Cell Surface Adhesion Proteins Research Articles

Mechanism of Porphyra Yezoensis Polysaccharides in Inhibiting Hyperoxalate-Induced Renal Injury and Crystal Deposition.

Oxidative damage to the kidneys is a primary factor in the occurrence of kidney stones. This study explores the inhibitory effect of Porphyra yezoensis polysaccharides (PYP) on oxalate-induced renal injury by detecting levels of oxidative damage, expression of adhesion molecules, and damage to intracellular organelles and revealed the molecular mechanism by molecular biology methods. Additionally, we validated the role of PYP in vivo using a crystallization model of hyperoxalate-induced rats. PYP effectively scavenged the overproduction of reactive oxygen species (ROS) in HK-2 cells, inhibited the adhesion of calcium oxalate (CaOx) crystals on the cell surface, unblocked the cell cycle, restored the depolarization of the mitochondrial membrane potential, and inhibited cell death. PYP upregulated the expression of antioxidant proteins, including Nrf2, HO-1, SOD, and CAT, while decreasing the expression of Keap-1, thereby activating the Keap1/Nrf2 signaling pathway. PYP inhibited CaOx deposition in renal tubules in the rat crystallization model, significantly reduced high oxalate-induced renal injury, decreased the levels of the cell surface adhesion proteins, improved renal function in rats, and ultimately inhibited the formation of kidney stones. Therefore, PYP, which has crystallization inhibition and antioxidant properties, may be a therapeutic option for the treatment of kidney stones.

CD44 Expression in Oral Lichen Planus and Related Lesions-An Immunohistochemical Study.

Cluster of differentiation 44 (CD44) is a cell surface adhesion protein involved in the progression and metastasis of oral squamous cell carcinoma. The current study aims to evaluate the expression of CD44 in oral lichen planus and related lesions and thereby assess the relative risk of malignant transformation of these lesions. Formalin-fixed paraffin-embedded tissue blocks of 10 oral lichen planus (Group 1), 10 oral lichenoid lesions (Group 2), 8 with oral lichen planus with dysplasia (Group 3), and 5 with lichenoid dysplasia (Group 4) were included in the study. Immunostaining was done for the tissue sections using CD44 mouse monoclonal antibody. Staining density, staining intensity, and immunoreactive scores of CD44 were evaluated in all four groups. Statistical analysis was done by Statistical Package for the Social Sciences® software and the Kruskal-Wallis test was used. CD44 staining pattern of lichenoid dysplasia and lichen planus with dysplasia changed from membranous to cytoplasmic. The membranous CD44 immunoreactivity was mild with a score of 2.25 for Group 3 and 1.6 for Group 4 whereas moderate for other groups with a P-value of 0.009. The cytoplasmic immunoreactivity was significantly high in Group 3 (5.3 ± 2.6) followed by Group 4 (3.2 ± 1.2), Group 2 (1 ± 1.8), and Group 1 (0.7 ± 1.3) with a P-value of 0.001. The CD44 membranous immunoreactivity scores were low while the cytoplasmic immunoreactivity was high in oral lichen planus with dysplasia and oral lichenoid dysplasia when compared to oral lichen planus and oral lichenoid lesions. CD44 immunostaining pattern can help in assessing the malignant transformation of oral lichen planus or lichenoid lesions.

Cerium oxide-based nanozyme suppresses kidney calcium oxalate crystal depositions via reversing hyperoxaluria-induced oxidative stress damage

Oxidative stress damage to renal epithelial cells is the main pathological factor of calcium oxalate calculi formation. The development of medicine that could alleviate oxidative damage has become the key to the prevention and treatment of urolithiasis. Herein, porous nanorods CeO2 nanoparticles (CNPs) were selected from CeO2 with different morphologies as an antioxidant reagent to suppress kidney calcium oxalate crystal depositions with excellent oxidation resistance due to its larger specific surface area. The reversible transformation from Ce3+ to Ce4+ could catalyze the decomposition of excess free radicals and act as a biological antioxidant enzyme basing on its strong ability to scavenge free radicals. The protection capability of CNPS against oxalate-induced damage and the effect of CNPS on calcium oxalate crystallization were studied. CNPS could effectively reduce reactive oxygen species production, restore mitochondrial membrane potential polarity, recover cell cycle progression, reduce cell death, and inhibit the formation of calcium oxalate crystals on the cell surface in vitro. The results of high-throughput sequencing of mRNA showed that CNPs could protect renal epithelial cells from oxidative stress damage caused by high oxalate by suppressing the expression gene of cell surface adhesion proteins. In addition, CNPS can significantly reduce the pathological damage of renal tubules and inhibit the deposition of calcium oxalate crystals in rat kidneys while having no significant side effect on other organs and physiological indicators in vivo. Our results provide a new strategy for CNPS as a potential for clinical prevention of crystalline kidney injury and crystal deposition.

Control of CDH1/E-Cadherin Gene Expression and Release of a Soluble Form of E-Cadherin in SARS-CoV-2 Infected Caco-2 Intestinal Cells: Physiopathological Consequences for the Intestinal Forms of COVID-19.

COVID-19 is the biggest pandemic the world has seen this century. Alongside the respiratory damage observed in patients with severe forms of the disease, gastrointestinal symptoms have been frequently reported. These symptoms (e.g., diarrhoea), sometimes precede the development of respiratory tract illnesses, as if the digestive tract was a major target during early SARS-CoV-2 dissemination. We hypothesize that in patients carrying intestinal SARS-CoV-2, the virus may trigger epithelial barrier damage through the disruption of E-cadherin (E-cad) adherens junctions, thereby contributing to the overall gastrointestinal symptoms of COVID-19. Here, we use an intestinal Caco-2 cell line of human origin which expresses the viral receptor/co-receptor as well as the membrane anchored cell surface adhesion protein E-cad to investigate the expression of E-cad after exposure to SARS-CoV-2. We found that the expression of CDH1/E-cad mRNA was significantly lower in cells infected with SARS-CoV-2 at 24 hours post-infection, compared to virus-free Caco-2 cells. The viral receptor ACE2 mRNA expression was specifically down-regulated in SARS-CoV-2-infected Caco-2 cells, while it remained stable in HCoV-OC43-infected Caco-2 cells, a virus which uses HLA class I instead of ACE2 to enter cells. It is worth noting that SARS-CoV-2 induces lower transcription of TMPRSS2 (involved in viral entry) and higher expression of B0AT1 mRNA (that encodes a protein known to co-express with ACE2 on intestinal cells). At 48 hours post-exposure to the virus, we also detected a small but significant increase of soluble E-cad protein (sE-cad) in the culture supernatant of SARS-CoV-2-infected Caco-2 cells. The increase of sE-cad release was also found in the intestinal HT29 cell line when infected by SARS-CoV-2. Beside the dysregulation of E-cad, SARS-CoV-2 infection of Caco-2 cells also leads to the dysregulation of other cell adhesion proteins (occludin, JAMA-A, zonulin, connexin-43 and PECAM-1). Taken together, these results shed light on the fact that infection of Caco-2 cells with SARS-CoV-2 affects tight-, adherens-, and gap-junctions. Moreover, intestinal tissues damage was associated to the intranasal SARS-CoV-2 infection in human ACE2 transgenic mice.

Potent suppression of neuroendocrine tumors and gastrointestinal cancers by CDH17CAR T cells without toxicity to normal tissues.

Gastrointestinal cancers (GICs) and neuroendocrine tumors (NETs) are often refractory to therapy after metastasis. Adoptive cell therapy using chimeric antigen receptor (CAR) T cells, though remarkably efficacious for treating leukemia, is yet to be developed for solid tumors such as GICs and NETs. Here we isolated a llama-derived nanobody, VHH1, and found that it bound cell surface adhesion protein CDH17 upregulated in GICs and NETs. VHH1-CAR T cells (CDH17CARTs) killed both human and mouse tumor cells in a CDH17-dependent manner. CDH17CARTs eradicated CDH17-expressing NETs and gastric, pancreatic and colorectal cancers in either tumor xenograft or autochthonous mouse models. Notably, CDH17CARTs do not attack normal intestinal epithelial cells, which also express CDH17, to cause toxicity, likely because CDH17 is localized only at the tight junction between normal intestinal epithelial cells. Thus, CDH17 represents a class of previously unappreciated tumor-associated antigens that is 'masked' in healthy tissues from attack by CAR T cells for developing safer cancer immunotherapy.

Dynamics of the Actin Cytoskeleton at Adhesion Complexes.

Simple SummaryAll cells in the human body experience force. To withstand these forces, cells rearrange and reinforce their cytoskeletons. In this review, we describe the structure of the actin cytoskeleton and its anchorage at points where cells adhere to the extracellular matrix and one another. We examine the current understanding for how the actin cytoskeleton is remodeled by force. This examination reveals that the response of cells to force is complex and highly coordinated.The shape of cells is altered to allow cells to adapt to their changing environments, including responding to internally generated and externally applied force. Force is sensed by cell surface adhesion proteins that are enriched in sites where cells bind to the extracellular matrix (focal adhesions) and neighboring cells (cell–cell or adherens junctions). Receptors at these adhesion sites stimulate intracellular signal transduction cascades that culminate in dramatic changes in the actin cytoskeleton. New actin filaments form, and/or new and existing filaments can be cleaved, branched, or bundled. Here, we discuss the actin cytoskeleton and its functions. We will examine the current understanding for how the actin cytoskeleton is tethered to adhesion sites. Finally, we will highlight recent studies describing how the actin cytoskeleton at these adhesion sites is remodeled in response to force.

Staphylococcus epidermidis Has Growth Phase Dependent Affinity for Fibrinogen and Resulting Fibrin Clot Elasticity.

Bacterial infection and thrombosis are highly correlated, especially in patients with indwelling medical devices. Coagulase-negative staphylococci, typified by Staphylococcus epidermidis, are a common cause of medical device infections owing to their biofilm forming capacity which provides protection from antibiotics and host immune response. Attention has been drawn to the interaction between S. epidermidis and host proteins, specifically fibrinogen. However, little is known regarding the impact of the transition from planktonic to biofilm forming phenotype on this interaction. Here we investigate the growth phase dependence of bacteria-fibrinogen interaction and the resulting effect on fibrin clot formation, structure, and mechanics. Flow cytometry demonstrated growth phase dependent affinity for fibrinogen. To mimic intravascular device seeding, we quantified the adhesion of S. epidermidis to a fibrinogen coated surface under continuous flow conditions in vitro. The bacterial deposition rate onto fibrinogen was significantly greater for stationary (5,360 ± 1,776 cells/cm2s) versus exponential phase (2,212 ± 264, cells/cm2 s). Furthermore, the expression of sdrG–a cell surface adhesion protein with specificity for fibrinogen–was upregulated ∼twofold in the stationary versus the exponential phase. Rheometry and confocal microscopy demonstrated that stationary phase S. epidermidis slows clot formation and generates a more heterogeneous fibrin network structure with greater elasticity (G′ = 5.7 ± 1.0 Pa) compared to sterile fibrinogen (G′ = l.5 ± 0.2 Pa), while exponential phase cells had little effect. This work contributes to the current understanding of the growth phase dependent regulation of bacterial virulence factors and the correlation between bacterial infection and thrombosis.

In Vitro Acute Ischemic Injury Altered Human Brain Endothelial MMP‐9 Activity and Associated Cell Surface Adhesion Protein Expression in Part by S1PR Type 1 Activation

Cerebrovascular endothelial cells are among the first cells to be impacted by acute ischemic stroke (AIS) which poses them as important therapeutic targets pivotal to the maintenance of blood brain barrier (BBB) integrity. Following AIS, matrix metalloproteinase 9 (MMP-9) proteolytic activity contributes to the loss in BBB integrity, increased hemorrhagic risk, and worse outcomes. We have previously shown that sphingosine-1-phosphate receptor (S1PR) ligands attenuated hypoxia plus glucose deprivation (HGD)-induced decreases in brain microvascular endothelial trans-endothelial resistance (TEER), suggesting a potential role for S1PR ligands on barrier function during AIS. Endothelial vascular cell adhesion molecule 1 (VCAM-1) is an inflammatory mediator and adhesion molecule that contributes to the loss of endothelial barrier integrity by serving as a scaffold for immune cell extravasation as well as a mediator for intracellular signals involved in MMP-9 activation. Cell surface adhesion receptor, CD44, likewise plays a key role in facilitating MMP-9 activity via co-localization at the endothelial surface. Thusly, we sought to investigate protein expression levels of VCAM-1, MMP-9, tight junctional proteins, (ZO-1 and claudin-5), as well as to characterize CD44 protein expression in cerebrovascular endothelial cells following HGD. Pediatric male human brain microvascular endothelial cells (HBMECs) were treated with ozanimod (selective S1PR type 1 ligand; 0.5 nM) at P7 and immediately exposed to normoxia (21% O2) or HGD (1% O2) for 3h, a timepoint determined from our previous TEER studies which revealed deterioration of the endothelial barrier as early as 3h. To verify S1PR type 1 dependency, selective blockade with antagonist W146 was applied to select experiments. We observed through standard immunoblotting that HGD induced a simultaneous decrease in ZO-1 and claudin-5 in parallel to increased expression of VCAM-1. Additionally, intracellular pro-MMP-9 (inactive enzymatic form) levels decreased following HGD and the ratio of active MMP-9 to pro-MMP-9 was increased. Ozanimod did not alter these responses. Using immunocytochemistry, anti-CD44 fluorescence was increased following HGD exposure, and ozanimod attenuated this response. S1PR type 1 blockade demonstrated receptor dependence. In conclusion, HGD induced endothelial activation characterized by increased adhesion molecule levels and involvement of MMP-9 activity potentially contributing to tight junction protein loss. Furthermore, S1PR type 1 ligands may in part attenuate HGD-induced BBB integrity loss by modulating the bioavailability of MMP-9 to disrupt endothelial barrier integrity. Mechanism(s) by which S1PR type 1 ligands regulate MMP-9 activity and its associated cell surface adhesion proteins (e.g. CD44 and VCAM-1) following ischemic-like injury warrants further investigation.

Development of a targeted nanoLC-MS/MS method for quantitation of residual toxins from Bordetella pertussis

Whooping cough is a highly contagious respiratory disease caused by Bordetella pertussis (B. pertussis) infection. Pertussis pathogenesis is driven by cell-surface adhesion proteins and secreted toxins; some of which have been harnessed for their immunogenic properties as purified antigen components in acellular vaccines. Two of these virulence factors, adenylate cyclase toxin (ACT) and dermonecrotic toxin (DNT), are protein toxins with potential for co-purification, and therefore must be monitored as process-related impurities during the development of acellular Pertussis vaccine candidates. Here we describe the development of a targeted nanoLC-MS/MS method for absolute quantitation of ACT and DNT in process intermediates from acellular Pertussis antigen purification. Starting from an in silico digest of the toxin sequences, a synthetic peptide screening approach was applied to systematically evaluate candidate sequences as surrogates for protein quantitation. Following refinement to a subset of sequences, absolutely quantified heavy-labelled (AQUA) peptides were implemented in a parallel reaction monitoring (PRM) workflow with limits of detection (LOD) and quantitation (LOQ) in the 12.5–25 amol (2−4 ng/mL) range on-column. In this work, we highlight a ‘standards-driven’ approach to surrogate peptide selection for protein quantitation. This strategy can be broadly applied in the absence of purified reference material and accelerate quantitative LC–MS method development across multiple sample matrices.

ITGA3 serves as a diagnostic and prognostic biomarker for pancreatic cancer.

Background and objective: ITGA3 is a cell surface adhesion protein that interacts with extracellular matrix proteins which function in cancer metastasis. We examined the relationship of pancreatic ITGA3 expression with the clinical and pathological characteristics of patients with pancreatic cancer.Methods: Data mining was used to analyze pancreatic cancer data from The Cancer Genome Atlas database. A Chi squared test was used to evaluate correlations of ITGA3 expression with clinical and pathological parameters. Receiver operating characteristic (ROC) analysis was used to evaluate the diagnostic performance of ITGA3 expression. Survival analysis and Cox regression analysis were used to examine the prognostic value of ITGA3 expression. Gene Set Enrichment Analysis (GSEA) was used to identify signaling pathways related to ITGA3 expression.Results: Pancreatic expression of ITGA3 was greater in patients with pancreatic cancer than those without cancer, and was also associated with histological type, histological grade, stage, T classification, vital status, and relapse. ROC analysis indicated that ITGA3 had significant diagnostic value, in that high expression correlated with poor overall survival and relapse-free survival, especially in patients with early-stage cancer. Cox analysis indicated that high ITGA3 expression was an independent prognostic factor for pancreatic cancer. GSEA analysis identified 9 signaling pathways that were enriched in the presence of high ITGA3 expression.Conclusion: Expression of ITGA3 can be used as a diagnostic and prognostic biomarker in pancreatic cancer.

The unique trimeric assembly of the virulence factor HtrA from Helicobacter pylori occurs via N-terminal domain swapping

Knowledge of the molecular mechanisms of specific bacterial virulence factors can significantly contribute to antibacterial drug discovery. Helicobacter pylori is a Gram-negative microaerophilic bacterium that infects almost half of the world's population, leading to gastric disorders and even gastric cancer. H. pylori expresses a series of virulence factors in the host, among which high-temperature requirement A (HpHtrA) is a newly identified serine protease secreted by H. pylori. HpHtrA cleaves the extracellular domain of the epithelial cell surface adhesion protein E-cadherin and disrupts gastric epithelial cell junctions, allowing H. pylori to access the intercellular space. Here we report the first crystal structure of HpHtrA at 3.0 Å resolution. The structure revealed a new type of HtrA protease trimer stabilized by unique N-terminal domain swapping distinct from other known HtrA homologs. We further observed that truncation of the N terminus completely abrogates HpHtrA trimer formation as well as protease activity. In the presence of unfolded substrate, HpHtrA assembled into cage-like 12-mers or 24-mers. Combining crystallographic, biochemical, and mutagenic data, we propose a mechanistic model of how HpHtrA recognizes and cleaves the well-folded E-cadherin substrate. Our study provides a fundamental basis for the development of anti-H. pylori agents by using a previously uncharacterized HtrA protease as a target.

Expression and Roles of Teneurins in Zebrafish.

The teneurins, also known as Ten-m/Odz, are highly conserved type II transmembrane glycoproteins widely expressed throughout the nervous system. Functioning as dimers, these large cell-surface adhesion proteins play a key role in regulating neurodevelopmental processes such as axon targeting, synaptogenesis and neuronal wiring. Synaptic specificity is driven by molecular interactions, which can occur either in a trans-homophilic manner between teneurins or through a trans-heterophilic interaction across the synaptic cleft between teneurins and other cell-adhesion molecules, such as latrophilins. The significance of teneurins interactions during development is reflected in the widespread expression pattern of the four existing paralogs across interconnected regions of the nervous system, which we demonstrate here via in situ hybridization and the generation of transgenic BAC reporter lines in zebrafish. Focusing on the visual system, we will also highlight the recent developments that have been made in furthering our understanding of teneurin interactions and their functionality, including the instructive role of teneurin-3 in specifying the functional wiring of distinct amacrine and retinal ganglion cells in the vertebrate visual system underlying a particular functionality. Based on the distinct expression pattern of all teneurins in different retinal cells, it is conceivable that the combination of different teneurins is crucial for the generation of discrete visual circuits. Finally, mutations in all four human teneurin genes have been linked to several types of neurodevelopmental disorders. The opportunity therefore arises that findings about the roles of zebrafish teneurins or their orthologs in other species shed light on the molecular mechanisms in the etiology of such human disorders.

Structural Basis for Teneurin Function in Circuit-Wiring: A Toxin Motif at the Synapse

Teneurins (TENs) are cell-surface adhesion proteins with critical roles in tissue development and axon guidance. Here, we report the 3.1-Å cryoelectron microscopy structure of the human TEN2 extracellular region (ECR), revealing a striking similarity to bacterial Tc-toxins. The ECR includes a large β barrel that partially encapsulates a C-terminal domain, which emerges to the solvent through an opening in themid-barrel region. An immunoglobulin (Ig)-like domain seals the bottom of the barrel while a β propeller is attached in a perpendicular orientation. We further show that an alternatively spliced region within the β propeller acts as a switch to regulate trans-cellular adhesion of TEN2 to latrophilin (LPHN), a transmembrane receptor known to mediate critical functions in the central nervous system. One splice variant activates trans-cellular signaling in a LPHN-dependent manner, whereas the other induces inhibitory postsynaptic differentiation. These results highlight the unusual structural organization of TENs giving rise to their multifarious functions.

High Expression of ITGA3 Promotes Proliferation and Cell Cycle Progression and Indicates Poor Prognosis in Intrahepatic Cholangiocarcinoma.

Integrin subunit alpha 3 (ITGA3) interacts with a beta 1 subunit to form a member of the integrin family. Integrins are heterodimeric integral membrane proteins that serve as cell surface adhesion proteins. In this research, we investigated the biological function of this protein in human intrahepatic cholangiocarcinoma (ICC) for the first time. Here, using Western blotting and immunohistochemistry assays, we discovered that ITGA3 was overexpressed in ICC cell lines and ICC patients. Moreover, we found ITGA3 expression correlated with several clinicopathological features, including tumor size, lymph node metastasis, and the TNM stage. Patients with high ITGA3 expression underwent a worse prognosis after complete resection compared with patients with low ITGA3 expression in terms of overall survival. Furthermore, we demonstrated that ITGA3 could significantly promote ICC cell proliferation and cell cycle progression in vitro. However, as a classical cell surface adhesion molecule, we found ITGA3 correlated negatively with the migration and invasion of ICC cell lines, which differs from other malignant tumors. Generally, these findings suggest that ITGA3 may play a role as a potential oncogene in ICC and suppression of ITGA3 expression may establish a novel target for guiding the therapy of ICC patients.

Covalent attachment and Pro-Pro endopeptidase (PPEP-1)-mediated release of Clostridium difficile cell surface proteins involved in adhesion.

In the past decade, Clostridium difficile has emerged as an important gut pathogen. This anaerobic, Gram-positive bacterium is the main cause of infectious nosocomial diarrhea. Whereas much is known about the mechanism through which the C. difficile toxins cause diarrhea, relatively little is known about the dynamics of adhesion and motility, which is mediated by cell surface proteins. This review will discuss the recent advances in our understanding of the sortase-mediated covalent attachment of cell surface (adhesion) proteins to the peptidoglycan layer of C.difficile and their release through the action of a highly specific secreted metalloprotease (Pro-Pro endopeptidase 1, PPEP-1). Specific emphasis will be on a model in which PPEP-1 and its substrates control the switch from a sessile to motile phenotype in C. difficile, and how this is regulated by the cyclic dinucleotide c-di-GMP (3'-5' cyclic dimeric guanosine monophosphate).

Analyzing Cell Surface Adhesion Remodeling in Response to Mechanical Tension Using Magnetic Beads.

Mechanosensitive cell surface adhesion complexes allow cells to sense the mechanical properties of their surroundings. Recent studies have identified both force-sensing molecules at adhesion sites, and force-dependent transcription factors that regulate lineage-specific gene expression and drive phenotypic outputs. However, the signaling networks converting mechanical tension into biochemical pathways have remained elusive. To explore the signaling pathways engaged upon mechanical tension applied to cell surface receptor, superparamagnetic microbeads can be used. Here we present a protocol for using magnetic beads to apply forces to cell surface adhesion proteins. Using this approach, it is possible to investigate not only force-dependent cytoplasmic signaling pathways by various biochemical approaches, but also adhesion remodeling by magnetic isolation of adhesion complexes attached to the ligand-coated beads. This protocol includes the preparation of ligand-coated superparamagnetic beads, and the application of define tensile forces followed by biochemical analyses. Additionally, we provide a representative sample of data demonstrating that tension applied to integrin-based adhesion triggers adhesion remodeling and alters protein tyrosine phosphorylation.

Mechanical Strength and Inhibition of the Staphylococcus aureus Collagen-Binding Protein Cna.

ABSTRACTThe bacterial pathogen Staphylococcus aureus expresses a variety of cell surface adhesion proteins that bind to host extracellular matrix proteins. Among these, the collagen (Cn)-binding protein Cna plays important roles in bacterium-host adherence and in immune evasion. While it is well established that the A region of Cna mediates ligand binding, whether the repetitive B region has a dedicated function is not known. Here, we report the direct measurement of the mechanical strength of Cna-Cn bonds on living bacteria, and we quantify the antiadhesion activity of monoclonal antibodies (MAbs) targeting this interaction. We demonstrate that the strength of Cna-Cn bonds in vivo is very strong (~1.2 nN), consistent with the high-affinity “collagen hug” mechanism. The B region is required for strong ligand binding and has been found to function as a spring capable of sustaining high forces. This previously undescribed mechanical response of the B region is of biological significance as it provides a means to project the A region away from the bacterial surface and to maintain bacterial adhesion under conditions of high forces. We further quantified the antiadhesion activity of MAbs raised against the A region of Cna directly on living bacteria without the need for labeling or purification. Some MAbs are more efficient in blocking single-cell adhesion, suggesting that they act as competitive inhibitors that bind Cna residues directly involved in ligand binding. This report highlights the role of protein mechanics in activating the function of staphylococcal adhesion proteins and emphasizes the potential of antibodies to prevent staphylococcal adhesion and biofilm formation.

Development of lifitegrast: a novel T-cell inhibitor for the treatment of dry eye disease.

Dry eye disease (DED) is a multifactorial disorder of the ocular surface characterized by symptoms of discomfort, decreased tear quality, and chronic inflammation that affects an estimated 20 million patients in the US alone. DED is associated with localized inflammation of the ocular surface and periocular tissues leading to homing and activation of T cells, cytokine release, and development of hyperosmolar tears. This inflammatory milieu results in symptoms of eye dryness and discomfort. Homing of T cells to the ocular surface is influenced by the binding of lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18; αLβ2), a cell surface adhesion protein, to its cognate ligand, intercellular adhesion molecule-1 (ICAM-1; CD54), which is expressed on inflamed ocular/periocular epithelium and vascular endothelium. LFA-1/ICAM-1 binding within the immunologic synapse enables both T-cell activation and cytokine release. Lifitegrast is a novel T-cell integrin antagonist that is designed to mimic the binding epitope of ICAM-1. It serves as a molecular decoy to block the binding of LFA-1/ICAM-1 and inhibits the downstream inflammatory process. In vitro studies have demonstrated that lifitegrast inhibits T-cell adhesion to ICAM-1-expressing cells and inhibits secretion of pro-inflammatory cytokines including interferon gamma, tumor necrosis factor alpha, macrophage inflammatory protein 1 alpha, interleukin (IL)-1α, IL-1β, IL-2, IL-4, and IL-6, all of which are known to be associated with DED. Lifitegrast has the potential to be the first pharmaceutical product approved in the US indicated for the treatment of both symptoms and signs of DED. Clinical trials involving over 2,500 adult DED patients have demonstrated that topically administered lifitegrast 5.0% ophthalmic solution can rapidly reduce the symptoms of eye dryness and decrease ocular surface staining with an acceptable long-term safety profile. The purpose of this review is to highlight the developmental story – from bench top to bedside – behind the scientific rationale, engineering, and clinical experience of lifitegrast for the treatment of DED.

Identification of G α 12 and G α 13 Determinants of Binding to Cell Adhesion Proteins

The trimeric G protein α-subunits Gα12 and Gα13 have 66% amino acid homology but only partial overlap in their signaling function and downstream target proteins. Previously we reported that Gα12 and Gα13 bind the cytoplasmic domain of several cadherins, and other investigators have shown Gα13, but not Gα12, to bind the cytoplasmic tail of β3-integrin. The switch I region of Gα13 has been implicated in binding to vascular endothelial (VE) cadherin and β3-integrin, suggesting key residues arose in the G12/13 class to allow interaction with these cell-surface adhesion proteins. To more precisely define determinants in the G12/13 class mediating these interactions, we utilized epitope-tagged forms of Gα12 and Gα13 to examine their relative affinity for epithelial (E), neural (N), and VE cadherin, as well as β3-integrin. Gα12 showed more robust interaction than Gα13 with both types of cell adhesion protein, and Gα12-to-Gα13 substitutions revealed several residues contributing to their different affinity for these proteins. Surprisingly, GFP tagging in the Gα12 helical domain disrupted binding to β3-integrin but not cadherins, whereas Gα12 with a myc tag at the same location bound robustly to both proteins. These findings suggest that helical domain residues contributing to β3-integrin binding are blocked by the larger GFP adduct but not the myc tag. Cassette substitutions in the Gα12 helical domain implicated several regions in binding cadherins, and these showed partial overlap with Gα12 mutations disruptive to β3-integrin binding. These findings provide structural details of G12/13 engagement of cadherins and integrins and suggest distinct mechanisms of interaction with these adhesion proteins. Supported by the GSK Foundation and Lineberger Comprehensive Cancer Center

Inhibition of LFA-1/ICAM-1 Interaction: A Therapeutic Strategy for Surmounting Inflammation

Leukocyte function associated antigen-1 (LFA-1, CD11a/CD18, αLβ2) is a cell surface adhesion protein expressed on leukocytes. Interaction of LFA-1 with its counter receptor intercellular adhesion molecule-1 (ICAM-1) promotes the migration of leukocytes to the site of inflammation and is responsible for a variety of cell adhesion events required for normal and pathological functions of the immune system. Thus, the LFA-1/ICAM-1 interaction is thought to play a role in the pathogenesis of inflammatory and auto-immune disease conditions such as psoriasis, multiple sclerosis, asthma, rheumatoid arthritis, inflammatory bowel disease and transplant rejection. LFA-1/ICAM-1 interaction inhibitors have been extensively studied as a therapeutic target for clinically important diseases; leading to the development of therapeutic antibodies, peptides, peptidomimetics, allosteric and competitive inhibitors. However, despite of promising preclinical results, the outcome of clinical trials with LFA-1/ICAM-1 interaction inhibitors has been inconsistent and met with limited clinical success. This article gives a brief account of rationale and efficacy of different LFA-1/ICAM-1 interaction inhibitors as promising therapeutic strategy for surmounting inflammatory disorders.