Role of ADAM17 in kidney disease : a clinical and experimental approach"

Les desintegrines i metal·loproteases (ADAMs) són una família de proteïnases que alliberen dominis proteics solubles en un procés conegut com a "shedding". En concret, l'ADAM17 és el membre de la família millor estudiat. Aquest, es troba altament expressat en les cèl·lules dels túbuls distals i augmenta la seva expressió en totes les cèl·lules del ronyó en la malaltia renal crònica (MRC), la nefropatia diabètica (ND) i la malaltia cardiovascular (CV), entre d'altres. Basant-nos en aquests antecedents, es van dissenyar tres estudis per aquesta tesi doctoral. En primer lloc, es va analitzar l'activitat circulant dels ADAMs en plasmes humans de la cohort NEFRONA que inclou pacients amb MRC sense història prèvia de malaltia CV. Es va avaluar l'associació entre l'activitat circulant dels ADAMs i variables clíniques basals, així com amb la progressió de la funció renal i els events CV. Per altra banda, es va estudiar l'efecte en el ronyó de la deleció de l'Adam17, a nivell endotelial (eAdam17) o tubular proximal (tAdam17), en la histologia, la modulació del sistema renina angiotensina (SRA), la inflamació i la fibrosis en un model murí de diabetis tipus 1. Finalment, en un cultiu 3D de cèl·lules tubulars proximals humanes, en condicions d'alta glucosa, també es va abordar l'efecte de la deleció de l'Adam17. L'estudi en humans va demostrar que l'activitat circulant dels ADAMs era un potent predictor de la progressió de MRC en els homes. A més, s'associava de forma independent amb els events CV en els pacients amb MRC. En relació amb els estudis experimentals, es va veure com la deleció específica de l'eAdam17 atenuava la fibrosis i la inflamació renal, mentre que la deleció del tAdam17 disminuïa la pèrdua podocitària, atenuava el SRA i disminuïa la infiltració per macròfags així com l'acumulació de α-SMA i col·lagen en el ronyó. El cultiu in vitro 3D va reforçar els resultats obtinguts en el model de ratolí tAdam17KO on es va observar una disminució de marcadors fibròtics en els esferoides amb deleció de l'Adam17. En conclusió, la manipulació de l'Adam17 podria ser considerada com una estratègia terapèutica en el tractament de la ND. En pacients amb MRC sense events CV previs, l'activitat circulant dels ADAMs podria utilitzar-se com a biomarcador de malalties renals i cardiovasculars.Las desintegrinas y metaloproteasas (ADAMs) son una familia de proteinasas que liberan dominios proteicos solubles en un proceso conocido como "shedding". En concreto, el ADAM17 es el miembro de la familia mejor estudiado. Éste se encuentra altamente expresado en las células de los túbulos distales y aumenta su expresión en todas las células del riñón en la enfermedad renal crónica (ERC), la nefropatía diabética (ND) y la enfermedad cardiovascular (CV), entre otras. Basándonos en estos antecedentes, se diseñaron tres estudios para esta tesis doctoral. En primer lugar, se analizó la actividad circulante de los ADAMs en plasmas humanos de la cohorte NEFRONA que incluye pacientes con ERC sin historia previa de enfermedad CV. Se evaluó la asociación entre la actividad circulante de los ADAMs y variables clínicas basales, así como la progresión de la función renal y los eventos CV. Por otro lado, se estudió el efecto en el riñón de la deleción del ADAM17 a nivel endotelial (eAdam17) o tubular proximal (tAdam17), en la histología, la modulación del sistema renina angiotensina (SRA), la inflamación y la fibrosis en un modelo murino de diabetes tipo 1. Finalmente, en un cultivo 3D de células tubulares proximales humanas, en condiciones de alta glucosa, también se abordó el efecto de la deleción del Adam17. El estudio en humanos demostró que la actividad circulante de los ADAMs era un potente predictor de la progresión de ERC en los hombres. Además, se asociaba de forma independiente con los eventos CV en los pacientes con ERC. En relación con los estudios experimentales, se vio como la deleción específica del eAdam17 atenuaba la fibrosis y la inflamación renal, mientras que la deleción del tAdam17 disminuía la pérdida de podocitos, atenuaba el SRA y disminuía la infiltración por macrófagos así como la acumulación de α-SMA y colágeno en el riñón. El cultivo in vitro 3D reforzó los resultados obtenidos en el modelo de ratón tAdam17KO donde se observó una disminución de marcadores fibróticos en los esferoides con deleción del Adam17. En conclusión, la manipulación del Adam17 podría ser considerada como una estrategia terapéutica en el tratamiento de la ND. En pacientes con ERC sin eventos CV previos, la actividad circulante de los ADAMs podría utilizarse como biomarcador de enfermedades renales y cardiovasculares.Disintegrins and metalloproteinases (ADAMs) are a family of proteinases that function as sheddases. Especially, ADAM17 has been the best studied family member. ADAM17 is highly expressed in renal distal tubules and is increased in the whole kidney, mainly in endothelial and proximal tubular cells, during chronic kidney disease (CKD), diabetic nephropathy (DN) and cardiovascular (CV) disease, among others. Given these premises, three studies have been proposed in this thesis. Firstly, baseline circulating ADAMs activity was analysed in human plasma samples from the NEFRONA Study, which includes CKD patients without previous history of CV disease. Baseline and prospective studies were performed to evaluate the association of circulating ADAMs activity with baseline clinical variables, and with the progression of renal function and CV outcomes. Secondly, the role of endothelial (eAdam17) and proximal tubular (tAdam17) Adam17 deletion in renal histology, modulation of the renin angiotensin system (RAS), renal inflammation and fibrosis was studied in a mouse model of type 1 Diabetes Mellitus. Thirdly, the effect of Adam17 deletion in an in vitro 3D cell culture from human proximal tubular cells under high glucose conditions was evaluated. The human study showed that circulating ADAMs activity is a powerful predictor of CKD progression in male patients. Furthermore, circulating ADAMs activity is independently associated with CV events in CKD patients. Within the experimental studies, eAdam17 deletion attenuates renal fibrosis and inflammation, whereas tAdam17 deletion decreases podocyte loss, attenuates the RAS, and decreases macrophage infiltration and α-SMA and collagen accumulation. The 3D in vitro cell culture reinforced the findings obtained in tAdam17KO mice with decreased fibrosis in the Adam17 knockout spheroids. In conclusion, the manipulation of Adam17 should be considered as a therapeutic strategy for treating DN. In CKD patients, without previous history of CV disease, circulating ADAMs activity can be useful as a biomarker of renal and CV outcomes

Role of ADAM17 in kidney disease

It is known that the renin-angiotensin system plays a major role in the pathophysiology of cardiovascular disease and renal injury. Within the renin-angiotensin system, angiotensin-converting enzyme 2 (ACE2) cleaves ANG II to generate ANG(1-7) peptide, which counteracts the adverse effects of ANG II accumulation. ACE2 can undergo cleavage or shedding to release the catalytically active ectodomain into the circulation by a disintegrin and metalloprotease (ADAM)17, also known as TNF-α-converting enzyme. ADAM17 is involved in many pathological processes such as cancer, inflammatory diseases, neurological diseases, cardiovascular diseases, atherosclerosis, diabetes, and hypertension. Clinical and experimental studies have shown that ADAM17 is involved in chronic kidney disease (CKD) with a proinflammatory and profibrotic role, suggesting that it could be an important mediator of CKD progression. ADAM17 inhibition attenuates fibrosis and inflammation, suggesting that its inhibition may be a possible new valuable therapeutic tool in fibrotic kidney disease treatment. In addition, in renal disease, some experimental studies have demonstrated that ADAM17 is differently expressed in the kidney. Thus, ADAM17 is highly expressed in distal renal tubules and increased in the whole kidney in diabetic models. In this article, we will review the role of ADAM17 under physiological and pathological conditions. We will mainly focus on the importance of ADAM17 in the context of CKD

Characterization of ACE and ACE2 Expression within Different Organs of the NOD Mouse

Renin angiotensin system (RAS) is known to play a key role in several diseases such as diabetes, and renal and cardiovascular pathologies. Its blockade has been demonstrated to delay chronic kidney disease progression and cardiovascular damage in diabetic patients. In this sense, since local RAS has been described, the aim of this study is to characterize angiotensin converting enzyme (ACE) and ACE2 activities, as well as protein expression, in several tissues of the non-obese diabetic (NOD) mice model. After 21 or 40 days of diabetes onset, mouse serums and tissues were analyzed for ACE and ACE2 enzyme activities and protein expression. ACE and ACE2 enzyme activities were detected in different tissues. Their expressions vary depending on the studied tissue. Thus, whereas ACE activity was highly expressed in lungs, ACE2 activity was highly expressed in pancreas among the studied tissues. Interestingly, we also observed that diabetes up-regulates ACE mainly in serum, lung, heart, and liver, and ACE2 mainly in serum, liver, and pancreas. In conclusion, we found a marked serum and pulmonary alteration in ACE activity of diabetic mice, suggesting a common regulation. The increase of ACE2 activity within the circulation in diabetic mice may be ascribed to a compensatory mechanism of RAS

Both specific endothelial and proximal tubular Adam17 deletion protect against diabetic nephropathy

ADAM17 is a disintegrin and metalloproteinase capable of cleaving the ectodomains of a diverse variety of molecules including TNF-α, TGF-α, L-selectin, and ACE2. We have previously demonstrated that renal ADAM17 is upregulated in diabetic mice. The role of endothelial (eAdam17) and proximal tubular (tAdam17) Adam17 deletion in renal histology, modulation of the renin angiotensin system (RAS), renal inflammation, and fibrosis was studied in a mouse model of type 1 Diabetes Mellitus. Moreover, the effect of Adam17 deletion in an in vitro 3D cell culture from human proximal tubular cells under high glucose conditions was evaluated. eAdam17 deletion attenuates renal fibrosis and inflammation, whereas tAdam17 deletion decreases podocyte loss, attenuates the RAS, and decreases macrophage infiltration, α-SMA and collagen accumulation. The 3D in vitro cell culture reinforced the findings obtained in tAdam17KO mice with decreased fibrosis in the Adam17 knockout spheroids. In conclusion, Adam17 deletion either in the endothelial or the tubular cells mitigates kidney injury in the diabetic mice by targeting different pathways. The manipulation of Adam17 should be considered as a therapeutic strategy for treating DN

Redefining the role of ADAM17 in renal proximal tubular cells and its implications in an obese mouse model of pre-diabetes

Acute and chronic kidney lesions induce an increase in A Disintegrin And Metalloproteinase domain 17 (ADAM17) that cleaves several transmembrane proteins related to inflammatory and fibrotic pathways. Our group has demonstrated that renal ADAM17 is upregulated in diabetic mice and its inhibition decreases renal inflammation and fibrosis. The purpose of the present study was to analyze how Adam17 deletion in proximal tubules affects different renal structures in an obese mice model. Tubular Adam17 knockout male mice and their controls were fed a high-fat diet (HFD) for 22 weeks. Glucose tolerance, urinary albumin-to-creatinine ratio, renal histology, and pro-inflammatory and pro-fibrotic markers were evaluated. Results showed that wild-type mice fed an HFD became obese with glucose intolerance and renal histological alterations mimicking a pre-diabetic condition; consequently, greater glomerular size and mesangial expansion were observed. Adam17 tubular deletion improved glucose tolerance and protected animals against glomerular injury and prevented podocyte loss in HFD mice. In addition, HFD mice showed more glomerular macrophages and collagen accumulation, which was prevented by Adam17 deletion. Galectin-3 expression increased in the proximal tubules and glomeruli of HFD mice and ameliorated with Adam17 deletion. In conclusion, Adam17 in proximal tubules influences glucose tolerance and participates in the kidney injury in an obese pre-diabetic murine model. The role of ADAM17 in the tubule impacts on glomerular inflammation and fibrosis

SARS-CoV-2 infection modulates ACE2 function and subsequent inflammatory responses in swabs and plasma of COVID-19 patients

Angiotensin converting enzyme 2 (ACE2) is a host ectopeptidase and the receptor for the SARS-CoV-2 virus, albeit virus-ACE2 interaction goes far beyond viral entry into target cells. Controversial data exists linking viral infection to changes in ACE2 expression and function, which might influence the subsequent induction of an inflammatory response. Here, we tested the significance of soluble ACE2 enzymatic activity longitudinally in nasopharyngeal swabs and plasma samples of SARS-CoV-2 infected patients, along with the induction of inflammatory cytokines. Release of soluble functional ACE2 increases upon SARS-CoV-2 infection in swabs and plasma of infected patients, albeit rapidly decreasing during infection course in parallel with ACE2 gene expression. Similarly, SARS-CoV-2 infection also induced the expression of inflammatory cytokines. These changes positively correlated with the viral load. Overall, our results demonstrate the existence of mechanisms by which SARS-CoV-2 modulates ACE2 expression and function, intracellular viral sensing and subsequent inflammatory response, offering new insights into ACE2 dynamics in the human upper respiratory tract and pointing towards soluble ACE2 levels as a putative early biomarker of infection severity

The GenoDiabMar registry: A collaborative research platform of type 2 diabetes patients

The GenoDiabMar registry is a prospective study that aims to provide data on demographic, biochemical, and clinical changes in type 2 diabetic (T2D) patients attending real medical outpatient consultations. This registry is also used to find new biomarkers related to the micro- and macrovascular complications of T2D, with a particular focus on diabetic nephropathy. With this purpose, longitudinal serum and urine samples, DNA banking, and data on 227 metabolomics profiles, 77 immunoglobulin G glycomics traits, and other emerging biomarkers were recorded in this cohort. In this study, we show a detailed longitudinal description of the clinical and analytical parameters of this registry, with a special focus on the progress of renal function and cardiovascular events. The main objective is to analyze whether there are differential risk factors for renal function deterioration between sexes, as well as to analyze cardiovascular events and mortality in this population. In total, 650 patients with a median age of 69 (14) with different grades of chronic kidney disease-G1-G2 (eGFR > 90-60 mL/min/1.73 m2) 50.3%, G3 (eGFR; 59-30 mL/min/1.73 m2) 31.4%, G4 (eGFR; 29-15 mL/min/1.73 m2) 10.8%, and G5 (eGFR < 15 mL/min/1.73 m2) 7.5%-were followed up for 4.7 (0.65) years. Regardless of albuminuria, women lost 0.93 (0.40-1.46) fewer glomerular filtration units per year than men. A total of 17% of the participants experienced rapid deterioration of renal function, 75.2% of whom were men, with differential risk factors between sexes-severe macroalbuminuria > 300 mg/g for men OR [IQ] 2.40 [1.29:4.44] and concomitant peripheral vascular disease 3.32 [1.10:9.57] for women. Overall mortality of 23% was detected (38% of which was due to cardiovascular etiology). We showed that kidney function declined faster in men, with different risk factors compared to women. Patients with T2D and kidney involvement have very high mortality and an important cardiovascular burden. This cohort is proposed as a great tool for scientific collaboration for studies, whether they are focused on T2D, or whether they are interested in comparing differential markers between diabetic and non-diabetic populations

Effects of pre-operative isolation on postoperative pulmonary complications after elective surgery: an international prospective cohort study an international prospective cohort study

We aimed to determine the impact of pre-operative isolation on postoperative pulmonary complications after elective surgery during the global SARS-CoV-2 pandemic. We performed an international prospective cohort study including patients undergoing elective surgery in October 2020. Isolation was defined as the period before surgery during which patients did not leave their house or receive visitors from outside their household. The primary outcome was postoperative pulmonary complications, adjusted in multivariable models for measured confounders. Pre-defined sub-group analyses were performed for the primary outcome. A total of 96,454 patients from 114 countries were included and overall, 26,948 (27.9%) patients isolated before surgery. Postoperative pulmonary complications were recorded in 1947 (2.0%) patients of which 227 (11.7%) were associated with SARS-CoV-2 infection. Patients who isolated pre-operatively were older, had more respiratory comorbidities and were more commonly from areas of high SARS-CoV-2 incidence and high-income countries. Although the overall rates of postoperative pulmonary complications were similar in those that isolated and those that did not (2.1% vs 2.0%, respectively), isolation was associated with higher rates of postoperative pulmonary complications after adjustment (adjusted OR 1.20, 95%CI 1.05–1.36, p = 0.005). Sensitivity analyses revealed no further differences when patients were categorised by: pre-operative testing; use of COVID-19-free pathways; or community SARS-CoV-2 prevalence. The rate of postoperative pulmonary complications increased with periods of isolation longer than 3 days, with an OR (95%CI) at 4–7 days or ≥ 8 days of 1.25 (1.04–1.48), p = 0.015 and 1.31 (1.11–1.55), p = 0.001, respectively. Isolation before elective surgery might be associated with a small but clinically important increased risk of postoperative pulmonary complications. Longer periods of isolation showed no reduction in the risk of postoperative pulmonary complications. These findings have significant implications for global provision of elective surgical care. We aimed to determine the impact of pre-operative isolation on postoperative pulmonary complications after elective surgery during the global SARS-CoV-2 pandemic. We performed an international prospective cohort study including patients undergoing elective surgery in October 2020. Isolation was defined as the period before surgery during which patients did not leave their house or receive visitors from outside their household. The primary outcome was postoperative pulmonary complications, adjusted in multivariable models for measured confounders. Pre-defined sub-group analyses were performed for the primary outcome. A total of 96,454 patients from 114 countries were included and overall, 26,948 (27.9%) patients isolated before surgery. Postoperative pulmonary complications were recorded in 1947 (2.0%) patients of which 227 (11.7%) were associated with SARS-CoV-2 infection. Patients who isolated pre-operatively were older, had more respiratory comorbidities and were more commonly from areas of high SARS-CoV-2 incidence and high-income countries. Although the overall rates of postoperative pulmonary complications were similar in those that isolated and those that did not (2.1% vs 2.0%, respectively), isolation was associated with higher rates of postoperative pulmonary complications after adjustment (adjusted OR 1.20, 95%CI 1.05–1.36, p = 0.005). Sensitivity analyses revealed no further differences when patients were categorised by: pre-operative testing; use of COVID-19-free pathways; or community SARS-CoV-2 prevalence. The rate of postoperative pulmonary complications increased with periods of isolation longer than 3 days, with an OR (95%CI) at 4–7 days or ≥ 8 days of 1.25 (1.04–1.48), p = 0.015 and 1.31 (1.11–1.55), p = 0.001, respectively. Isolation before elective surgery might be associated with a small but clinically important increased risk of postoperative pulmonary complications. Longer periods of isolation showed no reduction in the risk of postoperative pulmonary complications. These findings have significant implications for global provision of elective surgical care