Glomerular Vasculature Research Articles

APOL1 promotes endothelial cell activation beyond the glomerulus

Apolipoprotein L1 (APOL1) high-risk genotypes are associated with increased risk of chronic kidney disease (CKD) in people of West African ancestry. Given the importance of endothelial cells (ECs) in CKD, we hypothesized that APOL1 high-risk genotypes may contribute to disease via EC-intrinsic activation and dysfunction. Single cell RNA sequencing (scRNA-seq) analysis of the Kidney Precision Medicine Project dataset revealed APOL1 expression in ECs from various renal vascular compartments. Utilizing two public transcriptomic datasets of kidney tissue from African Americans with CKD and a dataset of APOL1-expressing transgenic mice, we identified an EC activation signature; specifically, increased intercellular adhesion molecule 1 (ICAM-1) expression and enrichment in leukocyte migration pathways. Invitro, APOL1 expression in ECs derived from genetically modified human induced pluripotent stem cells and glomerular ECs triggered changes in ICAM-1 and platelet endothelial cell adhesion molecule 1 (PECAM-1) leading to an increase in monocyte attachment. Overall, our data suggest the involvement of APOL1 as an inducer of EC activation in multiple renal vascular beds with potential effects beyond the glomerular vasculature.

The PDE5 inhibitor, vardenafil, ameliorates progressive pathological changes in a focal segmental glomerulosclerosis mouse model

AimsPhosphodiesterase 5 inhibitors (PDE5is) inhibit the hydrolysis of cyclic guanosine 5′-monophosphate in smooth muscle cells and are a widely known treatment for erectile dysfunction. Accumulating evidence also suggests that PDE5is exhibit potential benefits in cardiovascular and chronic kidney diseases. In this study, we examined the therapeutic effects of a PDE5i, vardenafil (VAR), in a focal segmental glomerulosclerosis (FSGS) mouse model. Materials and methodsFSGS was induced in BALB/c mice by the intravenous administration of Adriamycin (AD, 11 mg/kg of body weight). After 24 h, VAR (at 12.5 μg/ml) was given in drinking water ad libitum until the animals were sacrificed. At the end of the experiment, plasma and kidney samples were harvested to evaluate clinical parameters, histopathological changes, and alterations in transcriptome and protein expressions. Key findingsIn this study, VAR treatment attenuated the deterioration of proteinuria, renal dysfunction, and hypercholesterolemia in AD-induced FSGS. Treatment with VAR exhibited reductions in the severity of both glomerulosclerosis and tubulointerstitial injury in the histological analysis. In addition to relieving AD-induced podocyte loss, VAR also preserved endothelial cells in glomerular capillaries and ameliorated the accumulation of collagen fibers in the mesangial area and Bowman's capsule basement membrane. In addition, VAR showed an ability to suppress transforming growth factor-β-induced fibroblast-to-myofibroblast transdifferentiation. SignificanceOur data suggest that VAR exhibited reno-therapeutic effects via attenuating podocyte loss, preserving the integrity of the glomerular vasculature, and ameliorating fibrotic changes. These findings suggest that PDE5is might be a promising treatment modality for nephrotic syndrome.

Histological observation on the initial stage of vascular invasion into the secondary ossification of murine femoral epiphyseal cartilage.

It remains unknown whether the histology of vascular invasion during secondary ossification of epiphyseal cartilage is the same as that seen in primary ossification; we examined the initial processes of vascular invasion of secondary ossification in the murine femora. Many endomucin-immunoreactive blood vessels gathered at the central region of the articular surface, and buds of soft tissue, including glomerular loops of endomucin-immunoreactive blood vessels and TNALPase- immunopositive osteoblastic cells accompanied by TRAP-positive osteoclasts, had begun to invade the epiphyseal cartilage. The invading soft tissues formed cartilage canals displaying MMP9 immunoreactivity in the tip region, and cartilaginous collagen fibrils were not visible in the vicinity of the vascular wall of the blood vessels. Thus, the histological profile marked by invading glomerular vasculature and the erosion of the cartilage matrix near the vascular walls during secondary ossification differs from that seen during primary ossification.

NMDA Receptor Hypofunction in the Aging-Associated Malfunction of Peripheral Tissue.

Glutamatergic transmission through NMDA receptors (NMDARs) is important for the function of peripheral tissues. In the bone, NMDARs and its co-agonist, D-serine participate in all the phases of the remodeling. In the vasculature, NMDARs exerts a tonic vasodilation decreasing blood perfusion in the corpus cavernosum and the filtration rate in the renal glomerulus. NMDARs are relevant for the skin turnover regulating the proliferation and differentiation of keratinocytes and the formation of the cornified envelope (CE). The interference with NMDAR function in the skin leads to a slow turnover and repair. As occurs with the brain and cognitive functions, the manifestations of a hypofunction of NMDARs resembles those observed during aging. This raises the question if the deterioration of the glomerular vasculature, the bone remodeling and the skin turnover associated with age could be related with a hypofunction of NMDARs. Furthermore, the interference of D-serine and the effects of its supplementation on these tissues, suggest that a decrease of D-serine could account for this hypofunction pointing out D-serine as a potential therapeutic target to reduce or even prevent the detriment of the peripheral tissue associated with aging.

FC 016SPARSENTAN IMPROVES GLOMERULAR BLOOD FLOW AND AUGMENTS PROTECTIVE TISSUE REMODELING IN MOUSE MODELS OF FOCAL SEGMENTAL GLOMERULOSCLEROSIS (FSGS)

Abstract Background and Aims Preliminary preclinical and emerging clinical evidence indicates strong antiproteinuric actions of dual endothelin type A (ETA) and angiotensin II type 1 (AT1) receptor antagonism with sparsentan. These nephroprotective effects have been more pronounced in different experimental and clinical settings compared to current standard of care using an AT1 receptor blocker (ARB). Considering the broad spectrum of renal actions of endothelin (ET) and angiotensin II (Ang II), inhibition of both pathways using sparsentan is postulated to target multiple renal cell types via a variety of renoprotective mechanisms. The aim of this study was to determine glomerular action of sparsentan as compared to the ARB losartan (Los) by direct visualization of effects on renal hemodynamics and tissue remodeling in the intact living kidney. Method Intravital multiphoton microscopy (MPM) of the glomerular vasculature and filtration barrier structure and function was performed in genetically engineered mice combined with traditional urinalysis and histology-based phenotyping. Glomerular hemodynamic parameters (afferent and efferent arteriole (AA and EA) diameters and single nephron glomerular filtration rate (SNGFR)) and podocyte calcium entry, as a measure of cell injury, were quantitatively visualized in the FSGS model Pod-GCaMP5/Tomato TRPC6 transgenic mice (1.5 years of age), in which TRPC6 is overexpressed together with the calcium reporter GCaMP5 in podocytes. Single cell identification and fate tracking of cells of the renin lineage (CoRL) was performed over time using a second physiologic control mouse model, Ren1d-Confetti mice that feature a multicolor CFP/GFP/YFP/FP reporter. Three groups of mice in each model received treatment with either vehicle (CTRL), the ARB losartan (Los; 10 mg/kg/day), or sparsentan (120 mg/kg/day) for 6 weeks (FSGS model) or 2 weeks (control physiology model). Results Both Los and sparsentan treatment attenuated the acute ET + Ang II-induced elevation of podocyte calcium by ∼80%, and the development of albuminuria, and glomerulosclerosis and tissue fibrosis in the FSGS model. Notably, sparsentan prevented the ET + Ang II increases in podocyte calcium more than Los and was significantly more effective in dilating both AA and EA (Fig. 1A, B), increasing SNGFR (Fig. 1C), increasing capillary blood flow (2-fold; p<0.0001 vs. CTRL), and decreasing albuminuria (20%; p<0.05 vs. CTRL). Sparsentan also preserved p57+ podocyte number by 50% compared to Los (p<0.0001 vs. Los). Similarly, pretreatment with sparsentan was more effective in preventing glomerular arteriolar vasoconstriction induced by acute ET + Ang II iv injection compared to Los (p<0.05 vs. Los). Following a 2-week treatment in control healthy Ren1d-Confetti mice, sparsentan resulted in a more robust increase compared to Los in the number of Confetti+ cells, clones, and individual cells per clone in the glomeruli and AA (Fig. 1D-F). Renal tubule segments also showed active cellular remodeling in response to sparsentan. Conclusion Serial MPM imaging directly visualized several mechanisms underlying beneficial antiproteinuric and structural effects of sparsentan in both FSGS and in the normal mouse kidney and differences between dual ETA/AT1 receptor antagonism of sparsentan and a mono-selective ARB. The sparsentan-induced glomerular hemodynamic pattern was driven by both AA and EA dilation resulting in an increase in capillary blood flow. Compared to Los, sparsentan was more effective in attenuating ET/Ang II-induced podocyte injury and in activation of resident progenitor cells and tissue remodeling. These findings suggest multiple layers of renal protective actions by dual ETA and AT1 receptor antagonism.

Succinate Recapitulates a Diabetes-Like Phenotype of Renal Dysfunction and Injury in the Absence of Hyperglycemia

Background: Succinate is a well-known intermediate in the tricarboxylic acid (Krebs) cycle, where it is converted to form fumarate by succinate dehydrogenase from succinyl-CoA. Surprisingly, besides as a general metabolic intermediate, succinate can selectively activate orphan G receptor GPR91, which is highly expressed in the kidney, including glomerular vasculature, the macula densa (MD), proximal tubules, distal tubules, the cortical thick ascending limb of (cTAL) Henle’s loop, and the cortical as well as medullary collecting duct (CD). The level of succinate in patients with type 2 diabetes and obesity was much higher than healthy controls. However, the causative role of succinate in renal function is still unknown. Considering the excess succinate concentration, abundant expression of GPR91, and the importance of the kidney, we speculated that succinate could affect renal function. Methods: Male C57 BL/6 mice provided sterile drinking water containing 4% succinate ad libitum for 8 weeks and 12 weeks were used to investigate the effect of succinate on important renal functions. Results: Succinate administration markedly promoted water reabsorption via upregulating AQP2 protein and gene expression. Furthermore, Na + retention was significantly enhanced by succinate via increasing ENaC protein and gene expression. Moreover, succinate caused severe damage to the kidney, including glomerulus and proximal tubule, and tubulointerstitial fibrosis. These pathological changes were consistent with characteristics reported in diabetic kidney disease both in clinical patients and animal models. Conclusions: Other than a common metabolic intermediate in the Krebs cycle, elevated succinate in the circulation causes renal dysfunction and damage and recapitulates a diabetes-like phenotype of kidney disease, indicating that eliminating succinate from blood may improve clinic intervention in diabetic kidney disease. Funding Information: This study was supported by the National Natural Science Foundation of China (Grants 82070888, 82070882, 81770808, 81872165, 81701414, 81871211, and 81702879); National Key RD Guangdong Provincial Key RD Key Project of Nature Science Foundation of Guangdong Province, China (Grant 2019B1515120077); Guangdong Natural Science Fund (Grant 2019A1515011810, 2021A1515010434); Guangdong Science Technology Project (Grant 2017A020215075); Key Sci-Tech Research Project of Guangzhou Municipality, China (Grants 201803010017, 201807010069, and 202002020022); 2017 and 2019 Milstein Medical Asian American Partnership Foundation Research Project Award in Translational Medicine. Declaration of Interests: None. Ethics Approval Statement: All animal procedures were approved by the Animal Care and Use Committee of Sun Yat-Sen University (Ethics Committee of ZSSOM on Laboratory Animal Care No. 2020001017; Guangzhou, China).

Live Imaging of Monocyte Subsets in Immune Complex-Mediated Glomerulonephritis Reveals Distinct Phenotypes and Effector Functions.

Immune complexes within glomerular capillary walls cause crescentic GN (CrGN). Monocytes and macrophages are important in mediating CrGN, but little work has been done to phenotype the subpopulations involved and determine their respective contributions to glomerular inflammation. Live glomerular imaging using confocal microscopy monitored intravascular monocyte subset behavior during nephrotoxic nephritis (NTN) in a novel WKY-hCD68-GFP monocyte/macrophage reporter rat strain. Flow cytometry and qPCR further analyzed ex vivo the glomerular leukocyte infiltrate during NTN. Non-classical monocytes surveyed the glomerular endothelium via lymphocyte function-associated antigen 1 (LFA-1) in the steady state. During NTN, non-classical monocytes were recruited first, but subsequent recruitment and retention of classical monocytes was associated with glomerular damage. Monocytes recruited to the glomerular vasculature did not undergo transendothelial migration. This finding suggests that inflammation in immune complex-mediated CrGN is predominantly intravascular, driven by dynamic interactions between intravascular blood monocytes and the endothelium. Glomerular endothelium and non-classical monocytes overexpressed a distinct chemokine axis, which may orchestrate inflammatory myeloid cell recruitment and expression of damage mediators. Reduced classical monocyte recruitment in Lewis rats during NTN confirmed a role for CD16 in mediating glomerular damage. Monocyte subsets with distinct phenotypes and effector functions may be important in driving inflammation in experimental CrGN resulting from immune complexes formed within the glomerular capillary wall. LFA-1-dependent endothelial surveillance by non-classical monocytes may detect immune complexes through CD16, orchestrating the inflammatory response through intravascular retention of classical monocytes, which results in glomerular damage and proteinuria.

Tectorigenin attenuates diabetic nephropathy by improving vascular endothelium dysfunction through activating AdipoR1/2 pathway

Diabetic nephropathy (DN), a kind of microvascular complication, is a primary cause of end-stage renal disease worldwide. However, therapeutic drugs for DN treatment are still in lack. The glomerular endothelium is essential to maintain selective permeability of glomerular filtration barrier and glomerular vasculature function. Growing evidences show that endothelial dysfunction or injury is the initial stage of vascular damage in DN, which can be induced by hyperglycemia, lipotoxicity, and inflammation. Therefore, to improve the function of vascular endothelium in kidney is a key point for treatment of DN. As a plant isoflavone, tectorigenin (TEC) has attracted considerable attention due to its anti-proliferative and anti-inflammatory functions. However, whether TEC could inhibit the DN development remains unknown. In this study, we examined the effects of TEC on DN development in db/db mice, a type of genetic defect diabetic mice that can spontaneously develop into severe renal dysfunction. Intriguingly, TEC treatment restored diabetes-induced glucose and lipid metabolic disorder; and improved the deterioration of renal function, particularly the renal endothelium function in db/db mice. Additionally, TEC inhibited the renal inflammation via reducing macrophages infiltration and M1 polarization. Moreover, TEC inhibited lipopolysaccharide (LPS)-induced endothelial injury and M1 polarization in vitro. Mechanistically, TEC partially restored the reduction in expression of adiponectin receptor 1/2 (AdipoR1/2), pi-LKB1, pi-AMPKα, and PPARα in vitro and in vivo. Noteworthy, these beneficial pharmacological activities mediated by TEC were significantly attenuated after AdipoR1/2 knockdown by siRNA, indicating that AdipoR1/2 plays a critical role in protection against DN. Collectively, these results suggested that TEC have a potently effect for retarding type 2 diabetes–associated DN.

In Vivo Developmental Trajectories of Human Podocyte Inform In Vitro Differentiation of Pluripotent Stem Cell-Derived Podocytes

The renal corpuscle of the kidney comprises a glomerular vasculature embraced by podocytes and supported by mesangial myofibroblasts, which ensure plasma filtration at the podocyte-generated slit diaphragm. With a spectrum of podocyte-expressed gene mutations causing chronic disease, an enhanced understanding of podocyte development and function to create relevant invitro podocyte models is a clinical imperative. To characterize podocyte development, scRNA-seq was performed on human fetal kidneys, identifying distinct transcriptional signatures accompanying the differentiation of functional podocytes from progenitors. Interestingly, organoid-generated podocytes exhibited highly similar, progressive transcriptional profiles despite an absence of the vasculature, although abnormal gene expression was pinpointed in late podocytes. On transplantation into mice, organoid-derived podocytes recruited the host vasculature and partially corrected transcriptional profiles. Thus, human podocyte development is mostly intrinsically regulated and vascular interactions refine maturation. These studies support the application of organoid-derived podocytes to model disease and to restore or replace normal kidney functions.

Glomerular endothelial cell maturation depends on ADAM10, a key regulator of Notch signaling.

The principal function of glomeruli is to filter blood through a highly specialized filtration barrier consisting of a fenestrated endothelium, the glomerular basement membrane and podocyte foot processes. Previous studies have uncovered a crucial role of endothelial a disintegrin and metalloprotease 10 (ADAM10) and Notch signaling in the development of glomeruli, yet the resulting defects have not been further characterized nor understood in the context of kidney development. Here, we used several different experimental approaches to analyze the kidneys and glomeruli from mice lacking ADAM10 in endothelial cells (A10ΔEC mice). Scanning electron microscopy of glomerular casts demonstrated enlarged vascular diameter and increased intussusceptive events in A10ΔEC glomeruli compared to controls. Consistent with these findings, genes known to regulate vessel caliber (Apln, AplnR and Vegfr3) are significantly upregulated in A10ΔEC glomeruli. Moreover, transmission electron microscopy revealed the persistence of diaphragms in the fenestrae of A10ΔEC glomerular endothelial cells, which was corroborated by the elevated expression of the protein PLVAP/PV-1, an integral component of fenestral diaphragms. Analysis of gross renal vasculature by light sheet microscopy showed no major alteration of the branching pattern, indicating a localized importance of ADAM10 in the glomerular endothelium. Since intussusceptions and fenestrae with diaphragms are normally found in developing, but not mature glomeruli, our results provide the first evidence for a crucial role of endothelial ADAM10, a key regulator of Notch signaling, in promoting the development and maturation of the glomerular vasculature.

Prevention and management of diabetic retinopathy in STZ diabetic rats by Tinospora cordifolia and its molecular mechanisms

We investigated the potential of Tinospora cordifolia (TC) in treatment of diabetic retinopathy in STZ-induced rats due to its antihyperglycemic, angiogenic, antiinflammatory and antioxidant effects. The diabetic rats, treated for 24weeks with TC extract (250mg/kg), were evaluated for lenticular and fundus changes. Biochemical parameters were estimated and histopathological studies performed. TC significantly reduced blood glucose and glycated hemoglobin in treated rats. It prevented cataract development in treated group. Angiogenic markers VEGF and PKC increased in diabetic retina, which reduced significantly with TC. Anti-inflammatory parameters TNF-α and IL-1β elevated in diabetic group unlike that in treated group. TC also provided defense against depletion of antioxidant enzymes- glutathione and catalase. Histopathological studies revealed thickening of basement membrane of the retinal and glomerular vasculature of diabetic rat, but no basement membrane widening was seen in treated animals. Destruction of pancreatic islet structure was observed in diabetic group, but not in treated. Thus, TC reduces blood glucose and inhibits overexpression of angiogenic and inflammatory mediators, which are distinct markers of diabetic retinopathy. It also prevents retinal oxidative stress and restores antioxidant enzyme levels. These data provide evidence for the safety and potential effect of TC in the management of experimental diabetic retinopathy.

Mechanisms of HO-1 mediated attenuation of renal immune injury: a gene profiling study

Using a mouse model of immune injury directed against the renal glomerular vasculature and resembling human forms of glomerulonephritis (GN), we assessed the effect of targeted expression of the cytoprotective enzyme heme oxygenase (HO)-1. A human (h) HO-1 complementary DNAN (cDNA) sequence was targeted to glomerular epithelial cells (GECs) using a GEC-specific murine nephrin promoter. Injury by administration of antibody against the glomerular basement membrane (anti-GBM) to transgenic (TG) mice with GEC-targeted hHO-1 was attenuated compared with wild-type (WT) controls. To explore changes in the expression of genes that could mediate this salutary effect, we performed gene expression profiling using a microarray analysis of RNA isolated from the renal cortex of WT or TG mice with or without anti-GBM antibody-induced injury. Significant increases in expression were detected in 9 major histocompatibility complex (MHC)-class II genes, 2 interferon-γ (IFN-γ)-inducible guanosine triphosphate (GTP)ases, and 3 genes of the ubiquitin-proteasome system. The increase in MHC-class II and proteasome gene expression in TG mice with injury was validated by real-time polymerase chain reaction (PCR) or Western blot analysis. The observations point to novel mechanisms underlying the cytoprotective effect of HO-1 in renal immune injury.

Constructing Kidney-like Tissues from Cells Based on Programs for Organ Development: Toward a Method ofIn VitroTissue Engineering of the Kidney

The plausibility of constructing vascularized three-dimensional (3D) kidney tissue from cells was investigated. The kidney develops from mutual inductive interactions between cells of the ureteric bud (UB), derived from the Wolffian duct (WD), and the metanephric mesenchyme (MM). We found that isolated MMs were capable of inducing branching morphogenesis of the WD (an epithelial tube) in recombination cultures; suggesting that the isolated MM retains inductive capacity for WD-derived epithelial tubule cells other than those from the UB. Hanging drop aggregates of embryonic and adult renal epithelial cells from UB and mouse inner medullary collecting duct cell (IMCD) lines, which are ultimately of WD origin, were capable of inducing MM epithelialization and tubulogenesis with apparent connections (UB cells) and collecting duct-like tubules with lumens (IMCD). This supports the view that the collecting system can be constructed from certain epithelial cells (those ultimately of WD origin) when stimulated by MM. Although the functions of the MM could not be replaced by cultured mesenchymal cells, primary MM cells and one MM-derived cell line (BSN) produced factors that stimulate UB branching morphogenesis, whereas another, rat inducible metanephric mesenchyme (RIMM-18), supported WD budding as a feeder layer. This indicates that some MM functions can be recapitulated by cells. Although engineering of a kidney-like tissue from cultured cells alone remains to be achieved, these results suggest the feasibility of such an approach following the normal developmental progression of the UB and MM. Consistent with this notion, implants of kidney-like tissues constructed in vitro from recombinations of the UB and MM survived for over 5 weeks and achieved an apparently host-derived glomerular vasculature. Lastly, we addressed the issue of optimal macro- and micro-patterning of kidney-like tissue, which might be necessary for function of an organ assembled using a tissue engineering approach. To identify suitable conditions, 3D reconstructions of HoxB7-green fluorescent protein mouse rudiments (E12) cultured on a filter or suspended in a collagen gel (type I or type IV) revealed that type IV collagen 3D culture supports the deepest tissue growth (600 +/- 8 microm) and the largest kidney volume (0.22 +/- 0.02 mm(3)), and enabled the development of an umbrella-shaped collecting system such as occurs in vivo. Taken together with prior work (Rosines et al., 2007; Steer et al., 2002), these results support the plausibility of a developmental strategy for constructing and propagating vascularized 3D kidney-like tissues from recombinations of cultured renal progenitor cells and/or primordial tissue.

Study of Glomerular Vasculature in Teleosts with the Resin-Replica Method

Glomerular vasculature was investigated in the carpCyprinus carpio, the scorpionfishSebastiscus marmoratus, and the marine catfishPlotosus lineatus with the resin-replica method. An afferent arteriole was connected with a glomerulus in every fish. It was slender in the carp, whereas the scorpionfish and marine catfish possessed thick afferent arterioles. The glomerular capillaries were sinusoidal. The divergences, convergences, and windings of these capillaries were not well developed in any of the fish. The glomerular capillaries converged into an efferent arteriole in the carp and scorpionfish. In the marine catfish, on the other hand, most of the glomeruli had two efferent arterioles.

Antimyeloperoxidase antibodies rapidly induce α4-integrin–dependent glomerular neutrophil adhesion

AbstractPatients with antineutrophil cytoplasmic antibodies (ANCAs) frequently develop severe vasculitis and glomerulonephritis. Although ANCAs, particularly antimyeloperoxidase (anti-MPO), have been shown to promote leukocyte adhesion in postcapillary venules, their ability to promote adhesion in the glomerular vasculature is less clear. We used intravital microscopy to examine glomerular leukocyte adhesion induced by anti-MPO. In mice pretreated with LPS, 50 μg anti-MPO induced LFA-1–dependent adhesion in glomeruli. In concert with this finding, in mice pretreated with LPS, more than 80% of circulating neutrophils bound anti-MPO within 5 minutes of intravenous administration. However, even in the absence of LPS, more than 40% of circulating neutrophils bound anti-MPO in vivo, a response not seen in MPO−/− mice. In addition, a higher dose of anti-MPO (200 μg) induced robust glomerular leukocyte adhesion in the absence of LPS. The latter response was β2-integrin independent, instead requiring the α4-integrin, which was up-regulated on neutrophils in response to anti-MPO. These data indicate that anti-MPO antibodies bind to circulating neutrophils, and can induce glomerular leukocyte adhesion via multiple pathways. Lower doses induce adhesion only after an infection-related stimulus, whereas higher doses are capable of inducing responses in the absence of an additional inflammatory stimulus, via alternative adhesion mechanisms.

Anti‐myeloperoxidase antibodies rapidly induce α4 integrin‐dependent glomerular neutrophil adhesion

Despite the fact that anti‐myeloperoxidase antibodies (anti‐MPO) have been shown to promote leukocyte adhesion in flow‐based assays in vitro and in postcapillary venules in vivo, their role in promoting adhesion in the glomerular vasculature is less clear. We used renal intravital microscopy to examine glomerular leukocyte adhesion induced by anti‐MPO. In mice pretreated with LPS, 50 μg of anti‐MPO induced LFA‐1‐dependent adhesion in glomeruli. In concert with this finding, in mice pretreated with LPS, ~75% of circulating neutrophils bound anti‐MPO within 5 min of intravenous administration. However, even in the absence of LPS pretreatment, ~50% of circulating neutrophils bound anti‐MPO, a response not seen in MPO−/− mice. In addition, a higher dose of anti‐MPO (200 μg) induced robust glomerular leukocyte adhesion in the absence of LPS pretreatment. The latter response was β2‐integrin‐independent, instead requiring the α4‐integrin, which was upregulated on neutrophils in response to anti‐MPO. These data indicate that anti‐MPO antibodies are capable of inducing glomerular leukocyte adhesion via multiple pathways, with lower doses only inducing adhesion in mice receiving an infection‐related stimulus, and higher doses being capable of inducing responses in the absence of an additional inflammatory stimulus, via alternative adhesion mechanisms.Supported by the NHMRC (Australia) & Genzyme

Shuttling of calcium between endoplasmic reticulum and mitochondria in the renal vasculature

Shuttling of calcium between endoplasmic reticulum and mitochondria in the renal vasculature

A New Trick for an Old Dogma: ENaC Proteins as Mechanotransducers in Vascular Smooth Muscle

Myogenic constriction is a vasoconstriction of blood vessels to increases in perfusion pressure. In renal preglomerular vasculature, it is an established mechanism of renal blood flow autoregulation. Recently, myogenic constriction has been identified as an important protective mechanism, preventing the transmission of systemic pressure to the fragile glomerular vasculature. Although the signal transduction pathways mediating vasoconstriction are well known, how the increases in pressure trigger vasoconstriction is unclear. The response is initiated by pressure-induced stretch of the vessel wall and thus is dependent on mechanical signaling. The identity of the sensor detecting VSMC stretch is unknown. Previous studies have considered the role of extracellular matrix-integrin interactions, ion conduction units (channels and/or transporters), and the cytoskeleton as pressure detectors. Whether, and how, these structures fit together in VSMCs is poorly understood. However, a model of mechanotransduction in the nematode Caenorhadbditis elegans (C. elegans) has been established that ties together extracellular matrix, ion channels, and cytoskeletal proteins into a large mechanosensing complex. In the C. elegans mechanotransducer model, a family of evolutionarily conserved proteins, referred to as the DEG/ENaC/ASIC family, form the ion-conducting pore of the mechanotransducer. Members of this protein family are expressed in VSMC where they may participate in pressure detection. This review will address how the C. elegans mechanotransducer model can be used to model pressure detection in mammalian VSMCs and provide a new perspective to pressure detection in VSMCs.

Alterations of Cell Adhesion Molecules in Human Glomerular Endothelial Cells in Response to Nephritis-Associated Plasminogen Receptor

Background: Acute post-streptococcal glomerulonephritis (APSGN) is induced by glomerular deposition of nephritogenic streptococcal antigen-antibody complexes. Recently, a streptococcal antigen, nephritis-associated plasminogen receptor (NAPlr) was purified from ruptured streptococcal cell supernatants (RCS). However, the cellular and molecular mechanisms of NAPlr action on the glomerular vas culature are still unknown. Methods: Expression of cell adhesion molecules were measured by cellular ELISA (enzyme-linked immunosorbent assay), immunofluorescence microscopy and Western blot analysis. Results: RCS and NAPlr significantly decreased the PECAM-1 expression in human glomerular endothelial cells (HGECs) as compared to that in the control cells. Plasminogen treatment reversed the RCS or NAPlr-induced decrease of PECAM-1 expression and increase of MCP-1 expression. Immunofluorescent microscopy and Western blot analysis also showed that PECAM-1 expression in HGECs was downregulated upon treatment with RCS or NAPlr and this effect was reversed by plasminogen treatment. Furthermore, we found that tumor necrosis factor-α production in culture medium of HGECs was increased at the lower level when the culture system was treated with RCS. Conclusion: RCS and NAPlr modulated PECAM-1 expression and MCP-1 production in HGECs, indicating the involvement of NAPlr in inflammatory cell accumulation in glomerular tufts and functional abnormality of glomerular microvasculature such as hyperpermeability.

Compartment-Specific Expression and Function of the Chemokine IP-10/CXCL10 in a Model of Renal Endothelial Microvascular Injury

The recruitment of inflammatory cells into renal tissue, mainly T cells and monocytes, is a typical feature of various renal diseases such as glomerulonephritis, thrombotic angiopathies, allograft rejection, and vasculitis. T cells predominantly infiltrate the tubulointerstitium, whereas monocytes are present in the tubulointerstitial and glomerular compartment. Because chemokines play a pivotal role in leukocyte trafficking under inflammatory conditions, this study investigated whether a differential expression of chemokines contributes to the precise coordination of leukocyte subtype trafficking in a rat model of renal microvascular endothelial injury. Renal microvascular endothelial injury was induced in rats by selective renal artery perfusion with an anti-endothelial antibody. Induction of the disease led to severe glomerular and tubulointerstitial endothelial injury with subsequent upregulation of chemokines followed by inflammatory cell recruitment. Among the analyzed chemokine mRNA, IP-10/CXCL10 (119-fold), acting via CXCR3 on activated T cells, and MCP-1/CCL2 (65-fold), acting via CCR2 on monocytes, were by far the most strongly upregulated chemokines. In situ hybridization revealed that IP-10/CXCL10 mRNA was selectively expressed by endothelial cells in the tubulointerstitial area, co-localizing with infiltrating T cells. Despite extensive damage of glomerular vasculature, no IP-10/CXCL10 expression by glomerular endothelial cells was detected. MCP-1/CCL2 mRNA in contrast was detectable in the glomerulus and the tubulointerstitium. Treatment with a neutralizing anti-IP-10/CXCL10 antibody significantly reduced the number of infiltrating tubulointerstitial T cells without affecting monocyte migration and led to an improved renal function. Our study demonstrates a role of IP-10/CXCL10 on T cell recruitment in a rat model of renal endothelial microvascular injury. Furthermore, a differential chemokine expression profile by endothelial cells in different renal compartments was found. These findings are consistent with the hypothesis that functional heterogeneity of endothelial cells from different vascular sites exists and provide an insight into the molecular mechanisms that may mediate compartment-specific T cell and monocyte recruitment in inflammatory renal disease.