Subunits Alpha1 Research Articles

Lentiviral expression of wild-type LAMA3A restores cell adhesion in airway basal cells from children with epidermolysis bullosa

The hallmark of epidermolysis bullosa (EB) is fragile attachment of epithelia due to genetic variants in cell adhesion genes. We describe 16 EB patients treated in the Ear, Nose and Throat department of a tertiary pediatric hospital linked to the United Kingdom’s National EB unit between 1992 and 2023. Patients suffered a high degree of morbidity and mortality from laryngotracheal stenosis. Variants in laminin subunit alpha-3 (LAMA3) were found in 10/15 patients where genotype was available. LAMA3 encodes a subunit of the laminin-332 heterotrimeric extracellular matrix protein complex and is expressed by airway epithelial basal stem cells. We investigated the benefit of restoring wildtype LAMA3 expression in primary EB patient-derived basal cell cultures. EB basal cells demonstrated weak adhesion to cell culture substrates, but could otherwise be expanded similarly to non-EB basal cells. In vitro lentiviral overexpression of LAMA3A in EB basal cells enabled them to differentiate in air-liquid interface cultures, producing cilia with normal ciliary beat frequency. Moreover, transduction restored cell adhesion to levels comparable to a non-EB donor culture. These data provide proof-of-concept for a combined cell and gene therapy approach to treat airway disease in LAMA3-affected EB.

Childhood-related neural genotype-phenotype in ATP1A3 mutations: comprehensive analysis.

ATP1A3 is a gene that encodes the ATPase Na + /K + transporting subunit alpha-3 isoenzyme that is widely expressed in GABAergic neurons. It maintains metabolic balance and neurotransmitter movement. These pathways are essential for the proper functioning of the nervous system. A mutation in the ATP1A3 gene demonstrates remarkable genotype-phenotype heterogeneity. To provide insight into patients with ATP1A3 mutation. These cases were identified using next generation sequencing. The patients' clinical and genetic data were retrieved. Detailed revision of the literature was conducted to illustrate and compare findings. The clinical, genetical, neuroimaging, and electrophysiological data of all pediatric patients were extracted. The study included 14 females and 12 males in addition to two novel females cases. Their mean current age is 6.3 ± 4.24years. There were 11.54% preterm pregnancies with 5 cases reporting pregnancy complications. Mean age of seizure onset was 1.07 ± 1.06years. Seizure semiology included generalized tonic-clonic, staring spells, tonic-clonic, and others. Levetiracetam was the most frequently used Anti-seizure medication. The three most frequently reported classical symptoms included alternating hemiplegia of childhood (50%), cerebellar ataxia (50%), and optic atrophy (23.08%). Non-classical symptoms included dystonia (73.08%), paroxysmal dyskinesias (34.62%), and encephalopathy (26.92%). Developmental delay was reported among 84.62% in cognitive, 92.31% in sensorimotor, 80.77% in speech, and 76.92% in socioemotional. EEG and MRI were non-specific. Our study demonstrated high heterogeneity among patients with pathogenic variants in the ATP1A3 gene. Such variation is multifactorial and can be a predisposition of wide genetic and clinical variables. Many patients shared few similarities in their genetic map including repeatedly reported de novo, heterozygous, mutations in the gene. Clinically, higher females prevalence of atypical presentation was noted. These findings are validated with prior evidence and the comprehensive analysis in this study.

Trim46 knockout impaired neuronal architecture and caused hypoactive behavior in rats.

Tripartite motif (TRIM46) is a relatively novel protein that belongs to tripartite motif family. TRIM46 organizes parallel microtubule arrays on the axons, which are important for neuronal polarity and axonal function. TRIM46 is highly expressed in the brain, but its biological function in adults has not yet been determined. Trim46 knockout (KO) rat line was established using CRISPR/cas9. Trim46 KO rats had smaller hippocampus sizes, fewer neuronal dendritic arbors and dendritic spines, and shorter and more distant axon initial segment. Furthermore, the protein interaction between endogenous TRIM46 and FK506 binding protein 5 (FKBP5) in brain tissues was determined; Trim46 KO increased hippocampal FKBP5 protein levels and decreased hippocampal protein kinase B (Akt) phosphorylation, gamma-aminobutyric acid type A receptor subunit alpha1 (GABRA1) and glutamate ionotropic receptor NMDA type subunit 1 (NMDAR1) protein levels. Trim46 KO rats exhibited hypoactive behavioral changes such as reduced spontaneous activity, social interaction, sucrose preference, impaired prepulse inhibition (PPI), and short-term reference memory. These results demonstrate the significant impact of Trim46 KO on brain structure and behavioral function. This study revealed a novel potential association of TRIM46 with dendritic development and neuropsychiatric behavior, providing new insights into the role of TRIM46 in the brain.

CHRNA9 as a New Prognostic Marker and Potential Therapeutic Target in Glioma.

Background: The nicotinic acetylcholine receptor (nAChR) subunit alpha-9 (CHRNA9) is a unique cholinergic receptor, which is involved in tumor proliferation, apoptosis, metastasis and chemotherapy resistance. However, the correlation between the expression level of CHRNA9 in glioma and the clinical features and prognosis of glioma patients has not been clarified. The aim of this study was to verify the expression level of CHRNA9 in glioma and its effect on prognosis by bioinformatics methods. Methods: The RNA-seq data of glioma and normal samples were obtained from the TCGA and GTEx databases. Bioinformatics methods were utilized to analyze the differential expression of CHRNA9 between tumor samples and normal samples. The potential association between CHRNA9 and the clinicopathological features of glioma patients was also investigated. The Kaplan-Meier method and Cox regression were utilized to analyze the relationship between CHRNA9 expression level and survival time and prognostic value of glioma patients. Enrichment analysis was applied to predict gene function and signaling pathways associated with CHRNA9. Experimental verification was performed using tumor tissues and paracancerous tissues from glioma patients. Results: The results of bioinformatics analysis showed that the expression of CHRNA9 was increased in glioma tissues, correlating with poor prognosis and reduced patient survival time. Enrichment analysis suggested that CHRNA9 may interact with the JAK/STAT pathway. CHRNA9 was also found to be abnormally expressed in various other tumors and associated with the expression levels of numerous immune checkpoints in glioma. The findings from the analysis of clinical samples revealed that the expression levels of both mRNA and protein of CHRNA9 in glioma tissues were higher than those in paracancerous tissues. Similarly, the mRNA expression levels of STAT3, IL-6, and TNF-α, which are crucial factors in the STAT3 pathway, were elevated in glioma tissues compared to paracancerous tissues. Conclusion: CHRNA9 is a potential prognostic marker and immunotherapy target for glioma, with its mechanism of action potentially linked to the STAT3 pathway.

Mutational analysis of cytoplasmic domain of integrin subunit alpha-1 and its association with periapical wound healing after surgical endodontic treatment.

Numerous studies reported that the healing after surgical endodontic retreatment is influenced by multiple factors which include the genetic profile of the patient, epigenetics, and immune responses. The genes which are primarily responsible for the healing potential in different individuals are those which are involved in the regulation of the cytoskeleton and cellular adhesion which subsequently affects bone deposition and healing. Integrins are cell-surface molecules, possess a key role in the cytoskeleton and cellular adhesion. Integrin Subunit Alpha 1 (ITGA1) is one among the integrin family and helps in regulating the Epidermal Growth Factor receptor (EGFR) pathway, consequently affects proliferation and healing. The objectives of the study were to identify mutations in the cytoplasmic domain of Integrin Subunit Alpha 1 (ITGA1), to assess the expression of activated EGFR, EGFRPhospho and TC-PTP in the periapical wound and to correlate these mutations and expression patterns with periapical wound healing. Thirty-seven patients between ages 18-60 years reported chronic apical periodontitis of single-rooted anterior teeth with periapical radiolucency, equal or greater than 4 mm or periapical lesion in an open apex of single-rooted teeth due to trauma were included in the study from 01st June 2018 till 31st October 2019. Patients with persistent radiolucency after primary root canal treatment and endodontic retreatment were kept on follow-up for 3-4 months surgical endodontic treatment was performed in cases with persistent periapical lesions of 4mm or more in diameter. Periapical lesion sample was collected and used for (1) histo-pathological analysis after Hematoxylin & Eosin staining, (2) total DNA extraction for ITGA1 cytoplasmic domain mutational analysis and immunohistochemistry for EGFR and TCPTP. A positive correlation was observed between the expression levels of EGFRPhospho and the healing of periapical lesions. Moreover, a negative weak correlation was observed between the expression levels of EGFR and TCPTP and the healing of periapical lesions. Out of nine sequences of cytoplasmic domain of ITGA1 which were analyzed, none of them was detected with SNP. Higher expression levels of EGFRPhospho and lower expression levels of EGFR and TCPTP were associated with patients with good healing potential in periapical area. However, immunohistochemistry scores were statistically insignificant to draw any conclusion.

Size exclusion chromatography based proteomic and degradomic profiling of inflammasome-activated, murine bone marrow-derived dendritic cells highlights complex retention and release of cleavage products.

Coupling size exclusion chromatography (SEC) with mass spectrometry-based proteomics enables investigating protein complexes, with degradomic profiling providing deeper insights into complex-associated proteolytic processing and retaining of cleavage products. This study aims to map protein complex formation upon inflammasome activation in bone marrow-derived dendritic cells (BMDCs) from gasdermin D-deficient mice, focusing on proteolytic enzymes and truncated proteins in higher molecular weight complexes. Cultured BMDCs were primed with LPS and subsequently treated with nigericin or Val-boroPro (VbP). SEC-fractionated proteins were TMT-labelled and analyzed via liquid chromatography-tandem mass spectrometry (LC-MS/MS). We identified 6862 proteins and 70 802 peptides, including 14 714 semi-tryptic peptides indicating elevated endogenous proteolytic processing. The sequence motif of numerous cleavage sites maps to caspase-like activity. Inflammasome activation was corroborated by elevated levels of apoptosis-associated speck-like protein containing a CARD (ASC) in higher molecular weight (MW) fractions and increased IL-1β levels in low MW fractions upon nigericin or VbP treatment. The majority of truncated cleavage products remained within their corresponding, higher MW protein complexes while caspase-specific cleavage products of Rho-associated protein kinase 1, gelsolin, and AP-2 complex subunit alpha-2 dissociated to lower MW fractions. SEC profiles identified 174 proteases, with cell surface proteases forming high MW complexes, including ADAMs and DPP4 but not MMP14. VbP treatment led to the accumulation of ISG15 in low MW fractions while RNA polymerase II coactivator p15 shifted to higher MW fractions. This study demonstrates that SEC-coupled proteomics and degradomic profiling offer unique insights into protein complex dynamics and proteolytic processes upon inflammasome activation.

Potential application of bisoprolol derivative compounds as antihypertensive drugs: synthesis and in silico study.

Two bisoprolol derivatives, N-acetyl bisoprolol and N-formyl bisoprolol, belonging to the beta-blocker class of antihypertensive drugs, were synthesized using acetylation and formylation reactions. The yields of the reactions were determined to be 32.40% for N-acetyl bisoprolol and 20.20% for N-formyl bisoprolol. In silico methods such as molecular docking, molecular dynamics simulation and SwissADME prediction were employed to evaluate the potential of these bisoprolol derivatives as antihypertensive drugs. These methods were used to assess the interaction between the bisoprolol derivatives and various receptors associated with hypertension, including human angiotensin I-converting enzyme (PDB ID: 1O8A), renin (PDB ID: 2V0Z), beta-1 adrenergic receptors (PDB ID: 4BVN, 7BVQ), voltage-dependent L-type calcium channel subunit alpha-1S (PDB ID: 6JP5) and mineralocorticoid receptor (PDB ID: 6L88). Our results demonstrated the highest binding energy when bisoprolol and its derivatives bound to 4BVN, with binding energy values of 6.74 kcal mol-1, 7.03 kcal mol-1 and 7.63 kcal mol-1 for bisoprolol, N-acetyl bisoprolol and N-formyl bisoprolol, respectively. The stability of these complexes was confirmed by molecular dynamics simulations, with a root-mean-square deviation value of approximately 2. Furthermore, the SwissADME results indicated that both derivatives exhibited similar properties to the reference drug bisoprolol.

Synaptic Involvement of the Human Amygdala in Parkinson’s Disease

α-Synuclein, a protein mostly present in presynaptic terminals, accumulates neuropathologically in Parkinson’s disease in a 6-stage sequence and propagates in the nervous system in a prion-like manner through neurons and glia. In stage 3, the substantia nigra are affected, provoking motor symptoms and the amygdaloid complex, leading to different nonmotor symptoms; from here, synucleinopathy spreads to the temporal cortex and beyond. The expected increase in Parkinson's disease incidence accelerates the need for detection biomarkers; however, the heterogeneity of this disease, including pathological aggregates and pathophysiological pathways, poses a challenge in the search for new therapeutic targets and biomarkers. Proteomic analyses are lacking, and the literature regarding synucleinopathy, neural and glial involvement, and volume of the human amygdaloid complex is controversial. Therefore, the present study combines both proteomic and stereological probes. Data-independent acquisition-parallel accumulation of serial fragmentation proteomic analysis revealed a remarkable proteomic impact, especially at the synaptic level in the human amygdaloid complex in Parkinson’s disease. Among the 199 differentially expressed proteins, guanine nucleotide-binding protein G(i) subunit alpha-1 (GNAI1), elongation factor 1-alpha 1 (EEF1A1), myelin proteolipid protein (PLP1), neuroplastin (NPTN), 14-3-3 protein eta (YWHAH), gene associated with retinoic and interferon-induced mortality 19 protein (GRIM19), and orosomucoid-2 (ORM2) stand out as potential biomarkers in Parkinson’s disease. Stereological analysis, however, did not reveal alterations regarding synucleinopathy, neural or glial populations, or volume changes. To our knowledge, this is the first proteomic study of the human amygdaloid complex in Parkinson’s disease, and it identified possible biomarkers of the disease. Lewy pathology could not be sufficient to cause neurodegeneration or alteration of microglial and astroglial populations in the human amygdaloid complex in Parkinson’s disease. Nevertheless, damage at the proteomic level is manifest, showing up significant synaptic involvement.

Effects of Remimazolam on Intracellular Calcium Dynamics in Myotubes Derived from Patients with Malignant Hyperthermia and Functional Analysis of Type 1 Ryanodine Receptor Gene Variants.

Remimazolam is a novel general anesthetic and its safety in patients with malignant hyperthermia (MH) is unknown. We used myotubes derived from the skeletal muscle of patients with MH to examine the response to ryanodine receptor 1 (RYR1) agonist and remimazolam in MH-susceptible patients. Patients underwent muscle biopsy for the Ca2+-induced Ca2+ release (CICR) rate test, a diagnostic tool for MH in Japan. Ten patients had myotubes obtained from skeletal muscle cultures, and the genes associated with malignant hyperthermia in these patients were analyzed. The EC50 of caffeine, cresol, and remimazolam to induce intracellular calcium concentration change were compared between myotubes from CICR-negative genetic test patients and myotubes from other patients. Eight of the ten were CICR-positive, five of whom had RYR1 causative gene mutations or variants. Two patients had CICR-negative genetic tests, and as expected had the highest EC50 (the concentration of a drug that gives a half-maximal response) in response to caffeine, 4CmC and remimazolam. Three patients had a positive CICR but no known variants in RYR1 or CACNA1S (voltage-gated calcium channel subunit alpha1S). Myotubes in these patients had significantly lower EC50s for all agents than myotubes in CICR-negative patients. When myotubes from a patient who was CICR-negative and had no gene variant were used as a control, myotubes from CICR-positive patients were more hyper-responsive than controls to all stimulants used. The EC50 for remimazolam was lowest for myotubes from CICR-positive, RYR1-mutant patients, at 206 µM (corresponding to 123 µg/mL). The concentration was more than 80-times higher than the clinical concentration. RYR1 gene variants in R4645Q and W5020G were shown to be causative gene mutations for MH. Intracellular calcium in myotubes from MH patients are elevated at high concentrations of remimazolam but not at clinically used concentrations of remimazolam. Remimazolam appears to be safe to use in patients with MH.

An Overview of Schizotypal Personality Disorder: Etiology Treatment

The article summarizes and analyzes some of the previous studies on schizotypal personality disorder to explain the etiology, genetic aspect such as Catechol-O-methyltransferase (COMT which is situated on chromosome 22q11) and calcium voltage-gated channel subunit alpha1 C (CACNA1C rs1006737); environmental aspect such as risky factors affecting an unnormal brain development like and psychological trauma, and chronic stress, and the treatment (Metacognitive treatment; RuminationFocused Cognitive Behavioral Therapy; Medication treatment) of schizotypal personality disorder are also discussed. Both methods, Metacognitive treatment, and RuminationFocused Cognitive Behavioral Therapy, in their respective cases, improved symptoms in schizotypal personality disorder patients. However, although research on medication treatment shows that although some medicines, like Risperidone and Haloperidol, can also improve schizotypal personality disorder symptoms, they are also accompanied by the side effects. Moreover, the analysis shows that the present research on managing schizotypal personality disorder encounters issues such as insufficient sample size and absence of repeated experiments. These results of prior studies still need further verification before they can be used in practice.

Mechanism of Lingbao Huxin Dan in the treatment of bradyarrhythmia complicated with coronary heart disease: a network pharmacology analysis.

To investigate the mechanism of action of the Lingbao Huxin Dan in treating bradycardia arrhythmia with coronary heart disease (BA-CHD) by network pharmacology. The active ingredients of the Lingbao Huxin Dan were screened on the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform and Bioinformatics tools designed for the analysis of molecular mechanisms of Chinese medicine platform; target prediction was conducted with the SwissTargetPrediction database, and Cytoscape 3.8 was used to construct a drug ingredient-target network. The Genecards, Online Mendelian Inheritance in Man, and DrugBank databases were searched for disease targets. Venn plots were used to display the common targets of BA-CHD and active ingredients. The STRING platform was used to construct a protein-protein interaction network. The Metascape data platform was used for Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis to construct a signaling pathway network of the active ingredients of the Lingbao Huxin Dan. There were 121 active ingredients, 899 related targets, 39 targets important in BA-CHD and 14 targets which intersected between the active ingredients and BA-CHD. There were 27 core therapeutic ingredients, 153 biological processes, 18 cell ingredients and 20 molecular functions obtained by GO enrichment analysis. The KEGG pathway analysis yielded 19 signaling pathways. RBA-CHD may treat BA-CHD by regulating adrenergic receptor beta-1, alpha 1-α adrenergic receptor, calcium voltage-gated channel subunit alpha1 C, alpha-1β-adrenergic receptor, nitric oxide synthase 2, beta-2 adrenergic receptor, voltage-dependent calcium channel subunit alpha-2/delta-1, an- giotensin-converting enzyme, Raf-1 proto-oncogene serine/threonine-protein kinase, and other targets, potentially by affecting adrenergic receptor binding and calcium channel opening, to regulate the activity of cardiomyocytes.

Prohibitin1 facilitates viral replication by impairing the RIG-I-like receptor signaling pathway.

Type I interferon (IFN-I), produced by the innate immune system, plays an essential role in host antiviral responses. Proper regulation of IFN-I production is required for the host to balance immune responses and prevent superfluous inflammation. IFN regulatory factor 3 (IRF3) and subsequent sensors are activated by RNA virus infection to induce IFN-I production. Therefore, proper regulation of IRF3 serves as an important way to control innate immunity and viral replication. Here, we first identified Prohibitin1 (PHB1) as a negative regulator of host IFN-I innate immune responses. Mechanistically, PHB1 inhibited the nucleus import of IRF3 by impairing its binding with importin subunit alpha-1 and importin subunit alpha-5. Our study demonstrates the mechanism by which PHB1 facilitates the replication of multiple RNA viruses and provides insights into the negative regulation of host immune responses.

Generation of eight hiPSCs lines from two pathogenic variants in CACNA1A using the CRISPR-Cas9 gene editing technology

Developmental and epileptic encephalopathies (DEEs) are rare severe neurodevelopmental disorders with a cumulative incidence of 1:6.000 live births. Many epileptic conditions arise from single nucleotide variants in CACNA1A (calcium voltage-gated channel subunit alpha1 A), encoding the CaV2.1 calcium channel subunit. Human induced pluripotent stem cells (hiPSCs) are an optimal choice for modeling DEEs, as they can be differentiated in vitro into diverse neuronal subpopulations. Here, we report the generation of hiPSC lines with two pathogenic CACNA1A variants c.1767C > T, p. (Arg589Cys), referred to as R589C and c. 2139G > A, p.(Ala713Thr), referred to as A713T, previously associated with epilepsy. The variants were introduced into a hiPSC line from a healthy individual via CRISPR-Cas9 gene editing technology.

PAI1 inhibits the pathogenesis of primary focal hyperhidrosis by targeting CHRNA1

BackgroundPrimary focal hyperhidrosis (PFH) may be attributed to the up-regulation of the cholinergic receptor nicotinic alpha 1 subunit (CHRNA1) in eccrine glands. Plasminogen activator inhibitor-1 (PAI1, encoded by SERPINE1) is reported to inhibit the expression of CHRNA1, while the role of PAI1 in hyperhidrosis is unknown.MethodsSerpine1 KO mice, Serpine1-Tg mice, and wild type BALB/c mice were intraperitoneally injected with pilocarpine hydrochloride to induce PFH. Cisatracurium (CIS, antagonist of CHRNA1) or PAI-039 (small-molecule inhibitor of PAI1) was pre-administrated before the induction of hyperhidrosis. On the other hand, Chrna1-expressing AAV was constructed and administered to Serpine1-Tg mice with hydrochloride stimulation. Hydrochloride-related biomarkers, such as acetylcholine (ACH) in the serum, calcium voltage-gated channel subunit alpha1 C (CACNA1C), and aquaporin 5 (AQP5) in sweat glands of mice were assayed with ELISA, RT-PCR, and Western blot.ResultsThe administration of PAI-039 or Pai1 knock-out increased Chrna1 expression, sweat secretion, and hydrochloride-related biomarkers (ACH, CACNA1C, and AQP5) expression. On the other hand, CIS administration diminished the strengthened hyperhidrosis phenotype induced by Pai1 knock-out with decreased sweat gland secretion.ConclusionPAI1 inhibits CHRNA1-mediated hydrochloride-induced hyperhidrosis, with decreased sweat gland secretion and diminished ACH, AQP5, and CACNA1C expression. These results indicate the potential to utilize PAI1 to alleviate PFH.

Familial hemiplegic migraine in pediatric patients: A genetic, clinical, and follow-up study.

The aim of this study was to describe a cohort of pediatric patients with genetically confirmed familial hemiplegic migraine (FHM). The knowledge of genotype-phenotype correlations may suggest prognostic factors associated with severe phenotypes. Hemiplegic migraine is a rare disease and data concerning the pediatric population are even more rare as they are often extrapolated from mixed cohorts. We selected patients who met International Classification of Headache Disorders, third edition criteria for FHM, who had a molecular diagnosis, and whose first attack occurred under the age of 18 years. We enrolled nine patients (seven males and two females) first referred to our three centers. Three of the nine (33%) patients had calcium voltage-gated channel subunit alpha1 A (CACNA1A) mutations, five (55%) had ATPase Na+/K+ transporting subunit alpha 2 (ATP1A2) mutations, and one had both genetic mutations. The patients experienced at least one aura feature other than hemiplegia during the first attack. The mean (SD) duration of HM attacks in the sample was 11.3 (17.1) h; 3.8 (6.1) h in the ATP1A2 group, and 24.3 (23.5) h in the CACNA1A group. The mean (SD, range) duration of follow-up was 7.4 (2.2, 3-10) years. During the first year from the disorder's onset, only four patients had additional attacks. Over the course of follow-up, the attack frequency overall was 0.4 attacks/year without a difference between the two groups (CACNA1A and ATP1A2). The study data show that most of our patients with early-onset FHM experienced infrequent and non-severe attacks, which improved over time. Furthermore, the clinical course revealed neither the appearance of novel neurological disorders or a deterioration of basic neurological or cognitive functioning.

Anti-Cancer Roles of Probiotic-Derived P8 Protein in Colorectal Cancer Cell Line DLD-1.

A novel probiotics-derived protein, P8, suppresses the growth of colorectal cancer (CRC). P8 can penetrate the cell membrane via endocytosis and cause cell cycle arrest in DLD-1 cells through down-regulation of CDK1/Cyclin B1. However, neither the protein involved in the endocytosis of P8 nor the cell cycle arrest targets of P8 are known. We identified two P8-interacting target proteins [importin subunit alpha-4 (KPNA3) and glycogen synthase kinase-3 beta (GSK3β)] using P8 as a bait in pull-down assays of DLD-1 cell lysates. Endocytosed P8 in the cytosol was found to bind specifically to GSK3β, preventing its inactivation by protein kinases AKT/CK1ε/PKA. The subsequent activation of GSK3β led to strong phosphorylation (S33,37/T41) of β-catenin, resulting in its subsequent degradation. P8 in the cytosol was also found to be translocated into the nucleus by KPNA3 and importin. In the nucleus, after its release, P8 binds directly to the intron regions of the GSK3β gene, leading to dysregulation of GSK3β transcription. GSK3β is a key protein kinase in Wnt signaling, which controls cell proliferation during CRC development. P8 can result in a cell cycle arrest morphology in CRC cells, even when they are in the Wnt ON signaling state.

Functional characterization of KCNMA1 mutation associated with dyskinesia, seizure, developmental delay, and cerebellar atrophy

KCNMA1 located on chromosome 10q22.3, encodes the pore-forming α subunit of the ‘Big K+’ (BK) large conductance calcium and voltage-activated K + channel. Numerous evidence suggests the functional BK channel alterations produced by different KCNMA1 alleles may associate with different symptoms, such as paroxysmal non kinesigenic dyskinesia with gain of function and ataxia with loss of function. Functional classifications revealed two major patterns, gain of function and loss of function effects on channel properties in different cell lines. In the literature, two mutations have been shown to confer gain of function properties to BK channels: D434G and N995S. In this study, we report the functional characterization of a variant which was previously reported the whole exome sequencing revealed bi-allelic nonsense variation of the cytoplasmic domain of calcium-activated potassium channel subunit alpha-1 protein. To detect functional consequences of the variation, we parallely conducted two independent approaches. One is immunostaining using and the other one is electrophysiological recording using patch-clamp on wild-type and R458X mutant cells to detect the differences between wild-type and the mutant cells. We detected the gain of function effect for the mutation (NM_001161352.1 (ENST00000286628.8):c.1372C > T;Arg458*) using two parallel approaches. According to the result we found, the reported mutation causes the loss of function in the cell. It should be noted that in future studies, it can be thought that the functions of genes associated with channelopathies may have a dual effect such as loss and gain.

Prenatal dexamethasone exposure impaired vascular reactivity in adult male offspring cerebral arteries

BackgroundCerebrovascular disease is one of the leading causes of death worldwide. Middle cerebral artery (MCA) is the largest and most complex of cerebral arteries. The prenatal period is a critical time for development, which largely determines lifelong health. Clinically, glucocorticoids (GCs) administration to accelerate preterm fetal lung maturation has become standard practice. Prenatal GCs administration increases cardiovascular risks in offspring, but little is known regarding the side effects on offspring MCA function. ObjectiveWe investigated the alterations of MCA reactivity following prenatal GCs administration in postnatal offspring. Method and resultsPregnant Sprague-Dawley rats received synthetic GCs (dexamethasone, DEX) during the last week of pregnancy, and we examined vascular reactivity, cellular electrophysiology, and gene promoter epigenetic modifications in the male offspring MCA. Our results showed that prenatal DEX exposure increased the sensitivity of offspring MCA to Angiotensin II, which was resulted from the increased Cav1.2 (L-type Ca2+ channels subunit alpha1 C). Mechanistically, prenatal DEX exposure resulted in a transcriptionally active chromatin structure at the Cav1.2 gene promoter by altering histone modifications. This activation led to increased expression of vascular Cav1.2 gene, ultimately resulting in increased MCA contractility in offspring. ConclusionThe present study is the first to demonstrate that the adverse effects of prenatal GCs administration on cerebrovascular tone persist into adulthood, providing new insights into developmental origins of cerebrovascular disease.

Imbalance of multiple neurotransmitter pathways leading to depression-like behavior and cognitive dysfunction in the triple transgenic mouse model of Alzheimer disease.

Depression is among the most frequent psychiatric comorbid conditions in Alzheimer disease (AD). However, pharmacotherapy for depressive disorders in AD is still a big challenge, and the data on the efffcacy of current antidepressants used clinically for depressive symptoms in patients with AD remain inconclusive. Here we investigated the mechanism of the interactions between depression and AD, which we believe would aid in the development of pharmacological therapeutics for the comorbidity of depression and AD. Female APP/PS1/Tau triple transgenic (3×Tg-AD) mice at 24 months of age and age- and sex-matched wild-type (WT) mice were used. The shuttle-box passive avoidance test (PAT) were implemented to assess the abilities of learning and memory, and the open field test (OFT) and the tail suspension test (TST) were used to assess depression-like behavior. High-performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was used to detect the level of neurotransmitters related to depression in the hippocampus of mice. The data was identified by orthogonal projections to latent structures discriminant analysis (OPLS-DA). Most neurotransmitters exert their effects by binding to the corresponding receptor, so the expression of relative receptors in the hippocampus of mice was detected using Western blot. Compared to WT mice, 3×Tg-AD mice displayed significant cognitive impairment in the PAT and depression-like behavior in the OFT and TST. They also showed significant decreases in the levels of L-tyrosine, norepinephrine, vanillylmandelic acid, 5-hydroxytryptamine, and acetylcholine, in contrast to significant increases in 5-hydroxyindoleacetic acid, L-histidine, L-glutamine, and L-arginine in the hippocampus. Moreover, the expression of the alpha 1a adrenergic receptor (ADRA1A), serotonin 1A receptor (5HT1A), and γ-aminobutyric acid A receptor subunit alpha-2 (GABRA2) was significantly downregulated in the hippocampus of 3×Tg-AD mice, while histamine H3 receptor (H3R) expression was significantly upregulated. In addition, the ratio of phosphorylated cAMP-response element-binding protein (pCREB) and CREB was significantly decreased in the hippocampus of 3×Tg-AD mice than WT mice. We demonstrated in the present study that aged female 3×Tg-AD mice showed depression-like behavior accompanied with cognitive dysfunction. The complex and diverse mechanism appears not only relevant to the imbalance of multiple neurotransmitter pathways, including the transmitters and receptors of the monoaminergic, GABAergic, histaminergic, and cholinergic systems, but also related to the changes in L-arginine and CREB signaling molecules.

A pilot pharmacogenetic study of calcium channel blocker treatment of bipolar mania

Common genetic variants located in calcium channel genes are important markers of genetic susceptibility for bipolar disorder (BD). Previous clinical trials with Calcium Channel Blocker (CCB) medication improved mood stability for some BD patients. We hypothesize that manic patients who carried calcium channel risk variants would differentially benefit from treatment with CCBs. In this pilot study, 50 BD patients (Chinese: 39; US: 11) who were hospitalized for manic episodes were given add-on CCB treatment. We determined genotypes for each patient. There was a significant decrease in the Young Mania Rating Scale (YMRS) after add-on medication treatment. Of note, two intronic variants of the Calcium Voltage-Gated Channel Subunit Alpha1 B (CACNA1B) were associated with treatment outcomes for manic patients: rs2739258 and rs2739260. BD rs2739258/rs2739260 AG-allele carriers had a better treatment response with add-on CCB than those carrying the AA or GG genotypes by survival analysis. Although these findings did not pass multiple testing correction, this study suggests that single-nucleotide polymorphisms (SNPs) residing in calcium channel genes could be predictors for response to add-on CCB treatment of bipolar mania patients, and that calcium channel genes may be involved in treatment responses for BD.