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Urine proteomic analysis of cystinuric patients

ABSTRACT: Cystinuria is the most frequent monogenic cause of renal calculi. We previously described a urinary inflammatory signature in 8 cystinuric patients through quantitative proteomic analysis. Our data suggested an association of abnormal renal function and an inflammatory protein profile. The objectives of the present study were to investigate: i) the urinary proteomic profile in cystinuric patients without chronic kidney disease (CKD), and in a control group of healthy volunteers; ii) the potential effect of urine alkalinization on the urinary proteomic profile in cystinuric patients as this is the cornerstone of the preventive medical treatment. Urinary inflammatory profile was evaluated at baseline in the control group and in cystinuric patients not yet treated or for whom the alkalizing treatment or the cysteine- binding thiols have been discontinuated for at least 3 months and with an eGFR (using the CKD-EPI formula) greater than 60 ml/min/1.73m2 (so without CKD G3-G5). In cystinuric patients, change from baseline urinary inflammatory profile was evaluated after 3 months of alkalizing treatment. Urine proteome was analyzed by Nano- liquid chromatography coupled to high resolution mass spectrometry. Twenty-one cystinuric patients (12 men, median age [IQR] 30.0 years [25.0 – 44.0]), and 7 healthy volunteers (2 men, median age 43.1 years [31.0 – 53.9]) were included. Median eGFR in these two groups was 95 ml/min/1.73m2 [87.5 – 111.5] and 116 ml/min/1.73m2 [91.0 – 120.0] respectively (p=0.38). Healthy volunteers had no inflammatory signature. The inflammatory signature was present in the cystinuric patients at different degrees. A hierarchical clustering of the cystinuric patients according to the intensity of the profile of core inflammation signature (11 proteins) was performed. The signature could clearly separate 5 cystinuric patients with a high inflammation from other cystinuric patients and healthy volunteers. The alkalizing treatment was associated with a decrease in the intensity of the urinary inflammatory profile in all 5 cystinuric patients compared to the initial high level of urinary inflammation. A panel of 11 proteins was found as elevated in some cystinuric patients without CKD G3- G5 and could work as an early inflammatory signature. The alkalizing treatment was associated with a reduction of this urinary inflammation signature, suggesting that this panel of proteins could also be a new tool to monitor the treatment of cystinuric patient.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Urine

DISEASE(S): Cystinuria

SUBMITTER: Chiara guerrera

LAB HEAD: Chiara Guerrera

PROVIDER: PXD035631 | Pride | 2024-03-29

REPOSITORIES: Pride

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			Action	DRS
	190621_LFQ-Cys_n1-4_MaCO_PaOL1_1_Q30284.raw	Raw
	190621_LFQ-Cys_n1-4_MaCO_PaOL1_2_Q30285.raw	Raw
	190621_LFQ-Cys_n1-4_MaCO_PaOL1_3_Q30286.raw	Raw
	190621_LFQ-Cys_n1-4_MaCO_PaOL2_1_Q30293.raw	Raw
	190621_LFQ-Cys_n1-4_MaCO_PaOL2_2_Q30294.raw	Raw

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Project description:Pseudomonas aeruginosa is one of the most frequent pathogen dominant in complicated urinary tract infections (UTI). To unravel the adaptation strategies of P. aeruginosa to the conditions in the urinary tract and to define the underlying regulatory network an artificial growth system mimicking the conditions in the urinary tract was established. Transcriptome analyses were used to investigate the physiological status of P. aeruginosa under this conditions. We performed comparisons to identify genes induced under artificial urinary tract conditions to unravel the adaptive strategies and the underlying regulatory network used by Pseudomonas aeruginosa during urinary tract infections using Affimetrix GeneChips. Pseudomonas aeruginosa wild type strain PAO1 was grown in an artificial in vitro growth system mimicking the conditions in the urinary tract. Therefore, biofilms were grown on the surface of membrane filters placed on agar plates at 37 Â°C up to the late logarithmic state under aerobic and anaerobic conditions (incubated in an anaerobic beanch). An artificial urine medium (AUM) simulating the averaged urine of an human adult was used as nutrient souce. 10-fold diluted Luria Bertani (LB)-medium was used as reference medium. For growth under oxygen depletion the media were supplemented with 50 mM KNO3 to sustain anaerobic respiration. The biofilms were harveted at this time points and resuspsended in 0.9% (w/v) NaCl. The OD578 of biofilm suspension was 0.8 for all tested conditions. First comparison: Identification of genes induced or repressed under aerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown aerobically for 18 h to the late logarithmic phase in biofilms on AUM with the transcriptome profile of the PAO1 strain, which was grown aerobically for 18 h to the late logarithmic phase in biofilms on 10-fold diluted LB. Second comparison: Identification of genes induced or repressed under anaerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown anaerobically for 2 days up to the late logarithmic phase in biofilms on AUM supplemented with 50 mM nitrate with the transcriptome profile of the PAO1 strain, which was grown anaerobically for 2 days up to the late logarithmic phase in biofilms on 10-fold diluted LB supplemented with 50 mM nitrate.

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Urine proteomic analysis of cystinuric patients

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