Tracing angiotensin II's yin-yang effects on cardiovascular-renal pathophysiology.

Adverse changes in cardiovascular and renal systems are major contributors to overall morbidity and mortality. Human cardiovascular and renal systems exhibit a complex network of positive and negative feedback that is reflected in the control of vascular tone via angiotensin II (Ang II) based signaling. This review will examine in some depth, the multiple components and processes that control the status and reflect the health of these various cardiovascular and renal systems, such as pathways associated to monomeric G proteins, RhoA/Rho kinase system and ERK, oxidative stress and NO balance. It will specifically emphasize the "yin-yang" nature of Ang II signaling by comparing and contrasting the effects and activity of various systems, pathways and components found in hypertension to those found in Gitelman's and Bartter's syndromes (GS/BS), two rare autosomal recessive tubulopathies characterized by electrolytic imbalance, metabolic alkalosis, sodium wasting and prominent activation of the renin-angiotensin-aldosterone system. Notwithstanding the activation of the renin-angiotensin-aldosterone system, GS/BS are normo-hypotensive and protected from cardiovascular-renal remodeling and therefore can be considered the mirror image, the opposite of hypertension.

The Assessment of Renal Functional Reserve in ß-Thalassemia Major Patients by an Innovative Ultrasound and Doppler Technique: A Pilot Study.

Beta-thalassemia syndromes are the most common inherited monogenic disorders worldwide. The most common pathophysiologic and clinical renal disease manifestations of in ß-TM patients is the tubular dysfunctions related to iron overload, chronic anemia, and the need for chronic iron chelation therapy. The aim of this pilot study is to apply an innovative ultrasound and Doppler technique to assess the Renal Functional Reserve (RFR) in ß-TM patients, and to evaluate its reliability in iron overload tubulopathy. Ultrasound assessment of intra-parenchymal renal resistive index variation (IRRIV) has recently been proposed as a safe and reproducible technique to identify RFR presence. We define the preserved RFR when the Delta Renal Resistive Index (RRI) is >0.05 (baseline RRI­minimum RRI value during stress) in the Renal Stress Test (RST). Nineteen ß-TM patients were enrolled for this study. In our series, we found a strong negative correlation between mean ferritin values and Delta RRI (R = −0.51, p = 0.03). This pilot study suggested the RST as reliable tool for assessing the RFR by ultrasound. Specifically, RST could help in clinical practice suggesting the patient's management and iron chelation therapy.

[Tolvaptan in ADPKD patients at the University of Padova Nephrology Unit: impact on quality of life, efficacy and safety].

Autosomal dominant polycystic kidney disease (ADPKD) is responsible of the 10% of the dialysis patients. Tolvaptan is a consolidate option for treatment of ADPKD patients; it slows renal deterioration rate and cysts' growth, although its acquaretic effects often impact on quality of life (QoL) and treatment adherence. Few studies have documented the tolvaptan long term efficacy and safeness profiles and, mostly, the impact of treatment with tolvaptan on patients' QoL. Our study aimed to investigate in 13 ADPKD patients of our cohort the differences in terms of QoL before and after the start of treatment via a questionnaire based on SF-36 and PSQI tests, integrated with other original questions. In addition we have also examined the tolvaptan long term efficacy and safeness profiles. The results of our study show that tolvaptan does not significantly reduce patients QoL notwithstanding its expected acquaretic effects, the only reported side effects. Finally, the average annual renal deterioration rate was lower in patients treated with tolvaptan than in the others. Relevant limits of our study are the small number of selected patients and the relative short study duration. However, on one hand, the results of our study provide further information to the few data available in literature; on the other hand, they may serve as a useful working hypothesis for further studies with a larger number of patients enrolled and an extended study duration. They would demonstrate the absence of significant impact of tolvaptan on patients' QoL.

[Lymphocytic leukopenia in two patients affected by polycystic kidney disease waiting for renal transplantation].

Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease, responsible for 10% of patients on renal replacement therapy. The disease is well known to be associated with many extrarenal manifestations. Leukopenia may also be present, even if it is not commonly identified as a typical extrarenal manifestation. Herein we describe two case reports of ADPKD patients with leukopenia. The first case is about a 47-year-old patient affected by ADPKD, regularly treated with peritoneal dialysis, who showed a progressive reduction of white blood cell count, mostly of lymphocytes. Lymphocytic leukopenia was so severe that, when he was called for transplantation from a deceased donor, he was considered temporarily not eligible. We then describe a second ADPKD patient regularly treated with peritoneal dialysis, who had stable lymphopenia for years. Six years after starting PD, it was necessary to perform bone marrow aspirate to investigate the simultaneous presence of hypogammaglobulinemia together with M-protein and to exclude monoclonal gammopathy. All the exams performed did not show any significant results, the patients were re-included in the waiting list and one of them was transplanted. Given our experience and what is reported in the literature, there seems to be enough evidence to consider leukopenia as an extrarenal manifestation of ADPKD. However, the clinical significance of leukopenia in ADPKD patients is not known. It could be interesting to investigate the leucocytes' function and if ADPKD patients with leukopenia are more susceptible to infection, or not. Moreover, it would be very useful to analyze the relationship between such manifestation and genotype/phenotype.

[Reduced oxidative stress in ADPKD patients treated with tolvaptan].

Autosomal dominant polycystic kidney disease (ADPKD) is the most frequent monogenic kidney disease. It causes hypertension and progressive renal failure, both strictly linked to oxidative stress (OxSt). Treatment with tolvaptan is a consolidate option which slows renal deterioration rate, although the molecular mechanisms involved are not fully clarified. We evaluated the OxSt state in tolvaptan-treated ADPKD patients, untreated patients and healthy subjects. OxSt was assessed in 9 patients for each group as mononuclear cell protein expression, MYPT-1 phosphorylation state (Western blot) and heme oxygenase (HO-1) (ELISA). p22 phox protein expression was lower in tolvaptan treated ADPKD and controls compared to untreated patients: 0.86 ±0.15 d.u. p=0.015; 0.53 ±0.11, p<0.001; 1.42 ±0.11 respectively. The same was observed for phosphorylated MYPT-1: 0.68 ±0.09, p=0.013 and vs 0.47 ±0.13, p<0.001, 0.96 ±0.28, while HO-1 of untreated patients was significantly lower compared to treated and controls: 5.33 ±3.34 ng/mL, 2.08 ±0.79, p=0.012, 1.97 ±1.22, p=0.012. Tolvaptan-treated ADPKD patients have reduced OxSt, which might contribute to slowing down the loss of renal function.

Effects of Tolvaptan on Oxidative Stress in ADPKD: A Molecular Biological Approach.

Autosomal dominant polycystic disease (ADPKD) is the most frequent monogenic kidney disease. It causes progressive renal failure, endothelial dysfunction, and hypertension, all of which are strictly linked to oxidative stress (OxSt). Treatment with tolvaptan is known to slow the renal deterioration rate, but not all the molecular mechanisms involved in this effect are well-established. We evaluated the OxSt state in untreated ADPKD patients compared to that in tolvaptan-treated ADPKD patients and healthy subjects. OxSt was assessed in nine patients for each group in terms of mononuclear cell p22phox protein expression, NADPH oxidase key subunit, MYPT-1 phosphorylation state, marker of Rho kinase activity (Western blot) and heme oxygenase (HO)-1, induced and protective against OxSt (ELISA). p22phox protein expression was higher in untreated ADPKD patients compared to treated patients and controls: 1.42 ± 0.11 vs. 0.86 ± 0.15 d.u., p = 0.015, vs. 0.53 ± 0.11 d.u., p < 0.001, respectively. The same was observed for phosphorylated MYPT-1: 0.96 ± 0.28 vs. 0.68 ± 0.09 d.u., p = 0.013 and vs. 0.47 ± 0.13 d.u., p < 0.001, respectively, while the HO-1 expression of untreated patients was significantly lower compared to that of treated patients and controls: 5.33 ± 3.34 vs. 2.08 ± 0.79 ng/mL, p = 0.012, vs. 1.97 ± 1.22 ng/mL, p = 0.012, respectively. Tolvaptan-treated ADPKD patients have reduced OxSt levels compared to untreated patients. This effect may contribute to the slowing of renal function loss observed with tolvaptan treatment.

Genotype-phenotype correlation in Gordon's syndrome: report of two cases carrying novel heterozygous mutations.

Gordon's syndrome, known also as Pseudohypoaldosteronism type II is a rare inherited dominant form of low-renin hypertension associated with hyperkalemia and metabolic acidosis. Four genes related to the regulation of the NaCl co-symporter NCC have been discovered associated to Gordon phenotypes: WINK 1 and WINK4, which, along with WNK2 and WNK3, encode a family of WNK-kinases, and KLHL3 and CUL3 encoding respectively, Kelch-like 3 protein and cullin. Heterozygous mutations in these genes constitutively activate NCC leading to abnormally increased salt reabsorption and salt-sensitive hypertension. Thiazide diuretic is the recognized treatment for this condition. We report and discuss phenotypic and genetic heterogeneity of two patients with Gordon's syndrome carrying novel heterozygous mutations in the WNK1 and KLHL3 genes. A very rare variant in the SCNN1G gene encoding the γ subunit of epithelial sodium channel ENaC was also identified in one patient.

[Slowing progression of chronic kidney disease in polycistic kidney disease patients with tolvaptan: from guidelines to clinical practice].

Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease and accounts for∼10% of patients on renal replacement therapy. In the last decade, no specific treatment was available and only preventive measures could be put in place to delay the onset of ESRD. Following the results of the TEMPO 3:4 study, tolvaptan was approved in many countries, for the purpose of slowing the progression of renal insufficiency. In Italy tolvaptan is available since 2016 for patients with chronic kidney disease (CKD) stage 1-3, and since 2020 for patients with CKD stage 4, who fulfil the criteria of "rapid disease progression", according to the European recommendations. After this approval, Italian nephrology units have had to change their organization to be able to identify the patients eligible for the drug and to guarantee frequent patient monitoring. In this paper, we present our three-year experiences with tolvaptan, focusing on its safety profile and tolerability, but also on the high burden of care that such therapy represents not only for doctors, but also for patients. Strategies to implement remote monitoring may be useful to reduce the burden of assistance on one side, and the medicalization of ADPKD patients in the early stage of the disease, on the other.

Oxidative stress, inflammation, and peritoneal dialysis: A molecular biology approach.

The key role of oxidative stress (OxSt) and inflammation for the induction of cardiovascular disease, the leading cause of excess morbidity/mortality in chronic kidney disease and dialysis patients, is known and both the activations of NADPH oxidase and RhoA/Rho kinase (ROCK) pathway are pivotal for their effects. While specific hemodialysis procedures, such as hemodiafiltration with on-line reinfusion of ultrafiltrate and/or the use of vitamin E-coated dialyzers, are beneficial for OxSt and inflammation, studies in peritoneal dialysis (PD) are instead scarce and results seem not favorable. In nine patients under PD OxSt in terms of mononuclear cell protein level of p22phox (Western blot), subunit of NADPH oxidase, essential for the generation of OxSt, and MYPT-1 phosphorylation state (Western blot), a marker of ROCK activity, have been measured at the beginning and after 3 and 6 months of PD. Blood levels of interleukin 6 (IL-6), ferritin, and albumin have been considered for evaluating the inflammatory state. p22phox protein expression, MYPT-1-phosphorylation, and ferritin level were increased both at baseline vs healthy subjects (P = .02, P < .0001, P = .004, respectively) and vs baseline after 3 and 6 months of peritoneal dialysis (P = .007, P < .001, P = .004, respectively). Albumin was lower after 6 months of PD (P = .0014). IL-6 was increased at baseline vs reference values and remained unchanged at 3 and 6 months. OxSt and inflammation increase during PD confirming via molecular biology approach a report at biochemical level. To improve OxSt state in PD, a multitarget approach is necessary. It might include the use of more physiologic pH, low glucose degradation products, low lactate and iso-osmolar PD solutions, patients' strict glycemic control, optimal volume management, and antioxidant administration, such as N-acetylcysteine.

ACE2 and SARS-CoV-2 Infection Risk: Insights From Patients With Two Rare Genetic Tubulopathies, Gitelman's and Bartter's Syndromes.

COVID-19 is spreading globally with the angiotensin converting enzyme (ACE)-2 serving as the entry point of SARS-CoV-2 virus. This raised concerns how ACE2 and the Renin-Angiotensin (Ang)-System (RAS) are to be dealt with given their roles in hypertension and their involvement in COVID-19's morbidity and mortality. Specifically, increased ACE2 expression in response to treatment with ACE inhibitors (ACEi) and Ang II receptor blockers (ARBs) might theoretically increase COVID-19 risk by increasing SARS-CoV-2 binding sites. However, ACE2 is part of the protective counter-regulatory ACE2-Ang1-7-MasR axis, which opposes the classical ACE-AngII-AT1R regulatory axis. We used Gitelman's and Bartter's syndromes (GS/BS) patients, rare genetic tubulopathies that have endogenously increased levels of ACE2, to explore these issues. Specifically, 128 genetically confirmed GS/BS patients, living in Lombardia, Emilia Romagna and Veneto, the Northern Italy hot spots for COVID-19, were surveyed via telephone survey regarding COVID-19. The survey found no COVID-19 infection and absence of COVID-19 symptoms in any patient. Comparison analysis with the prevalence of COVID-19 in those regions showed statistical significance (p < 0.01). The results of the study strongly suggest that increased ACE2 does not increase risk of COVID-19 and that ACEi and ARBs by blocking excessive AT1R-mediated Ang II activation might favor the increase of ACE2-derived Ang 1-7. GS/BS patients' increased ACE2 and Ang 1-7 levels and their characteristic chronic metabolic alkalosis suggest a mechanism similar to that of chloroquine/hydroxychloroquine effect on ACE2 glycosylation alteration with resulting SARS-COV-2 binding inhibition and blockage/inhibition of viral entry. Studies from our laboratory are ongoing to explore GS/BS ACE2 glycosylation and other potential beneficial effects of BS/GS. Importantly, the absence of frank COVID-19 or of COVID-19 symptoms in the BS/GS patients cohort, given no direct ascertainment of COVID-19 status, suggest that elevated ACE2 levels as found in GS/BS patients at a minimum render COVID-19 infection asymptomatic and thus that COVID-19 symptoms are driven by ACE2 levels.

Clinical Evidence for the Choice of the Direct Oral Anticoagulant in Patients with Atrial Fibrillation According to Creatinine Clearance.

Atrial fibrillation (AF) often coexists with chronic kidney disease (CKD), which confer to the patient a higher risk of both thromboembolic and hemorrhagic events. Oral anticoagulation therapy, nowadays preferably with direct oral anticoagulants (DOACs), represents the cornerstone for ischemic stroke prevention in high-risk patients. However, all four available DOACs (dabigatran, apixaban, rivaroxaban and edoxaban) are eliminated by the kidneys to some extent. Reduced kidney function facilitates DOACs accumulation and, therefore, different dose reductions are required, with slight differences between American and European recommendations especially in case of severe renal impairment (creatinine clearance < 30 mL/min). Overall, the use of DOACs in patients with non-end stage CKD and AF is effective similarly to warfarin, showing a better safety profile. The management of thromboembolic risk among patients with AF on dialysis remains challenging, as warfarin effectiveness for stroke prevention in this population is questionable and retrospective data on apixaban need to be confirmed on a larger scale. In kidney transplant recipients, DOACs may provide a potentially safer option compared to warfarin, but co-administration with immunosuppressants is a matter of concern.

ROCK (RhoA/Rho Kinase) in Cardiovascular-Renal Pathophysiology: A Review of New Advancements.

Rho-associated, coiled-coil containing kinases (ROCK) were originally identified as effectors of the RhoA small GTPase and found to belong to the AGC family of serine/threonine kinases. They were shown to be downstream effectors of RhoA and RhoC activation. They signal via phosphorylation of proteins such as MYPT-1, thereby regulating many key cellular functions including proliferation, motility and viability and the RhoA/ROCK signaling has been shown to be deeply involved in arterial hypertension, cardiovascular-renal remodeling, hypertensive nephropathy and posttransplant hypertension. Given the deep involvement of ROCK in cardiovascular-renal pathophysiology and the interaction of ROCK signaling with other signaling pathways, the reports of trials on the clinical beneficial effects of ROCK's pharmacologic targeting are growing. In this current review, we provide a brief survey of the current understanding of ROCK-signaling pathways, also integrating with the more novel data that overall support a relevant role of ROCK for the cardiovascular-renal physiology and pathophysiology.

Intravenous ferric carboxymaltose for iron deficiency anemia in dialysis patients: Effect of a new protocol adopted for a hemodialysis limited assistance center.

Iron and erythropoietin deficiencies are determinants of anemia in chronic kidney disease. In hemodialysis (HD) patients, intravenous (IV) iron is associated with a greater hemoglobin (Hb) production and better erythropoietin response but may be associated to hypersensitivity reaction. After the 2013 European Medicines Agency report regarding early detection/management of iron allergic reactions, IV iron administration dramatically reduced in Italian Hemodialysis-Limited-Assistance-Centre (HD-CAL) where a physician is present only once a week. Objective of the study was providing an effective and secure IV iron administration protocol for HD-CAL patients. IV ferric carboxymaltose (FCM) administration was more effective and better tolerated than sodium ferric gluconate for iron deficiency correction and resolution of anemia in 24 patients undergoing HD in our HD-CAL. Six months of FCM IV treatment once a week increased ferritin and Hb compared to sodium ferric gluconate once a week leading to decreased erythropoietin consumption from 24 000 to 15 000 U/patient/week with an erythropoietin annual expense reduction. No blood transfusions, gastrointestinal intolerance or other adverse effects were reported. The FCM IV administration protocol for our HD-CAL patients was safe and no adverse events were reported, resulting in significantly increased ferritin, transferrin saturation, and Hb levels, reduction of erythropoietin requirements, and consequently reduction of erythropoietin expenses.

Rho Kinase Activity, Connexin 40, and Atrial Fibrillation: Mechanistic Insights from End-Stage Renal Disease on Dialysis Patients.

Evidence on cellular/molecular mechanisms leading to atrial fibrillation (AF) are scanty. Increased expression of Rho kinase (ROCK) and myosin-phosphatase-target subunit-1 (MYPT-1), ROCK activity's marker, were shown in AF patients, which correlated with connexin 40 (Cx40) expression, membrane protein of heart gap junctions, key for rapid action potential's cell-cell transfer. AF is the most frequent arrhythmia in dialysis patients who present increased MYPT-1 phosphorylation, which correlates with left ventricular (LV) mass. Given ROCK's established role in cardiovascular-renal remodeling, induction of impaired cell-to-cell coupling/potential conduction promoting AF initiation/perpetuation, we evaluated in dialysis patients with AF, MYPT-1 phosphorylation, Cx40 expression, and their relationships to support their involvement in AF. Mononuclear cells' MYPT-1 phosphorylation, Cx40 expression, and the ROCK inhibitor fasudil's effect were assessed in dialysis patients with AF (DPAFs), dialysis patients with sinus rhythm (DPs), and healthy subjects (C) (western blot). M-mode echocardiography assessed LV mass and left atrial systolic volume. DPAF's phospho-MYPT-1 was increased vs. that of DPs and C (1.57 ± 0.17 d.u. vs. 0.69 ± 0.04 vs. 0.51 ± 0.05 respectively, p < 0.0001). DP's phospho-MYPT-1 was higher vs. that of C, p = 0.009. DPAF's Cx40 was higher vs. that of DPs and C (1.23 ± 0.12 vs. 0.74 ± 0.03 vs. 0.69 ± 0.03, p < 0.0001). DPAF's phospho-MYPT-1 correlated with Cx40 (p < 0.001), left atrial systolic volume (p = 0.013), and LV mass (p = 0.014). In DPAFs, fasudil reduced MYPT-1 phosphorylation (p < 0.01) and Cx40 expression (p = 0.03). These data point toward ROCK and Cx40's role in the mechanism(s) leading to AF in dialysis patients. Exploration of the ROCK pathway in AF could contribute to AF generation's mechanistic explanations and likely identify potential pharmacologic targets for translation into treatment.

Ultrasound for the Clinical Management of Vascular Access Cannulation and Needle Position in Hemodialysis Patients.

A native arteriovenous fistula (AVF) is the vascular access of choice for hemodialysis (HD) treatment. Compared with other types of vascular access such as grafts and central venous catheters, it functions longer and is associated with a lower risk of complications. The aim of the study described here was to assess, in an HD population, the position of the fistula needles during an HD session and evaluate the role of ultrasound in the management of AVF puncture. Forty-five consecutive chronic HD patients with an AVF or an arteriovenous vascular graft were included in the study for ultrasound evaluation. Each patient underwent an ultrasound evaluation during HD treatment to assess the position of the needles inside the vascular access. The ultrasound evaluation revealed that 81.8% of the traditional needles were incorrectly adjacent to the vessel walls, in the absence of clinical symptoms or hemodynamic alterations detectable on the dialysis monitor. A greater frequency of malpositioning has been observed for needles in the arterial portion of the vascular access, closer to the anastomosis. The absence of clinically detectable signs of venipuncture-related complications does not ensure correct positioning of the needles within the AVF. Ultrasound evaluation may not only resolve suboptimal cannulation problems of new or complicated vascular accesses but may also be useful in the prevention of acute and chronic damage to the AVF.