Oxalate and Citrate in Humans - Response to Citrate

This is a single-center study that aims to learn more about how two compounds found in food, oxalate and citrate, interact in the body and may influence a person's chances of forming kidney stones. The study will examine changes in urinary oxalate and citrate levels after participants consume potassium citrate.

Administration of oral citrate without oxalate is the next step to determining a potential transporter linkage in humans as the mechanism driving the oxalate and citrate association. We will measure fractional excretion of oxalate and citrate. Both are freely filtered by the glomerulus and either secreted or reabsorbed (oxalate) or just reabsorbed (citrate). We will be able to assess urine excretions partitioned from the contribution of the filtered load and subsequent tubule handling by using fractional excretion.

Experimental Design: Studies will be carried out in the University of Chicago Medicine (UCM) Clinical Research Center (CRC).

Informed Consent Process: Subjects will be taken through the informed consent process by myself or the study research assistant. Subjects will be given ample time to discuss and ask questions. No study activities will occur until after the subject has signed the consent form.

In NSF (N=12) and CaOx SF (N=12), give a potassium citrate load and measure both urine and serum oxalate and citrate over six hours. Primary endpoint (hypothesis 1a) is change from baseline in fractional excretion of oxalate adjusted for change in fractional excretion citrate after oxalate load. Secondary endpoint (hypothesis 1b) is to compare change in fractional excretion of oxalate adjusted to fractional excretion oxalate in calcium SF versus NSF.

Rationale: There is an association between urine oxalate and citrate excretion. We have completed a study of giving an oxalate load to calcium SF and NSF and found that citrate goes up after an oxalate load and the change in oxalate and citrate was correlated. If this is due to a specific effect of oxalate and citrate transport in the proximal tubule, then administration of an oral citrate load should lead to increased urine oxalate excretion, and this increase may differ between NSF and SF. Therefore, 1a: an oral citrate load will increase fractional excretion of oxalate; and 1b: will raise fractional excretion of oxalate more in NSF than in non-obese SF. If true, this suggests a link at the level of proximal tubule transporter (SLC26A6/NaDC1) function. If false, other mechanisms such as dietary variation, must be responsible

Study Population and Recruitment: Enroll a total of 24 subjects (12 NSF, 12 SF) with equal numbers by sex over the first two years or the award. NSF controls will be recruited from our existing group of previous research participants and from Research Match and The New Normal research participant resources of the UCM Institute for Translational Medicine. Research Match and The New Normal are resources for finding individuals who are interested in participating in research studies. Queries can be used to identify eligible controls. Eligible NSF controls will complete one home 24-hour baseline urine collection at home to screen for any severe acid-base or oxalate abnormalities. SF participants (all of whom have 24-hour data available) will be recruited from the UCM Kidney Stone Clinic (both current and repository of previous patients). The UCM Kidney Stone Clinic is a large clinic with over 50 years of patients. Our group has a long history of successfully recruiting research participants from this source.

Inclusion Criteria:

SF: Age 18-70, history of at least one calcium-based kidney stone NSF: Age 18-70, no history of kidney stone, 24-hour urine oxalate within lab normal (\<50mg/day) Exclusion Criteria: History of primarily uric acid, cysteine, or struvite stones. History of severe acid-base abnormality, very low (less than 100mg/day) or very high (greater than 1500mg/day) urine citrate. Any controls or SF participants who cannot stop diuretic medication or alkali supplementation for the course of the study period. Chronic kidney disease (glomerular filtration rate \<75 ml/min/1.73m2.

NSF and SF with extreme levels of urine citrate will be excluded from this small initial study to reduce heterogeneity and remove focus from the extremes of citrate levels.

Protocol: This protocol was developed and adapted based on previously published methodologies,40-42 as well as a completed IRB (21-1349) approved study of an oxalate load in similar participants. We will select SF patients both on and not on alkali supplementation. Those on alkali supplementation will be asked to discontinue for two weeks prior to study day. For all patients, one week prior to study day, all vitamin C, multivitamins, calcium supplements, and diuretics will be held. Holding of diuretics and alkali supplementation will be done in discussion with patient's primary nephrologist or primary care physician to discuss safety in stopping these medications for the study period. One day prior to study day, participants will complete 24-hour urine collection and food frequency questionnaire at home. On study day, participants will be admitted to the CRC in a fasting state. Three baseline urine and blood specimens will be collected and height and weight will be measured. Participant will consume an oral potassium citrate (40 meq) load. Repeat timed urine (every 45 minutes) and blood specimens (every 45 minutes) will be obtained for 6 hours after the oral oxalate load. Blood collections will be approximately 8ml blood per draw (total of 9 blood draws). All participants will remain fasting until study completion. Patient will be given 500 cc of water at the start of study and water intake throughout the remainder of the study will be allowed ad lib. At study completion, all participants will be offered a boxed lunch.

Urine and Serum Measurements: 24-hour and timed urine collections will be completed. Instructions for home 24-hour urine collections will be provided. Urine will be assayed for oxalate and citrate as well as other kidney stone risk factor chemistries including urine volume, calcium, pH, uric acid, sodium, potassium, chloride, magnesium, phosphorus, urea, sulfate, ammonium, and creatinine. Supersaturation of calcium oxalate, calcium phosphate, and uric acid will be calculated using EQUIL2.43,44 These additional assays will provide data for future study. Serum sodium, potassium, chloride, bicarbonate, calcium, phosphorus, magnesium, creatinine, and calcium, citrate, and oxalate will be measured. All urine and serum measurements will be performed at the UCM Kidney Stone Laboratory except plasma oxalate which will be measured at University of Alabama-Birmingham (UAB). Serum and urine will be frozen at -80° C and stored for potential later assays.

Clinical variable collection: Clinical variables will include age, sex, height, weight, body mass index, chronic medical conditions, and medications. Clinical variables will be collected from the electronic health record and verified in interview by me or the research assistant.

Statistical Analysis: Change in urine oxalate, urine citrate, and fractional excretions of citrate and oxalate from baseline (time 1) to each time point will be calculated for each participant. Descriptive statistics and t-tests will be used to demonstrate the change in urine citrate and fractional excretion of citrate by change in oxalate at each time period for all participants and between SF and NSF. The levels of urine oxalate and citrate and fractional excretions of citrate and oxalate will be graphed over time. Longitudinal methods will be used to perform regression modeling of the association of urine oxalate by urine citrate for all patients and stratified by SF status. Variables to be considered for multivariate analysis include clinical (sex, body mass index) and urinary (GI anion) variables.

Sample Size and Statistical Power: Sample size is based on preliminary data and power to detect a difference between SF and NSF participants for mean change in citrate (mmol)/creatinine per oxalate (mmol)/creatinine (mmol). Preliminary data shows that the change in citrate (mmol)/creatinine per oxalate (mmol)/creatinine (mmol) was 3.0 (SD 1.5) for NSF and 0.3 (SD 0.1) for SF participants in multivariate models. Setting alpha=0.05 and power 80%, see the Table 2 for the mean difference that can be detected in citrate (mmol)/creatinine (mmol) per 1 mmol oxalate/creation (mmol) between SF and NSF. We will enroll 12 SF patients and 12 NSF controls (total=24).

Eligibility Criteria:
Inclusion Criteria:

* Stone formers (SF):

  * Age 18-70
  * History of at least one calcium-based kidney stone
* Non-stone formers (NSF):

  * Age 18-70
  * No history of kidney stone
  * 24-hour urine oxalate within lab normal (\<50mg/day)

Exclusion Criteria:

* History of primarily uric acid, cysteine, or struvite stones
* History of severe acid-base abnormality, very low (less than 100mg/day) or very high (greater than 1500mg/day) urine citrate
* Any controls or SF participants who cannot stop diuretic medication or alkali supplementation for the course of the study period
* Chronic kidney disease (glomerular filtration rate \<75 ml/min/1.73m2.
* NSF and SF with extreme levels of urine citrate will be excluded from this small initial study to reduce heterogeneity and remove focus from the extremes of citrate levels.