The sole currently approved malaria vaccine targets the circumsporozoite protein-the protein that densely coats the surface of sporozoites, the parasite stage deposited in the skin of the mammalian host by infected mosquitoes. However, this vaccine only confers moderate protection against clinical diseases in children, impelling a continuous search for novel candidates. In this work, we studied the importance of the membrane-associated erythrocyte binding-like protein (MAEBL) for infection by Plasmodium sporozoites. Using transgenic parasites and live imaging in mice, we show that the absence of MAEBL reduces Plasmodium berghei hemolymph sporozoite infectivity to mice. Moreover, we found that maebl knockout (maebl-) sporozoites display reduced adhesion, including to cultured hepatocytes, which could contribute to the defects in multiple biological processes, such as in gliding motility, hepatocyte wounding, and invasion. The maebl- defective phenotypes in mosquito salivary gland and liver infection were reverted by genetic complementation. Using a parasite line expressing a C-terminal myc-tagged MAEBL, we found that MAEBL levels peak in midgut and hemolymph parasites but drop after sporozoite entry into the salivary glands, where the labeling was found to be heterogeneous among sporozoites. MAEBL was found associated, not only with micronemes, but also with the surface of mature sporozoites. Overall, our data provide further insight into the role of MAEBL in sporozoite infectivity and may contribute to the design of future immune interventions.

A new cell surface-associated adhesive glycoprotein with a molecular weight of 70,000 was separated from differentiated rat ascites hepatoma cells forming cell islands in vivo (but not from undifferentiated rat ascites hepatoma cells present as single cells in vivo) and highly purified by chromatography; it was synthesized by the cells and localized on the cell surface. Its synthesis began to rise rapidly and reached its peak in 24 hr cultivation, i.e., a 10-fold increase. This substance induced not only aggregation but also adhesiveness of the cells characterized by junctional complexes including tight junctions, desmosomes, and intermediate junctions, closely resembling the frequency and distribution of junctional complexes observed on the above cell islands. Its potency was inhibited specifically by D-mannose and alpha-methyl-D-mannoside; the numbers of the binding sites per cell were calculated as 6 x 10(5). Its activity was concerned with the protein portion of the molecule, and not with the carbohydrate portion. Thus, it seemed reasonable that the adhesive glycoprotein may play a key role in the cell adhesiveness and island formation. In contrast, serum-associated adhesive glycoprotein, separated from normal rat serum, could aggregate the cells but not develop junctional complex.

Opticin is an extracellular matrix glycoprotein that we identified associated with the collagen network of the vitreous humor of the eye. Recently, we discovered that opticin possesses anti-angiogenic activity using a murine oxygen-induced retinopathy model: here, we investigate the underlying mechanism. Using an ex vivo chick chorioallantoic membrane assay, we show that opticin inhibits angiogenesis when stimulated by a range of growth factors. We show that it suppresses capillary morphogenesis, inhibits endothelial invasion, and promotes capillary network regression in three-dimensional matrices of collagen and Matrigel(TM). We then show that opticin binds to collagen and thereby competitively inhibits endothelial cell interactions with collagen via α(1)β(1) and α(2)β(1) integrins, thereby preventing the strong adhesion that is required for proangiogenic signaling via these integrins.

Hyaluronic acid (HA) is a natural polysaccharide with great biocompatibility for a variety of biomedical applications, such as tissue scaffolds, dermal fillers, and drug-delivery carriers. Despite the medical impact of HA, its poor adhesiveness and short-term in vivo stability limit its therapeutic efficacy. To overcome these shortcomings, a versatile modification strategy for the HA backbone has been developed. This strategy involves tethering phenol moieties on HA to provide both robust adhesiveness and intermolecular cohesion and can be used for oxidative crosslinking of the polymeric chain. However, a lack of knowledge still exists regarding the interchangeable phenolic adhesion and cohesion depending on the type of oxidizing agent used. Here, we reveal the correlation between phenolic adhesion and cohesion upon gelation of two different HA-phenol conjugates, HA-tyramine and HA-catechol, depending on the oxidant. For covalent/non-covalent crosslinking of HA, oxidizing agents, horseradish peroxidase/hydrogen peroxide, chemical oxidants (e.g., base, sodium periodate), and metal ions, were utilized. As a result, HA-catechol showed stronger adhesion properties, whereas HA-tyramine showed higher cohesion properties. In addition, covalent bonds allowed better adhesion compared to that of non-covalent bonds. Our findings are promising for designing adhesive and mechanically robust biomaterials based on phenol chemistry.

Mesenchymal stem cells natively circulating or delivered into the blood stream home to sites of injury. The mechanism of mesenchymal stem cell homing to sites of injury is poorly understood. We have shown that the development of apoptosis in endothelial cells stimulates endothelial cell adhesiveness for mesenchymal stem cells. Adhesion of mesenchymal stem cells to apoptotic endothelial cells depends on the activation of endothelial caspases and p38 MAPK. Activation of p38 MAPK in endothelial cells has a primary effect while the activation of caspases potentiates the mesenchymal stem cell adhesion. Overall, our study of the mesenchymal stem cell interaction with endothelial cells indicates that mesenchymal stem cells recognize and specifically adhere to distressed/apoptotic endothelial cells.

Anoikis is a type of apoptosis that occurs in response to the loss of adhesion to the extracellular matrix (ECM). Anoikis resistance is a critical mechanism in cancer and contributes to tumor metastasis. Nitric oxide (NO) is frequently upregulated in the tumor area and is considered an important player in cancer metastasis. The aim of this study was to evaluate the effect of NO on adhesiveness, invasiveness, and migration of anoikis-resistant endothelial cells. Here, we report that anoikis-resistant endothelial cells overexpress endothelial nitric oxide synthase. The inhibition of NO release in anoikis-resistant endothelial cells was able to decrease adhesiveness to fibronectin, laminin, and collagen IV. This was accompanied by an increase in cell invasiveness and migration. Furthermore, anoikis-resistant cell lines displayed a decrease in fibronectin and collagen IV protein expression after L-NAME treatment. These alterations in adhesiveness and invasiveness were the consequence of MMP-2 up-regulation observed after NO release inhibition. The decrease in NO levels was able to down-regulate the activating transcription factor 3 (ATF3) protein expression. ATF3 represses MMP-2 gene expression by antagonizing p53-dependent trans-activation of the MMP-2 promoter. We speculate that the increased release of NO by anoikis-resistant endothelial cells acted as a response to restrict the MMP-2 action, interfering in MMP-2 gene expression via ATF3 regulation. The up-regulation of nitric oxide by anoikis-resistant endothelial cells is an important response to restrict tumorigenic behavior. Without this mechanism, invasiveness and migration potential would be even higher, as shown after L-NAME treatment.

Kindlins are integrin-interacting proteins essential for integrin-mediated cell adhesiveness. In this study, we focused on the evolutionary origin and functional specialization of kindlins as a part of the evolutionary adaptation of cell adhesive machinery. Database searches revealed that many members of the integrin machinery (including talin and integrins) existed before kindlin emergence in evolution. Among the analyzed species, all metazoan lineages—but none of the premetazoans—had at least one kindlin-encoding gene, whereas talin was present in several premetazoan lineages. Kindlin appears to originate from a duplication of the sequence encoding the N-terminal fragment of talin (the talin head domain) with a subsequent insertion of the PH domain of separate origin. Sequence analysis identified a member of the actin filament-associated protein 1 (AFAP1) superfamily as the most likely origin of the kindlin PH domain. The functional divergence between kindlin paralogues was assessed using the sequence swap (chimera) approach. Comparison of kindlin 2 (K2)/kindlin 3 (K3) chimeras revealed that the F2 subdomain, in particular its C-terminal part, is crucial for the differential functional properties of K2 and K3. The presence of this segment enables K2 but not K3 to localize to focal adhesions. Sequence analysis of the C-terminal part of the F2 subdomain of K3 suggests that insertion of a variable glycine-rich sequence in vertebrates contributed to the loss of constitutive K3 targeting to focal adhesions. Thus emergence and subsequent functional specialization of kindlins allowed multicellular organisms to develop additional tissue-specific adaptations of cell adhesiveness.

Leukocyte cell recruitment into the vascular subendothelium constitutes an early event in the atherogenic process. As the effect of the constitutive androstane receptor (CAR) on leukocyte recruitment and endothelial dysfunction is poorly understood, this study investigated whether the role of CAR activation can affect this response and the underlying mechanisms involved. Under physiological flow conditions, TNFα-induced endothelial adhesion of human leukocyte cells was concentration-dependently inhibited by preincubation of human umbilical arterial endothelial cells with the selective human CAR ligand CITCO. CAR agonism also prevented TNFα induced VCAM-1 expression, as well as MCP-1/CCL-2 and RANTES/CCL-5 release in endothelial cells. Suppression of CAR expression with a small interfering RNA abrogated the inhibitory effects of CITCO on these responses. Furthermore, CITCO increased interaction of CAR with Retinoid X Receptor (RXR) and reduced TNFα-induced p38-MAPK/NF-κB activation. In vivo, using intravital microscopy in the mouse cremasteric microcirculation treatment with the selective mouse CAR ligand TCPOBOP inhibited TNFα-induced leukocyte rolling flux, adhesion, and emigration and decreased VCAM-1 in endothelium. These results reveal that CAR agonists can inhibit the initial inflammatory response that precedes the atherogenic process by targeting different steps in the leukocyte recruitment cascade. Therefore, CAR agonists may constitute a new therapeutic tool in controlling cardiovascular disease-associated inflammatory processes.

Atherosclerosis is an arterial inflammatory disease. The circulating level of the C-C chemokine ligand (CCL4) is increased in atherosclerotic patients. This study aimed to investigate whether CCL4 inhibition could retard the progression of atherosclerosis. In ApoE knockout mice, CCL4 antibody treatment reduced circulating interleukin-6 (IL-6) and tumor necrosis factor (TNF)-α levels and improved lipid profiles accompanied with upregulation of the liver X receptor. CCL4 inhibition reduced the atheroma areas and modified the progression of atheroma plaques, which consisted of a thicker fibrous cap with a reduced macrophage content and lower matrix metalloproteinase-2 and -9 expressions, suggesting the stabilization of atheroma plaques. Human coronary endothelial cells (HCAECs) and macrophages were stimulated with TNF-α or oxidized LDL (ox-LDL). The induced expression of E-selectin, vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) were attenuated by the CCL4 antibody or CCL4 si-RNA. CCL4 inhibition reduced the adhesiveness of HCAECs, which is an early sign of atherogenesis. CCL4 blockade reduced the activity of metalloproteinase-2 and -9 and the production of TNF-α and IL-6 in stimulated macrophages. The effects of CCL4 inhibition on down-regulating adhesion and inflammation proteins were obtained through the nuclear factor kappa B (NFκB) signaling pathway. The direct inhibition of CCL4 stabilized atheroma and reduced endothelial and macrophage activation. CCL4 may be a novel therapeutic target for modulating atherosclerosis.

The Anaerolineae lineage of Chloroflexi had been identified as one of the core microbial populations in anaerobic digesters; however, the ecological role of the Anaerolineae remains uncertain due to the scarcity of isolates and annotated genome sequences. Our previous metatranscriptional analysis revealed this prevalent population that showed minimum involvement in the main pathways of cellulose hydrolysis and subsequent methanogenesis in the thermophilic cellulose fermentative consortium (TCF).

Nature provides biomaterials that tend to be effective to control both their adhesive and cohesive properties. A catecholamine motif found in the marine mussels, the mytilus edulis foot protein, can play adhesiveness and cohesiveness. Particularly, acidic pH drives catechol (Cat) to have adhesive function, resulting in surface coating, while basic pH allows to enhance its cohesive properties, resulting in the formation of hydrogels. In this work, we demonstrated the usefulness of Cat-conjugated chondroitin sulfate (CS) as a platform for mesenchymal stem cell culture, utilizing the adhesive property of CS-Cat as coating for different substrates and the cohesive properties as hydrogel for cells encapsulation. To prepare the CS-Cat biopolymer, dopamine (DP) was coupled to the CS by carbodiimide coupling reaction and the Cat content was determined by UV-Vis spectroscopy (4.8 ± 0.6%). To demonstrate the adhesive properties of the biopolymer, PLA, PCL, TiO2, and SiO2 substrates were immersed in CS-Cat solution (pH < 2). Following the coating, the surfaces became highly hydrophilic, exhibiting a contact angle less than 35°. Also, in the presence of an oxidizing agent at pH 8, CS-Cat solution immediately became a hydrogel, as shown by inverted-vial test. Finally, immortalized TERT human mesenchymal stem cells (Y201) confirmed the high cytocompatibility of the biopolymer. The CS-Cat coating significantly enabled the Y201 adhesion onto PLA substrates, while the prepared hydrogel demonstrated to be a suitable environment for the encapsulation of cells as suitable bioink for further bioprinting applications.

Plasmodium vivax, the major cause of malaria in Latin America, has a large subtelomeric multigene family called vir. In the P. vivax genome, about 20% of its sequences are vir genes. Vir antigens are grouped in subfamilies according to their sequence similarities and have been shown to have distinct roles and subcellular locations. However, little is known about vir subfamilies, especially when comes to their functions.

A major deficit in tissue engineering strategies is the lack of materials that promote angiogenesis, wherein endothelial cells from the host vasculature invade the implanted matrix to form new blood vessels. To determine the material properties that regulate angiogenesis, we have developed a microfluidic in vitro model in which chemokine-guided endothelial cell sprouting into a tunable hydrogel is followed by the formation of perfusable lumens. We show that long, perfusable tubes only develop if hydrogel adhesiveness and degradability are fine-tuned to support the initial collective invasion of endothelial cells and, at the same time, allow for matrix remodeling to permit the opening of lumens. These studies provide a better understanding of how cell-matrix interactions regulate angiogenesis and, therefore, constitute an important step towards optimal design criteria for tissue-engineered materials that require vascularization.

Gutta-percha's lack of adhesion has been presented as a drawback to avoid gaps at sealer/gutta-percha interface. Plasma treatments have been scarcely assessed on gutta-percha surfaces as a method of enhancing adhesiveness. This study aimed to evaluate the effect of low-pressure Argon and Oxygen plasma atmospheres on conventional and bioceramic gutta-percha standardized smooth discs, assessing their roughness, surface free energy, chemical structure, and sealer wettability. A Low-Pressure Plasma Cleaner by Diener Electronic (Zepto Model) was used. Different gases (Argon or Oxygen), powers (25 W, or 50 W), and exposure times (30 s, 60 s, 120 s, or 180 s) were tested in control and experimental groups. Kruskal-Wallis and Student's t-test were used in data analysis. Statistically significant differences were detected when P < 0.05. Both gases showed different behaviors according to the parameters selected. Even though chemical changes were detected, the basic molecular structure was maintained. Argon or Oxygen plasma treatments favoured the wetting of conventional and bioceramic gutta-perchas by Endoresin and AH Plus Bioceramic sealers (P < 0.001). Overall, the functionalization of gutta-percha surfaces with Argon or Oxygen plasma treatments can increase roughness, surface free energy and wettability, which might improve its adhesive properties when compared to non-treated gutta-percha.

Dynamic regulation of integrin adhesiveness is required for immune cell-cell interactions and leukocyte migration. Here, we investigate the relationship between cell adhesion and integrin microclustering as measured by fluorescence resonance energy transfer, and macroclustering as measured by high resolution fluorescence microscopy. Stimuli that activate adhesion through leukocyte function-associated molecule-1 (LFA-1) failed to alter clustering of LFA-1 in the absence of ligand. Binding of monomeric intercellular adhesion molecule-1 (ICAM-1) induced profound changes in the conformation of LFA-1 but did not alter clustering, whereas binding of ICAM-1 oligomers induced significant microclustering. Increased diffusivity in the membrane by cytoskeleton-disrupting agents was sufficient to drive adhesion in the absence of affinity modulation and was associated with a greater accumulation of LFA-1 to the zone of adhesion, but redistribution did not precede cell adhesion. Disruption of conformational communication within the extracellular domain of LFA-1 blocked adhesion stimulated by affinity-modulating agents, but not adhesion stimulated by cytoskeleton-disrupting agents. Thus, LFA-1 clustering does not precede ligand binding, and instead functions in adhesion strengthening after binding to multivalent ligands.

Reproductive strategies and spawning habits play key roles in the evolution of endemic East Asian cyprinids. However, the molecular mechanisms underlying the regulation of spawning habits are not well understood. We recently identified zona pellucida (Zp) as the top differentially expressed protein between East Asian cyprinids that produce adhesive and semi-buoyant eggs, suggesting that Zp protein may play important roles in the regulation of egg type. In this work, we generated transgenic zebrafish in which oocyte-specific expression of zp genes from rare minnow ( Gobiocypris rarus), an East Asian cyprinid laying adhesive eggs, was driven by a zebrafish zp3.2 gene promoter. We found that the transgenic eggs obtained partial adhesiveness and exhibited alteration in hydration and buoyancy. Abnormal metabolism of vitellogenin (VTG) may contribute to enhanced hydration and/or buoyancy. Our work shows that expression of the exogenous zp3a gene from an adhesive-egg producing fish is sufficient to induce changes in both egg adhesiveness and buoyancy in zebrafish, emphasizing the important role of zp genes in the regulation of spawning habits. Our results thus provide new insights into how endemic East Asian cyprinids may have adapted to the Yangtze river-lake system via changes in spawning habits.

Platelet-rich plasma (PRP) has attracted much attention for the treatment of articular cartilage defects or wounds due to its intrinsic content of growth factors relevant for tissue repair. However, the short residence time of PRP in vivo, due to the action of lytic enzymes, its weak mechanical properties and the consequent short-term release of bioactive factors has restricted its application and efficacy. The present work aimed at designing new formulation strategies for PRP, based on the use of platelet concentrate (PC)-loaded hydrogels or interpenetrating polymer networks, directed at improving mechanical stability and sustaining the release of bioactive growth factors over a prolonged time-span. The interpenetrating hydrogels comprised two polymer networks interlaced on a molecular scale: (a) a first covalent network of thermosensitive and biodegradable vinyl sulfone bearing p(hydroxypropyl methacrylamide-lacate)-polyethylene glycol triblock copolymers, tandem cross-linked by thermal gelation and Michael addition when combined with thiolated hyaluronic acid, and (b) a second network composed of cross-linked fibrin. The PC-loaded hydrogels, instead, was formed only by network (a). All the designed and successfully synthesized formulations greatly increased the stability of PRP in vitro, leading to significant increase in degradation time and storage modulus of PRP gel. The resulting viscoelastic networks showed the ability to controllably release platelet derived growth factor and transforming growth factr β1, and to improve the tissue adhesiveness of PRP. The newly developed hydrogels show great potential for application in the field of wound healing, cartilage repair and beyond.

The incidence of infective endocarditis in patients with bioprosthetic heart valves is over 100 times that of the general population with S. aureus recognized as the causative organism in approximately 1/3 of cases. In this study, (1) the microbicidal and virucidal effect of a polyphenolic solution was carefully evaluated. The same solution was then adopted for the treatment of a commercial bioprosthetic heart valve model for (2) the assessment of inhibition of S. aureus adhesiveness.

Retinal adhesiveness and subretinal fluid absorption was studied in Dutch rabbit eyes given intravitreal injections of hemicholinium-3 (HC-3) which causes loss of photoreceptor outer segments and retinal pigment epithelial (RPE) damage. After HC-3 administration, some areas of the fundus showed pigmentary changes and others appeared normal. Small, non-rhegmatogenous retinal detachments were made in both areas. Within 2-5 days after HC-3 injection, only in the areas of visible damage, subretinal fluid spread laterally to make very flat retinal detachments, and the fluid absorbed very quickly. At later intervals, absorption was slower than normal, presumably because of scarring and RPE metabolic damage. HC-3 provides an experimental technique for transiently weakening retinal adhesiveness in vivo but its use as a model must account for the effects of both outer segment and RPE damage.

Immune cells have the ubiquitous capability to migrate disregarding the adhesion properties of the environment, which requires a versatile adaptation of their adhesiveness mediated by integrins, a family of specialized adhesion proteins. Each subtype of integrins has several ligands and several affinity states controlled by internal and external stimuli. However, probing cell adhesion properties on live cells without perturbing cell motility is highly challenging, especially in vivo. Here, we developed a novel in vitro method using micron-size beads pulled by flow to functionally probe the local surface adhesiveness of live and motile cells. This method allowed a functional mapping of the adhesiveness mediated by VLA-4 and LFA-1 integrins on the trailing and leading edges of live human T lymphocytes. We show that cell polarization processes enhance integrin-mediated adhesiveness toward cell rear for VLA-4 and cell front for LFA-1. Furthermore, an inhibiting crosstalk of LFA-1 toward VLA-4 and an activating crosstalk of VLA-4 toward LFA-1 were found to modulate cell adhesiveness with a long-distance effect across the cell. These combined signaling processes directly support the bistable model that explains the emergence of the versatile guidance of lymphocyte under flow. Molecularly, Sharpin, an LFA-1 inhibitor in lymphocyte uropod, was found involved in the LFA-1 deadhesion of lymphocytes; however, both Sharpin and Myosin inhibition had a rather modest impact on adhesiveness. Quantitative 3D immunostaining identified high-affinity LFA-1 and VLA-4 densities at around 50 and 100 molecules/μm2 in basal adherent zones, respectively. Interestingly, a latent adhesiveness of dorsal zones was not grasped by immunostaining but assessed by direct functional assays with beads. The combination of live functional assays, molecular imaging, and genome editing is instrumental to characterizing the spatiotemporal regulation of integrin-mediated adhesiveness at molecular and cell scales, which opens a new perspective to decipher sophisticated phenotypes of motility and guidance.