Lerman, Lilach O.

Scattered tubular-like cells (STCs), dedifferentiated renal tubular epithelial cells, contribute to renal self-healing, but severe injury might blunt their effectiveness. We hypothesized that ischemic renovascular disease (RVD) induces senescence in STC and impairs their reparative potency. CD24+/CD133+ STCs were isolated from swine kidneys after 16 weeks of RVD or healthy controls. To test their reparative capabilities in injured kidneys, control or RVD-STC (5×10 5 ) were prelabeled and injected into the aorta of 2 kidneys, 1-clip (2k,1c) mice 2 weeks after surgery. Murine renal function and oxygenation were studied in vivo 2 weeks after injection using micro-magnetic resonance imaging, and fibrosis, tubulointerstitial injury, capillary density, and expression of profibrotic and inflammatory genes ex vivo. STC isolated from swine RVD kidneys showed increased gene expression of senescence and senescence-associated secretory phenotype markers and positive SA-β-gal staining. Delivery of normal pig STCs in 2k,1c mice improved murine renal perfusion, blood flow, and glomerular filtration rate, and downregulated profibrotic and inflammatory gene expression. These renoprotective effects were blunted using STC harvested from RVD kidneys, which also failed to attenuate hypoxia, fibrosis, tubular injury, and capillary loss in injured mouse 2k,1c kidneys. Hence, RVD may induce senescence in endogenous STC and impair their reparative capacity. These observations implicate cellular senescence in the pathophysiology of ischemic kidney disease and support senolytic therapy to permit self-healing of senescent kidneys.

Abstract BACKGROUND Renovascular hypertension (RVH) often manifest with metabolic syndrome (MetS) as well. Coexisting MetS and hypertension increases cardiovascular morbidity and mortality, but the mechanisms underlying cardiac injury remain unknown. We hypothesized that superimposition of MetS induces myocardial mitochondrial damage, leading to cardiac injury and dysfunction in swine RVH. METHODS Pigs were studied after 16 weeks of diet-induced MetS with or without RVH (unilateral renal artery stenosis), and Lean controls (n = 6 each). Systolic and diastolic cardiac function were assessed by multidetector CT, and cardiac mitochondrial morphology (electron microscopy) and myocardial function in tissue and isolated mitochondria. RESULTS Body weight was similarly higher in MetS groups vs. Lean. RVH groups achieved significant stenosis and developed hypertension. Mitochondrial matrix density and adenosine triphosphate production were lower and H2O2 production higher in RVH groups vs. Lean and MetS. Lean + RVH (but not MetS + RVH) activated mitophagy, which was associated with decreased myocardial expression of mitophagy-related microRNAs. MetS groups exhibited higher numbers of intermitochondrial junctions, which could have prevented membrane depolarization/activation of mitophagy in MetS + RVH. Cardiac fibrosis, hypertrophy (increased left ventricular muscle mass), and diastolic function (decreased E/A ratio) were greater in MetS + RVH vs. Lean + RVH. CONCLUSIONS MetS+RVH induces myocardial mitochondrial damage and dysfunction. MetS + RVH failed to activate mitophagy, resulting in greater cardiac remodeling, fibrosis, and diastolic dysfunction. Mitochondrial injury and impaired mitophagy may constitute important mechanisms and therapeutic targets to ameliorate cardiac damage and dysfunction in patients with coexisting MetS and RVH.

Abstract Background Heart failure (HF) is a debilitating disease and is associated with significant morbidity and mortality, and costs to health care systems. Monitoring the impact of therapy remotely holds the potential to reduce HF-related hospitalizations, improve quality of life, and optimize the use of the limited resources. Voice signal is an emerging non-invasive biomarker that has been associated with a number of disease states. We previously identified a significant relationship between specific vocal biomarkers and coronary artery disease, and recently extended these observations by showing that a pre-specified voice biomarker was associated with increased mortality and re-hospitalization in patients with HF. Purpose In the current study, we evaluated the association between a vocal biomarker derived from voice signal analysis and invasively measured indices of pulmonary hypertension (PH). We hypothesized that the pre-specified voice biomarker might be associated with hemodynamic indices reflective of PH that are known to be linked to HF-severity, and that predict outcomes such as HF-related hospitalization and death. Methods The study population included patients referred for an invasive cardiac hemodynamic study between January 2017 and December 2018. Subjects had their voice signal recorded to their smartphone on three separate occasions prior to the cardiac study. A pre-established numeric vocal biomarker was derived from each recording, and the mean vocal biomarker calculated for each patient. Patients were a priori divided into two groups: those with high pulmonary arterial pressure (PAP) defined as ≥35 mmHg consistent with moderate or greater PH, versus those with a lower PAP. Results Eighty three patients, mean age 61.6±15.1 years, 37 (44.6%) male, were included in the study. The intraclass correlation coefficient for the vocal biomarker in all patients was 0.83 implying very good agreement between values. Patients with a high mean PAP (≥35 mmHg) had significantly higher values of the voice biomarker compared to those with a lower mean PAP (0.74±0.85 vs. 0.43±0.86, p=0.008). Patients with a high pulmonary vascular resistance (PVR) defined as a PVR ≥1.7 Wood Units had significantly higher values of the voice biomarker compared to those with a lower PVR (0.62±0.83 vs. 0.33±0.90, p=0.026). Multivariable logistic regression showed that an increase in the voice biomarker by 1 unit was significantly associated with a high PAP, odds ratio (OR) 2.31, 95% CI 1.05–5.07, p=0.038, and with borderline significance with a high PVR, OR 2.14, 95% CI 0.94–4.87, p=0.07. Conclusion The current study shows a relationship between a noninvasive vocal biomarker derived from voice signal analysis and invasively derived hemodynamic indices related to PH obtained during cardiac catheterization. These results may have important and practical clinical implications for telemedicine and remote monitoring of patients with HF and PH. Funding Acknowledgement Type of funding source: Private grant(s) and/or Sponsorship. Main funding source(s): Mayo Foundation; Beyond Verbal Communications

Background. Coexisting metabolic syndrome (MetS) and renal artery stenosis (RAS) are linked to poor renal outcomes. Mesenchymal stem/stromal cell- (MSC-) derived extracellular vesicles (EVs) from lean animals show superior ability to repair the experimental MetS+RAS kidney compared to EVs from MetS pig MSCs. We hypothesized that MetS leads to selective packaging in porcine EVs of microRNAs capable of targeting mitochondrial genes, interfering with their capacity to repair the MetS+RAS kidney. Methods. Five groups of pigs ( n = 7 each) were studied after 16 weeks of diet-induced MetS and RAS (MetS+RAS) and MetS+RAS 4 weeks after a single intrarenal delivery of EVs harvested from allogeneic adipose tissue-derived MSCs isolated from Lean or MetS pigs, and Lean or MetS sham controls. Single-kidney blood flow (RBF) and glomerular filtration rate (GFR) were assessed in vivo with multidetector CT, whereas EV microRNA cargo, renal tubular mitochondrial structure and bioenergetics, and renal injury pathways were assessed ex vivo. Results. microRNA sequencing revealed 19 dysregulated microRNAs capable of targeting several mitochondrial genes in MetS-EVs versus Lean-EVs. Lean- and MetS-EVs were detected in the stenotic kidney 4 weeks after administration. However, only MetS-EVs failed to improve renal mitochondrial density, structure, and function or attenuate oxidative stress, tubular injury, and fibrosis. Furthermore, Lean-EVs but not MetS-EVs restored RBF and GFR in MetS+RAS. Conclusion. MetS alters the cargo of mitochondria-related microRNAs in swine MSC-derived EVs, which might impair their capacity to repair the poststenotic kidney in MetS+RAS. These observations may contribute to develop approaches to improve the efficacy of MSC-EVs for patients with MetS.

Introduction: Recent evidence has demonstrated that Clonal Hematopoiesis of Indeterminate Potential (CHIP) increases the risk of cardiovascular events. However, the exact role played by CHIP in early stages of coronary atherosclerosis, characterized by coronary endothelial dysfunction remains to be elucidated. Hypothesis: The current study was designed to test the hypothesis that the presence of CHIP in peripheral blood cells is associated with coronary endothelial dysfunction and increased levels of inflammatory markers. Methods: Next generation sequencing was used to detect mutations among patients who had coronary endothelial dysfunction and control group (normal coronary endothelial function). Endothelial dysfunction was defined by ≥ 20% decrease in coronary artery diameter or ≤50% increase in blood flow in response to acetylcholine injection compared to baseline. Plasma cytokine levels of Interleukin (IL)-6 and IL-8 were also assessed. Results: Clonal hematopoiesis relevant gene mutations were found in 1 out of 64 patients with normal endothelial function (1.5%) and 11 out of 119 cases in the endothelial dysfunction group (9.2%) (P = 0.04) with ASXL1 (Additional sex combs-like 1) being the most frequent gene (4.2%) mutated. Cytokine analysis demonstrated that mutations in ASXL1, DNMT3A (DNA methyltransferase 3A) and TET2 (Ten-eleven-translocation-2) in the endothelial dysfunction group were associated with increased levels of IL-6 and IL-8 (P = 0.001, P = 0.003; respectively). Conclusions: The current study supports a potential role for CHIP in mechanisms of coronary artery disease starting at early stage of atherosclerosis. Furthermore, enhanced expression levels of IL-6 and IL-8 seem to be related to mutations in DNMT3A, ASXL1 and TET2 genes, more than other gene mutations relevant CHIP.

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.

Chronic kidney disease is a global health care burden, yet clinically-proven treatments are limited. Low-intensity shockwave, which utilizes ≈10% of the energy levels used in clinically indicated shockwave lithotripsy, is a promising technique to ameliorate ischemia and regenerate tissues. It has been demonstrated to improve healing in tissues such as bone, muscle, myocardium, and kidney via several mechanisms, particularly through promoting neovascularization. Low-intensity shockwave stimulates mechanoreceptors located primarily in endothelial and proximal tubular cells and subsequently upregulates vascular endothelial growth factors. This, in turn, promotes angiogenesis and ameliorates renal hypoxia, inflammation, and fibrosis, and ultimately preserves renal function. Furthermore, low-intensity shockwave can stimulate release of homing factors to attract endothelial progenitor or stem cells into injured kidneys for tissue repair. These effects may be beneficial in several kidney disease models, including renal artery stenosis, diabetic kidney disease, and various chronic kidney diseases, although most studies reported to date have been performed in animal models. Because of its low energy intensity, the procedure is relatively tolerable and safe, yet, more clinical studies are needed to establish its efficacy beyond currently existing strategies. Therefore, low-intensity shockwave therapy emerges as an alternative therapeutic approach that may offer a promising noninvasive intervention for treating renal diseases. Registration— URL: https://www.clinicaltrials.gov ; Unique identifier: NCT02515461; NCT03602807; and NCT03445247.

Background Elevated levels of serum homocysteine, via impaired nitric oxide production, and coronary microvascular dysfunction are associated with increased risk of major adverse cardiovascular events. However, whether serum homocysteine levels and coronary microvascular endothelial dysfunction (CMED) are linked remains unknown. Methods and Results This study included 1418 patients with chest pain or an abnormal functional stress test and with nonobstructive coronary artery disease (<40% angiographic stenosis), who underwent CMED evaluation with functional angiography and had serum homocysteine levels measured. Patients were classified as having normal microvascular function versus CMED. Patients in the CMED group (n=743; 52%) had higher mean age (52.1±12.2 versus 50.0±12.4 years; P <0.0001), higher body mass index (29.1 [25.0–32.8] versus 27.5 [24.2–32.4]; P =0.001), diabetes mellitus (12.5% versus 9.4%; P =0.03), and fewer women (63.5% versus 68.7%; P =0.04) compared with patients in the normal microvascular function group. However, they had lower rates of smoking history, and mildly lower low‐density lipoprotein cholesterol levels. Serum homocysteine levels were significantly higher in patients with CMED, and the highest quartile of serum homocysteine level (>9 µmol/L) was an independent predictor of CMED (odds ratio, 1.34 [95% CI, 1.03–1.75]; P =0.03) after adjustment for age; sex; body mass index; chronic kidney disease (CKD); diabetes mellitus; smoking exposure; low‐density lipoprotein cholesterol; high‐density lipoprotein cholesterol and triglycerides; and aspirin, statin, and B vitamin use. Conclusions Patients with CMED have significantly higher levels of serum homocysteine. Elevated serum homocysteine levels were associated with a significantly increased odds of an invasive diagnosis of CMED. The current study supports a potential role for homocysteine for diagnosis and target treatment in the patients with early coronary atherosclerosis.