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Platelets are critical mediators of hemostasis and thrombosis. Platelets circulate as discs in their resting form but change shape rapidly upon activation by vascular damage and/or soluble agonists such as thrombin. Platelet shape change is driven by a dynamic remodeling of the actin cytoskeleton. Actin filaments interact with the protein myosin, which is phosphorylated on the myosin light chain (MLC) upon platelet activation. Actin-myosin interactions trigger contraction of the actin cytoskeleton, which drives platelet spreading and contractile force generation. Filamin A (FLNA) is an actin cross-linking protein that stabilizes the attachment between subcortical actin filaments and the cell membrane. In addition, FLNA binds multiple proteins and serves as a critical intracellular signaling scaffold. Here, we used platelets from mice with a megakaryocyte/platelet-specific deletion of FLNA to investigate the role of FLNA in regulating platelet shape change. Relative to controls, FLNA-null platelets exhibited defects in stress fiber formation, contractile force generation, and MLC phosphorylation in response to thrombin stimulation. Blockade of Rho kinase (ROCK) and protein kinase C (PKC) with the inhibitors Y27632 and bisindolylmaleimide (BIM), respectively, also attenuated MLC phosphorylation; our data further indicate that ROCK and PKC promote MLC phosphorylation through independent pathways. Notably, the activity of both ROCK and PKC was diminished in the FLNA-deficient platelets. We conclude that FLNA regulates thrombin-induced MLC phosphorylation and platelet contraction, in a ROCK- and PKC-dependent manner., (© 2024 The Author(s).)

Background: Periventricular nodular heterotopia (PVNH) is a neuronal migration disorder caused by the inability of neurons to move to the cortex. Patients with PVNH often experience epilepsy due to ectopic neuronal discharges. Most cases of PVNH are associated with variations in filamin A ( FLNA ), which encodes an actin-binding protein. However, the underlying pathological mechanisms remain unclear., Methods: Next-generation sequencing was performed to detect variants in the patient with PVNH, and the findings were confirmed using Sanger sequencing. Iterative threading assembly refinement was used to predict the structures of the variant proteins, and the search tool for the retrieval of interacting genes/proteins database was used to determine the interactions between FLNA and motility-related proteins. An induced pluripotent stem cell (iPSC) line was generated as a disease model by reprogramming human peripheral blood mononuclear cells. The FLNA expression in iPSCs was assessed using western blot and quantitative real-time polymerase chain reaction (qRT-PCR). Immunofluorescence analysis was performed to determine the arrangement of F-actin., Results: A novel FLNA frameshift variant (NM_001456.3: c.1466delG, p. G489Afs*9) was identified in a patient with PVNH and epilepsy. Bioinformatic analysis indicated that this variation was likely to impair FLNA function. Western blot and qRT-PCR analysis of iPSCs derived from the patient's peripheral blood mononuclear cells revealed the absence of FLNA protein and mRNA. Immunofluorescence analysis suggested an irregular arrangement and disorganization of F-actin compared to that observed in healthy donors., Conclusion: Our findings indicate that the frameshift variant of FLNA (NM_001456.3: c.1466delG, p. G489Afs*9) impairs the arrangement and organization of F-actin, potentially influencing cell migration and causing PVNH., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Xue, Wang, Peng, Feng, Guan and Hao.)

Communication in the form of nonverbal, social vocalization, or crying is evolutionary conserved in mammals and is impaired early in human infants that are later diagnosed with autism spectrum disorder (ASD). Defects in infant vocalization have been proposed as an early sign of ASD that may exacerbate ASD development. However, the neural mechanisms associated with early communicative deficits in ASD are not known. Here, we expressed a constitutively active mutant of Rheb (Rheb S16H ), which is known to upregulate two ASD core pathways, mTOR complex 1 (mTORC1) and ERK1/2, in Layer (L) 2/3 pyramidal neurons of the neocortex of mice of either sex. We found that cellular mosaic expression of Rheb S16H in L2/3 pyramidal neurons altered the production of isolation calls from neonatal mice. This was accompanied by an expected misplacement of neurons and dendrite overgrowth, along with an unexpected increase in spine density and length, which was associated with increased excitatory synaptic activity. This contrasted with the known decrease in spine density in Rheb S16H neurons of 1-month-old mice. Reducing the levels of the actin cross-linking and adaptor protein filamin A (FLNA), known to be increased downstream of ERK1/2, attenuated dendrite overgrowth and fully restored spine properties, synaptic connectivity, and the production of pup isolation calls. These findings suggest that upper-layer cortical pyramidal neurons contribute to communicative deficits in a condition known to affect two core ASD pathways and that these mechanisms are regulated by FLNA., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

The incidence of intrahepatic cholangiocarcinoma (ICC) is steadily rising, and it is associated with a high mortality rate. Clinical samples were collected to detect the expression of HSPB8 and BAG3 in ICC tissues. ICC cells were cultured and transfected with plasmids that overexpressed or silenced specific genes to investigate the impact of gene expression alterations on cell function. qPCR and Western blot techniques were utilized to measure gene and protein expression levels. A wound healing assay was conducted to assess cell migration ability. The Transwell assay was used to assess cell invasion ability. Co-IP was used to verify the binding relationship between HSPB8 and BAG3. The effects of HSPB8 and BAG3 on lung metastasis of tumors in vivo were verified by constructing a metastatic tumor model. Through the above experiments, we discovered that the expressions of HSPB8 and BAG3 were up-regulated in ICC tissues and cells, and their expressions were positively correlated. The metastatic ability of ICC cells could be promoted or inhibited by upregulating or downregulating the expression of BAG3. Furthermore, the HSPB8-BAG3 chaperone complex resulted in the abnormal degradation of Filamin A by activating autophagy. Increased expression of Filamin A inhibits the migration and invasion of ICC cells. Overexpression of HSPB8 and BAG3 in vivo promoted the lung metastasis ability of ICC cells. The HSPB8-BAG3 chaperone complex promotes ICC cell migration and invasion by regulating CASA-mediated degradation of Filamin A, offering insights for enhancing ICC therapeutic strategies.

Background: Thoracic aortic aneurysm (TAA) is associated with significant morbidity and mortality. Although individuals with family histories of TAA often undergo clinical molecular genetic testing, adults with nonsyndromic TAA are not typically evaluated for genetic causes. We sought to understand the genetic contribution of both germline and somatic mosaic variants in a cohort of adult individuals with nonsyndromic TAA at a single center., Methods and Results: One hundred eighty-one consecutive patients <60 years who presented with nonsyndromic TAA at the Massachusetts General Hospital underwent deep (>500×) targeted sequencing across 114 candidate genes associated with TAA and its related functional pathways. Samples from 354 age- and sex-matched individuals without TAA were also sequenced, with a 2:1 matching. We found significant enrichments for germline (odds ratio [OR], 2.44, P =4.6×10 -6 [95% CI, 1.67-3.58]) and also somatic mosaic variants (OR, 4.71, P =0.026 [95% CI, 1.20-18.43]) between individuals with and without TAA. Likely genetic causes were present in 24% with nonsyndromic TAA, of which 21% arose from germline variants and 3% from somatic mosaic alleles. The 3 most frequently mutated genes in our cohort were FLNA (encoding Filamin A), NOTCH3 (encoding Notch receptor 3), and FBN1 (encoding Fibrillin-1). There was increased frequency of both missense and loss of function variants in TAA individuals., Conclusions: Likely contributory dominant acting genetic variants were found in almost one quarter of nonsyndromic adults with TAA. Our findings suggest a more extensive genetic architecture to TAA than expected and that genetic testing may improve the care and clinical management of adults with nonsyndromic TAA.

Filamin A is an essential protein in the cell cytoskeleton because of its actin binding properties and unique homodimer rod-shaped structure, which organises actin into three-dimensional orthogonal networks imperative to cell motility, spreading and adhesion. Filamin A is subject to extensive posttranslational modification (PTM) which serves to co-ordinate cellular architecture and to modulate its large protein-protein interaction network which is key to the protein's role as a cellular signalling hub. Characterised PTMs include phosphorylation, irreversible cleavage, ubiquitin mediated degradation, hydroxylation and O-GlcNAcylation, with preliminary evidence of tyrosylation, carbonylation and acetylation. Each modification and its relation to filamin A function will be described here. These modifications are often aberrantly applied in a range of diseases including, but not limited to, cancer, cardiovascular disease and neurological disease and we discuss the concept of target specific PTMs with novel therapeutic modalities. In summary, our review represents a topical 'one-stop-shop' that enables understanding of filamin A function in cell homeostasis and provides insight into how a variety of modifications add an extra level of Filamin A control., (© 2024 The Author(s).)

Introduction: Microvesicles (MV) released by endothelial cells (EC) following injury or inflammation contain tissue factor (TF) and mediate communication with the underlying smooth muscle cells (SMC). Ser253-phosphorylated TF co-localizes with filamin A at the leading edge of migrating SMC. In this study, the influence of endothelial-derived TF-MV, on human coronary artery SMC (HCASMC) migration was examined., Methods and Results: MV derived from human coronary artery EC (HCAEC) expressing TF Wt accelerated HCASMC migration, but was lower with cytoplasmic domain-deleted TF. Furthermore, incubation with TF Asp253 -MV, or expression of TF Asp253 in HCASMC, reduced cell migration. Blocking TF-factor VIIa (TF-fVIIa) procoagulant/protease activity, or inhibiting PAR2 signaling on HCASMC, abolished the accelerated migration. Incubation with fVIIa alone increased HCASMC migration, but was significantly enhanced on supplementation with TF. Neither recombinant TF alone, factor Xa, nor PAR2-activating peptide (SLIGKV) influenced cell migration. In other experiments, HCASMC were transfected with peptides corresponding to the cytoplasmic domain of TF prior to stimulation with TF-fVIIa. Cell migration was suppressed only when the peptides were phosphorylated at position of Ser253. Expression of mutant forms of filamin A in HCASMC indicated that the enhancement of migration by TF but not by PDGF-BB, was dependent on the presence of repeat-24 within filamin A. Incubation of HCASMC with TF Wt -MV significantly reduced the levels of Smoothelin-B protein, and upregulated FAK expression., Discussion: In conclusion, Ser253-phosphorylated TF and fVIIa released as MV-cargo by EC, act in conjunction with PAR2 on SMC to promote migration and may be crucial for normal arterial homeostasis as well as, during development of vascular disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Featherby and Ettelaie.)

Triple-negative breast cancer is a rare but highly heterogeneous breast cancer subtype with a limited choice of specific treatments. Chemotherapy remains the only efficient treatment, but its side effects and the development of resistance consolidate the urgent need to discover new targets. In TNBC, filamin A expression correlates to grade and TNM stage. Accordingly, this protein could constitute a new target for this BC subtype. Even if most of the data indicates its direct involvement in cancer progression, some contrasting results underline the need to deepen the studies. To elucidate a possible function of this protein as a TNBC marker, we summarized the main characteristic of filamin A and its involvement in physiological and pathological processes such as cancer. Lastly, we scrutinized its actions in triple-negative breast cancer and highlighted the need to increase the number of studies useful to better clarify the role of this versatile protein as a marker and target in TNBC, alone or in "collaboration" with other proteins with a relevant role in this BC subgroup., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Despite ongoing research in the field of breast cancer, the morbidity rates indicate that the disease remains a significant challenge. While patients with primary tumors have relatively high survival rates, these chances significantly decrease once metastasis begins. Thus, exploring alternative approaches, such as targeting proteins overexpressed in malignancies, remains significant. Filamin A (FLNa), an actin-binding protein (ABP), is involved in various cellular processes, including cell migration, adhesion, proliferation, and DNA repair. Overexpression of the protein was confirmed in samples from patients with numerous oncological diseases such as prostate, lung, gastric, colorectal, and pancreatic cancer, as well as breast cancer. Although most researchers concur on its role in promoting breast cancer progression and aggressiveness, discrepancies exist among studies. Moreover, the precise mechanisms through which FLNa affects cell migration, invasion, and even cancer progression remain unclear, highlighting the need for further research. To evaluate FLNa's potential as a therapeutic target, we have summarized its roles in breast cancer.

Somatostatin receptors (SSTs) are widely expressed in pituitary tumors and neuroendocrine neoplasms (NENs) of different origins, i.e. the gastrointestinal tract and the thorax (lungs and thymus), thus representing a well-established target for medical treatment with SST ligands (SRLs). However, the response to SRLs is highly heterogeneous between tumors. Two main factors can contribute to this variability: (i) the differential SST expression among tumor types and (ii) the differential expression/modulation of the SST-related intracellular machinery. In this literature review, we provide an overview of available data on the variable expression of SSTs in pituitary tumors and NENs, together with the resulting clinical implications. Moreover, we aim to describe the complex intracellular machinery involved in SST signaling and trafficking. Particularly, we will focus on β-arrestins and describe their role in receptor internalization and recycling, as well as the various functions of these scaffold molecules in tumor pathogenesis and progression. This review highlights the interplay between membrane receptors and intracellular machinery, together with its role in determining the clinical behavior of the tumor and the response to treatment in patients with pituitary tumors or NENs.

Background: Upon vessel injury, platelets adhere to exposed matrix constituents via specific membrane receptors, including the von Willebrand factor receptor glycoprotein (GP)Ib-IX-V complex and integrins β1 and β3. In platelets, the Fes/CIP4-homology Bin-Amphiphysin-Rvs protein PACSIN2 associates with the cytoskeletal and scaffolding protein filamin A (FlnA), linking GPIbα and integrins to the cytoskeleton., Objectives: Here we investigated the role of PACSIN2 in platelet function., Methods: Platelet parameters were evaluated in mice lacking PACSIN2 and platelet integrin β1., Results: Pacsin2 -/- mice displayed mild thrombocytopenia, prolonged bleeding time, and delayed thrombus formation in a ferric chloride-mediated carotid artery injury model, which was normalized by injection of control platelets. Pacsin2 -/- platelets formed unstable thrombi that embolized abruptly in a laser-induced cremaster muscle injury model. Pacsin2 -/- platelets had hyperactive integrin β1, as evidenced by increased spreading onto surfaces coated with the collagen receptor α2β1-specific peptide GFOGER and increased binding of the antibody 9EG7 directed against active integrin β1. By contrast, Pacsin2 -/- platelets had normal integrin αIIbβ3 function and expressed P-selectin normally following stimulation through the collagen receptor GPVI or with thrombin. Deletion of platelet integrin β1 in Pacsin2 -/- mice normalized platelet count, hemostasis, and thrombus formation. A PACSIN2 peptide mimicking the FlnA-binding site mediated the pull-down of a FlnA rod 2 construct by integrin β7, a model for integrin β-subunits., Conclusions: Pacsin2 -/- mice displayed severe thrombus formation defects due to hyperactive platelet integrin β1. The data suggest that PACSIN2 binding to FlnA negatively regulates platelet integrin β1 hemostatic function., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Mitral valve prolapse (MVP) is common among heart valve disease patients, causing severe mitral regurgitation (MR). Although complications such as cardiac arrhythmias and sudden cardiac death are rare, the high prevalence of the condition leads to a significant number of such events. Through next-generation gene sequencing approaches, predisposing genetic components have been shown to play a crucial role in the development of MVP. After the discovery of the X-linked inheritance of filamin A, autosomal inherited genes were identified. In addition, the study of sporadic MVP identified several genes, including DZIP1, TNS1, LMCD1, GLIS1, PTPRJ, FLYWCH, and MMP2. The early screening of these genetic predispositions may help to determine the patient population at risk for severe complications of MVP and impact the timing of reconstructive surgery. Surgical mitral valve repair is an effective treatment option for MVP, resulting in excellent short- and long-term outcomes. Repair rates in excess of 95% and low complication rates have been consistently reported for minimally invasive mitral valve repair performed in high-volume centers. We therefore conceptualize a potential preventive surgical strategy for the treatment of MVP in patients with genetic predisposition, which is currently not considered in guideline recommendations. Further genetic studies on MVP pathology and large prospective clinical trials will be required to support such an approach.

Due to the current limited knowledge about the role of filamin A (FLNA) in pituitary tumour progression, we aimed to analyse FLNA expression levels and its impact on aggressive markers of pituitary neuroendocrine tumours (PitNETs), using an integrative approach of in vivo and in vitro models and human samples. An increase in the expression levels of FLNA was observed in the advanced tumoural stages of the hyperplastic adenomatous pituitary model, concomitant with a decrease in cell proliferation and with a modification in the subcellular localisation of this protein. Similarly, overexpression of FLNA in the somatolactotropic GH3 cell line induced a decrease in the cell proliferation, promoted a migratory phenotype, increased invasion activity, and decreased the prolactin secretion. Cyclin D1 (CCND1) and cyclin-dependent kinase 4 (CDK4) expression increased in both models in correlation with the increase observed in FLNA levels. When human tissues were analysed a significant increase of FLNA was observed in PitNETs compared to normal pituitary gland, with heterogeneous intracellular localisation. Higher levels of FLNA expression were observed in tumours with invasive characteristics. These results underline the crucial roles of FLNA as a modulator of pathological markers and as a potential prognostic marker in pituitary tumours.

The insulin-like growth factor 2 (IGF2) promotes cell growth by overactivating the IGF system in an autocrine loop in adrenocortical carcinomas (ACCs). The cytoskeleton protein filamin A (FLNA) acts as a repressor of IGF2 mitogenic signalling in ACC cells. The aims of this study were to test FLNA expression by immunohistochemistry in 119 ACCs and 26 adrenocortical adenomas (ACAs) and to evaluate its relationship with clinicopathological features and outcome in ACCs. We found that 71.4% of ACCs did not express FLNA, whereas FLNA absence was a rare event in ACAs (15.4%, p < 0.001 vs. ACCs). In addition, the expression of FLNA was associated with a less aggressive tumour behaviour in ACCs. Indeed, the subgroup of ACCs with high FLNA showed a lower ENSAT stage, Weiss score, and S-GRAS score compared to ACCs with low FLNA expression ( p < 0.05). Moreover, patients with high FLNA had a longer overall survival than those with low FLNA ( p < 0.05). In conclusion, our data suggest that FLNA may represent a "protective" factor in ACCs, and the integration of FLNA immunohistochemical expression in ACC tissues along with other clinical and molecular markers could be helpful to improve diagnostic accuracy and prognosis prediction in ACCs.

In resting platelets, the 17 th domain of filamin a (FLNa17) constitutively binds to the platelet membrane glycoprotein Ibα (GPIbα) at its cytoplasmic tail (GPIbα-CT) and inhibits the downstream signal activation, while the binding of ligand and blood shear force can activate platelets. To imitate the pull force transmitted from the extracellular ligand of GPIbα and the lateral tension from platelet cytoskeleton deformation, two pulling modes were applied on the GPIbα-CT/FLNa17 complex, and the molecular dynamics simulation method was used to explore the mechanical regulation on the affinity and mechanical stability of the complex. In this study, at first, nine pairs of key hydrogen bonds on the interface between GPIbα-CT and FLNa17 were identified, which was the basis for maintaining the complex structural stability. Secondly, it was found that these hydrogen bonding networks would be broken down and lead to the dissociation of FLNa17 from GPIbα-CT only under the axial pull force; but, under the lateral tension, the secondary structures at both terminals of FLNa17 would unfold to protect the interface of the GPIbα-CT/FLNa17 complex from mechanical damage. In the range of 0~40 pN, the increase of pull force promoted outward-rotation of the nitrogen atom of the 563 rd phenylalanine (PHE 563 -N) at GPIbα-CT and the dissociation of the complex. This study for the first time revealed that the extracellular ligand-transmitted axial force could more effectively relieve the inhibition of FLNa17 on the downstream signal of GPIbα than pure mechanical tension at the atomic level, and would be useful for further understanding the platelet intracellular force-regulated signal pathway.

Developmental dysplasia of the hip (DDH), characterized by acetabular deformity that manifests from loose ligaments to complete dislocation of the hip, can cause notable pain and dysfunction and lead to hip dislocation, secondary fractures, scoliosis, and osteoarthritis of hip. Variants in FLNA may produce a spectrum of malformations in multiple organs, especially the skeleton. This study aimed to identify the genetic etiologies of DDH patients and provide genetic testing information for further diagnosis and treatment of DDH. We recruited a Chinese woman with DDH and her family members. Whole-exome sequencing was used to identify the patient's genetic etiologies. Protein models were used to analyze the pathogenic mechanism of the identified variants. A novel variant (c.3493T>G, p.C1165G) of FLNA was detected. The structural models of the mutant FLNA protein indicated that the variant would lose its sulfhydryl side chain and destroy the attraction between benzene rings and sulfhydryl. We reported a novel variant (c.3493T>G, p.C1165G) of FLNA in a Chinese woman with DDH. Our research outcome enriches the gene pool for hip dysplasia and emphasizes the pathogenicity of sulfhydryl side chain disruption in FLNA .

Prolonged exposure to arsenic can cause nonalcoholic steatohepatitis (NASH). The NOD-like receptor protein 3 (NLRP3) inflammasome plays an essential role in the process of NASH. However, the mechanism by which arsenic promotes NLRP3 expression remains unclear. Three-month NaAsO 2 gavage led to the nuclear factor-κB (NF-κB) signaling pathway activation and NASH. Additionally, NaAsO 2 upregulated the level of Filamin A (FLNA) and pyroptosis, thereby activating the NLRP3 inflammasome in SD rat liver. Using FLNA siRNA, NASH-associated inflammation and pyroptosis were clearly mitigated by reducing activation of the NLRP3 inflammasome. Furthermore, arsenic treatment facilitated activation of the NF-κB signaling pathway and promoted p-p65 translocation into the nucleus. Chromatin immunoprecipitation (Ch-IP) assay indicated that FLNA promoted p65 binding to the NLRP3 gene and upregulated the transcription of NLRP3, ultimately leading to pyroptosis and NASH. Our findings indicate that FLNA and pyroptosis are strongly associated with NASH induced by NaAsO 2 . Collectively, the findings of this study indicated that FLNA mediates NF-κB signaling pathway-induced activation of the NLRP3 inflammasome and ultimately activates pyroptosis and NASH upon NaAsO 2 exposure. This information may be useful for improving therapeutic strategies against arsenic-induced NASH., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The decades-old cholinergic hypothesis of Alzheimer's disease (AD) led to clinical testing and FDA approval of acetylcholinesterase inhibitor drugs. Subsequently, the α7 nicotinic acetylcholine receptor (α7nAChR) was proposed as a new drug target for enhancing cholinergic neurotransmission. Nearly simultaneously, soluble amyloid β 1-42 (Aβ 42 ) was shown to bind α7nAChR with picomolar affinity to activate kinases that hyperphosphorylate tau, the precursor to tau-containing tangles. Multiple biopharmaceutical companies explored α7nAChR as a drug target for AD, mostly to enhance neurotransmission. Directly targeting α7nAChR proved to be a drug development challenge. The ultra-high-affinity interaction between Aβ 42 and α7nAChR posed a significant hurdle for direct competition in the AD brain. The receptor rapidly desensitizes, undermining efficacy of agonists. Drug discovery approaches therefore included partial agonists and allosteric modulators of α7nAChR. After substantial effort, numerous drug candidates were abandoned due to lack of efficacy or drug-related toxicities. As alternatives, proteins interacting with α7nAChR were sought. In 2016, a novel nAChR regulator was identified, but no drug candidates have emerged from this effort. In 2012, the interaction of filamin A with α7nAChR was shown to be critical to Aβ 42 's toxic signaling via α7nAChR, presenting a new drug target. The novel drug candidate simufilam disrupts the filamin A-α7nAChR interaction, reduces Aβ 42 's high-affinity binding to α7nAChR, and suppresses Aβ 42 's toxic signaling. Early clinical trials of simufilam showed improvements in experimental CSF biomarkers and indications of cognitive improvement in mild AD patients at 1 year. Simufilam is currently in phase 3 clinical trials as a disease-modifying treatment for AD., (© 2023 Wiley Periodicals LLC.)

Down syndrome is the most common chromosomal abnormality in humans. Patients with Down syndrome have hematologic disorders, including mild to moderate thrombocytopenia. In case of Down syndrome, thrombocytopenia is not associated with bleeding, and it remains poorly characterized regarding molecular mechanisms. We investigated the effects of overexpression of Dyrk1A, an important factor contributing to some major Down syndrome phenotypes, on platelet number and bleeding in mice. Mice overexpressing Dyrk1A have a decrease in platelet number by 20%. However, bleeding time was found to be reduced by 50%. The thrombocytopenia and the decreased bleeding time observed were not associated to an abnormal platelet receptors expression, to a defect of platelet activation by ADP, thrombin or convulxin, to the presence of activated platelets in the circulation or to an abnormal half-life of the platelets. To propose molecular mechanisms explaining this discrepancy, we performed a network analysis of Dyrk1A interactome and demonstrated that Dyrk1A, fibronectin and fibrinogen interact indirectly through two distinct clusters of proteins. Moreover, in mice overexpressing Dyrk1A, increased plasma fibronectin and fibrinogen levels were found, linked to an increase of the hepatic fibrinogen production. Our results indicate that overexpression of Dyrk1A in mice induces decreased bleeding consistent with increased plasma fibronectin and fibrinogen levels, revealing a new role of Dyrk1A depending on its indirect interaction with these two proteins., (© 2023 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Moyamoya angiopathy (MMA) is an uncommon cerebrovascular disease characterized by a progressive steno-occlusive lesion of the internal carotid artery and the compensatory development of an unstable network of collateral vessels. These vascular hallmarks are responsible for recurrent ischemic/hemorrhagic strokes. Surgical treatment represents the preferred procedure for MMA patients, and indirect revascularization may induce a spontaneous angiogenesis between the brain surface and dura mater (DM), whose function remains rather unknown. A better understanding of MMA pathogenesis is expected from the molecular characterization of DM. We performed a comprehensive, label-free, quantitative mass spectrometry-based proteomic characterization of DM. The 30 most abundant identified proteins were located in the extracellular region or exosomes and were involved in extracellular matrix organization. Gene ontology analysis revealed that most proteins were involved in binding functions and hydrolase activity. Among the 30 most abundant proteins, Filamin A is particularly relevant because considering its well-known biochemical functions and molecular features, it could be a possible second hit gene with a potential role in MMA pathogenesis. The current explorative study could pave the way for further analyses aimed at better understanding such uncommon and disabling intracranial vasculopathy.

Introduction: Implicated in both aging and Alzheimer's disease (AD), mammalian target of rapamycin (mTOR) is overactive in AD brain and lymphocytes. Stimulated by growth factors such as insulin, mTOR monitors cell health and nutrient needs. A small molecule oral drug candidate for AD, simufilam targets an altered conformation of the scaffolding protein filamin A (FLNA) found in AD brain and lymphocytes that induces aberrant FLNA interactions leading to AD neuropathology. Simufilam restores FLNA's normal shape to disrupt its AD-associated protein interactions. Methods: We measured mTOR and its response to insulin in lymphocytes of AD patients before and after oral simufilam compared to healthy control lymphocytes. Results: mTOR was overactive and its response to insulin reduced in lymphocytes from AD versus healthy control subjects, illustrating another aspect of insulin resistance in AD. After oral simufilam, lymphocytes showed normalized basal mTOR activity and improved insulin-evoked mTOR activation in mTOR complex 1, complex 2, and upstream and downstream signaling components (Akt, p70S6K and phosphorylated Rictor). Suggesting mechanism, we showed that FLNA interacts with the insulin receptor until dissociation by insulin, but this linkage was elevated and its dissociation impaired in AD lymphocytes. Simufilam improved the insulin-mediated dissociation. Additionally, FLNA's interaction with Phosphatase and Tensin Homolog deleted on Chromosome 10 (PTEN), a negative regulator of mTOR, was reduced in AD lymphocytes and improved by simufilam. Discussion: Reducing mTOR's basal overactivity and its resistance to insulin represents another mechanism of simufilam to counteract aging and AD pathology. Simufilam is currently in Phase 3 clinical trials for AD dementia., Competing Interests: This basic research was supported in part by supplies and funding from Cassava Sciences, Inc. LB is an employee and shareholder of Cassava Sciences, as was the late NF. H-YW is an employee of City University of New York School of Medicine, is a long-time consultant and scientific advisor to Cassava Sciences, owns an insignificant financial interest (less than ½ of 1%) in the equity of Cassava Sciences, and has consulted to various pharmaceutical companies over the past 20 years. LB and H-YW are inventors on simufilam patents. None of the authors are due patent royalties. This basic research was conducted in non-GCP compliant facilities. ZP and K-CL are employees of City University of New York School of Medicine. BN, TD, and JP are investigators in the clinical trial protocols. Author BN is employed by IMIC, Inc. Authors TD and JP are employed by Cognitive Clinical Trials., (Copyright © 2023 Wang, Pei, Lee, Nikolov, Doehner, Puente, Friedmann and Burns.)

Social determinants of health (SDoH) include socioeconomic, environmental, and psychological factors that impact health. Neighborhood socioeconomic deprivation (NSD) and low individual-level socioeconomic status (SES) are SDoH that associate with incident heart failure, stroke, and cardiovascular mortality, but the underlying biological mechanisms are not well understood. Previous research has demonstrated an association between NSD, in particular, and key components of the neural-hematopoietic-axis including amygdala activity as a marker of chronic stress, bone marrow activity, and arterial inflammation. Our study further characterizes the role of NSD and SES as potential sources of chronic stress related to downstream immunological factors in this stress-associated biologic pathway. We investigated how NSD, SES, and catecholamine levels (as proxy for sympathetic nervous system activation) may influence monocytes which are known to play a significant role in atherogenesis. First, in an ex vivo approach, we treated healthy donor monocytes with biobanked serum from a community cohort of African Americans at risk for CVD. Subsequently, the treated monocytes were subjected to flow cytometry for characterization of monocyte subsets and receptor expression. We determined that NSD and serum catecholamines (namely dopamine [DA] and norepinephrine [NE]) associated with monocyte C-C chemokine receptor type 2 (CCR2) expression (p < 0.05), a receptor known to facilitate recruitment of monocytes towards arterial plaques. Additionally, NSD associated with catecholamine levels, especially DA in individuals of low SES. To further explore the potential role of NSD and the effects of catecholamines on monocytes, monocytes were treated in vitro with epinephrine [EPI], NE, or DA. Only DA increased CCR2 expression in a dose-dependent manner (p < 0.01), especially on non-classical monocytes (NCM). Furthermore, linear regression analysis between D2-like receptor surface expression and surface CCR2 expression suggested D2-like receptor signaling in NCM. Indicative of D2-signaling, cAMP levels were found to be lower in DA-treated monocytes compared to untreated controls (control 29.78 pmol/ml vs DA 22.97 pmol/ml; p = 0.038) and the impact of DA on NCM CCR2 expression was abrogated by co-treatment with 8-CPT, a cAMP analog. Furthermore, Filamin A (FLNA), a prominent actin-crosslinking protein, that is known to regulate CCR2 recycling, significantly decreased in DA-treated NCM (p < 0.05), indicating a reduction of CCR2 recycling. Overall, we provide a novel immunological mechanism, driven by DA signaling and CCR2, for how NSD may contribute to atherogenesis. Future studies should investigate the importance of DA in CVD development and progression in populations disproportionately experiencing chronic stress due to SDoH., Competing Interests: None

Aims: Degenerative mitral valve dystrophy (MVD) leading to mitral valve prolapse is the most frequent form of MV disease, and there is currently no pharmacological treatment available. The limited understanding of the pathophysiological mechanisms leading to MVD limits our ability to identify therapeutic targets. This study aimed to reveal the main pathophysiological pathways involved in MVD via the multimodality imaging and transcriptomic analysis of the new and unique knock-in (KI) rat model for the FilaminA-P637Q (FlnA-P637Q) mutation associated-MVD., Methods and Results: Wild-type (WT) and KI rats were evaluated morphologically, functionally, and histologically between 3-week-old and 3-to-6-month-old based on Doppler echocardiography, 3D micro-computed tomography (microCT), and standard histology. RNA-sequencing and Assay for Transposase-Accessible Chromatin (ATAC-seq) were performed on 3-week-old WT and KI mitral valves and valvular cells, respectively, to highlight the main signalling pathways associated with MVD. Echocardiographic exploration confirmed MV elongation (2.0 ± 0.1 mm vs. 1.8 ± 0.1, P = 0.001), as well as MV thickening and prolapse in KI animals compared to WT at 3 weeks. 3D MV volume quantified by microCT was significantly increased in KI animals (+58% vs. WT, P = 0.02). Histological analyses revealed a myxomatous remodelling in KI MV characterized by proteoglycans accumulation. A persistent phenotype was observed in adult KI rats. Signalling pathways related to extracellular matrix homeostasis, response to molecular stress, epithelial cell migration, endothelial to mesenchymal transition, chemotaxis and immune cell migration, were identified based on RNA-seq analysis. ATAC-seq analysis points to the critical role of transforming growth factor-β and inflammation in the disease., Conclusion: The KI FlnA-P637Q rat model mimics human myxomatous MVD, offering a unique opportunity to decipher pathophysiological mechanisms related to this disease. Extracellular matrix organization, epithelial cell migration, response to mechanical stress, and a central contribution of immune cells are highlighted as the main signalling pathways leading to myxomatous MVD. Our findings pave the road to decipher underlying molecular mechanisms and the specific role of distinct cell populations in this context., Competing Interests: Conflict of interest: None declared., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Filamin A (FLNA) is an actin cross-linking protein that mediates mechanotransduction. Force-dependent conformational changes of FLNA molecule expose cryptic binding site of FLNA, allowing interaction with partners such as integrin, smoothelin, and fimbacin. Here, we identified La-related protein 4 (LARP4) as a new FLNA mechanobinding partner. LARP4 specifically interacts with the cleft formed by C and D strands of immunoglobulin-like repeat 21 (R21) which is blocked by A strand of R20 without force. We validated the interaction between LARP4 and FLNA R21 both in vivo and in vitro . We also determined the critical amino acid that is responsible for the interaction and generated the non-FLNA-binding mutant LARP4 (F277A in human: F273A in mouse Larp4) that disrupts the interaction. Fluorescence recovery after photobleaching (FRAP) of GFP-labeled LARP4 in living cells demonstrated that mutant LARP4 diffuses faster than WT LARP4. Proximity ligation assay (PLA) also confirmed their interaction and disruption of actin polymerization diminishes the interaction. Data mining of RNAseq analysis of LARP4 knockdown (KD) HEK293T cells suggested that LARP4 is involved in morphogenesis and cell motility. Consistent with this prediction, we found that KD of LARP4 increases cell migration speed and expression of the F277A mutant LARP4 in LARP4-KD cells also leads to a higher cell migration speed compared to WT LARP4. These results demonstrated that the LARP4 interaction with FLNA regulates cell migration., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Mao and Nakamura.)

Non-alcoholic steatohepatitis (NASH) is a chronic and progressive liver disease characterized by steatosis, inflammation, and fibrosis. Filamin A (FLNA), an actin-binding protein, is involved in various cell functions, including the regulation of immune cells and fibroblasts. However, its role in the development of NASH through inflammation and fibrogenesis is not fully understood. In this study, we found that FLNA expression was increased in liver tissues of patients with cirrhosis and mice with non-alcoholic fatty liver disease (NAFLD)/NASH and fibrosis. Immunofluorescence analysis showed that FLNA was primarily expressed in macrophages and hepatic stellate cells (HSCs). Knocking down of FLNA by specific shRNA in phorbol-12-myristate-13-acetate (PMA)-derived THP-1 macrophages reduced lipopolysaccharide (LPS)-stimulated inflammatory response. The decreased mRNA levels of inflammatory cytokines and chemokines and suppression of the STAT3 signaling were observed in FLNA-downregulated macrophages. In addition, knockdown of FLNA in immortalized human hepatic stellate cells (LX-2 cells) resulted in decreased mRNA levels of fibrotic cytokines and enzymes involved in collagen synthesis, as well as increased levels of metalloproteinases and pro-apoptotic proteins. Overall, these results suggest that FLNA may contribute to the pathogenesis of NASH through its role in the regulation of inflammatory and fibrotic mediators., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The immunoglobulin-like cell adhesion molecule CLMP is a member of the CAR family of cell adhesion proteins and is implicated in human congenital short-bowel syndrome (CSBS). CSBS is a rare but very severe disease for which no cure is currently available. In this review, we compare data from human CSBS patients and a mouse knockout model. These data indicate that CSBS is characterized by a defect in intestinal elongation during embryonic development and impaired peristalsis. The latter is driven by uncoordinated calcium signaling via gap junctions, which is linked to a reduction in connexin43 and 45 levels in the circumferential smooth muscle layer of the intestine. Furthermore, we discuss how mutations in the CLMP gene affect other organs and tissues, including the ureter. Here, the absence of CLMP produces a severe bilateral hydronephrosis-also caused by a reduced level of connexin43 and associated uncoordinated calcium signaling via gap junctions.

Filamin A (FLNA) is an actin crosslinking protein that mediates mechanotransduction. External and internal mechanical forces, through the actin cytoskeleton, can induce conformational changes of the FLNA molecule to expose cryptic binding sites for its binding partners. Here, we identified Ras GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) as a new FLNA mechanobinding partner. Unlike other FLNA binding partners to the mechanosensing domain repeat 21 (R21), G3BP1 requires an additional neighboring repeat R22 to interact. We demonstrated that their interaction occurs in the cytosol of living cells in an actin polymerization-dependent manner. We also mapped the FLNA-binding site on G3BP1 and found that a F360A point mutation in the RNA recognition motif disrupts the interaction. RNA interfered with the FLNA-G3BP1 interaction, and FLNA did not localize in RNA-rich stress granules (SGs). Disruption of the interaction was sufficient to promote phase-separated SG formation, and arsenite treatment further stimulated the formation of SGs. Taken together, these data identify G3BP1 as a new mechanobinding protein that interacts with the FLNA mechanosensing domain R21 and suggest that SG formation is partially regulated by mechanical force., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Filamin A (FLNA) is a high molecular weight cytoskeleton protein important for cell locomotion. A relationship between FLNA mutations and pulmonary arterial hypertension (PAH) has previously been reported; however, the detailed mechanism remains unclear. The present study aimed to explore the role of FLNA in vascular smooth muscle cells during the development of PAH. Smooth muscle cell (SMC)‑specific FLNA‑deficient mice were generated and the mice were then exposed to hypoxia for 28 days to build the mouse model of PAH. Human pulmonary arterial smooth muscle cells (PASMCs) were also cultured and transfected with FLNA small interfering RNA or overexpression plasmids to investigate the effects of FLNA on PASMC proliferation and migration. Notably, compared with control individuals, the expression levels of FLNA were increased in lung tissues from patients with PAH, and it was obviously expressed in the PASMCs of pulmonary arterioles. FLNA deficiency in SMCs attenuated hypoxia‑induced pulmonary hypertension and pulmonary vascular remodeling. In vitro studies suggested that absence of FLNA impaired PASMC proliferation and migration, and produced lower levels of phosphorylated (p)‑PAK‑1 and RAC1 activity. However, FLNA overexpression promoted PASMC proliferation and migration, and increased the expression levels of p‑PAK‑1 and RAC1 activity. The present study highlights the role of FLNA in pulmonary vascular remodeling; therefore, it could serve as a potential target for the treatment of PAH.

Mitral valve prolapse (MVP) is a common condition affecting 2-3% of the general population, and the most complex form of valve pathology, with a complication rate up to 10-15% per year in advanced stages. Complications include mitral regurgitation which can lead to heart failure and atrial fibrillation, but also life-threatening ventricular arrhythmia and cardiovascular death. Sudden death has been recently brought to the forefront of MVP disease, increasing the complexity of management and suggesting that MVP condition is not properly understood. MVP can occur as part of syndromic conditions such as Marfan syndrome, but the most common form is non-syndromic, isolated or familial. Although a specific X-linked form of MVP was initially identified, autosomal dominant inheritance appears to be the primary mode of transmission. MVP can be stratified into myxomatous degeneration (Barlow), fibroelastic deficiency, and Filamin A-related MVP. While FED is still considered a degenerative disease associated with aging, myxomatous MVP and FlnA-MVP are recognized as familial pathologies. Deciphering genetic defects associated to MVP is still a work in progress; although FLNA , DCHS1 , and DZIP1 have been identified as causative genes in myxomatous forms of MVP thanks to familial approaches, they explain only a small proportion of MVP. In addition, genome-wide association studies have revealed the important role of common variants in the development of MVP, in agreement with the high prevalence of this condition in the population. Furthermore, a potential genetic link between MVP and ventricular arrhythmia or a specific type of cardiomyopathy is considered. Animal models that allow to advance in the genetic and pathophysiological knowledge of MVP, and in particular those that can be easily manipulated to express a genetic defect identified in humans are detailed. Corroborated by genetic data and animal models, the main pathophysiological pathways of MVP are briefly addressed. Finally, genetic counseling is considered in the context of MVP., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Delwarde, Capoulade, Mérot, Le Scouarnec, Bouatia-Naji, Yu, Huttin, Selton-Suty, Sellal, Piriou, Schott, Dina and Le Tourneau.)

The loss of function (LOF) due to mutations in the Filamin A (FLNA) gene may result in abnormality of the FLNA protein. Of the many clinical syndromes, this condition may produce chronic lung disease, which usually presents and is diagnosed in the infant/toddler age group. Its clinical pattern may mimic broncho-pulmonary dysplasia. It is part of the entities included in childhood interstitial lung disease group of disorders. We are herein reporting a patient that was diagnosed with FLNA-associated lung disease at 11 years of age. This case provides a unique insight into the long-term course of lung disease in this illness and broadens our understanding of the spectrum of its presentation. Although the patient had symptoms early in life, the diagnosis was not entertained because of the rarity of the disorder, its atypical and clinically mild presentation, and discontinuous care due to parents moving to different cities for employment reasons. Her presentation to our institution was for pneumonia. Due to highly unusual chest X-ray images, asthenia, and early clubbing, an extensive workup included further imaging and a lung biopsy. The final diagnosis was confirmed by the detection of FLNA LOF gene mutation., (© 2022 Wiley Periodicals LLC.)

Platelets are anucleate cells that are essential for hemostasis and wound healing. Upon activation of the cell surface receptors by their corresponding extracellular ligands, platelets undergo rapid shape change driven by the actin cytoskeleton; this shape change reaction is modulated by a diverse array of actin-binding proteins. One actin-binding protein, filamin A (FLNA), cross-links and stabilizes subcortical actin filaments thus providing stability to the cell membrane. In addition, FLNA binds the intracellular portion of multiple cell surface receptors and acts as a critical intracellular signaling scaffold that integrates signals between the platelet's plasma membrane and the actin cytoskeleton. This mini-review summarizes how FLNA transduces critical cell signals to the platelet cytoskeleton., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 De Silva, Hong, Falet and Kim.)

Rationale: Variable disease course and outcomes have been reported in children's interstitial lung disease associated with FLNA (Filamin A gene) variants. Objectives: To further delineate long-term respiratory outcomes and identify potential contributing factors to severe disease course. Methods: We retrospectively collected longitudinal data from three centers on nine cases (one male) with FLNA variants and early respiratory disease onset (within the first 24 mo of life). Clinical, radiographic, and histopathologic data were analyzed, focusing on cardiorespiratory disease course. Results: All required early respiratory support (three invasive ventilation, three noninvasive ventilation, three supplemental oxygen), and all experienced frequent severe infective respiratory exacerbations. Three died in infancy from refractory respiratory failure and pulmonary hypertension (PH). The six surviving individuals were 3, 10, 11, 15, 18, and 33 years old at time of reporting. The extent of functional respiratory impairment decreased with age; at last follow-up, there were no individuals on home invasive ventilation, one on nocturnal noninvasive ventilation, four on oxygen, and one on no respiratory support. Spirometry consistently demonstrated moderate to severe obstructive defects (forced expiratory volume in 1 s/forced vital capacity [FVC] z -score, -3.76 to -1.77; percent predicted FVC, 31.5% to 92.1%). Seven required PH treatment in early childhood (7/9), and three of the survivors (3/6) still receive treatment. Radiologic and histopathologic findings were consistent among cases. Conclusions: Early mortality was common, but many survivors stabilized even after severe symptoms in infancy. All survivors had persistent obstructive defects on spirometry, and half have persistent or recurrent PH. These typical findings are suggestive of this rare diagnosis and should prompt consideration of genetic testing.

A rare case of extensive multiple idiopathic cervical root resorption with potential genetic predisposition was presented. A heathy 19-year-old Chinese male with no contributory medical or family/social history complained of pain during mastication that lasted for several months. Oral examination identified 7 missing teeth and external cervical root resorption involving 9 teeth. Comparison of orthopantomograms taken in May 2021 and February 2022 identified that cervical root resorption occurred in 22 teeth. Resorption commenced at the cementoenamel junction and progressed rapidly over the 9-month period. Laboratory test results were within normal limits. Trio-based whole-exome sequencing showed a missense mutation c.5630 C > T in the filamin A (FLNA) gene at chromosome X of the subject. This is suggestive of the possibility of sex-linked recessive inheritance. This is the first study to report FLNA mutation in human subjects with cervical root resorption involving multiple teeth., (Copyright © 2022 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Sinus of Valsalva aneurysm (SVA) is a rare cardiovascular disease with male predominance. Recently, an association with aortic aneurysm and SVA has been revealed in periventricular nodular heterotopia patients with loss-of-function Filamin A (FLNA) mutations, which were located on chromosome X and almost exclusively affect females.Among patients hospitalized for aortic surgery with aortic root diameter over 4.0 cm, next-generation sequencing was performed to investigate 30 candidate genes related to inherited aortic aneurysm syndromes and familial thoracic aortic aneurysm and dissection. The present report reviewed an electronic case database and identified two female cases of unruptured SVA with heterozygous FLNA truncating mutations.Case 1 displaying a rare SVA phenotype involving left and noncoronary sinus harbored a nonsense variant p.Tyr1720Ter/c.5160C > G. Case 2 displayed right and noncoronary SVA with predominantly enlarged right coronary sinus, posterior mitral valve prolapse, and harbored a frameshift variant p.Val1724fs*68/c.5171&#95;5172delTG. Both novel mutations resulted in the premature termination of filamin A with the loss of functional Rod 2 and dimerization region.The present report raised the possibility of the presence of a cardiovascular onset form in the spectrum of FLNA hereditary diseases. The association between SVA and loss-of-function FLNA mutations indicates a unique etiology and pathogenesis among female patients, which requires further investigation to establish the linkage between FLNA variants and a wide spectrum of phenotypes.

Introduction: Alzheimer's disease (AD) is a multifactorial disorder diagnosed through the assessment of amyloid-beta (Aβ) and tau protein depositions. Filamin A (FLNA) could be a key partner of both Aβ and tau pathological processes and may be an important contributor to AD progression. The main aim of this study was to describe the differences in FLNA levels across clinicopathologic groups., Methods: From parietal cortex samples of 57 individuals (19 with no cognitive impairment (NCI), 19 mild cognitively impaired (MCI) and 19 with dementia) from the Religious Orders Study (ROS), we quantified total tau, phosphorylated tau (pTau), FLNA, synaptophysin, vesicular acetylcholine transporters (VAChT) and choline acetyltransferase (ChAT) by Western blot. Aβ42 and neuritic plaques (NP) were quantified by ELISA and Bielschowsky silver impregnation, respectively. AD staging was determined using ABC method combining Thal, Braak and the CERAD staging. From this, clinicopathologic stages of AD were established by subdividing subjects with neuropathological AD between preclinical AD, prodromal AD and AD dementia (ADD). Receiver operating characteristics analyses were performed to predict AD neuropathology from FLNA quantifications., Results: Insoluble FLNA was significantly and positively correlated with Aβ42, NP, Thal stages, ABC scores and AD clinicopathologic stages ( p  < 0.05 False discovery rate-corrected). No correlation of FLNA with tau measures was found. Insoluble FLNA levels were significantly higher in the prodromal AD, ADD and intermediate ABC groups. This was consistent with significantly lower levels of soluble FLNA specifically in prodromal AD. Insoluble (AUC: 0.830) and soluble FLNA levels (AUC: 0.830) as well as the ratio of soluble over insoluble FLNA (AUC: 0.852), were excellent predictors of prodromal AD among subjects with MCI from the ROS cohort., Discussion: We observed opposite level changes between insoluble and soluble FLNA in prodromal AD. As this stage coincides with the appearance of cognitive symptoms, this may be a key event in the transition from preclinical to prodromal AD. Insoluble FLNA could be useful to identify prodromal AD among subjects with an MCI, indicating that it might be a hallmark of prodromal AD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Aumont, Tremblay, Levert, Bennett, Calon and Leclerc.)

Adenosine to inosine (A to I) editing is mediated by adenosine deaminases acting on RNA (ADAR) enzymes. Inosines are interpreted as guanosines by the translational machinery. Consequently, A to I editing in mRNAs can lead to their recoding and the formation of proteins not encoded in the genome. Filamin A is an actin-crosslinking protein. A to I editing in the filamin pre-mRNA leads to the exchange of a glutamine to an arginine in a highly interactive domain of the protein. However, the consequences of this editing event are still poorly understood. Here we show, using transgenic mice expressing either constitutively edited or constitutively uneditable filamin A that filamin A editing critically controls angiogenesis in tumors but also in a mouse ischemia model. Hyper-editing reduces angiogenesis, while hypoediting leads to increased angiogenesis, possibly by altering vascular endothelial growth factor receptor 2 (VEGFR2) turnover. Further, FLNA editing of the tumor itself seemingly affects its metastatic potential by changing its interaction with the extracellular matrix. We therefore identify filamin A editing as a critical component for angiogenesis, tumor growth, and metastasis formation., Competing Interests: M.F.J. and the Medical University of Vienna have filed a European patent for the therapeutic use of FLNA editing to control tumor growth, inflammation, and angiogenesis., (© 2022 The Author(s).)

A-to-I RNA editing by ADARs is an abundant epitranscriptomic RNA-modification in metazoa. In mammals, Flna pre-mRNA harbours a single conserved A-to-I RNA editing site that introduces a Q-to-R amino acid change in Ig repeat 22 of the encoded protein. Previously, we showed that FLNA editing regulates smooth muscle contraction in the cardiovascular system and affects cardiac health. The present study investigates how ADAR2-mediated A-to-I RNA editing of Flna affects actin crosslinking, cell mechanics, cellular adhesion and cell migration. Cellular assays and AFM measurements demonstrate that the edited version of FLNA increases cellular stiffness and adhesion but impairs cell migration in both, mouse fibroblasts and human tumour cells. In vitro, edited FLNA leads to increased actin crosslinking, forming actin gels of higher stress resistance. Our study shows that Flna RNA editing is a novel regulator of cytoskeletal organisation, affecting the mechanical property and mechanotransduction of cells., (© 2022 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Objective: To study the radiosensitization effect of curcumin, a natural product with anti-inflammatory and anti-cancer properties, in bladder cancer cells and identify the specific role of FLNA gene in that process., Methods: CCK-8 method was initially adopted to identify the proper interventional concentration of curcumin. T24 bladder cancer cells were subjected to CCK-8, flow cytometry, and colony formation assay to study the cell biological behaviors under different interventions. γ-H2AX test was performed to test the level of damage in T24 cells. RT-qPCR and Western blot were conducted to measure FLNA mRNA and protein levels., Results: Low-dose curcumin (10, 20 μM) following X-ray exposure resulted in increased DNA damage, augmented apoptosis, and reduced proliferation of T24 cells. Certain radiosensitization was demonstrated when curcumin was applied at 10 μM. Additionally, elevation of FLNA gene and protein levels was also indicated upon combination treatment., Conclusion: Low-dose curcumin has certain radiosensitization effect in bladder cancer, where FLNA plays a certain regulatory role., Competing Interests: Declaration of Conflicting Interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s) 2022.)

Pathogenic variants of the X-linked FLNA gene encoding filamin A protein have been associated with a wide spectrum of symptoms, including the recently described pulmonary phenotype with childhood-onset panlobular emphysema. We describe three female patients from two families with novel heterozygous FLNA variants c.5837&#95;2del and c.508C > T. Analysis of immunofluorescence of peripheral blood smears and platelet function was performed for all patients. FLNA-negative platelets were observed, suggesting that these variants result in the loss of a functional protein product. All three patients also had periventricular nodular heterotopia and panlobular emphysema. However, they had considerably milder symptoms and later age of onset than in the previously reported cases. Therefore, patients with pathogenic FLNA variants should be studied actively for lung involvement even in the absence of pronounced respiratory symptoms. Conversely, any patient with unexplained panlobular emphysema should be analyzed for pathogenic FLNA variants. We also suggest that immunofluorescence analysis is a useful tool for investigating the pathogenicity of novel FLNA variants., (© 2022 The Authors. American Journal of Medical Genetics Part A published by Wiley Periodicals LLC.)

Glycoprotein (GP) Ibα shedding mediated by ADAM17 (a disintegrin and metalloproteinase 17) plays an important role in negatively regulating platelet function and thrombus formation. However, the mechanism of GPIbα shedding remains elusive. Here, we show that jasplakinolide (an actin-polymerizing peptide)-induced actin polymerization results in GPIbα shedding and impairs platelet function. Thrombin and A23187-induced GPIbα shedding is increased by jasplakinolide; in contrast, GPIbα shedding is reduced by a depolymerization regent (cytochalasin B). We find that actin polymerization activates calpain leading to filamin A hydrolyzation. We further demonstrate that the interaction of filamin A with the cytoplasmic domain of GPIbα plays a critical role in regulating actin polymerization-induced GPIbα shedding. Taken together, these data demonstrate that actin polymerization regulates ADAM17-mediated GPIbα shedding, suggesting a novel strategy to negatively regulate platelet function.

Adrenocorticotropic hormone (ACTH)-secreting pituitary tumors mainly express somatostatin receptor 5 (SSTR5) since SSTR2 is downregulated by the elevated levels of glucocorticoids that characterize patients with Cushing's disease (CD). SSTR5 is the molecular target of pasireotide, the only approved pituitary tumor-targeted drug for the treatment of CD. However, the molecular mechanisms that regulate SSTR5 are still poorly investigated. This review summarizes the experimental evidence supporting the role of the cytoskeleton actin-binding protein filamin A (FLNA) in the regulation of SSTR5 expression and signal transduction in corticotroph tumors. Moreover, the correlations between the presence of somatic USP8 mutations and the expression of SSTR5 will be reviewed. An involvement of glucocorticoid-mediated β-arrestins modulation in regulating SSTRs expression and function in ACTH-secreting tumors will also be discussed., Competing Interests: The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported., (© The authors.)

Background: Filamin (FLN) regulates many cell functions through its scaffolding activity cross-linking cytoskeleton and integrins. FLN was shown to inhibit integrin activity, but the exact mechanism remains unclear., Objectives: The aim of this study was to evaluate the role of filamin A (FLNa) subdomains on the regulation of integrin αIIbβ3 signaling., Methods: Three FLNa deletion mutants were overexpressed in the erythro-megakaryocytic leukemic cell line HEL: Del1, which lacks the N-terminal CH1-CH2 domains mediating the FLNa-actin interaction; Del2, lacking the Ig-like repeat 21, which mediates the FLNa-β3 interaction; and Del3, lacking the C-terminal Ig repeat 24, responsible for FLNa dimerization and interaction with the small Rho guanosine triphosphatase involved in actin cytoskeleton reorganisation. Fibrinogen binding to HEL cells in suspension and talin-β3 proximity in cells adherent to immobilized fibrinogen were assessed before and after αIIbβ3 activation by the protein kinase C agonist phorbol 12-myristate 13-acetate., Results: Our results show that FLNa-actin and FLNa-β3 interactions negatively regulate αIIbβ3 activation. Moreover, FLNa-actin interaction represses Rac activation, contributing to the negative regulation of αIIbβ3 activation. In contrast, the FLNa dimerization domain, which maintains Rho inactive, was found to negatively regulate αIIbβ3 outside-in signaling., Conclusion: We conclude that FLNa negatively controls αIIbβ3 activation by regulating actin polymerization and restraining activation of Rac, as well as outside-in signaling by repressing Rho., (© 2022 The Authors. Research and Practice in Thrombosis and Haemostasis published by Wiley Periodicals LLC on behalf of International Society on Thrombosis and Haemostasis (ISTH).)

Membrane proteins often cluster in nanoscale membrane domains (lipid rafts) that coalesce into ceramide-rich platforms during cell stress, however the clustering mechanisms remain uncertain. The cystic fibrosis transmembrane conductance regulator (CFTR), which is mutated in cystic fibrosis (CF), forms clusters that are cholesterol dependent and become incorporated into long-lived platforms during hormonal stimulation. We report here that clustering does not involve known tethering interactions of CFTR with PDZ domain proteins, filamin A or the actin cytoskeleton. It also does not require CFTR palmitoylation but is critically dependent on membrane lipid order and is induced by detergents that increase the phase separation of membrane lipids. Clustering and integration of CFTR into ceramide-rich platforms are abolished by the disease mutations F508del and S13F and rescued by the CFTR modulators elexacaftor plus tezacaftor. These results indicate CF therapeutics that correct mutant protein folding restore both trafficking and normal lipid interactions in the plasma membrane. This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

Osteoporosis (OP) is a worldwide prevalent chronic metabolic bone disease, causing by a disruption of the balance between bone resorption and formation. Estrogen deficiency and aging are the main causes for disturbances in bone remodeling activity and bone loss, however, the mechanisms underlying bone remodeling regulation require clarification if novel targets for OP treatment are to be identified. In this investigation, we showed that filamin A (FLNA) accumulated in osteoblasts (OBs) and osteoclasts (OC) in bone from human OP samples, and mice with age-related and postmenopausal OP. FLNA negatively modulated in vitro osteogenic differentiation and positively promoted RANKL-induced osteoclastic differentiation. Mechanistically, FLNA interacted with low-density lipoprotein receptor-related proteins 6 (LRP6) to inhibit β-catenin expression, and enhanced nuclear factor of activated T cell c1 (NFATc1)-dependent osteoclastogenic gene expression to inhibit osteogenesis, and promote osteoclastogenesis. Inhibiting FLNA with calpeptin activated WNT/β-catenin signaling, resulting in prominent protective effects of bone loss in ovariectomy (OVX)-induced postmenopausal OP mice. Our findings revealed that FLNA not only participated in OP pathogenesis, but could be a new target to stimulate bone formation and inhibit bone resorption. Targeting FLNA with calpeptin may be a promising therapeutic approach for postmenopausal OP in the future., (Copyright © 2021. Published by Elsevier Inc.)

Objective: To investigate whether the selective cyclooxygenase-2 enzyme inhibitors celecoxib has protective effect on the liver of rats with type 2 diabetes mellitus (T2DM) combined with nonalcoholic steatohepatitis (NASH) via inhibiting the expression of Rho/ROCK pathway. Methods: Forty male SD rats were randomly divided into four groups: type 2 diabetes mellitus combined with nonalcoholic steatohepatitis (T2DM-NASH) group, T2DM-NASH + celecoxib group, control group, and control+celecoxib group. The T2DM-NASH and T2DM-NASH + celecoxib groups were fed with high-sugar and fat diet, and the control group and control + celecoxib group were fed with basal diet (25 kJ/kg). Four weeks later, streptozotocin (STZ, 30 mg/kg) was intraperitoneally injected into the NASH group and T2DM-NASH + celecoxib group to induce T2DM model, and the control group and control + celecoxib group were intraperitoneally injected with isovolumic citric acid-sodium citrate buffer. Four weeks after STZ injection, the T2DM-NASH + celecoxib group and the control + celecoxib group were gavaged with celecoxib (10 mg·kg·d) dissolved in normal saline for 4 weeks, and the remaining two groups of rats were gavaged with isovolumic normal saline for 4 weeks. Animals were sacrificed at the end of the 12- weeks, and the liver tissue was collected. Liver pathological changes were observed by HE staining. The expressions of RhoA, RhoA, ROCK1 and ROCK2 proteins in liver were detected by immunohistochemistry and western blot. The expressional condition of RhoA, ROCK1 and ROCK2 mRNA in liver were detected by real-time quantitative PCR. The differences were compared between protein and mRNA expression among the groups by analysis of variance and t-test. Results: Compared with the control group and the control + celecoxib group, the liver tissue of the T2DM-NASH group and the T2DM-NASH + celecoxib group had severe steatosis, and there was partial inflammatory cell infiltration under the light microscope. The expression levels of RhoA, ROCK1 and ROCK2 protein and mRNA were significantly increased ( P < 0.05) in each liver tissue, while liver steatosis was reduced to certain extent in T2DM-NASH + celecoxib group than T2DM-NASH group, and the expression levels of RhoA, ROCK1 and ROCK2 protein and mRNA were decreased in each liver tissue of T2DM-NASH group ( P < 0.05). Conclusion: The selective cyclooxygenase-2 enzyme inhibitors celecoxib has a protective effect on the liver of rats with T2DM-NASH, and its effect may be achieved by inhibiting the expression of Rho/ROCK pathway.