Sleep Problems Influence Emotional/Behavioral Symptoms and Repetitive Behavior in Preschool-Aged Children With Autism Spectrum Disorder in the Unique Social Context of China.

Sleep disturbances are common in people with autism spectrum disorder (ASD), but research on this topic is still limited in China. In the current study, we evaluated the prevalence of sleep problems in preschool-aged children with ASD and to examine the correlations between sleep disturbances and emotional/behavioral symptoms and repetitive behavior in the unique social context of China. This study recruited 475 preschool-aged children aged 3-6 years old, including 252 children with ASD (mean age 5.13 ± 1.15, 80.6% male) and 223 age-matched typically developing (TD) children (mean age 5.12 ± 0.97, 74.9% male). The parents of all children completed a sociodemographic questionnaire and the Childhood Sleep Habits Questionnaire. The parents of 114 ASD children completed the Strengths and Difficulties Questionnaire (SDQ) and the Repetitive Behavioral Questionnaire-2 (RBQ-2). The prevalence of sleep problems in preschool-aged children with ASD in this study was 81.7%, which was higher than that in TD children (61.0%). The scores for bedtime resistance, sleep anxiety, sleep duration, parasomnias, and sleep onset delay in the ASD group were significantly higher than those in the TD group (t=-7.664, P=0.000; t=-10.477, P=0.000; t=-4.133, P=0.000; Z=-3.916, P=0.000; Z=-7.093, P=0.000; respectively). Sleep onset delay explained 17.3% of the variance (adjusted R2 = 0.173) in the total SDQ score of children with ASD, and bedtime resistance explained a large proportion of total RBQ-2 score variance (adjusted R2 = 0.206). The high rate of sleep disturbances in preschool-aged children with ASD emphasizes the importance of screening for sleep problems in this population. Attention should also be directed toward formulating good sleep hygiene practices for preschool-aged children in the particular social context and cultural setting of China.

Sensory Processing Problems and Comorbidities in Chinese Preschool Children with Autism Spectrum Disorders.

Little is known about the relationship between sensory processing problems and sleep disturbances, emotional and behavioral problems and mealtime behavioral problems in Chinese children with autism spectrum disorders (ASDs). This study examined those relationships in Chinese preschool children with ASD using a case-control design. Atypical sensory processing was associated with increased risks of sleep disturbances, emotional and behavioral problems, and abnormal mealtime behaviors in the children with ASD, whereas sensory processing problems were significantly correlated with abnormal mealtime behaviors only in the typically developing children. Based on our findings, clinicians must collect information about sensory problems when a child with ASD experiences sleep disturbances and emotional and behavioral problems or presents abnormal mealtime behaviors.

[Core prescription excavation of launched Chinese patent medicines for treating different syndromes of angina pectoris].

To analyze the medication features and regularity of prescriptions of traditional Chinese medicines in treating patients with coronary heart disease angina pectoris based on the launched Chinese patent medicines. In the article,we collected all of the launched Chinese patent medicines for treating coronary heart disease and angina pectoris from the Chinese patent medicine value assessment information database,and set up a medical record normalized database,then carried out the classification of syndromes. The medication features and prescription rules for angina pectoris were analyzed by frequency statistics and association rules(IBM SPSS Modeler 14. 1 Premiums software,Apriori algorithm). Finally,a total of 170 prescriptions were selected,and 197 Chinese herbs were included,involving to totally 11 syndrome types,in which blood stasis syndrome,Qi stagnation and blood stasis syndrome,Qi deficiency and blood stasis syndrome,Qi-Yin deficiency and blood stasis syndrome were the main syndrome types. The frequency of single-herb medicines for the four main syndrome types,the combination of commonly used medicines,and the core prescriptions were summarized. After comparing the core prescriptions of the four syndrome types,we could analyze the medication features and prescription rules. In conclusion,the therapeutic principle is blood-activating and stasis-dissolving,and consideration was also given to promoting Qi,invigorating Qi and resuscitation and invigorating Qi-Yin. The main medicines include Danshen(Salvia Miltiorrhizae Radix et Rhizoma) and Chuan-xiong(Chuanxiong Rhizoma). According to different types of syndromes,Chinese herbal medicines are added,such as Jiangxiang(Dalbergiae Odoriferae Lignum), Chishao(Paeoniaeradix Rubra), Sanqi(Notoginseng Radix et Rhizoma), Honghua(Carthami Flos),Bingpian(Borneolum Syntheticum),Renshen(Ginseng Radix et Rhizoma). Frequency statistics and association rules are combined to explore the medication features and core prescriptions,which provide ideas for the treatment of angina pectoris and the development of new drugs.

Two single-nucleotide polymorphisms of the RELN gene and symptom-based and developmental deficits among children and adolescents with autistic spectrum disorders in the Tianjin, China.

Increasing evidence has revealed that genetic variants in Reelin (RELN) gene, especially single-nucleotide polymorphisms (SNPs), correlate with autistic spectrum disorders (ASD) risk; however, no consensus have been reached. This study aimed to provide additional evidence for the association between two SNPs of RELN (i.e., rs736707, rs2229864) and ASD risk, as well as the relationship between RELN gene and symptom-based and developmental deficits of ASD patients in Chinese Han children and adolescents. 157 ASD subjects and 256 typical development (TD) controls were genotyped by TaqMan® genotyping assay. ASD patients were assessed by Childhood Autism Rating Scale (CARS), Autism Behavior Checklist (ABC), and Early Childhood Development Questionnaire (ECDQ). We found that SNP rs2229864 was associated with the genetic predisposition of ASD, whereas a negative association between SNP rs2229864 and symptom-based and developmental features was detected. In contrast, RELN rs736707 correlated with the sensory subscale of the ABC, the relating subscale of the ABC and the total score of ABC, although we did not detect a significant association between SNP rs736707 and ASD risk. Furthermore, a significant rs736707-rs2229864 haplotype was detected. Individuals with a CC haplotype were more likely to have ASD, but individuals with a CT haplotype had more chance be TD controls. Further studies using more samples and including more gene variants in RELN are warranted to confirm our results.

[Absorption enhancing effects and safety of PAMAM dendrimers on liquiritin].

PAMAM dendrimer is one of the most widely studied dendrimers in recent years, which has a large number of functional groups on the surface and cavities inside, specific three-dimensional structure and good biocompatibility, permeability and stability. It has been widely applied in drug and gene carrier fields and may become a new absorption enhancer. In order to study the absorption enhancing effects of PAMAM dendrimers, liquiritin was selected as the model drug, with the protection of spleen and liver, detoxification and other functions, but it had not been widely used in clinical application because of its difficult absorption, first pass effect, and low bioavailability. This topic was based on the two main determinants (solubility and permeability) of intestinal absorption in the body, researched the physicochemical properties of liquiritin, analyzed the transport volume of liquiritin with or without PAMAM dendrimers by using Caco-2 cell model, and analyzed the cytotoxicity of PAMAM dendrimers on Caco-2 cells by MTT experiments. These results showed that 0.1% of the G4 generation PAG can promote the absorption of liquiritin safely and effectively, and it was suitable for further development into a new type of pharmaceutical excipients.

Nuclear receptor retinoid-related orphan receptor α deficiency exacerbates high-fat diet-induced cardiac dysfunction despite improving metabolic abnormality.

Retinoid-related orphan receptor α (RORα), a member of the metabolic nuclear receptor superfamily, plays a vital regulatory role in circadian rhythm and metabolism. Here, we investigated the role of RORα in high-fat diet (HFD)-induced cardiac impairments and the underlying mechanisms involved. RORα-deficient stagger mice (sg/sg) and wild type (WT) littermates were fed with either standard diet or HFD. At 20weeks after HFD treatment, RORα deficiency resulted in significantly decreased body weight gain, improved dyslipidemia and ameliorated insulin resistance (evaluated by blood biochemical and glucose/insulin tolerance tests) compared with WT control. However, compared with HFD-treated WT mice, HFD-treated sg/sg mice exhibited significantly augmented myocardial hypertrophy, cardiac fibrosis (wheat germ agglutinin, masson trichrome and sirius red staining) and cardiac dysfunction (echocardiography and hemodynamics). Mechanistically, RORα deficiency impaired mitochondrial biogenesis and function. Additionally, RORα deficiency resulted in inhibition of the AMPK-PGC1α signaling pathway. In contrast, cardiomyocyte-specific RORα overexpression ameliorated myocardial hypertrophy, fibrosis and dysfunction by restoring AMPK-PGC1α signaling, and subsequently normalizing mitochondrial biogenesis. These findings demonstrated for the first time that nuclear receptor RORα deficiency aggravated HFD-induced myocardial dysfunction at least in part by impairing mitochondrial biogenesis in association with disrupting AMPK-PGC1α signaling. This article is part of a Special Issue entitled: Genetic and epigenetic control of heart failure - edited by Jun Ren and Megan Yingmei Zhang.

Reproductive toxicity and gender differences induced by cadmium telluride quantum dots in an invertebrate model organism.

Sexual glands are key sites affected by nanotoxicity, but there is no sensitive assay for measuring reproductive toxicity in animals. The aim of this study was to investigate the toxic effects of cadmium telluride quantum dots (CdTe-QDs) on gonads in a model organism, Bombyx mori. After dorsal vein injection of 0.32 nmol of CdTe-QDs per individual, the QDs passed through the outer membranes of gonads via the generation of ROS in the membranes of spermatocysts and ovarioles, as well as internal germ cells, thereby inducing early germ cell death or malformations via complex mechanisms related to apoptosis and autophagy through mitochondrial and lysosomal pathways. Histological observations of the gonads and quantitative analyses of germ cell development showed that the reproductive toxicity was characterized by obvious male sensitivity. Exposure to QDs in the early stage of males had severe adverse effects on the quantity and quality of sperm, which was the main reason for the occurrence of unfertilized eggs. Ala- or Gly-conjugated QDs could reduce the nanotoxicity of CdTe-QDs during germ cell development and fertilization of their offspring. The results demonstrate that males are preferable models for evaluating the reproductive toxicity of QDs in combined in vivo/in vitro investigations.

Effect of glucagon-like peptide-1 on major cardiovascular outcomes in patients with type 2 diabetes mellitus: A meta-analysis of randomized controlled trials.

The effect of glucagon-like peptide-1 (GLP-1) treatment in patients with type 2 diabetes mellitus (T2DM) remains controversial. The purpose of this study was to compare the effect of GLP-1 and placebo/conventional antidiabetic agents on cardiovascular risk in T2DM patients. PubMed, EmBase and the Cochrane Library were searched to identify its eligible studies as well as manual searches for the reliability of this study. All eligible trials were performed in T2DM patients who received GLP-1 therapy or placebo/conventional antidiabetic agents. The reported outcomes included major cardiovascular events (MACE), and total mortality. Of 490 identified studies, we included 13 trials reporting data on 11,943 T2DM patients. Overall, the pooled results suggested that GLP-1 therapy has no or little effect on MACE (RR: 0.99; 95% CI: 0.88-1.12; P=0.872) and total mortality (RR: 0.90; 95% CI: 0.70-1.15; P=0.399). Furthermore, sensitivity analysis indicated that GLP-1 was associated with lower incidence of total mortality (RR: 0.28; 95% CI: 0.08-0.93; P=0.037). We concluded that GLP-1 therapy was not associated with MACE and total mortality compared with placebo or antidiabetic agents.

Impact of fluorescent silicon nanoparticles on circulating hemolymph and hematopoiesis in an invertebrate model organism.

Silicon nanoparticles (SiNPs) have attractive potential applications in biological and medical fields, and yet their impact on animals is still controversial, and there have been no reports of their effects on hematopoiesis. In this study, the effects of SiNPs on hemocytes and hematopoiesis were investigated by administering SiNPs via a vascular injection into an invertebrate model, the silkworm. Our results show that the ability of SiNPs to enter different types of circulating hemocytes and their impact on those hemocytes differed significantly. Rapid accumulation of SiNPs was observed in granulocytes, oenocytoids, and spherulocytes, which have immune functions in the circulating hemolymph, whereas SiNPs did not easily enter prohemocytes, which can differentiate into granulocytes, oenocytoids, and spherulocytes and replenish them. The SiNPs that entered the hemocytes initiated autophagy and apoptosis via the lysosomal/mitochondrial pathway. High-dose SiNPs weakly stimulated lysosomal activity in hematopoietic organs, but did not lead to a significant increase in reactive oxygen species or severe autophagy or apoptosis in the organ tissues. We suggest that the damage caused by high-dose SiNPs to hematopoiesis is self-healing, because few SiNPs entered the hematopoietic stem cells in the circulating hemolymph, so the damage to the hematopoietic tissues was limited.

Ubiquitin-Specific Protease 4 Is an Endogenous Negative Regulator of Pathological Cardiac Hypertrophy.

Dysregulation of the ubiquitin proteasome system components ubiquitin ligases and proteasome plays an important role in the pathogenesis of cardiac hypertrophy. However, little is known about the role of another ubiquitin proteasome system component, the deubiquitinating enzymes, in cardiac hypertrophy. Here, we revealed a crucial role of ubiquitin specific protease 4 (USP4), a deubiquitinating enzyme prominently expressed in the heart, in attenuating pathological cardiac hypertrophy and dysfunction. USP4 levels were consistently decreased in human failing hearts and in murine hypertrophied hearts. Adenovirus-mediated gain- and loss-of-function approaches indicated that deficiency of endogenous USP4 promoted myocyte hypertrophy induced by angiotensin II in vitro, whereas restoration of USP4 significantly attenuated the prohypertrophic effect of angiotensin II. To corroborate the role of USP4 in vivo, we generated USP4 global knockout mice and mice with cardiac-specific overexpression of USP4. Consistent with the in vitro study, USP4 depletion exacerbated the hypertrophic phenotype and cardiac dysfunction in mice subjected to pressure overload, whereas USP4 transgenic mice presented ameliorated pathological cardiac hypertrophy compared with their control littermates. Molecular analysis revealed that USP4 deficiency augmented the activation of the transforming growth factor ß-activated kinase 1 (TAK1)-(JNK1/2)/P38 signaling in response to hypertrophic stress, and blockage of TAK1 activation abolished the pathological effects of USP4 deficiency in vivo. These findings provide the first evidence for the involvement of USP4 in cardiac hypertrophy, and shed light on the therapeutic potential of targeting USP4 in the treatment of cardiac hypertrophy.

Targeting and retention enhancement of quantum dots decorated with amino acids in an invertebrate model organism.

The use of quantum dots (QDs) in biological imaging applications and targeted drug delivery is expected to increase. However, the efficiency of QDs in drug targeting needs to be improved. Here, we show that amino acids linked to CdTe QDs significantly increased the targeted transfer efficiency and biological safety in the invertebrate model Bombyx mori. Compared with bare QDs530, the transfer efficiency of Ala- and Gly-conjugated QDs (QDs530-Ala and QDs530-Gly) in circulatory system increased by 2.6 ± 0.3 and 1.5 ± 0.3 times, and increased by 7.8 ± 0.9 and 2.9 ± 0.2 times in target tissue silk glands, respectively, after 24 h of QDs exposure. Meanwhile, the amount of conjugated QDs decreased by (68.4 ± 4.4)% and (46.7 ± 9.1)% in the non-target tissue fat body, and the speed at which they entered non-target circulating blood cells significantly decreased. The resultant QDs530-Ala revealed a better structural integrity in tissues and a longer retention time in hemolymph than that of QDs530 after exposure via the dorsal vessel. On the other hand, QDs530-Ala significantly reduced the toxicity to hemocytes, silk gland, and fat body, and reduced the amount of reactive oxygen species (ROS) in tissues.

Does Periconceptional Fish Consumption by Parents Affect the Incidence of Autism Spectrum Disorder and Intelligence Deficiency? A Case-control Study in Tianjin, China.

OBJECTIVE: This study aimed to explore the association between periconceptional fish consumption by parents and autism spectrum disorder (ASD) and intelligence deficiency (ID). METHODS: A case-control study was conducted through a questionnaire with 108 ASD cases, 79 ID cases, and 108 controls. The ASD and ID cases were students from special educational schools in Tianjin from 2012 to 2014. The age- and sex-matched controls were from a high school, three primary schools, and a kindergarten in Tianjin. Multivariate logistic regression was performed. RESULTS: Paternal habit of eating hairtail before fertilization, maternal preference for fruits during pregnancy, and maternal habit of eating grass carp during pregnancy were preventive factors for ASD. Paternal habit of drinking alcohol before fertilization was a risk factor for ID, whereas maternal preference for fruits during pregnancy and maternal habit of eating crucian carp during pregnancy were protective factors for ID. CONCLUSION: Parental fish consumption is beneficial for the prevention of ASD and ID. Meanwhile, the protective effects of fish consumption on ASD and ID differ. More attention should be paid to the combined effect of other food when eating fish.

Association between Prenatal Environmental Factors and Child Autism: A Case Control Study in Tianjin, China.

OBJECTIVE: To investigate the association between autism and prenatal environmental risk factors. METHODS: A case-control study was conducted among 193 children with autism from the special educational schools and 733 typical development controls matched by age and gender by using questionnaire in Tianjin from 2007 to 2012. Statistical analysis included quick unbiased efficient statistical tree (QUEST) and logistic regression in SPSS 20.0. RESULTS: There were four predictors by QUEST and the logistic regression analysis, maternal air conditioner use during pregnancy (OR=0.316, 95% CI: 0.215-0.463) was the single first-level node (χ²=50.994, P=0.000); newborn complications (OR=4.277, 95% CI: 2.314-7.908) and paternal consumption of freshwater fish (OR=0.383, 95% CI: 0.256-0.573) were second-layer predictors (χ²=45.248, P=0.000; χ²=24.212, P=0.000); and maternal depression (OR=4.822, 95% CI: 3.047-7.631) was the single third-level predictor (χ²=23.835, P=0.000). The prediction accuracy of the tree was 89.2%. CONCLUSION: The air conditioner use during pregnancy and paternal freshwater fish diet might be beneficial for the prevention of autism, while newborn complications and maternal depression might be the risk factors.

Abnormal transsulfuration metabolism and reduced antioxidant capacity in Chinese children with autism spectrum disorders.

Autism spectrum disorder (ASD) is a neurological disorder that presents a spectrum of qualitative impairments in social interaction, communication, as well as restricted and stereotyped behavioral patterns, interests, and activities. Several studies have suggested that the etiology of ASD can be partly explained by oxidative stress. However, the implications of abnormal transsulfuration metabolism and oxidative stress, and their relation with ASD are still unclear. The purpose of this study was to evaluate several transsulfuration pathway metabolites in Chinese participants diagnosed with ASD, to better understand their role in the etiology of this disorder. Fifty children (39 male, 11 female) diagnosed with ASD and 50 age- and gender-matched non-ASD children (i.e., control group) were included in this study. This prospective blinded study was undertaken to assess transsulfuration and oxidative metabolites, including levels of homocysteine (Hcy), cysteine (Cys), total glutathione (tGSH), reduced glutathione (GSH), oxidized glutathione (GSSG), and glutathione ratio (GSH/GSSG). The clinical severity of ASD was evaluated with the Childhood Autism Rating Scale (CARS), and the autistic children's present behavior was measured by the Autism Behavior Checklist (ABC). The results indicated that Hcy and GSSG levels were significantly higher in children diagnosed with ASD, Cys, tGSH and GSH levels as well as the GSH/GSSG ratio showed remarkably lower values in ASD children compared to control subjects. Hcy levels correlated significantly with increasing CARS scores and GSSG levels in children with ASD. Our results suggest that an abnormal transsulfuration metabolism and reduced antioxidant capacity (i.e., hyperhomocysteinemia and increased oxidative stress), and Hcy level appears to have a potentially negative impact on clinical severity of autistic disorder.

Effect of exogenous methanol on glycolate oxidase and photorespiratory intermediates in cotton.

Application of methanol (MeOH) inhibits photorespiration and enhances growth and yield in C3 plants. However, the underlying cellular and molecular mechanisms are not clear. In this study, we investigated the effects of foliar application of MeOH (30%, v/v) on glycolate oxidase (GO) activity and photorespiratory intermediates in cotton leaves in a field experiment. MeOH treatment significantly inhibited GO activity (by 30% compared with the controls). We also found that endogenous glyoxylate, a photorespiratory intermediate, increased and glycine decreased significantly in MeOH-treated plants. Serine increased significantly in MeOH-treated plants. These results thus demonstrated that exogenous MeOH can modulate GO activity and the production of photorespiratory intermediates, and sheds new lights on our current understanding of how exogenous MeOH inhibits photorespiration and enhances the growth and yield of C3 plants such as cotton.

Role of RASSF1A promoter methylation in the pathogenesis of ovarian cancer: a meta-analysis.

OBJECTIVE: The aim of the current meta-analysis was to comprehensively assess the role of RASSF1A promoter methylation in the pathogenesis of ovarian cancer. METHOD: A range of electronic databases were searched: Web of Science (1945-2013), the Cochrane Library Database (Issue 12, 2013), PubMed (1966-2013), EMBASE (1980-2013), CINAHL (1982-2013), and the Chinese Biomedical Database (1982-2013) without language restrictions. Meta-analysis was conducted using the STATA 12.0 software. The crude odds ratio (OR) with its corresponding 95% confidence interval (CI) was calculated. RESULTS: Twelve clinical cohort studies with a total of 739 ovarian cancer patients were included in the current meta-analysis. The results of our meta-analysis suggested that the frequency of RASSF1A promoter methylation in cancer tissues was higher compared with benign, adjacent, and normal tissues (cancer tissues vs. benign tissues: OR=9.92, 95% CI: 7.67-12.82, p<0.001; cancer tissues vs. adjacent tissues: OR=68.15, 95% CI: 39.30-118.18, p<0.001; cancer tissues vs. normal tissues: OR=30.71, 95% CI: 23.12-40.80, p<0.001; respectively). Subgroup analysis based on ethnicity and sample types revealed that RASSF1A gene methylation was closely associated with the pathogenesis of ovarian cancer in all subgroups (all p<0.05). CONCLUSION: Our findings indicated that abnormal RASSF1A promoter methylation may be strongly correlated with the pathogenesis of ovarian cancer.

Hematopoiesis toxicity induced by CdTe quantum dots determined in an invertebrate model organism.

Quantum dots (QDs) have gained significant attention due to their superior optical properties and wide usage in biological and biomedical studies. In recent years, there has been intense concern regarding the in vivo toxicity of QDs. This study was undertaken to examine the toxicity of CdTe QDs on hematopoiesis in an invertebrate model organism, Bombyx mori. Vascular injection of sub-lethal doses of QDs in B. mori larvae caused time- and dose-dependent damage in the hematopoietic organ and hematocytes. QDs with the maximum emission wavelength of 530 nm (QDs530) were quickly observed in cystocytes and plasmacytes, and gradually bleached their green fluorescence, followed by a decrease in peripheral hematocytes. Additionally, the proportion of abnormal hematocytes increased. In marked contrast, QDs with the maximum emission wavelength of 720 nm (QDs720) were quickly surrounded by hematocytes and subsequently enriched in cystocytes like the human's leukocytes, but with weaker cytotoxicity. QDs exposure promoted the mitotic nucleus in prohemocytes and hematocytes similar to peripheral blood stem cells in humans, but aggravated apoptosis. A decrease in hematopoiesis was accompanied by shrinkage and death of hematopoietic organs via an increase in reactive oxygen species. QDs with smaller size resulted in more severe hematopoiesis toxicity.

KIF5B promotes the forward transport and axonal function of the voltage-gated sodium channel Nav1.8.

Nav1.8 is a tetrodotoxin-resistant voltage-gated sodium channel selectively expressed in primary sensory neurons. Peripheral inflammation and nerve injury induce Nav1.8 accumulation in peripheral nerves. However, the mechanisms and related significance of channel accumulation in nerves remains unclear. Here we report that KIF5B promotes the forward transport of Nav1.8 to the plasma membrane and axons in dorsal root ganglion (DRG) neurons of the rat. In peripheral inflammation induced through the intraplantar injection of complete Freund's adjuvant, increased KIF5 and Nav1.8 accumulation were observed in the sciatic nerve. The knock-down of KIF5B, a highly expressed member of the KIF5 family in DRGs, reduced the current density of Nav1.8 in both cultured DRG neurons and ND7-23 cells. Overexpression of KIF5B in ND7-23 cells increased the current density and surface expression of Nav1.8, which were abolished through brefeldin A treatment, whereas the increases were lost in KIF5B mutants defective in ATP hydrolysis or cargo binding. Overexpression of KIF5B also decreased the proteasome-associated degradation of Nav1.8. In addition, coimmunoprecipitation experiments showed interactions between the N terminus of Nav1.8 and the 511-620 aa sequence in the stalk domain of KIF5B. Furthermore, KIF5B increased Nav1.8 accumulation, Nav1.8 current, and neuronal excitability detected in the axons of cultured DRG neurons, which were completely abolished by the disruption of interactions between KIF5B and the N terminus of Nav1.8. Therefore, our results reveal that KIF5B is required for the forward transport and axonal function of Nav1.8, suggesting a mechanism for axonal accumulation of Nav1.8 in inflammatory pain.

Endosome-mediated retrograde axonal transport of P2X3 receptor signals in primary sensory neurons.

Neurotrophins and their receptors adopt signaling endosomes to transmit retrograde signals. However, the mechanisms of retrograde signaling for other ligand/receptor systems are poorly understood. Here, we report that the signals of the purinergic (P)2X(3) receptor, an ATP-gated ion channel, are retrogradely transported in dorsal root ganglion (DRG) neuron axons. We found that Rab5, a small GTPase, controls the early sorting of P2X(3) receptors into endosomes, while Rab7 mediates the fast retrograde transport of P2X(3) receptors. Intraplantar injection and axonal application into the microfluidic chamber of α, ß-methylene-ATP (α, ß-MeATP), a P2X selective agonist, enhanced the endocytosis and retrograde transport of P2X(3) receptors. The α, ß-MeATP-induced Ca(2+) influx activated a pathway comprised of protein kinase C, rat sarcoma viral oncogene and extracellular signal-regulated protein kinase (ERK), which associated with endocytic P2X(3) receptors to form signaling endosomes. Disruption of the lipid rafts abolished the α, ß-MeATP-induced ERK phosphorylation, endocytosis and retrograde transport of P2X(3) receptors. Furthermore, treatment of peripheral axons with α, ß-MeATP increased the activation level of ERK and cAMP response element-binding protein in the cell bodies of DRG neurons and enhanced neuronal excitability. Impairment of either microtubule-based axonal transport in vivo or dynein function in vitro blocked α, ß-MeATP-induced retrograde signals. These results indicate that P2X(3) receptor-activated signals are transmitted via retrogradely transported endosomes in primary sensory neurons and provide a novel signaling mechanism for ligand-gated channels.

Transmembrane segments prevent surface expression of sodium channel Nav1.8 and promote calnexin-dependent channel degradation.

The voltage-gated sodium channel (Na(v)) 1.8 contributes substantially to the rising phase of action potential in small dorsal root ganglion neurons. Na(v)1.8 is majorly localized intracellularly and its expression on the plasma membrane is regulated by exit from the endoplasmic reticulum (ER). Previous work has identified an ER-retention/retrieval motif in the first intracellular loop of Na(v)1.8, which prevents its surface expression. Here we report that the transmembrane segments of Na(v)1.8 also cause this channel retained in the ER. Using transferrin receptor and CD8α as model molecules, immunocytochemistry showed that the first, second, and third transmembrane segments in each domain of Na(v)1.8 reduced their surface expression. Alanine-scanning analysis revealed acidic amino acids as critical factors in the odd transmembrane segments. Furthermore, co-immunoprecipitation experiments showed that calnexin interacted with acidic amino acid-containing sequences through its transmembrane segment. Overexpression of calnexin resulted in increased degradation of those proteins through the ER-associated degradation pathway, whereas down-regulation of calnexin reversed the phenotype. Thus our results reveal a critical role and mechanism of transmembrane segments in surface expression and degradation of Na(v)1.8.