Mass Spectrometry-Based Proteomic Analysis of Potential Host Proteins Interacting with GP5 in PRRSV-Infected PAMs.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a typical immunosuppressive virus causing a large economic impact on the swine industry. The structural protein GP5 of PRRSV plays a pivotal role in its pathogenicity and immune evasion. Virus-host interactions play a crucial part in viral replication and immune escape. Therefore, understanding the interactions between GP5 and host proteins are significant for porcine reproductive and respiratory syndrome (PRRS) control. However, the interaction network between GP5 and host proteins in primary porcine alveolar macrophages (PAMs) has not been reported. In this study, 709 GP5-interacting host proteins were identified in primary PAMs by immunoprecipitation coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Bioinformatics analysis revealed that these proteins were involved in multiple cellular processes, such as translation, protein transport, and protein stabilization. Subsequently, immunoprecipitation and immunofluorescence assay confirmed that GP5 could interact with antigen processing and presentation pathways related proteins. Finally, we found that GP5 may be a key protein that inhibits the antigen processing and presentation pathway during PRRSV infection. The novel host proteins identified in this study will be the candidates for studying the biological functions of GP5, which will provide new insights into PRRS prevention and vaccine development.

MicroRNA-376b-3p Promotes Porcine Reproductive and Respiratory Syndrome Virus Replication by Targeting Viral Restriction Factor TRIM22.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major economically significant pathogen and has evolved several strategies to evade host antiviral response and provide favorable conditions for survival. In the present study, we demonstrated that a host microRNA, miR-376b-3p, was upregulated by PRRSV infection through the viral components, nsp4 and nsp11, and that miR-376b-3p can directly target tripartite motif-containing 22 (TRIM22) to impair its anti-PRRSV activity, thus facilitating the replication of PRRSV. Meanwhile, we found that TRIM22 induced degradation of the nucleocapsid protein (N) of PRRSV by interacting with N protein to inhibit PRRSV replication, and further study indicated that TRIM22 could enhance the activation of the lysosomal pathway by interacting with LC3 to induce lysosomal degradation of N protein. In conclusion, PRRSV increased miR-376b-3p expression and hijacked the host miR-376b-3p to promote PRRSV replication by impairing the antiviral effect of TRIM22. Therefore, our finding outlines a novel strategy of immune evasion exerted by PRRSV, which is helpful for better understanding the pathogenesis of PRRSV. IMPORTANCE Porcine reproductive and respiratory syndrome virus (PRRSV) causes enormous economic losses each year in the swine industry worldwide. MicroRNAs (miRNAs) play important roles during viral infections via modulating the expression of viral or host genes at the posttranscriptional level. TRIM22 has recently been identified as a key restriction factor that inhibited the replication of a number of human viruses, such as HIV, encephalomyocarditis virus (ECMV), hepatitis C virus (HCV), HBV, influenza A virus (IAV), and respiratory syncytial virus (RSV). In this study, we showed that host miR-376b-3p could be upregulated by PRRSV and functioned to impair the anti-PRRSV role of TRIM22 to facilitate PRRSV replication. Meanwhile, we found that TRIM22 inhibited the replication of PRRSV by interacting with viral N protein and accelerating its degradation through the lysosomal pathway. Collectively, the findings reveal a novel mechanism that PRRSV used to exploit the host miR-376b-3p to evade antiviral responses and provide new insight into the study of virus-host interactions.

Diabetes, fasting blood glucose and the efficacy of remote ischaemic conditioning: A secondary analysis of the RICAMIS trial.

AIM: To investigate whether diabetes and fasting blood glucose (FBG) levels affect the efficacy of remote ischaemic conditioning (RIC) using the database included in the Remote Ischaemic Conditioning for Acute Moderate Ischaemic Stroke (RICAMIS) trial. METHODS: A total of 1707 patients were enrolled in this post hoc study, including 535 patients with diabetes and 1172 without diabetes. Each group was further divided into RIC and control subgroups. The primary outcome was excellent functional outcome, defined as a modified Rankin Scale (mRS) score of 0 to 1 at 90 days. The difference in the proportion of patients with excellent functional outcome between the RIC subgroup and control subgroup was compared in diabetic and non-diabetic patients, respectively, and the interactions of treatment assignment with diabetes status and FBG were evaluated. RESULTS: Compared with the control group, RIC produced a significantly higher proportion of patients with excellent functional outcome in the non-diabetic group (70.5% vs. 63.2%; odds ratio [OR] 1.487, 95% confidence interval [CI] 1.134-1.949; P = 0.004), while a similar, but not significant difference was observed in the diabetic group (65.3% vs. 59.8%; OR 1.424, 95% CI 0.978-2.073; P = 0.065). Similar results were observed in patients with normal FBG levels (69.3% vs. 63.7%; OR 1.363, 95% CI 1.011-1.836; P = 0.042) and those with high FBG levels (64.2% vs. 58%; OR 1.550, 95% CI 1.070-2.246; P = 0.02). Furthermore, we did not find an interaction effect of intervention (RIC or control) by different diabetes status or FBG levels on clinical outcomes (P > 0.05 for all). However, diabetes (OR 0.741, 95% CI 0.585-0.938; P = 0.013) and high FBG (OR 0.715, 95% CI 0.553-0.925; P = 0.011) were independently associated with functional outcomes in patients overall. CONCLUSION: Diabetes and FBG levels did not influence the neuroprotective effect of RIC in acute moderate ischaemic stroke, although diabetes and high FBG levels were independently associated with functional outcomes.

Efficacy of remote ischemic conditioning does not differ between men and women: A secondary analysis of the RICAMIS study.

BACKGROUND AND PURPOSE: The present study aimed to determine sex difference in clinical outcomes after Remote Ischemic Conditioning for Acute Moderate Ischemic Stroke (RICAMIS). METHODS: In this secondary analysis of the RICAMIS study, eligible patients aged 18 years or older with acute moderate ischemic stroke who received remote ischemic conditioning (RIC) within 48 h of stroke onset were divided into two groups: men and women. The primary endpoint was an excellent functional outcome, defined as a modified Rankin Scale score of 0-1 at 90 days. Binary logistic regression analyses and generalized linear models were used. RESULTS: Of 1707 eligible patients, 34% (579) were women. Women had a higher burden of hypertension and diabetes, and less alcohol and smoking consumption than men. The mean systolic blood pressure and blood glucose level at randomization were higher in women than in men. Compared with the control group, RIC was associated with an increased rate of primary endpoint in men (unadjusted odds ratio [OR] = 1.277; 95% confidence interval [CI] 0.933-1.644; p = 0.057) and women (unadjusted OR = 1.454; 95% CI 1.040-2.032; p = 0.028). Furthermore, a higher absolute risk difference in primary endpoint between control and RIC groups was found in women (9.2%) than in men (5.7%), but there was no significant interaction effect between sex and intervention on primary outcome (p interaction = 0.545). CONCLUSION: Compared with men, women may have a higher probability of excellent functional outcomes at 90 days in the RIC group than in the control group; however, no interaction effect between sex and intervention was found.

Melatonin attenuates lung ischemia-reperfusion injury through SIRT3 signaling-dependent mitophagy in type 2 diabetic rats.

Background: Lung ischemia-reperfusion injury (LIRI) remains the major cause of primary lung dysfunction after lung transplantation. Diabetes mellitus (DM) is an independent risk factor for morbidity and mortality following lung transplantation. Mitochondrial dysfunction is recognized as a key mediator in the pathogenesis of diabetic LIRI. Melatonin has been reported to be a safe and potent preserving mitochondrial function agent. This study aimed at investigating the potential therapeutic effect and mechanisms of melatonin on diabetic LIRI. Methods: High-fat-diet-fed streptozotocin-induced type 2 diabetic rats were exposed to melatonin, with or without administration of the SIRT3 short hairpin ribonucleic acid (shRNA) plasmid following a surgical model of ischemia-reperfusion injury of the lung. Lung function, inflammation, oxidative stress, cell apoptosis, and mitochondrial function were examined. Results: The SIRT3 signaling and mitophagy were suppressed following diabetic LIRI. Treatment with melatonin markedly induced mitophagy and restored SIRT3 expression. Melatonin treatment also attenuated subsequent diabetic LIRI by improving lung functional recovery, suppressing inflammation, decreasing oxidative damage, diminishing cell apoptosis, and preserving mitochondrial function. However, either administration of SIRT3 shRNA or an autophagy antagonist 3-methyladenine (3-MA) suppressing mitophagy, and compromised the protective action of melatonin. Conclusion: Data indicated that melatonin attenuates diabetic LIRI through activation of SIRT3 signaling-mediated mitophagy.

The Host E3-Ubiquitin Ligase TRIM28 Impedes Viral Protein GP4 Ubiquitination and Promotes PRRSV Replication.

Porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), is a highly pathogenic porcine virus that brings tremendous economic losses to the global swine industry. PRRSVs have evolved multiple elegant strategies to manipulate the host proteins and circumvent against the antiviral responses to establish infection. Therefore, the identification of virus-host interactions is critical for understanding the pathogenesis of PRRSVs. Tripartite motif protein 28 (TRIM28) is a transcriptional co-repressor involved in the regulation of viral and cellular transcriptional programs; however, its precise role in regulating PRRSV infection remains unknown. In this study, we found that the mRNA and protein levels of TRIM28 were up-regulated in PRRSV-infected porcine alveolar macrophages (PAMs) and MARC-145 cells. Ectopic TRIM28 expression dramatically increased viral yields, whereas the siRNA-mediated knockdown of TRIM28 significantly inhibited PRRSV replication. Furthermore, we used a co-immunoprecipitation (co-IP) assay to demonstrate that TRIM28 interacted with envelope glycoprotein 4 (GP4) among PRRSV viral proteins. Intriguingly, TRIM28 inhibited the degradation of PRRSV GP4 by impeding its ubiquitination. Taken together, our work provides evidence that the host E3-ubiquitin ligase TRIM28 suppresses GP4 ubiquitination and is important for efficient virus replication. Therefore, our study identifies a new host factor, TRIM28, as a potential target in the development of anti-viral drugs against PRRSV.

Cytoplasmic Cargo Receptor p62 Inhibits Avibirnavirus Replication by Mediating Autophagic Degradation of Viral Protein VP2.

Selective autophagy regulates the degradation of cytoplasmic cargos, such as damaged organelles, invading pathogens, and aggregated proteins. Furthermore, autophagy is capable of degrading avibirnavirus, but the mechanism responsible for this process is unclear. Here, we show that autophagy cargo receptor p62 regulates the degradation of the avibirnavirus capsid protein VP2. Binding of p62 to VP2 enhances autophagic induction and promotes autophagic degradation of viral protein VP2. Further study showed that the interaction of p62 with viral protein VP2 is dependent on ubiquitination at the K411 site of VP2 and the ubiquitin-associated domain of p62. Mutation analysis showed that the K411R mutation of viral protein VP2 prohibits its p62-mediated degradation. Consistent with this finding, p62 lacking the ubiquitin-associated domain or the LC3-interacting region no longer promoted the degradation of VP2. Virus production revealed that the knockout of p62 but not the overexpression of p62 promotes the replication of avibirnavirus. Collectively, our findings suggest that p62 mediates selective autophagic degradation of avibirnavirus protein VP2 in a ubiquitin-dependent manner and is an inhibitor of avibirnavirus replication.IMPORTANCE Avibirnavirus causes severe immunosuppression and mortality in young chickens. VP2, the capsid protein of avibirnavirus, is responsible for virus assembly, maturation, and replication. Previous study showed that avibirnavirus particles could be engulfed into the autophagosome and degradation of virus particles took apart. Selective autophagy is a highly specific and regulated degradation pathway for the clearance of damaged or unwanted cytosolic components and superfluous organelles as well as invading microbes. However, whether and how selective autophagy removes avibirnavirus capsids is largely unknown. Here, we have shown that selective autophagy specifically clears ubiquitinated avibirnavirus protein VP2 by p62 recognition and that p62 is an inhibitor of avibirnavirus replication, highlighting the role of p62 as a potential drug target for mediating the removal of ubiquitinated virus components from cells.

Lactic acid promotes metastatic niche formation in bone metastasis of colorectal cancer.

BACKGROUND: To investigate the effect of lactic acid (LA) on the progression of bone metastasis from colorectal cancer (CRC) and its regulatory effects on primary CD115 (+) osteoclast (OC) precursors. METHODS: The BrdU assay, Annexin-V/PI assay, TRAP staining and immunofluorescence were performed to explore the effect of LA on the proliferation, apoptosis and differentiation of OC precursors in vitro and in vivo. Flow cytometry was performed to sort primary osteoclast precursors and CD4(+) T cells and to analyze the change in the expression of target proteins in osteoclast precursors. A recruitment assay was used to test how LA and Cadhein-11 regulate the recruitment of OC precursors. RT-PCR and Western blotting were performed to analyze the changes in the mRNA and protein expression of genes related to the PI3K-AKT pathway and profibrotic genes. Safranin O-fast green staining, H&E staining and TRAP staining were performed to analyze the severity of bone resorption and accumulation of osteoclasts. RESULTS: LA promoted the expression of CXCL10 and Cadherin-11 in CD115(+) precursors through the PI3K-AKT pathway. We found that CXCL10 and Cadherin-11 were regulated by the activation of CREB and mTOR, respectively. LA-induced overexpression of CXCL10 in CD115(+) precursors indirectly promoted the differentiation of osteoclast precursors through the recruitment of CD4(+) T cells, and the crosstalk between these two cells promoted bone resorption in bone metastasis from CRC. On the other hand, Cadherin-11 mediated the adhesion between osteoclast precursors and upregulated the production of specific collagens, especially Collagen 5, which facilitated fibrotic changes in the tumor microenvironment. Blockade of the PI3K-AKT pathway efficiently prevented the progression of bone metastasis caused by lactate. CONCLUSION: LA promoted metastatic niche formation in the tumor microenvironment through the PI3K-AKT pathway. Our study provides new insight into the role of LA in the progression of bone metastasis from CRC. Video Abstract.

Overexpression of circ_0021739 in Peripheral Blood Mononuclear Cells in Women with Postmenopausal Osteoporosis Is Associated with Reduced Expression of microRNA-194-5p in Osteoclasts.

BACKGROUND Postmenopausal osteoporosis, a common disease among elderly women, is linked to estrogen deficiency, mechanical loading, and genotype. Circular RNAs (circRNAs) are formed through reverse splicing of the splice donor at the 3' end and the splice accepter at the 5' end in pre-mRNA and have been shown to be involved in the development of multiple diseases. Based on their high sequence conservation and stability, circRNAs may be useful biomarkers in different diseases. However, the roles of circRNAs in postmenopausal osteoporosis remain incompletely understood. MATERIAL AND METHODS Fifty-three postmenopausal women were assigned to either the postmenopausal osteoporosis group (n=28) or the control group (n=25). Reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) analysis was performed to determine the differential expression of circRNAs between the 2 groups. Receiver-operating characteristic (ROC) curve analysis was conducted to evaluate the clinical diagnostic value of circRNA. Prediction of the binding sites between circRNA and miRNAs was conducted using miRanda and RNAhybrid. The function of the circRNA in osteoclastogenesis was determined by circRNA overexpression followed by tartrate-resistant acid phosphatase staining and RT-qPCR analysis. RESULTS Among 4 circRNAs previously identified by RNA-sequencing analysis as differentially expressed in patients with postmenopausal osteoporosis, only hsa_circ_0021739 showed a significant difference in expression between the groups and was downregulated in patients with postmenopausal osteoporosis. The hsa_circ_0021739 expression level was determined to be correlated with the lumbar vertebra, femur, and forearm T-scores. Overexpression of hsa_circ_0021739 decreased the level of hsa-miR-502-5p and inhibited the differentiation of osteoclasts. CONCLUSIONS The circRNA hsa_circ_0021739 is a potential blood biomarker for postmenopausal osteoporosis. In addition, hsa-miR-502-5p is a likely target of hsa_circ_0021739, which acts to regulate the differentiation of osteoclasts.

Neuronal LRP4 regulates synapse formation in the developing CNS.

The low-density lipoprotein receptor-related protein 4 (LRP4) is essential in muscle fibers for the establishment of the neuromuscular junction. Here, we show that LRP4 is also expressed by embryonic cortical and hippocampal neurons, and that downregulation of LRP4 in these neurons causes a reduction in density of synapses and number of primary dendrites. Accordingly, overexpression of LRP4 in cultured neurons had the opposite effect inducing more but shorter primary dendrites with an increased number of spines. Transsynaptic tracing mediated by rabies virus revealed a reduced number of neurons presynaptic to the cortical neurons in which LRP4 was knocked down. Moreover, neuron-specific knockdown of LRP4 by in utero electroporation of LRP4 miRNA in vivo also resulted in neurons with fewer primary dendrites and a lower density of spines in the developing cortex and hippocampus. Collectively, our results demonstrate an essential and novel role of neuronal LRP4 in dendritic development and synaptogenesis in the CNS.

SUMO1 Modification Facilitates Avibirnavirus Replication by Stabilizing Polymerase VP1.

SUMOylation is a posttranslational modification that has crucial roles in diverse cellular biological pathways and in various viral life cycles. In this study, we found that the VP1 protein, the RNA-dependent RNA polymerase of avibirnavirus infectious bursal disease virus (IBDV), regulates virus replication by SUMOylation during infection. Our data demonstrated that the polymerase VP1 is efficiently modified by small ubiquitin-like modifier 1 (SUMO1) in avibirnavirus-infected cell lines. Mutation analysis showed that residues 404I and 406I within SUMO interaction motif 3 of VP1 constitute the critical site for SUMO1 modification. Protein stability assays showed that SUMO1 modification enhanced significantly the stability of polymerase VP1 by inhibiting K48-linked ubiquitination. A reverse genetic approach showed that only IBDV with I404C/T and I406C/F mutations of VP1 could be rescued successfully with decreased replication ability. Our data demonstrated that SUMO1 modification is essential to sustain the stability of polymerase VP1 during IBDV replication and provides a potential target for designing antiviral drugs targeting IBDV.IMPORTANCE SUMOylation is an extensively discussed posttranslational modification in diverse cellular biological pathways. However, there is limited understanding about SUMOylation of viral proteins of IBDV during infection. In the present study, we revealed a SUMO1 modification of VP1 protein, the RNA-dependent RNA polymerase of avibirnavirus infectious bursal disease virus (IBDV). The required site of VP1 SUMOylation comprised residues 404I and 406I of SUMO interaction motif 3, which was essential for maintaining its stability by inhibiting K48-linked ubiquitination. We also showed that IBDV with SUMOylation-deficient VP1 had decreased replication ability. These data demonstrated that the SUMOylation of IBDV VP1 played an important role in maintaining IBDV replication.

Ubiquitination Is Essential for Avibirnavirus Replication by Supporting VP1 Polymerase Activity.

Ubiquitination is critical for several cellular physical processes. However, ubiquitin modification in virus replication is poorly understood. Therefore, the present study aimed to determine the presence and effect of ubiquitination on polymerase activity of viral protein 1 (VP1) of avibirnavirus. We report that the replication of avibirnavirus is regulated by ubiquitination of its VP1 protein, the RNA-dependent RNA polymerase of infectious bursal disease virus (IBDV). In vivo detection revealed the ubiquitination of VP1 protein in IBDV-infected target organs and different cells but not in purified IBDV particles. Further analysis of ubiquitination confirms that VP1 is modified by K63-linked ubiquitin chain. Point mutation screening showed that the ubiquitination site of VP1 was at the K751 residue in the C terminus. The K751 ubiquitination is independent of VP1's interaction with VP3 and eukaryotic initiation factor 4A II. Polymerase activity assays indicated that the K751 ubiquitination at the C terminus of VP1 enhanced its polymerase activity. The K751-to-R mutation of VP1 protein did not block the rescue of IBDV but decreased the replication ability of IBDV. Our data demonstrate that the ubiquitination of VP1 is crucial to regulate its polymerase activity and IBDV replication.IMPORTANCE Avibirnavirus protein VP1, the RNA-dependent RNA polymerase, is responsible for IBDV genome replication, gene expression, and assembly. However, little is known about its chemical modification relating to its polymerase activity. In this study, we revealed the molecular mechanism of ubiquitin modification of VP1 via a K63-linked ubiquitin chain during infection. Lysine (K) residue 751 at the C terminus of VP1 is the target site for ubiquitin, and its ubiquitination is independent of VP1's interaction with VP3 and eukaryotic initiation factor 4A II. The K751 ubiquitination promotes the polymerase activity of VP1 and unubiquitinated VP1 mutant IBDV significantly impairs virus replication. We conclude that VP1 is the ubiquitin-modified protein and reveal the mechanism by which VP1 promotes avibirnavirus replication.

Overexpression of Dnmt3a ameliorates diabetic muscle atrophy by modulating the Pten/Akt pathway.

NEW FINDINGS: What is the central question of this study? Does Dnmt3a play a crucial role in regulating diabetic muscle atrophy? What is the main finding and its importance? Muscle atrophy is one of the major long-term complications of diabetes mellitus. However, little is known about the molecular mechanism involved. In this paper, we demonstrated that Dnmt3a overexpression effectively improves the diabetic muscle health in mice and documented the underlying mechanisms. DNMT3A might become a promising target to prevent muscle atrophy in patients with diabetes. ABSTRACT: Muscle atrophy is one of the major long-term complications of diabetes mellitus, which greatly affects the mobility of patients. Epigenetic processes mediated by DNA methyltransferases (DNMTs) play crucial roles in the locomotor system, but little is known about the functions of DNMTs in diabetic muscle atrophy. Here, we investigated the function of Dnmt3a in diabetic muscle atrophy and explored the mechanisms involved. Adeno-associated virus AAV2 overexpressing Dnmt3a or its vector control was injected into the tibialis anterior muscle of streptozotocin-induced diabetic mice. Muscle mass and muscle cross-sectional area were used to evaluate muscle atrophy. In vitro, adeno-associated virus AAV2 overexpressing Dnmt3a or its vector control was transfected into C2C12 myoblasts. Horse serum was used to induce differentiation and palmitate to stimulate the C2C12 myoblasts. The expressions of myogenic regulatory factors were examined by real-time PCR and western blot analysis. Overexpression of Dnmt3a attenuated muscle atrophy in diabetic mice and promoted myotube formation of C2C12 myoblasts. Overexpression of Dnmt3a restored the expressions of myogenic regulatory factors atrogin-1, MuRF1, Pax7, Myod1 and myogenin, both in vivo and in vitro. Moreover, overexpression of Dnmt3a activated the phosphorylation of Akt by inhibiting the activation of Pten. This study demonstrates that overexpression of Dnmt3a prevents diabetic muscle atrophy by modulating the Pten/Akt pathway.

Long noncoding RNA LINC00511 regulates the proliferation, apoptosis, invasion and autophagy of trophoblast cells to mediate pre-eclampsia progression through modulating the miR-31-5p/homeobox protein A7 axis.

AIM: Pre-eclampsia (PE) is the usual complication during pregnancy. Long noncoding RNAs are essential regulatory factors in many diseases. Nevertheless, the role of LINC00511 in the development of PE has not been fully elucidated. METHODS: The expression of LINC00511, homeobox protein A7 (HOXA7) and miR-31-5p was determined by quantitative real-time polymerase chain reaction. The levels of HOXA7 protein and autophagy-related proteins were measured by western blot analysis. Besides, cell proliferation was evaluated using cell counting kit 8 and colony formation assays. The apoptosis and invasion of cells were detected via flow cytometry and transwell assay, respectively. Further, the interaction between miR-31-5p and LINC00511 or HOXA7 was confirmed by dual-luciferase reporter assay. RESULTS: The LINC00511 and HOXA7 expression levels were decreased in placental tissues of PE patients, and the expression levels of both were positively correlated. LINC00511 knockdown suppressed proliferation, invasion and autophagy, while enhanced apoptosis in trophoblast cells. Meanwhile, the elevated HOXA7 expression promoted proliferation, invasion, autophagy, and inhibited the apoptosis of trophoblast cells. Besides, overexpression of HOXA7 also could reverse the effect of LINC00511 knockdown on the biological function of trophoblast cells. Further experiments confirmed that miR-31-5p could be sponged by LINC00511 and could target HOXA7. Also, miR-31-5p mimic could invert the promoting effect of LINC00511 overexpression on the biological function of trophoblast cells. CONCLUSION: LINC00511 expression was crucial for maintaining the normal function of trophoblast cells, and the decreased its expression might promote the progress of PE, which might provide some theoretical strategies for reducing the development of PE.

Residential exposure to natural outdoor environments and general health among older adults in Shanghai, China.

BACKGROUND: Exposure to natural outdoor environments (NOE) has been shown to be beneficial to older adults' health and functioning, yet this assertion has rarely been tested in China. We investigated the relationships between exposure to NOE and older adults' self-rated health in Shanghai, China and examined whether these relationships varied by sex, age, education and hukou status. METHOD: This cross-sectional study used micro-data sample of the 2010 Shanghai population census, including 7962 older adults nested within 3345 neighbourhoods. Self-rated health was the outcome variable. Four NOE exposure indicators were calculated for each neighbourhood: the amount of surrounding greenness/blueness and proximity to large green/blue spaces. Multilevel logistic regression was employed to explore the association between natural outdoor environment exposure and self-rated health, adjusting for individual-level and neighbourhood-level covariates. Stratified analyses were used to examine variations by sex, age, education and hukou status. RESULTS: Older adults living in neighbourhoods with higher surrounding greenness and higher proximity to both green spaces and blue spaces were more likely to report good health. Residential surrounding blueness was not significantly related to self-rated health. Females, those aged 60-69 years, those who had elementary school or junior high school education and those with non-local hukou benefit more from residential surrounding greenness, and those aged 70-79 years and who had elementary school or junior high school education benefit more from residential proximity to blue spaces. CONCLUSIONS: Higher residential greenness and proximity to both green spaces and blue spaces were associated with better self-rated health, particularly for females, younger older adults, the low educated and non-local hukou holders. Our findings suggest that urban green spaces and urban blue spaces have different effects on health among Chinese older adults and that the assessment of exposure matters to the investigation of NOE-health relationships.

Protective effects of apigenin on LPS-induced endometritis via activating Nrf2 signaling pathway.

Endometritis is a puzzling disease that often associates with severe pelvic pain. In this study, we aimed to detect whether apigenin had protective effect against LPS-induced endometritis, if so, the underlying mechanism was further investigated. Apigenin was administrated 1 h before LPS treatment. The levels of inflammatory cytokines were measured by ELISA. The expression of NF-κB and Nrf2 were detected by Western blot analysis. The results showed that LPS treatment induced severe histological alteration of uterus and this change was attenuated by the treatment of apigenin. Apigenin significantly attenuated LPS-induced MPO activity, MDA content, and inflammatory cytokines TNF-α and IL-1ß production. LPS-induced NF-κB activation was suppressed by apigenin. Furthermore, apigenin elevated the expression of Nrf2 and HO-1 in uterine tissues. In conclusion, the present study demonstrated that apigenin protected against LPS-induced endometritis through activation of Nrf2 signaling pathway and inhibition of NF-κB signaling pathway.

Galactose-Functionalized Double-Hydrophilic Block Glycopolymers and Their Thermoresponsive Self-Assembly Dynamics.

Glycopolymers with large galactose units are attractive in biological processes because of their ability to selectively recognize lectin proteins. Recently, thermoresponsive double-hydrophilic block glycopolymers (TDHBGs) have been designed, which allow sugar residues to expose or hide via the lower critical solution temperature (LCST)-type phase transition. In this work, we first synthesize a new type of TDHBGs, composed of a thermoresponsive poly(di(ethylene glycol)methyl ether methacrylate) block and a galactose-functionalized, poly(6- O-vinyladipoyl-d-galactose) (POVNG) block. The LCST can be tuned by varying the size of the POVNG block. Then, we have systematically investigated their thermoresponsive self-assembly behavior, using static and dynamic light scattering techniques, combined with transmission electron microscopy (TEM) imaging. It is found that the TDHBGs possess both micellization and LCST-type transition, and there exist strong interactions between them, depending on the concentration and structure of the TDHBGs. It is particularly interesting that for the same type of TDHBGs under different conditions, such interactions result in rich morphologies of the formed micelles (or nanoparticles) such as spheres, hollow spheres, prolate ellipsoids, crystal-like, and so on, thus potentially enriching their biological applications by noting that they are hepatoma-targeting glycopolymers.

Decrease in newly generated oligodendrocytes leads to motor dysfunctions and changed myelin structures that can be rescued by transplanted cells.

NG2-glia in the adult brain are known to proliferate and differentiate into mature and myelinating oligodendrocytes throughout lifetime. However, the role of these newly generated oligodendrocytes in the adult brain still remains little understood. Here we took advantage of the Sox10-iCreERT2 x CAG-eGFP x Esco2fl/fl mouse line in which we can specifically ablate proliferating NG2-glia in adult animals. Surprisingly, we observed that the generation of new oligodendrocytes in the adult brain was severely affected, although the number of NG2-glia remained stable due to the enhanced proliferation of non-recombined cells. This lack of oligodendrogenesis led to the elongation of the nodes of Ranvier as well as the associated paranodes, which could be locally rescued by myelinating oligodendrocytes differentiated from transplanted NG2-glia deriving from wildtype mice. Repetitive measurements of conduction velocity in the corpus callosum of awake animals revealed a progressive deceleration specifically in the mice lacking adult oligodendrogenesis that resulted in progressive motor deficits. In summary, here we demonstrated for the first time that axon function is not only controlled by the reliable organization of myelin, but also requires a dynamic and continuous generation of new oligodendrocytes in the adult brain. GLIA 2016;64:2201-2218.

Crosstalk between 6-OHDA-induced autophagy and apoptosis in PC12 cells is mediated by phosphorylation of Raf-1/ERK1/2.

Parkinson's disease (PD) is a degenerative brain disorder characterized by motor symptoms and loss of dopaminergic (DA) neurons in the substantia nigra. The mechanisms for DA cell death in PD have been extensively investigated using PC12 cells treated with a dopamine neurotoxin 6-hydroxydopamine (6-OHDA). 6-OHDA may induce both autophagy and apoptosis in PC12 cells. However, it remains unclear whether crosstalk occurs between autophagy and apoptosis in PC12 cells treated with 6-OHDA and whether Raf-1/ERK1/2 and their phosphorylation status play a role in autophagy. In this study, we used MDC staining assay and flow cytometry and found that 6-OHDA induced autophagy in PC12 cells. This induction was inhibited by the autophagy inhibitor 3-MA. Our electron microscopy observations also supported 6-OHDA induced autophagy in PC12 cells. Apoptosis of PC12 cells was increased with inhibition of autophagy by 3-MA. In addition, Inhibition of Raf-1 resulted in a decreased 6-OHDA-induced autophagy rate among PC12 cells. Phosphorylation levels of Raf-1 and ERK1/2 were increased in PC12 cells treated with 6-OHDA and inhibited by co-treatment with 6-OHDA and 3-MA. These data suggest that crosstalk between 6-OHDA-induced apoptosis and autophagy in PC12 cells may be regulated via the Raf-1/ERK1/2 signaling pathway. Our data suggest a mechanism for 6-OHDA toxicity in PC12 cells, contributing to our understanding of the pathogenesis of PD.

Formation of cholinergic synapse-like specializations at developing murine muscle spindles.

Muscle spindles are complex stretch-sensitive mechanoreceptors. They consist of specialized skeletal muscle fibers, called intrafusal fibers, which are innervated in the central (equatorial) region by afferent sensory axons and in both polar regions by efferent γ-motoneurons. We show that AChRs are concentrated at the γ-motoneuron endplate as well as in the equatorial region where they colocalize with the sensory nerve ending. In addition to the AChRs, the contact site between sensory nerve ending and intrafusal muscle fiber contains a high concentration of choline acetyltransferase, vesicular acetylcholine transporter and the AChR-associated protein rapsyn. Moreover, bassoon, a component of the presynaptic cytomatrix involved in synaptic vesicle exocytosis, is present in γ-motoneuron endplates but also in the sensory nerve terminal. Finally, we demonstrate that during postnatal development of the γ-motoneuron endplate, the AChR subunit stoichiometry changes from the γ-subunit-containing fetal AChRs to the ε-subunit-containing adult AChRs, similar and approximately in parallel to the postnatal subunit maturation at the neuromuscular junction. In contrast, despite the onset of ε-subunit expression during postnatal development the γ-subunit remains detectable in the equatorial region by subunit-specific antibodies as well as by analysis of muscle spindles from mice with genetically-labeled AChR γ-subunits. These results demonstrate an unusual maturation of the AChR subunit composition at the annulospiral endings and suggest that in addition to the recently described glutamatergic secretory system, the sensory nerve terminals are also specialized for cholinergic synaptic transmission, synaptic vesicle storage and exocytosis.