Outer membrane vesicles (OMVs) are vesicle-like spherical structures formed spontaneously by Gram-negative bacteria (Bitto and Kaparakis-Liaskos, 2017; Toyofuku et al., 2019) during normal growth (Schwechheimer and Kuehn, 2015). OMVs secretion is a strain-specific and selective process (Jan, 2017). OMVs generated from the Gram-negative bacterial outer membranes are spherical lipid bilayer nanostructures range in size from 20 to 300 nm (Jan, 2017). As illustrated in Figure 1, they comprise a range of parent bacterial-derived components (Li et al., 2020), which are mostly enzymes, bacterial-specific antigens, virulence factors, and PAMPs such as LPS, peptidoglycan, lipoproteins, bacterial DNA, RNA (Kaparakis-Liaskos and Ferrero, 2015). The outermost layer of OMVs includes the majority of the pathogenic compounds found in Gram-negative bacteria, and it is composed of three layers: lipid A, core polysaccharide layer, and O-antigen (Hao et al., 2015). OMVs from Gram-negative bacteria have lipopolysaccharide and outer membrane proteins incorporated in their membranes (Bitto and Kaparakis-Liaskos, 2017).

OMVs are often formed by blebbing of the outer membrane (Kulp et al., 2015; Orench-Rivera and Kuehn, 2016; Roier et al., 2016), in which the cell wall component, peptidoglycan production, is disrupted or hydrophobic molecules are injected into the outer membrane, damaging the cell envelope, and OMVs protrude from the outer membrane area. Disruption of the crosslink between the peptidoglycan and the outer membrane, resulting in their separation. Moreover, the buildup of peptidoglycan fragments or misfolded proteins has been observed in the literature, resulting in bulking pressure on the outer membrane and, eventually, OMVs freedom (Henriquez and Falciani, 2023). The population-sensing quinolone signaling (PQS) molecules induce OMVs production by inserting into the outer membrane and mediating its packing and transport, and when PQS in the outer leaflets is cumulative thought to change curvature (Florez et al., 2017). Insertion of hexadecane, 1-phenylbutadiene, and toluene into the outer membrane’s hydrophobic region causes changes in membrane curvature and stimulation of OMVs production (Shetty and Hickey, 2014). Because the inner membrane is still intact, cytoplasmic components cannot come into direct contact with these OMVs (See Figure 1B; Table 1). Gentamicin and mucin also promote OMVs formation (Fulsundar et al., 2014). Hyper vesiculation was caused by deacylation of lipid A and concomitant outer membrane remodeling in Salmonella typhimurium, as well as accumulation of phospholipids in the outer leaflet of the outer membrane of Haemophilus influenzae and Vibrio cholera (Elhenawy et al., 2016; Roier et al., 2016). Iron restriction can regulate OMVs formation in Escherichia coli, V. cholera and H. influenzae by affecting the expression of phospholipid transport genes (Roier et al., 2016).

Outer and inner membrane vesicles (OIMVs) can be produced by explosive cell lysis and blebbing. Endolysin is commonly utilized by ds-DNA phages to split their bacterial hosts, and DNA damage stress induces endolysin expression, resulting in peptidoglycan layer degradation (Turnbull et al., 2016). In contrast to OMVs, peptidoglycan is degraded, cells explode, and membrane fragments aggregate and self-assemble into outer and inner membrane vesicles (OIMVs) and explosive outer membrane vesicles (EOMVs) (See Table 1). Meanwhile, CMVs are released because of phage-triggered cell lysis (Tzipilevich et al., 2017). Under hypoxic condition, explosive cell lysis can be induced (Toyofuku et al., 2014). Endolysin weakens the bacterial peptidoglycan layer. The inner membrane extrudes into the peripheral cytoplasm, allowing inner cytoplasmic matters like DNA entering the vesicles, which ultimately extrude from the bacterial inner cytoplasmic to extracellular space with the surrounding outer membrane. The bacteria produce both EOMVs and OIMVs, but the OIMVs contain more DNA (see Figure 1). The SOS reaction causes phage-encoded endolysins to be expressed to accelerate vesicle formation, and this pathway appears to be the predominant route of OIMV production (Bauwens et al., 2017). Exposure to DNA-damaging chemicals or UV radiation, as well as quinolones such as ciprofloxacin, might trigger the SOS response (Bernier et al., 2013).

Gram-positive bacteria Bacillus subtilis are capable of releasing cytoplasmic membrane vesicles containing PBSX phage particles. PBSX-encoded endolysins (Toyofuku et al., 2017) or cell wall-weakening antibiotics such as β-lactam (Biagini et al., 2015) lead a peptidoglycan layer to form a pore through which the cytoplasmic membrane protrudes to produce CMVs. Endolysins in thick-walled Gram-positive bacteria promote partial hydrolysis of the bacterial cell wall, a condition known as “bubbling cell death.” CMVs formation in surrounding bacteria are triggered by endolysin released after death (Toyofuku et al., 2017; see Figure 1D).

This is a tubular protrusion of the outer membrane of Gram-negative bacteria or a localized cleavage of the Gram-positive cell wall resulting in a protrusion of the cytoplasmic membrane (Dubey et al., 2016), which frequently decorates on the surface of the resulting cells as well connects with cells to allow the exchange of diverse cellular components (Baidya et al., 2018). This tubular structure has received less attention, been less studied, and the precise method of its development is still unknown.

Bacterial extracellular vesicles are somewhat different from extracellular vesicles (EVs). EVs are tiny biological nano-vesicles, approximately 30–3,000 nm in diameter, released into most extracellular matrix and biological fluids (Théry et al., 2018). Among the classifications of vesicles, the most common are exosomes, microvesicles, and apoptotic vesicles (Gavard, 2023). EVs contain parental cell-derived proteins, mRNA, miRNA, lipids, and small molecule metabolites, etc., which can be transported to the recipient cells for action, mediating intercellular communication (van Niel et al., 2022). Thus, EVs regulate various critical physiological and pathological processes, such as immune regulation. They can be used as disease diagnostic markers, therapeutic agents, drug targets, and drug carriers (Zhu et al., 2017; Xu et al., 2018). EVs are essential in antimicrobial defense, allergy, autoimmune, and antitumor immune responses, contributing to organ development and progress in cancer therapy.

For membrane vesicle separation, Ultracentrifugation (UC) has been deemed the method of choice (Momen-Heravi et al., 2013). It works by applying centrifugal force to a solution of macromolecules and exploiting the buoyancy of the particles to deposit them in the order of their density. As the speed of centrifugal force increases, apoptotic vesicles, and cellular debris, as well as loosened vesicles, pellet one after another. Centrifugation at 300–400 × g for 10 min allows for cells precipitation, cell debris removal at 2,000 × g, and removal biopolymers and apoptotic vesicles at 10,000 × g (Lobb et al., 2015). The vesicles in the supernatant are precipitated by ultracentrifugation at 100,000-200,000 × g for 2 h (Xu et al., 2016). The effectiveness of centrifugal separation affected by several factors, including gravity acceleration (g), rotor settling angle, rotation radius, pelleting efficiency (rotor and tube k-factor), and viscosity of the separation fluid (Livshits et al., 2015; Abramowicz et al., 2016). As a result, when using and correcting the ultracentrifugation protocol to obtain less mix other impurities vesicles, these influencing factors should be considered.

To increase the intensity of particle separation, Density gradient ultracentrifugation (DGUC) is based on buoyant density (Hogan et al., 2014). This technique can separate subcellular components like mitochondria, peroxisomes, and nucleosomes. To form a gradient, density gradient ultracentrifugation uses either continuous density gradients (formed during centrifugation or pre-centrifugation) or stepwise gradients (density increases in a discrete manner) (Webber and Clayton, 2013). However, the final vesicle product may contain significant non-MV protein contamination using this method (See Table 2). Two double sucrose layers containing 1 and 2 mol/L sucrose can be used with DGUC (Raj et al., 2012). MVs are fractionated into the layer containing 1 mol/L sucrose, whereas the layer containing 2 mol/L sucrose contains more giant vesicles. The standard density gradient ultracentrifugation protocol with two sucrose layers produces a higher purity MVs preparation (Lobb et al., 2015; Abramowicz et al., 2016).

Based on same-diameter pores and a narrow range of pore size distribution, Ultrafiltration (UF) is used to simplify the separation of specific-size particles. Researchers frequently combine MVs separation with microfiltration or ultrafiltration. Ultrafiltration, in particular, can exist in successive stages of ultracentrifugation (Campoy et al., 2016). Ultrafiltration has been shown to separate MVs preparations with low levels of MVs proteins, such as aquaporins and nephrin, from those with high levels of non-MV proteins, such as albumin and 1-antitrypsin (Kim et al., 2022). Ultrafiltration is a quick, easy method that does not require any expensive equipment (Lobb et al., 2015). However, Alvarez et al. (2012) demonstrated that ultrafiltration produces less MVs and less pure RNA (including microRNA) than ultracentrifugation and PEG precipitation (See Table 2).

Size-exclusive chromatography (SEC) employs gentle physical conditions to efficiently separate the gel from a large number of soluble macromolecules present in the biological sample, while preserving vesicle integrity and structure (Taylor and Shah, 2015). SEC gelation separates molecules with different hydrodynamic radii, and can also separate MVs from plasma, urine protein complexes, and lipoproteins (Gamez-Valero et al., 2016). Small hydrodynamic radii components of the sample can pass through the pores, resulting in delayed elution, whereas large hydrodynamic components are prevented from entering the pores. However, in order to obtain MVs samples free of protein and lipoprotein impurities, MVs samples must be pretreated with ultracentrifugation or ultrafiltration. Immunoblot analysis of water channel protein-2, a typical microbubble protein, revealed that the chromatographic method is capable of separating a relatively large fraction when compared to the classical (Balaj et al., 2015).

Proteins, nucleic acids, viruses, and other small particles have long been precipitated by using precipitation polyethylene glycols (PEGs) of varying molecular weights. This method reduces the solubility of compounds in super hydrophilic polymers like polyethylene glycol solutions. Water-free polymers hold water and push insoluble components out of the solution. Samples containing MVs are typically incubated with a precipitation solution containing a polymer (e.g., PEG 8000) at low temperatures and for a relatively long period of time (overnight), after which the MV-rich precipitate is separated by low-speed centrifugation or filtration (Zeringer et al., 2015). PEG precipitation is a simple and fast method that does not distort MVs, can work in the physiological pH range, and is less dependent on ion concentration (Popović and de Marco, 2018). PEG-separated MVs have particle sizes is comparable to ultracentrifugation, ultrafiltration, and chromatography. At the same time, the number of MVs, specific protein molecules, and RNAs is typically much higher (Taylor and Shah, 2015; Andreu et al., 2016). The main disadvantage of this method is that the sample becomes contaminated with insoluble protein aggregates, viruses, and other particles (Lobb et al., 2015).

The method involves precipitating protein in acetone while retaining hydrophobic vesicles in the supernatant (Gallart-Palau et al., 2016). Supernatants containing MVs fractions were concentrated in a vacuum concentrator after samples were spiked four times the volume of cold acetone (−20°C). Western blot assays have revealed that PROSPR-isolated MVs had higher levels of expression of relevant markers than ultracentrifugation (Gallart-Palau et al., 2015). However, cold acetone causes a decrease in enzyme activity. Some scholars have also used saturated ammonium sulfate to precipitate proteins to extract MVs, and although the yield was not high, the enzyme activity was largely unaffected (Cui et al., 2021).

After describing the formation of MVs and the extraction method, the versatile of MVs will be described next. MVs can mediate microbe-microbe and host–microbe interactions, increasing viability and pathogenicity. Furthermore, they may activate macrophages and neutrophils, inhibit antigen presentation by dendritic cells, disrupt with the regulation of T and B cells, stimulate the secretion of inflammatory factors, and even cause cell death. The transport, virulence, inflammation, and immune interactions of MVs with host cells come next.

MVs are capable of delivering virulence factors and PAMPs. Pathogenic MVs can transfer toxins and PAMPs to host cells, altering host defense mechanism and regulating immunological responses, resulting in infectious diseases (Yoon, 2016). The most prevalent pathogenic factors detected in MVs are virulence factors, which convey a specific virulence factor to specific sections or distal areas of the host cell (As shown in Figure 2B). Pseudomonas aeruginosa MVs can transport bioactive compounds such as alkaline phosphatase, phospholipase-C, β-lactamase, and Cif into the cytoplasm (Schertzer and Whiteley, 2013). Pseudomonas aeruginosa MVs, according to Bomberger et al. (2009), can convey virulence factors to airway epithelial cells. Furthermore, the principal pathogenic toxins of V. cholera, including as CT and TCP, can be delivered to host cells in a physiologically active state via MVs (Elluri et al., 2014). Toxins are released by cells during MVs internalization and translocation, stimulating DNA-damaging processes and eventually leading to cell death (Bielaszewska et al., 2018). MVs may transfer lipids and other membrane components between cells simultaneously (Alves et al., 2016), providing them a significant edge in becoming drug carriers. CAGA protein can be transported to cellular endosomes and lysosomes by H. pylori outer membrane vesicles can transport via unique absorption processes (Parker and Keenan, 2012).

Bacterial adhesins in MVs compete for binding to host cells (Zhang et al., 2019a). Bacterial adherence to host cells is mediated by MV-carried sticky molecules such as Ata, BabA, SabA, OmpA, and others. MVs then enter the cell as previously stated (see Figure 2A). They bind to pattern recognition receptors (PRRs) once inside the body, activating the immune system and boosting the development of inflammatory reactions. The primary PRRs linked with MVs are Toll-like receptors (TLRs) on the cell surface and NOD-like receptors (NLRs) (Kaparakis-Liaskos and Ferrero, 2015) (see Figure 2A). Porphyromonas gingivalis MVs release adhesins (Tanabe et al., 2008), which drive cells to clump and promote biofilm formation in gingival plaque. MVs, for example, promote bacterial adhesion to the intestinal and respiratory epithelia, allowing microorganisms to resist physical clearance (Kim et al., 2013).

Following bacterial invasion, the bacteria will instantly produce massive amounts of MVs, which will encapsulate antibiotics and virulence factors and expel them from the body as a result of temperature fluctuations, oxidative stress, antibiotics, and other conditions. This method successfully eliminates hazardous components in bacteria while maintaining the interconnectedness of the bacterial membrane, hence safeguarding microorganisms (Romanowski et al., 2021). Lysogenic phages break down bacteria by recognizing target receptors on the cell membrane. MVs operate as decoys for lysogenic phages and antibacterial chemicals, providing immediate antibacterial protection (Balhuizen et al., 2021). Helicobacter pylori MVs aid pathogen immune evasion by boosting intracellular immunosuppressive cytokines and suppressing the inflammatory responses (Winter et al., 2014). Porphyromonas gingivalis MVs contain porphyrin peptide acylcarnitine deiminase (PPAD), which inhibit complement factor C5a function and permit P. gingivalis immune evasion (Bielecka et al., 2014).

Actin-dependent macropinocytosis is driven by the polymerization of actin rings in the subcellular membrane, which takes the form of a circular fold protrusion that finally closes at the top and encompasses some of the extracellular area (Bloomfield and Kay, 2016). Shigella fowleri has been demonstrated to employ this pathway during host cell invasion (Weiner et al., 2016) (shown by Figure 2A, route1).

The creation of clathrin-mediated endocytosis is caused by the formation of clathrin-encapsulated pits of up to 200 nm in diameter (shown by Figure 2A, route 2). In contrast to macrophage drinking, ligand attachment to cell surface receptors can trigger internalization (Rewatkar et al., 2015). Many bacterial virulence factors, including Shiga toxin, cholera toxin, and the gingival silver pain adhesin of Porphyromonas gingivalis, have been shown to enter host cells via clathrin-protein-mediated endocytosis (Neilands et al., 2019). Another mechanism by which Haemophilus pylori MVs can invade human gastric epithelial cells is clathrin-protein-mediated endocytosis (Parker et al., 2010). Although multiple investigations have connected this pathway MVs entry, suitable ligands have yet to be identified. If these interactions are necessary for MVs internalization, then identifying the components involved could lead to the design of inhibitors that minimize infection by preventing the distribution of virulence factors associated with MVs (O’Donoghue and Krachler, 2016).

Pseudomonas aeruginosa, Campylobacter jejuni, E. coli, and S. typhimurium all use caveolae protein-mediated endocytosis (shown by Figure 2A, route 3)to entry cells (Machado et al., 2012). It is believed that bacteria internalized by vesicles, as opposed to bacteria absorbed by grid protein-coated pits, are able to avoid being transported to lysosomes and subsequently being destroyed (O’Donoghue and Krachler, 2016). This suggests that pathogens may prefer this approach.

Numerous investigations have demonstrated that lipid rafts facilitate MVs penetration (shown by Figure 2A, route 4). Sphingolipid and cholesterol are abundant in the structural regions known as plasma membrane lipid rafts (Mulcahy et al., 2014). The cholesterol-rich regions of the bilayer aggregate and cause the membrane to bend, which leads to host cells invasions and particle entry into the cells (Wang et al., 2022). Early research established that the enterotoxin-producing E. coli MVs enter host cells via lipid rafts at different temperatures (Kesty et al., 2004). The MVs of P. gingivalis, H. influenzae, P. aeruginosa, and C. jejuni also enter their host epithelial cells using lipid rafts. Another investigation confirmed that H. pylori MVs use a cholesterol-dependent method to invade host cells (Olofsson et al., 2014).

Despite structural differences between the bilayers of MVs and host eukaryotic cells, membrane fusion has been described as a method for MVs entry into host cells (shown by Figure 2A, route 5). Using membrane-bound fluorescent dyes like rhodamine R-18, this has been shown to take place preferentially in the lipid raft structural domain in Legionella pneumophila (Jager et al., 2015).

By attaching to PRRs on the surface of immune cells via PAMPs, MVs can innate immune responses. MVs also activate intrinsic host sensing mechanisms (e.g., the inflammatory vesicle pathway) (Pathirana and Kaparakis-Liaskos, 2016), and in particular, MVs increase caspase-11-mediated atypical inflammatory vesicle responses via MVs bound LPS (Chen et al., 2018). Once internalized by host epithelial cells, MVs produced by bacteria such as Helicobacter pylori, Neisseria gonorrhoeae, and P. aeruginosa are recognized by the cytoplasmic immune receptor NOD1, making them susceptible to NOD1-dependent adaptive immune responses (Kaparakis et al., 2010; Irving et al., 2014).

MVs decrease DC responsiveness to cytokines associated with secondary bacterial antigens while inducing dendritic cell (DC) maturation and promoting antigen presentation (Kaparakis-Liaskos and Ferrero, 2015; Schetters et al., 2019) (see Figure 3). Neisseria meningitides MVs can aid in the development of DCs and encourage the synthesis of inflammatory mediators, including Salmonella spp. MVs cause DCs to express CD80, CD86, and MHC class II molecules. C-type lectin receptor (CLR) can facilitate DC attachment to integrin CD11c during antigen presentation, triggering the organism’s intrinsic and acquired immunity (Demento et al., 2011). Salmonella-derived MVs drive dendritic cells to express CD86 and TNF-α and IL-12 in the major histocompatibility complex (Imayoshi et al., 2011). Salmonella typhimurium MVs aid in the growth and maturation of BMDCs, which are inflammatory cytokine-producing cells generated from mouse bone marrow (Alaniz et al., 2007). By attaching to the host factor bactericidal permeability-increasing protein (BPI), N. meningitides MVs can facilitate their transport and internalization to dendritic cells (Schultz et al., 2007). A NOD1-dependent mechanism that polarizes T cells toward inflammatory Th2/Th17 responses is used by V. cholera O395 MVs to control epithelial pro-inflammatory responses and activate dendritic cells (Chatterjee and Chaudhuri, 2013).

MVs stimulates the expression of PMN (polymorphonuclear neutrophil) mRNA and produces pro-inflammatory cytokines and chemokines, such as TNF-α, IL-1β, IL-8, macrophage inflammatory protein 1α (MIP-1α), and MIP-1β. Neisseria meningitides MVs provoke an inflammatory reaction and PMN stimulation (Lapinet et al., 2000). Cytotoxic necrosis factor type 1 (CNF1), which hinders PMN chemotaxis and phagocytosis, is present in both non-pathogenic and uropathogenic E. coli MVs (Davis et al., 2006). In the lungs of mice, neutrophil migration was accelerated by E. coli MVs (Lee et al., 2018). MVs work with oral epithelial PRRs to activate cells and release inflammatory factors, such as IL-6, IL-8, MCP-1. These factors then entice neutrophils to infiltrate periodontal tissues, where they can eventually lead to the degeneration of periodontal tissues and the onset of chronic periodontitis (Cecil et al., 2019).

MVs in vivo testing result in beneficial cellular and humoral immune reactions (Prados-Rosales et al., 2011). The MVs of P. gingivalis diminish the expression of HLA-DR and MHC-II molecules in endothelial cells, consequently limiting antigen presentation and adaptive immunological responses in peripheral blood and inhibiting the influence on capillary formation (Cecil et al., 2019).

MVs contain a large number of PAMPs that stimulate the immune system, and APCs are necessary for the formation of adaptive immunological responses. APCs are activated by PAMPs found in MVs that bind with the appropriate pattern recognition receptors on APCs (Bachmann and Jennings, 2010). MVs have the necessary size to allow their entrance into lymphatic channels and efficient APCs absorption. Antigens delivered by MVs are presented by APCs (Liu et al., 2018; Tan et al., 2018). MVs with alloantigen on their surface, as revealed by Kuipers et al. (2015), handled antigens well and triggered cellular immunological response, which the physical binding of PAMPs may have enhanced. Inhibit antigen presentation and class antigen expression (Bomberger et al., 2014).

The suppression of T-cell immunity by MVs is manifested by the inhibition of cytokine secretion and T-cell proliferation. The mechanism of T cell suppression by H. pylori MVs was discovered to be stimulating monocytes to overexpress COX-2, IL-6, and IL-10 (Hock et al., 2017), inhibiting CD4+ T cells proliferation and inducing T cells apoptosis, resulting in immune modulation (Winter et al., 2014). Bacteroides fragilis MV-derived polysaccharides are sensed by intestinal dendritic cells via TLR2, promoting IL-10 expression, which then improves the function of regulatory T cells and suppresses intestinal inflammation (Chu et al., 2016). MVs activate immune tolerant cells from the intestine, which regulate other tissue-resident T cells and inflammation (Winter et al., 2014).

B-cell activation and antibody production are aided by MVs. MVs carry parental bacteria’s virulence molecules, giving them immunomodulatory capabilities to reduce B-cell resistance in vivo (Kuipers et al., 2018). MVs from Moraxella catarrhalis and H. influenzae could promote B cell proliferation (Deknuydt et al., 2014) and significantly increase IgG and IgM secretion from B cells by cross-linking with immunoglobulin, IgD BCR, or binding to TLR9 receptor. The IgD-binding (MID) protein on Moraxella MVs is required for B-cell receptor uptake (Perez Vidakovics et al., 2010), induction of IL-6 production, and increased IgM production following MVs B-cell uptake.

MVs can enter the cytoplasm of macrophages, leading to cellular inflammation and even death (Cecil et al., 2017; Fleetwood et al., 2017; Deo et al., 2018). Soluble VacA in H. pylori outer membrane vesicles induces apoptosis in gastric epithelial cells via a mitochondria-dependent pathway and generates reactive oxygen species (ROS), which induce cells to form autophagic vesicles, resulting in apoptosis (Yahiro et al., 2015). Macrophage death typically occurs 20 h after MVs treatment, whereas macrophages die rapidly within 35–48 h (Deo et al., 2020). The MVs of Gonococcal also trigger caspase-11-dependent apoptosis. Purified MV-mediated AMPK activation acts as an early warning system, initiating autophagy prior to bacterial invasion (Losier and Russell, 2019). MVs deliver LPS to the cytoplasm, stimulate IL-1β expression through caspase-11 dependence, and cause inflammation-induced programmed cellular scorching (Vanaja et al., 2016).

MVs avoid the degradation of encapsulated contents and enable for long-distance transport in vivo (Schertzer and Whiteley, 2013), and MVs can transport large amounts of nucleic acids to their functionally relevant destination. The findings indicated that MVs could enter the intestinal barrier and cause systemic inflammation via epithelial bypass (Jones et al., 2020). Pathogenic the LT intolerable toxin may be packaged by E. coli as MVs, while the CT cholera toxin can be packaged by V. cholerae (Chatterjee and Chaudhuri, 2011). These toxins cause watery diarrhea by altering intracellular cAMP levels, which causes cells to discharge more water into the colon (Kopic and Geibel, 2010). Helicobacter pylori MVs can cause micronuclei formation, altered iron metabolism and oxidative stress in human gastric epithelial cells (Chitcholtan et al., 2008). In a sulfate esterase-dependent way, MVs produced by commensal gut bacteria B can across the intestinal epithelial barrier of colitis-prone mice, causing intestinal inflammation (Hickey et al., 2015). By their virulence factors, E. coli MVs stimulate the host immune system, inducing sepsis by prompting host macrophages to generate IL-6 and TNF-α (Kim et al., 2018). Exposure of macrophage to MVs from Legionella pneumophila, Neisseria gonorrhoeae, uropathogenic E. coli, and P. aeruginosa promotes mitochondrial apoptosis and NLRP3 inflammatory vesicle activation (Deo et al., 2020). MVs produced by the intracellular pathogen, L. pneumophila, causing production of IL-6, IL-8, IFN-γ, MCP-1, and G-CSF, etc. Helicobacter pylori MVs increase the release of anti-inflammatory IL-10 from human peripheral blood mononuclear cells (PBMCs) (Fleetwood et al., 2017) while decreasing cytokine reactions, a result similarly found when P. gingivalis MVs interact with macrophages.

Porphyromonas gingivalis secrets MVs containing gingival pain circulate distant organs (Qing et al., 2020). MVs isolated from P. gingivalis can inhibit TLR4 and mTOR signaling, rendering monocytes nonresponsive to live bacteria. In addition, these MVs have been demonstrated to downregulate the expression of anti-atherogenic endothelial-type nitric oxide synthase (eNOS), while upregulating the expression of inducible nitric oxide synthase (iNOS) (Jia et al., 2015). Both foam cell production and platelet aggregation were induced by this MVs, both of which can contribute to the development of cardiovascular disease (Qi et al., 2003).

The addition of the MVs fraction of strain TK1402, which has a high biofilm-forming capability relative to other strains and is highly related with the generation of MVs, improves biofilm formation in of H. pylori strain (Yonezawa et al., 2009). Porphyromonas gingivalis MVs have been shown to increase the aggregation and adhesion of several other oral microbes in dental plaque biofilms, demonstrating the potential for MVs from one organism to promote the adhesion of another in biofilms (Kamaguchi et al., 2003). The attachment of whole-cell hyphae to epithelial cells was facilitated by the release of MVs carrying P. gingivalis (Inagaki et al., 2006). Research suggested that P. gingivalis MVs play a role in shaping the bacteria profile of periodontal plaque.

Pseudomonas aeruginosa vesicles stimulate the non-phagocytic NOD1 response as well as the production IL-8 synthesis in lung epithelial cells. MVs from other respiratory infections have been shown to trigger cytokine production. Klebsiella pneumoniae MVs cause epithelial cells to produce pro-inflammatory IL-8 and IL-1β after intratracheal injection, mimicking the inflammatory response in a neutropenic animal model of the disease (Bauman and Kuehn, 2006). Staphylococcus aureus MVs can migrate from the oropharynx to the skeletal system and enter osteoblasts and synovial cells via internalization (Chen et al., 2018), including cytogenic factors, such as GM-CSF and IL-6, to cause inflammation and bone tissues destruction. MVs produced by the intracellular pathogen L. pneumophila, causing production of IL-6, IL-8, IFN-γ, MCP-1, and G-CSF, etc.