Textor, Stephen C.

Introduction: Morbidity and mortality attributable to hypertension are higher in African American (AAEH) compared to Caucasian essential hypertensive (EH) patients, possibly related to a differential effect on vascular injury and repair. While circulating endothelial progenitor cells (EPC) preserve endothelial integrity IEC detach from sites of injury and represent markers of vascular damage. We hypothesized that plasma levels of IEC and inflammatory markers would be higher in AAEH compared to EH patients. METHODS: Inferior vena cava levels of CD34+/KDR+ (EPC) and VAP-1+ (IEC) cells were measured by FACS in EH and AAEH under fixed sodium intake (150 mEq/d) and blockade of the renin-angiotensin-system, and compared to systemic levels in normotensive control subjects (n=19 each). Systemic levels of inflammatory cytokines and EPC homing factors were measured by Luminex. Results: Blood pressure, serum creatinine, lipids, antihypertensive medications, and EPC levels did not differ between EH and AAEH patients. Circulating IEC were higher in AAEH, and inversely correlated with EPC levels (Figure). Systemic levels of inflammatory cytokines and EPC homing factors were higher in AAEH compared to EH patients (Table), and correlated directly with IEC. Conclusion: Despite preserved kidney function and controlled BP, circulating inflammatory markers were elevated in AAEH and correlated with increased IEC and decreased EPC levels. Increased release of cytokines and IEC in AAEH may impair EPC reparative capacity and predispose to hypertensive vascular injury. This process may aggravate vascular damage and accelerate hypertension-related morbidity/mortality rates in AAEH.

Background: Insulin-like growth factor binding protein7 (IGFBP-7) and tissue inhibitor of metalloproteinase-2 (TIMP-2) reflect G1-cell cycle arrest and are used as biomarkers for AKI. Recent studies show that these biomarkers rise in ischemic conditions and suggest that they actually may limit the severity of AKI. We tested the hypothesis that renal vein [IGFBP-7]*[TIMP-2] correlate with reductions in renal blood flow (RBF) and post-stent single kidney (SK)-GFR changes in patients with RVD undergoing contrast-based imaging and stent revascularization Methods: Inpatient studies were performed during 150 mEq Na+ intake and ACE/ARB Rx in patients with hemodynamically severe RVD (n=29, Doppler velocity = 318 ± 100cm/sec, and eGFR= 34.7 ± 11.7 mL/ min) scheduled for renal artery stenting, and compared to essential hypertensive (EH) healthy controls (n=32). Cortical and medullary RBFs (by multidetector CT) and renal vein levels of IGFBP-7 and TIMP-2 were measured before renal artery stenting and 3 months later Results: Pre-stenting IGFBP-7 and TIMP-2 levels were elevated in RVD compared to EH (18.5±2. vs 15.7±1.5 and 97.4±23.1vs 62.7±9.2 ng/mL respectively, P <0.0001). Baseline renal vein levels of IGFBP-7 * TIMP-2 correlated inversely with pre-stent RBF (r= - 0.53, P=0.01) and directly with the change (%) in SK-GFR observed 3 months after stenting Conclusion: IGFBP-7 and TIMP-2 levels are elevated in patients with chronic RVD as a function of the baseline reduction in RBF, and associate with subsequent improvement in SK-GFR 3 months after revascularization. These data are consistent with renovascular occlusion inducing cell-cycle arrest that serves to protect kidney function.

Significance Statement Renal artery stenosis (RAS) engenders stenotic-kidney ischemia, dysfunction, and injury, but whether these are mediated by cellular senescence has not been elucidated. INK-ATTAC transgenic mice, high-resolution imaging, and unbiased single-cell RNA sequencing of murine kidneys demonstrated cellular senescence as an important mechanism of progressive injury to epithelial/stromal cells within poststenotic kidneys. Both p16-specific and broad quercetin/dasatinib interventions to blunt senescence improved renal function and structure, underscoring the central role of senescence in the pathogenesis. Furthermore, this mechanism was conserved in human subjects with RAS. These observations reveal new mechanisms that contribute to the pathogenesis of chronic ischemic renal injury, and support the development of senolytic therapy to reduce senescent cell burden and delay renal injury. Background Peripheral vascular diseases may induce chronic ischemia and cellular injury distal to the arterial obstruction. Cellular senescence involves proliferation arrest in response to stress, which can damage neighboring cells. Renal artery stenosis (RAS) induces stenotic-kidney dysfunction and injury, but whether these arise from cellular senescenceand their temporal pattern remain unknown. Methods Chronic renal ischemia was induced in transgenic INK-ATTAC and wild type C57BL/6 mice by unilateral RAS, and kidney function ( in vivo micro-MRI) and tissue damage were assessed. Mouse healthy and stenotic kidneys were analyzed using unbiased single-cell RNA-sequencing. To demonstrate translational relevance, cellular senescence was studied in human stenotic kidneys. Results Using intraperitoneal AP20187 injections starting 1, 2, or 4 weeks after RAS, selective clearance of cells highly expressing p16 Ink4a attenuated cellular senescence and improved stenotic-kidney function; however, starting treatment immediately after RAS induction was unsuccessful. Broader clearance of senescent cells, using the oral senolytic combination dasatinib and quercetin, in C57BL/6 RAS mice was more effective in clearing cells positive for p21 ( Cdkn1a ) and alleviating renal dysfunction and damage. Unbiased, single-cell RNA sequencing in freshly dissociated cells from healthy and stenotic mouse kidneys identified stenotic-kidney epithelial cells undergoing both mesenchymal transition and senescence. As in mice, injured human stenotic kidneys exhibited cellular senescence, suggesting this process is conserved. Conclusions Maladaptive tubular cell senescence, involving upregulated p16 ( Cdkn2a ), p19 ( Cdkn2d ), and p21 ( Cdkn1a ) expression, is associated with renal dysfunction and injury in chronic ischemia. These findings support development of senolytic strategies to delay chronic ischemic renal injury.

Mesenchymal stem/stromal cells (MSCs) facilitate repair in experimental diabetic kidney disease (DKD). However, the hyperglycemic and uremic milieu may diminish regenerative capacity of patient-derived therapy. We hypothesized that DKD reduces human MSC paracrine function. Adipose-derived MSC from 38 participants with DKD and 16 control subjects were assessed for cell surface markers, trilineage differentiation, RNA sequencing (RNA-seq), in vitro function (coculture or conditioned medium experiments with T cells and human kidney cells [HK-2]), secretome profile, and cellular senescence abundance. The direction of association between MSC function and patient characteristics were also tested. RNA-seq analysis identified 353 differentially expressed genes and downregulation of several immunomodulatory genes/pathways in DKD-MSC versus Control-MSC. DKD-MSC phenotype, differentiation, and tube formation capacity were preserved, but migration was reduced. DKD-MSC with and without interferon-γ priming inhibited T-cell proliferation greater than Control-MSC. DKD-MSC medium contained higher levels of anti-inflammatory cytokines (indoleamine 2,3-deoxygenase 1 and prostaglandin-E2) and prorepair factors (hepatocyte growth factor and stromal cell–derived factor 1) but lower IL-6 versus control-MSC medium. DKD-MSC medium protected high glucose plus transforming growth factor-β–exposed HK-2 cells by reducing apoptotic, fibrotic, and inflammatory marker expression. Few DKD-MSC functions were affected by patient characteristics, including age, sex, BMI, hemoglobin A1c, kidney function, and urine albumin excretion. However, senescence-associated β-galactosidase activity was lower in DKD-MSC from participants on metformin therapy. Therefore, while DKD altered the transcriptome and migratory function of culture-expanded MSCs, DKD-MSC functionality, trophic factor secretion, and immunomodulatory activities contributing to repair remained intact. These observations support testing of patient-derived MSC therapy and may inform preconditioning regimens in DKD clinical trials.

Ischemic nephropathy reflects progressive loss of kidney function due to large vessel atherosclerotic occlusive disease. Recent studies indicate that this process is characterized by microvascular rarefaction, increased tissue hypoxia and activation of inflammatory processes of tissue injury. This review summarizes the rationale and application of functional MR imaging to evaluate tissue oxygenation in human subjects that defines the limits of renal adaptation to reduction in blood flow, development of increasingly severe tissue hypoxia and recruitment of inflammatory injury pathways in ischemic nephropathy. Human mesenchymal stromal/stem cells (MSC) are capable of modifying angiogenic pathways and immune responses, but the potency of these effects vary between individuals and various clinical characteristics including age and chronic kidney disease and levels of hypoxia. We summarize recently completed first-in-human studies applying intrarenal infusion of autologous adipose-derived MSC in human subjects with ischemic nephropathy that demonstrate a rise in blood flow and reduction in tissue hypoxia consistent with partial repair of microvascular injury, even without restoring main renal arterial blood flow. Inflammatory biomarkers in the renal vein of post-stenotic kidneys fell after MSC infusion. These changes were associated with modest but significant dose-related increments in kidney function. These data provide support a role for autologous MSC in repair of microvascular injury associated with tissue hypoxia.

AbstractBackgroundThe medium‐ to long‐term outcomes of living kidney donors with hypertension compared to normotensive donors are not well understood, especially with the recent changes in hypertension guidelines.MethodsWe studied a cohort of 950 living kidney donors using different definitions of hypertension based on either ≥140/90 or ≥130/80 mmHg thresholds and based on either office or ambulatory blood pressure readings. Microstructural features on kidney biopsy at the time of donation were compared using different definitions of hypertension.ResultsAfter adjusting for years of follow‐up, age, sex, and baseline eGFR, hypertension (by any definition) did not significantly predict an eGFR < 45 ml/min/1.73 m2 at a median follow‐up of 10 years postdonation, though there was a borderline association with ambulatory blood pressure ≥ 130/80 mmHg predicting a 40% decline in eGFR (OR = 1.53, 1.00–2.36; p = .051). Proteinuria was predicted by office blood pressure ≥ 140/90 mmHg and by nondipper profile on nocturnal ambulatory blood pressure measurements. At the time of donation, larger glomeruli and arterial hyalinosis on biopsy were associated with hypertension defined by either ≥140/90 or ≥130/80 mmHg (by office or ambulatory measurements). Nocturnal nondipper status was associated with larger glomeruli size but not arteriolar hyalinosis when compared to dippers.ConclusionsIn programs that accept donors with controlled hypertension, various definitions of hypertension are associated with histological findings in the donated kidney, but none predict a clinically significant decline in kidney function 10 years after donation. These data support allowing healthy individuals with controlled hypertension to donate a kidney. However, donors with office hypertension (≥140/90 mmHg) and nondippers (regardless of hypertension status) are at greater long‐term risk for proteinuria, and particularly for these donors, longer follow‐up is warranted.

AbstractPercutaneous transluminal renal angioplasty (PTRA) has been used to treat renovascular disease (RVD), a chronic condition characterized by renal ischemia and metabolic abnormalities. Mitochondrial injury has been implicated as a central pathogenic mechanism in RVD, but whether it can be reversed by PTRA remains uncertain. We hypothesized that PTRA attenuates mitochondrial damage, renal injury, and dysfunction in pigs with coexisting renal artery stenosis (RAS) and metabolic syndrome (MetS). Four groups of pigs (n = 6 each) were studied after 16 weeks of diet‐induced MetS and RAS (MetS + RAS), MetS + RAS treated 4 weeks earlier with PTRA, and Lean and MetS Sham controls. Single‐kidney renal blood flow (RBF) and glomerular filtration rate (GFR) were assessed in vivo with multidetector computed tomography, and renal tubular mitochondrial structure and function and renal injury ex vivo. PTRA successfully restored renal artery patency, but mean arterial pressure remained unchanged. Stenotic kidney RBF and GFR, which fell in MetS + RAS compared to MetS, rose after PTRA. PTRA attenuated MetS + RAS‐induced mitochondrial structural abnormalities in tubular cells and peritubular capillary endothelial cells, decreased mitochondrial H202 production, and increased renal cytochrome‐c oxidase‐IV activity and ATP production. PTRA also improved cortical microvascular and peritubular capillary density and ameliorated tubular injury and tubulointerstitial fibrosis in the poststenotic kidney. Importantly, renal mitochondrial damage correlated with poststenotic injury and dysfunction. Renal revascularization attenuated mitochondrial injury and improved renal hemodynamics and function in swine poststenotic kidneys. This study suggests a novel mechanism by which PTRA might be relatively effective in ameliorating mitochondrial damage and improving renal function in coexisting MetS and RAS.

AbstractCell stress may give rise to insuperable growth arrest, which is defined as cellular senescence. Stenotic kidney (STK) ischemia and injury induced by renal artery stenosis (RAS) may be associated with cellular senescence. Mesenchymal stem cells (MSCs) decrease some forms of STK injury, but their ability to reverse senescence in RAS remains unknown. We hypothesized that RAS evokes STK senescence, which would be ameliorated by MSCs. Mice were studied after 4 weeks of RAS, RAS treated with adipose tissue‐derived MSCs 2 weeks earlier, or sham. STK senescence‐associated β‐galactosidase (SA‐β‐Gal) activity was measured. Protein and gene expression was used to assess senescence and the senescence‐associated secretory phenotype (SASP), and staining for renal fibrosis, inflammation, and capillary density. In addition, senescence was assessed as p16+ and p21+ urinary exosomes in patients with renovascular hypertension (RVH) without or 3 months after autologous adipose tissue‐derived MSC delivery, and in healthy volunteers (HV). In RAS mice, STK SA‐β‐Gal activity increased, and senescence and SASP marker expression was markedly elevated. MSCs improved renal function, fibrosis, inflammation, and capillary density, and attenuated SA‐β‐Gal activity, but most senescence and SASP levels remained unchanged. Congruently, in human RVH, p21+ urinary exosomes were elevated compared to HV, and only slightly improved by MSC, whereas p16+ exosomes remained unchanged. Therefore, RAS triggers renal senescence in both mice and human subjects. MSCs decrease renal injury, but only partly mitigate renal senescence. These observations support exploration of targeted senolytic therapy in RAS.

The significance of peristenotic collateral circulation (PCC) development around a stenotic renal artery is unknown. We tested the hypothesis that PCC is linked to loss of kidney function and recovery potential in patients with atherosclerotic renovascular disease (ARVD). Thirty-four patients with ARVD were assigned to medical-therapy with or without revascularization based on clinical indications. The PCC was visualized using multidetector computed tomography and defined relative to segmental arteries in patients with essential hypertension. PCC number before and 3 months after treatment was correlated with various renal parameters. Thirty-four stenotic kidneys from 30 patients were analyzed. PCC number correlated inversely with kidney volume. ARVD–stenotic kidneys with baseline PCC (collateral ARVD [C-ARVD], n=13) associated with elevated 24-hour urine protein and stenotic kidney vein level of tumor necrosis factor-α, lower single-kidney volume and blood flow, and greater hypoxia than in stenotic kidneys with no PCC (no collateral ARVD [NC-ARVD], n=17). Revascularization (but not medical-therapy alone) improved stenotic kidney function and reduced inflammation in both NC-ARVD and C-ARVD. In C-ARVD, revascularization also increased stenotic kidney volume, blood flow, and oxygenation to levels comparable to NC-ARVD, and induced PCC regression. However, revascularization improved systolic blood pressure, plasma renin activity, and filtration fraction only in NC-ARVD. Therefore, patients with C-ARVD have greater kidney dysfunction, atrophy, hypoxia, and inflammation compared with patients with NC-ARVD, suggesting that PCC does not effectively protect the stenotic kidney in ARVD. Renal artery revascularization improved in C-ARVD stenotic kidney function, but not hypertension or renin-angiotensin system activation. These observations may help direct management of patients with ARVD.