The most important pro-angiogenic factor is VEGF. It also regulates endothelial cell function through the induction of nitric oxide, promoting vasodilation, decreased vascular tone, and reduced blood pressure. In the kidney, VEGF is located mainly in the podocyte, playing an important role in the maintenance and stability of the glomerular filtration barrier (18, 19). Situations that cause a reduction in VEGF expression are associated with endothelial and podocyte damage, leading to proteinuria, hypertension, and the TMA as a more severe presentation (18). The main scenarios associated with reduced VEGF expression are the use of anti-VEGF drugs, such as for the treatment of neoplasms, and preeclampsia, caused by a disproportionate increase in soluble fms-like tyrosine kinase-1 (produced in the process of inadequate placentation), which binds to circulating VEGF and placental growth factor, thus preventing them from interacting with their endothelial receptors (18). In recent years, many studies have also demonstrated the involvement of VEGF in the progression of chronic kidney disease and as a prognostic marker in glomerulopathies, including lupus nephritis and membranous nephropathy (19–22).

Angiotensin II (AngII) is the main effector of the RAS, being responsible for activating several mechanisms involved in vasoconstriction, as well as pro-oxidant and inflammatory pathways that affect endothelial cell function (23). The effects of AngII activity occur through the action of AngII receptors, mainly the angiotensin type 1 receptor (AT₁R). After binding to AT₁R, AngII exerts vasoconstrictive effects (from the release of Ca²⁺, leading to an increase in vascular tone), together with prothrombotic, pro-oxidant, and antifibrinolytic effects, as well as stimulating the expression of pro-inflammatory, atherogenic, and fibrogenic factors (24, 25).

It is known that AT₁R is present in various cells of the body, especially in the smooth muscle of arteries and arterioles. In the kidney, it is expressed not only in arteries and the endothelium but also in the medulla, proximal tubular epithelium (where it plays a role in sodium reabsorption), podocytes, and glomerular mesangial cells (26). Vasoconstriction of the efferent arteriole, caused by AngII, leads to increased intraglomerular pressure, resulting in proteinuria and glomerular sclerosis. In addition to the hemodynamic effect, the inflammatory stimulus is associated with progression to fibrosis and progression of kidney disease. There is increased expression of AngII and AT₁R in the renal interstitial cells of patients with progressive glomerulopathies, mainly in those with interstitial fibrosis (27). It is known that atrophy and tubulointerstitial fibrosis are important markers of the progression of kidney disease in many glomerulopathies. The use of AngII receptor blockers and AngII-converting enzyme inhibitors has proven benefits in effectively and safely reducing the progression of renal fibrosis, improving blood pressure control, as well as cardiovascular and renal outcomes (28–30).

The association between vascular injury and interstitial fibrosis could explain the fact that glomerulopathies with ischemic vascular lesions, such as IgAN, membranous glomerulopathy, FSGS, and LN, have worse prognoses than glomerulopathies without such lesions. Increased AT₁R expression plays a fundamental role in renal fibrogenesis, which also involves the participation of ET-1, another potent vasoconstrictor present in progressive glomerulopathies, the blockade of which reveals a renoprotective effect (31). Studies have shown that AngII acts to increase ET-1–induced vasoconstriction by increasing the expression of the ET-1 receptor and the binding between ET-1 and its receptor (32).

In addition, AngII regulates and increases the expression of transient receptor potential channel C6, a calcium-dependent channel associated with the slit diaphragm. In animal models, it has been shown that this increased activity through AT₁R leads to cytoskeletal disruption, podocyte damage, and glomerulosclerosis (33).

In 1999, Wallukat et al. identified the previously unknown agonist angiotensin II type 1 receptor autoantibody (AT1-AA) in the circulation of pregnant women with preeclampsia (34). It is primarily a member of the immunoglobulin G₃ antibody subclass that has an agonistic action in the AT₁R, thus exerting vasoconstrictive effects similar to those of AngII (25). Depending on the clinical scenario, it can present as one of the other immunoglobulin G subclasses. This antibody binds with high affinity to AT₁R, promoting permanent stimulation. It is known that renin, angiotensin, and aldosterone levels are reduced in pregnant women with preeclampsia. Activation of AT₁R by AT₁-AA would therefore explain the occurrence of systemic vasoconstriction and hypertension in preeclampsia, even with low levels of RAS components (25), as represented in Figure 2 .

Various studies have corroborated the correlation between AT₁-AA positivity and the occurrence of preeclampsia. Experimental animal models of pregnancy have shown that AT₁-AA injection triggers the manifestation of maternal preeclampsia syndrome, with hypertension, proteinuria, and glomerular endotheliosis, the main renal histopathological finding of endothelial injury in preeclampsia (35). Subsequent studies implicated AT₁-AA in vascular transplant rejection, malignant hypertension, glomerulopathies, scleroderma and other scenarios, as well as showing that it is associated with accelerated vascular senescence (34, 36–38).

Some studies have analyzed the role of AT₁-AA in glomerulopathies. In a study of patients with LN, the AT₁-AA positivity rate was 66% (39). In that study, AT₁-AA levels were higher among the patients without immunosuppression and with other signs of disease activity such as complement consumption and high anti-DNA titers. In a subsequent study (40), the association between AT₁-AA and vascular damage was analyzed in patients with LN. In that study, renal biopsies showed that medial layer hypertrophy and subintimal fibrosis were greater in the vessels of AT₁-AA–positive patients than in those of AT₁-AA–negative patients. One report described the case of an AT₁-AA–positive patient with LN who underwent kidney transplantation and developed non-human leukocyte antigen antibody-mediated rejection, together with collapsing FSGS (41). The authors reported that the patient presented a good response to treatment with plasmapheresis, immunoglobulin, and AT₁R blockade. It has been demonstrated that the levels of AT₁-AA are significantly higher in patients with LN than in those with other primary glomerulopathies, such as membranous glomerulonephritis, IgAN, and FSGS (38), although no significant differences have been detected in comparison with patients with cytoplasmic or perinuclear antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis. It is known that severe vascular injury, such as that provoked by TMA, is associated with worse outcomes in LN. The incidence of TMA is greater in LN with higher levels of activity, which somehow provides an immunological stimulus for vascular damage (42). The presence of AT₁-AA is correlated with higher levels of immunological activity in patients with LN (38). Therefore, AT₁-AA might play a central role in intense immunological stimulation and vascular damage.

Although it was originally described as an endothelium-derived vasoconstrictor, ET-1 is now known to play a role in cell proliferation, water–sodium balance, acid–base balance, tissue injury, fibrosis, and the progression of kidney disease, depending on which receptor is activated (43). The two main receptors are endothelin receptor type A (through which ET-1 promotes vasoconstriction, cell proliferation, and matrix accumulation) and endothelin receptor type B (through which ET-1 promotes antiproliferative, antifibrotic, and vasodilatory effects), being the type A expressed predominantly in vascular smooth muscle cells and the type B in vascular smooth muscle cells and endothelial cells, but both of which are also found in the mesangium, podocytes and tubules (44). Plasma ET-1 is increased in conditions of inflammation and vascular damage, correlating with albuminuria, and a high plasma ET-1 level is also an independent predictor of vascular dysfunction in chronic kidney disease (43). Treatment with selective endothelin A receptor blockers has been shown to provide a significant reduction in proteinuria, making it a useful tool in the treatment of FSGS, IgAN, and chronic kidney disease, mainly in combination with an AT₁R antagonist (45–48).

The complement system is a crucial part of the innate immune response, being responsible for clearing microorganisms, damaged cells, and immune complexes, as well as promoting inflammation and attacking the cell membranes of pathogens (49). The complement system can be activated through the lectin, classical, and alternative pathways leading to the generation of the C5b9, known as the membrane attack complex, as reviewed by Yoshida Y et al. (50). On the endothelial cell surface, C5b9 stimulates the secretion of von Willebrand factor, stimulates endothelial prothrombinase activity, and induces tissue factor expression. This process can involve nuclear factor kappa B, inflammatory cytokines, and culminates in an intense state of inflammation and microvascular coagulation (51).

Various proteins act as complement system regulators to prevent exacerbated activation of the complement system and protect autologous tissues, such as glomerular endothelial cells, against attack by the complement system. The main ones are membrane cofactor protein (MCP/CD46), Complement Factor H (CFH), Complement Factor I (CFI), C4b-binding protein, CR1, decay accelerating factor (DAF/CD55), and CD59 (50).

In aHUS, the dysfunction of regulatory or activating factors is associated with the pathogenesis of vascular damage, and the dosage of some components can show the complement system activation, although only genetics tests can define the constitutive complement system dysregulation (52). CFH is a fluid-phase protein, but it is also found on the cell surface, binding to glycosaminoglycans and sialic acid. In fact, CFH dysfunction is detected in aHUS and C3G, suggesting that it plays an important role in the pathogenesis of TMA in these glomerular disease (50). In some glomerulopathies, the serum level of many factors can be associated with active disease, such as C3a and C5a in ANCA-vasculitis, and soluble C5b-9 in C3G (52). MCP, CR1, CD55, and CD59 are expressed in the glomerulus and can be altered in several glomerular disorders, such as IgAN, lupus nephritis, membranous nephropathy, and primary membranoproliferative glomerulonephritis (53). More details on the mechanisms of complement-endothelial interactions in glomerulopathies are described in section 5.

Under physiological conditions, the endothelium maintains the environment in a more anticoagulant state by producing regulatory factors and through the action of the undamaged glycocalyx. The glycocalyx disruption exposes tissue factor, which triggers the coagulation cascade (1).

The coagulation system works together with platelets, leukocytes, and the complement system when endothelial cells are damaged. The coagulation and complement systems share common proteolytic pathways that enable an inflammatory response. In endothelial cells and neutrophils, C5a and C5b-9 induce tissue factor expression, initiating the extrinsic coagulation cascade. In addition, C5a induces secretion of von Willebrand factor and P-selectin, as well as increasing neutrophil adhesion in endothelial cells in culture. In addition, coagulation factors can activate the complement system at different levels (50). This interaction can be observed clinically in glomerular diseases associated with intense manifestations of TMA and antiphospholipid syndrome, including some with thrombomodulin and plasminogen variants (54).

The endothelial compartment comprises circulating endothelial cells (CECs), endothelial cell-derived microparticles (EMPs), and endothelial progenitor cells (EPCs). After endothelial injury, it is crucial that there is a balance between CECs and EMPs (released into the bloodstream after endothelial injury)—and the capacity for endothelial repair by EPCs. The activation of EMPs is also closely related to the activation of the alternative complement pathway, which can favor the development of TMA (55). The EPCs promote angiogenesis and microvascular repair through the production of VEGF, fibroblast growth factor 2, and angiopoietin (1).

In the setting of vascular injury with endothelial dysfunction, regardless of the etiology, markers of injury/activation (circulating endothelial cells and EMPs) are at high levels, while endothelial repair markers (endothelial progenitor cells and circulating angiogenic cells) are at low levels (1).

Classically, TMA is defined as a pathological lesion characterized by endothelial damage and the formation of microthrombi in small vessels (68). It can manifest clinically as microangiopathic hemolytic anemia, thrombocytopenia and ischemia (68). In the kidney, the acute finding of fibrin thrombus (often accompanied by fragmented red blood cells) is not always present. However, the presence of other morphological changes constitutes the aspect of microangiopathy. For example, in the active/acute phase the following lesions can be present: in glomeruli—endothelial edema, mesangiolysis ( Figure 3A ), and microaneurysms; in arterioles—endothelial/intimal edema, intramural fibrin, and myocyte necrosis; and in arteries—myxoid intimal edema and intramural fibrin. Electron microscopy (EM) shows glomerular endothelial cells with loss of fenestrations, fibrin tactoids with fragmented red blood cells and platelets, and expansion of the inner lamina. In the chronic phase, the main findings on light microscopy are as follows (68–70): double-contour sign in the glomerular capillary basement membrane ( Figure 3B ) - which appears in the EM as interposed cells with deposition of new matrix material; arteriolar hyalinosis; and thickening/fibrosis of the arterial intima, with an “onion-skin” appearance ( Figure 3C ). In the absence of a fibrin thrombus and of clinical signs of a TMA syndrome, there is still no consensus regarding how many of these other lesions must be present to confirm TMA or whether they would serve only to describe microangiopathy alone with or without thrombus. In such a situation, the diagnosis of TMA would be presumptive because other lesions are typically present in the classic form.

Arteriolar hyalinosis, also known as hyaline arteriolosclerosis, is the deposition of proteinaceous material with a hyaline appearance in the subendothelial region of arterioles (78). That deposition results in thickening of the vessel wall and narrowing of the lumen, leading to ischemia ( Figure 4A ). Arteriolar hyalinosis can be observed in healthy individuals as they age. However, it occurs earlier in patients with hypertension or diabetes mellitus.

Arteriosclerosis is characterized by thickening of the intimal layer by smooth muscle fibers or fibroblasts, collagen fibers and the fundamental connective tissue that causes narrowing of the vessel lumen ( Figure 4B ). It does not have the homogeneous appearance characteristic of arteriolar hyalinosis, with which it should not be confused (78). It is also commonly found in elderly individuals, individuals with hypertension, and individuals with diabetes.