Contribution of ApoB-100/SORT1-Mediated Immune Microenvironment in Regulating Oxidative Stress, Inflammation, and Ferroptosis After Spinal Cord Injury.

This study aims to explore the impacts of ApoB-100/SORT1-mediated immune microenvironment during acute spinal cord injury (SCI), and to investigate the potential mechanism. CB57BL/6 mice underwent moderate thoracic contusion injury to establish the SCI animal model, and received ApoB-100 lentivirus injection to interfere ApoB-100 level. Functional recovery was assessed using the Basso, Beattie, and Bresnahan (BBB) score and footprint analysis. Transmission electron microscopy was applied to observe the ultrastructure of the injured spinal cord tissue. Hematoxylin-eosin (HE) staining and Perls staining were conducted to assess histological changes and iron deposition. Biochemical factor and cytokines were detected using their commercial kits. M1/M2 macrophage markers were detected by immunofluorescence assay in vivo and by flow cytometry in vitro. HT22 neurons were simulated by lipopolysaccharide (LPS), followed by incubation with polarized macrophage medium to simulate the immune microenvironment of injured spinal cord in vitro. The local immune microenvironment is changed in SCI mice, accompanied with the occurrence of oxidative stress and the elevation of both M1 and M2 macrophages. Knockdown of ApoB-100 ameliorates oxidative stress and lipid disorder, and inhibits inflammation and ferroptosis in SCI mice. Importantly, knockdown of ApoB-100 can partly restrict M1 macrophages but does not change M2 macrophage proportion in SCI mice. Further, M1 macrophages are observed to attenuate the inflammatory response, oxidative stress, and ferroptosis levels of LPS-induced HT22 cells, which is further strengthened by SORT1 knockdown. Blockage of ApoB-100/SORT1-mediated immune microenvironment plays a protective role against SCI via inhibiting oxidative stress, inflammation, lipid disorders, and ferroptosis, providing novel insights of the targeted therapy of SCI.

VIRMA promotes neuron apoptosis via inducing m6A methylation of STK10 in spinal cord injury animal models.

BACKGROUND: Spinal cord injury (SCI) occurs as a devastating neuropathic disease. The role of serine-threonine kinase 10 (STK10) in the development of SCI remains unclear. OBJECTIVE: This study aimed to investigate the action of m6A methylation on STK10 in the apoptosis of spinal cord neurons in the pathogenesis of SCI and the possible underlying mechanisms. METHODS: Rat model of SCI was established and subsequently evaluated for motor function, pathological conditions, and apoptosis of spinal cord neurons. And the effects of overexpression of STK10 on neuronal cells in animal models of spinal cord injury and glyoxylate deprivation (OGD) cell models were evaluated. m6A2Target database and SRAMP database were used to predict the m6A methylation sites of STK10. The methylation kits were used to detect overall m6A methylation. Finally, the interaction between STK10 and vir like m6A methyltransferase associated (VIRMA) was explored in animal and cellular models. RESULTS: STK10 is markedly decreased in spinal cord injury models and overexpression of STK10 inhibits neuronal apoptosis. VIRMA can induce m6A methylation of STK10. VIRMA is over-expressed in spinal cord injury models and negatively regulates the expression of STK10. m6A methylation and apoptosis of neuronal cells are reduced by the knockdown of VIRMA and STK10 shRNA have shown the opposite effects. CONCLUSIONS: VIRMA promotes neuronal apoptosis in spinal cord injury by regulating STK10 m6A methylation.

Effect of Different Accumulative Temperate Zones in Heilongjiang on Glycine Soja Metabolites as Analyzed by Non-Target Metabolomics.

To study the effect of growth temperature on the nutritional components and metabolites of the wild soybean (Glycine soja), we analyzed the nutritional components and metabolic gases of the wild soybean in six accumulated temperature regions of the Heilongjiang Province, China, by gas chromatography-time-of-flight mass spectrometry (GC-TOF-MS). A total of 430 metabolites, including organic acids, organic oxides, and lipids, were identified and analyzed using multivariate statistical analysis, orthogonal partial least squares discriminant analysis, principal component analysis, and cluster analysis. Eighty-seven metabolites significantly differed in the sixth accumulated temperature region compared with the other five accumulated temperature regions. The 40 metabolites (such as threonine (Thr) and lysine (Lys)) were found to be elevated in soybeans from the sixth accumulated temperature zone compared with the other five accumulated temperature zones. Through analyzing the metabolic pathways of these metabolites, amino acid metabolism had the greatest influence on wild soybean quality. The results of the amino acid analysis were consistent with those of the GC-TOF-MS and showed that amino acids in wild soybeans from the sixth accumulated temperature zone significantly differed from those of the other zones. Threonine and lysine were the main substances driving these differences. The growth temperature affected the type and concentrations of metabolites in wild soybeans, and the GC-TOF-MS analysis of the effect of growth temperature on wild soybean metabolites was shown to be feasible.

A novel role of lactate: Promotion of Akt-dependent elongation of microglial process.

As a key component of the innate immune system, over-activation of microglia that occurs in nervous system diseases is usually accompanied by retraction of their branched processes. Reversal of microglial process retraction is a potential strategy to prevent neuroinflammation. In our previous studies, we reported some molecules that can promote the elongation of microglial processes under in vitro and in vivo conditions, such as butyrate, ß-hydroxybutyrate, sulforaphane, diallyl disulfide, compound C, and KRIBB11. Here, we found that lactate, a molecule that mimics endogenous lactic acid and has been shown to suppress neuroinflammation, reversibly triggered significant elongations of processes in microglia under cultured and in vivo conditions. Pretreatment with lactate also prevented lipopolysaccharide (LPS)-induced shortening of microglial processes under cultured and in vivo conditions, pro-inflammatory responses in primary cultured microglia and prefrontal cortex, and depression-like behaviors in mice. Mechanistic studies revealed that incubation with lactate increased phospho-Akt levels in primary cultured microglia and inhibition of Akt blocked the pro-elongation effect of lactate on the microglial process under cultured and in vivo conditions, suggesting that the regulatory effect of lactate on the microglial process is dependent on activation of Akt. Inhibition of Akt also abolished the preventive effect of lactate on LPS-induced inflammatory responses in primary cultured microglia and prefrontal cortex and on LPS-induced depression-like behaviors in mice. Overall, these results demonstrate that lactate can induce Akt-mediated elongation of the microglial process, which appropriately contributes to the inhibition of microglia-mediated neuroinflammation.

Role of macrophage polarization in osteoarthritis (Review).

Osteoarthritis (OA) is a disease involving the whole joint that seriously reduces the living standards of individuals. Traditional treatments include physical therapy, administration of anti-inflammatory and analgesic drugs and injection of glucocorticoids or hyaluronic acid into the joints. However, these methods have limited efficacy and it is difficult to reverse the progression of OA, therefore it is urgent to find new effective treatment methods. Immune microenvironment is significant in the occurrence and development of OA. Recent studies have shown that macrophages are important targets for the treatment of OA. Macrophages are polarized into M1 pro-inflammatory phenotype and M2 anti-inflammatory phenotype under stimulation of different factors, which release and regulate inflammatory response and cartilage growth. Accumulating studies have tried to alleviate OA by regulating macrophage homeostasis. The present study summarized the related studies, discuss the mechanism of various therapeutic reagents on OA, expound the molecular mechanism of drug effect on OA and attempted to provide clues for the treatment of OA.

Percutaneous vertebral-disc plasty for thoracolumbar very severe osteoporotic vertebral compression fractures: A randomized controlled study.

Purpose: To compare the clinical outcomes and radiological parameters of patients undergoing percutaneous vertebroplasty (PVP) versus those undergoing percutaneous vertebral-disc plasty (PVDP) for back pain, segmental instability, and kyphosis due to thoracolumbar very severe osteoporotic vertebral compression fractures (vsOVCFs). Methods: This prospective randomized controlled study included elderly patients with thoracolumbar vsOVCFs. All the patients were randomly allocated into the PVP group (who underwent conventional PVP) and the PVDP group (who underwent PVP combined percutaneous cement discoplasty). The visual analogue scale (VAS), Oswestry Disability Index (ODI), local kyphosis angle, and disc height were recorded preoperatively and postoperatively. Results: Significant postoperative improvements in the VAS, ODI, and the local kyphosis angle (LKA) were shown, compared with the preoperative values in both groups (p < 0.05). The average VAS, ODI, and LKA for patients in the PVP group were increased compared to those in the PVDP group observed at the last follow-up (p < 0.05). The DHA, DHP, and LKA were seen to be maintained in the PVDP group at the last follow-up (p > 0.05). The change was significantly lower in the PVDP group at the last follow-up in those parameters (p < 0.05). Conclusion: PVDP may be a feasible and effective technique for the treatment of very severe OVCFs, that can restore intervertebral height, provide segmental stabilizing and relieve back pain in the short term.

Ubiquitin ligase Triad1 promotes neurite outgrowth by inhibiting MDM2-mediated ubiquitination of the neuroprotective factor pleiotrophin.

Spinal cord injury (SCI) is the most severe result of spine injury, but no effective therapy exists to treat SCI. We have previously shown that the E3 ubiquitin ligase Two RING fingers and DRIL 1 (Triad1) promotes neurite outgrowth after SCI. However, the mechanism by which Triad1 affects neuron growth and the potential involvement of its ubiquitination activity is unclear. Neuroprotective cytokine pleiotrophin (PTN) can promote microglia proliferation and neurotrophic factor secretion to achieve neuroprotection. We find using immunostaining and behavioral assays in rats that the expression of Triad1 and the PTN was peaked at 1 day after SCI and Triad1 improved motor function and histomorphological injury after SCI. We show using flow cytometry and astrocyte/neuronal coculture assays that Triad1 overexpression promoted PTN protein levels, neurotrophic growth factor (NGF) expression, brain-derived neurotrophic factor (BDNF) expression, astrocyte and neuronal viability, and neurite outgrowth but suppressed astrocyte apoptosis, while shRNA-mediated knockdown of Triad1 and PTN had the opposite effects. Ubiquitin ligase murine double mutant 2 (MDM2) has previously been demonstrated to participate in the process of neurite outgrowth and mediate ubiquitination of p53. Furthermore, we demonstrate overexpression of MDM2 downregulated PTN protein levels, NGF expression and BDNF expression in astrocytes, and inhibited neurite outgrowth of neurons. In addition, MDM2 facilitated PTN ubiquitination, which was reversed by Triad1. Finally, we show simultaneous sh-PTN and MDM2 overexpression attenuated the neurite outgrowth-promoting effect of Triad1 overexpression. In conclusion, we propose Triad1 promotes astrocyte-dependent neurite outgrowth to accelerate recovery after SCI by inhibiting MDM2-mediated PTN ubiquitination.

Upregulation of P2Y12 inhibits chondrocyte apoptosis in lumbar osteoarthritis through the PI3K/AKT signaling pathway.

Lumbar facet osteoarthritis (FJOA) is a major cause of severe lower back pain and disability worldwide. However, the mechanism underlying cartilage degeneration in FJOA remains unclear. The purpose of this study was to investigate the regulation and mechanism of P2Y12 on chondrocyte apoptosis in FJOA. The experimental rats were randomly divided into non-operation (n = 20) and operation groups (n = 20). In the operation group, Sodium iodoacetate (MIA, Sigma, 200 mg/mL) was injected into the right L4/5 facet process using a blunt nanoneedle 26 (WPI, Sarasota, FL, USA) under the control of an injection pump. The final injection volume was 5µL and the injection rate was 2µL/min. The facet joint was removed four weeks after surgery. After the operation, samples were stored at -80 °C until further use, whereby the right facet joints in each group were tested. Hematoxylin and eosin (HE) and iron-red solid green staining were used to observe the degeneration of articular chondrocytes in rats. Immunohistochemistry and western blotting were used to observe the expressions of P2Y12, Matrix metalloproteinase 13 (MMP13), Collagen II (COL2), and other cartilage degeneration and apoptosis-related genes. Co-localization of P2Y12-cleaved caspase-3 in the apoptosis model was detected by dual-standard immunofluorescence staining. Apoptosis was also detected by flow cytometry and TUNEL assay.P2Y12 is highly expressed in OA cartilage tissue, and inhibits IL-1ß -induced chondrocyte apoptosis through PI3K/AKT signaling pathway, thus playing a certain protective role on cartilage.

Wnt16 protects chondrocytes from lumbar facet joint osteoarthritis through the Wnt/ß-catenin pathway in low back pain patients.

PURPOSE: Low back pain (LBP) is a long-lasting and chronic symptom without any exact cause. This study attempts to propose a new staging system based on the original grading system combined with pathological results and clinical symptoms to better clarify the dynamic evolution of LBP related to cartilage degeneration during facet joint osteoarthritis (FJOA). To explore a potential target for diagnosis, treatment, and drug intervention of facet joint osteoarthritis related LBP via protecting chondrocytes. MATERIALS AND METHODS: All the facet joints were divided into 4 groups according to our new degenerative staging system based on Weishaupt grade, CT and MRI. Collect the facet joint samples from patients whom suffered lumbar fusion surgery for lumbar disc herniation. Molecular biology experiments were used to explore the effect of Wnt16 on the degeneration of facet joints. Micro-CT examination and pain stimulation test checked the biological function of Wnt16 in rats. RESULTS: Wnt16 was significantly increased and more aggregated in the facet joint chondrocytes in the Phase III and Phase IV, which is consistent with the pathological findings of cartilage degeneration (OARSI). We found that Wnt16 participated in the regulation of FJOA via Wnt/ß-catenin pathway in vitro, which was inhibited by specific inhibitor DKK1. The rats, rich expressed Wnt16, showed higher paw withdrawal thresholds and prolonged paw withdrawal latency to FJOA related LBP. Micro-CT examination for the lumbar spine of rats showed Wnt16 protected the chondrocytes from FJOA. CONCLUSIONS: This study defined a new staging system for LBP related cartilage degeneration of facet joint based on the original grading system combined with pathological results and clinical symptoms. Wnt16 is expected to be a potential target for treatment of FJOA via protecting chondrocytes.

Knockdown of TRAF6 inhibits chondrocytes apoptosis and inflammation by suppressing the NF-κB pathway in lumbar facet joint osteoarthritis.

Tumor necrosis factor receptor-associated factor 6 (TRAF6), a regulator of NF-κB signaling, has been discovered recently to be probably related to osteoarthritis, while the function of TRAF6 in lumbar facet joint osteoarthritis(FJOA)still remains unknown. The aim of this study was to probe the specific function of TRAF6 in chondrocytes and its connection with the pathophysiology of FJOA. We found upregulation of TRAF6 in FJOA cartilage by western blot analysis. In vitro, we stimulated immortalized human chondrocytes by LPS to establish the cells apoptosis model. Western blot analysis demonstrated that levels of TRAF6 and cleaved caspase-3/8 in the chondrocyte injury model increased significantly. Knockdown of TRAF6 suppressed the expression of matrix metallopeptidase-13 (MMP-13) and interleukin-6 (IL-6) induced by LPS, and alleviated cell apoptosis. Meanwhile, western blot and immunofluorescent staining demonstrated that IκBα degradation and p65 nuclear transportation were also inhibited, revealing that knockdown of TRAF6 suppressed activation of the NF-κB pathway in LPS-induced chondrocytes apoptosis model. Collectively, our findings suggest that TRAF6 plays a crucial role in FJOA development by regulating NF-κB signaling pathway. Knockdown of TRAF6 may supply a potential therapeutic strategy for FJOA.

Sprouty2 Inhibits Migration and Invasion of Fibroblast-Like Synoviocytes in Rheumatoid Arthritis by Down-regulating ATF2 Expression and Phosphorylation.

Activating transcription factor 2(ATF2), a transcription factor belonging to the AP-1 family, plays an important role in inflammation. However, its biological functions and underlying molecular mechanisms in rheumatoid arthritis (RA) remain unclear. Western blot and immunohistochemistry were used to identify the expression of ATF2 and Sprouty2(SPRY2) in RA synovial tissues. SW982 cells were stimulated by TNF-α to establish an in vitro RA fibroblast-like synoviocyte (RA-FLS) model. Transwell and monolayer wound-healing were used to detect cell migration and invasion. RNA interference (si-ATF2) and adenovirus vector (Ad-SPRY2) methods were employed to manipulate ATF2 or SPRY2 expression in SW982 cells. The protein expression and phosphorylation levels in SW982 cells were evaluated by western blot. ATF2 expression and phosphorylation were upregulated in the RA synovial tissues. In RA-FLS model, ATF2 expression and phosphorylation were increased in a time-dependent manner. ATF2 knockdown inhibited the migration and invasion of RA-FLS model, reducing the inflammatory factors, which was consistent with the influence on cell behaviors caused by SPRY2 overexpression. Moreover, SPRY2 overexpression inhibited the TNF-α-induced phosphorylation of ERK and ATF2 in SW982 cells. The high expression and phosphorylation of ATF2 promoted migration and invasion of RA-FLSs. SPRY2 might inhibited the inflammatory responses of RA-FLSs via suppressing ERK-ATF2 pathway.

Novel Role of HAX-1 in Neurons Protection After Spinal Cord Injury Involvement of IRE-1.

Spinal cord injury (SCI) is one of the diseases with high probability of causing disability in human beings, and there is no reliable treatment at present. Neuronal apoptosis is a vital component of secondary injury and plays a critical role in the development of neurological dysfunction after spinal cord injury. In this study, we found that the expression and distribution of HAX-1 in neurons increased 1 day after SCI. PC12 cells overexpressing HAX-1 showed decreased apoptosis and PC12 cells are more likely to undergo apoptosis after down-regulating HAX-1, which was confirmed via TUNEL experiments. We found GRP94 showed the same trend as HAX-1 in expression and interacted with HAX-1 and IRE-1 in both spinal cord tissue and PC12 cells, and this interaction seems to be enhanced after SCI. When the expression of HAX-1 was up-regulated, GRP94 also increased, but IRE-1 did not change at all. Further studies showed that overexpression of HAX-1 decreased the expression of pIRE-1, rather than IRE-1, and downstream proteins of the IRE signaling pathway (Caspase12, pJNK and CHOP) were significantly reduced, and vice versa. In animals treated with HAX-1 expressing adenovirus there are more neuronal cells remaining in the damaged spinal cord tissue, and hindlimb motor function of rats was significantly improved. So, we speculate that HAX-1 might play a role in protecting neurons from apoptosis after SCI by regulating the IRE-1 signaling pathway via promoting the dissociation of GRP94 from IRE-1. This may provide a theoretical basis and a potential therapeutic target for clinical improvement of neural function recovery after SCI.

Interaction between C/EBPß and RUNX2 promotes apoptosis of chondrocytes during human lumbar facet joint degeneration.

The pathophysiological changes in cartilage are a crucial feature of lumbar facet joint (LFJ) degeneration and arthritis. However, the molecular mechanism of human LFJ degeneration remains largely defined. This study aimed to examine the changes in chondrocytes at different stages of degenerative LFJ using hematoxylin and eosin and Safranin O staining. The significant loss of chondrocytes in grades 2 and 3 of LFJs was observed. The expression levels of CCAAT enhancer binding protein ß (C/EBPß), Runt-related transcription factor 2 (RUNX2), and matrix metalloproteinase 13 (MMP13) also increased with the aggravation of degeneration (4.89, 5.77, and 6.3 times by Western blot). In vitro, chondrocytes scraped from the LFJs during surgery were stimulated by interleukin (IL)-1ß to establish the injury model. The association of C/EBPß and RUNX2 with active caspase-3 on chondrocytes was analyzed. The high expression level of C/EBPß, RUNX2, and MMP13 was consistent with that of caspase-3, which reached a peak after 36 h of stimulation. Immunofluorescence suggested that C/EBPß, RUNX2, and MMP13 co-labeled with active caspase-3. Moreover, immunoprecipitation data prompted that C/EBPß was able to interact with RUNX2. The knockdown of C/EBPß significantly decreased the expression levels of MMP13 and active caspase-3 (2.48 and 2.89 times as detected by Western blot analysis) and inhibited chondrocyte apoptosis, which was further demonstrated using flow cytometry. Taken together, the findings of this study uncovered that C/EBPß could interact with RUNX2 to induce chondrocyte apoptosis in human LFJ degeneration by regulating the expression of MMP13.

Chest computed tomography findings of coronavirus disease 2019 (COVID-19) pneumonia.

OBJECTIVE: To retrospectively analyze the chest computed tomography (CT) features in patients with coronavirus disease 2019 (COVID-19) pneumonia. METHODS: From January 9, 2020, to February 26, 2020, totally 56 laboratory-confirmed patients with COVID-19 underwent chest CT. For 40 patients, follow-up CT scans were obtained. The CT images were evaluated for the number, type and distribution of the opacity, and the affected lung lobes. Furthermore, the initial CT scan and the follow-up CT scans were compared. RESULTS: Forty patients (83.6%) had two or more opacities in the lung. Eighteen (32.7%) patients had only ground-glass opacities; twenty-nine patients (52.7%) had ground-glass and consolidative opacities; and eight patients (14.5%) had only consolidation. A total of 43 patients (78.2%) showed two or more lobes involved. The opacities tended to be both in peripheral and central (30/55, 54.5%) or purely peripheral distribution (25/55, 45.5%). Fifty patients (90.9%) had the lower lobe involved. The first follow-up CT scans showed that twelve patients (30%) had improvement, 26 (65%) patients had mild-moderate progression, and two patients (5%) had severe progression with "white lungs." The second follow-up CT showed that 22 patients (71%) showed improvement compared with the first follow-up CT, four patients (12.9%) had aggravated progression, and five patients (16.1%) showed unchanged radiographic appearance. CONCLUSIONS: The common CT features of COVID-19 pneumonia are multiple lung opacities, multiple types of the opacity (ground-glass, ground-glass and consolidation, and consolidation alone), and multiple lobes especially the lower lobe involved. Follow-up CT could demonstrate the rapid progression of COVID-19 pneumonia (either in aggravation or absorption). KEY POINTS: • The predominant CT features of COVID-19 pneumonia are multiple ground-glass opacities with or without consolidation and, with both lungs, multiple lobes and especially the lower lobe affected. • CT plays a crucial role in early diagnosis and assessment of COVID-19 pneumonia progression. • CT findings of COVID-19 pneumonia may not be consistent with the clinical symptoms or the initial RT-PCR test results.

Enhanced osteogenic activity and antibacterial ability of manganese-titanium dioxide microporous coating on titanium surfaces.

Titanium (Ti) and its alloys are widely used in clinical practice as preferred materials for bone tissue repair and replacement because of their good mechanical properties; however, as Ti lacks biological activity, clinical application has been limited. Herein, we prepared a manganese-titanium dioxide (Mn-TiO2) microporous biotic coating on Ti surfaces by micro-arc oxidation (MAO). The coating showed good surface topography and was uniformly doped with Mn, and the Mn ions were slowly released. In vitro, the Mn-TiO2 microporous biotic coating promoted the adhesion, proliferation, differentiation, and mineralization of MC3T3-E1 osteoblasts. Moreover, in vivo experiments showed that the coating promoted early osseointegration. We also conducted a preliminary investigation to explore the molecular mechanism underlying the regulation of the function of osteoblasts by the coating. Furthermore, we found that the coating could inhibit the growth of Escherichia coli in vitro, demonstrating reliable antibacterial ability. To conclude, Mn-TiO2 microporous biotic coating can improve the biological activity of Ti implants, which can potentially improve their clinical applications.

Triad1 regulates the expression and distribution of EHD1 contributing to the neurite outgrowth of neurons after spinal cord injury.

Traumatic spinal cord injury is a common and severe complication after an accident. As we all know that neurite outgrowth of neurons is difficult after a spinal cord injury. Endosome system is associated with cargoes transportation and contributes in promoting the neuronal capability for neurite outgrowth. EH domain-containing protein 1 (EHD1) transports proteins through the endosome system, especially in the recycling endosomes and regulating the neurite outgrowth. In mammalian cells, the involvement of the ubiquitin-proteasome system in endosomal sorting has been well established. Two RING fingers and a DRIL (double RING finger-linked) 1 (Triad1) plays an important role in membrane trafficking and its mutant results in the wrong accumulation of receptors in endosomes and plasma membrane. In this current study, we reasonably integrated the results of the above research and investigated the regulating function of Triad1 to EHD1 following the spinal cord injury. We characterized the upregulated expression and distribution of Triad1 and EHD1 in the neurons after SCI and declared the interaction between Triad1 with EHD1 both in vitro and in vivo. Triad1 regulated the interaction between itself and the full-length or EH domain of EHD1, which influenced the neurite outgrowth of PC12 cells. Our data delineate a novel interaction between Triad1 and EHD1 that may contribute to the regulation of neurite outgrowth for neurons after the spinal cord injury.

Dyrk1A promotes the proliferation, migration and invasion of fibroblast-like synoviocytes in rheumatoid arthritis via down-regulating Spry2 and activating the ERK MAPK pathway.

Fibroblast-like synoviocytes (FLSs) play an essential role in rheumatoid arthritis (RA) by promoting synovitis, pannus growth and cartilage/bone destruction. Increased proliferation, migration and invasion of FLSs greatly contribute to RA initiation and progression. Dual-specificity tyrosine-regulated kinase 1A (Dyrk1A), a serine/threonine kinase, regulates MAPK pathway activation, and governs the proliferation and differentiation of neuronal progenitor cells and cancer cells. Till now, the expression and possible function of Dyrk1A in RA FLSs have not been explored. In this study, we detected an increased expression of Dyrk1A both in the synovial tissues of RA patients and in a TNF-α-induced FLSs activation model. CCK-8 and Edu assays revealed that Dyrk1A knockdown inhibited TNF-α-induced FLSs proliferation. Moreover, inhibiting Dyrk1A expression apparently prevented the migration and invasion capability of FLSs accompanied by a decreased MMP-3 and -9 expression. To investigate the molecular mechanism through which Dyrk1A modulates FLSs activities, we evaluated the effects of Dyrk1A on Spry2, a negativity modulator of ERK MAPK pathway. Western blot assay demonstrated that Dyrk1A silencing significantly increased Spry2 expression and suppressed the phosphorylation of ERK in TNF-α-treated FLSs. Taken together, our results indicated that Dyrk1A might promote FLSs proliferation, migration and invasion by suppressing Spry2 expression and activating the ERK MAPK signaling pathway in RA.

HAX1 is associated with neuronal apoptosis and astrocyte proliferation after spinal cord injury.

HS1-associated protein X-1 (HAX1) is a class of multifunctional protein, participated in various physiological processes such as cell apoptosis, proliferation and motility. However, the HAX1 expression and function in the spinal cord injury (SCI) pathological process have not been investigated. In our current research, the rat model of SCI was established, and then we explored the possible role of HAX1 after SCI. The results of western blot indicated that HAX1 was present in sham operated control group and significantly elevated at 3 days post SCI, then declined gradually. Immunohistochemical studies indicated HAX1 expression was enhanced significantly in white and gray matter at 3 days post SCI compared with sham operated group. Double immunofluorescence staining showed the proportion of cells, double-labeled HAX1 and neurons, astrocytes, increased significantly at 3 days post SCI. In addition, co-localization of HAX1/active caspase-3 and HAX1/PCNA was tested in cells. Furthermore, over-expression of HAX1 inhibited neuronal apoptosis in vitro, and in astrocytes HAX1 silencing could down-regulate PCNA expression post LPS treatment. Meanwhile, CCK8 assay showed that knockdown of HAX1 could inhibit the astrocyte proliferation. In summary, our data indicated that HAX1 might play significant roles in pathological process of neuronal apoptosis and astrocyte proliferation during SCI.

Up-regulation of TRAF2 inhibits chondrocytes apoptosis in lumbar facet joint osteoarthritis.

Tumor necrosis factor receptor-associated factor 2 (TRAF2) has been demonstrated that it plays a significant role in cell death receptor signal transduction. The purpose of this study was to investigate the expression of TRAF2 and its possible role in FJOA. We observed an up-regulation of TRAF2 in FJOA by immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) compared to normal tissues. In vitro, we used TNF-α to stimulate Human SW1353 chondrosarcoma cells to establish the chondrocytes injury model. Western blot analysis revealed significant expression of TRAF2 and cleaved caspase-3/8 in SW1353 cells. Co-localization of TRAF2/cleaved caspase-3/8 was detected in the cells injury model by double-labeling immunofluorescent staining. We demonstrated a possible anti-apoptotic effect of TRAF2 in chondrocyte apoptosis in FJOA by knockdown of its expression with siRNA. Moreover, TRAF2 knockdown was demonstrated to enhance TNF-α-induced apoptosis by flow cytometry assay. In conclusion, our results show that the up-regulation of TRAF2 may play an important role in the inhibition of chondrocyte apoptosis of FJOA.

E3 Ubiquitin Ligase c-cbl Inhibits Microglia Activation After Chronic Constriction Injury.

E3 ubiquitin ligase c-Caritas B cell lymphoma (c-cbl) is associated with negative regulation of receptor tyrosine kinases, signal transduction of antigens and cytokine receptors, and immune response. However, the expression and function of c-cbl in the regulation of neuropathic pain after chronic constriction injury (CCI) are unknown. In rat CCI model, c-cbl inhibited the activation of spinal cord microglia and the release of pro-inflammatory factors including tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL-1ß) and interleukin 6 (IL-6), which alleviated mechanical and heat pain through down-regulating extracellular signal-regulated kinase (ERK) pathway. Additionally, exogenous TNF-α inhibited c-cbl protein level vice versa. In the primary microglia transfected with c-cbl siRNA, when treated with TNF-α or TNF-α inhibitor, the corresponding secretion of IL-1ß and IL-6 did not change. In summary, CCI down-regulated c-cbl expression and induced the activation of microglia, then activated microglia released inflammatory factors via ERK signaling to cause pain. Our data might supply a novel molecular target for the therapy of CCI-induced neuropathic pain.