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To evaluate the occurrence and define the aetiology of osteopenia in children receiving highly active antiretroviral therapy (HAART).Bone mineral density (BMD) of total body and lumbar spine (L2–L4) was assessed by dual-energy X-ray absorptiometry in 40 children vertically infected with HIV: 35 taking HAART and five naive to any antiretroviral treatment (untreated). Six HAART-treated children showed clinical evidence of lipodystrophy. N-terminal propeptide of type-I procollagen (PINP), bone-specific alkaline phosphatase (BALP) and N-terminal telopeptide of type I collagen (NTx) were measured. Results were compared with those obtained in 314 healthy controls. Differences between HIV-positive and healthy children and within the HIV-positive group were assessed by multivariate analyses, controlling for confounding variables (age, sex, weight and height).HAART-treated children showed lower spine BMD values than untreated (P = 0.045) and healthy (P = 0.004) children and lower total body BMD values than untreated (P = 0.012) and healthy (P < 0.0001) children. Spine and total body BMD were similar between untreated and healthy children. Total body BMD was lower (P < 0.005) in HAART-treated children with lipodystrophy than in untreated patients, while children on HAART but without lipodystrophy had intermediate values. BALP, PINP and NTx were similar among untreated and healthy children. HAART-treated children had higher BALP levels than healthy (P = 0.0007) and untreated (P = 0.045) children. PINP values showed the same trend as BALP. HAART-treated children had higher NTx urine levels than healthy (P < 0.0001) and untreated (P = 0.041) children.HAART seems a new risk factor for life-long osteoporosis in children. An increased rate of bone turnover causes BMD decrease. Severity of osteopenia seems to be related to lipodystrophy.