**1. Introduction**

Lately, research has been increasingly focused on understanding of the biology of extracellular vesicles (EVs). Finding a more accurate name to define and classify EVs remains an open and, at the same time, a real challenge in the scientific world.

There are many reasons why it is difficult to find a very precise name for EVs: they are secreted by near all cell types in living organisms; the mechanisms through which they are released into the biological fluids are different and multiple; moreover, they have different sizes (30–2000 nm in diameter) which make the methods of obtaining and analyzing them to be diverse, but at the same time, some of them are slightly controversial. Once released from the cells, EVs are not inert particles, but they have complex functions in both physiological and pathological processes due to their specific cargo and factors stimulating their secretion. Thus, EVs are now viewed as early noninvasive biomarkers for various disorders in order to establish a correct diagnosis, but they also can be real targets for an effective treatment and, at the same time, valuable tools for treating several diseases such as diabetic cardiovascular diseases.

## **2. Terminology and biogenesis pathways of extracellular vesicles**

EVs are a large term used to define a variety of membrane-limited vesicles involved in the intercellular communication. A nomenclature has been proposed but there are still numerous papers using different terms for EVs [1–3]. The EVs comprise different types of vesicles, and based on the size, morphology, and mechanism of biogenesis, they are largely classified as: **exosomes and ectosomes**, also referred as shedding microvesicles (MVs) or microparticles (MPs) [4].

As for the apoptotic bodies, the researchers' opinions are different; some of them think that they can be included in the EV category and others do not include them. Apoptotic bodies result from cells undergoing programmed cell death (apoptosis) and could be identified in EV probes [5]. The large cellular fragments resulted from apoptosis are phagocyted by neighboring cells and recycled; therefore, they should not be regarded as EVs involved in intercellular communication.

**Exosomes** (50–100 nm) have been described since 1980s as "exfoliated membrane vesicles," which may serve as a physiologic function occurring in many normal and neoplastic cells [6]. An ultrastructural study [7] showed that about 50 nm small vesicles are exocyted from multivesicular bodies (MVBs) after receptormediated endocytosis. For reticulocytes, exosomes' exocytosis determines the loss of transferrin receptors during red cell maturation [8].

MVBs (**Figure 1A–D**) of 0.5–1 μm large vesicles containing 2–50 small intraluminal smaller vesicles belong to the endolysosomal compartment. This pleomorphic cellular compartment comprises early and late endosomes where a highly controlled molecular sorting mechanism drives MVBs to the lysosomes or to the extracellular space. During endosome maturation into late endosomes, inward budding from the limiting membrane of the endosome leads to the formation of intraluminal vesicles in MVBs [9]. Usually, MVBs fuse with lysosomes, the terminal compartment of the endocytic pathway, where they are digested and the final components are recycled. Some MVBs can fuse with the plasma membrane and their intralumenal vesicles are released from cells as exosomes. The process by which the fate of endosomal content is determined is not fully understood [10, 11]. Accumulating evidence suggests that the release of EVs often serves as an alternative disposal pathway to the overloaded lysosomes [12, 13]. This mechanism may be involved in a vascular calcification [14].

It is demonstrated that exosomes are not only cell specific but also they carry RNAs between cells and play major roles in intercellular communication [15]. How

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the extracellular space.

*Part One: Extracellular Vesicles as Valuable Players in Diabetic Cardiovascular Diseases*

RNAs reach the MVB vesicles is not clear, but it is supposed that cytosolic RNAs are taken up into intraluminal vesicles undergoing inward budding from the limiting

*Transmission electron microscopy of the extracellular vesicles in diabetic kidney. (A) Multivesicular body (MVB) with intraluminal vesicles in the cytoplasm of endothelial cell. (B) Numerous extracellular vesicles (square area) present between vascular smooth muscles cells (VSMCs) in vascular media. (C) Multivesicular* 

*cargos (MVCs) released by an endothelial cell (E) into the lumen of a peritubular capillary.* 

*(D) Multivesicular cargos (MVCs) released by a circulating lymphocyte (Ly).*

**Ectosomes (MVs or MPs)** are slightly larger vesicles (100–500 nm) compared with exosomes and are also cell specific as they are released from plasma membrane by budding. Ectosomes do not require exocytosis as they are generated by outward budding of a plasma membrane domain, which enclose a cargo gathered at the cytosolic face. The detachment of the ectosomes from the donor cells involves contraction of cortical actin beneath the plasma membrane [17]. These plasma membrane-derived vesicles are also reported to carry RNAs and proteins as an

Multivesicular cargos (**Figure 1A–D**) have also been described as EVs with a particular appearance: clustered vesicles (80–200 nm) shielded by plasma membrane [18]. This type of EVs has been described as mediating bone mineralization [19], vascular calcifications [20], or intercellular communication between telocytes [18], which often surround the vessels [21]. In our experience, endothelial cells (ECs) from diabetic kidney often release multivesicular cargos in the vascular lumen (**Figure 1C**). Possible mechanism of multivesicular cargo biogenesis based on electron microscopy images [18, 19] involves an initial aggregation of vesicles in the cortical cytoplasm which further will bulge a segment of the plasma membrane. Finally, gathered vesicles are released into the extracellular space as a cargo shielded by plasma membrane. The dissolution of the shielding membrane of the multivesicular cargo will release individual or grouped cytoplasmic-derived vesicles into

*DOI: http://dx.doi.org/10.5772/intechopen.85225*

membrane of the MVBs [9, 16].

**Figure 1.**

effective mechanism for intercellular communication.

*Part One: Extracellular Vesicles as Valuable Players in Diabetic Cardiovascular Diseases DOI: http://dx.doi.org/10.5772/intechopen.85225*

#### **Figure 1.**

*Extracellular Vesicles and Their Importance in Human Health*

vascular diseases.

(MPs) [4].

communication.

There are many reasons why it is difficult to find a very precise name for EVs: they are secreted by near all cell types in living organisms; the mechanisms through which they are released into the biological fluids are different and multiple; moreover, they have different sizes (30–2000 nm in diameter) which make the methods of obtaining and analyzing them to be diverse, but at the same time, some of them are slightly controversial. Once released from the cells, EVs are not inert particles, but they have complex functions in both physiological and pathological processes due to their specific cargo and factors stimulating their secretion. Thus, EVs are now viewed as early noninvasive biomarkers for various disorders in order to establish a correct diagnosis, but they also can be real targets for an effective treatment and, at the same time, valuable tools for treating several diseases such as diabetic cardio-

**2. Terminology and biogenesis pathways of extracellular vesicles**

EVs are a large term used to define a variety of membrane-limited vesicles involved in the intercellular communication. A nomenclature has been proposed but there are still numerous papers using different terms for EVs [1–3]. The EVs comprise different types of vesicles, and based on the size, morphology, and mechanism of biogenesis, they are largely classified as: **exosomes and ectosomes**, also referred as shedding microvesicles (MVs) or microparticles

As for the apoptotic bodies, the researchers' opinions are different; some of them think that they can be included in the EV category and others do not include them. Apoptotic bodies result from cells undergoing programmed cell death (apoptosis) and could be identified in EV probes [5]. The large cellular fragments resulted from apoptosis are phagocyted by neighboring cells and recycled; therefore, they should not be regarded as EVs involved in intercellular

**Exosomes** (50–100 nm) have been described since 1980s as "exfoliated membrane vesicles," which may serve as a physiologic function occurring in many normal and neoplastic cells [6]. An ultrastructural study [7] showed that about 50 nm small vesicles are exocyted from multivesicular bodies (MVBs) after receptormediated endocytosis. For reticulocytes, exosomes' exocytosis determines the loss

MVBs (**Figure 1A–D**) of 0.5–1 μm large vesicles containing 2–50 small intraluminal smaller vesicles belong to the endolysosomal compartment. This pleomorphic cellular compartment comprises early and late endosomes where a highly controlled molecular sorting mechanism drives MVBs to the lysosomes or to the extracellular space. During endosome maturation into late endosomes, inward budding from the limiting membrane of the endosome leads to the formation of intraluminal vesicles in MVBs [9]. Usually, MVBs fuse with lysosomes, the terminal compartment of the endocytic pathway, where they are digested and the final components are recycled. Some MVBs can fuse with the plasma membrane and their intralumenal vesicles are released from cells as exosomes. The process by which the fate of endosomal content is determined is not fully understood [10, 11]. Accumulating evidence suggests that the release of EVs often serves as an alternative disposal pathway to the overloaded lysosomes [12, 13]. This mechanism may be involved in a vascular

It is demonstrated that exosomes are not only cell specific but also they carry RNAs between cells and play major roles in intercellular communication [15]. How

of transferrin receptors during red cell maturation [8].

**112**

calcification [14].

*Transmission electron microscopy of the extracellular vesicles in diabetic kidney. (A) Multivesicular body (MVB) with intraluminal vesicles in the cytoplasm of endothelial cell. (B) Numerous extracellular vesicles (square area) present between vascular smooth muscles cells (VSMCs) in vascular media. (C) Multivesicular cargos (MVCs) released by an endothelial cell (E) into the lumen of a peritubular capillary. (D) Multivesicular cargos (MVCs) released by a circulating lymphocyte (Ly).*

RNAs reach the MVB vesicles is not clear, but it is supposed that cytosolic RNAs are taken up into intraluminal vesicles undergoing inward budding from the limiting membrane of the MVBs [9, 16].

**Ectosomes (MVs or MPs)** are slightly larger vesicles (100–500 nm) compared with exosomes and are also cell specific as they are released from plasma membrane by budding. Ectosomes do not require exocytosis as they are generated by outward budding of a plasma membrane domain, which enclose a cargo gathered at the cytosolic face. The detachment of the ectosomes from the donor cells involves contraction of cortical actin beneath the plasma membrane [17]. These plasma membrane-derived vesicles are also reported to carry RNAs and proteins as an effective mechanism for intercellular communication.

Multivesicular cargos (**Figure 1A–D**) have also been described as EVs with a particular appearance: clustered vesicles (80–200 nm) shielded by plasma membrane [18]. This type of EVs has been described as mediating bone mineralization [19], vascular calcifications [20], or intercellular communication between telocytes [18], which often surround the vessels [21]. In our experience, endothelial cells (ECs) from diabetic kidney often release multivesicular cargos in the vascular lumen (**Figure 1C**). Possible mechanism of multivesicular cargo biogenesis based on electron microscopy images [18, 19] involves an initial aggregation of vesicles in the cortical cytoplasm which further will bulge a segment of the plasma membrane. Finally, gathered vesicles are released into the extracellular space as a cargo shielded by plasma membrane. The dissolution of the shielding membrane of the multivesicular cargo will release individual or grouped cytoplasmic-derived vesicles into the extracellular space.
