 6.1 cm (F), 1.8 cm (M) versus 1.8 cm (F), 83° (M) versus 82° (F), and 13° (M) versus 12° (F), respectively. CONCLUSIONS: This retrospective study indicates that the radiologically anatomic measurements for the S3 foramen have unique characteristics. Our study provides an efficient and straightforward implantation protocol for SNM, improving its use in China.

Preoperative prediction of axillary sentinel lymph node burden with multiparametric MRI-based radiomics nomogram in early-stage breast cancer.

OBJECTIVES: To develop and validate a multiparametric MRI-based radiomics nomogram for pretreatment predicting the axillary sentinel lymph node (SLN) burden in early-stage breast cancer. METHODS: A total of 230 women with early-stage invasive breast cancer were retrospectively analyzed. A radiomics signature was constructed based on preoperative multiparametric MRI from the training dataset (n = 126) of center 1, then tested in the validation cohort (n = 42) from center 1 and an external test cohort (n = 62) from center 2. Multivariable logistic regression was applied to develop a radiomics nomogram incorporating radiomics signature and predictive clinical and radiological features. The radiomics nomogram's performance was evaluated by its discrimination, calibration, and clinical use and was compared with MRI-based descriptors of primary breast tumor. RESULTS: The constructed radiomics nomogram incorporating radiomics signature and MRI-determined axillary lymph node (ALN) burden showed a good calibration and outperformed the MRI-determined ALN burden alone for predicting SLN burden (area under the curve [AUC]: 0.82 vs. 0.68 [p < 0.001] in training cohort; 0.81 vs. 0.68 in validation cohort [p = 0.04]; and 0.81 vs. 0.58 [p = 0.001] in test cohort). Compared with the MRI-based breast tumor combined descriptors, the radiomics nomogram achieved a higher AUC in test cohort (0.81 vs. 0.58, p = 0.005) and a comparable AUC in training (0.82 vs. 0.73, p = 0.15) and validation (0.81 vs. 0.65, p = 0.31) cohorts. CONCLUSION: A multiparametric MRI-based radiomics nomogram can be used for preoperative prediction of the SLN burden in early-stage breast cancer. KEY POINTS: • Radiomics nomogram incorporating radiomics signature and MRI-determined ALN burden outperforms the MRI-determined ALN burden alone for predicting SLN burden in early-stage breast cancer. • Radiomics nomogram might have a better predictive ability than the MRI-based breast tumor combined descriptors. • Multiparametric MRI-based radiomics nomogram can be used as a non-invasive tool for preoperative predicting of SLN burden in patients with early-stage breast cancer.

Differentiation between high-grade gliomas and solitary brain metastases: a comparison of five diffusion-weighted MRI models.

BACKGROUND: To compare the diagnostic performance of neurite orientation dispersion and density imaging (NODDI), mean apparent propagator magnetic resonance imaging (MAP-MRI), diffusion kurtosis imaging (DKI), diffusion tensor imaging (DTI) and diffusion-weighted imaging (DWI) in distinguishing high-grade gliomas (HGGs) from solitary brain metastases (SBMs). METHODS: Patients with previously untreated, histopathologically confirmed HGGs (n = 20) or SBMs (n = 21) appearing as a solitary and contrast-enhancing lesion on structural MRI were prospectively recruited to undergo diffusion-weighted MRI. DWI data were obtained using a q-space Cartesian grid sampling procedure and were processed to generate parametric maps by fitting the NODDI, MAP-MRI, DKI, DTI and DWI models. The diffusion metrics of the contrast-enhancing tumor and peritumoral edema were measured. Differences in the diffusion metrics were compared between HGGs and SBMs, followed by receiver operating characteristic (ROC) analysis and the Hanley and McNeill test to determine their diagnostic performances. RESULTS: NODDI-based isotropic volume fraction (Viso) and orientation dispersion index (ODI); MAP-MRI-based mean-squared displacement (MSD) and q-space inverse variance (QIV); DKI-generated radial, mean diffusivity and fractional anisotropy (RDk, MDk and FAk); and DTI-generated radial, mean diffusivity and fractional anisotropy (RD, MD and FA) of the contrast-enhancing tumor were significantly different between HGGs and SBMs (p < 0.05). The best single discriminative parameters of each model were Viso, MSD, RDk and RD for NODDI, MAP-MRI, DKI and DTI, respectively. The AUC of Viso (0.871) was significantly higher than that of MSD (0.736), RDk (0.760) and RD (0.733) (p < 0.05). CONCLUSION: NODDI outperforms MAP-MRI, DKI, DTI and DWI in differentiating between HGGs and SBMs. NODDI-based Viso has the highest performance.

Quantitative dynamic contrast-enhanced MR imaging can be used to predict the pathologic stages of oral tongue squamous cell carcinoma.

BACKGROUND: To investigate whether quantitative dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) pharmacokinetic parameters can be used to predict the pathologic stages of oral tongue squamous cell carcinoma (OTSCC). METHODS: For this prospective study, DCE-MRI was performed in participants with OTSCC from May 2016 to June 2017. The pharmacokinetic parameters, including Ktrans, Kep, Ve, and Vp, were derived from DCE-MRI by utilizing a two-compartment extended Tofts model and a three-dimensional volume of interest. The postoperative pathologic stage was determined in each patient based on the 8th AJCC cancer staging manual. The quantitative DCE-MRI parameters were compared between stage I-II and stage III-IV lesions. Logistic regression analysis was used to determine independent predictors of tumor stages, followed by receiver operating characteristic (ROC) analysis to evaluate the predictive performance. RESULTS: The mean Ktrans, Kep and Vp values were significantly lower in stage III-IV lesions compared with stage I-II lesions (p = 0.013, 0.005 and 0.011, respectively). Kep was an independent predictor for the advanced stages as determined by univariate and multivariate logistic analysis. ROC analysis showed that Kep had the highest predictive capability, with a sensitivity of 64.3%, a specificity of 82.6%, a positive predictive value of 81.8%, a negative predictive value of 65.5%, and an accuracy of 72.5%. CONCLUSION: The quantitative DCE-MRI parameter Kep can be used as a biomarker for predicting pathologic stages of OTSCC.

Discrimination between benign and malignant breast lesions using volumetric quantitative dynamic contrast-enhanced MR imaging.

OBJECTIVE: To determine the diagnostic performance of volumetric quantitative dynamic contrast-enhanced MRI (qDCE-MRI) in differentiation between malignant and benign breast lesions. METHODS: DCE-MRI was performed in 124 patients with 136 breast lesions. Quantitative pharmacokinetic parameters Ktrans, Kep, Ve, Vp and semi-quantitative parameters TTP, MaxCon, MaxSlope, AUC were obtained by using a two-compartment extended Tofts model and three-dimensional volume of interest. Morphologic features (lesion size, margin, internal enhancement pattern) and time-signal intensity curve (TIC) type were also assessed. Logistic regression analysis was used to determine predictors of malignancy, followed by receiver operating characteristics (ROC) analysis to evaluate the diagnostic performance. RESULTS: qDCE parameters (Ktrans, Kep, Vp, TTP, MaxCon, MaxSlope and AUC), morphological parameters and TIC type were significantly different between malignant and benign lesions (P≤0.001). Multivariate logistic regression analyses showed that Ktrans, Kep, MaxSlope, size, margin and TIC type were independent predictors of malignancy. The diagnostic accuracy of logistic models based on qDCE parameters alone, morphological features plus TIC type, and all parameters combined was 94.9%, 89.0%, and 95.6% respectively. CONCLUSION: qDCE-MRI can be used to improve diagnostic differentiation between benign and malignant breast lesions in relation to morphology and kinetic analysis. KEY POINTS: • qDCE-MRI parameters are useful for discriminating between malignant and benign breast lesions. • K trans , K ep and MaxSlope were independent predictors of breast malignancy. • qDCE-MRI has a better diagnostic ability than morphology and kinetic analysis. • qDCE-MRI can be used to improve the diagnostic accuracy of breast malignancy.

DTI metrics can be used as biomarkers to determine the therapeutic effect of stem cells in acute peripheral nerve injury.

PURPOSE: To determine the role of diffusion tensor imaging (DTI) metrics as biomarkers for the therapeutic effects of mesenchymal stem cells (MSCs) in acute peripheral nerve injury. MATERIALS AND METHODS: Forty-four adult rats received subepineurial microinjection of MSCs (n = 22) or phosphate buffered saline (PBS, n = 22) 1 week after the sciatic nerve trunk crush injury. Sequential fat-suppressed T2-weighted imaging, T2 measurement, DTI and sciatic nerve functional assessment were performed at a 3.0 Tesla MR unit over an 8-week follow-up, with histological assessments performed at regular intervals. The sciatic nerve function index, T2 value, and DTI metrics, including fractional anisotropy (FA), axial diffusivity, radial diffusivity (RD), and mean diffusivity values of the distal stumps of crushed nerves were measured and compared between the two groups. RESULTS: Nerves treated with MSCs showed better functional recovery and exhibited more pronounced nerve regeneration compared with nerves treated with PBS. T2 values in nerves treated with MSCs or PBS showed a similar change pattern (P = 0.174), while FA and RD values in nerves treated with MSCs showed more rapid return (one week earlier) to baseline level than nerves treated with PBS (P = 0.045; 0.035). Nerves treated with MSCs had higher FA and lower RD values than nerves treated with PBS during the period from 2 to 3 weeks after surgery (P ≤ 0.0001, 0.004; P = 0.004, 0.006). CONCLUSION: FA and RD values derived from DTI might be used as sensitive biomarkers for detecting the therapeutic effect of stem cells in acute peripheral nerve crush injuries. LEVEL OF EVIDENCE: 2 J. Magn. Reson. Imaging 2017;45:855-862.

CT-guided nucleoplasty with radiofrequency energy for the treatment of lumbar disk herniation.

STUDY DESIGN: A clinical randomized controlled trial. OBJECTIVE: This study sought to compare the clinical effectiveness of CT-guided nucleoplasty, CT-guided nucleoplasty combined with nerve root injection, and CT-guided transforaminal lumbar epidural injections in treating patients with contained lumbar disk herniation and leg pain, which are caused by radicular encroachment. SUMMARY OF BACKGROUND DATA: Lumbar disk herniation is the most common cause of nerve root pain. The conservative treatment is proved to be effective for the majority of these patients, and the remaining patients are not ideal surgical candidates. Studies have found that minimally invasive percutaneous disk procedures may be preferable to open surgery in certain clinical situations. However, nucleoplasty in treating contained lumbar disk herniation and leg pain caused by radicular encroachment is still a controversy. DESIGN: A total of 97 patients with leg pain and MRI evidence of small-sized or medium-sized herniated disks correlating with the symptoms participated in the study. The patients were randomly allocated into 3 groups: the CT-guided nucleoplasty group (N=33), the CT-guided nucleoplasty with nerve root injection group (N=35), and CT-guided transforaminal lumbar epidural injections group (N=29). Numeric Rating Scale (NRS) pain score and Oswestry Disability Index (ODI) values were applied at pretreatment and 1 week, 1 month, 3 months, and 12 months at posttreatment. RESULTS: There were statistically significant decreases (P=0.000) in the NRS and ODI scores for all posttreatment time points when compared with the pretreatment values in all the 3 groups. The average NRS and ODI results for the transforaminal lumbar epidural injections group were significantly higher than those for the other 2 groups at 3 and 12 months posttreatment (P<0.05). The combination of nucleoplasty with nerve root injection produced a significantly greater reduction in the NRS and ODI scores when compared with nucleoplasty at 1 week (P=0.000 for NRS and P=0.004 for ODI) and 1 month (P=0.000 for NRS and P=0.007 for ODI) after the treatment. CONCLUSIONS: The results of this study suggest that CT-guided nucleoplasty with radiofrequency energy is a relative effective and safe technique for treating leg pain caused by radicular encroachment. Furthermore, nucleoplasty combined with nerve root injection had achieved a significant greater improvement in pain management and functional level in short term (within 1 mo) after treatment than nucleoplasty alone.

Local Injection of Rapamycin-Loaded Pcl-Peg Nanoparticles for Enhanced Tendon Healing in Rotator Cuff Tears via Simultaneously Reducing Fatty Infiltration and Drug Toxicity.

As a common cause of shoulder pain, rotator cuff tears (RCTs) are difficult to treat clinically because of their unsatisfactory prognosis due to the fatty infiltration caused by muscle-derived stem cells (MDSCs). Previous studies have found that rapamycin (RAPA) can inhibit fatty infiltration. However, systemic administration of RAPA may cause complications such as infection and nausea, while local administration of RAPA may lead to the cytotoxicity of tendon cells, affecting the healing of rotator cuffs. In this study, biocompatible and clinically approved polycaprolactone-polyethylene glycol (PCL-PEG) is formulated into an injectable nanoparticle for the sustained release of RAPA. The results indicate that the RAPA/PCL-PEG nanoparticles (NPs) can efficiently prolong the release of RAPA and significantly reduce the cytotoxicity of tendon cells caused by RAPA. The study of the fatty infiltration model in rats with delayed rotator cuff repair shows that weekly intraarticular injection of RAPA/PCL-PEG NPs can more effectively reduce the fatty infiltration and muscle atrophy of rat rotator cuffs and leads to better mechanical properties and gait improvements than a daily intraarticular injection of RAPA. These findings imply that local injection of RAPA/PCL-PEG NPs in the shoulder joints can be a potential clinical option for RCTs patients with fatty infiltration.

A two-step neural network-based guiding system for obtaining reliable radiographs for critical shoulder angle measurement.

Background: The critical shoulder angle (CSA) has been reported to be highly associated with rotator cuff tears (RCTs) and an increased risk of RCT re-tears. However, the measurement of the CSA is greatly affected by the malpositioning of the shoulder. To address this issue, a two-step neural network-based guiding system was developed to obtain reliable CSA radiographs, and its feasibility and accuracy was evaluated. Methods: A total of 1,754 shoulder anteroposterior (AP) radiographs were retrospectively acquired to train and validate a two-step neural network-based guiding system to obtain reliable CSA radiographs. The study included patients aged 18 years or older who underwent X-rays and/or computed tomography (CT) scans of the shoulder. Patients who had undergone shoulder surgery, had a confirmed fracture, or were diagnosed with a musculoskeletal tumor or glenoid defect were excluded from the study. The system consisted of a two-step neural network that in the first step, localized the region of interest of the shoulder, and in the second step, classified the radiography according to type [i.e., 'forward' when the non-overlapping coracoid process is above the glenoid rim, 'backward' when the non-overlapping coracoid process is below or aligned with the glenoid rim, a ratio of the transverse to longitudinal diameter of the glenoid projection (RTL) ≤0.25, or a RTL >0.25]. The performance of the model was assessed in an offline, prospective manner, focusing on the sensitivity and specificity for the forward, backward, RTL ≤0.25, or RTL >0.25 types (denoted as SensF, B, -, + and SpecF, B, -, +, respectively), and Cohen's kappa was also reported. Results: Of 273 cases in the offline prospective test, the SensF, SensB, Sens-, and Sens+ were 88.88% [95% confidence interval (CI): 50.67-99.41%], 94.11% (95% CI: 82.77-98.47%), 96.96% (95% CI: 91.94-99.02%), and 95.06% (95% CI: 87.15-98.40%), respectively. The SpecF, SpecB, Spec-, and Spec+ were 98.48% (95% CI: 95.90-99.51%), 99.55% (95% CI: 97.12-99.97%), 95.04% (95% CI: 89.65-97.81%), and 97.39% (93.69-99.03%), respectively. A high classification rate (93.41%; 95% CI: 89.14-96.24%) and almost perfect agreement (Cohen's kappa: 0.903, 95% CI: 0.86-0.95) were achieved. Conclusions: The guiding system can rapidly and accurately classify the types of AP shoulder radiography, thereby guiding the adjustment of patient positioning. This will facilitate the rapid obtainment of reliable CSA radiography to measure the CSA on proper AP radiographs.

Small molecules inhibit the interaction of Nrf2 and the Keap1 Kelch domain through a non-covalent mechanism.

Keap1 binds to the Nrf2 transcription factor to promote its degradation, resulting in the loss of gene products that protect against oxidative stress. While cell-active small molecules have been identified that modify cysteines in Keap1 and effect the Nrf2 dependent pathway, few act through a non-covalent mechanism. We have identified and characterized several small molecule compounds that specifically bind to the Keap1 Kelch-DC domain as measured by NMR, native mass spectrometry and X-ray crystallography. One compound upregulates Nrf2 response genes measured by a luciferase cell reporter assay. The non-covalent inhibition strategy presents a reasonable course of action to avoid toxic side-effects due to non-specific cysteine modification.

Inhibition of the PI3K/AKT pathway reduces tumor necrosis factor-alpha production in the cellular response to wear particles in vitro.

Joint replacement is the most effective treatment for end-stage osteoarticular disease. However, macrophage-mediated aseptic loosening of joint prosthesis severely hampers the clinical effects of joint replacement. Until now, the mechanism by which macrophages regulate the secretion of inflammatory cytokines after particle stimulation is not clear. It is well known that the PI3K/AKT pathway participates in multiple cellular processes, including cell growth, survival, and inflammation. However, whether the PI3K/AKT pathway participates in the proinflammatory response of macrophages after particle stimulation and secondary aseptic loosening is still unknown. In this study, ceramic and titanium particles of different sizes were prepared to stimulate macrophages. LY294002, a specific inhibitor of PI3K, was pretreated prior to particle stimulation. The expression of tumor necrosis factor-alpha (TNF-α) and all the subunits of PI3K and AKT were detected by real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blot. The result showed that LY294002 could suppress the RNA and protein expression of TNF-α in RAW264.7 cells after stimulation of different particles. The subunits of PI3K (p110ß and p85ß), followed by activation of phosphor-AKT (Ser473), participated in the regulation of activating macrophages by wear particles, ultimately resulting in the secretion of TNF-α.

A comparative metabolomics analysis of domestic yak (Bos grunniens) milk with human breast milk.

Yaks are tough animals living in Tibet's hypoxic stress environment. However, the metabolite composition of yak milk and its role in hypoxic stress tolerance remains largely unexplored. The similarities and differences between yak and human milk in hypoxic stress tolerance are also unclear. This study explored yak colostrum (YC) and yak mature milk (YMM) using GC-MS, and 354 metabolites were identified in yak milk. A comparative metabolomic analysis of yak and human milk metabolites showed that over 70% of metabolites were species-specific. Yak milk relies mainly on essential amino acids- arginine and essential branched-chain amino acids (BCAAs): L-isoleucine, L-leucine, and L-valine tolerate hypoxic stress. To slow hypoxic stress, human breast milk relies primarily on the neuroprotective effects of non-essential amino acids or derivates, such as citrulline, sarcosine, and creatine. In addition, metabolites related to hypoxic stress were significantly enriched in YC than in YMM. These results reveal the unique metabolite composition of yak and human milk and provide practical information for applying yak and human milk to hypoxic stress tolerance.

TAK-715 alleviated IL-1ß-induced apoptosis and ECM degradation in nucleus pulposus cells and attenuated intervertebral disc degeneration ex vivo and in vivo.

BACKGROUND: Intervertebral disc degeneration (IDD) is one of the most common disorders related to the spine. Inflammation, apoptosis and extracellular matrix (ECM) degradation contribute to disc degeneration in nucleus pulposus cells (NPCs). This study focused on the role and mechanism of the p38 inhibitor TAK-715 in intervertebral disc degeneration. METHODS: NPCs were treated with IL-1ß to mimic apoptosis, followed by the addition of TAK-715. It was determined that apoptosis, inflammatory mediators (COX-2), inflammatory cytokines (HMGB1), and ECM components (collagen II, MMP9, ADAMTS5, and MMP3) existed in NPCs. In addition, the p38MAPK signaling pathways were examined. The role of TAK-715 in vivo was determined by acupuncture-induced intervertebral disc degeneration. Following an intradiscal injection of TAK-715, MRI and a histopathological analysis were conducted to assess the degree of degeneration. RESULTS: IL-1ß-induced apoptosis was alleviated by TAK-715 in vitro, and antiapoptotic proteins were upregulated. Furthermore, TAK-715 blocked IL-1ß-induced inflammatory mediator production (COX-2) and inflammatory cytokine production (HMGB1) and degraded the ECM (collagen II, MMP9, ADAMTS5, and MMP3). By inhibiting the phosphorylation of p38, TAK-715 exerted its effects. In a rat tail model, TAK-715 ameliorates puncture-induced disc degeneration based on MRI and histopathology evaluations. CONCLUSION: TAK-715 attenuated intervertebral disc degeneration in vitro and in vivo, suggesting that it might be an effective treatment for IDD.

Amifostine-Loaded Nanocarrier Traverses the Blood-Brain Barrier and Prevents Radiation-Induced Brain Injury.

Radiation-induced brain injury (RIBI) is a severe, irreversible, or even life-threatening cerebral complication of radiotherapy in patients with head and neck tumors, and there is no satisfying prevention and effective treatment available for these patients. Amifostine (AMF) is a well-known free radical scavenger with demonstrated effectiveness in preventing radiation-induced toxicity. However, the limited permeability of AMF across the blood-brain barrier (BBB) when administered intravenously reduces the effectiveness of AMF in preventing RIBI. Herein, we construct a nanoparticle (NP) platform for BBB delivery of AMF. AMF is conjugated with 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-n-[poly(ethylene glycol)]-hydroxy succinamide [DSPE-PEG-NHS, PEG M 2000], and the product is DSPE-PEG-AMF. Then, the nanoparticles (DAPP NPs) were formed by self-assembly of poly(lactic-co-glycolic acid) (PLGA), DSPE-PEG-AMF, and polysorbate 80 (PS 80). PEG shields the nanoparticles from blood clearance by the reticuloendothelial system and lengthens the drug circulation time. PS 80 is used to encapsulate nanoparticles for medication delivery to the brain. The results of our study showed that DAPP NPs were able to effectively penetrate the blood-brain barrier (BBB) in healthy C57BL/6 mice. Furthermore, in a well-established mouse model of X-knife-induced brain injury, treatment with DAPP NPs (corresponding to 250 mg/kg AMF) was found to significantly reduce the volume of brain necrosis compared to mice treated with AMF (250 mg/kg). Importantly, the use of DAPP NPs was also shown to significantly mitigate the effects of radiation-induced neuronal damage and glial activation. This work presents a convenient brain-targeted AMF delivery system to achieve effective radioprotection for the brain, providing a promising strategy with tremendous clinical translation potential.

Fumarates promote cytoprotection of central nervous system cells against oxidative stress via the nuclear factor (erythroid-derived 2)-like 2 pathway.

Oxidative stress is central to the pathology of several neurodegenerative diseases, including multiple sclerosis, and therapeutics designed to enhance antioxidant potential could have clinical value. The objective of this study was to characterize the potential direct neuroprotective effects of dimethyl fumarate (DMF) and its primary metabolite monomethyl fumarate (MMF) on cellular resistance to oxidative damage in primary cultures of central nervous system (CNS) cells and further explore the dependence and function of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway in this process. Treatment of animals or primary cultures of CNS cells with DMF or MMF resulted in increased nuclear levels of active Nrf2, with subsequent up-regulation of canonical antioxidant target genes. DMF-dependent up-regulation of antioxidant genes in vivo was lost in mice lacking Nrf2 [Nrf2(-/-)]. DMF or MMF treatment increased cellular redox potential, glutathione, ATP levels, and mitochondrial membrane potential in a concentration-dependent manner. Treating astrocytes or neurons with DMF or MMF also significantly improved cell viability after toxic oxidative challenge in a concentration-dependent manner. This effect on viability was lost in cells that had eliminated or reduced Nrf2. These data suggest that DMF and MMF are cytoprotective for neurons and astrocytes against oxidative stress-induced cellular injury and loss, potentially via up-regulation of an Nrf2-dependent antioxidant response. These data also suggest DMF and MMF may function through improving mitochondrial function. The clinical utility of DMF in multiple sclerosis is being explored through phase III trials with BG-12, which is an oral therapeutic containing DMF as the active ingredient.

Fumaric acid esters exert neuroprotective effects in neuroinflammation via activation of the Nrf2 antioxidant pathway.

Inflammation and oxidative stress are thought to promote tissue damage in multiple sclerosis. Thus, novel therapeutics enhancing cellular resistance to free radicals could prove useful for multiple sclerosis treatment. BG00012 is an oral formulation of dimethylfumarate. In a phase II multiple sclerosis trial, BG00012 demonstrated beneficial effects on relapse rate and magnetic resonance imaging markers indicative of inflammation as well as axonal destruction. First we have studied effects of dimethylfumarate on the disease course, central nervous system, tissue integrity and the molecular mechanism of action in an animal model of chronic multiple sclerosis: myelin oligodendrocyte glycoprotein induced experimental autoimmune encephalomyelitis in C57BL/6 mice. In the chronic phase of experimental autoimmune encephalomyelitis, preventive or therapeutic application of dimethylfumarate ameliorated the disease course and improved preservation of myelin, axons and neurons. In vitro, the application of fumarates increased murine neuronal survival and protected human or rodent astrocytes against oxidative stress. Application of dimethylfumarate led to stabilization of the transcription factor nuclear factor (erythroid-derived 2)-related factor 2, activation of nuclear factor (erythroid-derived 2)-related factor 2-dependent transcriptional activity and accumulation of NADP(H) quinoline oxidoreductase-1 as a prototypical target gene. Furthermore, the immediate metabolite of dimethylfumarate, monomethylfumarate, leads to direct modification of the inhibitor of nuclear factor (erythroid-derived 2)-related factor 2, Kelch-like ECH-associated protein 1, at cysteine residue 151. In turn, increased levels of nuclear factor (erythroid-derived 2)-related factor 2 and reduced protein nitrosylation were detected in the central nervous sytem of dimethylfumarate-treated mice. Nuclear factor (erythroid-derived 2)-related factor 2 was also upregulated in the spinal cord of autopsy specimens from untreated patients with multiple sclerosis. In dimethylfumarate-treated mice suffering from experimental autoimmune encephalomyelitis, increased immunoreactivity for nuclear factor (erythroid-derived 2)-related factor 2 was detected by confocal microscopy in neurons of the motor cortex and the brainstem as well as in oligodendrocytes and astrocytes. In mice deficient for nuclear factor (erythroid-derived 2)-related factor 2 on the same genetic background, the dimethylfumarate mediated beneficial effects on clinical course, axon preservation and astrocyte activation were almost completely abolished thus proving the functional relevance of this transcription factor for the neuroprotective mechanism of action. We conclude that the ability of dimethylfumarate to activate nuclear factor (erythroid-derived 2)-related factor 2 may offer a novel cytoprotective modality that further augments the natural antioxidant responses in multiple sclerosis tissue and is not yet targeted by other multiple sclerosis therapies.