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Volume overload has been shown to be an independent risk factor for mortality in patients receiving chronic dialysis, but data in non-dialysis patients are scarce. Therefore we evaluated the prognostic value of extracellular fluid (ECF) volume for chronic kidney disease (CKD) progression and mortality in a prospective hospital-based cohort with CKD stage 1-4 (NephroTest Study). ECF (scaled to body surface area) and the measured glomerular filtration rate (mGFR) were determined using the distribution volume and clearance of 51Cr-EDTA, respectively. Cause-specific Cox and linear mixed-effect regression models were used to analyze the association of ECF with end-stage kidney disease (ESKD) and mortality, and with mGFR decline, respectively. The 1593 patients were mean age 58.8 years, 67% were men, mean mGFR of 43.6 mL/min/1.73m2 and mean ECF 15.1 L/1.73m2. After a median follow-up of 5.3 years, ESKD occurred in 324 patients and 185 patients died before ESKD. In multivariable analysis, ECF was significantly associated with the risk of ESKD (hazard ratio per 1L/1.73m2 increase: 1.14; 95% confidence interval [1.07; 1.21]) and with a faster GFR decline (adjusted mean difference in mGFR slope per 1L/1.73m2 increase -0.14 [-0.23; -0.05] mL/min/year). The relationship of ECF with mortality was non-linear and not significant (per 1L/1.73m2 increase 0.92, [0.73; 1.16]), below 15L/1.73m2, but significant (1.28; [1.14-1.45]) above 15L/1.73m2. Thus, in this large cohort of carefully phenotyped patients with CKD, ECF was an independent risk factor of CKD progression and mortality. Hence, close monitoring and treatment of fluid overload are important for the clinical management of patients with non-dialysis CKD.

International audience; Increased urinary oxalate excretion (hyperoxaluria) promotes the formation of calcium oxalate crystals. Monogenic diseases due to hepatic enzyme deficiency result in chronic hyperoxaluria, promoting end-stage renal disease in children and young adults. Ethylene glycol poisoning also results in hyperoxaluria, promoting acute renal failure and frequently death. Stiripentol is an antiepileptic drug used to treat children affected by Dravet syndrome. It has been shown to inhibit neuronal lactate dehydrogenase 5 enzyme. As this isoenzyme is also the last step of hepatic oxalate production, we hypothesized that stiripentol would potentially reduce hepatic oxalate production and urine oxalate excretion. In vitro, stiripentol decreased the synthesis of oxalate by hepatocytes in a dose-dependent manner. In vivo, oral administration of stiripentol significantly reduced urine oxalate excretion in rats. Stiripentol protected the kidneys against calcium oxalate crystal deposits in acute ethylene glycol intoxication and chronic calcium oxalate nephropathy models. In both models, stiripentol significantly improved renal function. Patients affected by Dravet syndrome and treated with stiripentol had a lower urine oxalate excretion than control patients. A young girl affected by severe type I hyperoxaluria received stiripentol for several weeks, and urine oxalate excretion decreased by two-thirds. Stiripentol is a promising potential therapy against genetic hyperoxaluria and ethylene glycol poisoning.

Currently an evidence-based approach to nephrolithiasis is hampered by a lack of randomized controlled trials. Thus, there is a need for common platforms for data sharing and recruitment of patients to interventional studies. A first step in achieving this objective would be to share practice methods and protocols for subsequent standardization in what is still a heterogeneous clinical field. Here, we present the results of a pilot survey performed across 24 European clinical kidney stone centers. The survey was distributed by a voluntary online questionnaire circulated between June 2017 and January 2018. About 46% of centers reported seeing on average 20 or more patients per month. Only 21% adopted any formal referral criteria. Centers were relatively heterogeneous in respect of the definition of an incident stone event. The majority (71%) adopted a formal follow-up scheme; of these, 65% included a follow-up visit at 3 and 12 months, and 41% more than 12 months. In 79% of centers some kind of imaging was performed systematically. 75% of all centers performed laboratory analyses on blood samples at baseline and during follow-up. All centers performed laboratory analyses on 24-h urine samples, the majority (96%) at baseline and during follow-up. There was good correspondence across centers for analyses performed on 24-h urine samples, although the methods of 24-h urine collection and analysis were relatively heterogeneous. Our survey among 24 European stone centers highlights areas of homogeneity and heterogeneity that will be investigated further. Our aim is the creation of a European network of stone centers sharing practice patterns and hosting a common database for research and guidance in clinical care.

A symposium on kidney stones and mineral metabolism held on December 2017 in Brussels, Belgium was the first international multidisciplinary conference of the International Collaborative Network on Kidney Stones and Mineral Metabolism. This meeting addressed epidemiology, underlying pathophysiological mechanisms, genetics, pathological, as well as clinical and research topics. The participants included clinicians and recognized experts in the field from Europe and the United States interacted closely during the symposium which promoted a chance to explore new frontiers in the field of kidney stone disease. This manuscript summarizes some of the major highlights of the meeting., info:eu-repo/semantics/published

Green tea is widely used as a &lsquo, &rsquo, healthy&rsquo, beverage due to its high level of antioxidant polyphenol compounds. However tea is also known to contain significant amount of oxalate. The objective was to determine, in a cross-sectional observational study among a population of 273 hypercalciuric stone-formers referred to our center for metabolic evaluation, whether daily green tea drinkers (n = 41) experienced increased stone risk factors (especially for oxalate) compared to non-drinkers. Stone risk factors and stone composition were analyzed according to green tea status and sex. In 24-h urine collection, the comparison between green tea drinkers and non-drinkers showed no difference for stone risk factors such as urine oxalate, calcium, urate, citrate, and pH. In females, the prevalence of calcium oxalate dihydrate (COD) and calcium phosphate stones, assessed by infrared analysis (IRS) was similar between green tea drinkers and non-drinkers, whereas prevalence of calcium oxalate monohydrate (COM) stones was strikingly decreased in green tea drinkers (0% vs. 42%, p = 0.04), with data in accordance with a decreased oxalate supersaturation index. In males, stone composition and supersaturation indexes were similar between the two groups. Our data show no evidence for increased stone risk factors or oxalate-dependent stones in daily green tea drinkers.

International audience; BACKGROUND: Sarcoidosis, characterized by epithelioid granulomas, is considered to be caused by a complex interplay between genetics and environmental agents. It has been hypothesized that exogenous inorganic particles as crystalline silica could be a causal or adjuvant agent in sarcoidosis onset.OBJECTIVES: To investigate the location, frequency and physicochemical characteristics of foreign materials and mineral tissue deposits in the granulomatous area of cutaneous sarcoidosis.METHODS: Skin biopsies (n = 14) from patients diagnosed with cutaneous sarcoidosis (mean age 43 years; 11 patients with extracutaneous involvement) were investigated using polarized light examination (PLE), μFourier Transform Infra-Red (μFT-IR) spectroscopy and Field Emission Scanning Electron Microscopy coupled with Energy Dispersive X-ray Spectroscopy (FE-SEM/EDX).RESULTS: Combined PLE, μFT-IR, FE-SEM/EDX analysis allowed to characterize mineral deposits in 7/14 biopsies (50%). It identified crystalline silica (SiO2 ) inside granulomas in three biopsies and calcite (CaCO3 ) at their periphery in 4.CONCLUSION: This study emphasizes the need of using combined methods for assessment of mineral deposits in granulomatous diseases. According to the location and characteristics of deposits, we can hypothesize that SiO2 particles contribute to the granuloma formation, whereas CaCO3 deposits are related to the granuloma biology. However, the significance of the association between SiO2 deposits and sarcoidosis is still disputed.

International audience; Importance: Calcific uremic arteriolopathy (CUA), a rare, potentially fatal, disease with calcium deposits in skin, mostly affects patients with end-stage renal disease who are receiving dialysis. Chemical composition and structure of CUA calcifications have been poorly described.Objectives: To describe the localization and morphologic features and determine the precise chemical composition of CUA-related calcium deposits in skin, and identify any mortality-associated factors.Design, Setting, and Participants: A retrospective, multicenter cohort study was conducted at 7 French hospitals including consecutive adults diagnosed with CUA between January 1, 2006, and January 1, 2017, confirmed according to Hayashi clinical and histologic criteria. Patients with normal renal function were excluded. For comparison, 5 skin samples from patients with arteriolosclerosis and 5 others from the negative margins of skin-carcinoma resection specimens were also analyzed.Main Outcomes and Measures: Localization and morphologic features of the CUA-related cutaneous calcium deposits were assessed with optical microscopy and field-emission-scanning electron microscopy, and the chemical compositions of those deposits were evaluated with μ Fourier transform infrared spectroscopy, Raman spectroscopy, and energy dispersive radiographs.Results: Thirty-six patients (median [range] age, 64 [33-89] years; 26 [72%] female) were included, and 29 cutaneous biopsies were analyzed. Calcific uremic arteriolopathy and arteriolosclerosis skin calcifications were composed of pure calcium-phosphate apatite. Calcific uremic arteriolopathy vascular calcifications were always circumferential, found in small to medium-sized vessels, with interstitial deposits in 22 (76%) of the samples. A thrombosis, most often in noncalcified capillary lumens in the superficial dermis, was seen in 5 samples from patients with CUA. Except for calcium deposits, the vessel structure of patients with CUA appeared normal, unlike thickened arteriolosclerotic vessel walls. Twelve (33%) patients died of CUA.Conclusions and Relevance: Calcific uremic arteriolopathy-related skin calcifications were exclusively composed of pure calcium-phosphate apatite, localized circumferentially in small to medium-sized vessels and often associated with interstitial deposits, suggesting its pathogenesis differs from that of arteriolosclerosis. Although the chemical compositions of CUA and arteriolosclerosis calcifications were similar, the vessels' appearances and deposit localizations differed, suggesting different pathogenetic mechanisms.

International audience; Calpains, intracellular proteases specifically inhibited by calpastatin, play a major role in neoangiogenesis involved in tumor invasiveness and metastasis. They are partly exteriorized via the ATP-binding cassette transporter A1(ABCA1) transporter, but the importance of this process in tumor growth is still unknown. The aim of our study was to investigate the role of extracellular calpains in a model of melanoma by blocking their extracellular activity or exteriorization. In the first approach, a B16-F10 model of melanoma was developed in transgenic mice expressing high extracellular levels of calpastatin. In these mice, tumor growth was inhibited by ∼ 3-fold compared with wild-type animals. In vitro cytotoxicity assays and in vivo tumor studies have demonstrated that this protection was associated with a defect in tumor neoangiogenesis. Similarly, in wild-type animals given probenecid to blunt ABCA1 activity, melanoma tumor growth was inhibited by ∼ 3-fold. Again, this response was associated with a defect in neoangiogenesis. In vitro studies confirmed that probenecid limited endothelial cell migration and capillary formation from vascular explants. The observed reduction in fibronectin cleavage under these conditions is potentially involved in the response. Collectively, these studies demonstrate that probenecid, by blunting ABCA1 activity and thereby calpain exteriorization, limits melanoma tumor neoangiogenesis and invasiveness.

International audience; Alexander Randall described eight decades ago a heterogeneous nucleation process at the tip of renal papillae giving birth to calcium oxalate stones. Kidney stones were for the first time described to originate from calcium phosphate plaques growing in the interstitial tissue, breaking urothelium and then promoting calcium oxalate crystals aggregation at their contact. During the next decades, few studies were dedicated to these Randall's plaques but the increasing incidence of calcium oxalate kidney stones, the development of endoscopic procedures allowing plaques visualization and series evidencing the high proportion of calcium oxalate stones generated on these plaques renewed interest in Randall's plaque. Although some progress has been carried during the past two decades, the origin of Randall' plaques, their composition, their role in kidney stones epidemic, their specific affinity for some crystalline phases, and their potential deleterious effects in kidney tissues are among the challenges that lie ahead.

Nephrotoxicity is a specific and serious adverse effect of cisplatin which limits its utilization. Here we present an experimental approach to decipher its mechanisms focusing on the renal distribution of Pt and Zn in cisplatin exposed mice kidneys. We combined Synchrotron Radiation (S.R.) μX-ray fluorescence with statistical treatments. μX-ray fluorescence data were collected on a set of mice biopsies after cisplatin, oxaliplatin and carboplatin injections. Even if this investigation is based on a limited number of samples, these preliminary data seem to reveal a stronger correlation between the spatial repartition of Pt and Zn for cisplatin than for the other Pt containing drugs. The cisplatin injection induced a redistribution of medullary Zn across the corticomedullary junction where histological lesions develop. These results were confirmed by evaluation of Pearson's and Manders' co-localization coefficients. These data suggest that Zn is involved in the nephrotoxicity of cisplatin. This could lead to new diagnosis, physiopathological and even therapeutical approaches.

Tubulo-interstitial microcalcifications in renal transplant are described with a wide difference of incidence (4–78%) according to time and goal of biopsies. Currently, staining procedures are used to deduce the composition of crystals and speculate about their aetiologies. Here we test the contribution of infrared microspectroscopy (IR-MS) in understanding kidney transplant crystal deposits. First, microcalcifications observed in 118 allograft biopsies are studied by IR-MS. The Fourier transform infrared signal shows that a major proportion (92%) of calcium phosphate crystals is in the pure or mixed form. Next, we compare 50 patients with calcifications to 100 without calcifications and show persistent hyperparathyroidism and tubular cell vacuolization as circumstances of crystal deposition. Finally, the graduation level of calcification by IR-MS appears to be correlated with the graft outcome. Graft survival seems to be worse in case of high microcalcification detection by IR-MS. These preliminary data suggest IR-MS as a great tool for clinicians to diagnose, characterize, and quantify microcalcifications in kidney allografts.

International audience; Background: Recent epidemiologic investigations have identified an association between type 2 diabetes and uric acid kidney stones. This association was more apparent in women than in men. However, male patients are more prone than women to form uric acid stones in upper and lower urinary tract. In addition, uric acid stone morphology may be different according to stone location. Finally, it was shown that uric acid stone prevalence is increasing with the patient’s age. Aim of the study: To compare uric acid crystal size as determined by powder neutron diffraction with clinical data and the gender of patients. Material and methods: Uric acid stones from 43 patients (24 males, 19 females) identified by Fourier transform infrared spectroscopy were investigated using Environmental Scanning Electron Microscopy and Powder neutron diffraction.Results: Uric acid anhydrous was the main crystalline form of the stones. The mean size of the crystals was 91.3 ± 28.5 nm. No significant differences were found regarding uric acid crystal size in the stones by comparison to the stone location or the patient’s age. However, particle sizes of uric acid kidney stones were significantly different between male and female patients (84.7 ± 5.3 vs. 140.2 ± 6.7 nm, p < 0.000003) in the absence of diabetes mellitus. Interestingly, when type 2 diabetes appeared, this structural difference between male and female vanished (76.1 ± 3.9 vs. 78.8 ± 4.2 nm, not significant). Thus, the complete set of structural data is in line with observations regarding epidemiological data. Some explanations based on supersaturation are discussed.

Excessive growth of pulmonary arterial (PA) smooth muscle cells (SMCs) is a major component of PA hypertension (PAH). The calcium-activated neutral cysteine proteases calpains 1 and 2, expressed by PASMCs, contribute to PH but are tightly controlled by a single specific inhibitor, calpastatin. Our objective was to investigate calpastatin during pulmonary hypertension (PH) progression and its potential role as an intracellular and/or extracellular effector. We assessed calpains and calpastatin in patients with idiopathic PAH and mice with hypoxic or spontaneous (SM22-5HTT(+) strain) PH. To assess intracellular and extracellular roles for calpastatin, we studied effects of the calpain inhibitor PD150606 on hypoxic PH in mice with calpastatin overexpression driven by the cytomegalovirus promoter (CMV-Cast) or C-reactive protein (CRP) promoter (CRP-Cast), inducing increased calpastatin production ubiquitously and in the liver, respectively. Chronically hypoxic and SM22-5HTT(+) mice exhibited increased lung calpastatin and calpain 1 and 2 protein levels and activity, both intracellularly and extracellularly. Prominent calpastatin and calpain immunostaining was found in PASMCs of remodeled vessels in mice and patients with PAH, who also exhibited increased plasma calpastatin levels. CMV-Cast and CRP-Cast mice showed similarly decreased PH severity compared with wild-type mice, with no additional effect of PD150606 treatment. In cultured PASMCs from wild-type and CMV-Cast mice, exogenous calpastatin decreased cell proliferation and migration with similar potency as PD150606 and suppressed fibronectin-induced potentiation. These results indicate that calpastatin limits PH severity via extracellular mechanisms. They suggest a new approach to the development of treatments for PH.

RESUME Alexander Randall, un urologue americain qui exercait au debut du XXe siecle, a identifie des plaques de phosphate de calcium presentes a la pointe des papilles renales comme etant a l’origine des calculs renaux. Ces plaques qui portent desormais sont nom naissent dans l’interstitium des papilles renales, se developpent en rompant l’urothelium puis une fois au contact de l’urine permettent la nucleation heterogene de cristaux d’oxalate de calcium monohydrate (whewellite) et la croissance de calculs oxalocalciques. Initialement identifiees sur des series autopsiques, ces plaques ont suscite un interet au milieu du siecle dernier puis un relatif oubli jusqu’a ces deux dernieres decennies. Les progres de l’urologie et notamment le developpement de l’ureteroscopie souple ont permis de visualiser les plaques sur les papilles et leur lien avec les calculs renaux. D’autre part, l’analyse attentive des calculs oxalocalciques expulses par les patients depuis une trentaine d’annees revele qu’un nombre croissant de calculs presente une ombilication papillaire avec des residus de plaque, en particulier chez les jeunes adultes, suggerant que les plaques de Randall sont a l’origine d’un nombre croissant de calculs oxalocalciques chez des sujets de plus en plus jeunes. La question de la responsabilite des plaques de Randall dans l’augmentation de l’incidence de la maladie lithiasique est posee. L’identification des mecanismes genetiques et environnementaux responsables de leur formation, le detail de leur composition et la mise en evidence de leur site initial de formation dans la papille sont des enjeux d’autant plus importants que ces plaques ont un impact clinique chez des millions de personnes dans le monde.

Most mouse kidney stone models induce nephrocalcinosis rather than urolithiasis. The aim of our study was to find an accelerated experimental model in order to study the early events of stone formation, that is, at the time of crystal binding to intrarenal urothelium. C57B6 mice exposed to vitamin D supplements and water containing hydroxyl-L-proline, ammonium chloride and calcium chloride were studied for 42 days. A group receiving urothelial cell mitogen Fibroblast Growth Factor 7 (FGF7) was compared to control group receiving saline. Calcium oxalate monohydrate (COM) crystals were detected in urines by day 2 and within urinary spaces in specialized fornix areas in both groups as soon as day 14 with enhanced deposits in FGF7 group compared to controls at day 21. Urothelial cells proliferation, uroplakin III downregulation and de novo expression of osteopontin receptor CD44 detected in FGF7 group, were delayed in the control group (day 42). Crystal aggregates within specialized fornix areas by day 42 were located in urinary spaces but also within and under a multilayered metaplastic urothelium, simultaneous to macrophages influx. Point of note, administration of a normal diet by day 21 was responsible for a spontaneous crystal clearance. Our data show that under supersaturation conditions, urothelial cell proliferation and calcium oxalate crystal retention occur within specialized fornix areas. Enhanced crystal deposits following FGF7 administration suggest that urothelium proliferation would be a relevant trigger for renal stone formation.

Background Pseudoxanthoma elasticum (PXE) is a genetic disease caused by mutations in the ABCC6 gene that result in low pyrophosphate levels and subsequent progressive soft tissue calcifications. PXE mainly affects the skin, retina, and arteries. However, many patients with PXE experience kidney stones. We determined the prevalence of this pathology in patients with PXE and examined the possible underlying mechanisms in murine models. Methods We conducted a retrospective study in a large cohort of patients with PXE and analyzed urine samples and kidneys from Abcc6 −/− mice at various ages. We used Yasue staining, scanning electron microscopy, electron microscopy coupled to electron energy loss spectroscopy, and Fourier transform infrared microspectroscopy to characterize kidney calcifications. Results Among 113 patients with PXE, 45 (40%) had a past medical history of kidney stones. Five of six computed tomography scans performed showed evidence of massive papillary calcifications (Randall plaques). Abcc6 −/− mice spontaneously developed kidney interstitial apatite calcifications with aging. These calcifications appeared specifically at the tip of the papilla and formed Randall plaques similar to those observed in human kidneys. Compared with controls, Abcc6 −/− mice had low urinary excretion of pyrophosphate. Conclusions The frequency of kidney stones and probably, Randall plaque is extremely high in patients with PXE, and Abcc6 −/− mice provide a new and useful model in which to study Randall plaque formation. Our findings also suggest that pyrophosphate administration should be evaluated for the prevention of Randall plaque and kidney stones.

Background Aortic stiffness assessed by carotid-femoral pulse wave velocity (CF-PWV) is a predictor of mortality in several populations. However, little is known in kidney transplant recipients. Our objectives were to evaluate the ability of CF-PWV measured 3 months following transplantation to predict mortality, graft loss and its potential links to measured Glomerular Filtration Rate (mGFR) or kidney graft microvasculature parameters. Methods The study is based on a monocentric retrospective cohort including 220 adult kidney graft recipients evaluated three months after transplantation. CF-PWV measures, clinical, laboratory and histological data performed at 3 (M3) and 12 months (M12) following transplantation were retrospectively collected. The two primary endpoints were all-cause mortality and occurrence of end stage renal disease (ESRD) defined by initiation of dialysis. Results After a median follow up of 5.5 years [1.9; 8.8], death and graft loss occurred in 10 and 12 patients respectively. M3 CF-PWV was an independent mortality risk factor (HR = 1.29 [1.03; 1.61]; p = 0.03), despite no aortic stiffness variation during the first year of transplantation. Of notice, M3 CF-PWV was not associated with M12 mGFR or ESRD outcome. Graft microcirculation assessed by Banff vascular fibrous intimal thickening score (cv) worsened between M3 and M12 (p = 0.01), but no link was found with CF-PWV, mGFR or ESRD outcome. Surprisingly, acute rejections at M3 were associated after adjustment with mortality (p = 0.03) but not ESRD. Conclusion Aortic stiffness measured 3 months after kidney transplantation is a strong predictor of mortality with no obvious influence on kidney graft microvasculature or graft loss.

International audience; Most of kidney stones are supposed to originate from Randall's plaque at the tip of the papilla or from papillary tubular plugs. Nevertheless, the frequency and the composition of crystalline plugs remain only partly described. The objective was to assess the frequency, the composition and the topography of papillary plugs in human kidneys. A total of 76 papillae from 25 kidneys removed for cancer and without stones were analysed by immunohistochemistry combined with Yasue staining, field emission-scanning electron microscopy and Fourier transformed infrared micro-spectroscopy. Papillary tubular plugs have been observed by Yasue staining in 23/25 patients (92%) and 52/76 papillae (68%). Most of these plugs were made of calcium phosphate, mainly carbonated apatite and amorphous calcium phosphate, and rarely octacalcium phosphate pen-tahydrate. Calcium and magnesium phosphate (whitlockite) have also been observed. Based upon immunostaining coupled to Yasue coloration, most of calcium phosphate plugs were located in the deepest part of the loop of Henle. Calcium oxalate monohydrate and dihydrate tubular plugs were less frequent and stood in collecting ducts. At last, we observed calcium phosphate plugs deforming and sometimes breaking adjacent collecting ducts. Papillary tubular plugging, which may be considered as a potential first step toward kidney stone formation, is a very frequent setting, even in kidneys of non-stone formers. The variety in their composition and the distal precipitation of calcium oxalate suggest that plugs may occur in various conditions of urine supersaturation. Plugs were sometimes associated with collecting duct deformation.

International audience; The estimated lifetime risk of nephrolithiasis is growing nowadays, and the formation of kidney stones is frequently promoted by hypercalciuria. Vitamin D, and especially its active metabolite calcitriol, increase digestive calcium absorption—as urinary calcium excretion is directly correlated with digestive calcium absorption, vitamin D metabolites could theoretically increase calciuria and promote urinary stone formation. Nevertheless, there was, until recently, low evidence that 25-hydroxyvitamin D serum levels would be correlated with kidney stone formation, even if high calcitriol concentrations are frequently observed in hypercalciuric stone formers. Low 25-hydroxyvitamin D serum levels have been associated with a broad spectrum of diseases, leading to a huge increase in vitamin D prescription in the general population. In parallel, an increased frequency of kidney stone episodes has been observed in prospective studies evaluating vitamin D alone or in association with calcium supplements, and epidemiological studies have identified an association between high 25-hydroxyvitamin D serum levels and kidney stone formation in some groups of patients. Moreover, urinary calcium excretion has been shown to increase in response to vitamin D supplements, at least in some groups of kidney stone formers. It seems likely that predisposed individuals may develop hypercalciuria and kidney stones in response to vitamin D supplements.

Introduction L’endocardite infectieuse (EI) est une pathologie rare mais frequemment compliquee d’insuffisance renale (IR). Parmi les mecanismes physiopathologiques decrits, la nephropathie microcristalline aux cristaux d’antibiotiques semble etre sous evaluee. L’objectif de notre etude est de decrire la prevalence et l’impact sur le pronostic renal des cristaux urinaires d’antibiotique ainsi que d’identifier les facteurs associes a leur survenue. Methodes Tous les patients admis pour suspicion d’EI sur une periode de 12 mois et traites par antibiotherapie ont beneficie d’une ou plusieurs cristalluries. Cette analyse etait realisee par microscopie en lumiere polarise et spectroscopie infrarouge sur urines fraiches. Le critere de jugement principal etait la presence de cristaux d’antibiotique. Les criteres de jugement secondaires etaient la survenue d’une IRAigue durant l’hospitalisation et le DFG a j14. Un modele de regression mixte a ete applique afin de prendre en compte le caractere longitudinal des cristalluries realisees. Resultats obtenus ou attendus 65 cristalluries issues de 34 patients ont ete realisees. Le delai median de realisation de la 1e cristallurie etait de 5[3–6]jours. 18/65 prelevements issus de 14/34 patients sont revenus positifs. La creatininemie concomitante de l’analyse urinaire etait de 99[65–206]μmol/L. La presence de cristaux etait associee a une concentration serique d’amoxicilline elevee (OR = 1,02[1,01–1,05]) et un pH urinaire bas (OR = 0,79[0,62–0,94]). Une IRAigue etait observee chez 18/34 des patients. Aucune association entre une cristallurie positive et la survenue d’une IRA n’etait retrouve (p = 0,9). A j14, 11/34 patients avaient un DFG Conclusion Chez les patients traites par antibiotherapie pour EI, la survenue de cristaux urinaire d’antibiotiques est frequemment observee et associe au pH urinaire et a la concentration serique d’antibiotique. Il n’a pas ete observe d’association avec le pronostic renal. Des etudes supplementaires sont necessaires pour confirmer ces resultats.

Summary The management of sickle cell nephropathy (SCN) at an early stage is an important issue to prevent renal and cardiovascular morbidity and mortality. This study aimed to evaluate in this population, whether angiotensin converting enzyme inhibitors (ACEIs) treatment could exert a cardio-renal protection in a SCN cohort. Forty-two SCN patients (urine albumin:creatinine ratio (ACR) > 10 mg/mmol) were treated with ACEIs for 6 months, then evaluated for ACR, measured glomerular filtration rate (mGFR) together with haematological and cardiovascular parameters. A 1-month washout was also performed in order to differentiate short- and long-term ACEIs effects. A decrease in ACR baseline value (>30%) was detected in 62% of cases (mean ACR: 46·4 ± 7·6 and 26·4 ± 3·9 mg/mmol at baseline and 6 months respectively; P = 0·002), whereas mGFR values were unchanged. ACR decrease was detected at 1 month following ACEI initiation (32·9 ± 6·9, P = 0·02) with a persistent trend after withdrawal (P = 0·08). ACEIs also decreased diastolic blood pressure (P = 0·007), pulse wave velocity (P = 0·01), tricuspid regurgitation velocity (TRV; P = 0·04), asymmetric dimethyl arginine (ADMA: P = 0·001) and haemoglobin (P = 0·01) while conventional haemolytic biomarkers were unchanged. Our data suggest that ACEIs are safe and effective at decreasing albuminuria in sickle cell patients with a beneficial effect on specific mortality risk factors, such as TRV and asymmetric dimethyl arginine.

International audience; Vancomycin is a widely prescribed antibiotic, but the exact nature of vancomycin-associated nephrotoxicity is unclear, in particular when considering the frequent coadministration of aminoglycosides. We describe here the initial case of a 56-year-old woman with normal renal function developing unexplained ARF without hypovolemia after administration of vancomycin without coadministration of aminoglycosides. Studying the patient’s renal biopsy specimen, we ascertained that obstructive tubular casts composed of noncrystal nanospheric vancomycin aggregates entangled with uromodulin explained the vancomycin-associated ARF. We developed in parallel a new immunohistologic staining technique to detect vancomycin in renal tissue and confirmed retrospectively that deleterious vancomycin-associated casts existed in eight additional patients with acute tubular necrosis in the absence of hypovolemia. Concomitant high vancomycin trough plasma levels had been observed in each patient. We also reproduced experimentally the toxic and obstructive nature of vancomycin-associated cast nephropathy in mice, which we detected using different in vivo imaging techniques. In conclusion, the interaction of uromodulin with nanospheric vancomycin aggregates represents a new mode of tubular cast formation, revealing the hitherto unsuspected mechanism of vancomycin-associated renal injury.

This paper gathers different investigations dedicated to pathological calcifications made of apatite in order to define the structural characteristics, which display a clinical significance. Such approach helps the clinician to define the characterisation tools, which have to be combined to obtain such structural characteristics in order to establish a possible relationship between the structural characteristics of the pathological calcifications and the disease. The fact that it is now possible to perform such experiments at the subcellular scale gives new opportunities for a large number of diseases associated to pathological calcification. Moreover, new diagnostic tools based on these characterisation techniques, quite sensitive to analyze microcrystals, can be defined.

Introduction Le Pseudoxanthome elastique (PXE) est une maladie genetique recessive liee a une mutation du gene ABCC6, exprime dans le foie et le rein. Cette mutation est a l’origine d’un defaut systemique de production de pyrophosphate, un inhibiteur de la mineralisation. Les patients atteints de PXE developpent des calcifications de la peau, de la retine et des arteres mais la presence de calcifications renales chez ces patients a ete tres peu documentee. Patients/Materiels et methodes Une etude retrospective sous forme de questionnaire a ete conduite parmi 113 patients atteints de PXE afin d’evaluer la prevalence de la maladie lithiasique et de ses complications. Les scanners renaux et analyses biologiques de 6 patients atteints de PXE et de lithiase ont egalement ete analyses. Enfin, une etude a ete menee in vivo a l’aide d’un modele murin de PXE (souris Abcc6−/−), dont la biologie urinaire et les calcifications renales ont ete analysees entre l’âge de 2 mois et 2 ans. Chez certains animaux la fonction hepatique d’ABCC6 a ete restauree par transfection d’un gene fonctionnel humain, avec des animaux controles transfectes par un gene mute non fonctionnel. Les calcifications ont ete etudiees par la coloration de Yasue, par microscopie electronique a balayage et a transmission et par micro-spectrometrie infrarouge. Observation/Resultats Parmi les 113 patients ayant repondu au questionnaire, 45 (40 %) ont ete atteints de colique nephretique liee a des calculs renaux. Les scanners ont mis en evidence des calcifications papillaires massives chez 5 patients sur 6. Les souris Abcc6−/− ont developpe spontanement des calcifications avec l’âge, manifestes apres 12 mois et massives a 24 mois, localisees electivement a la pointe des papilles renales, interstitielles, faites de phosphate de calcium amorphe et d’apatite, soit toutes les caracteristiques de la plaque de Randall humaine. L’expression de la proteine ABCC6 fonctionnelle dans le foie de ces animaux a permis de restaurer la production de pyrophosphate et de prevenir le developpement de la plaque de Randall dans le rein. Discussion Le deficit en pyrophosphate circulant lie a une mutation ou une deletion d’ABCC6 est suffisant pour induire des calcifications papillaires massives, des plaques de Randall qui sont la premiere etape de la formation ulterieure des calculs urinaires chez l’homme. Conclusion Il s’agit du premier modele murin mimant reellement les caracteristiques de la plaque de Randall humaine. Le traitement par pyrophosphates exogenes, potentiellement realisable chez l’homme, pourrait ainsi prevenir le developpement de ces calcifications pathologiques.

Introduction La nephropathie a 2,8 di-hydroxyadenine (2,8 DHA) est une maladie severe, secondaire a une mutation autosomique recessive du gene qui code pour l’adenine phosphoribosyl transferase (APRT). Elle se manifeste par la survenue de lithiase renale et/ou par la precipitation intratubulaire de 2,8 DHA et conduit a la maladie renale chronique, recidivant sur le greffon renal. Le traitement preventif repose sur l’administration d’inhibiteurs de la xanthine oxydase. L’objectif de cette etude etait d’evaluer l’impact de ces depots cristallins sur l’inflammation tissulaire renale. Patients/materiels et methodes Nous avons analyse 17 biopsies renales realisees chez 12 patients atteints de mutations de l’APRT (10 femmes, 2 hommes ; 12 greffons et 5 reins natifs). Nous avons realise des immunomarquages des cellules inflammatoires. En parallele, nous avons etudie in vitro la reponse inflammatoire induite par des cristaux de DHA (et des cristaux d’urate de sodium comme controles positifs) sur une lignee de macrophages murins RAW 264.7, notamment la secretion de cytokines (Elisa et qPCR). Resultats Les cristaux de DHA avaient un aspect tres variable selon les prelevements : irreguliers (n = 17), spheriques (n = 16), ou en anneau (n = 10). Leur aspect en lumiere polarisee etait radial dans 100 % des biopsies (n = 17), en aiguilles (n = 7) ou en croix de malte (n = 6). Leur localisation etait intratubulaire (n = 16), intra-epitheliale (n = 17) ou interstitielle (n = 12). L’ensemble des prelevements comportait des lesions de necrose tubulaire (n = 17), souvent une atrophie tubulaire et une fibrose interstitielle (n = 14), et une inflammation interstitielle diffuse (n = 9). Des lesions granulomateuses ont ete observees dans 88 % des biopsies (n = 15), avec presence de cellules geantes multinucleees englobant les cristaux et entoures de fibrose interstitielle. Les macrophages etaient repartis de facon stereotypee autour des cristallites (M1 au centre, M2 a l’exterieur). In vitro, les cristaux de DHA ont induit la formation de granulomes (macrophages « geants », avec aspect dendritique, cellules multinuclees), a partir de la 72e heure. L’exposition des macrophages aux cristaux de 2,8 DHA a induit une augmentation de secretion de cytokines, notamment de TNF-alpha. Conclusion La nephropathie a 2,8 DHA est une maladie granulomateuse, ou l’inflammation tissulaire joue probablement un role majeur dans la degradation de la fonction renale, qui n’est donc probablement pas liee qu’a l’obstruction tubulaire. Des modeles in vivo vont permettre de determiner si l’ajout de traitements anti-inflammatoires (ciblant le TNF-alpha) peut ameliorer la fonction renale en synergie avec les inhibiteurs de la xanthine oxydase.

Introduction Le Hungry Bone Syndrome (HBS) est defini par une hypocalcemie persistante pendant plusieurs jours apres l’effondrement des taux de PTH, par un processus actif de remineralisation osseuse. C’est une complication de la parathyroidectomie realisee dans le cadre des hyperparathyroidies primaires, ou autonomisees chez l’insuffisant renal. Un HBS peut etre observe a minima apres transplantation renale. Nous decrivons ici un cas extremement severe de HBS dans les suites d’une transplantation renale. Patients/Materiels et methodes/Observation Une patiente de 36 ans, Malienne, hemodialysee depuis 5 ans, beneficie d’une transplantation renale d’un donneur decede quatre semaines apres une parathyroidectomie subtotale (hyperplasie diffuse). Le jour de la greffe la calcemie corrigee est a 2,18 mmol/L et la PTH a 51 ng/L (n = 17–65 pg/mL) avec des dosages de 25-OH D3 et de calcitriol normaux. Les suites sont marquees par une reprise immediate de fonction avec un nadir de creatininemie a 124 μmol/L a j10. Elle consulte a j21 de la greffe pour crampes, spasmes de la face et du cou. On note une calcemie corrigee a 1,21 mmol/L, ionisee a 0,58 mmol/L (n = 1,14–1,31 mmol/L), un taux de parathormone inapproprie a 35 pg/mL, une hyperphosphoremie transitoire a 2 mmol/L, et une hypomagnesemie a 0,66 mmol/L. La calciurie, la magnesurie et la phosphaturie sont effondrees, le calcitriol est augmente, sous alfacalcidol (172 pg/mL, 17 En depit d’apports calciques per os de plusieurs grammes par jour et d’un traitement par alfacalcidol a pres de 2 μg/jour, la calcemie ne peut etre maintenue a un taux suffisant pour eviter les symptomes qu’au prix d’une perfusion de calcium, avec des apports IV allant jusqu’a 12 grammes/jour sur voie centrale. Durant plus de 3 semaines, la calciurie reste effondree et la calcemie reste basse en depit des apports massifs de calcium, traduisant un flux osseux entrant de calcium extremement eleve. Le TEP-scanner au fluorure de sodium objective une intense captation intense et diffuse du traceur par le squelette axial et peripherique proximal. Resultats/Discussion/Conclusion La conjonction dans le temps de deux etiologies de HBS, la parathyroidectomie et la transplantation renale, ont abouti a une hypocalcemie profonde et symptomatique necessitant des doses massives de calcium IV en plus des apports per os de calcium et de vitamine D active de facon prolongee. Une surveillance rapprochee de la calcemie et de la calciurie doit etre realisee dans les suites d’une transplantation renale suivant une parathyroidectomie recente.

Introduction L’amoxicilline (AMX) est un des antibiotiques les plus prescrits et sa toxicite renale est peu decrite. La presence de cristaux d’AMX dans les urines au cours de cas d’insuffisance renale aigue (IRA) associees a l’AMX a fait evoquer un mecanisme de precipitation intra-tubulaire. Cependant, il n’existe pas de description de cristaux d’AMX dans le parenchyme renal. Methodes Nous decrivons deux patients presentant une IRA secondaire a l’AMX et ayant beneficie d’une biopsie renale. Nous avons ensuite elabore un modele de toxicite chez le rongeur. Nous decrivons ces deux cas de nephropathie associee a l’AMX et l’analyse de leurs biopsies renales avec spectrophotometrie infra-rouge (SPIR) et microscopie electronique a balayage (MEB). Dans un deuxieme temps, differentes doses d’AMX (de 200 a 3000 mg/kg/jour) ont ete injectees par voie intra-peritoneale pendant 48 heures aux rats, et jusqu’a 9 jours aux souris. Nous avons analyse au sacrifice la cristallurie, la fonction renale et l’histologie des rongeurs. Resultats obtenus ou attendus Deux patients de 58 et 68 ans hospitalises pour une meningite bacterienne ont presente une IRA a l’AMX. Ils recevaient 12 g/jour et 25 g/jour respectivement en intraveineux. L’analyse en microscopie optique des biopsies revele dans les deux cas une tubulopathie severe avec des lesions de nephrose osmotique. Dans un des cas, l’analyse SPIR combinee a la MEB confirme la presence d’un depot intra-tubulaire d’AMX. La cristallurie n’etait pas disponible pour ces deux cas. In vivo, l’injection aux rongeurs d’AMX a posologie croissante aboutit a une IRA avec necrose tubulaire aigue et nephrose osmotique. On ne retrouve pas de cristaux d’AMX intraparenchymateux chez le rongeur ni de cristallurie. Conclusion Ces deux cas suggerent qu’il peut exister une toxicite tubulaire directe de l’AMX avec des lesions de nephrose osmotique reproduites dans le modele animal. On decrit, pour la premiere fois, un depot intratubulaire d’AMX chez l’Homme suggerant une precipitation intratubulaire associee.

Introduction Grâce a une prise en charge precoce, l’esperance de vie des patients drepanocytaires homozygotes augmente expliquant l’incidence des complications chroniques liees a cette maladie. La prevalence de la maladie renale chronique (MRC) serait de 85 % chez les patients de plus de 65 ans drepanocytaires. L’objectif de notre etude est de determiner les facteurs de risque d’apparition d’une MRC stade 2. Methodes Etude prospective observationnelle monocentrique d’une cohorte de 688 patients suivis entre 2007 et 2018 dans un centre de drepanocytose. Un total de 660 patients ayant un eDFG > 90 ml/min/1.73 m2 (CKD EPI) ont ete inclus. L’evenement etudie etait l’apparition d’une MRC stade 2 (DFG Resultats obtenus ou attendus Le delai median de suivi etait de 3,7 ans (IQR 0,76–7,36). L’âge median a l’inclusion etait de 22 ans (IQR 19–29), avec 377 (57,12 %) femmes. Une hyperfiltration glomerulaire (DFG > 130 ml/min/1,73m2) etait presente chez 380 patients (57,58 %), une microalbuminurie et une macroalbuminurie chez 218 (33,70 %) et 83 (12,83 %) patients respectivement. Les pressions arterielles systoliques et diastoliques moyennes (± deviation standard) etaient basses ; respectivement 114,8 mmHg (± 13,6) et 66,0 (± 9,7) mmHg. Les facteurs associes a l’apparition d’une MRC stade 2 etaient (analyse multivariee) : la proteinurie (pour 100 mg/mmol creatinine) HR : 1,24 [1,07–1,45] p = 0,005, la pression arterielle diastolique (PAD) (pour 10 mmHg) HR : 1,35 [1,03–1,76] p = 0,029 et les LDH HR : 1,22 [1,04–1,43] p = 0,015, le DFG initial HR : 0,92 [0,90–0,95] p Conclusion Notre etude montre une augmentation significative du risque de degradation de la fonction renale dans les cas de macroalbuminurie, d’augmentation de la PAD et d’hemolyse chronique intense. Ces resultats suggerent l’importance d’un controle tensionnel dans cette population avec des valeurs cibles particulierement basses.

International audience; Michel Daudon is a worldwide recognized biologist who performed several major breakthroughs in the field of urolithiasis. It is indeed difficult to address his outstanding personality and his great contributions to urolithiasis. Michel Daudon studied many aspects of urolithiasis, from urine crystals to kidney stone morphoconstitutional analysis. He is the author of more than 500 scientific articles on urolithiasis, and more generally on pathological calcifications, including several books and nice articles in the New England Journal of Medicine, The Lancet, Kidney International, among many others. Michel Daudon is not only a prolific writer; his broad fields of interests range from chemistry to biology or renal pathology, but also clinical bedside subjects explain his success and his attractive way of explaining pathophysiological processes. He has already given more than 300 lectures, in France as abroad.

Clinical interest for the identification of the crystalline phase of CaOx, either monohydrate (COM) or dihydrate (COD), in crystalluria analysis is debated. In the present study, we simultaneously determined calcium and oxalate concentration and identified the two crystalline phases of CaOx crystals in 1288 first morning urine samples from 407 CaOx stone formers with crystalluria.The mean concentration of oxalate ions was higher in urine samples containing COM than in those containing COD crystals, and conversely the mean concentration of calcium ions was markedly higher in urine samples containing COD than in those containing COM crystals. COM crystals were predominant (93%) in urine samples with a Ca/Ox molar ratio 14 (i.e. markedly hypercalciuric). In the intermediate Ca/Ox values, mixed COD and COM crystals were present. Logistic regression analysis showed that increasing the concentration of calcium resulted in a higher risk of COD than of COM crystal formation, whereas increasing oxalate concentration was associated with a greater COM than COD crystal formation. In conclusion, this study demonstrates the calcium-dependence of COD and the oxalate-dependence of COM crystal formation. Thus, a crystalluria mainly composed of COD suggests a search for conditions associated with hypercalciuria whereas an abundant crystalluria made of COM crystals should prompt a search for heavy hyperoxaluria and noteworthy primary hyperoxaluria.

Biopsies of native or transplanted kidneys in patients suffering chronic or acute renal failure are commonly stained for tissue examination and search for possible crystal deposits which are then identified by polarizing microscopy and staining by von Kossa's method revealing mainly calcium deposits. Renal biopsies presumably containing crystal deposits were analyzed with a Spotlight 400 μFTIR (Fourier Transform Infra Red) imaging system in the mid-infrared spectral range to obtain infrared maps of tissue slides at high spatial resolution, down to 10 microns. When required, an optional ATR imaging accessory was used, improving the spatial resolution by a factor four, down to 3 microns at 1000 cm−1. Among the 685 renal biopsies, 72% contained abnormal non-proteic material. Among them, 2.16% contained drug crystals (triamterene, N-acetylsulfadiazine, ciprofloxacine, indinavir, atazanavir, foscarnet, and vancomycine). We focused on foscarnet and atazanavir deposits. In the case of foscarnet-induced renal failure, two types of crystals were found in one patient. They were located in different parts of the nephron: sodium and/or calcium phosphonoformate crystals within glomerules and carbapatite in the proximal tubular cells. By contrast, atazanavir was only found in the tubular lumen of the nephron.A precise identification of crystal deposits is essential for the diagnosis of an unexplained renal failure. Common histological procedures clearly fail to identify crystals deposits accurately. In most cases, light and polarizing microscopic examination should be completed by FTIR analysis.

International audience; Severe hypertension can lead to malignant hypertension (MH) with renal thrombotic microangiopathy and hemolysis. The role of plasma heme release in this setting is unknown. We aimed at evaluating the effect of a mild plasma heme increase by hemin administration in angiotensin II (AngII)-mediated hypertensive rats. Prevalence of MH and blood pressure values were similar in AngII and AngII + hemin groups. MH rats displayed a decreased renal blood flow (RBF), increased renal vascular resistances (RVR), and increased aorta and interlobar arteries remodeling with a severe renal microcirculation assessed by peritubular capillaries (PTC) rarefaction. Hemin-treated rats with or without AngII displayed also a decreased RBF and increased RVR explained only by PCT rarefaction. In AngII rats, RBF was similar to controls (with increased RVR). PTC density appeared strongly correlated to tubular damage score (rho = −0.65, p

International audience; Introduction: The presence of pathological calcifications, which can be either concretions or ectopic call for physicochemical characterisation techniques in order to define a significant diagnosis. The aim of this review is to present a set of characterisation techniques able to describe at the micrometer scale their structural and chemical characteristics and show their place at the hospital.Method: Results already published in the last ten years based on characterisation techniques present in laboratories or specific to large-scale instruments are presented. Their usefulness for the clinician is discussed.Results: The presence and role of heavy metals in urinary stones through data collected through μX-ray fluorescence is debated. If these data suggest a simple substitution process of calcium, recent data suggest that weddellite is associated to a Zn-rich environment, partly favoured by an inflammation process. Investigation on the chemistry and the structure of unusual deposits in kidney biopsies show a great chemical diversity of ectopic calcifications. Such diversity shows that staining procedures to characterize such deposits are obsolete. Finally, several diseases which can be assessed through techniques specific to large-scale instruments and defined by the clinician are presented.Conclusion: The chemical and structural complexity of pathological calcifications call for a characterization through physicochemical techniques. Only such approach allows the clinician to define a significant diagnosis and to care the patient.; Introduction: Les calcifications pathologiques rénales qu’elles s’apparentent à des concrétions ou à des calcifications tissulaires impliquent de savoir déterminer correctement les composés chimiques qui les composent afin de porter un diagnostic menant à une prise en charge adéquate. Le but de cet article de revue est de dépeindre les techniques physicochimiques aptes à réaliser cette caractérisation structurale et chimique à l’échelle du micromètre et de montrer leur insertion au sein même de l’hôpital.Méthode: Les résultats acquis ces dix dernières années utilisant des techniques de caractérisation de laboratoire ou spécifiques aux grands instruments comme le synchrotron Soleil ou le laboratoire Léon-Brillouin sont présentés. Leur pertinence dans l’activité du clinicien a été examinée.Résultats: Le rôle des éléments traces dans la pathogenèse des calculs rénaux au travers de données acquises par fluorescence X est discuté. Si leurs propriétés catalytiques ne sont pas confirmées par les données expérimentales, l’analyse fine a dévoilé certaines particularités intéressantes concernant les teneurs en Zn des deux composés d’oxalate de calcium, i.e. whewellite et weddellite. En effet, les calculs de weddellite présentent des bipyramides aux angles et arêtes vifs susceptibles d’induire une inflammation tissulaire. L’analyse morphoconstitutionnelle des biopsies rénales dévoile une grande diversité chimique puisque 22 corps différents ont été identifiés. Cette diversité indique que les méthodes de coloration dont l’anatomopathologiste dispose sont insuffisantes pour caractériser ces corps. Enfin, nous énumérons différentes problématiques abordées par les grands instruments et définies par le clinicien.Conclusion: La complexité structurale et la diversité chimique des calcifications pathologiques invitent à la mise en œuvre de techniques de caractérisation physicochimiques. Seule cette approche permet au clinicien la pose d’un diagnostic fiable et donc une prise en charge adéquate du patient.

International audience; BackgroundBiopsies of native or transplanted kidneys in patients suffering chronic or acute renal failure are commonly stained for tissue examination and search for possible crystal deposits which are then identified by polarizing microscopy and staining by von Kossa’s method revealing mainly calcium deposits. Aim of the studyRevisiting the nature of crystal deposits in kidney tissue sections by infrared microscopy. Method 205 renal biopsies presumably containing crystal deposits were analyzed with the Spotlight 400 FTIR imaging System in the mid infrared spectral range to obtain infrared maps of tissue slides at high spatial resolution, down to 10 microns.Results Based on infrared analysis, we identified crystals in all biopsies including 78.8% calcium-containing crystals, 15.3% purine crystals of which 90.5% of 2,8-dihydroxyadenine, and 5.9% drugs crystals. Among birefringent crystals observed under polarized light, we identified 60.2% of calcium oxalate monohydrate, 2.6% of lipids, 13.1% of 2,8-dihydroxyadenine, 7.8% of uric acids and urates, 5.9% of drug crystals (triamterene, N-acetylsulfadiazine, ciprofloxacine, indinavir monohydrate, atazanavir); and among non refringent crystals: 27% of carbapatite, 14.6% of amorphous carbonated calcium phosphate, 3.9% of drug crystals (foscarnet, vancomycine) and 1.5% of other phosphates. Overall, we identified 26 different types of crystals. Surprisingly, we found mixtures of crystalline phases in a high proportion (23.8%) of biopsy samples.DiscussionCrystals may be related to various pathological conditions. Identification of dihydroxyadenine prompt to treat the patient with allopurinol. In transplanted patients, we found a negative correlation between the amount of calcium phosphate deposits and the graft survival. The high occurrence of mixed crystals may be a marker for successive episodes of kidney dysfunction related to different mechanisms. We therefore suggest that FTIR microspectroscopy is a major diagnostic tool for crystal identification and should be proposed to any patient with a past history of stone disease and presenting with an unexplained renal failure of the native or grafted kidney.Conclusion A precise identification of crystal deposits in the kidney tissue may totally modify the diagnosis of an unexplained kidney dysfunction. Common histological procedures clearly fail to identify accurately crystals deposits and should be completed by infrared analysis.

International audience; Background: Composition, etiology and consequences of microcalcifications in kidney allograft are unclear. Here we test the contribution of infrared microspectroscopy (IR-MS) to the kidney transplant crystal deposits understanding.Materials and methods: We compare 55 patients with calcifications (C+) to 100 without calcification (C-). Biopsies were evaluated by microscopy examination. In case of visualization of crystalloid deposits, an IR-MS analysis was performed with an IN10MX microscope to identify chemical composition and to assess the spatial distribution and quantification of the calcifications (score 1 to 4). Clinical and biochemical data and graft outcome were compared between both populations.Results: Biopsies were performed at 5.2 ± 6 months and 6.9 ± 7 months after transplantation respectively in C- and C+ groups. Microcalcifications were calcium phosphates in 91%. Crystal deposits abundance was considered moderate (score 1-2) for 45 (82%) patients and high (score 3-4) for 10 (18%) patients. Microcalcifications were associated with a higher 3 months post-graft parathyroid hormone concentration (C- 123 ±59 pg/ml; C+ 211±177; p=0.003). A high score of crystal deposits was associated with a poor graft function as judged by the rate of eGRF

International audience; Randall‘s plaques are found in a large number of calcium oxalate stone formers today. From an epidemiological point of view, we observed a constant increase in the proportion of calcium oxalate stones developed from a Randall’s plaque in France over the past forty years. Stones developed on a Randall‘s plaque typically present a small depressed zone (named umbilication) corresponding to the tip of the papilla and containing some material detached from the plaque. Thus, by examining the morphology of the stone, we can determine the part of urinary calculi initiated from a Randall’s plaque. The proportion of Randall plaque-related stones was assessed over 3 time periods: 1989–1991, 1999–2001, and 2009–2011. Because modern techniques used to remove urinary calculi from the urinary tract often use laser or ultrasounds or shock waves, Randall’s plaque is often lost during the treatment. However, around 50% of all stones are spontaneously passed and 10-15% of stones are removed from the urinary tract without fragmentation. So, we examined the morphology of 30,149 non fragmented calcium oxalate stones and we found that 10,282 harbored Randall plaque residues (34.1%). The prevalence of Randall plaque-related stones increased dramatically during the past years. In young women, 17% of calcium oxalate stones were associated with Randall plaque during the 1989–1991 period, but the proportion rose to 59% 20 years later (P

International audience; Vitamin D supplementation in humans should be accompanied by calcium administration to avoid bone demineralization through vitamin D receptor signaling. Here we analyzed whether long-term exposure of rats to vitamin D supplementation, with or without a calcium-rich diet, would promote kidney stone formation. Four groups of rats received vitamin D alone (100,000 UI/kg/3 weeks), a calcium-enriched diet alone, both vitamin D supplementation and calcium-enriched diet, or a standard diet (controls) for 6 months. Serum and urine parameters and crystalluria were monitored. Kidney stones were assessed by 3-dimensional micro-computed tomography, infrared spectroscopy, von Kossa/Yasue staining, and field emission scanning electron microscopy. Although serum calcium levels were similar in the 4 groups, rats receiving vitamin D had a progressive increase in urinary calcium excretion over time, especially those receiving both calcium and vitamin D. However, oral calcium supplementation alone did not increase urinary calcium excretion. At 6 months, rats exposed to both calcium and vitamin D, but not rats exposed to calcium or vitamin D alone, developed significant apatite kidney calcifications (mean volume, 0.121 mm3). Thus, coadministration of vitamin D and increased calcium intake had a synergistic role in tubular calcifications or kidney stone formation in this rat model. Hence, one should be cautious about the cumulative risk of kidney stone formation in humans when exposed to both vitamin D supplementation and high calcium intake.

International audience; Calcium oxalate nephrolithiasis is a common ailment. Frequent risk factors associated with are low diuresis, dietary imbalance, inherited or acquired metabolic disorders. Another important factor, namely the presence of a mineral deposit made of apatite at the surface of the papilla, named Randall's plaque (RP), has been recently underlined. In most cases, RP which serves as a nidus for kidney stone formation is made of calcium phosphate apatite (CA). However, RP does not seem to be composed exclusively of CA. We would like to assess the case of RP where sodium hydrogen urate monohydrate (NaUr) is also present in its chemical composition. To attain this goal, a set of experiments including Environmental Scanning Electron Microscopy (ESEM) and Synchrotron Radiation Fourier Transform InfraRed (SR-FTIR) has been performed to analyze papillae of six kidneys randomly selected after they were surgically removed for cancer. NaUr crystals were found in two samples. We show through ESEM an usual morphology of RP present at the surface of kidney stones in the presence of NaUr. Moreover, we discuss the presence of NaUr in the renal parenchyma and its spatial repartition with CA. The complete set of data indicates that different biochemical mechanisms are probably involved in the pathogenesis of RP. The next step will be to establish a significant relationship between these physicochemical data and the clinical and biochemical data of the patients.

International audience; Learning from Mother Nature constitutes a major intellectual movement encompassing music, architecture and chemistry. In this contribution, we would like to assess the symmetry and the interface present in pathological calcifications in order to propose a bio-inspired approach. We present an overview on the observations made on kidney stone growth at the macroscopic scale. The results are discussed at the molecular scale and examples are proposed to implement the biological asymmetrical growth in pathological calcifications in biomimetic strategies such as in in vitro microfluidic experiments.

International audience; Crystal formation in kidney tissue is increasingly recognized as a major cause of severe or acute renal failure. Kidney biopsies are currently performed and analyzed using different staining procedures. Unfortunately, none of these techniques are able to distinguish the different Ca phosphates (e.g., amorphous or nanostructured Ca phosphate apatite, octacalcium phosphate, brushite…) or Ca oxalates (whewellite and weddellite). Moreover, the crystal’s morphology, a structural parameter proven as a major information to the clinician regarding kidney stones, is not taken into account. Such major limitations call for a different research approach, based on physicochemical techniques. Here we propose classical observations through field-emission scanning electron microscopy experiments combined with energy dispersive spectroscopy as well as measurements through Raman and μFourier transform Infra-Red Spectroscopy. If necessary, in the case of microcrystals, observations using cutting edge technology such as Synchrotron Radiation (SR) – FTIR or SR-UV visible spectroscopy can be subsequently performed on the same sample. Taken together this set of diagnostic tools will help clinicians gather information regarding the nature and the spatial distribution at the subcellular scale of different chemical phases present in kidney biopsies as well as on the crystal morphology and therefore obtain more precise diagnosis.

International audience; L’inflammation est une réaction aiguë de défense de l’organisme face à une agression par exemple d’origine infectieuse ou traumatique. Elle permet la restitution de tissus morphologiquement et fonctionnellement normaux. Néanmoins, pour éviter que son développement se perpétue et conduise lui-même à de nouvelles lésions, il est nécessaire qu’elle se résolve rapidement. L’interruption de la réaction inflammatoire suppose que l’afflux de cellules et la libération ou la fonction de médiateurs inflammatoires soient progressivement limités. C’est le rôle des médiateurs anti-inflammatoires, protéiques et lipidiques, qui sont synthétisés très tôt après l’apparition des premières lésions.Un très bel exemple de résolution de l’inflammation est donné par la cascade de médiateurs lipidiques décrite par Charles Serhan. L’initiation de la réaction inflammatoireest caractérisée par la synthèse, à partir de l’acide arachidonique, de médiateurs (prostaglandine E2 [PGE2] et leucotriène B4 [LTB4]) quifavorisent l’afflux de polynucléaires neutrophiles. Un changement (en anglais, switch) dans le métabolisme de l’acide arachidonique se produit ensuite, conduisant à la synthèse préférentielle d’un nouveau dérivé lipidique, la lipoxine, qui en limitant l’afflux de polynucléaires neutrophiles eten favorisant leur apoptose et leur élimination par des macrophages permet la résolution de l’inflammation. Au cours de ces dix dernières années, notre groupe a mis en évidence un nouvel exemple de switch, cette fois dans l’activité d’une protéase, la calpaïne, qui, de pro-inflammatoire dans le cytoplasme, devient anti-inflammatoire et réparatrice, simplement en passant du cytosol au milieu extracellulaire de la cellule qui la produit.

International audience; BackgroundAlexander Randall identified calcium phosphate plaques in renal papillae as the origin of kidney stones. However, little is known about the early steps of Randall’s plaque formation, preceding the onset of urolithiasis. Our objectives were to characterize the composition and the initial formation site of incipient Randall’s plaque.Material and MethodsFifty-four healthy papillae from kidneys removed for cancer have been analyzed by immunohistochemistry and Von Kossa staining, Field Emission-Scanning Electron Microscopy with Energy Dispersive X-ray analysis, µ-Fourier Transform Infrared Spectroscopy, Cryo-Transmission Electron Microscopy coupled to Selected-Area Electron Diffraction and Electron Energy Loss Spectroscopy. ResultsIncipient Randall’s plaque has been observed in 72.7% of kidneys. Carbonated apatite was the main component of microcalcifications altogether with amorphous calcium phosphate and whitlockite. Incipient plaques stood in the deepest part of the papillae, around the loop of Henle tip but also vasa recta (respectively 62.4 % and 37.2 % of microcalcifications). Microcalcifications were often made of several nanocrystals inside organic material looking like microvesicles. ConclusionIncipient Randall’s plaque is frequent and appears at the tip of renal papillae, around the hairpin structure of the loop of Henle and vasa recta as well. Nanoscale analyses suggest a local nucleation process promoting nanocrystal growth in a supersaturated milieu.

International audience; BackgroundMost mice kidney stone models induce nephrocalcinosis rather than urolithiasis.Materials and methodsC57B6 mice under vitamin D and water containing 4% hydroxyl-L-proline, ammonium chloride (0.28M) and calcium (0.25%) were follow up until day 42. A group receiving Fibroblast Growth Factor 7 (FGF7 i.p. once a week), a urothelial cell mitogen was compared to control group. Localization and identification of crystal deposits were performed with polarizing microscopy, infrared imager and Scanning Electron Microscopy. Urothelial and tubular phenotypes were studied by immunohistochemistry, immunofluorescence and RT PCR.ResultsCalcium oxalate monhydrate (COM) deposits in fornices were detected in all kidneys as soon as day 14 with very few crystals in tubules. On day 21, crystal number was increased in FGF7 compared to control group. Though osteopontin (OPN) was unregulated and detected in urothelial cells, de novo expression of CD44 (osteopontin receptor) receptor was induced only in proliferating urothelial cells either induced by FGF7 or by long standing urine crystal exposure altogether with a loss of apical uroplakines III expression. ConclusionOur model seems interesting to study in KO mice, in order to investigate the critical events leading to urolithiasis. Our data show that urothelial cells proliferation promotes renal crystal retention especially within fornices, probably through different processes involving specific and non-specific crystal adhesion, and thus appears per se as a relevant risk factor for stone formation.

International audience; Renal stone incidence has progressively increased in industrialized countries but the implication of Randall’s plaque in this epidemic remains unknown. Our objectives were to determine whether Randall’s plaque-related stones prevalence increased during the past decades after having analyzed 30,149 intact stones containing mainly calcium oxalate since 1989 (cross-sectional study), and to identify determinants associated with Randall’s plaque-related stones in patients (case-control study). The proportion of Randall’s plaque-related stones was assessed over three time periods: 1989-1991, 1999-2001 and 2009-2011. Moreover, we analyzed clinical and biochemical parameters of 105 patients affected by calcium oxalate stones, with or without plaque.Of 30,149 calcium oxalate stones, 10,282 harboured Randall’s plaque residues (34.1%). The prevalence of Randall’s plaque-related stones increased dramatically during the past years. In young females, 17% of calcium oxalate stones were associated with Randall’s plaque during the 1989-1991 period but the proportion rose to 59% twenty years later (p

International audience; The effect of mixing calcium and oxalate precursors by diffusion at miscible liquid interfaces on calcium oxalate crystalline phases, and in physiological conditions (concentrations and flow rates), is studied using a microfluidic channel. This channel has similar dimensions as the collection duct in human kidneys and serves as a biomimetic model in order to understand the formation of pathological microcalcifications.