Alpha1 Subunit Research Articles

Abstract 13166: Paraoxonase-3 Regulation of Cardiotonic Steroids Mediates Renal Injury and Dysfunction in a Dahl Salt Sensitive Model of Chronic Kidney Disease

Background: Paraoxonases (Pons) belong to a family of hydrolytic enzymes with 3 distinct isoforms: Pon 1, 2, and 3. While the physiologic substrates for Pons are unclear, their lactonase activity is considered central to their native physiological role. We have previously demonstrated an association between diminished Pon lactonase activity and worse clinical outcomes in volume expanded states such as chronic kidney disease (CKD). Furthermore, we have identified a novel reno-protective role for Pon3 in hypertensive renal disease. However, the underlying mechanism(s) are unknown. Cardiotonic steroids (CTS) are natriuretic hormones that are increased in CKD and promote renal inflammation and fibrosis via binding of their 17β-lactone ring to the alpha-1 subunit of the Na + /K + -ATPase (NKA-α-1). Therefore, we tested the hypothesis that reduction of Pon3 lactonase activity would lead to increased renal injury and dysfunction in a CTS induced rat model of CKD. Methods/Results: Experiments were performed on Dahl salt-sensitive rats (SS rats) and Pon3 knock-out rats on the Dahl salt-sensitive background (SS- Pon3 KO). First, CTS levels in the urine of SS control and SS- Pon3 KO rats were quantified using mass spectrometry. We found that SS- Pon3 KO rats had 59 times greater increase in 24-hour urine CTS levels vs SS rats. Next, to examine the role of Pon3 in mediating CTS induced renal injury, ten-week-old, age-matched male SS control and SS- Pon3 KO rats were administered the CTS telocinobufagin (TCB, 100 μg/kg/day s.q) for four weeks followed by assessment of blood pressure (tail cuff plethysmography) and renal function. We found that SS- Pon3 KO rats following TCB administration had significantly increased 24-hour urinary protein excretion (UPE) (50.73 ± 3.59vs 25.29 ± 2.07, p<0.0001), as well as decreased FITC-Sinistrin monitored glomerular filtration rate (GFR) (0.80 ± 0.10 vs 1.16 ± 0.05, p<0.01) compared to SS control rats, despite similar degrees of hypertension. Conclusion: These findings suggest that Pon3 plays a critical role in regulating CTS mediated renal dysfunction in vivo.

A quantitative trait variant in Gabra2 underlies increased methamphetamine stimulant sensitivity.

Psychostimulant (methamphetamine, cocaine) use disorders have a genetic component that remains mostly unknown. We conducted genome-wide quantitative trait locus (QTL) analysis of methamphetamine stimulant sensitivity. To facilitate gene identification, we employed a Reduced Complexity Cross between closely related C57BL/6 mouse substrains and examined maximum speed and distance traveled over 30 min following methamphetamine (2 mg/kg, i.p.). For maximum methamphetamine-induced speed following the second and third administration, we identified a single genome-wide significant QTL on chromosome 11 that peaked near the Cyfip2 locus (LOD = 3.5, 4.2; peak = 21 cM [36 Mb]). For methamphetamine-induced distance traveled following the first and second administration, we identified a genome-wide significant QTL on chromosome 5 that peaked near a functional intronic indel in Gabra2 coding for the alpha-2 subunit of the GABA-A receptor (LOD = 3.6–5.2; peak = 34–35 cM [66–67 Mb]). Striatal cis-expression QTL mapping corroborated Gabra2 as a functional candidate gene underlying methamphetamine-induced distance traveled. CRISPR/Cas9-mediated correction of the mutant intronic deletion on the C57BL/6J background to the wild-type C57BL/6NJ allele was sufficient to reduce methamphetamine-induced locomotor activity toward the wild-type C57BL/6NJ-like level, thus validating the quantitative trait variant (QTV). These studies show the power and efficiency of Reduced Complexity Crosses in identifying causal variants underlying complex traits. Functionally restoring Gabra2 expression decreased methamphetamine stimulant sensitivity and supports preclinical and human genetic studies implicating the GABA-A receptor in psychostimulant addiction-relevant traits. Importantly, our findings have major implications for studying psychostimulants in the C57BL/6J strain—the gold standard strain in biomedical research.

CONGENITAL MUSCULAR DYSTROPHIES: EP.70 Exploring the role of genetic modifiers in a mild LAMA2-RD case associated with a LAMA2 loss-of-function mutation

Merosin-deficient congenital muscular dystrophy (LAMA2-RD) is a neuromuscular disorder caused by mutations in the LAMA2 gene, coding for the alpha2 subunit of laminin-211 (merosin). LAMA2-RD patients carrying LAMA2 loss-of-function mutations lack merosin and their invariably severe clinical phenotype is characterised by the inability to acquire ambulation. A wider and often milder spectrum of disease severity results from missense mutations if they allow the production of a partially functional protein.

Induced pluripotent stem cells for defining Parkinsonian patient subtypes: a further step toward precision medicine.

Induced pluripotent stem cells for defining Parkinsonian patient subtypes: a further step toward precision medicine.

The expression of the alpha1 subunit of Na+/K+-ATPase is related to tumor development and clinical outcomes in gastric cancer.

The Na+/K+-ATPase alpha1 subunit (ATP1A1) is a critical component of Na+/K+-ATPase (NKA), a membrane pump that maintains a low intracellular Na+/K+ ratio and retains cellular volume and osmolarity. ATP1A1 was recently implicated in tumor behavior. Therefore, the present study investigated the role of ATP1A1 in patients with gastric cancer (GC). Knockdown experiments were conducted on human GC cell lines using ATP1A1 siRNA, and its effects on proliferation, the cell cycle, apoptosis, and cellular movement were examined. Gene expression profiling was performed by a microarray analysis. Primary tumor samples from 192 GC patients who underwent gastrectomy were subjected to an immunohistochemical analysis. High ATP1A1 expression levels were observed in NUGC4 and MKN74 cells. Cell proliferation was suppressed and apoptosis was induced by the siRNA-induced knockdown of ATP1A1. The microarray analysis showed that knockdown of ATP1A1 leads to the up-regulated expression of genes involved in the interferon (IFN) signaling pathway, such as STAT1, STAT2, IRF1, and IRF9. Furthermore, the depletion of ATP1A1 altered the phosphorylation of the MAPK pathway. The immunohistochemical analysis revealed that the expression of ATP1A1 was associated with the histological type, venous invasion, and the pathological T stage. Furthermore, the prognostic analysis showed a relationship between high ATP1A1 expression levels and poor postoperative survival. ATP1A1 appears to regulate tumor progression by altering IFN signaling, and high ATP1A1 expression levels were associated with poor postoperative survival in GC patients. The present results provide novel insights into the function of ATP1A1 as a mediator and/or biomarker of GC.

Reduced Expression of GABA A Receptor Alpha2 Subunit Is Associated With Disinhibition of DYT-THAP1 Dystonia Patient-Derived Striatal Medium Spiny Neurons.

DYT-THAP1 dystonia (formerly DYT6) is an adolescent-onset dystonia characterized by involuntary muscle contractions usually involving the upper body. It is caused by mutations in the gene THAP1 encoding for the transcription factor Thanatos-associated protein (THAP) domain containing apoptosis-associated protein 1 and inherited in an autosomal-dominant manner with reduced penetrance. Alterations in the development of striatal neuronal projections and synaptic function are known from transgenic mice models. To investigate pathogenetic mechanisms, human induced pluripotent stem cell (iPSC)-derived medium spiny neurons (MSNs) from two patients and one family member with reduced penetrance carrying a mutation in the gene THAP1 (c.474delA and c.38G > A) were functionally characterized in comparison to healthy controls. Calcium imaging and quantitative PCR analysis revealed significantly lower Ca2+ amplitudes upon GABA applications and a marked downregulation of the gene encoding the GABAA receptor alpha2 subunit in THAP1 MSNs indicating a decreased GABAergic transmission. Whole-cell patch-clamp recordings showed a significantly lower frequency of miniature postsynaptic currents (mPSCs), whereas the frequency of spontaneous action potentials (APs) was elevated in THAP1 MSNs suggesting that decreased synaptic activity might have resulted in enhanced generation of APs. Our molecular and functional data indicate that a reduced expression of GABAA receptor alpha2 subunit could eventually lead to limited GABAergic synaptic transmission, neuronal disinhibition, and hyperexcitability of THAP1 MSNs. These data give pathophysiological insight and may contribute to the development of novel treatment strategies for DYT-THAP1 dystonia.

Temporal-parietal-occipital epilepsy in GEFS+ associated with SCN1A mutation.

Most families with genetic epilepsy with febrile seizures plus show a mutation in the sodium channel alpha1 subunit gene, however, but there is much phenotypic heterogeneity and focal epilepsy remains relatively rare. Here, we report a family with electroclinical features indicative of temporal-parietal-occipital carrefour epilepsy with common occurrence of post-ictal migraine. We studied a four-generation family including nine affected subjects by means of EEG and MRI. Genetic testing was performed by targeted re-sequencing (gene panel). In most patients, seizure semiology included cognitive, autonomic, and emotional symptoms, eventually evolving towards sensory visual phenomena. Focal sensory vestibular seizures and changes in body perception were also reported in some cases. Post-ictal migraine was common, occurring in five out of the six (83%) epilepsy patients. A missense mutation (c.1130 G>A; p.R377Q) affecting the S5-S6 segment (pore region) of the sodium channel alpha1 subunit was identified in all affected and four unaffected subjects. Temporal-parietal-occipital carrefour epilepsy is part of the genetic epilepsy with febrile seizures plus spectrum. The electroclinical features in this family support the involvement of a genetically impaired neural network. High prevalence of post-ictal migraine suggests the role of posterior brain areas in the clinical expression of this gene defect.

ALTERED EXPRESSION OF LUNG EPITHELIAL ION AND FLUID TRANSPORTERS DURING PROGRESSION OF PULMONARY HYPERTENSION IN THE MONOCROTALINE MODEL

Objective: Recent studies have evaluated a significant incidence of pleural effusions in patients with idiopathic and heritable pulmonary arterial hypertension (PAH) co-diagnosed with isolated right-sided heart failure (RHF). Coincidence of pleural effusions and RHF in these patients significantly increased mortality rate during follow-up. However, mechanism of pleural effusions formation has not been fully explained. Therefore, we examined expression of lung epithelial ion and fluid transporters responsible for alveolar fluid clearance (AFC), which may contribute to lung fluid accumulation under pathological conditions and potentially formation of pleural effusions. Design and method: 12-weeks old male Wistar rats were subcutaneously injected with monocrotaline (60 mg/kg, MCT) or vehicle (CON). The relative expression of AFC transporters: Na+/K+-ATPase alpha1, alpha2, beta1, beta3 subunits, epithelial sodium channel (ENaC) alpha, beta, gama subunits, cystic fibrosis transmembrane conductance regulator (CFTR) were determined by RT-qPCR 1, 2, 4 weeks after MCT application and also in end-stage disease (animals sacrificed after acute health deterioration, ∼ 4 weeks after MCT application) where pleural effusions were observed. Results: Increase in lung weight was observed from second week after MCT application and was more profound in end-stage disease (by 26 % compared to fourth week, p < 0.05). A downregulation of all three pore-forming subunits of ENaC was observed in end-stage disease, accompanied by downregulation of small beta3 subunit of Na+/K+-ATPase. Expression of catalytic alpha2 subunit of Na+/K+-ATPase gradually decreased with disease progression from first week after MCT (-80% in end-stage vs. CON, p < 0.05). On the other hand, we observed upregulation of CFTR (by 80%) and beta1 subunit of Na+/K+ATPase (by 79% vs. CON, p < 0.05) mainly in end-stage disease. Conclusions: Downregulation of lung epithelial ion and fluid transporters contributing to AFC is predominantly limited to higher stages of PH with a marked downregulation in end-stage. These expression alterations may contribute to overall lung weight increase observed in end-stage disease. However, observed early alpha2 subunit of Na+/K+-ATPase downregulation could participate on the pathogenesis of PH while upregulation of beta1 subunit and CFTR in end-stage disease could represent a compensatory mechanism.

IN SILICO DRUG REPOSITIONING OF TOPIRAMATE AS AN ANTIHYPERTENSIVE BY CONVERGING KIDNEY TRANSCRIPTOMIC, EPIGENOMIC AND GENOMIC DATA

Objective: No new antihypertensive medication has been introduced to clinical practice since 2007. Mendel's second law states that genotype is allocated randomly at conception therefore, short nucleotide variants acting in cis can be utilised to deduce the pharmacological action on the same protein as in an RCT. Here, we investigate new therapeutic opportunities for hypertension through exploiting the druggable genome. Design and method: We conducted expression(e), splicing(s) and methylation(m) quantitative trait loci (QTL) analysis on 430 human kidneys from 5 studies [TRANScriptome of renaL humAn TissuE (TRANSLATE) study, Tissue Cancer Genome Atlas (TCGA), moleculAr analysis of human kiDney-Manchester renal tIssue pRojEct (ADMIRE) study, Molecular analysis of mechanisms regulating gene expression in post-ischemic injury to renal allograft (REPAIR) and Renal gEne expreSsion and PredispOsition to cardiovascular and kidNey Disease (RESPOND)]. Through their intersection with blood pressure variants from all previous genome wide association studies (GWAS) we identified genetic variants partnering with protein-coding kidney e-Genes, s-Genes and m-Genes. We then mapped the proteins encoded by these genes onto a set of licensed drugs or compounds with bioactivities against these targets in-silico. Results: Altogether, 479 informative genetic variants mapping onto 918 protein coding genes were identified. Of these, 201 (21.9%) genes were druggable. We found 209 unique associations with 44 (21.9%) protein coding genes. Of these, 23 associations acting on 7 drug targets were precisely concordant (captopril associated with ACE gene) and 16 associations (4 targets) had the indication association within the same disease area [for example, ambrisentan (an endothelin receptor antagonist) acting on a BP GWAS gene EDNRA]. 3 (1.3%) drugs were identified as repurposing opportunities for hypertension based on clinical evidence and directionality of effect on gene activity. Of those, topiramate [target for the alpha-subunit of Gamma-Aminobutyric Acid Type A receipt Alpha2 Subunit (GABRA2) gene], was deemed as clinically strongest drug repurposing opportunity for hypertension. The alpha subunit of this receptor is coded by GABRA2 gene, which is a hypertension risk gene identified in our study. Conclusions: Integration of pharmacological databases with -omic data of tissue of key relevance to hypertension may uncover novel drug repurposing opportunities.

Neurotrophin receptor activation rescues cognitive and synaptic abnormalities caused by hemizygosity of the psychiatric risk gene Cacna1c

Genetic variation in CACNA1C, which encodes the alpha-1 subunit of CaV1.2 L-type voltage-gated calcium channels, is strongly linked to risk for psychiatric disorders including schizophrenia and bipolar disorder. To translate genetics to neurobiological mechanisms and rational therapeutic targets, we investigated the impact of mutations of one copy of Cacna1c on rat cognitive, synaptic and circuit phenotypes implicated by patient studies. We show that rats hemizygous for Cacna1c harbour marked impairments in learning to disregard non-salient stimuli, a behavioural change previously associated with psychosis. This behavioural deficit is accompanied by dys-coordinated network oscillations during learning, pathway-selective disruption of hippocampal synaptic plasticity, attenuated Ca2+ signalling in dendritic spines and decreased signalling through the Extracellular-signal Regulated Kinase (ERK) pathway. Activation of the ERK pathway by a small-molecule agonist of TrkB/TrkC neurotrophin receptors rescued both behavioural and synaptic plasticity deficits in Cacna1c+/− rats. These results map a route through which genetic variation in CACNA1C can disrupt experience-dependent synaptic signalling and circuit activity, culminating in cognitive alterations associated with psychiatric disorders. Our findings highlight targeted activation of neurotrophin signalling pathways with BDNF mimetic drugs as a genetically informed therapeutic approach for rescuing behavioural abnormalities in psychiatric disorder.

Alterations in nicotinic receptor alpha5 subunit gene differentially impact early and later stages of cocaine addiction: a translational study in transgenic rats and patients.

Cocaine addiction is a chronic and relapsing disorder with an important genetic component. Human candidate gene association studies showed that the single nucleotide polymorphism (SNP) rs16969968 in the α5 subunit (α5SNP) of nicotinic acetylcholine receptors (nAChRs), previously associated with increased tobacco dependence, was linked to a lower prevalence of cocaine use disorder (CUD). Three additional SNPs in the α5 subunit, previously shown to modify α5 mRNA levels, were also associated with CUD, suggesting an important role of the subunit in this pathology. To investigate the link between this subunit and CUD, we submitted rats knockout for the α5 subunit gene (α5KO), or carrying the α5SNP, to cocaine self-administration (SA) and showed that the acquisition of cocaine-SA was impaired in α5SNP rats while α5KO rats exhibited enhanced cocaine-induced relapse associated with altered neuronal activity in the nucleus accumbens. In addition, we observed in a human cohort of patients with CUD that the α5SNP was associated with a slower transition from first cocaine use to CUD. We also identified a novel SNP in the β4 nAChR subunit, part of the same gene cluster in the human genome and potentially altering CHRNA5 expression, associated with shorter time to relapse to cocaine use in patients. In conclusion, the α5SNP is protective against CUD by influencing early stages of cocaine exposure while CHRNA5 expression levels may represent a biomarker for the risk to relapse to cocaine use. Drugs modulating α5 containing nAChR activity may thus represent a novel therapeutic strategy against CUD.

Targeting the cholinergic anti-inflammatory pathway with vagus nerve stimulation in patients with Covid-19?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), at the origin of the worldwide COVID-19 pandemic, is characterized by a dramatic cytokine storm in some critical patients with COVID-19. This storm is due to the release of high levels of pro-inflammatory cytokines such as interleukin (IL)-1 β, IL-6, tumor necrosis factor (TNF), and chemokines by respiratory epithelial and dendritic cells, and macrophages. We hypothesize that this cytokine storm and the worsening of patients’ health status can be dampened or even prevented by specifically targeting the vagal-driven cholinergic anti-inflammatory pathway (CAP). The CAP is a concept that involves an anti-inflammatory effect of vagal efferents by the release of acetylcholine (ACh). Nicotinic acetylcholine receptor alpha7 subunit (α7nAChRs) is required for ACh inhibition of macrophage-TNF release and cytokine modulation. Hence, targeting the α7nAChRs through vagus nerve stimulation (VNS) could be of interest in the management of patients with SARS-CoV-2 infection. Indeed, through the wide innervation of the organism by the vagus nerve, especially the lungs and gastrointestinal tract, VNS appears as a serious candidate for a few side effect treatment that could dampen or prevent the cytokine storm observed in COVID-19 patients with severe symptoms. Finally, a continuous vagal tone monitoring in patients with COVID-19 could be used as a predictive marker of COVID-19 illness course but also as a predictive marker of response to COVID-19 treatment such as VNS or others.

Abstract 185: Collagen VI Alpha-3 Expression Is Higher In The Superficial Epigastric Adipose Tissue Of Former Smokers Than In Non-smokers

Purpose: Collagen VI (Col6) is an extracellular matrix (ECM) protein that plays a role in adipose tissue architecture by providing a framework for cellular connections to the ECM. Its overexpression in adipose tissue is associated with metabolic derangements such as insulin resistance and tissue fibrosis. The alpha-3 subunit of Collagen VI (Col6a3) has specifically been implicated in insulin resistance and inflammatory pathways via a cleaved mediator known as endotrophin. While previous clinical literature has shown that former smokers have an increased risk of wound complications following major surgery (including surgical site infection, disruption, delayed healing, and hernia, among others), the molecular basis for this is unclear. Moreover, the Col6a3 subunit has not been studied in this setting. Our study sought to examine the expression of Col6a3 among adipose tissue collected from obese and overweight patients following massive weight loss, and to characterize differences among patients based on preoperative comorbidities. Methods: Patients undergoing abdominoplasty following massive weight loss were consented for tissue donation for laboratory examination. The abdominal tissue samples were sampled in three distinct anatomic locations, the epigastric, periumbilical and hypogastric regions. Samples were collected both deep and superficial to Scarpa’s fascia in each location. mRNA was extracted from each sample for RT-qPCR. Results of the qPCR assay were normalized locally to the reference gene beta-actin, and globally to the same periumbilical depot per patient. Gene expression levels were calculated using the 2-delta-delta fold-change method. Average gene expression levels were compared among different cohorts based on comorbidities. Results: To date, eleven (11) patients have undergone abdominoplasty and were included in the study. All patients were female. Average BMI at the time of abdominoplasty was 30.98. There were five (5) former smokers, six (6) non-smokers, and zero (0) current smokers. Average time since cessation of smoking was 29 months prior to abdominoplasty, with an average of 13.8 pack-years smoked. There was significantly higher Col6a3 gene expression in the epigastric region superficial to Scarpa’s fascia among former smokers when compared to the same depot in non-smokers (2.1-fold higher expression than the reference depot versus 0.1-fold, p = 0.02). There was no difference in mean adipocyte size between former smokers and non-smokers in this depot (p=0.97). Conclusion: Col6a3 gene expression in the superficial epigastrium is higher among former smokers than in non-smokers. This finding suggests another possible role of Col6a3, as a mediator of tissue fibrosis in former smokers. This, along with its association with insulin resistance, may implicate Col6a3 as the molecular culprit in wound complications among obese patients who are former smokers. Long-term follow up of this cohort is warranted for evaluation of wound complications.

Chicken cognin interactome and the memory of filial imprinting.

Visual imprinting is a learning process whereby young animals come to prefer a visual stimulus after exposure to it (training). The intermediate medial mesopallium in the domestic chick forebrain is critical for visual imprinting and contributes to molecular regulation of memory formation. Criteria used to infer that a change following training is learning-related have been formulated and published. Cognin (protein disulphide isomerase) is one of several identified plasma membrane and mitochondrial proteins that are upregulated in a learning-related way 24 hours after training. Since virtually nothing is known about the cognin interactome, we have used immunoaffinity chromatography and mass spectrometry to identify proteins that interact with cognin in the cytoplasmic and plasma membrane-mitochondrial fractions. As the learning-related upregulation of cognin has been shown to occur in the plasma membrane-mitochondrial fraction and not in the cytoplasmic fraction, we studied the effect of training on three cognin-interacting partners in the plasma membrane-mitochondrial fraction: the b5 subunit of mitochondrial ATP synthase and the alpha-2 and alpha-3 subunits of sodium-potassium ATPase. Learning-related upregulation was found in the left intermediate medial mesopallium 24 hours after training for the b5 subunit of mitochondrial ATP synthase and the alpha-2 subunit of sodium-potassium ATPase. The hemispheric asymmetry revealed here is consistent with the predominance of many other learning-related effects in the left intermediate medial mesopallium. The alpha-2 subunit of sodium-potassium ATPase is mainly expressed in astrocytes, supporting a role for these glial cells in memory.

The nicotinic receptor alpha5 coding polymorphism rs16969968 as a major target in disease: Functional dissection and remaining challenges.

Nicotinic acetylcholine receptors (nAChRs) are major signalling molecules in the central and peripheral nervous system. Over the last decade, they have been linked to a number of major human psychiatric and neurological conditions, like smoking, schizophrenia, Alzheimer's disease and many others. Human Genome-Wide Association Studies (GWAS) have robustly identified genetic alterations at a locus of chromosome 15q to several of these diseases. In this review, we discuss a major coding polymorphism in the alpha5 subunit, referred to as α5SNP, and its functional dissection in vitro and in vivo. Its presence at high frequency in many human populations lends itself to pharmaceutical intervention in the context of 'positive allosteric modulators' (PAMs). We will present the prospects of this novel treatment, and the remaining challenges to identify suitable molecules.

Alternating Hemiplegia of Childhood: Understanding the Genotype-Phenotype Relationship of ATP1A3 Variations.

Alternating hemiplegia of childhood (AHC) is a rare neurological disorder affecting children with an onset before 18 months. Diagnostic clues include transient episodes of hemiplegia alternating in the laterality or quadriparesis, nystagmus and other paroxysmal attacks as tonic and dystonic spells. Epilepsy is also a common feature. In the past, a great effort has been done to understand the genetic basis of the disease leading to the discovery of mutations in the ATP1A3 gene encoding for the alpha3 subunit of Na+/K+ATPase, a protein already related to another disease named Rapid Onset Dystonia Parkinsonism (RDP). ATP1A3 mutations account for more than 70% of cases of AHC. In particular, three hotspot mutations account for about 60% of all cases, and these data have been confirmed in large population studies. Specifically, the p.Asp801Asn variant has been found to cause 30–43% of all cases, p.Glu815Lys is responsible for 16–35% of cases and p.Gly947Arg accounts for 8–15%. These three mutations are associated with different clinical phenotype in terms of symptoms, severity and prognosis. In vitro and in vivo models reveal that a crucial role of Na+/K+ATPase pump activity emerges in maintaining a correct membrane potential, survival and homeostasis of neurons. Herein, we attempt to summarize all clinical, genetic and molecular aspects of AHC considering ATP1A3 as its primary disease-causing determinant.

Influence of the integrin alpha-1 subunit and its relationship with high-fat diet upon extracellular matrix synthesis in skeletal muscle and tendon.

Integrins are important for mechanosensation in tissue and play, together with nutrition, a role in regulating extracellular matrix (ECM) in skeletal muscle and tendon. Integrin receptors are dimers that consist of an α and β subunit and bridge extracellular and intracellular signals. The present study investigates whether the deletion of the integrin receptor α1 subunit influences collagen and other matrix proteins in the musculotendinous tissue and whether it causes any compensatory changes in other integrin subunits in C57BL/6J mice. In addition, we study whether a high-fat diet (HFD) influences these responses in muscle or tendon. Mice on a HFD had a higher number of non-enzymatic cross-links in skeletal muscle ECM and increased gene expression of collagen and other extracellular matrix proteins. In contrast to gene expression, total collagen protein content was decreased by HFD in the muscle with no change in tendon. Integrin α1 subunit knockout resulted in a decrease of collagen type I and III, TGF-β1 and IGF-1 gene expression in muscle of HFD mice but did not affect total collagen protein compared with wild-type (WT) littermates in either muscle or tendon. There was no compensatory increase in the genes that express other integrin subunits. In conclusion, HFD induced a significant increase in expression of ECM genes in muscle. On the protein level, HFD resulted in a lower collagen content in muscle. Tendons were unaffected by the diet. Deletion of the integrin α1 subunit did not affect collagen protein or gene expression in muscle or tendon.

Mechanism of tanshinones and phenolic acids from Danshen in the treatment of coronary heart disease based on co-expression network

BackgroundThe tanshinones and phenolic acids in Salvia miltiorrhiza (also named Danshen) have been confirmed for the treatment of coronary heart disease (CHD), but the action mechanisms remain elusive.MethodsIn the current study, the co-expression protein interaction network (Ce-PIN) was used to illustrate the differences between the tanshinones and phenolic acids of Danshen in the treatment of CHD. By integrating the gene expression profile data and protein-protein interactions (PPIs) data, the Ce-PINs of tanshinones and phenolic acids were constructed. Then, the Ce-PINs were analyzed by gene ontology enrichment analyzed based on the optimal algorithm.ResultsIt turned out that Danshen is able to treat CHD by regulating the blood circulation, immune response and lipid metabolism. However, phenolic acids may regulate the blood circulation by Extracellular calcium-sensing receptor (CaSR), Endothelin-1 receptor (EDNRA), Endothelin-1 receptor (EDNRB), Kininogen-1 (KNG1), tanshinones may regulate the blood circulation by Guanylate cyclase soluble subunit alpha-1 (GUCY1A3) and Guanylate cyclase soluble subunit beta-1 (GUCY1B3). In addition, both the phenolic acids and tanshinones may regulate the immune response or inflammation by T-cell surface glycoprotein CD4 (CD4), Receptor-type tyrosine-protein phosphatase C (PTPRC).ConclusionThrough the same targets of the same biological process and different targets of the same biological process, the tanshinones and phenolic acids synergistically treat coronary heart disease.

CaV1.2 and CaV1.3 voltage-gated L-type Ca2+ channels in rat white fat adipocytes.

L-type channel antagonists are of therapeutic benefit in the treatment of hyperlipidaemia and insulin resistance. Our aim was to identify L-type voltage-gated Ca2+ channels in white fat adipocytes, and determine if they affect intracellular Ca2+, lipolysis and lipogenesis. We used a multidisciplinary approach of molecular biology, confocal microscopy, Ca2+ imaging and metabolic assays to explore this problem using adipocytes isolated from adult rat epididymal fat pads. CaV1.2, CaV1.3 and CaV1.1 alpha1, beta and alpha2delta subunits were detected at the gene expression level. The CaV1.2 and CaV1.3 alpha1 subunits were identified in the plasma membrane at the protein level. Confocal microscopy with fluorescent antibodies labelled CaV1.2 in the plasma membrane. Ca2+ imaging revealed that the intracellular Ca2+ concentration, [Ca2 +]i was reversibly decreased by removal of extracellular Ca2+, an effect mimicked by verapamil, nifedipine and Co2+, all blockers of L-type channels, whereas the Ca2+ channel agonist BAY-K8644 increased [Ca2+]i. The finding that the magnitude of these effects correlated with basal [Ca2+]i suggests that adipocyte [Ca2+]i is controlled by L-type Ca2+ channels that are constitutively active at the adipocyte depolarized membrane potential. Pharmacological manipulation of L-type channel activity modulated both basal and catecholamine-stimulated lipolysis but not insulin-induced glucose uptake or lipogenesis. We conclude that white adipocytes have constitutively active L-type Ca2+ channels which explains their sensitivity of lipolysis to Ca2+ channel modulators. Our data suggest CaV1.2 as a potential novel therapeutic target in the treatment of obesity.

Nicotine Suppresses the Invasiveness of Human Trophoblasts by Downregulation of CXCL12 Expression through the Alpha-7 Subunit of the Nicotinic Acetylcholine Receptor

Smoke exposure during pregnancy has detrimental effects upon numerous fetal and neonatal outcomes. Nicotine (the main component of tobacco) has been suggested to affect placental development. During placental development, efficient invasion by trophoblasts is required for establishment of the fetus–maternal circulation. In this study we explored the regulation of trophoblast invasion by nicotine. An immortalized first trimester extravillous trophoblast cell line (HTR-8/SVneo cells) was used for all the experiments, which were treated by nicotine, methyllycaconitine, and C-X-C motif chemokine ligand 12 (CXCL12). Total RNA and protein were used to study the expressions of nicotinic acetylcholine receptors (nAChRs), and transwell assay was used to study invasiveness. Changes of RNA expression due to nicotine treatment were detected by RNA sequence. Level of CXCL12 mRNA was verified by quantitative PCR. We showed that HTR-8/SVneo expressed subunits α2–4, α7, α9, β1, and β2 of nAChRs. Nicotine downregulated CXCL12 expression and inhibited trophoblast invasion. Methyllycaconitine, as an antagonist of the α7 homopolymer, blocked the inhibitory effect of nicotine. CXCL12 could rescue the nicotine-induced inhibitory effect on invasion of HTR-8/SVneo cells. These results suggest that the α7 subunit of the nAChR has important roles in modulating trophoblast invasion through CXCL12.