Lerman, Lilach O.

Abstract Aims We aimed to test the hypothesis that the presence of clonal hematopoiesis of indeterminate potential (CHIP) in peripheral blood cells is associated with coronary endothelial dysfunction, enhanced inflammatory markers, and major adverse cardiovascular events (MACE).Methods and results We compared targeted next generation sequencing (35 CHIP related genes) between patients with coronary endothelial dysfunction (n = 123) and controls (n=65). Coronary endothelial dysfunction was defined by ≥ 20% decrease in coronary artery diameter (CAD) or ≤ 50% increase in coronary blood flow (CBF) in response to acetylcholine injection compared to baseline. Plasma cytokine levels of Interleukin (IL)-6 and IL-8 were also assessed. Patients were subsequently followed for 12.2 ± 4.3 years. Clonal hematopoiesis relevant gene mutations were found in 1 individual in normal endothelial function group (1.5%) and 11 cases in endothelial dysfunction group (9.3%) (p = 0.04). Additionally, CHIP mutations were associated with an increased risk of MACE (OR = 4.08, P = 0.04). Mutations in ASXL1, DNMT3A and TET2 in the endothelial dysfunction group were also associated with increased levels of IL-6 and IL-8 (P = 0.001, P = 0.003; respectively).Conclusion The current study demonstrates a high frequency of CHIP in patients with coronary endothelial dysfunction as well as an association between mutations in three most common epigenetic regulator genes and increased levels of IL-6 and IL-8. Therefore it infers a probable relationship between CHIP, endothelial dysfunction and cardiovascular adverse events.

Aims This study uniquely explored the relationship between coronary microvascular function and exercise haemodynamics using concurrent invasive testing. Methods and results Fifty‐one consecutive patients with unexplained cardiac exertion symptoms, non‐obstructive coronary artery disease and normal left ventricular ejection fraction (>50%) underwent haemodynamic exercise assessment and concurrent coronary reactivity testing. Heart failure with preserved ejection fraction (HFpEF) was defined as a pulmonary arterial wedge pressure (PAWP) ≥15 mmHg at rest and/or ≥25 mmHg at peak exercise. Endothelium‐independent coronary microvascular dysfunction (CMD) was defined as a coronary flow reserve (CFR) ≤2.5, while endothelium‐dependent CMD was defined as ≤50% increase in coronary blood flow (CBF) in response to intracoronary acetylcholine infusions. Patients with HFpEF ( n = 22) had significantly lower CFR (2.5 ± 0.6 vs. 3.2 ± 0.7; P = 0.0003) and median %CBF increase in response to intracoronary acetylcholine [1 (−35; 34) vs. 64 (−4; 133); P = 0.002] compared to patients without HFpEF ( n = 29). PAWP was significantly higher in patients with endothelium‐independent CMD compared to controls during both rest and exercise. This significant elevation was only present during exercise in patients with endothelium‐dependent CMD compared to controls. CFR had significant inverse correlations with PAWP at rest (r = −0.31; P = 0.03) and peak exercise (r = −0.47, P = 0.001). CFR also had positive correlations with maximal exercise capacity (in W/kg) (r = 0.33, P = 0.02). Conclusions Coronary microvascular function is inversely associated with filling pressures, particularly during exercise. Both types of CMD are associated with higher filling pressures at peak exercise. These findings underscore the potential mechanism and therapeutic target for CMD and HFpEF.

BackgroundRenal fibrosis is a common pathway in tubulointerstitial injury and a major determinant of renal insufficiency. Collagen deposition, a key feature of renal fibrosis, may serve as an imaging biomarker to differentiate scarred from healthy kidneys.PurposeTo test the feasibility of using quantitative magnetization transfer (qMT), which assesses tissue macromolecule content, to measure renal fibrosis.Study TypeProspective.Animal ModelFifteen 129S1 mice were studied 4 weeks after either sham (n = 7) or unilateral renal artery stenosis (RAS, n = 8) surgeries.Field Strength/SequenceMagnetization transfer (MT)‐weighted images were acquired at 16.4T using an MT‐prepared fast‐low‐angle‐shot sequence. Renal B0, B1, and T1 maps were also acquired, using a dual‐echo gradient echo, an actual flip angle, and inversion recovery method, respectively.AssessmentA two‐pool model was used to estimate the bound water fraction (f) and other tissue imaging biomarkers. Masson's trichrome staining was subsequently performed ex vivo to evaluate renal fibrosis.Statistical TestsComparisons of renal parameters between sham and RAS were performed using independent samples t‐tests. Pearson's correlation was conducted to investigate the relationship between renal fibrosis by histology and the qMT‐derived bound pool fraction f.ResultsThe two‐pool model provided accurate fittings of measured MT signal. The qMT‐derived f of RAS kidneys was significantly increased compared to sham in all kidney zones (renal cortex [CO], 7.6 ± 2.4% vs. 4.6 ± 0.6%; outer medulla [OM], 8.2 ± 4.2% vs. 4.2 ± 0.9%; inner medulla [IM] + P, 5.8 ± 1.6% vs. 2.9 ± 0.6%, all P < 0.05). Measured f correlated well with histological fibrosis in all kidney zones (CO, Pearson's correlation coefficient r = 0.95; OM, r = 0.93; IM + P, r = 0.94, all P < 0.05).Data ConclusionThe bound pool fraction f can be quantified using qMT at 16.4T in murine kidneys, increases significantly in fibrotic RAS kidneys, and correlates well with fibrosis by histology. Therefore, qMT may constitute a valuable tool for measuring renal fibrosis in RAS.Level of Evidence1Technical Efficacy Stage3

Significance Statement Predicting outcomes on the basis of renal histology after a radical nephrectomy has been limited to studies with small sample sizes, short follow-ups, and only a descriptive characterization of histology. In this study of 936 patients followed up for a median of 6.4 years after establishing a new baseline kidney function following radical nephrectomy, the authors used morphometric methods to quantitatively characterize microstructural features on large renal wedge sections. Findings of larger nephrons and more extensive glomerulosclerosis and interstitial fibrosis predicted progressive CKD; interstitial fibrosis also predicted mortality from causes other than cancer. These findings provide insight into the important microstructural features of “generic” CKD in patients without a specific kidney disease and support the use of quantitative methods to evaluate renal histology after a radical nephrectomy to determine patients’ long-term risks. Background Nephron hypertrophy and nephrosclerosis may be important determinants of CKD and mortality. However, studies of outcomes associated with these microstructural features have been limited to small tissue specimens from patients selected for either good kidney health or known kidney disease. Methods To determine whether microstructural features are predictive of progressive CKD and mortality outcomes, we studied patients who underwent a radical nephrectomy for a tumor. Large wedge sections of renal parenchyma distal to the tumor were stained and scanned into high-resolution images; we annotated the cortex and all glomeruli to calculate glomerular volume, cortex volume per glomerulus, and percentage of globally sclerotic glomeruli. Morphometric measurements also included percentages of artery luminal stenosis and interstitial fibrosis/tubular atrophy (IF/TA) of the cortex. At follow-up visits every 6–12 months, we determined which patients experienced progressive CKD (defined as dialysis, kidney transplantation, or a 40% decline from postnephrectomy eGFR). Cox models for these outcomes were adjusted for age, sex, body mass index, hypertension, diabetes, smoking, eGFR, and proteinuria. Results Among 936 patients (mean age, 64 years; postnephrectomy baseline eGFR, 48 ml/min per 1.73 m 2 ), 117 progressive CKD events, 183 noncancer deaths, and 116 cancer deaths occurred during a median follow-up of 6.4 years. Larger glomerular volume, larger cortex per glomerulus, and higher percentage of globally sclerotic glomeruli or IF/TA predicted progressive CKD. Higher percentage IF/TA also predicted noncancer mortality. Microstructural features did not predict cancer mortality or recurrence. Conclusions After a radical nephrectomy, larger nephrons and nephrosclerosis predicted progressive CKD, and IF/TA predicted noncancer mortality. Morphometric analysis of renal parenchyma can predict noncancer clinical events in patients long after their radical nephrectomy.

Kidney diseases secondary to several pathogeneses affect millions of people worldwide and have become increasingly recognized as a global public health problem. Recent evidence suggests that cellular senescence plays an important role in the pathogenesis of different forms of renal damage, including acute and chronic kidney disease, and renal transplantation. Renal senescence involves cell cycle arrest and affects several cellular pathways, manifesting in downregulation of klotho, elevated expression of cyclin-dependent kinase inhibitors, cellular telomere shortening, and oxidative stress. Furthermore, senescent cells might induce kidney injury by paracrine release of inflammatory factors. Yet, cellular senescence may be renoprotective during development and in some models of renal diseases, reflecting the yin/yang duality of cellular senescence. This review provides an overview of the role of this emerging player in renal injury, with emphasis on new findings of cellular senescence.

Objectives Magnetization transfer imaging is a novel noninvasive tool for the detection of kidney fibrosis, but its association with kidney function and hemodynamics is unclear. Renovascular disease (RVD) associated with metabolic derangements elicits renal fibrosis and is often unresponsive to percutaneous transluminal angioplasty (PTRA), but tools to predict therapeutic success are unavailable. We hypothesized that magnetization transfer imaging predicts kidney recovery after PTRA in swine with unilateral RVD. Materials and Methods Stenotic kidney (STK) and contralateral kidney magnetization transfer ratios (MTRs; M t/M 0) were measured at 3.0-T magnetic resonance imaging, at offset frequencies of 600 and 1000 Hz, before and 1 month post-PTRA in 7 RVD pigs. Stenotic kidney MTR was correlated to renal perfusion, renal blood flow (RBF), and glomerular filtration rate (GFR), determined using multidetector computed tomography and with ex vivo renal fibrosis (trichrome staining). Untreated RVD (n = 6) and normal pigs (n = 7) served as controls. Results Renovascular disease induced hypertension and renal dysfunction. Blood pressure and renal perfusion were unchanged post-PTRA, but GFR and RBF increased. Baseline cortical STK-MTR predicted post-PTRA renal perfusion and RBF, and MTR changes associated inversely with changes in perfusion and normalized GFR. Stenotic kidney MTR at 600 Hz showed closer association with renal parameters, but both frequencies predicted post-PTRA cortical fibrosis. Conclusions Renal STK-MTR, particularly at 600 Hz offset, is sensitive to hemodynamic changes after PTRA in swine RVD and capable of noninvasively predicting post-PTRA kidney perfusion, RBF, and fibrosis. Therefore, STK-MTR may be a valuable tool to predict renal hemodynamic and functional recovery, as well as residual kidney fibrosis after revascularization in RVD.

Background We examined feasibility of a unique approach towards gaining insight into heritable risk for early atherosclerosis: surveying gene expression by endothelial cells from living subjects. Methods and Results Subjects aged <50 years (mean age, 37; range, 22–49) without obstructive coronary artery disease underwent coronary reactivity testing that identified them as having normal or abnormal coronary endothelial function. Cultures of Blood Outgrowth Endothelial Cells (BOEC) from 6 normal and 13 abnormal subjects passed rigorous quality control and were used for microarray assessment of gene expression. Of 9 genes differentially expressed at false discovery rate <0.1%, we here focus upon abnormal subjects having elevated expression of HMGB1 (high mobility group box 1) which we unexpectedly found to be linked to low LAMC1 (laminin gamma 1) expression. This linkage was corroborated by 3 of our past studies and confirmed bio‐functionally. Compared with normal BOEC, abnormal BOEC released 13±3‐fold more HMGB1 in response to lipopolysaccharide; and they deposited one tenth as much LAMC1 into collagen subendothelial matrix during culture. Clinical follow‐up data are provided for 4 normal subjects (followed 13.4±0.1 year) and for 12 abnormal subjects (followed 9.1±4.5 years). Conclusions The known pathogenic effects of high‐ HMGB1 and low‐ LAMC1 predict that the combination would biologically converge upon the focal adhesion complex, to the detriment of endothelial shear responsiveness. This gene expression pattern may comprise a heritable risk state that promotes early coronary atherosclerosis. If so, the testing could be applied even in childhood, enabling early intervention. This approach offers a way to bridge the information gap between genetics and clinical phenotype.

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.