Integrated network analysis and experimental verification of the mechanisms employed by Compound Jixuecao Decoction to improve endoplasmic reticulum stress and apoptosis in chronic renal failure.

ETHNOPHARMACOLOGICAL RELEVANCE: Compound Jixuecao Decoction (CJD) is a traditional Chinese herbal medicine prescribed in China to treat chronic renal failure (CRF). Previous studies have shown that CJD affects cell apoptosis and proliferation. However, the mechanism of its renal protective action has not been characterized. AIM OF THE STUDY: To explore the mechanism(s) underlying the effect of CJD on endoplasmic reticulum stress (ERS) and apoptosis in the treatment of CRF using network pharmacology, molecular docking, molecular dynamics simulations, and in vivo studies. MATERIALS AND METHODS: The compounds comprising CJD were extracted from the Traditional Chinese Medicine Systems Pharmacology Database. A Swiss target prediction database and similarity integration approach were employed to identify potential targets of these components. The GeneCards and DisGeNET databases were used to identify targets associated with CRF, apoptosis, and ERS. The STRING database was employed to analyze the protein-protein interactions (PPIs) associated with drug-disease crossover. A chemical composition-shared target network was established, and critical pathways were identified through gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses. The Protein Data Bank database was used to search key proteins, while molecular docking and dynamics simulations were performed between the top four CJD active ingredients and proteins involved in apoptosis and ERS in CRF. Subsequent in vivo studies using a 5/6 nephrectomy rat model of CRF were performed to verify the findings. RESULTS: The 80 compounds identified in CJD yielded 875 target genes, of which 216 were potentially related to CRF. PPI network analysis revealed key targets via topology filtering. Enrichment analysis, molecular docking, and molecular dynamics simulation results suggested that CJD primarily targets mitofusin-2 (MFN2), B-cell lymphoma-2 (BCL2), BAX, protein kinase RNA-like ER kinase (PERK), and C/EBP homologous protein (CHOP) during CRF treatment. In vivo, CJD significantly increased the abundance of MFN2, BCL2, and significantly reduced the abundance of BAX, PERK, CHOP proteins in kidney tissues, indicating that CJD could improve apoptosis and ERS in CRF rats. CONCLUSIONS: This study provides evidence that CJD effectively delays CFR through modulation of the MFN2 and PERK-eIF2α-ATF4-CHOP signaling pathways.

Cataract-related variant R114C increases ßA3-crystallin susceptibility to environmental stresses by disrupting the protein senior structure.

Congenital cataract is a major cause of childhood blindness worldwide, with crystallin mutations accounting for over 40 % of gene-mutation-related cases. Our research focused on a novel R114C mutation in a Chinese family, resulting in bilateral coronary cataract with blue punctate opacity. Spectroscopic experiments revealed that ßA3-R114C significantly altered the senior structure, exhibiting aggregation, and reduced solubility at physiological temperature. The mutant also displayed decreased resistance and stability under environmental stresses such as UV irradiation, oxidative stress, and heat. Further, cellular models confirmed its heightened sensitivity to environmental stresses. These data suggest that the R114C mutation impairs the hydrogen bond network and structural stability of ßA3-crystallin, particularly at the boundary of the second Greek-key motif. This study revealed the pathological mechanism of ßA3-R114C and may help in the development of potential treatment strategies for related cataracts.

Patronin regulates presynaptic microtubule organization and neuromuscular junction development in Drosophila.

Synapses are fundamental components of the animal nervous system. Synaptic cytoskeleton is essential for maintaining proper neuronal development and wiring. Perturbations in neuronal microtubules (MTs) are correlated with numerous neuropsychiatric disorders. Despite discovering multiple synaptic MT regulators, the importance of MT stability, and particularly the polarity of MT in synaptic function, is still under investigation. Here, we identify Patronin, an MT minus-end-binding protein, for its essential role in presynaptic regulation of MT organization and neuromuscular junction (NMJ) development. Analyses indicate that Patronin regulates synaptic development independent of Klp10A. Subsequent research elucidates that it is short stop (Shot), a member of the Spectraplakin family of large cytoskeletal linker molecules, works synergistically with Patronin to govern NMJ development. We further raise the possibility that normal synaptic MT polarity contributes to proper NMJ morphology. Overall, this study demonstrates an unprecedented role of Patronin, and a potential involvement of MT polarity in synaptic development.

Network pharmacology and in vitro experimental verification to reveal the mechanism of Astragaloside IV against kidney ischemia-reperfusion injury.

Ischemic acute kidney injury (AKI) is a prevalent disorder among hospitalized patients worldwide. Astragaloside IV (AS-IV) has been shown to protect against ischemic AKI. However, the specific effects and mechanisms of AS-IV on alleviating kidney ischemia-reperfusion (I/R) injury remain unclear. The objective of this research was to elucidate the regulatory targets and mechanisms through which AS-IV protects kidney I/R injury. A combination of network pharmacology, molecular docking, molecular dynamics (MD) simulation, pharmacodynamic study and Western blot were employed to explore the underlying mechanisms. Network pharmacology revealed that ferroptosis was a potential mechanism of AS-IV against kidney I/R injury. Molecular docking and MD simulations demonstrated strong binding affinity between the GPX4/SLC7A11 and AS-IV. The experimental verification demonstrated that AS-IV improved cell proliferation, decreased the level of ROS and Fe2+, and increased the expressions of GPX4 and SLC7A11 as same as Ferrostatin-1 in OGD/R-injured HUVECs. In conclusion, AS-IV had a significant inhibition on ferroptosis in kidney I/R injury, providing a new perspective for drug development on kidney I/R injury. Definitely, further exploration in vivo is necessary to fully understand whether AS-IV alleviates kidney I/R injury through inhibiting endothelial ferroptosis.

SCFRMF mediates degradation of the meiosis-specific recombinase DMC1.

Meiotic recombination requires the specific RecA homolog DMC1 recombinase to stabilize strand exchange intermediates in most eukaryotes. Normal DMC1 levels are crucial for its function, yet the regulatory mechanisms of DMC1 stability are unknown in any organism. Here, we show that the degradation of Arabidopsis DMC1 by the 26S proteasome depends on F-box proteins RMF1/2-mediated ubiquitination. Furthermore, RMF1/2 interact with the Skp1 ortholog ASK1 to form the ubiquitin ligase complex SCFRMF1/2. Genetic analyses demonstrate that RMF1/2, ASK1 and DMC1 act in the same pathway downstream of SPO11-1 dependent meiotic DNA double strand break formation and that the proper removal of DMC1 is crucial for meiotic crossover formation. Moreover, six DMC1 lysine residues were identified as important for its ubiquitination but not its interaction with RMF1/2. Our results reveal mechanistic insights into how the stability of a key meiotic recombinase that is broadly conserved in eukaryotes is regulated.

Insight into Pathogenic Mechanism Underlying the Hereditary Cataract Caused by ßB2-G149V Mutation.

Congenital cataracts account for approximately 5-20% of childhood blindness worldwide and 22-30% of childhood blindness in developing countries. Genetic disorders are the primary cause of congenital cataracts. In this work, we investigated the underlying molecular mechanism of G149V point missense mutation in ßB2-crystallin, which was first identified in a three-generation Chinese family with two affected members diagnosed with congenital cataracts. Spectroscopic experiments were performed to determine the structural differences between the wild type (WT) and the G149V mutant of ßB2-crystallin. The results showed that the G149V mutation significantly changed the secondary and tertiary structure of ßB2-crystallin. The polarity of the tryptophan microenvironment and the hydrophobicity of the mutant protein increased. The G149V mutation made the protein structure loose and the interaction between oligomers was reduced, which decreased the stability of the protein. Furthermore, we compared ßB2-crystallin WT and the G149V mutant with their biophysical properties under environmental stress. We found that the G149V mutation makes ßB2-crystallin more sensitive to environmental stresses (oxidative stress, UV irradiation, and heat shock) and more likely to aggregate and form precipitation. These features might be important to the pathogenesis of ßB2-crystallin G149V mutant related to congenital cataracts.

Spatiotemporal formation of the large vacuole regulated by the BIN2-VLG module is required for female gametophyte development in Arabidopsis.

In Arabidopsis thaliana, female gametophyte (FG) development is accompanied by the formation and expansion of the large vacuole in the FG; this is essential for FG expansion, nuclear polar localization, and cell fate determination. Arabidopsis VACUOLELESS GAMETOPHYTES (VLG) facilitates vesicular fusion to form large vacuole in the FG, but the regulation of VLG remains largely unknown. Here, we found that gain-of-function mutation of BRASSINOSTEROID INSENSITIVE2 (BIN2) (bin2-1) increases VLG abundance to induce the vacuole formation at stage FG1, and leads to abortion of FG. Loss-of-function mutation of BIN2 and its homologs (bin2-3 bil1 bil2) reduced VLG abundance and mimicked vlg/VLG phenotypes. Knocking down VLG in bin2-1 decreased the ratio of aberrant vacuole formation at stage FG1, whereas FG1-specific overexpression of VLG mimicked the bin2-1 phenotype. VLG partially rescued the bin2-3 bil1 bil2 phenotype, demonstrating that VLG acts downstream of BIN2. Mutation of VLG residues that are phosphorylated by BIN2 altered VLG stability and a phosphorylation mimic of VLG causes similar defects as did bin2-1. Therefore, BIN2 may function by interacting with and phosphorylating VLG in the FG to enhance its stability and abundance, thus facilitating vacuole formation. Our findings provide mechanistic insight into how the BIN2-VLG module regulates the spatiotemporal formation of the large vacuole in FG development.

Establishment of a novel axon pruning model of Drosophila motor neuron.

Developmental neuronal pruning is a process by which neurons selectively remove excessive or unnecessary neurite without causing neuronal death. Importantly, this process is widely used for the refinement of neural circuits in both vertebrates and invertebrates, and may also contribute to the pathogenesis of neuropsychiatric disorders, such as autism and schizophrenia. In the peripheral nervous system (PNS), class IV dendritic arborization (da) sensory neurons of Drosophila, selectively remove the dendrites without losing their somas and axons, while the dendrites and axons of mushroom body (MB) γ neuron in the central nervous system (CNS) are eliminated by localized fragmentation during metamorphosis. Alternatively, dendrite pruning of ddaC neurons is usually investigated via live-cell imaging, while dissection and fixation are currently used for evaluating MB γ neuron axon pruning. Thus, an excellent model system to assess axon specific pruning directly via live-cell imaging remains elusive. Here, we report that the Drosophila motor neuron offers a unique advantage for studying axon pruning. Interestingly, we uncover that long-range projecting axon bundle from soma at ventral nerve cord (VNC), undergoes degeneration rather than retraction during metamorphosis. Strikingly, the pruning process of the motor axon bundle is straightforward to investigate via live imaging and it occurs approximately at 22 h after pupal formation (APF), when axon bundles are completely cleared. Consistently, the classical axon pruning regulators in the Drosophila MB γ neuron, including TGF-ß signaling, ecdysone signaling, JNK signaling, and the ubiquitin-proteasome system are also involved in governing motor axon pruning. Finally, our findings establish an unprecedented axon pruning mode that will serve to systematically screen and identify undiscovered axon pruning regulators. This article has an associated First Person interview with the first author of the paper.

A novel cataract-causing mutation Ile82Met of γA crystallin trends to aggregate with unfolding intermediate.

Cataract is the most common pathogenic ophthalmic disease leading to blindness in children worldwide. Genetic disorder is the leading cause of congenital cataract, among which crystallin mutations have a high incidence. There are few reports on γA-crystallin, one critical member of crystallin superfamilies. In this study, we identified a novel pathogenic mutation (Ile82Met) in γA-crystallin from a three-generation Chinese family with cataract, and investigated the potential molecular mechanism in detail. To elucidate the pathogenic mechanism of I82M mutant, spectroscopic and solubility experiments were performed to determine the difference between the purified γA-crystallin wild type (WT) and I82M mutant under both physiological conditions and environmental stresses (UV irradiation, thermal denaturation or chemical denaturation). The I82M mutant did not affect the secondary/tertiary structure of monomeric γA-crystallin under physiological status, but decreased protein stability and increased aggregatory potency under the stressful treatment. Surprisingly, the chemical denaturation caused I82M to switch from the two-state unfolding of γA-crystallin to three-state unfolding involving an unfolding intermediate. This study expands the genetic variation map of cataract, and provides novel insights into the pathomechanism, in particular, filling in a gap in the understanding of γA-crystallin mutants causing cataract.

Cataract-Causing S93R Mutant Destabilized Structural Conformation of ßB1 Crystallin Linking With Aggregates Formation and Cellular Viability.

Cataract, opacity of the eye lens, is the leading cause of visual impairment worldwide. The crucial pathogenic factors that cause cataract are misfolding and aggregation of crystallin protein. ßB1-crystallin, which is the most abundant water-soluble protein in mammalian lens, is essential for lens transparency. A previous study identified the missense mutation ßB1-S93R being responsible for congenital cataract. However, the exact pathogenic mechanism causing cataract remains unclear. The S93 residue, which is located at the first Greek-key motif of ßB1-crystallin, is highly conserved, and its substitution to Arginine severely impaired hydrogen bonds and structural conformation, which were evaluated via Molecular Dynamic Simulation. The ßB1-S93R was also found to be prone to aggregation in both human cell lines and Escherichia coli. Then, we isolated the ßB1-S93R variant from inclusion bodies by protein renaturation. The ßB1-S93R mutation exposed more hydrophobic residues, and the looser structural mutation was prone to aggregation. Furthermore, the S93R mutation reduced the structural stability of ßB1-crystallin when incubated at physiological temperature and made it more sensitive to environmental stress, such as UV irradiation or oxidative stress. We also constructed a ßB1-S93R cellular model and discovered that ßB1-S93R was more sensitive to environmental stress, causing not only aggregate formation but also cellular apoptosis and impaired cellular viability. All of the results indicated that lower solubility and structural stability, sensitivity to environmental stress, vulnerability to aggregation, and impaired cellular viability of ßB1-S93R might be involved in cataract development.

An F-box protein ACOZ1 functions in crossover formation by ensuring proper chromosome compaction during maize meiosis.

Meiosis is an essential reproductive process to create new genetic variation. During early meiosis, higher order chromosome organization creates a platform for meiotic processes to ensure the accuracy of recombination and chromosome segregation. However, little is known about the regulatory mechanisms underlying dynamic chromosome organization in plant meiosis. Here, we describe abnormal chromosome organization in zygotene1 (ACOZ1), which encodes a canonical F-box protein in maize. In acoz1 mutant meiocytes, chromosomes maintain a leptotene-like state and never compact to a zygotene-like configuration. Telomere bouquet formation and homologous pairing are also distorted and installation of synaptonemal complex ZYP1 protein is slightly defective. Loading of early recombination proteins RAD51 and DMC1 is unaffected, indicating that ACOZ1 is not required for double strand break formation or repair. However, crossover formation is severely disturbed. The ACOZ1 protein localizes on the boundary of chromatin, rather directly to chromosomes. Furthermore, we identified that ACOZ1 interacts with SKP1 through its C-terminus, revealing that it acts as a subunit of the SCF E3 ubiquitin/SUMO ligase complex. Overall, our results suggest that ACOZ1 functions independently from the core meiotic recombination pathway to influence crossover formation by controlling chromosome compaction during maize meiosis.

Cataract-causing mutation R48C increases γA-crystallin susceptibility to oxidative stress and ultraviolet radiation.

Among all congenital cataracts caused by genetic mutations, approximately half are caused by a mutation in crystallin genes, and accounts the leading cause of blindness in children globally. In this study, we investigated the underlying molecular mechanism of R48C mutation (c.142C > T; p.[Arg48Cys]) of γA-crystallin in a Mexican-Mestizo descent family causing congenital cataracts. We purified γA-crystallin wild-type (WT) and R48C mutant and compared their structural characteristics and biophysical properties by Spectroscopic experiments and environmental stress (oxidative stress, ultraviolet irradiation, pH disorders, thermal shock, or chemical denaturation). The R48C mutant did not affect the secondary and tertiary structure of monomer γA-crystallin, nor did it affect its stability to heat shock and chemicals. However, the R48C mutant destroys the oxidative stability of γA-crystallin, which makes the protein more prone to aggregation and precipitation under oxidative conditions. These might be the pathogenesis of γA-crystallin R48C mutant related to congenital cataract and help to develop anti-cataract strategies from the perspective of γA-crystallin.

Congenital cataract-causing mutation ßB1-L116P is prone to amyloid fibrils aggregation and protease degradation with low structural stability.

Congenital cataract, a common disease with lens opacification, causes blindness in the newborn worldwide and is mainly caused by abnormal aggregation of crystallin. As the main structural protein in the mammalian lens, ßB1-crystallin has an important role in the maintenance of lens transparency. Recently, the L116P mutation in ßB1-CRY was found in a Chinese family with congenital nuclear cataracts, while its underlying pathogenic mechanism remains unclear. In the current study, the ßB1 wild-type protein was purified, and the mutated form, ßB1-L116P, was examined for examining the effect on structural stability and susceptibility against environmental stresses. Our results reveal low solubility and structural stability of ßB1-L116P at physiological temperature, which markedly impaired the protein structure and the oligomerization of ßB1-crystallin. Under guanidine hydrochloride-induced denaturing conditions, ßB1-L116P mutation perturbed the protein unfolding process, making it prone to amyloid fibrils aggregation. More importantly, the L116P mutation increased susceptibility of ßB1-crystallin against UV radiation. ßB1-L116P overexpression led to the formation of more serious intracellular aggresomes under UV radiation or oxidative stress. Furthermore, the ßB1-L116P mutation increased the sensitivity to the proteolysis process. These results indicate that the low structural stability, susceptibility to amyloid fibrils aggregation, and protease degradation of ßB1-L116P may contribute to cataract development and associated symptoms.

Fast Approximation of Coherence for Second-Order Noisy Consensus Networks.

It has been recently established that for second-order consensus dynamics with additive noise, the performance measures, including the vertex coherence and network coherence defined, respectively, as the steady-state variance of the deviation of each vertex state from the average and the average steady-state variance of the system, are closely related to the biharmonic distances. However, direct computation of biharmonic distances is computationally infeasible for huge networks with millions of vertices. In this article, leveraging the implicit fact that both vertex and network coherence can be expressed in terms of the diagonal entries of pseudoinverse L2† of the square of graph Laplacian, we develop a nearly linear-time algorithm to approximate all diagonal entries of L2† , which has a theoretically guaranteed error for each diagonal entry. The key ingredient of our approximation algorithm is an integration of the Johnson-Lindenstrauss lemma and Laplacian solvers. Extensive numerical experiments on real-life and model networks are presented, which indicate that our approximation algorithm is both efficient and accurate and is scalable to large-scale networks with millions of vertices.

Coherence Scaling of Noisy Second-Order Scale-Free Consensus Networks.

A striking discovery in the field of network science is that the majority of real networked systems have some universal structural properties. In general, they are simultaneously sparse, scale-free, small-world, and loopy. In this article, we investigate the second-order consensus of dynamic networks with such universal structures subject to white noise at vertices. We focus on the network coherence HSO characterized in terms of the H2 -norm of the vertex systems, which measures the mean deviation of vertex states from their average value. We first study numerically the coherence of some representative real-world networks. We find that their coherence HSO scales sublinearly with the vertex number N . We then study analytically HSO for a class of iteratively growing networks-pseudofractal scale-free webs (PSFWs), and obtain an exact solution to HSO, which also increases sublinearly in N , with an exponent much smaller than 1. To explain the reasons for this sublinear behavior, we finally study HSO for Sierpinski gaskets, for which HSO grows superlinearly in N , with a power exponent much larger than 1. Sierpinski gaskets have the same number of vertices and edges as the PSFWs but do not display the scale-free and small-world properties. We thus conclude that the scale-free, small-world, and loopy topologies are jointly responsible for the observed sublinear scaling of HSO.

Cataract-causing G91del mutant destabilised ßA3 heteromers formation linking with structural stability and cellular viability.

BACKGROUND/AIMS: Congenital cataracts, which are genetically heterogeneous eye disorders, result in visual loss in childhood around the world. CRYBA1/BA3 serves as an abundant structural protein in the lens, and forms homomers and heteromers to maintain lens transparency. In previous study, we identified a common cataract-causing mutation, ßA3-glycine at codon 91 (G91del) (c.271-273delGAG), which deleted a highly conserved G91del and led to perinuclear zonular cataract. In this study, we aimed to explore the underlying pathogenic mechanism of G91del mutation. METHODS: Protein purification, size-exclusion chromatography, spectroscopy and molecular dynamics simulation assays were used to investigate the effects on the heteromers formation and the protein structural properties of ßA3-crystallin caused by G91del mutation. Intracellular ßA3-G91del overexpression, MTT (3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide) and cell apoptosis were used to investigate the cellular functions of ßA3-G91del. RESULTS: ßA3-crystallin and ßB2-crystallin could form heteromers, which have much more stable structures than ßA3 homomers. Interestingly, ßA3/ßB2 heteromers improved their resistance against the thermal stress and the guanidine hydrochloride treatment. However, the pathogenic mutation ßA3-G91del destroyed the interaction with ßB2, and thereby decreased its structural stability as well as the resistance of thermal or chemical stress. What's more, the ßA3-G91del mutation induced cell apoptosis and escaped from the protection of ßB2-crystallin. CONCLUSIONS: ßA3/ßB2 heteromers play an indispensable role in maintaining lens transparency, while the ßA3-G91del mutation destabilises heteromers formation with ßB2-crystallin, impairs cellular viability and induces cellular apoptosis. These all might contribute to cataract development.

The cohesin loader SCC2 contains a PHD finger that is required for meiosis in land plants.

Cohesin, a multisubunit protein complex, is required for holding sister chromatids together during mitosis and meiosis. The recruitment of cohesin by the sister chromatid cohesion 2/4 (SCC2/4) complex has been extensively studied in Saccharomyces cerevisiae mitosis, but its role in mitosis and meiosis remains poorly understood in multicellular organisms, because complete loss-of-function of either gene causes embryonic lethality. Here, we identified a weak allele of Atscc2 (Atscc2-5) that has only minor defects in vegetative development but exhibits a significant reduction in fertility. Cytological analyses of Atscc2-5 reveal multiple meiotic phenotypes including defects in chromosomal axis formation, meiosis-specific cohesin loading, homolog pairing and synapsis, and AtSPO11-1-dependent double strand break repair. Surprisingly, even though AtSCC2 interacts with AtSCC4 in vitro and in vivo, meiosis-specific knockdown of AtSCC4 expression does not cause any meiotic defect, suggesting that the SCC2-SCC4 complex has divergent roles in mitosis and meiosis. SCC2 homologs from land plants have a unique plant homeodomain (PHD) motif not found in other species. We show that the AtSCC2 PHD domain can bind to the N terminus of histones and is required for meiosis but not mitosis. Taken together, our results provide evidence that unlike SCC2 in other organisms, SCC2 requires a functional PHD domain during meiosis in land plants.

Protein Phosphatase 2A B'α and B'ß Protect Centromeric Cohesion during Meiosis I.

During meiosis, the stepwise release of sister chromatid cohesion is crucial for the equal distribution of genetic material to daughter cells, enabling generation of fertile gametophytes. However, the molecular mechanism that protects centromeric cohesion from release at meiosis I is unclear in Arabidopsis (Arabidopsis thaliana). Here, we report that the protein phosphatase 2A regulatory subunits B'α and B'ß participate in the control of sister chromatid separation. The double mutant b'αß exhibited severe male and female sterility, caused by the lack of a nucleus or presence of an abnormal nucleus in mature microspores and embryo sacs. 4',6-Diamidino-2-phenylindole staining revealed unequal amounts of DNA in the mononuclear microspores. Transverse sections of the anthers revealed unevenly sized tetrads with or without a nucleus, suggesting a defect in meiocyte meiosis. An analysis of chromosome spreads showed that the sister chromatids separated prematurely at anaphase I in b'αß Immunoblotting showed that AtRECOMBINATION DEFECTIVE8 (AtREC8), a key member of the cohesin complex, was hyperphosphorylated in b'αß anthers and pistils during meiosis but hypophosphorylated in the wild type. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation assays showed that B'α and B'ß interact specifically with AtREC8, AtSHUGOSHIN1 (AtSGO1), AtSGO2, and PATRONUS1. Given that B'α was reported to localize to the centromere in meiotic cells, we propose that protein phosphatase 2A B'α and B'ß are recruited by AtSGO1/2 and PATRONUS1 to dephosphorylate AtREC8 at the site of centromere cohesion to shield it from cleavage until anaphase II, contributing to the balanced separation of sister chromatids at meiosis.

Porphyromonas gingivalis-Induced MIF Regulates Intercellular Adhesion Molecule-1 Expression in EA.hy926 Cells and Monocyte-Endothelial Cell Adhesion Through the Receptors CD74 and CXCR4.

Porphyromonas gingivalis (P. gingivalis) is an important pathogen that contributes to periodontal disease and causes infections that promote the progression of atherosclerosis. Our previous studies showed that macrophage migration inhibitory factor (MIF) facilitates monocyte adhesion to endothelial cells by regulating the expression of intercellular adhesion molecule-1 (ICAM-1) in P. gingivalis-infected endothelial cells. However, the detailed pathological role of MIF has yet to be elucidated in this context. To explore the functional receptor(s) of MIF that underlie its participation in the pathogenesis of atherosclerosis, we investigated the expression of the chemokine receptors CD74 and CXCR4 in endothelial cells, both of which were shown to be involved in the adhesion of monocytes to endothelial cells pretreated with P. gingivalis. Furthermore, the formation of a MIF, CD74, and CXCR4 ligand-receptor complex was revealed by our immunofluorescence staining and coimmunoprecipitation results. By interacting with the CD74/CXCR4 receptor complex, MIF may act as a crucial regulator of monocyte-endothelial cell adhesion and promote the atherosclerotic plaque formation induced by P. gingivalis.

Porphyromonas gingivalis ATCC 33277 promotes intercellular adhesion molecule-1 expression in endothelial cells and monocyte-endothelial cell adhesion through macrophage migration inhibitory factor.

BACKGROUND: Porphyromonas gingivalis (P. gingivalis), one of the main pathogenic bacteria involved in periodontitis, induces the expression of intercellular adhesion molecule - 1 (ICAM-1) and monocyte-endothelial cell adhesion. This effect plays a pivotal role in atherosclerosis development. Macrophage migration inhibitory factor (MIF) is a multifunctional cytokine and critically affects atherosclerosis pathogenesis. In this study, we tested the involvement of MIF in the P. gingivalis ATCC 33277-enhanced adhesive properties of endothelial cells. RESULTS: Endothelial MIF expression was enhanced by P. gingivalis ATCC 33277 infection. The MIF inhibitor ISO-1 inhibited ICAM-1 production in endothelial cells, and monocyte-endothelial cell adhesion was induced by P. gingivalis ATCC 33277 infection. However, the addition of exogenous human recombinant MIF to P. gingivalis ATCC 33277-infected endothelial cells facilitated monocyte recruitment by promoting ICAM-1 expression in endothelial cells. CONCLUSIONS: These experiments revealed that MIF in endothelial cells participates in the pro-atherosclerotic lesion formation caused by P. gingivalis ATCC 33277 infection. Our novel findings identify a more detailed pathological role of P. gingivalis ATCC 33277 in atherosclerosis.