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Trichomonas vaginalis is a protist parasite that causes trichomoniasis, a sexually transmitted disease. Metronidazole is the current treatment for trichomoniasis. However, this drug can provoke severe side effects, and some strains present resistance, making the development of alternative treatments for trichomoniasis urgent.We investigate the use of essential oil obtained from Dracocephalum kotschyi on T. vaginalis. D. kotschyi has antispasmodic and analgesic properties and is well known in Iran.The essential oil was obtained by hydrodistillation from 1000 g of the powdered plant. Gas chromatography-mass spectrometry analysis was used for the chemical composition of the essential oil, and 11 substances were identified, corresponding to 91.5% of the oil. Copaene (22.15%), Methyl geranate (16.31%), Geranial (13.78%) and Carvone (11.34%) were the main substances. A cell viability test was used to determine the percentage of growth inhibition (GI%) and the half-maximal inhibitory concentration (IC50) on T. vaginalis after incubation with the prepared essential oil.The oil induced an IC50 of 84.07 µg/ml after 24 h contact with trophozoites. Cytotoxicity was determined by MTT assay on the J774.A1 haematopoietic cell line. In addition, the initial stage of apoptosis was assayed using the fluorescein isothiocyanate Annexin V Apoptosis Detection Kit. Evaluation of the in vitro anti-trichomonal properties of D. kotschyi essential oils showed that it effectively induces apoptosis on T. vaginalis between 100 and 700 µg/ml after 48 h without toxicity on haematopoietic cells, suggesting that D. kotschyi essential oil can induce programmed death in T. vaginalis.The anti-trichomonal properties of D. kotschyi essential oil indicate that they could be suitable for new pharmacologic studies after new tests with human vaginal epithelial cells.

COVID-19 is a respiratory disease of worldwide importance as it has brought enormous health problems to the world’s population. The best-known way of transmission of the virus is through aerosolization. However, research is needed to explore other transmission routes. Researchers hypothesized that arthropods could transmit SARs-CoV-2. This study is aimed at reviewing research on arthropods as possible reservoirs and/or vectors of SARS-CoV-2, the causative agent of COVID-19. Following PRISMA guidelines, we conducted a systematic review using several electronic databases/academic searches with the search terms “arthropods,” “coronavirus,” and “transmission.” A total of 64 unique articles were identified, of which 58 were included in the review. The SARS-CoV-2 virus is tiny and invisible to the naked eye, and its presence in stools, droplets, and surfaces was detected. One doubt is whether insects can transmit the virus from one place to another. Thus, a healthy carrier of the COVID-19 virus can be at the root of the contamination of their community or their family through the transport of the virus by insects from the interior (flies, cockroaches, etc.) from their feces and food surfaces. Hygiene care within communities and families becomes a prime factor. Coronavirus infection is a significant public health problem around the world. The prevention and control of outbreaks remain very important, even with the production of new vaccines. The main option to achieve this is the proper management of the transmission of the virus. The registry of infected people is currently the basis for the transmission of COVID-19. However, questions about the possibility of infection from other sources and its prevention are not receiving adequate attention. Numerous studies have shown the possibility that SARS-COV-2 fragments could have a longer life than shed respiratory droplets. Also, this virus is larger than those of other coronavirus families.

Trichomonas vaginalis is a protozoan that causes human trichomoniasis, the most common non-viral sexually transmitted infection (STI) affecting approximately 278 million people worldwide. The current treatment for trichomoniasis is based on 1-(2-hydroxyethyl)-2-methyl-5-nitroimidazole, known as metronidazole (MTZ). Although effective in clearing the parasite infection, MTZ is related to provoking severe side effects, and it is not recommended during pregnancy. In addition, some strains present resistance to 5'-nitroimidazoles, making urgent the development of alternative drugs for trichomoniasis. Amiodarone, an antiarrhythmic drug, exerts a significant anti-parasite effect, mainly due to its interference with calcium homeostasis and the biosynthesis of sterols. Therefore, we decided to test the effect of amiodarone and two other related compounds (amioder and dronedarone) on T. vaginalis. Our observations show that amiodarone stimulated, rather than inhibited, parasite growth, induced cell aggregation, and glycogen accumulation. Furthermore, the other two compounds displayed anti-parasite activity with IC50 of 3.15 and 11 µM, respectively, and the apoptosis-like process killed the cells. In addition, cells exhibited morphological changes, including an effect on hydrogenosomes structure.

The parasitic protozoan Giardia intestinalis, the causative agent of giardiasis, presents a stable and elaborated cytoskeleton, which shapes and supports several intracellular structures, including the ventral disc, the median body, the funis, and four pairs of flagella. Giardia trophozoite is the motile form that inhabits the host small intestine and attaches to epithelial cells, leading to infection. The ventral disc is considered one important element of adhesion to the intestinal cells. It is adjacent to the plasma membrane in the ventral region of the cell and consists of a spiral layer of microtubules and microribbons. In this work, we studied the organization of the cytoskeleton in the ventral disc of G. intestinalis trophozoites using high-resolution scanning electron microscopy or helium ion microscopy in plasma membrane-extracted cells. Here, we show novel morphological details about the arrangement of cross-bridges in different regions of the ventral disc. Results showed that the disc is a non-uniformly organized structure that presents specific domains, such as the margin and the ventral groove region. High-resolution scanning electron microscopy allowed observation of the labeling pattern for several anti-tubulin antibodies using secondary gold particle-labeled antibodies. Labeling in the region of the emergence of the microtubules and supernumerary microtubules using an anti-acetylated tubulin antibody was observed. Ultrastructural analysis and immunogold labeling for gamma-tubulin suggest that disc microtubules originate from a region bounded by the bands of the banded collar and merge with microtubules formed at the perinuclear region. Actin-like filaments and microtubules of the disc are associated, showing an interconnection between elements of the cytoskeleton of the trophozoite.

The parasitic diseases represent the most important health risk, especially in underdeveloped countries where they have a deep impact on public health. Trichomoniasis is a prevalent non-viral sexually transmitted disease, and a significant amount of new cases are identified each year globally. Furthermore, the infection is linked with serious concerns such as pregnancy outcomes, infertility, predisposition to cervical and prostate cancer, and increased transmission and acquisition of HIV. The therapy is restricted, adverse effects are often observed, and resistance to the drugs is emerging. Based on this, a new treatment for trichomoniasis is necessary. Natural products represent a rich source of bioactive compounds, and even today, they are used in the search for new drugs. Additionally, natural products provide a wide variety of leadership structures that can be used by the pharmaceutical industry as a template in the development of new drugs that are more effective and have fewer or no undesirable side effects compared to current treatments. This review focuses on the medicinal plants that possess anti-trichomonal activity in vitro or in vivo. An electronic database search was carried out covering the last three decades, i.e., 1990–2020. The literature search revealed that almost a dozen isolated phytoconstituents are being explored globally for their anti-trichomonal activity. Simultaneously, many countries have their own traditional or folk medicine for trichomoniasis that utilizes their native plants, as a whole, or even extracts. This review focuses mainly on the human parasite Trichomonas vaginalis. However, at some points mention is also made to Tritrichomonas foetus that causes trichomoniasis in animals of high veterinary and economical interest. We will focus on the plants and plant-based compounds and their anti-trichomonal activity. The literature search highlighted that there are abundant compounds that possess anti-trichomonal activity; however, in-depth in-vivo evaluation of compounds and their clinical evaluation has not been undertaken. There is a critical need for new anti-trichomonal compounds, and focused research on phytoconstituents can provide the way forward., Graphical abstract Image 1

Giardia comprises one genus with several morphologically distinct species described in mammals (including humans, marsupials, rodents), birds, and amphibians. This group of protists provokes diarrhoea diseases in humans and animals worldwide. Transmission of the parasite occurs through the faecal-oral route. Regarding the presence of Giardia in invertebrates, some works have shown that flies can transmit Giardia cysts by contact and transport between contaminated faeces and food. In this way, flies would eventually transmit this parasite. To date, Giardia's presence in the gut of other invertebrates has not been described in the literature. Here we show by first time, using scanning electron microscopy, the presence of Giardia-like trophozoites in the gut of termite Heterotermes tenuis. Two groups of Giardia were found based exclusively on the size and the flange shape of the protozoa: one presented eight flagella, a ventral disc, size, and shape very similar to Giardia intestinalis. In contrast, other cells were smaller and showed some differences in the external morphology. We cannot exclude the possibility that they correspond to the same species and that these differences result from protozoan heterogeneity.

Giardia intestinalis is a parasitic protozoan that inhabits its vertebrate hosts' upper small intestine and is the most common cause of waterborne diarrhoea worldwide. Giardia trophozoites present few organelles, and among them, they possess peripheral vesicles (PVs), which are considered an endosomal–lysosomal system. All experimental procedures carried out until now indicate that Giardia ingests macromolecules by fluid-phase and receptor-mediated endocytic pathways. Still, there is no description concerning the interaction and ingestion of large materials. Here, we tested Giardia's capacity to interact with large particles; once, in vivo, it inhabits an environment with a microbiota. We tested protozoan interaction with yeasts, bacteria, latex beads, ferritin and albumin, in different times of interaction and used several microscopy techniques (light microscopy, scanning electron microscopy and transmission electron microscopy) to follow their fate. Giardia interacted with all of the materials we tested. Projections of the plasma membrane similar to pseudopods were seen. As albumin, small markers were found in the PVs while the larger materials were not seen there. Large vacuoles containing large latex beads were detected intracellularly. Thus, we observed that: (1) Giardia interacts with large materials; (2) Giardia can display an amoeboid shape and exhibit membrane projections when in contact with microorganisms and large inorganic materials; (3) the region of the exit of the ventral flagella is very active when in contact with large materials, although all cell surface also present activity in the interactions; (4) intracellular vacuoles, which are not the PVs, present ingested large beads.

Giardia intestinalis has unique characteristics, even in the absence of certain organelles. For instance, Golgi and mitochondria are not found. On the other hand, there is a network of peripheral vacuoles (PVs) and mitosomes. The endoplasmic reticulum (ER), nuclear membrane, peroxisomes, and lipid bodies are present. The peripheral vacuole system seems to play several simultaneous roles. It is involved in the endocytic activity of the trophozoite but also has characteristics of early and late endosomes and even lysosomes, establishing a connection with the ER. Some of the PVs contain small vesicles, acting as multivesicular bodies, including the release of exosomes. The mitosomes are surrounded by two membranes, divide during mitosis, and are distributed throughout the cell. They do not contain DNA, enzymes involved in the citric acid cycle, respiratory chain, or ATP synthesis. However, they contain the iron-sulfur complex and transporters as TOM and TIM. Some mitosomes are linked to flagellar axonemes through a fibrillar connection. During encystation, two types of larger cytoplasmic vesicles appear. One originating from the ER contains the cyst wall proteins. Another contains carbohydrates. Both migrate to the cell periphery and fuse with plasma membrane secreting their contents to give rise to the cell wall.

Giardiasis is an intestinal disease caused by the parasite protozoan Giardia intestinalis. For more than five decades, the treatment of this disease has been based on compounds such as nitroimidazoles and benzimidazoles. The parasite's adverse effects and therapeutic failure are largely recognized. Therefore, it is necessary to develop new forms of chemotherapy treatment against giardiasis. Lysine deacetylases (KDACs), which remove an acetyl group from lysine residues in histone and non-histone proteins as tubulin, are found in the Giardia genome and can become an interesting option for giardiasis treatment. In the present study, we evaluated the effects of 4-[(10H-phenothiazin-10-yl)methyl]-N-hydroxybenzamide, a new class I/II KDAC inhibitor, on G. intestinalis growth, cytoskeleton, and ultrastructure organization. This compound decreased parasite proliferation and viability and displayed an IC

Giardia intestinalis presents an intriguing endomembrane system, which includes endoplasmic reticulum and peripheral vesicles (PVs). The PVs have previously been considered to be organelles that display early and late endosomal and lysosomal properties. Some of these vesicles accumulate macromolecules ingested by the protozoan and show acid phosphatase activity. It has been previously shown that the parasite releases microvesicles, which contribute to giardiasis pathogenesis; however, the vesicles’ origin and the way in which they are released by the parasite still remain unclear. In this study, we induced the parasites to encyst in vitro and analyzed these events using advanced electron microscopy techniques, including focused ion beam and electron microscopy tomography followed by three-dimensional reconstruction, in order to better understand protozoal multivesicular body (MVB) biogenesis. In addition, we performed an ultrastructural analysis of phosphatase activity during differentiation. We demonstrated that some vegetative trophozoites’ PVs exhibited morphological characteristics of MVBs with a mean diameter of 50 nm, containing intraluminal vesicles (ILVs).

The use of nanoparticles (NPs) in the healthcare market is growing exponentially, due to their unique physicochemical properties. Titanium dioxide nanoparticles (TiO2 NPs) are used in the formulation of sunscreens, due to their photoprotective capacity, but interactions of these particles with skin cells on the nanoscale are still unexplored. In the present study we aimed to determine whether the initial nano–biological interactions, namely the formation of a nano–bio-complex (other than the protein corona), can predict rutile internalization and intracellular trafficking in primary human fibroblasts and keratinocytes. Results showed no significant effect of NPs on fibroblast and keratinocyte viability, but cell proliferation was possibly compromised due to nano–bio-interactions. The bio-complex formation is dependent upon the chemistry of the biological media and NPs’ physicochemical properties, facilitating NP internalization and triggering autophagy in both cell types. For the first time, we observed that the intracellular traffic of NPs is different when comparing the two skin cell models, and we detected NPs within multivesicular bodies (MVBs) of keratinocytes. These structures grant selected input of molecules involved in the biogenesis of exosomes, responsible for cell communication and, potentially, structural equilibrium in human tissues. Nanoparticle-mediated alterations of exosome quality, quantity and function can be another major source of nanotoxicity.

Toxoplasma gondii is a protozoan parasite that is the causative agent of toxoplasmosis, an infection with high prevalence worldwide. Most of the infected individuals are either asymptomatic or have mild symptoms, but T. gondii can cause severe neurologic damage and even death of the fetus when acquired during pregnancy. It is also a serious condition in immunodeficient patients. The life-cycle of T. gondii is complex, with more than one infective form and several transmission pathways. In two animated videos, we describe the main aspects of this cycle, raising questions about poorly or unknown issues of T. gondii biology. Original plates, based on electron microscope observations, are also available for teachers, students and researchers. The main goal of this review is to provide a source of learning on the fundamental aspects of T. gondii biology to students and teachers contributing for better knowledge and control on this important parasite, and unique cell model. In addition, drawings and videos point to still unclear aspects of T. gondii lytic cycle that may stimulate further studies. Graphical Abstract

Giardia intestinalis, the causative agent of giardiasis, has complex cytoskeleton organization with structures involved in motility, adhesion, cell division, and cell differentiation. Microtubules are key components of the cytoskeleton and are the main elements of the ventral disc, median body, funis, in addition to four pairs of flagella. These cytoskeletal elements are basically stable microtubule arrangements. Although tubulins are the main proteins of these elements, molecular and biochemical analyses of Giardia trophozoites have revealed the presence of several new and not yet characterized proteins in these structures, which may contribute to their nanoarchitecture (mainly in the ventral disc). Despite these findings, morphological data are still required for understanding the organization and biogenesis of the cytoskeletal structures. In the study of this complex and specialized network of filaments in Giardia, two distinct and complementary approaches have been used in recent years: (a) transmission electron microscopy tomography of conventionally processed as well as cryo-fixed samples and (b) high-resolution scanning electron microscopy and helium ion microscopy in combination with new plasma membrane extraction protocols. In this review we include the most recent studies that have allowed better understanding of new Giardia components and their association with other filamentous structures of this parasite, thus providing new insights in the role of the cytoskeletal structures and their function in Giardia trophozoites.

Giardia intestinalis, the causative agent of giardiasis, has complex cytoskeleton organization with structures involved in motility, adhesion, cell division, and cell differentiation. Microtubules are key components of the cytoskeleton and are the main elements of the ventral disc, median body, funis, in addition to four pairs of flagella. These cytoskeletal elements are basically stable microtubule arrangements. Although tubulins are the main proteins of these elements, molecular and biochemical analyses of Giardia trophozoites have revealed the presence of several new and not yet characterized proteins in these structures, which may contribute to their nanoarchitecture (mainly in the ventral disc). Despite these findings, morphological data are still required for understanding the organization and biogenesis of the cytoskeletal structures. In the study of this complex and specialized network of filaments in Giardia, two distinct and complementary approaches have been used in recent years: (a) transmission electron microscopy tomography of conventionally processed as well as cryo-fixed samples and (b) high-resolution scanning electron microscopy and helium ion microscopy in combination with new plasma membrane extraction protocols. In this review we include the most recent studies that have allowed better understanding of new Giardia components and their association with other filamentous structures of this parasite, thus providing new insights in the role of the cytoskeletal structures and their function in Giardia trophozoites.

Background information The costa is a prominent striated fiber that is found in protozoa of the Trichomonadidae family that present an undulating membrane. It is composed primarily of proteins that have not yet been explored. In this study, we used cell fractionation to obtain a highly enriched costa fraction whose structure and composition was further analyzed by electron microscopy and mass spectrometry. Results Electron microscopy of negatively stained samples revealed that the costa, which is a periodic structure with alternating electron-dense and electron-lucent bands, displays three distinct regions, named the head, neck and body. Fourier transform analysis showed that the electron-lucent bands present sub-bands with a regular pattern. An analysis of the costa fraction via one- and two-dimensional electrophoresis and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) allowed the identification of 54 hypothetical proteins. Fourteen of those proteins were considered to be major components of the fraction. Conclusions The costa of T. foetus is a complex and organized cytoskeleton structure made of a large number of proteins which is assembled into filamentous structures. Some of these proteins exhibit uncharacterized domains and no function related according to gene ontology, suggesting that the costa structure may be formed by a new class of proteins that differ from those previously described in other organisms. Seven of these proteins contain prefoldin domains displaying coiled-coil regions. This propriety is shared with proteins of the striated fibers of other protozoan as well as in intermediate filaments. Significance Our observations suggest the presence of a new class of the cytoskeleton filaments in T. foetus. We believe that our data could auxiliate in determine the specific locations of these proteins in the distinct regions that compose the costa, as well as to define the functional roles of each component. Therefore, our study will help in the better understanding of the organization and function of this structure in unicellular organisms. This article is protected by copyright. All rights reserved

The mitosome is a double-membrane enveloped organelle that is found in few unicellular eukaryotes, one of which is the human intestinal parasitic protist Giardia intestinalis , which also lacks mitochondria and peroxisomes. This flagellated protist grows in vitro as trophozoites and under some conditions, differentiates into cysts, which are characterized by the absence of externalized flagella, a round shape, and the presence of a cyst wall. The presence and distribution of mitosomal proteins, such as giardial iron-sulfur cluster protein ( GiIscU ), heat-shock protein 70 (mit-HSP70) and giardial chaperonin 60 ( GiCpn60 ), during the process of trophozoite-to-cyst transformation was tracked using confocal laser scanning microscopy and western blotting. During the early stages of the differentiation process (∼12 h), there was a significant decrease in the extent of chaperone labeling in the cells, which disappeared after 21 h but was recovered during the cyst stage; IscU labeling remained present throughout the differentiation process. This finding was confirmed by mRNA expression analysis, thus indicating that a process modulates the expression of mitosomal chaperones during the G . intestinalis life cycle. Microscopy techniques, such as structured illumination and electron tomography, revealed a novel profile for central mitosomes, as well as the presence of both rounded and elongated mitosomes.

Acute graft-versus-host disease (aGVHD) is the main complication of allogeneic hematopoietic stem cell transplantation, and many efforts have been made to overcome this important limitation. We showed previously that G-CSF treatment generates low-density splenic granulocytes that inhibit experimental aGVHD. In this article, we show that aGVHD protection relies on incoming IL-10+ neutrophils from G-CSF–treated donor spleen (G-Neutrophils). These G-Neutrophils have high phagocytic capacity, high peroxide production, low myeloperoxidase activity, and low cytoplasmic granule content, which accounts for their low density. Furthermore, they have low expression of MHC class II, costimulatory molecules, and low arginase1 expression. Also, they have low IFN-γ, IL-17F, IL-2, and IL-12 levels, with increased IL-10 production and NO synthase 2 expression. These features are in accordance with the modulatory capacity of G-Neutrophils on regulatory T cell (Treg) generation. In vivo, CD25+ Treg depletion shortly after transplantation with splenic cells from G-CSF–treated donors blocks suppression of aGVHD, suggesting Treg involvement in the protection induced by the G-Neutrophils. The immunocompetence and specificity of the semiallogeneic T cells, long-term after the bone marrow transplant using G-Neutrophils, were confirmed by third-party skin graft rejection; importantly, a graft-versus-leukemia assay showed that T cell activity was maintained, and all of the leukemic cells were eliminated. We conclude that G-CSF treatment generates a population of activated and suppressive G-Neutrophils that reduces aGVHD in an IL-10– and Treg-dependent manner, while maintaining immunocompetence and the graft versus leukemia effect.

Trichomonas vaginalis is a protozoan parasite that causes trichomoniasis in humans, the most prevalent non-viral sexually transmitted disease (STD). Imidazole compounds are used for the treatment of trichomoniasis, and metronidazole is the most commonly prescribed. However, these compounds can lead to parasite resistance and unwanted side effects. Therefore, there is a need for an alternative treatment for this disease. Here, we explored the potential of clotrimazole (CTZ) and zinc compounds, as well as CTZ complexed with zinc salts ([1] acetate [Zn(CTZ)2(Ac)2] and [2] a chloride [Zn(CTZ)2Cl2] complexes) against T. vaginalis. We synthesized the zinc complexed CTZ compounds and determined their concentration values that inhibited parasite growth by 50% (IC50). We used scanning and transmission electron microscopy to visualize the ultrastructural alterations induced by CTZ and their zinc complexes. The incubation of the parasites with [Zn(CTZ)2(Ac)2] complex inhibited their growth, yielding an IC50 of 4.9 µm. Moreover, there were changes in the shape of treated parasites, including the formation of surface projections that subsequently detached from the cell, in addition to changes in the hydrogenosomes, endoplasmic reticulum and Golgi complex. We found [Zn(CTZ)2(Ac)2] to be a highly effective compound against T. vaginalis in vitro, suggesting its potential utility as an alternative chemotherapy for trichomoniasis.

Proteasomal proteolysis is required for a wide range of cellular processes, including protein quality control, cell cycle progression, cell death and metabolic adaptation to environment changes or stress responses. Proteasome inhibitors are useful compounds for determining the roles of proteasome in eukaryotic cells. Here, we investigated the effects of gliotoxin, a proteasome inhibitor, on the cell growth, replication, ultrastructure, DNA integrity and proteasomal proteolytic activity of the protist parasite Tritrichomonas foetus. The effect of gliotoxin on the transformation of T. foetus to endoflagellar form (EFF), also known as pseudocyst, was investigated. Gliotoxin inhibited the culture growth, arrested cell cycle, and provoked a trichomonacidal effect in a dose-dependent manner. Parasites treated with gliotoxin displayed features typical of cell death, such as membrane blebbing, concentric membrane whorls containing remnants of organelles, intense cytosolic and nuclear vacuolisation, chromatin condensation, DNA fragmentation, cytoplasmic disintegration and plasma membrane disruption. The proteasomal peptidase activity was inhibited by gliotoxin in a dose-dependent manner. Gliotoxin treatment also induced an irreversible EFF transformation in a dose/time-dependent manner. We compared morphological characteristics between gliotoxin- and cold-induced EFF parasites. Our results suggest that gliotoxin could induce EFF transformation by a mechanism distinct from that provoked by cold temperature. This study further contributes to a better understanding of the role of proteasome system in cell cycle, cell death and EFF transformation in T. foetus.

The unique composition of tumor-produced extracellular matrix (ECM) can be a determining factor in changing the profile of endothelial cells in the tumor microenvironment. As the main receptor for ECM proteins, integrins can activate a series of signaling pathways related to cell adhesion, migration, and differentiation of endothelial cells that interact with ECM proteins. We studied the direct impact of the decellularized ECM produced by a highly metastatic human melanoma cell line (MV3) on the activation of endothelial cells and identified the intracellular signaling pathways associated with cell differentiation. Our data show that compared to the ECM derived from a human melanocyte cell line (NGM-ECM), ECM produced by a melanoma cell line (MV3-ECM) is considerably different in ultrastructural organization and composition and possesses a higher content of tenascin-C and laminin and a lower expression of fibronectin. When cultured directly on MV3-ECM, endothelial cells change morphology and show increased adhesion, migration, proliferation, and tubulogenesis. Interaction of endothelial cells with MV3-ECM induces the activation of integrin signaling, increasing FAK phosphorylation and its association with Src, which activates VEGFR2, potentiating the receptor response to VEGF. The blockage of αvβ3 integrin inhibited the FAK-Src association and VEGFR activation, thus reducing tubulogenesis. Together, our data suggest that the interaction of endothelial cells with the melanoma-ECM triggers integrin-dependent signaling, leading to Src pathway activation that may potentiate VEGFR2 activation and up-regulate angiogenesis. J. Cell. Physiol. 231: 2464-2473, 2016. © 2016 Wiley Periodicals, Inc.