Dynamic shape-shifting of the single-celled eukaryotic predator Lacrymaria via unconventional cytoskeletal components.

Eukaryotic cells depend on dynamic changes in shape to fulfill a wide range of cellular functions, maintain essential biological processes, and regulate cellular behavior. The single-celled, predatory ciliate Lacrymaria exhibits extraordinary dynamic shape-shifting using a flexible "neck" that can stretch 7-8 times the length of its body to capture prey. The molecular mechanism behind this morphological change remains a mystery. We have observed that when in an active state, Lacrymaria repeatedly extends and contracts its neck to enable 360-degree space search and prey capture. This remarkable morphological change involves a unique actin-myosin system rather than the Ca2+-dependent system found in other contractile ciliates. Two cytoskeletons are identified in the cortex of the Lacrymaria cell, namely the myoneme cytoskeleton and the microtubule cytoskeleton. The myoneme cytoskeleton is composed of centrin-myosin proteins, exhibiting distinct patterns between the neck and body, with their boundary seemingly associated with the position of the macronucleus. A novel giant protein forming a ladder-like structure was discovered as a component of the microtubule cytoskeleton. Thick centrin-myosin fibers are situated very close to the right side of the ladders in the neck but are far away from such structures in the body. This arrangement enables the decoupling of the neck and body. Plasmodium-like unconventional actin has been discovered in Lacrymaria, and this may form highly dynamic short filaments that could attach to the giant protein and myosin, facilitating coordination between the two cytoskeletons in the neck. In summary, this fascinating organism employs unconventional cytoskeletal components to accomplish its extraordinary dynamic shape-shifting.

Protein P5 of pear chlorotic leaf spot-associated virus is a pathogenic factor that suppresses RNA silencing and enhances virus movement.

Pear chlorotic leaf spot-associated virus (PCLSaV) is a newly described emaravirus that infects pear trees. The virus genome consists of at least five single-stranded, negative-sense RNAs. The P5 encoded by RNA5 is unique to PCLSaV. In this study, the RNA silencing suppression (RSS) activity of P5 and its subcellular localization were determined in Nicotiana benthamiana plants by Agrobacterium tumefaciens-mediated expression assays and green fluorescent protein RNA silencing induction. Protein P5 partially suppressed local RNA silencing, strongly suppressed systemic RNA silencing and triggered reactive oxygen species accumulation. The P5 self-interacted and showed subcellular locations in plasmodesmata, endoplasmic reticulum and nucleus. Furthermore, P5 rescued the cell-to-cell movement of a movement defective mutant PVXΔP25 of potato virus X (PVX) and enhanced the pathogenicity of PVX. The N-terminal 1-89 amino acids of the P5 were responsible for the self-interaction ability and RSS activity, for which the signal peptide at positions 1-19 was indispensable. This study demonstrated the function of an emaravirus protein as a pathogenic factor suppressing plant RNA silencing to enhance virus infection and as an enhancer of virus movement.

Two plant membrane-shaping reticulon-like proteins play contrasting complex roles in turnip mosaic virus infection.

Positive-sense RNA viruses remodel cellular cytoplasmic membranes as the membranous sources for the formation of viral replication organelles (VROs) for viral genome replication. In plants, they traffic through plasmodesmata (PD), plasma membrane-lined pores enabling cytoplasmic connections between cells for intercellular movement and systemic infection. In this study, we employed turnip mosaic virus (TuMV), a plant RNA virus to investigate the involvement of RTNLB3 and RTNLB6, two ER (endoplasmic reticulum) membrane-bending, PD-located reticulon-like (RTNL) non-metazoan group B proteins (RTNLBs) in viral infection. We show that RTNLB3 interacts with TuMV 6K2 integral membrane protein and RTNLB6 binds to TuMV coat protein (CP). Knockdown of RTNLB3 promoted viral infection, whereas downregulation of RTNLB6 restricted viral infection, suggesting that these two RTNLs play contrasting roles in TuMV infection. We further demonstrate that RTNLB3 targets the α-helix motif 42LRKSM46 of 6K2 to interrupt 6K2 self-interactions and compromise 6K2-induced VRO formation. Moreover, overexpression of AtRTNLB3 apparently promoted the selective degradation of the ER and ER-associated protein calnexin, but not 6K2. Intriguingly, mutation of the α-helix motif of 6K2 that is required for induction of VROs severely affected 6K2 stability and abolished TuMV infection. Thus, RTNLB3 attenuates TuMV replication, probably through the suppression of 6K2 function. We also show that RTNLB6 promotes viral intercellular movement but does not affect viral replication. Therefore, the proviral role of RTNLB6 is probably by enhancing viral cell-to-cell trafficking. Taken together, our data demonstrate that RTNL family proteins may play diverse complex, even opposite, roles in viral infection in plants.

Influence of the P6 effector protein of Cauliflower mosaic virus (CaMV) on the sustained expression and subcellular localization of the CaMV movement protein.

The P6 protein of cauliflower mosaic virus (CaMV) is a multifunctional protein that forms the electron dense, amorphous inclusion bodies that accumulate in the cytoplasm and has been shown to physically interact with all other CaMV proteins, including the CaMV movement protein (P1). In this study, we have investigated the subcellular localization of the P6 and P1 proteins in transient expression assays in Nicotiana benthamiana, as well as the influence of P6 on the expression and subcellular localization of P1. A version of P6 tagged with RFP was shown to envelop the endoplasmic reticulum (ER), whereas P1 tagged with RFP was shown to induce the fragmentation of the ER. Co-expression of P6 with P1 led to an enhancement of the spatial and temporal expression of P1, with a shift from expression through the plasma membrane and interior of the cell to punctate spots associated with the cell wall.

Rare Variant in MRC2 Associated With Familial Supraventricular Tachycardia and Wolff-Parkinson-White Syndrome.

BACKGROUND: Accessory pathways are a common cause of supraventricular tachycardia (SVT) and can lead to sudden cardiac death in otherwise healthy children and adults when associated with Wolff-Parkinson-White syndrome. The goal of this study was to identify genetic variants within a large family with structurally normal hearts affected by SVT and Wolff-Parkinson-White syndrome and determine causality of the gene deficit in a corresponding mouse model. METHODS: Whole exome sequencing performed on 2 distant members of a 3-generation family in which multiple members were affected by SVT or Wolff-Parkinson-White pattern (preexcitation) on ECG identified MRC2 as a candidate gene. Serial electrocardiograms, intracardiac electrophysiology studies, echocardiography, optical mapping studies, and histology were performed on both Mrc2 mutant and WT (wild-type) mice. RESULTS: A rare HET (heterozygous) missense variant c.2969A>G;p.Glu990Gly (E990G) in MRC2 was identified as the leading candidate gene variant segregating with the cardiac phenotype following an autosomal-dominant Mendelian trait segregation pattern with variable expressivity. In vivo electrophysiology studies revealed reentrant SVT in E990G mice. Optical mapping studies in E990G mice demonstrated abnormal retrograde conduction, suggesting the presence of an accessory pathway. Histological analysis of E990G mouse hearts showed a disordered ECM (extracellular matrix) in the annulus fibrosus. Finally, Mrc2 knockdown in human cardiac fibroblasts enhanced accelerated cell migration. CONCLUSIONS: This study identified a rare nonsynonymous variant in the MRC2 gene in individuals with familial reentrant SVT, Wolff-Parkinson-White ECG pattern, and structurally normal hearts. Furthermore, Mrc2 knock-in mice revealed an increased incidence of reentrant SVT and bypass tract formation in the setting of preserved cardiac structure and function.

Self-extinguishing relay waves enable homeostatic control of human neutrophil swarming.

Neutrophils collectively migrate to sites of injury and infection. How these swarms are coordinated to ensure the proper level of recruitment is unknown. Using an ex vivo model of infection, we show that human neutrophil swarming is organized by multiple pulsatile chemoattractant waves. These waves propagate through active relay in which stimulated neutrophils trigger their neighbors to release additional swarming cues. Unlike canonical active relays, we find these waves to be self-terminating, limiting the spatial range of cell recruitment. We identify an NADPH-oxidase-based negative feedback loop that is needed for this self-terminating behavior. We observe near-constant levels of neutrophil recruitment over a wide range of starting conditions, revealing surprising robustness in the swarming process. This homeostatic control is achieved by larger and more numerous swarming waves at lower cell densities. We link defective wave termination to a broken recruitment homeostat in the context of human chronic granulomatous disease.

Potato spindle tuber viroid (PSTVd) loop 27 mutants promote cell-to-cell movement and phloem unloading of the wild type: Insights into RNA-based viroid interactions.

Variations in infection progression with concurrent or prior infections by different viruses, viroids, or their strains are evident, but detailed investigations into viroid variant interactions are lacking. We studied potato spindle tuber viroid intermediate strain (PSTVd-I) to explore variant interactions. Two mutants, U177A/A182U (AU, replication- and trafficking-competent) and U178G/U179G (GG, replication-competent but trafficking-defective) on loop 27 increased cell-to-cell movement of wild-type (WT) PSTVd without affecting replication. In mixed infection assays, both mutants accelerated WT phloem unloading, while only AU promoted it in separate leaf assays, suggesting that enhancement of WT infection is not due to systemic signals. The mutants likely enhance WT infection due to their loop-specific functions, as evidenced by the lack of impact on WT infection seen with the distantly located G347U (UU) mutant. This study provides the first comprehensive analysis of viroid variant interactions, highlighting the prolonged phloem unloading process as a significant barrier to systemic spread.

Sizzled (Frzb3) physically interacts with noncanonical Wnt ligands to inhibit gastrulation cell movement.

The coordinated movement of germ layer progenitor cells reaches its peak at the dorsal side, where the Bmp signaling gradient is low, and minimum at the ventral side, where the Bmp gradient is high. This dynamic cell movement is regulated by the interplay of various signaling pathways. The noncanonical Wnt signaling cascade serves as a pivotal regulator of convergence and extension cell movement, facilitated by the activation of small GTPases such as Rho, Rab, and Rac. However, the underlying cause of limited cell movement at the ventral side remains elusive. To explore the functional role of a key regulator in constraining gastrulation cell movement at the ventral side, we investigated the Bmp4-direct target gene, sizzled (szl), to assess its potential role in inhibiting noncanonical Wnt signaling. In our current study, we demonstrated that ectopic expression of szl led to gastrulation defects in a dose-dependent manner without altering cell fate specification. Overexpression of szl resulted in decreased elongation of Activin-treated animal cap and Keller explants. Furthermore, our immunoprecipitation assay unveiled the physical interaction of Szl with noncanonical Wnt ligand proteins (Wnt5 and Wnt11). Additionally, the activation of small GTPases involved in Wnt signaling mediation (RhoA and Rac1) was diminished upon szl overexpression. In summary, our findings suggest that Bmp4 signaling negatively modulates cell movement from the ventral side of the embryo by inducing szl expression during early Xenopus gastrulation.

Clinical significance of downregulated NISCH expression in skin cutaneous melanoma: Modulation of tumor cell invasion, migration, and EMT via PAK1 inhibition.

OBJECTIVE: This study aimed to investigate the expression and functional role of NISCH in skin cutaneous melanoma (SKCM), exploring its association with clinical characteristics and its potential impact on human skin melanoma cell behavior. METHODS: The research assessed differential NISCH expression in SKCM tissues using the GEPIA (Gene Expression Profiling Interactive Analysis) database and validated these findings through immunohistochemical staining of 45 clinical samples. To affirm NISCH expression at the cellular level, three human skin melanoma cell lines (RPMI-7951, A375, MEL-5), and the human normal skin cell line HEMa underwent quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blotting. Transwell experiments evaluated the migration and invasion capabilities of RPMI-7951 and A375 cells post-transduction with NISCH or PAK1 lentiviral activation particles. Additionally, qRT-PCR analysis of epithelial-mesenchymal transition (EMT)-related gene expression (Vimentin, E-cadherin, N-cadherin) was conducted in A375 and RPMI-7951 cells. RESULTS: SKCM tissues exhibited significantly reduced NISCH expression compared to normal tissues. Immunohistochemical analysis revealed predominant nuclear localization of NISCH in melanoma cells, with reduced expression significantly correlating with sex, advanced stage, and lymph node metastasis. Melanoma cell lines displayed lower NISCH expression levels compared to normal skin cells. Functional experiments showcased that NISCH overexpression suppressed p-PAK1/PAK1, while PAK1 upregulation notably increased melanoma cell migration, invasion, and induced EMT. Remarkably, NISCH overexpression counteracted PAK1-induced effects on EMT, migration, and invasion in melanoma cells. CONCLUSION: NISCH may significantly influence the aggressive behavior of SKCM cells via the PAK1 pathway, making it a potential therapeutic target for managing melanoma metastasis.

Modelling non-local cell-cell adhesion: a multiscale approach.

Cell-cell adhesion plays a vital role in the development and maintenance of multicellular organisms. One of its functions is regulation of cell migration, such as occurs, e.g. during embryogenesis or in cancer. In this work, we develop a versatile multiscale approach to modelling a moving self-adhesive cell population that combines a careful microscopic description of a deterministic adhesion-driven motion component with an efficient mesoscopic representation of a stochastic velocity-jump process. This approach gives rise to mesoscopic models in the form of kinetic transport equations featuring multiple non-localities. Subsequent parabolic and hyperbolic scalings produce general classes of equations with non-local adhesion and myopic diffusion, a special case being the classical macroscopic model proposed in Armstrong et al. (J Theoret Biol 243(1): 98-113, 2006). Our simulations show how the combination of the two motion effects can unfold. Cell-cell adhesion relies on the subcellular cell adhesion molecule binding. Our approach lends itself conveniently to capturing this microscopic effect. On the macroscale, this results in an additional non-linear integral equation of a novel type that is coupled to the cell density equation.

Innovative Arylimidazole-Fused Phytovirucides via Carbene-Catalyzed [3+4] Cycloaddition: Locking Viral Cell-To-Cell Movement by Out-Competing Virus Capsid-Host Interactions.

The control of potato virus Y (PVY) induced crop failure is a challengeable issue in agricultural chemistry. Although many anti-PVY agents are designed to focus on the functionally important coat protein (CP) of virus, how these drugs act on CP to inactivate viral pathogenicity, remains largely unknown. Herein, a PVY CP inhibitor -3j (S) is disclosed, which is accessed by developing unusually efficient (up to 99% yield) and chemo-selective (> 99:1 er in most cases) carbene-catalyzed [3+4] cycloaddition reactions. Compound -3j bears a unique arylimidazole-fused diazepine skeleton and shows chirality-preferred performance against PVY. In addition, -3j (S) as a mediator allows ARG191 (R191) of CP to be identified as a key amino acid site responsible for intercellular movement of virions. R191 is further demonstrated to be critical for the interaction between PVY CP and the plant functional protein NtCPIP, enabling virions to cross plasmodesmata. This key step can be significantly inhibited through bonding with the -3j (S) to further impair pathogenic behaviors involving systemic infection and particle assembly. The study reveals the in-depth mechanism of action of antiviral agents targeting PVY CP, and contributes to new drug structures and synthetic strategies for PVY management.

Juvenile hormones direct primordial germ cell migration to the embryonic gonad.

Germ cells are essential to sexual reproduction. Across the animal kingdom, extracellular signaling isoprenoids, such as retinoic acids (RAs) in vertebrates and juvenile hormones (JHs) in invertebrates, facilitate multiple processes in reproduction. Here we investigated the role of these potent signaling molecules in embryonic germ cell development, using JHs in Drosophila melanogaster as a model system. In contrast to their established endocrine roles during larval and adult germline development, we found that JH signaling acts locally during embryonic development. Using an in vivo biosensor, we observed active JH signaling first within and near primordial germ cells (PGCs) as they migrate to the developing gonad. Through in vivo and in vitro assays, we determined that JHs are both necessary and sufficient for PGC migration. Analysis into the mechanisms of this newly uncovered paracrine JH function revealed that PGC migration was compromised when JHs were decreased or increased, suggesting that specific titers or spatiotemporal JH dynamics are required for robust PGC colonization of the gonad. Compromised PGC migration can impair fertility and cause germ cell tumors in many species, including humans. In mammals, retinoids have many roles in development and reproduction. We found that like JHs in Drosophila, RA was sufficient to impact mouse PGC migration in vitro. Together, our study reveals a previously unanticipated role of isoprenoids as local effectors of pre-gonadal PGC development and suggests a broadly shared mechanism in PGC migration.

Dopamine receptor D2 regulates genes involved in germ cell movement and sperm motility in rat testes†.

The function of dopamine receptor D2 (D2R) is well associated with sperm motility; however, the physiological role of D2R present on testicular cells remains elusive. The aim of the present study is to delineate the function of testicular D2R. Serum dopamine levels were found to decrease with age, whereas testicular D2R expression increased. In rat testicular sections, D2R immunolabeling was observed in interstitial cells, spermatogonia, spermatocytes and mature elongated spermatids, whereas tyrosine hydroxylase immunolabeling was selectively detected in Leydig cells. In vitro seminiferous tubule culture following bromocriptine (D2R agonist) treatment resulted in decreased cAMP levels. Microarray identified 1077 differentially expressed genes (511 up-regulated, 566 down-regulated). The majority of differentially expressed genes were present in post-meiotic cells including early and late spermatids, and sperm. Gene ontology elucidated processes related to extra-cellular matrix to be enriched and was supported by differential expression of various collagens and laminins, thereby indicating a role of dopamine in extra-cellular matrix integrity and transport of spermatids across the seminiferous epithelium. Gene ontology and enrichment map also highlighted cell/sperm motility to be significantly enriched. Therefore, genes involved in sperm motility functions were further validated by RT-qPCR. Seven genes (Akap4, Ccnyl1, Iqcf1, Klc3, Prss55, Tbc1d21, Tl18) were significantly up-regulated, whereas four genes (Dnah1, Dnah5, Clxn, Fsip2) were significantly down-regulated by bromocriptine treatment. The bromocriptine-stimulated reduction in seminiferous tubule cyclic AMP and associated changes in spermatid gene expression suggests that dopamine regulates both spermatogenesis and spermiogenesis within the seminiferous epithelium, and spermatozoa motility following spermiation, as essential processes for fertility.

The functions of triple gene block proteins and coat protein of apple stem pitting virus in viral cell-to-cell movement.

Apple stem pitting virus is a species in the genus Foveavirus in the family Betaflexiviridae. Apple stem pitting virus (ASPV) commonly infects apple and pear plants grown worldwide. In this study, by integrating bimolecular fluorescence complementation, split-ubiquitin-based membrane yeast two-hybrid, and Agrobacterium-mediated expression assays, the interaction relationships and the subcellular locations of ASPV proteins TGBp1-3 and CP in Nicotiana benthamiana leaf cells were determined. Proteins CP, TGBp1, TGBp2, and TGBp3 were self-interactable, and TGBp2 played a role in the formation of perinuclear viroplasm and enhanced the colocalization of TGBp3 with CP and TGBp1. We found that the plant microfilament and endoplasmic reticulum structures were involved in the production of TGBp3 and TGBp2 vesicles, and their disruption decreased the virus accumulation level in the systemic leaves. The TGBp3 motile vesicles functioned in delivering the viral ribonucleoprotein complexes to the plasma membrane. Two cysteine residues at sites 35 and 49 of the TGBp3 sorting signal were necessary for the diffusion of TGBp3-marked vesicles. Furthermore, our results revealed that TGBp1, TGBp2, and CP could increase plasmodesmal permeability and move to the adjacent cells. This study demonstrates an interaction network and a subcellular location map of four ASPV proteins and for the first time provides insight into the functions of these proteins in the movement of a foveavirus.

Integrated bioinformatics approaches to investigate alterations in transcriptomic profiles of monkeypox infected human cell line model.

BACKGROUND: The recent re-emergence of the monkeypox (mpox) epidemic in nonendemic regions has raised concerns regarding a potential global outbreak. The mpox virus (MPV) is a smallpox-like virus belonging to the genus Orthopoxvirus (family: Poxviridae). Although studies suggest that MPV infection suppresses the Toll-like receptor-3- and tumor necrosis factor-α-related signaling pathways, whether MPV regulates other immune-related pathways remains unclear. METHODS: In this study, two distinct temporal patterns were used for establishing an MPV-infected human immortal epithelial cancer cell line (HeLa). These two durations 2 and 12 h of incubation were selected to identify the coregulated genes and pathways affected by MPV infection. RESULTS: The use of the Gene Ontology framework, Kyoto Encyclopedia of Genes and Genome database, and MetaCore software yielded valuable insights. Specifically, various pathways were found to be enriched in HeLa cells infected with MPV for 2 and 12 h. These pathways included Notch, CD40, CD95, hypoxia-inducible factor-1-α, interleukin (IL)- 1, IL-6, phosphoinositide 3-kinase, nuclear factor-κB, mitogen-activated protein kinase, and oxidative stress-induced signalling pathways. Clusters and pathways of metabolism and viral replication cycles were significantly associated with the 2-hour infection group. This association was identified based on the regulation of genes such as HSPG2, RHPN2, MYL1, ASPHD2, CA9, VIPR1, SNX12, MGC2752, SLC25A1, PEX19, and AREG. Furthermore, clusters and pathways related to immunity and cell movement were found to be associated with the 12-hour infection group. This association was identified based on the regulation of genes such as C1orf21, C19orf48, HRK, IL8, GULP1, SCAND2, ATP5C1, FEZ1, SGSH, TACC2, CYP4X1, MMP1, CPB1, P2RY13, WDR27, PRPF4, and ENDOD1. CONCLUSIONS: This study can improve our understanding of the mechanisms underlying the pathophysiology and post-infection sequelae of mpox. Our findings provide valuable insights into the various modes of MPV infection.

Myosin 1e deficiency affects migration of 4T1 breast cancer cells.

Metastasis of breast cancer cells to distant tissue sites is responsible for the majority of deaths associated with breast cancer. Previously we have examined the role of class I myosin motor protein, myosin 1e (myo1e), in cancer metastasis using the Mouse Mammary Tumor Virus-Polyoma Middle T Antigen (MMTV-PyMT) mouse model. Mice deficient in myo1e formed tumors with a more differentiated phenotype relative to the wild-type mice and formed no detectable lung metastases. In the current study, we investigated how the absence of myo1e affects cell migration and invasion in vitro, using the highly invasive and migratory breast cancer cell line, 4T1. 4T1 cells deficient in myo1e exhibited an altered morphology and slower rates of migration in the wound-healing and transwell migration assays compared to the WT 4T1 cells. While integrin trafficking and Golgi reorientation did not appear to be altered upon myo1e loss, we observed lower rates of focal adhesion disassembly in myo1e-deficient cells, which could help explain the cell migration defect.

Properties of Plant Virus Protein Encoded by the 5'-Proximal Gene of Tetra-Cistron Movement Block.

To move from cell to cell through plasmodesmata, many plant viruses require the concerted action of two or more movement proteins (MPs) encoded by transport gene modules of virus genomes. A tetra-cistron movement block (TCMB) is a newly discovered transport module comprising four genes. TCMB encodes three proteins, which are similar to MPs of the transport module known as the "triple gene block", and a protein unrelated to known viral MPs and containing a double-stranded RNA (dsRNA)-binding domain similar to that found in a family of cell proteins, including AtDRB4 and AtHYL1. Here, the latter TCMB protein, named vDRB for virus dsRNA-binding protein, is shown to bind both dsRNA and single-stranded RNA in vitro. In a turnip crinkle virus-based assay, vDRB exhibits the properties of a viral suppressor of RNA silencing (VSR). In the context of potato virus X infection, vDRB significantly decreases the number and size of "dark green islands", regions of local antiviral silencing, supporting the VSR function of vDRB. Nevertheless, vDRB does not exhibit the VSR properties in non-viral transient expression assays. Taken together, the data presented here indicate that vDRB is an RNA-binding protein exhibiting VSR functions in the context of viral infection.

Long-term, real-time and label-free live cell image processing and analysis based on a combined algorithm of CellPose and watershed segmentation.

Developing a rapid and quantitative method to accurately evaluate the physiological abilities of living cells is critical for tumor control. Many experiments have been conducted in the field of biology in an attempt to measure the proliferation and movement abilities of cells, but existing methods cannot provide real-time and objective data for label-free cells. The quantitative imaging technique, including an automatic segmentation algorithm for individual label-free cells, has been a breakthrough in this regard. In this study, we develop a combined automatic image processing algorithm of CellPose and watershed segmentation for the long-term and real-time imaging of label-free cells. This method shows strong reliability in cell identification regardless of cell densities, allowing us to obtain accurate information about the number and proliferation ability of the target cells. Additionally, our results also suggest that this method is a reliable way to assess real-time data on drug cytotoxicity, cell morphology, and cell movement ability.

Reduced Brain Cortex Angiogenesis in the Offspring of the Preeclampsia-Like Syndrome.

BACKGROUND: Children from pregnancies affected by preeclampsia have an increased risk of cognitive and behavioral alterations via unknown pathophysiology. We tested the hypothesis that preeclampsia generated reduced brain cortex angiogenesis in the offspring. METHODS: The preeclampsia-like syndrome (PELS) mouse model was generated by administering the nitric oxide inhibitor NG-nitroarginine methyl ester hydrochloride. Confirmatory experiments were done using 2 additional PELS models. While in vitro analysis used mice and human brain endothelial cells exposed to serum of postnatal day 5 pups or umbilical plasma from preeclamptic pregnancies, respectively. RESULTS: We report significant reduction in the area occupied by blood vessels in the motor and somatosensory brain cortex of offspring (postnatal day 5) from PELS compared with uncomplicated control offspring. These data were confirmed using 2 additional PELS models. Furthermore, circulating levels of critical proangiogenic factors, VEGF (vascular endothelial growth factor), and PlGF (placental growth factor) were lower in postnatal day 5 PELS. Also we found lower VEGF receptor 2 (KDR [kinase insert domain-containing receptor]) levels in mice and human endothelial cells exposed to the serum of postnatal day 5 PELS or fetal plasma of preeclamptic pregnancies, respectively. These changes were associated with lower in vitro angiogenic capacity, diminished cell migration, larger F-actin filaments, lower number of filopodia, and lower protein levels of F-actin polymerization regulators in brain endothelial cells exposed to serum or fetal plasma of offspring from preeclampsia. CONCLUSIONS: Offspring from preeclampsia exhibited diminished brain cortex angiogenesis, associated with lower circulating VEGF/PlGF/KDR protein levels, impaired brain endothelial migration, and dysfunctional assembly of F-actin filaments. These alterations may predispose to structural and functional alterations in long-term brain development.

The role of TMEM26 in disrupting tight junctions and activating NF-κB signaling to promote epithelial-mesenchymal transition in esophageal squamous cell carcinoma.

OBJECTIVES: Metastasis is one of the biggest challenges in the management of Esophageal Squamous Cell Carcinoma (ESCC), of which molecular mechanisms remain elusive. The present study aimed to explore the roles and underlying mechanisms of Transmembrane protein 26 (TMEM26) in ESCC. METHOD: TMEM26 expressions in tumorous and adjacent tissues from patients with ESCC and in normal esophageal epithelial and ESCC cell lines were detected by immunostaining and western blotting, respectively. The Epithelial-Mesenchymal Transition (EMT), a critical process during metastasis, was investigated by wound healing and Transwell assays, and EMT-related proteins were examined after the TMEM26 alteration in ESCC cell lines. NF-κB signaling activation and Tight Junction (TJ) protein expression were analyzed by western blotting and immunofluorescence, respectively. In vivo verification was performed on the liver metastatic murine model. RESULTS: Compared with non-cancerous esophageal tissues and cells, the TMEM26 expression level was higher in ESCC samples and cell lines, where the plasma membrane localization of TMEM26 was observed. The EMT-related processes of ESCC cells were suppressed by RNAi depletion of TMEM26 but aggravated by TMEM26 overexpression. Mechanistically, TMEM26 promoted NF-κB signaling to accelerate EMT in ESCC cells. The plasma membrane presentation and assembly of TJ proteins were impaired by TMEM26. CONCLUSION: Overall, TMEM26 acts as a critical determinant for EMT in ESCC cells by disrupting TJ formation and promoting NF-κB signaling, which may be a potential therapeutic target for treating metastatic ESCC.