Pathophysiology of mesangial expansion in diabetic nephropathy: mesangial structure, glomerular biomechanics, and biochemical signaling and regulation

Mesangial structure and function

The mesangium was first discovered and described by Zimmerman in 1929 as consisting of mesangial cells and mesangial matrix [10]. Studies by Kimmelsteil and Wilson revealed the importance of this region by discovering the formation of nodules in diabetic patients, the first structural lesion to be identified in the kidney of the diabetic patient [11]. Electron microscope studies have since revealed the structural-functional relationship of this region to the surrounding glomerular tuft. In this section, we examine the structural and functional relationships in the mesangium between the mesangial cell, matrix, and GBM to show that the mesangial cell has functions important to the biomechanical homeostasis of the glomerulus. Mesangial cell dysfunction results in the disruption of the biomechanical homeostasis, which is a concept that is usually ignored in favor of biochemical homeostasis. In order to understand how glomerular failure occurs in diabetic nephropathy, it is imperative to understand the biomechanical structure and function of the mesangial cell and matrix in health and disease.

Structure of the mesangial cell

Early investigations uniquely identified the mesangial cell from the rest of the glomerular cells by its central position within the glomerulus [12]. Later, it was found to be associated with matrix synthesis in disease and thus distinguished from neighboring endothelial and visceral epithelial cells [12]. With the advent of the electron microscope, the mesangial cell was further characterized by its unique cytoplasmic structure that enables it to carry out its function [13]. Like most eukaryotic cells, the mesangial cell contains a nucleus, mitochondria, a golgi apparatus, and endoplasmic reticulum; however, its golgi apparatus and endoplasmic reticulum are underdeveloped, and its cytoplasm sparsely populated [13, 14].

In the quiescent state, the mesangial cell does not have a robust protein or matrix synthesis system [14]. In addition to the quiescent phenotype, the mesangial cell exhibits activated and hypertrophied phenotypes. The quiescent phenotype is characterized by a stellate cell shape, a low proliferation rate, and a sparse cytoplasm. The activated phenotype is characterized by an elongated cell shape, high proliferation rate, alpha smooth muscle actin (α-sma) expression, and synthesis of interstitial collagens (Table 1) [1417]. The activated cell phenotype is usually associated with diseased conditions where increased proliferation rates and synthesis of interstitial collagens are observed [15]. The activated mesangial cell is reminiscent of myofibroblasts in other tissues, which exhibit characteristics of both smooth muscle cells and fibroblasts such as the expression the of α-sma, responsiveness to vasoactive agents, and production of interstitial collagens [15, 17]. The hypertrophied phenotype is characterized by a lowered proliferation rate, a high expression of α-sma, and a polygonal cell shape [14, 18, 19].

Table 1 Markers and mediators of different mesangial cell phenotypes. MC: mesangial cell, FCS: fetal calf serum, TGF- β: transforming growth factor β, PDGF- β: platelet derived growth factor β, Coll: collagen. Note: there is disagreement in the literature regarding the expression level of α-sma by differerent mesangial cell phenotypes; here, we show that through two rows in the table for the corresponding expression levels

The most distinct features of mesangial cells are the cytoplasmic processes that extend out from the core of each mesangial cell towards the capillary lumen (Fig. 1) [1214, 17]. These processes contain cytoplasmic fibrils that are 7-10 nm in diameter [17] and enable the cell and its cytoplasmic processes to attach to the surrounding matrix and GBM. The ends of these cytoplasmic processes branch out around the capillary walls creating wide contact areas between the mesangial processes and endothelial cells [13]. These processes, however, are not attached to the endothelial membrane nor are they connected by specialized intermembrane connections [13]. Instead, these branched processes wedged in between the GBM and the endothelial membrane are tightly attached to the GBM by microfibrils or indirectly connected through the mesangial matrix [13].

Fig. 1figure 1

Mesangial cell structure with its processes extending out to multiple capillaries. MC: mesangial cell, MM: mesangial matrix, P: podocytes, EC: endothelial cells, C: capillaries, GBM: glomerular basement membrane. The small arrows indicate the mesangial cell/GBM connections that oppose the hydrostatic pressures shown by long red arrow. The T-shaped marking indicates the mesangial cell process where the body of the T-shaped marking represents the axis of the mesangial processes, and the head of the T-shaped marking indicates the mesangial angles. The mesangial cell occupies a majority of the mesangium and is closely attached to the GBM directly by microfibrillar attachments or indirectly to the GBM via the mesangial matrix. The T-shaped mesangial processes are firmly embedded in between the capillaries and the GBM and enable the mesangial cell to maintain glomerular structure. Modified after [75] with permission

The mesangial processes enable the mesangial cell to carry out its most important function—the stabilization of the glomerular capillary structure. Without such stabilization, the glomerulus would collapse and fail to carry out filtration. The discovery of these processes also fueled a longstanding debate about the role of mesangial cell contractility in regulating the filtration rate of the glomerulus [17, 2022]. Although, less debated today, the hypothesis that mesangial cells and their processes can regulate filtration rate has not been fully discarded.

Structure and composition of the mesangial matrix

Early investigators of the mesangial region had the notion that the mesangial matrix was just an extension of the glomerular basement membrane surrounding the capillaries [12]. This mistake was understandable given that both the basement membrane and the mesangial matrix have similar composition. However, further studies revealed key differences between the mesangial matrix and basement membrane that correspond to the structural-functional relationships of both the basement membrane and the matrix.

The glomerular basement membrane is mainly composed of high-density collagen IV, laminin, and the proteoglycan heparan sulfate [23]. In contrast, the mesangial matrix is composed of low-density collagen IV, various proteoglycans including heparan sulfate, high concentration of fibronectin, and a low concentration of laminin [17].

These differences in composition result in significant differences in the properties and behavior of basement membrane and mesangial matrix, which enable the two to carry out their specialized functions. The high density of collagen IV in basement membranes provides structural integrity and elasticity to the GBM to be able to withstand the high capillary pressures. In contrast, the low-density collagen and the presence of water-binding proteoglycans [17] enable the mesangial matrix to oppose both expansile forces and compressive forces. The high concentration of glycoproteins such as fibronectin enable the binding of the cell to the structural components of the matrix [13, 24].

The mesangial matrix is gel-like [12] and contains extensive structural elements that are important for its function. Electron microscopy studies describe a fine meshwork of non-collagenous microfibrils (11-15 nm diameter) indirectly attaching the basement membrane to the mesangial cell processes [13]. The indirect attachments are augmented by direct attachments between the GBM and the mesangial cell [25]. The extensive structural elements and the intimate connection between the GBM and the mesangial cell enable the maintenance of the glomerular structure [22]. Without such frequent close attachments, hydrostatic pressures across the perimesangial GBM cause mesangial expansion [25].

Although the mesangial matrix is not considered a basement membrane, its composition indicates that it is more like a basement membrane than an interstitial matrix. It does not contain the major components found in interstitial matrices such as collagen I and collagen III, but rather consists of collagen IV, the main component of basement membrane matrix [17]. In diabetic nephropathy, however, the composition of the matrix is altered where even collagen III is observed [26].

Function of the mesangial cell and matrix

An important mesangial cell function is the maintenance of glomerular structure. Unlike the bulk of the vasculature, glomerular capillaries do not have a basement membrane that completely encircles the capillary lumen (Fig. 1), as it is blocked by a portion of the capillary lumen that is exposed to the mesangium. In other vasculature, the expansile forces due to blood flow are counterbalanced by wall tension provided by the vascular wall and the active contractile elements within the vascular wall [22]. However, due to the incomplete wrapping of the GBM around the capillary lumen, the GBM cannot develop wall tension independently. Likewise, the endothelial lining of the capillary lumen is incapable of providing wall tension because it is very thin and highly fenestrated [13]. Thus, the mesangial cell acts as an anchor for the GBM to be able to provide wall tension to resist the distending forces arising within the capillary [22]. This mechanism is reliant on the presence of strong connections between the mesangial cell, its matrix, and the GBM [22, 27]. Failure of these connections lead to capillary ballooning and bulging [12, 22, 27] and displacement of capillary towards urinary space leading to podocyte effacement or lesion formation [27]. Perfusion of glomeruli with high pressures breaks mesangial and GBM connections, eventually leading to capillary dilation [22]. Hypertension and hypertrophy, dysfunctions commonly associated with diabetic patients [28], can contribute to the mechanical failure of the mesangium [22].

Hypertension can also increase the stress experienced by the mesangial cell/GBM connections along the axis of the mesangial cell processes. Although, these mesangial cell/GBM connections are weak and susceptible to failure, they occur frequently enough to be able to oppose the distending forces of the hydrostatic pressure in the mesangial region [25]. However, increases in hydrostatic pressures can break these mesangial cell/GBM connections resulting in mesangial expansion [22, 25].

One of the hypothesized functions of mesangial cells was the regulation of GFR [21] via a reduction in capillary diameter [17]. In vivo studies of GFR indicated that the infusion of vasoactive agents led to a 50 percent reduction in the GFR [21]. The hypothesis was further supported by in vitro experiments that showed that mesangial cells contracted when exposed to vasoactive agents such as angiotensin II (Ang II) [29]. Electron microscope studies provided the mechanisms by which mesangial cell contraction could lead to a reduction in capillary diameter and thus GFR [13]. However, further electron microscope studies revealed that mesangial cell contraction could only reduce capillary diameter by 10 percent, which could not account for the 50 percent reduction in GFR that was observed in vivo [22]. Additionally, the in vivo studies showed significant increase in the efferent arteriolar resistance, which could have mediated the decrease in GFR [21]. It is well known today that the vasoconstriction and vasodilation of the glomerular arterioles mediated through vasoactive agents regulate glomerular hemodynamics, which directly impacts GFR [30]. Although the regulation of GFR is mainly mediated by arteriolar constriction and dilation, the involvement of mesangial cells in the regulation of GFR has not been completely discarded and awaits full elucidation [22].

Overall, the structure of the mesangial cell in relation to the GBM and the capillaries indicates that the mesangial cell’s functions are biomechanical in nature. As such in disease conditions such as diabetic nephropathy, dysfunction of the mesangial cell can lead to a failure of the mesangial cell to carry out its biomechanical functions. However, the extent to which the mesangial cell’s biomechanical functions are altered and how they, in turn, affect overall glomerular function is not well known.

Progression of mesangial expansion in diabetic nephropathy

Mesangial expansion is one of the key structural changes observed in the glomerulus 5-7 years after the onset of diabetes in humans [31, 32]. Prior to mesangial expansion, GBM thickening and kidney hypertrophy are the earliest observed structural changes [28, 3234]. However, metabolic and hemodynamic alterations such as hyperglycemia and hypertension generally precede and occur concurrently with the observed structural alterations [35].

Mesangial expansion occurs as a consequence of aberrant mesangial cell proliferation, mesangial matrix accumulation, and hypertrophy induced by the diabetic state. It is now well established that metabolic and hemodynamic alterations within the kidney are the primary causes of kidney failure in diabetes and by extension the causes of abnormal structural changes such as mesangial expansion. In vivo and in vitro models of the diabetic state have elucidated more clearly how metabolic and hemodynamic alterations lead to mesangial expansion. The results indicate that the biomechanical link of the mesangial cell to the glomerular capillaries enable hemodynamic alterations to play a role in the progression of mesangial expansion and that hemodynamic alterations exert their influence through metabolic derangements in the mesangial cell. Thus, the two are closely intertwined and augment each other in the progression of mesangial expansion.

The role of hyperglycemia in mediating matrix protein accumulation

Matrix protein accumulation is characteristic of mesangial expansion. Immunohistochemical studies showed that native collagens such as collagen IV, V, and VI and non-native collagens such as collagen III accumulated in the mesangium during diabetic nephropathy [26]. In vitro studies showed that the incubation of mesangial cells in high glucose concentrations resulted in the accumulation of matrix components such as collagen I, collagen IV, fibronectin, and laminin (Table 2) [3639]. Further studies elucidated that either the increased synthesis of collagen [37, 40] or the inhibition of collagen degradation [38] led to collagen accumulation. It is possible that short-term accumulation of collagen is mediated by increased synthesis and that long-term accumulation is mediated by the inhibition of collagen degradation. A thorough investigation elucidated that high glucose conditions did not cause the diabetic milieu, but that the increased uptake of glucose was the cause of the matrix accumulation, which was mediated by the upregulation of Glut-1 receptors [40].

Table 2 The role of hyperglycemia in matrix protein accumulation in in vitro studies. The effect of hyperglycemia on upregulation of matrix proteins is reduced after long periods of incubation. TGF- β: transforming growth factor - β, N.S.: not significant, -: indicates no data. A single arrow pointing upwards indicates a statistically significant increase in the respective protein expression, and a double arrow indicates a substantially larger increase in protein expression relative to control

Hyperglycemia-induced matrix accumulation is mediated by TGF- β. TGF- β is significantly upregulated during the diabetic milieu in in vitro studies [38, 41], diabetic animal models [42, 43], and human diabetes [43] and has been repeatedly implicated in mediating matrix accumulation [38, 41, 42]. TGF- β’s role in matrix accumulation [44, 45], the intracellular signaling pathways involved [

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