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We review the morpho-functional basis of the different types of angiogenesis and report our observations, including the formation of angiogenesis-related secondary structures. First of all, we consider the following issues: a) conceptual differences between angiogenesis and vasculogenesis, b) incidence of angiogenesis in pre- and postnatal life, c) regions of vascular tree with angiogenic capacity, d) cells (endothelial cells, pericytes, CD34+ adventitial stromal cells of the microvasculature and inflammatory cells) and extracellular matrix components involved in angiogenesis, e) events associated with angiogenesis, f) different types of angiogenesis, including sprouting and intussusceptive angiogenesis, and other angiogenic or vascularization forms arising from endothelial precursor cells (postnatal vasculogenesis), vasculogenesis mimicry, vessel co-option and piecemeal angiogenesis. Subsequently, we consider the specific morpho-functional characteristics of each type of angiogenesis. In sprouting angiogenesis, we grouped the events in three phases: a) activation phase, which includes vasodilation and increased permeability, EC, pericyte and CD34+ adventitial stromal cell activation, and recruitment and activation of inflammatory cells, b) sprouting phase, encompassing EC migration (concept and characteristics of endothelial tip cells, tip cell selection, lateral inhibition, localized filopodia formation, basal lamina degradation and extracellular changes facilitating EC migration), EC proliferation (concept of endothelial stalk cells), pericyte mobilization, proliferation, recruitment and changes in CD34+ adventitial stromal cells and inflammatory cells, tubulogenesis, formation of a new basal lamina, and vascular anastomosis with capillary loop formation, and c) vascular remodelling and stabilization phase (concept of phalanx cells). Subsequently, the concept, incidence, events and mechanisms are considered in the other forms of angiogenesis. Finally, we contribute the formation of postnatal angiogenesis-related secondary structures: a) intravascular structures through piecemeal angiogenesis, including intravascular papillae in vessel tumours and pseudotumours (intravascular papillary endothelial hyperplasia, vascular transformation of the sinus in lymph nodes, papillary intralymphatic angioendothelioma or Dabska tumour, retiform hemangioendothelioma, hemangiosarcoma and lymphangiosarcoma), vascular septa in hemorrhoidal veins and intravascular projections in some tumours; b) arterial intimal thickening; c) intravascular tumours and pseudotumours (e.g. intravenous pyogenic granulomas and intravascular myopericytoma); d) vascular glomeruloid proliferations; and e) pseudopalisading necrosis in glioblastoma multiform.

We review the morphofunctional characteristics of CD34+ stromal fibroblastic/fibrocytic cells (CD34+ SFCs) and report our observations. We consider the following aspects of CD34+ SFCs: A) The confusing terms applied to this cell type, often combining the prefix CD34 with numerous names, including fibroblasts, fibrocytes, dendrocytes, keratocytes, telocytes and stromal, dendritic, adventitial, supraadventitial, perivascular, paravascular and delimiting cells; B) Changes in their immunophenotype, e.g., loss of CD34 expression and gain of other markers, such as those defining mesenchymal and derivate cells (myofibroblasts, osteoblasts, chondroblasts, adipocytes); C) Morphology (elongated or triangular cell body and thin, moniliform, bipolar or multipolar cytoplasmic processes), immunohistochemistry (co-expression of and changes in molecular expression) and structure (characteristics of nucleus and cytoplasmic organelles, and points of contact and junctions in quiescent and activated stages by light and electron microscopy); D) Location and distribution in the vessels (adventitia or external layer), in the tissues (connective, adipose, blood, muscle and nervous) and in the organs and systems (skin, oral cavity and oropharynx, respiratory, digestive, urinary, male, female, endocrine and lymphoid systems, serosal and synovial membranes, heart, eye and meninges); E) Origin from the mesoderm and cranial neural crest in the embryo, and from stem cells (themselves or other cells) and/or peripheral blood pluripotent stem cells (circulating progenitor cells) in post-natal life; F) Functions, such as synthesis of different molecules, progenitor of mesenchymal cells, immunomodulation, parenchymal regulation (growth, maturation and differentiation of adjacent cells), induction of angiogenesis, scaffolding support of other cells and phagocytic properties. Since CD34+ SFCs are the main reservoir of tissue mesenchymal cells (great mesenchymal potential, probably higher than that proposed for pericytes and other stromal cells), we dedicate a broad section to explain their in vivo behaviour during proliferation and differentiation in different physiologic and pathologic conditions, in addition to their characteristics in the human tissues of origin (adult stem cell niches); G) Involvement in pathological processes, e.g., repair (regeneration and repair through granulation tissue), fibrosis, tumour stroma formation and possible CD34+ SFC-derived tumours (e.g., solitary fibrous tumour, dermatofibrosarcoma protuberans, giant cell fibroblastoma, nuchal-type fibroma, mammary and extramammary myofibroblastoma, spindle and pleomorphic cell lipoma, and elastofibroma) and H) Clinical and therapeutic implications.

The administration of prostaglandin (PG) E2, triacetylglycerol and glycerol induce the formation of numerous vascular buds arising from the femoral vein, as previously demonstrated by our group. In the present study, a great number of peg-and-socket junctions (PSJs) between smooth muscle cells (SMCs) (providing the pegs) and ECs (forming the sockets) were demonstrated. At the first stage, days 1 to 3, PSJs connect subendothelial penetrating processes from activated SMCs with activated ECs of the intima. Subsequently, during angiogenesis (days 4 to 6), SMCs, showing transitional aspects with pericytes, also form PSJs with intimal ECs, but also new PSJs between SMCs and sprouting ECs in the media layer were now observed. Immunohistochemically, α-smooth muscle actin (α-SMA) and H-caldesmon are positive in the cytoplasm of the SMCs, showing a higher expression in pegs. Desmin, however, although it is also positive in the cytoplasm of the SMCs, is negative in the pegs. The expression of CD34 in ECs reveals abundant positive folding that appears to correspond to the sockets. The peculiar expression of caldesmon, whose isoforms may contribute to the regulation of cell motility, and to vasculogenesis and angiogenesis, may have a role in the different mechanisms by which PSJs act in the vein wall.

We review the morphofunctional characteristics of pericytes and report our observations. After a brief historical background, we consider the following aspects of pericytes: A) Origin in embryonic vasculogenesis (mesenchymal stem cells, neurocrest and other possible sources) and in embryonic and postnatal life angiogenesis (pre-existing pericytes, fibroblast/ myofibroblasts and circulating progenitor cells). B) Location in pericytic microvasculature and in the other blood vessels (including transitional cell forms and absence in lymphatic vessels), incidence (differences depending on species, topographical location, and type and stage of vessels) and distribution (specific polarities) in blood vessels. C) Morphology (cell body, and longitudinal and circumferential cytoplasmic processes), structure (nucleus, cytoplasmic organelles and distribution of microtubules, intermediate filaments and microfilaments) and surface (caveolae system). D) Basement membrane disposition, formation, components and functions. E) Contacts with endothelial cells (ECs) (peg and socket arrangements, adherent junctions and gap junctions) and with basal membrane (adhesion plaques). F) Molecular expression (pericyte marker identification). G) Functions, such as vessel stabilization, regulation of vascular tone and maintenance of local and tissue homeostasis (contractile capacity and vessel permeability regulation), matrix protein synthesis, macrophage-like properties, immunological defense, intervention in coagulation, participation in mechanisms that regulate the quiescent and angiogenic stages of blood vessels (including the behaviour of pericytes during sprouting angiogenesis and intussuceptive vascular growth, as well as pericyte interactions with endothelium and other cells, and with extracellular matrix) and plasticity, as progenitor cells with great mesenchymal potential, originating other pericytes, fibroblast/myofibroblasts, preadipocytes, chondroblasts, osteoblasts, odontoblasts, vascular smooth muscle and myointimal cells. This mesenchymal capacity is seen in a broad section on the perivascular mesenchymal cell niche hypothesis and in the concept of pericyte and EC "marriage and divorce". H) Peculiar pericyte types, such as hepatic stellate cells (Ito cells), bone marrow reticular cells and mesangial cells. I) Involvement in pathological processes, such as repair through granulation tissue, pericyte-derived tumors, tumor angiogenesis and tumoral cell metastasis, diabetic microangiopathy, fibrosis, atherosclerosis and calcific vasculopathy, lymphedema distichiasis, chronic venous insufficiency, pulmonary hypertension, Alzheimer disease and multiple sclerosis. J) Clinical and therapeutic implications (de-stabilization of vessels or formation of a stable vasculature).

In occluded femoral artery segments, intimal thickening occurred and abundant neovascularization from the surrounding microcirculation developed. Under these conditions, the contribution of vasa-vasorum as a source of supplementary population of cells during the early intimal thickening formation was studied. Using a technique that specifically labels venules, predominantly postcapillary venules, a marker-Monastral Blue B-was used as a tracer to follow the pericyte, endothelial cell and monocyte/macrophage lineages. In the first two days of the experiment, the marker was restricted to the wall of the periarterial microcirculation, being incorporated by endothelial cells, pericytes and some monocytes/macrophages crossing the venule walls. Later, the marker continues to be observed in some of the following cells: endothelial cells and pericytes of the newly-formed vessels, fibroblast-like cells, transitional cells between pericytes and fibroblast-like cells, macrophages migrating into the interstitium, myointimal cells and neoendothelial cells of the arterial lumen. These findings provide evidence that, during arterial intimal thickening formation in occluded arterial segments, the periarterial microvascularization contributes, in addition to recruited macrophages, newly-formed endothelial cells and a supplementary population of fibroblast-like cells and myointimal cells.

This study assesses the action of hypercortisolism on the hormone and peptide periadenoma region of removed ACTH-producing microadenoma. Our findings show that cortisol excess affects both ACTH and GH production, with no immunoreaction for these hormones. The remaining pituitary hormones (TSH, FSH and PRL) and POMC-derived peptides (betaEnd, alphaMSH and betaMSH) were not modified. Likewise, we observed pituitary immunoreactive cells for Neurotensin (NT), Intestinal vasoactive peptide (VIP), Substance P (SP) and Angiotensin-II (Ang-II). The colocalization demonstrated that NT was expressed in thyrotrope and gonadotrope cells, VIP in gonadotrope cells and SP in corticotrope cells. The results about Ang-II were inconclusive. On the other hand, immunoreaction for the NPY and Gal peptides were not present. In the adenomatous cells, the peptide NT is present in ACTH cells as well as SP. These results suggest a peptide regulation of pituitary cells in the pathological state that can differ between normal and tumoural cells of the same pituitary.

Adult stem cells (ASC)--able to self renew and to intervene in maintaining the structural and functional integrity of their original tissue--can express greater plasticity than traditionally attributed to them, adopting functional phenotypes and expression profiles of cells from other tissues. Therefore, they could be useful to regenerative medicine and tissue engineering. Transit-amplifying cells (TAC) are committed progenitors among the ASC and their terminally differentiated daughter cells. The ASC reside in a specialized physical location named niche, which constitutes a three-dimensional microenviroment where ASC and TAC are protected and controlled in their self-renewing capacity and differentiation. The niche can be located near or far from the recruitment point, requiring a short or long-distance cellular migration, respectively. This paper briefly reviews the current status of research about ASC plasticity, transdifferentiation, fusion and functional adaptation mechanisms. Subsequently, ASC and TAC occurrence, characteristics and location have been considered in the skin, cornea, respiratory tract, teeth, gastrointestinal tract, liver, pancreas, salivary glands, kidney, breast, prostate, endometrium, mesenchyma, bone marrow, skeletal and cardiac muscle, nervous system and pituitary gland. Moreover, the role of cancer ASC has also been revised.

An intense and significant neovascularization, with numerous capillaries growing into the media layer of the rat femoral artery, was demonstrated when glycerol was administered into the interstitium between the femoral vein and the femoral artery. The maximum microvascularization was observed at days 7 and 9 after glycerol administration. Afterwards, involution of the majority of the newly-formed microvessels in the arterial wall occurred. Other substances containing glycerol in their molecules, such as triacetyl-glycerol and tributyril-glycerol, failed to produce significant neovascularization in the media layer of the femoral artery. Neovascularization of the arterial wall was preceded by a considerable decrease in the number of the smooth muscle cells, which experienced apoptosis and necrobiosis, disappearing in extense areas of the arterial segment affected by glycerol. Coinciding with neovascularization and microvascular involution, repopulation of the media layer by smooth muscle cells was observed.

The neovascularization of the arterial wall in human and experimental pathology has been demonstrated. The occlusion of the of the rat femoral artery is a suitable model for the study of these angiogenesis processes. Newly formed capillaries growing into the arterial wall have been described in this model. The origin of these ingrowing capillaries has been attribute to the preformed surrounding venules and capillaries. The contribution of the adjacent femoral vein with a supplementary population of vascular sprouts could also be possible. To test this hypothesis in half of the occluded arteries, the adventitia was removed from the side facing the femoral vein. Between 1 and 3 days after surgery several alterations were found both in the endothelial cells and the smooth muscle cells of the tunica media. Between 3 and 6 days, solid or canalized endothelial sprouts were observed arising from the femoral vein. By days 4 and 6, newly formed capillaries grew into the adventitia and tunica media of the femoral artery. Some of them, penetrated the internal elastic lamina. This microvascular penetration from the femoral vein was more prominent in the area of the ostium of the collateral and when the adventitia was removed. Some ingrowing capillaries were in continuity with the endothelial cells of the arterial neointima. At days 7 and 8, regressing capillaries were observed in the neomicrovasculature network between artery and vein, with a selective loss of the smaller vessels. From day 9 onwards, fewer and larger vascular channels were present between the femoral vein and the femoral artery. An arterial neolumen contained what appeared to be circulating "fresh" blood. Quantitatively, the venous neocapillary density increased from days 4 to 6 and then declined significantly by day 8. The arterial neocapillary density increased form days 4 to 8 and declined significantly by day 12. Moreover, both densities were significantly greater when the arterial adventitia was removed. The perfusion with barium solution showed the presence of the contrast material in the newly formed vessels, the lumen of the femoral vein, and the neolumen of the occluded arterial segment. The present findings indicate that putative angiogenic molecules released form the occluded arterial segment may reach the adjacent wall of the vein inducing neovascularization from it. The vein vascular sprouts are connected to the ingrowing capillaries in the occluded arterial wall and to the neocapillaries form the preexisting pericytic microvasculature. When the arterial adventitia were removed up to 2 times greater vein neocapillary's density was observed suggesting an easily access of the putative angiogenic factors to the vein.

The specific contribution of the proximal and distal nerve stumps across an 8 mm gap within silicone chamber regeneration models was studied. For this, proximal and distal (Group A), distal and distal (Group B) and proximal and proximal (Group C) nerve stumps were placed in opposite ends of silicone chambers. In all the groups, a tissue cable forms between the nerve stumps, demonstrating that, without distinction, proximal or distal stumps can stimulate the growth of other proximal or distal stumps. Furthermore, in Group B, the newly formed pseudo-nerve, in the absence of regenerating axons, contains a number of Schwann cells significantly similar to Group A, which confirms that proliferation and migration of Schwann cells do not require axonal presence or contact. Likewise, the findings demonstrate that, with the exception of the axons, the distal stump contributes to the peripheral nerve regeneration in the same way as the proximal stump. Finally, when proximal stumps are placed in both the opposite ends of the silicone chamber, Schwann cells and regenerating axons grow into the chamber gap from both inserts, and myelination also proceeds from both ends to the centre of the chambers.

A hundred years after the first description, many aspects of pericytes remain to be examined. Mesenchymal in origin, pericytes form an incomplete envelopment around the endothelial cells and within the microvascular basement membrane of capillaries and postcapillary venules. Morphologically, they appear as long, slender, polymorphic cells, showing an elongated cell body, from which arise longitudinal and circumferential branches. Cell bodies and cytoplasmic processes of pericytes, as well as the endothelial cells, are enveloped by the same basal lamina, except for where they make direct contacts with each other. The pericyte/endothelial cell contacts are peg and socket, adhesion plaques and gap junctions, making up structural mechanisms for force transmission and a possible receptor system for cells, in which the pericyte and endothelial cells respond to secondary signals generated in the other cells. Electron microscopic studies have revealed an elaborate network of cytoplasmic filaments. Pericyte intermediate filament proteins show species and tissue differences, expressing vimentin or vimentin and desmin. The pericytes also express protein typical of contractile cells, i.e. smooth muscle-specific isoforms of actin and myosin, cyclic GMP-protein kinase and tropomyosin. A gradual transition is observed between pericytes and smooth muscle cells in both terminal arterioles and venules. Several general functions for the pericytes have been postulated: contractability; permeability regulator; integrity maintainer; endothelial cell growth modulator; and cell progenitor with considerable mesenchymal potential.

Segments of rat femoral arteries, with one collateral each, occluded between ligatures and dissected from surrounding tissue, developed intimal thickening, with or without ligation of their collaterals. Numerous newly-formed capillaries from the surrounding arterial microcirculation growing into the adventitia, tunica media and intimal thickening were demonstrated by means of serial longitudinal sections, predominantly in the ostium of the collateral. When the ligatures were applied without damaging the microcirculation surrounding the artery and the normal continuity of the adventitial vessels was unchanged, earlier presence of intimal thickening was observed. When the fibrous layers of the adventitia were removed at the moment of the arterial ligation, the continuity between newly-formed vessels of the neoadventitia and those growing into the media and neointima was much more evident. It was then noted that the pericytes constituted a major component of the intimal thickening. The introduction of contrast material in microcirculation confirmed the connections between newly-formed adventitial and intimal vessels. At the beginning of the experiment, autoradiographic studies showed an increased DNA synthesis in the cells of preformed postcapillary venules and capillaries of surrounding arterial microcirculation and later in those of the newly-formed vessels growing into the arterial wall. These results indicate that newly-formed capillaries derived from surrounding arterial microcirculation penetrate the wall of the occluded arterial segments and contribute to the intimal thickening formation. It is likely that the pericytes and endothelial cells (EC) of these ingrowing vessels are sources of myointimal cells at the intimal thickening and of endothelium at the luminal surface, respectively.