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Nephrotoxicity, including electrolytic disorders and acute kidney injury (AKI), limits the clinical dosage and utility of platinated antineoplastics such as cisplatin. Cisplatin nephrotoxicity embodies a tubulopathy involving the medullary S2 and S3 segments of the proximal and the distal tubules. Higher dosage extends damage over the cortical S1 segment and intensifies overall injury. However, the standard diagnosis based on plasma creatinine as well as novel injury biomarkers lacks enough pathophysiological specificity. Further granularity in the detection of renal injury would help understand the implications of individual damage patterns needed for personalized patient handling. In this article, we studied the association of urinary ganglioside GM2 activator protein (GM2AP) with the patterns of tubular damage produced by 5 and 10â¯mg/kg cisplatin in rats. Our results show that GM2AP appears in the urine only following damage to the cortical segment of the proximal tubule. The information provided by GM2AP is not redundant with but distinct and complementary to that provided by urinary neutrophil gelatinase-associated lipocalin (NGAL). Similarly, treatment with 150â¯mg/kg/day gentamicin damages the renal cortex and increases GM2AP urinary excretion; whereas renal ischemia, which does not affect the cortex, has no effect on GM2AP. Because of the key role of the cortical proximal tubule in renal function, we contend GM2AP as a potential diagnostic biomarker to stratify AKI patients according to the underlying damage and follow their evolution and prognosis. Prospectively, urinary GM2AP may help grade the severity of platinated antineoplastic nephrotoxicity by forming part of a non-invasive liquid biopsy.
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Acute kidney frailty is a premorbid condition of diminished renal functional reserve that predisposes to acute kidney injury; this condition results from subclinical wear or distortion of renal homeostatic responses that protect the renal excretory function. Knowledge of its pathophysiological basis is critical for the development of diagnostic and therapeutic strategies that allow for prophylactic intervention and disease prevention.
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Fragilidade , Humanos , Rim , HomeostaseRESUMO
Acute kidney injury (AKI) is a syndrome of sudden renal excretory dysfunction with severe health consequences. AKI etiology influences prognosis, with pre-renal showing a more favorable evolution than intrinsic AKI. Because the international diagnostic criteria (i.e., based on plasma creatinine) provide no etiological distinction, anamnestic and additional biochemical criteria complement AKI diagnosis. Traditional, etiology-defining biochemical parameters, including the fractional excretion of sodium, the urinary-to-plasma creatinine ratio and the renal failure index are individually limited by confounding factors such as diuretics. To minimize distortion, we generated a composite biochemical criterion based on the congruency of at least two of the three biochemical ratios. Patients showing at least two ratios indicative of intrinsic AKI were classified within this category, and those with at least two pre-renal ratios were considered as pre-renal AKI patients. In this study, we demonstrate that the identification of intrinsic AKI by a collection of urinary injury biomarkers reflective of tubular damage, including NGAL and KIM-1, more closely and robustly coincide with the biochemical than with the anamnestic classification. Because there is no gold standard method for the etiological classification of AKI, the mutual reinforcement provided by the biochemical criterion and urinary biomarkers supports an etiological diagnosis based on objective diagnostic parameters.
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Injúria Renal Aguda , Rim , Humanos , Injúria Renal Aguda/diagnóstico , Injúria Renal Aguda/etiologia , Biomarcadores , CreatininaRESUMO
Diagnosis of cardiac surgery-associated acute kidney injury (CSA-AKI), a syndrome of sudden renal dysfunction occurring in the immediate post-operative period, is still sub-optimal. Standard CSA-AKI diagnosis is performed according to the international criteria for AKI diagnosis, afflicted with insufficient sensitivity, specificity, and prognostic capacity. In this article, we describe the limitations of current diagnostic procedures and of the so-called injury biomarkers and analyze new strategies under development for a conceptually enhanced diagnosis of CSA-AKI. Specifically, early pathophysiological diagnosis and patient stratification based on the underlying mechanisms of disease are presented as ongoing developments. This new approach should be underpinned by process-specific biomarkers including, but not limited to, glomerular filtration rate (GFR) to other functions of renal excretion causing GFR-independent hydro-electrolytic and acid-based disorders. In addition, biomarker-based strategies for the assessment of AKI evolution and prognosis are also discussed. Finally, special focus is devoted to the novel concept of pre-emptive diagnosis of acquired risk of AKI, a premorbid condition of renal frailty providing interesting prophylactic opportunities to prevent disease through diagnosis-guided personalized patient handling. Indeed, a new strategy of risk assessment complementing the traditional scores based on the computing of risk factors is advanced. The new strategy pinpoints the assessment of the status of the primary mechanisms of renal function regulation on which the impact of risk factors converges, namely renal hemodynamics and tubular competence, to generate a composite and personalized estimation of individual risk.
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Acute kidney injury (AKI) poses an increased risk factor for new AKI episodes, progression to chronic kidney disease, and death. A worsened evolution has been linked to an incomplete renal repair beyond the apparent functional recovery based on plasma creatinine (pCr) normalization. However, structural sequelae pass largely unnoticed due to the absence of specific diagnostic tools. The urinary kidney injury molecule 1 (KIM-1) participates in renal tissue damage and repair and is proposed as a biomarker of early and subclinical AKI. Thus, we study in this paper the evolution of KIM-1 urinary excretion alongside renal tissue sequelae after an intrinsic AKI episode induced by cisplatin in Wistar rats. Creatinine clearance, pCr, proteinuria and the fractional excretion of Na+ and glucose were used to monitor renal function. Renal tissue damage was blindly scored in kidney specimens stained with hematoxylin-eosin and periodic acid-Schiff. KIM-1 urinary excretion and renal mRNA expression were also assessed. Finally, we analyzed urinary KIM-1 in patients apparently recovered from AKI. Our results show that, after the normalization of the standard markers of glomerular filtration and tubular function, the extent of persistent histological findings of tissue repair correlates with the renal expression and urinary level of KIM-1 in rats. In addition, KIM-1 is also elevated in the urine of a significant fraction of patients apparently recovered from an AKI. Besides its potential utility in the early and subclinical diagnosis of renal damage, this study suggests a new application of urinary KIM-1 in the non-invasive follow-up of renal repair after AKI.
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Nephrotoxicity is a major cause of intrinsic acute kidney injury (AKI). Because renal tissue damage may occur independently of a reduction in glomerular filtration rate and of elevations in plasma creatinine concentration, so-called injury biomarkers have been proposed to form part of diagnostic criteria as reflective of tubular damage independently of renal function status. We studied whether the urinary level of NGAL, KIM-1, GM2AP, t-gelsolin, and REGIIIb informed on the extent of tubular damage in rat models of nephrotoxicity, regardless of the etiology, moment of observation, and underlying pathophysiology. At a time of overt AKI, urinary biomarkers were measured by Western blot or ELISA, and tubular necrosis was scored from histological specimens stained with hematoxylin and eosin. Correlation and regression studies revealed that only weak relations existed between biomarkers and tubular damage. Due to high interindividual variability in the extent of damage for any given biomarker level, urinary injury biomarkers did not necessarily reflect the extent of the underlying tissue injury in individual rats. We contended, in this work, that further pathophysiological contextualization is necessary to understand the diagnostic significance of injury biomarkers before they can be used for renal tubular damage severity stratification in the context of nephrotoxic and, in general, intrinsic AKI.
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Injúria Renal Aguda , Rim , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/diagnóstico , Animais , Biomarcadores , Creatinina , Taxa de Filtração Glomerular , Rim/patologia , Lipocalina-2/urina , RatosRESUMO
Acute kidney injury (AKI) is a risk factor for new AKI episodes, chronic kidney disease, cardiovascular events and death, as renal repair may be deficient and maladaptive, and activate proinflammatory and profibrotic signals. AKI and AKI recovery definitions are based on changes in plasma creatinine, a parameter mostly associated to glomerular filtration, but largely uncoupled from renal tissue damage. The evolution of structural and functional repair has been incompletely described. We thus aimed at identifying subclinical sequelae persisting after recovery from cisplatin-induced AKI in rats. Compared to controls, after plasma creatinine recovery, post-AKI kidneys showed histological alterations and attendant susceptibility to new AKI episodes. Tubular function (assessed by the furosemide stress test, FST) also remained affected. Lingering parenchymal and functional subclinical alterations were paralleled by tapering, but abnormally high levels of urinary albumin, transferrin, insulin-like growth factor-binding protein 7 (IGFBP7), tissue inhibitor of metalloproteinases-2 (TIMP-2) and, especially, the [TIMP-2]*[IGFBP7] product. As subclinical surrogates of incomplete renal recovery, the FST and the urinary [TIMP-2]*[IGFBP7] product provide two potential diagnostic tools to monitor the sequelae and kidney vulnerability after the apparent recovery from AKI.
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Injúria Renal Aguda/urina , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/urina , Inibidor Tecidual de Metaloproteinase-2/urina , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/patologia , Animais , Antineoplásicos/toxicidade , Biomarcadores/urina , Cisplatino/toxicidade , Rim/efeitos dos fármacos , Rim/patologia , Rim/fisiopatologia , Masculino , Ratos , Ratos WistarRESUMO
INTRODUCTION: Acute kidney injury (AKI) is a threatening, multiaetiological syndrome encompassing a variety of forms and damage patterns. AKI lacks sufficiently specific diagnostic tools to evaluate the distinct combination of pathophysiological events underlying each case, which limits personalized and optimized handling. Therefore, a pathophysiological diagnosis based on new urinary biomarkers is sought for practical (readiness and noninvasiveness) and conceptual reasons, as the urine is a direct product of the kidneys. However, biomarkers found in the urine may also have extrarenal origin, thus conveying pathophysiological information from other organs or tissues. Urinary plasminogen activator inhibitor-1 (PAI-1) has been associated to AKI, although its origin and traffic to the urine are not known. METHODS: Herein, we studied the blood or renal origin of urinary PAI-1 (uPAI-1) in experimental AKI in Wistar rats, by means of the in situ renal perfusion method. For this purpose, urine was collected while the kidneys of rats with AKI showing increased uPAI-1 excretion, and controls, were in situ perfused with a saline solution. RESULTS: Our results show that during perfusion, PAI-1 remained in the urine of AKI rats, suggesting that renal cells shed this protein directly to the urine. PAI-1 is also significantly increased in the urine of AKI patients. Its low correlation with other urinary markers such as NGAL or NAG suggests that PAI-1 provides complementary and distinct phenotypical information. CONCLUSION: In conclusion, uPAI-1 is a biomarker produced by damaged kidneys following AKI, whose precise pathophysiological meaning in AKI needs to be further investigated.
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Injúria Renal Aguda/urina , Túbulos Renais , Inibidor 1 de Ativador de Plasminogênio/urina , Adulto , Idoso , Animais , Biomarcadores/urina , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ratos , Ratos WistarRESUMO
Deficient recovery from acute kidney injury (AKI) has immediate and long-term health, clinical and economic consequences. Pre-emptive recovery estimation may improve nephrology referral, optimize decision making, enrollment in trials, and provide key information for subsequent clinical handling and follow-up. For this purpose, new biomarkers are needed that predict outcome during the AKI episode. We hypothesized that damage pattern-specific biomarkers are expected to more closely associate to outcome within distinct subpopulations (i.e. those affected by specific pathological processes determining a specific outcome), as biomarker pleiotropy (i.e. associated to phenomena unrelated to AKI) introduced by unselected, heterogeneous populations may blur statistics. A panel of urinary biomarkers was measured in patients with AKI and their capacity to associate to normal or abnormal recovery was studied in the whole cohort or after sub-classification by AKI etiology, namely pre-renal and intrinsic AKI. A combination of urinary GM2AP and TCP1-eta best associates with recovery from AKI, specifically within the sub-population of renal AKI patients. This two-step strategy generates a multidimensional space in which patients with specific characteristics (i.e. renal AKI patients with good or bad prognosis) can be identified based on a collection of biomarkers working serially, applying pathophysiology-driven criteria to estimate AKI recovery, to facilitate pre-emptive and personalized handling.
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Injúria Renal Aguda/urina , Biomarcadores/urina , Chaperonina com TCP-1/urina , Proteína Ativadora de G(M2)/urina , Injúria Renal Aguda/genética , Injúria Renal Aguda/patologia , Linhagem da Célula/genética , Feminino , Humanos , Rim/patologia , Masculino , Pessoa de Meia-IdadeRESUMO
Simultaneous administration of certain antihypertensive (renin-angiotensin system inhibitors and diuretics) and nonsteroidal anti-inflammatory drugs (NSAIDs) is associated with a renal toxicity syndrome known as "triple whammy" acute kidney injury (TW-AKI), yet poorly characterized at the pathophysiological level, as no specific experimental model exists on which to conduct preclinical research. Herein, we generated and characterized a rat model of TW-AKI (0.7 mg/kg/day trandolapril +400 mg/kg/day ibuprofen +20 mg/kg/day furosemide). Double treatments involving the NSAID caused a subclinical acute kidney injury, as they reduced glomerular filtration rate to a significant but not sufficient extent to increase Crpl concentration. Only the triple treatment generated an overt AKI with increased Crpl provided that animals were under partial water ingestion restriction. Histological examination revealed no evidence of tissue renal injury, and no proteinuria or makers of renal damage were detected in the urine. These findings, along with a normal fractional excretion of sodium and glucose, indicated that these drug combinations produce a prerenal type of AKI. In fact, blood pressure and renal blood flow were also reduced (most markedly following the triple combination), although renal dysfunction was more pronounced than expected for the corresponding pressure drop, supporting a key pathological role of the interference with renal autoregulation mechanisms. In summary, prerenal TW-AKI only occurs when volemia is challenged (i.e., by furosemide in partially water-deprived animals) under the effects of renin-angiotensin system inhibitors and NSAIDs. This model will facilitate further pathophysiological knowledge for a better diagnosis and clinical handling of this syndrome.
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Injúria Renal Aguda/induzido quimicamente , Inibidores da Enzima Conversora de Angiotensina/efeitos adversos , Anti-Inflamatórios não Esteroides/efeitos adversos , Modelos Animais de Doenças , Diuréticos/efeitos adversos , Animais , Pressão Sanguínea/efeitos dos fármacos , Quimioterapia Combinada/efeitos adversos , Furosemida/efeitos adversos , Ibuprofeno/efeitos adversos , Indóis/efeitos adversos , Masculino , Ratos Wistar , Circulação Renal/efeitos dos fármacosRESUMO
Neutrophil gelatinase-associated lipocalin (NGAL) is a secreted low-molecular weight iron-siderophore-binding protein. NGAL overexpression in injured tubular epithelia partly explains its utility as a sensitive and early urinary biomarker of acute kidney injury (AKI). Herein, we extend mechanistic insights into the source and kinetics of urinary NGAL excretion in experimental AKI. Three models of experimental AKI were undertaken in adult male Wistar rats; renal ischemia-reperfusion injury (IRI) and gentamicin (G) and cisplatin (Cisp) nephrotoxicity. Alongside standard histological and biochemical assessment of AKI, urinary NGAL excretion rate, plasma NGAL concentration, and renal NGAL mRNA/protein expression were assessed. In situ renal perfusion studies were undertaken to discriminate direct shedding of NGAL to the urine from addition of NGAL to the urine secondary to alterations in the tubular handling of glomerular filtrate-derived protein. Renal NGAL expression and urinary excretion increased in experimental AKI. In acute studies in both the IRI and G models, direct renal perfusion with Kreb's buffer eliminated urinary NGAL excretion. Addition of exogenous NGAL to the Kreb's buffer circuit, reestablishment of perfusion with systemic blood or reperfusion with renal vein effluent restored high levels of urinary NGAL excretion. Urinary NGAL excretion in AKI arises in large proportion from reduced reabsorption from the glomerular filtrate. Hence, subclinical cellular dysfunction could increase urinary NGAL, particularly in concert with elevations in circulating prerenal NGAL and/or pharmacological inhibition of tubular reabsorption. More granular interpretation of urinary NGAL measurements could optimize the scope of its clinical utility as a biomarker of AKI.
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Injúria Renal Aguda/urina , Túbulos Renais/metabolismo , Lipocalina-2/urina , Reabsorção Renal , Traumatismo por Reperfusão/urina , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/genética , Injúria Renal Aguda/fisiopatologia , Animais , Biomarcadores/urina , Cisplatino , Modelos Animais de Doenças , Gentamicinas , Túbulos Renais/fisiopatologia , Lipocalina-2/genética , Masculino , Ratos Wistar , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/fisiopatologia , Fatores de Tempo , Regulação para CimaRESUMO
Acute kidney injury (AKI) is a serious syndrome with increasing incidence and health consequences, and high mortality rate among critically ill patients. Acute kidney injury lacks a unified definition, has ambiguous semantic boundaries, and relies on defective diagnosis. This, in part, is due to the absence of biomarkers substratifying AKI patients into pathophysiological categories based on which prognosis can be assigned and clinical treatment differentiated. For instance, AKI involving acute tubular necrosis (ATN) is expected to have a worse prognosis than prerenal, purely hemodynamic AKI. However, no biomarker has been unambiguously associated with tubular cell death or is able to provide etiological distinction. We used a cell-based system to identify TCP1-eta in the culture medium as a noninvasive marker of damaged renal tubular cells. In rat models of AKI, TCP1-eta was increased in the urine co-relating with renal cortical tubule damage. When kidneys from ATN rats were perfused in situ with Krebs-dextran solution, a portion of the urinary TCP1-eta protein content excreted into urine disappeared, and another portion remained within the urine. These results indicated that TCP1-eta was secreted by tubule cells and was not fully reabsorbed by the damaged tubules, both effects contributing to the increased urinary excretion. Urinary TCP1-eta is found in many etiologically heterogeneous AKI patients, and is statistically higher in patients partially recovered from severe AKI. In conclusion, urinary TCP1-eta poses a potential, substratifying biomarker of renal cortical damage associated with bad prognosis.
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Injúria Renal Aguda/urina , Chaperonina com TCP-1/urina , Túbulos Renais/metabolismo , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/patologia , Injúria Renal Aguda/fisiopatologia , Animais , Apoptose , Biomarcadores/urina , Estudos de Casos e Controles , Linhagem Celular , Modelos Animais de Doenças , Diagnóstico Precoce , Túbulos Renais/patologia , Túbulos Renais/fisiopatologia , Masculino , Valor Preditivo dos Testes , Prognóstico , Ratos Wistar , Eliminação Renal , UrináliseRESUMO
Nephrotoxicity is the main limitation to the dosage and anticancer efficacy of cisplatin. Cisplatin produces tubular epithelial cell apoptosis and necrosis depending on the concentration of the drug. Protection from cisplatin nephrotoxicity must therefore tackle both cell death modes. For its ability to reduce cisplatin reactivity, in addition to its antioxidant effect, we tested and found that N-acetylcysteine (NAC) was most effective at inhibiting cisplatin cytotoxicity. NAC has no significant effect on cell death induced by either cycloheximide or Fas activation, indicating a rather selective action. Pt-DNA-binding experiments suggest that the differential effectiveness of NAC is due to its capacity to quench cisplatin reactivity inside the cell. NAC abolishes cisplatin-induced apoptosis, and transforms the necrosis induced by high concentrations of cisplatin into apoptosis. In fact, NAC allows the anti-apoptotic molecule Bcl-2 to reduce the cell death caused by pro-necrotic concentrations of cisplatin, to a significantly greater extent than in the absence of NAC. In rats, a dosage of NAC that significantly ameliorates cisplatin nephrotoxicity, has little effect on gentamicin nephrotoxicity. These characteristics provide NAC with a rationale as a potential nephroprotectant specifically tailored to and especially effective for therapeutic courses with platinated antineoplastics, which prompts to deepening into further preclinical knowledge, and to initiate clinical studies with NAC and mixed therapies composed of NAC and antiapoptotic drugs.
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Acetilcisteína/farmacologia , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Cisplatino/toxicidade , Sequestradores de Radicais Livres/farmacologia , Necrose/induzido quimicamente , Animais , Caspases/análise , Caspases/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Células Jurkat , Nefropatias/induzido quimicamente , Nefropatias/patologia , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Ratos , Ratos WistarRESUMO
Pre-renal acute kidney injury (AKI) results from glomerular haemodynamic alterations leading to reduced glomerular filtration rate (GFR) with no parenchymal compromise. Renin-angiotensin system inhibitors, such as angiotensin-converting enzyme inhibitors (ACEIs), angiotensin receptor antagonists (ARAs), non-steroidal anti-inflammatory drugs (NSAIDs) and diuretics, are highly prescribed drugs that are frequently administered together. Double and triple associations have been correlated with increased pre-renal AKI incidence, termed "double whammy" and "triple whammy", respectively. This article presents an integrative analysis of the complex interplay among the effects of NSAIDs, ACEIs/ARAs and diuretics, acting alone and together in double and triple therapies. In addition, we explore how these drug combinations alter the equilibrium of regulatory mechanisms controlling blood pressure (renal perfusion pressure) and GFR to increase the odds of inducing AKI through the concomitant reduction of blood pressure and distortion of renal autoregulation. Using this knowledge, we propose a more general model of pre-renal AKI based on a multi whammy model, whereby several factors are necessary to effectively reduce net filtration. The triple whammy was the only model associated with pre-renal AKI accompanied by a course of other risk factors, among numerous potential combinations of clinical circumstances causing hypoperfusion in which renal autoregulation is not operative or is deregulated. These factors would uncouple the normal BP-GFR relationship, where lower GFR values are obtained at every BP value.
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Injúria Renal Aguda/etiologia , Pressão Sanguínea/fisiologia , Modelos Teóricos , Injúria Renal Aguda/epidemiologia , Injúria Renal Aguda/fisiopatologia , Antagonistas de Receptores de Angiotensina/administração & dosagem , Antagonistas de Receptores de Angiotensina/efeitos adversos , Inibidores da Enzima Conversora de Angiotensina/administração & dosagem , Inibidores da Enzima Conversora de Angiotensina/efeitos adversos , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/efeitos adversos , Pressão Sanguínea/efeitos dos fármacos , Diuréticos/administração & dosagem , Diuréticos/efeitos adversos , Taxa de Filtração Glomerular , Humanos , Incidência , Sistema Renina-Angiotensina/efeitos dos fármacos , Fatores de RiscoRESUMO
Early detection of hypertensive end-organ damage and secondary diseases are key determinants of cardiovascular prognosis in patients suffering from arterial hypertension. Presently, there are no biomarkers for the detection of hypertensive target organ damage, most outstandingly including blood vessels, the heart, and the kidneys.We aimed to validate the usefulness of the urinary excretion of the serine protease kallikrein-related peptidase 9 (KLK9) as a biomarker of hypertension-induced target organ damage.Urinary, plasma, and renal tissue levels of KLK9 were measured by the Western blot in different rat models of hypertension, including angiotensin-II infusion, DOCA-salt, L-NAME administration, and spontaneous hypertension. Urinary levels were associated to cardiovascular and renal injury, assessed by histopathology. The origin of urinary KLK9 was investigated through in situ renal perfusion experiments.The urinary excretion of KLK9 is increased in different experimental models of hypertension in rats. The ACE inhibitor trandolapril significantly reduced arterial pressure and the urinary level of KLK9. Hypertension did not increase kidney, heart, liver, lung, or plasma KLK9 levels. Hypertension-induced increased urinary excretion of KLK9 results from specific alterations in its tubular reabsorption, even in the absence of overt nephropathy. KLK9 urinary excretion strongly correlates with cardiac hypertrophy and aortic wall thickening.KLK9 appears in the urine in the presence of hypertension as a result of subtle renal handling alterations. Urinary KLK9 might be potentially used as an indicator of hypertensive cardiac and vascular damage.
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Hipertensão/metabolismo , Calicreínas/sangue , Calicreínas/urina , Nefropatias/sangue , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Animais , Pressão Arterial , Biomarcadores , Pressão Sanguínea , Doenças Cardiovasculares/sangue , Modelos Animais de Doenças , Expressão Gênica , Indóis/farmacologia , Rim/metabolismo , Masculino , Reação em Cadeia da Polimerase , Ratos , Ratos WistarRESUMO
BACKGROUND/AIMS: Defective tissue repair underlies renal tissue degeneration during chronic kidney disease (CKD) progression. Unbalanced presence of TGF-ß opposes effective cell proliferation and differentiation processes, necessary to replace damaged epithelia. TGF-ß also retains arrested cells in a fibrotic phenotype responsible for irreversible scarring. In order to identify prospective molecular targets to prevent the effect of TGF-ß during CKD, we studied the signaling pathways responsible for the antiproliferative effect of this cytokine. METHODS: Tubule epithelial HK2 and MDCK cells were treated with TGF-ß (or not as control) to study cell proliferation (by MTT), cell signaling (by Western blot), cell cycle (by flow cytometry) and apoptosis (DNA fragmentation). RESULTS: TGF-ß fully activates the ALK-5 receptor pathway, whereas it has no effect on the ALK-1 and MAPK pathways in both HK2 and MDCK cells. Interestingly, TGF-ß exerts an antiproliferative effect only on MDCK cells, through a cytostatic effect in G0/G1. Inhibition of the ALK-5 pathway with SB431542 prevents the cytostatic effect of TGF-ß on MDCK cells. CONCLUSION: Activation of the ALK-5 pathway is not sufficient for the antiproliferative effect of TGF-ß. The presence of undetermined permissive conditions or absence of undetermined inhibitory conditions seems to be necessary for this effect. The ALK-5 pathway appears to provide targets to modulate fibrosis, but further research is necessary to identify critical circumstances allowing or inhibiting its role at modulating tubule epithelial cell proliferation and tubule regeneration in the context of CKD progression.
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Proliferação de Células/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Animais , Benzamidas/farmacologia , Linhagem Celular , Fragmentação do DNA/efeitos dos fármacos , Dioxóis/farmacologia , Cães , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Túbulos Renais/citologia , Células Madin Darby de Rim Canino , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Proteína Smad2/metabolismoRESUMO
The histological substrate of many forms of intrinsic acute kidney injury (AKI) has been classically attributed to tubular necrosis. However, more recent studies indicate that necrosis is not the main form of cell death in AKI and that other forms such as apoptosis, regulated necrosis (i.e. necroptosis and parthanatos), autophagic cell death and mitotic catastrophe, also participate in AKI and that their contribution depends on the cause and stage of AKI. Herein, we briefly summarize the main characteristics of the major types of cell death and we also critically review the existing evidence on the occurrence of different types of cell death reported in the most common experimental models of AKI and human specimens. We also discuss the pathophysiological mechanisms linking tubule epithelial cell death with reduced glomerular filtration, azotaemia and hydroelectrolytic imbalance. For instance, special relevance is given to the analysis of the inflammatory component of some forms of cell death over that of others, as an important and differential pathophysiological determinant. Finally, known molecular mechanisms and signalling pathways involved in each cell death type pose appropriate targets to specifically prevent or reverse AKI, provided that further knowledge of their participation and repercussion in each AKI syndrome is progressively increased in the near future.
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Urinary differential proteomics is used to study renal pathophysiological mechanisms, find novel markers of biological processes and renal diseases, and stratify patients according to proteomic profiles. The proteomic procedure determines the pathophysiological meaning and clinical relevance of results. Urine samples for differential proteomic studies are usually normalized by protein content, regardless of its pathophysiological characteristics. In the field of nephrology, this approach translates into the comparison of a different fraction of the total daily urine output between proteinuric and nonproteinuric samples. Accordingly, alterations in the level of specific proteins found by this method reflect the relative presence of individual proteins in the urine; but they do not necessarily show alterations in their daily excretion, which is a key parameter for the understanding of the pathophysiological meaning of urinary components. For renal pathophysiology studies and clinical biomarker identification or determination, an alternative proteomic concept providing complementary information is based on sample normalization by daily urine output, which directly informs on changes in the daily excretion of individual proteins. This is clinically important because daily excretion (rather than absolute or relative concentration) is the only self-normalized way to evaluate the real meaning of urinary parameters, which is also independent of urine concentration.
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Nefropatias/urina , Proteinúria/urina , Animais , Humanos , Rim/fisiopatologia , Proteoma/metabolismo , ProteômicaRESUMO
Cisplatin is a chemotherapeutic drug widely used against a variety of cancers. Its clinical utility is severely limited by its toxicity, which mainly affects, but is not limited to, the inner ear and renal tubules. Cisplatin toxicity is determined by target tissue and cell accumulation, subcellular handling and trafficking through diverse subcellular structures, and interaction with macromolecules. Cisplatin accumulates and stresses different organelles from which delay signaling is activated, including mitochondria, lysosomes, the endoplasmic reticulum, the nucleus, the cell membrane and cytoskeleton, and can also be found in the cytosol. This article critically summarizes the available information in order to establish the connection among its known subcellular effects in a hierarchical and integrative framework. Cisplatin causes different types of cell death in a concentration-dependent manner. Knowledge of the events and signaling leading to the different phenotypes is also intertwined within the model, within the scope of the potential utility of this information in the improvement of the pharmacotoxicological profile of this drug. Perspectives for the key aspects that need to be addressed by future investigation are also outlined.
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Antineoplásicos/toxicidade , Cisplatino/toxicidade , Animais , Transporte Biológico , Membrana Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Cisplatino/farmacocinética , Citoesqueleto/efeitos dos fármacos , Citosol/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Humanos , Lisossomos/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacosRESUMO
Cisplatin is a chemotherapeutic drug whose cytotoxicity is key to its therapeutic and side effects. Nephrotoxicity, mainly due to renal tubular injury, poses its most important therapeutic limitation. Tubular necrosis is derived from epithelial cell death by apoptosis and necrosis in the proximal and distal tubuli. The mode of cell death has been related to drug concentration, with necrosis occurring with high concentrations and apoptosis with lower concentrations. To fully understand the toxic effects of cisplatin to potentially improve its pharmaco-toxicological profile, it is necessary to unravel the cellular events and signaling pathways implicated in the appearance of both modes of cell death. We used cultured human lymphoma and renal tubule cells to investigate the biochemical and phenotypic characteristics of the death mode induced by increasing concentrations of cisplatin. Our results indicate that pronecrotic concentrations of cisplatin early activate the apoptotic machinery, which is in turn directly blocked by cisplatin at the level of effector caspases. Aborted apoptosis induces a death phenotype lacking some typical characteristics of this process, which more closely resembles necrosis. Furthermore, unidentified Bcl-2- and mitochondria-independent pathways are induced by pronecrotic and not by proapoptotic concentrations of cisplatin. Cisplatin-induced cell necrosis is the result of an aborted apoptosis at the level of effector caspases. Yet, Bcl-2-independent effects lead to cell death, which may pose potential targets for pharmacological intervention aimed at reducing cisplatin nephrotoxicity.