Natriuretic Effect of TwoWeeks ofDapagliflozin Treatment inPatientsWith Type 2 Diabetes andPreservedKidneyFunctionDuringStandardized Sodium Intake:Results of the DAPASALT TrialDiabetes Care 2021;44:440–447 | https://doi.org/10.2337/dc20-2604
OBJECTIVE
Sodium–glucose cotransporter 2 (SGLT2) inhibitors reduce the risk for heart failurehospitalization potentially by inducing sodium excretion, osmotic diuresis, andplasmavolumecontraction. Fewstudieshave investigated thishypothesis, butnonehave assessed cumulative sodium excretion with SGLT2 inhibition during stan-dardized sodium intake in patients with type 2 diabetes.
RESEARCH DESIGN AND METHODS
TheDAPASALT trialwasamechanistic, nonrandomized,open-label study inpatientswith type 2 diabetes with preserved kidney function on a controlled standardizedsodium diet (150 mmol/day). It evaluated the effects of dapagliflozin on sodiumexcretion, 24-h bloodpressure, and extracellular, intracellular, and plasma volumesat the start of treatment (ST) (days 2–4), end of treatment (ET) (days 12–14), andfollow-up (FU) (days 15–18).
RESULTS
Fourteenpatientswere included in the efficacy analysis.Mean (SD)baseline sodiumexcretion (150 [32] mmol/24-h) did not significantly change during treatment(change at ST: 27.0 mmol/24-h [95% CI222.4, 8.4]; change at ET: 2.1 mmol/24-h[228.8, 33.0]). Mean baseline 24-h systolic blood pressure was 128 (10) mmHg andsignificantly reducedatST (26.1mmHg [29.1,23.1];P<0.001)andET (27.2mmHg[210.0,24.3]; P < 0.001). Dapagliflozin did not significantly alter plasma volume orintracellular volume,whileextracellular volumechangedatST (20.7 L [21.3,20.1];P5 0.02). As expected, 24-h urinary glucose excretion significantly increasedduringdapagliflozin treatment and reversed during FU.
CONCLUSIONS
During standardized sodium intake, dapagliflozin reduced blood pressure withoutclear changes in urinary sodium excretion, suggesting that factors other thannatriuresis and volume changes may contribute to the blood pressure–loweringeffects.
1Amsterdam Diabetes Center, Department ofInternal Medicine, Academic Medical Center,VU University Medical Center, Amsterdam, theNetherlands2BioPharmaceuticals R&D, AstraZeneca, Gothen-burg, Sweden3BioPharmaceuticals R&D, AstraZeneca, Gaithers-burg, MD4Department of Clinical Pharmacy and Pharma-cology, University of Groningen, University Med-icalCenterGroningen,Groningen, theNetherlands
Data from recent trials have shown thatsodium–glucose cotransporter 2 (SGLT2)inhibitors, such as dapagliflozin, improvecardiovascular (CV) and kidney outcomes,including heart failure hospitalizationsand progression of kidney disease, inpatients with and without type 2 di-abetes and at different stages of chronickidney disease (1–5). The underlyingmechanisms for these CV benefits havebeen studied extensively but remain in-completely understood. A main hypoth-esis is that SGLT2 inhibitors confer CVprotection through hemodynamic actions,including volume contraction secondaryto glycosuria and natriuresis, leading toreduced cardiac pre- and afterload (6,7).Previous studies of varying designs
in healthy volunteers or patients withtype 2 diabetes have indeed suggestedthat SGLT2 inhibitors cause transientincreases in 24-h sodium excretion (8–11)along with reductions in plasma andextracellular volume (12,13). Studies inpatients with heart failure showed eitherno change (14) or a modest increase insodium excretion in parallel with a re-duction in plasma volume (15). The priorstudies were limited by the fact thatsodium excretion was estimated fromspot urine samples instead of 24-h urinecollections, or dietary intake of sodiumwas not recorded or standardized. Todate, no studies have formally assessedcumulative sodium excretion of SGLT2inhibitors together with changes in plasmavolume and extracellular volume in pa-tients with type 2 diabetes during con-trolled sodium intake.Therefore, the aim of this open-label,
mechanistic study was to assess the ef-fect of dapagliflozinonnatriuresis, plasmavolume, extracellular and intracellularvolume, and 24-h blood pressure regu-lation in patients with type 2 diabetesand preserved kidney function duringstrictly controlled sodium intake. Wehypothesized that SGLT2 inhibition in-creases urinary sodium excretion andosmotic diuresis and thereby affects bothvolume status and blood pressure.
RESEARCH DESIGN AND METHODS
This phase IV, multicenter, open-label,mechanistic interventional study wasconducted between July 2017 andMarch2020. Originally, the study had threestrata consisting of patients with type2 diabetes and impaired kidney func-tion (stratum 1), type 2 diabetes and
preserved kidney function (stratum 2),and no diabetes but impaired kidneyfunction (stratum3), with a total samplesize of 51 patients (17 patients perstratum). Here, we present data on thecompleted stratum of patients with type2 diabetes and preserved kidney function(stratum 2) who were recruited at theAmsterdam University Medical Center(AUMC), location VU University MedicalCenter (VUMC), and at ZiekenhuisgroepTwente, Almelo, the Netherlands.
Study PopulationPatients were recruited from study data-bases and by advertisements in localnewspapers. Eligible patients were Cau-casian, Asian, or Middle Eastern malesand females of no child-bearing potentialand aged 18 to #80 years. Eligible pa-tients had a diagnosis of type 2 diabeteswith glycated hemoglobin (HbA1c) rang-ing from 6.5% (48 mmol/mol) to ,10%(,86 mmol/mol) and were treated witha stable dose of metformin, sulfonylurea(SU), or a combination of metformin andSU as standard of care for at least 3months before enrollment. Patients hadto have a preserved kidney functiondefined as an estimated glomerular filtra-tion rate between.90 and#130mL/min/1.73 m2 for patients aged #59 years, be-tween .85 and #130 mL/min/1.73 m2
for patients aged 60–69 years, and be-tween .75 and #130 mL/min/1.73 m2
for patients aged$70 years. In addition,a stable dose of an angiotensin receptorblocker (ARB) for at least 6 weeks wasrequired to create a homogeneous co-hort using a similar class of agents toinhibit the renin-angiotensin-aldoste-rone system. For inclusion, patients hadto have a stable 24-h urinary sodiumexcretion on 2 successive days (,20%difference between days 23 and 22).Exclusion criteria were a history of un-stable or rapidly progressing kidney dis-ease, albumin-to-creatinine ratio .1,000mg/g, symptoms of urinary retention,use of a pacemaker or other implantedelectronic devices, type 1diabetes, bloodpressure $180/110 mmHg, and CV/vascular disease within 3 months beforescreening. Use of any glucose-loweringdrugs (except metformin and SU), ACEinhibitors, or nonsteroidal anti-inflam-matory drugs was not allowed duringthe study; treatment with diuretics dur-ing the study or within 2 weeks beforethe study also was not allowed. Written
informed consent was obtained from allpatients before any trial-related activ-ities. The study protocol, protocol amend-ments, and all other protocol-specificdocumentswere reviewed and approvedby local authorities and the medicalethical review boards of the participat-ing centers. The study complied with theDeclaration of Helsinki and Good ClinicalPractice guidelines.
InterventionEligible patients received dapagliflozin10-mg tablets once daily for 146 1 days.Patients were instructed to take theirstudy medication in the morning duringthe treatment period.
Outcome MeasuresThe overarching objective was to assessthe effect of dapagliflozin on change in24-h sodium excretion. The primary endpoint of change in 24-h sodium excretionwas defined as the change in averagesodium excretion at baseline (average ofdays 23 to 21) relative to the averagesodium excretion at start of treatment(ST) (average of days 2–4) on the basis ofearlier clinical studies (10). Secondaryobjectives included the effect of dapagli-flozin on changes in 24-h sodium excre-tion from baseline to end of treatment(ET) (days12–14)and fromETto follow-up(FU) (days 15–17). Additional secondaryend points of 24-h glucose excretion,systolic blood pressure (SBP), plasmavolume, and extracellular volume weremeasured at baseline, ST, ET, and FU.Change in intracellular volume, bodyweight, 24-h urinary aldosterone, and N-terminal pro-B-type natriuretic peptide(NT-proBNP) were exploratory end pointsmeasured at the same time points.Change from baseline in fractional ex-cretion of endogenous lithium, a surro-gate for sodium reabsorption in theproximal tubule, was a post hoc explor-atory end point.
Procedures and FU Visits
Run-in Period (Day 26 to Day 21)
Patients received food boxes (deliveredby Sodexo, Rotterdam, the Netherlands)with a daily sodium content of 150mmol,starting on day 26, and were requiredto adhere to the dietary requirementsthroughout the period of the study up toand including day 18 (Supplementary Fig.1). Food questionnaires were required tobe completedby theparticipants starting
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from day 26 to record any deviation inintake from the provided food boxes andliquid intake. Patients were allowed toconsume nonstudy food products, butthese were restricted to products thatdid not contain sodium. These foodproducts were also recorded in the foodquestionnaires. Participants were giventhe following oral and written instruc-tions about how to collect 24-h urinesamples: refrain from strenuous exerciseduring thecollectionperiods, collect 24-hurine in dedicated containers, and storecollections in a refrigerator until deliveryto the clinic. Twenty-four-hour urine sam-ples were collected on day23 to day21(baseline). Patients in whom 24-h sodiumexcretion did not differ by.20% betweenday23 to day22 were considered to beadherent to the dietary requirementsand could proceed to the active treat-ment period. On day 21, 24-h ambula-tory blood pressure monitoring (ABPM)(iCardiacTechnologies, Pittsford,NY)wasinitiated.
Treatment Period (Day 1 to Day 14)
In-patient study visits were scheduled atdays 1, 4, 5, 13, and 14. In total, fourconsecutive 24-h urine samples werecollected at days 1–4 followed by threeconsecutive 24-h urine collections atdays 12–14. Ondays 4 and 13, 24-h ABPMmeasurements were initiated. Bloodsamples were obtained in the fastingconditionon themorningofdays1,4, and14. Plasma volume assessment and bio-impedance spectroscopy (BIS) (Imped-iMed Limited, Pinkenba, Queensland,Australia) for assessment of extracellularvolume and intracellular volume wereperformed on days 1, 4, and 14. Plasmavolume was measured using the indoc-yanine green (ICG) (Verdye; DiagnosticGreen, Ascheim-Dornach, Germany) in-dicator dilution method. ICG is a water-soluble dye that binds to plasma proteins(mainly albumin) and is a marker of theplasma volume distribution space. Afteran intravenous bolus injection of ICG0.25mg/kg over a period of 1min, bloodsamples were obtained every 30 s from2 to 5 min as described previously (16,17).
FU Period (Day 15 to Day 19)
Three consecutive 24-h urine sampleswere collected at days 15–17 (FU). A finalin-patient study visit was scheduled atday 18 during which plasma volume andBIS assessments were performed. At thefinal in-patient study visit, blood samples
were obtained in the fasting condition,and 24-h ABPM was initiated.
Laboratory MeasurementsAll samples were measured by standardin-house assays at COVANCE (Geneva,Switzerland), MLMMedical Laboratories(Monchengladbach, Germany) (ICG), Uni-versity Medical Center Groningen (urineand serum lithium by a validated induc-tively coupled plasmamass spectrometryassay), and AUMC and ZiekenhuisgroepTwente (HbA1c, creatinine, urinary so-dium, and glucose).
Statistical AnalysisSample size calculations were performedfor each of the three strata individually.With a sample size of 15 patients andassuming an SD of 25 mmol/24 h inchange from average baseline in 24-hsodium excretion, the study had 80%power to detect an increase in 24-hsodium excretion of at least 20 mmol/24 h from baseline with dapagliflozin at atwo-sided a-level of 0.05. Under theseconventions, the minimum detectabledifference in change from average base-line in 24-h sodium excretion is ;13.6mmol/24 h. To account for early discon-tinuationbecauseofthecomplexprotocoland high demand on study participants,we enrolled 17 patients. Baseline charac-teristics were summarized using meanand SD or proportion. A longitudinal re-peated-measures analysis was used forthe change versus the baseline value. Themodel included a fixed effect of timepoint, interaction term between timepoint and baseline, and continuous base-line value as covariates. An unstructuredcovariance matrix structure was used tomodel correlations among the repeatedmeasurements. Point estimates and cor-responding 95% CIs for the least squaresmeans at each time point were derived.P , 0.05 was considered to indicatestatistical significance. Analyseswere per-formed using SAS 9.4 statistical software.
RESULTS
Participant CharacteristicsThirty-one patients were enrolled inthe study, of whom 17 started treat-mentwith dapagliflozin. Before databaselock, one patient was excluded fromanalysis because of a .20% differencein urinary sodium excretion betweendays 23 and 22 and another becauseof missing urine volume for 24-h urine
collection at day22. After database lock,one additional patient was excluded be-cause of nonadherence to dapagliflozin.Efficacy analyses were therefore per-formed in 14 patients, and their baselinecharacteristics are reported in Table 1.Efficacy data for the 15 patients includedbefore database lock are reported inSupplementary Table 1. Safety was as-sessed in the 17 patients who startedtreatment.
Effect of Dapagliflozin on UrinaryExcretion of Sodium, Glucose, Volume,and LithiumMean (SD) 24-h sodium excretion atbaseline was 150 (32) mmol/24-h, con-firming successful design and adherenceto the diet. Dapagliflozin treatment didnot significantly change 24-h sodiumexcretion (urinary 24-h sodium change atST:27.0 mmol/24-h [95% CI222.4, 8.4;P 5 0.34]; change at ET: 2.1 mmol/24-h[228.8, 33.0; P 5 0.89]) (Fig. 1A). Com-paredwith ET, 24-h sodium excretion didnot changeduring FU (changeatdays 15–17: 213.8 mmol/24-h [230.9, 3.2]; P 50.10) (Fig. 1A).Mean24-hurinary sodiumexcretion from day 21 to day 1 tendedtoward an increase (27mmol/24-h [210,63]) (Supplementary Table 2).
Mean (SD) baseline urinary glucoseexcretionwas1.9 (2.2)mmol/24-h,which,as expected, significantly increasedduringdapagliflozin treatment (urinary glucosechange at ST: 351mmol/24-h [95%CI273,428;P,0.0001];changeatET:322mmol/24-h [231, 413; P , 0.0001]) (Fig. 1B).Mean 24-h glucose excretion signifi-cantly decreased at FU comparedwith ET(change at days 15–17:2208mmol/24-h[2244, 2172]; P , 0.0001) (Fig. 1B).
At baseline,mean (SD) 24-hurine volumewas 2,285 (587) mL/24-h, which did notstatistically significantly change duringtreatment with dapagliflozin (urinary vol-umechangeatST:52mL/24-h[95%CI2234,338; P 5 0.70]; change at ET: 80 mL/24-h[2217, 378; P5 0.57]) (Fig. 1C). During FU,urine volume also did not change comparedwith ET (change at days 15–17: 2165 mL/24-h [2363, 33]; P 5 0.10) (Fig. 1C).
Mean (SD) baseline fractional lithiumexcretion was 18.1% (5.3%), which sig-nificantly increased during dapagliflozintreatment (urinary fractional lithiumchangeat ST: 5.0% [95% CI 1.3, 8.6; P 5 0.013];change at ET: 4.6% [1.2, 8.0; P5 0.013])(Fig. 1D). During FU, fractional lithiumexcretion returned in the direction of
442 Effects of Dapagliflozin on Natriuresis Diabetes Care Volume 44, February 2021
baseline (change at FU: 21.8% [25.0,1.4]; P 5 0.25) (Fig. 1D).
Plasma Volume, Extracellular Volume,and Intracellular VolumeMean (SD) plasma volume at baselinewas 3.75 (0.8) L, remained stable at ST(mean change 20.19 L [95% CI 20.9,0.5]; P5 0.58) (Fig. 2A) and tended to belower at ET (mean change20.45 L [21.1,0.2];P50.16).DuringFU,plasmavolumesignificantly increased compared withET, and the magnitude of the effect wasidentical to the decrease observed frombaseline to ET (change at day 18: 0.43 L[0.0, 0.8]; P 5 0.049) (Fig. 2A).At baseline, mean (SD) extracellular
volume was 20.9 (4.5) L, which was sig-nificantly reduced after ST but not at ET(changeatday4:20.7L[95%CI21.3,20.1;P 5 0.02]; change at day 14: 20.1 L[20.5, 0.4; P 5 0.73]) (Fig. 2B). Meanextracellular volume during FU did notchange compared with ET (change at day18: 0.2 L [20.1, 0.5]; P5 0.21) (Fig. 2B).Mean (SD) intracellular volumeatbase-
line was 26.6 (5.7) L, which did not sig-nificantly change during treatment withdapagliflozin (mean change at ST:20.3 L[95% CI 21.0, 0.4; P 5 0.36]; meanchange at ET: 0.04 L [20.7, 0.7;P50.91])(Fig. 2C). Change in intracellular volumeat FU compared with ET was 20.3 L(20.7, 20.0; P 5 0.08) (Fig. 2C).
Effect of Dapagliflozin on 24-h BloodPressure ProfilesMean (SD) 24-h SBP at baseline was 128(10) mmHg, which significantly changedduring dapagliflozin treatment (meanchange at ST: 26.1 mmHg [95% CI 29.1,23.1; P50.001];mean change at ET:27.2mmHg [210.1, 24.3; P , 0.001]) andremained reduced during FU comparedwith ET (0.7mmHg [22.0, 3.4];P50.56)(Fig. 2D). Treatment with dapagliflozinalso reduced 24-h diastolic blood pres-sure (DBP) (Fig. 2E).
Anthropometrics and HormonesMean (SD) body weight at baseline was98.7 (15.9) kg, which was significantlyreduced during dapagliflozin treatment(mean change at ST: 20.8 kg [95% CI21.2,20.5; P, 0.001]; mean change atET: 21.8 kg [22.5, 21.1; P , 0.001]).During FU, body weight returned in thedirection of baseline (change from ET toend of FU: 0.4 kg [0.0, 0.9]; P 5 0.07)(Table 2).Mean 24-h urinary aldosteroneexcretion was 14.0 (8.4) mg/24-h at base-line, which was significantly increased atST (mean change at day 4: 3.6 mg/24-h[2.2, 5.1]; P, 0.001), while no differenceswere observed at ET (mean change atday14: 0.8mg/24-h [22.4, 3.9];P50.60)(Table 2). Mean NT-proBNP at baselinewas 3.5 (2.6) pmol/L and did not changeduring dapagliflozin treatment (change
at ST:20.4 pmol/L [22.0, 1.2; P5 0.62];change at ET: 1.0 pmol/L [21.2, 3.3; P50.32]) (Table 2). BNP and hematocrit alsodid not changeduring dapagliflozin treat-ment (Table 2).
Adverse EventsDapagliflozin was generally well toler-ated. There were no serious adverseevents, and no participants discontin-ued dapagliflozin after treatment com-menced (Supplementary Table 3).
CONCLUSIONS
Thefindings fromtheDAPASALT (AnOpenLabel, Phase IV, Mechanistic, Three-ArmStudy to Evaluate the Natriuretic Effect of2-Week Dapagliflozin treatment in Type 2Diabetes Mellitus Patients with EitherPreserved or Impaired Renal Functionand Non-Diabetics with Impaired RenalFunction) trial show that during standard-ized conditions, including a strictly stan-dardized sodium intakewithoverall goodcompliance with the diet and stable useof ARBs, 24-h sodium excretion did notchange in diuretic-naive patients withtype 2 diabetes and preserved kidneyfunction.Despite similar sodiumbalance,SBP acutely decreased, which persistedduring the 2-week treatment period.Extracellular volume was reduced after4 days of treatment, but this initial re-duction dissipated after 14 days of treat-ment. Taken together, thesedata suggestthat the blood pressure–lowering effectduring treatment with SGLT2 inhibitorsmay be sodium excretion independent.
SGLT2 inhibitors exert their effect onthe proximal tubule of the kidney byblocking glucose and sodium reabsorp-tion (18), resulting in glucosuria and anincrease in sodium and chloride deliveryat the juxtaglomerular apparatus. Indeed,we observed an increase in fractionallithium excretion, a proxy for reducedsodium reabsorption at the proximaltubule, during dapagliflozin treatment.Enhanced natriuresis could be expectedas a consequence of inhibition of prox-imal sodium reabsorption. The lack ofincreased natriuresis in our trial suggeststhat compensatory mechanisms in thedistal tubule may be involved to com-pensate for the proximal decrease insodium reabsorption. Increased activityof the renin-angiotensin-aldosterone systemand, consequently, aldosterone-regulatedincreased sodium reabsorption throughepithelial sodium channels might contribute
Table 1—Baseline characteristics
Characteristic Study participants (n 5 14)
Age (years) 63.9 (7.9)
Male sex, n (%) 9 (64.3)
Race, n (%)White 13 (92.9)Asian 1 (7.1)
Diabetes duration (years) 10.2 (5.2)
Body weight (kg) 98.7 (15.9)
BMI (kg/m2) 31.9 (4.2)
Fasting plasma glucose (mmol/L) 8.1 (1.4)
HbA1c (%) 7.2 (0.6)
HbA1c (mmol/mol) 55 (6.6)
SBP (mmHg) 128.6 (13.6)
DBP (mmHg) 74.7 (7.5)
eGFR (CKD-EPI) (mL/min/1.73 m2) 94.3 (10.9)
UACR (mg/mmol), median (25th–75th percentile) 0.8 (0.5–2.8)
Hemoglobin (g/L) 137.9 (13.0)
Hematocrit (L/L) 0.4 (0.0)
Metformin, n (%) 14 (100)
SU derivative, n (%) 5 (35.7)
Data are mean (SD) unless otherwise indicated. Blood pressure recorded in supine position. CKD-EPI, Chronic Kidney Disease Epidemiology Collaboration; eGFR, estimated glomerular filtrationrate; UACR, urinary albumin-to-creatinine ratio.
care.diabetesjournals.org Scholtes and Associates 443
to this, as reflected by the increase inurinary aldosteroneexcretionat STdespiteall patients using a stable dose of ARBs.After drug discontinuation, we observed atrend toward sodium and volume reten-tion. This might reflect that activation ofcompensatory mechanisms in the distaltubule may take .4 days to resolve.In contrast to our findings, clinical
studies in patients with type 2 diabetesreported that SGLT2 inhibitors induce tran-sient increases in urine volume (8,9,19,20)and natriuresis (8–11). Indeed, a model-based prediction of the DAPASALT studyalso suggested that a transient increasein natriuresis can be expected at ST (21).However, few experimental and clinicalstudieswereperformedduring standard-ized sodium intake, which is essentialto properly assess natriuretic effects. Astudy in healthy volunteers receiving astandardized sodium diet and sodiumtablets aimed at daily sodium intake of110mmol reportedmodest natriuresis in
the first 6 h following dapagliflozin ad-ministration. However, no baseline as-sessment was performed, and therefore,actual 6-h sodium excretion was deter-minedbutnochanges frombaselinecouldbe determined. In addition, poor tol-erance to the sodium tablets was ob-served. Both limitations hamper theinterpretation of the results (10). An-other study in patients with type 2 di-abetes and heart failure reported asimilar acute increase in urinary sodiumexcretion over 6 h, but sodium intakewas not standardized, fluid administra-tion was high, and 24-h urine sampleswere not collected (15). The currentstudy investigated the effect of dapagli-flozin on cumulative urinary sodium ex-cretion in well-controlled patients withtype 2 diabetes and preserved kidneyfunction following a strictly controlledsodium diet. The primary end point waschosenon thebasis of earlier studies thatsuggested that changes in natriuresis
would occur 2–4 days following dapagli-flozin administration (10,15). Acute effectsduring the first hours after dapagliflozinadministration were not assessed, and it ispossible thatamodest transient increase insodium excretion was present but notdetected. We observed a modest increasein 24-h urinary sodium excretion at day 1.Whether this increase is real or can beattributed to biological variation in sodiumexcretion, which has been reported pre-viously in a well-controlled salt balancestudy, is difficult to ascertain (22). Never-theless, evenwhen theacute and transientincrease in natriuresis reflects a genuineeffect, it is questionable whether themag-nitude is sufficient for the substantial andpersistent reduction in blood pressure.
Under healthy circumstances, up to180 g/day of glucose is filtered by thekidney glomerulus, and virtually all of itis subsequently reabsorbed in the prox-imal convoluted tubule predominantlyby SGLT2 (23). Treatment with SGLT2
444 Effects of Dapagliflozin on Natriuresis Diabetes Care Volume 44, February 2021
inhibitors increases glucose concentra-tion in the distal nephron, resulting ina decrease in the osmotic gradient be-tween the tubular fluid and interstitium.This reduces passive water reabsorption,resulting in osmotic diuresis (13,14). Inthe current study, glycosuria was con-sistently increased, although we did notobserve a clear effect on urine volume,which contrasts thefindings from in silicomodels (21). A previous study showedthat dapagliflozin caused a larger in-crease in urine compared with plasmaosmolality, making it less likely that
osmotic diuresis explains the observedhemodynamic effects in our study (24).Given that glycosuria is persistent duringtreatment with dapagliflozin while theactual fluid balance of the participantswas negative, the kidney likely uses othermechanisms to concentrate the urine andenhance water reabsorption. Antidiuretichormone–regulated renal water conser-vation could possibly be involved. Toconserve water, antidiuretic hormoneindirectly increases renal urea absorp-tion, thereby increasing urea-driven urineconcentration, because urea is the most
efficient organic osmolyte to concentrateurine and prevent dehydration (25–27).Preclinical studies have shown that da-pagliflozin increases levels of the ureatransporter UT-A1 in rats, supporting thishypothesis (28).
If the observed reductions in systemicblood pressure in this study are not(osmotic) diuretic related, other effectsthat could contribute include reductionsin arterial stiffness, inhibition of thesympathetic nervous system (29–31), orrestoration of endothelial function. In-deed, in a previous study, dapagliflozin
Figure 2—Changes in plasma volume (A), extracellular volume (B), intracellular volume (C), SBP (D), and DBP (E) frombaseline (BL) to ST, fromBL to ET,and from ET to FU.
Table 2—Changes in anthropometrics, hormones, and hematocrit
Baseline value Change at ST* Change at ET* Change at FU†
Parameter Mean (SD) Mean (95% CI) P value Mean (95% CI) P value Mean (95% CI) P value
reduced pulse wave velocity 2 days afterinitiation, a time course of effect similarto our study (32,33). Another compart-ment that could influence extracellularvolume and blood pressure is the endo-thelial surface layer or glycocalyx (34,35).Damage to theendothelial glycocalyx hasbeen observed in patients with type 2diabetes, and SGLT2 inhibition has beenshown to restore the structural integrityof the glycocalyx in vitro and in vivo(35–37).In the current study, plasma volume
did not statistically significantly changeduring treatment with dapagliflozin inpatients who had no heart failure orvolume overload at baseline, althoughthemagnitudeofeffect inour small studywas consistent with the reduction inplasma volume observed in a prior studywith dapagliflozin (12). In addition, dur-ing FU, plasma volume significantly in-creased compared with ET, which couldreflect a real effect of dapagliflozin onplasmavolume. Extracellular volumewasacutely reducedafter4days,whilewedidnot observe a clear effect on intracellularvolume. Because extracellular volume isthe sumofplasmavolumeand interstitialvolume, this may suggest that althoughnot directly measured, a proportionallylarger decrease in interstitial volumeoccurred, which is consistent with theresults suggested by Hallow et al. (13) ina modeling study of SGLT2 inhibitoreffects. Importantly, heart failure is char-acterized by interstitial fluid accumula-tion and sodium retention, leading toperipheral and pulmonary edema, whichmay lead to differential effects of dapa-gliflozin on natriuresis and plasma vol-ume in this population. The ability toselectively reduce interstitial fluid maybe a unique feature of SGLT2 inhibitorscompared with diuretics and has beensuggested in amodeling study comparingdapagliflozin with the loop diuretic bu-metanide. Although both drugs wereassociated with a reduction in interstitialfluid, dapagliflozin induced little or nochange in blood volume, whereas bume-tanide was associated with greater re-ductions in intravascular volume (6,13).However, so far, no clinical data haveconfirmed this notion. A differential ef-fect in regulating interstitial fluid may beparticularly important in patients withheart failure inwhom, inmany instances,arterial intravascular contraction is pres-ent and often provoked by diuresis.
However, the fact that the effect ofextracellular fluid contraction dissipatedduring prolonged treatment in our studywould suggest a physiological adaptiveresponse of the kidney to maintain bodyfluid volume, at least in patients withtype 2 diabetes and preserved kidneyfunction. Whether our findings can beextrapolated to patients with heart fail-ure requires further study.
We acknowledge some limitations. Incontrast to other studies with SGLT2inhibitors,wedidnotobservean increasein hematocrit. The lack of effect onhematocrit may be explained by themultiple blood samplings performeddur-ing the relatively short study. To studythe effect of dapagliflozin on natriuresisand systemic hemodynamics, we delib-erately performed this study in a homo-geneous cohort of patients with type 2diabetes and preserved kidney functionwho were all using an ARB withoutconcomitant diuretic treatment, therebyavoiding disease heterogeneity and po-tential confoundingbypreexistingkidneydamage. The carefully selected cohortmay limit the generalizability of ourfindings. In addition, extracellular andintracellular volumes were assessed byBIS, which is not the gold standardmethod to assess body composition.Nevertheless extracellular and intracel-lular volume assessed by BIS correlateswell with the reference bromide method(38). In addition, although the study wassufficiently powered for the primary out-come, the size of our study cohort wassmall, limiting the precision of the effectestimates for some of the secondaryend points. Finally, the open-label de-sign does not allow definitive conclu-sions. Our results should be consideredhypothesis generating.
In conclusion, we have demonstratedthat treatment with dapagliflozin lowersblood pressure without clear effects onsodium excretion in patients with type 2diabetes and preserved kidney function.Together with the absence of prolongedchanges in plasma volume, extracellularvolume, and intracellular volume, thissuggests that other nondiuretic-relatedeffects cannot be ruled out in the CVprotective effects conferred by SGLT2inhibitors.
Acknowledgments. The authors are extremelygrateful to the participants who volunteered in
the study. The authors acknowledge the help ofdietician Esther Pekel for developing the foodmenus and the following study nurses and assis-tantswhowere indispensable in theprocessofdatacollection: Jeannette Boerop, Ingrid Knufman, andRenee de Meijer (Diabetes Center, Departmentof Internal Medicine, AUMC, location VUMC).The authors also thank the study teammembersat AstraZeneca. Finally, the authors thank ParitaSheth (inScience Communications, London, U.K.)for assistance with figure preparation and edit-ing; this support was funded by AstraZeneca.Funding and Duality of Interest. The DAPA-SALT studywas funded by AstraZeneca.M.H.A.M.hasactedasaspeaker/consultant forAstraZeneca,Eli Lilly, Novo Nordisk, and Sanofi; all honorariaare paid to his employer (AUMC, locationVUMC). P.J.G., C.K., A.H., and N.A. are employeesand shareholders at AstraZeneca. D.H.v.R. hasacted as a consultant and received honorariafromBoehringer Ingelheim, Eli Lilly,Merck, NovoNordisk, Sanofi, and AstraZeneca and has re-ceived researchoperating funds fromBoehringerIngelheim-Lilly Diabetes Alliance, AstraZeneca,and Novo Nordisk; all honoraria are paid to hisemployer (AUMC, location VUMC). H.J.L.H. isconsultant for AbbVie, AstraZeneca, Bayer, Boeh-ringer Ingelheim, Chinook, CSL Pharma, GileadSciences, Janssen, Merck, Mundipharma, Mitsu-bishi Tanabe, Novo Nordisk, and Retrophin.He received research support from AbbVie,AstraZeneca, Boehringer Ingelheim, and Janssen.No other potential conflicts of interest relevantto this article were reported.Author Contributions. R.A.S., M.H.A.M.,M.J.B.v.B., A.H., D.H.v.R., and H.J.L.H. were in-volved in data collection. R.A.S., D.H.v.R., andH.J.L.H. wrote the drafts of the manuscript. P.J.G.,C.K., D.H.v.R., and H.J.L.H. designed the study. Allauthors were involved in the data analysis andinterpretation and participated in critical reviewof the manuscript drafts and approved the finalversion for submission.R.A.S., D.H.v.R., andH.J.L.H. are the guarantors of this work and, as such,had full access toall thedata in the studyand takeresponsibility for the integrity of the data and theaccuracy of the data analysis.Prior Presentation. Parts of this study werepresented at the European Society of Cardi-ology Congress, 2020–The Digital Experience,29 August–1 September, 2020.
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