Bone Mineral Density Decline According to Renal Tubular Dysfunction and Phosphaturia in Tenofovir-exposed HIV-infected Patients

José L. Casado; Carmen Santiuste; Monica Vazquez; Sara Bañón; Marta Rosillo; Ana Gomez; María J. Perez-Elías; Carmen Caballero; José M. Rey; Santiago Moreno


AIDS. 2016;30(9):1423-1431. 

In This Article


This is a longitudinal evaluation of HIV-infected patients included in an ongoing cohort study of renal and bone involvement. From July 2012, we included in this cohort HIV-infected patients who were older than 18 years, without a previous diagnosis of renal disease, and who were not receiving, nor expecting to receive, nephrotoxic drugs (including nonsteroidal anti-inflammatory drugs or anticancer chemotherapy), other drugs that could alter tubular secretion of TDF (diclofenac and sildenafil), or in treatment with drugs that alter bone mineralization (corticosteroids and bisphosphonates). To analyse the prevalence and evolution of BMD, we included in this subanalysis those HIV-infected patients initially receiving a cART regimen containing TDF, and who had at least two successive BMD measurements with the condition of not experiencing any change in cART.

The study was approved by our institutional review board (EC 248/12), and written informed consent was obtained from every participant. The study was registered at (NCT 02116751).


Demographic data (age, sex, and BMI), HIV-related data (risk practice, time of HIV infection, nadir and baseline CD4+ cell count, HIV-1 RNA level, and previous cART), and hepatitis C virus (HCV) coinfection status were collected for each patient at the time of inclusion. A previous diagnosis of hypertension or diabetes mellitus was recorded in all the cases. Routine laboratory tests, including serum creatinine, phosphate, and CD4+ cell counts and HIV-1 RNA levels (Versant HIV-1 RNA 1.0 –kPCR-, Siemens Diagnostics, Munich, Germany; quantification limit 37 copies/ml) were available every 3–4 months since cART initiation and were collected from the history (mean, 9 determinations). As part of the study, serum glucose, creatinine, phosphate, bone-specific alkaline phosphatase (BALP), 25 dihydroxyvitamin D (25OHD), PTH, and urinary protein, albumin, creatinine, uricosuria, phosphaturia, glycosuria, β-2-microglobulin (B2M), and retinol-binding protein (RBP) in the first urine of the morning after overnight fast, were performed before and at inclusion visit. Estimated glomerular filtration rate (eGFR) was calculated using the chronic kidney disease (CKD)-epi equation.[14]

BMD of spine and hip was measured consecutively at inclusion and during follow-up by dual X-ray absorptiometry (DXA) using the same Hologic densitometer (Hologic 4500; Bedford, Massachusetts, USA) after daily calibration with 'gold standard' reference spine and hip phantoms with anatomically correct contours to read BMD within 1%.

Serum 25OHD was determined by chemiluminescent microparticle immunoassay (Architect 25OHD; Abbott Diagnostics, Germany). Serum intact PTH was determined by electrochemiluminescence immunoassay (Cobas e411, Roche Diagnostics, Indianapolis, Indiana, USA). Urine total protein was measured by using benzethonium chloride method and urine albumin by immunoturbidimetric assay. Both were expressed as a ratio to creatinine in mg/g (uPCR, protein to creatinine ratio; uACR, albumin to creatinine ratio). Urine creatinine concentration was determined by an enzymatic method. Phosphaturia and uricosuria were determined as fractional excretion (FE) of phosphate and uric acid: (urine phosphate or uric acid × serum creatinine)/(urine creatinine × plasma phosphate or uric acid) × 100. Urinary concentrations of B2M and RBP were measured by immunonephelometry (BN II system, Siemens Healthcare Diagnostics, Germany). Concentration of both proteins was standardized to a urinary creatinine of 1 g/l (μg/g), and values were log transformed.


Vitamin D deficiency was defined as a value less than 20 ng/ml. Secondary hyperparathyroidism was defined as serum PTH more than 65 pg/ml, in the absence of hypercalcemia. Hypophosphataemia was defined as a serum phosphate 2.5 mg/dl or less.

Proteinuria was defined as uPCR more than 100 mg/dl (11.3 mg/mmol), macroalbuminuria as uACR more than 300 mg/g, and microalbuminuria as uACR 30–300 mg/g, respectively. Hyperphosphaturia of tubular origin was defined as fractional excretion of phosphate more than 20%, or more than 10% in the presence of hypophosphataemia. Hyperuricosuria was defined as a fractional excretion of uric acid at least 10%.[15] Glycosuria was defined as a value more than 100 mg/dl (5.56 mmol/l) in presence of normal glycaemia (plasma glucose <125 mg/dl). Tubular dysfunction was defined as the presence of at least two tubular abnormalities known to be part of Fanconi's syndrome: hyperphosphaturia, nondiabetic glycosuria, hyperuricosuria, or/and proteinuria. As both albuminuria and tubular proteinuria were expected to be included in proteinuria, and serum phosphate could be related to phosphaturia, they were not considered in the definition.

CKD was established if eGFR value was less than 60 ml/min per 1.73 m2 in two determinations (CKD stage 3 or higher).[16] According to BMD at the femoral neck and spine (L1–L4), the T scores were calculated as a standard deviation score compared with BMD at peak bone mass (age 30), and categorized according to the World Health Organization criteria as osteoporosis if less than -2.5 SD and osteopenia when the T score less than -1.0 SD.[17] The primary study outcome was the change in the value of BMD during TDF treatment, according to the presence of different tubular abnormalities. Time at risk began upon TDF initiation.

Statistical Analysis

Characteristics of patients according to the presence of reduced BMD were compared for statistical significance using the Mann–Whitney U-test for continuous variables and the χ2 test for categorical variables. Analysis of paired observations was performed using the Wilcoxon rank t-test. Correlations between the different tubular abnormalities and BMD values were assessed using Spearman correlation coefficient. A linear regression model was created utilizing BMD changes at the femoral neck during follow-up as the dependent variable, considered as numerical (g/m2), adjusted for potential confounders by using the backward stepwise method. Age, sex, and BMI were considered to be required covariates. All P-values were two-sided and a value of P < 0.05 was considered statistically significant.