Mitochondrial DNA content, an inaccurate biomarker of mitochondrial alteration in human immunodeficiency virus-related lipodystrophy.

Lipoatrophy is a prevalent side effect of antiretroviral treatment of human immunodeficiency virus (HIV) infection. Its mechanisms are still disputed but include mitochondrial toxicity and, in particular, mitochondrial DNA (mtDNA) depletion induced by nucleoside reverse transcriptase inhibitors. To obtain an integrated evaluation of the mitochondrial alteration in lipoatrophy, we investigated the DNA, RNA, and protein levels in 15 samples of abdominal subcutaneous adipose tissue from HIV-infected patients with peripheral lipoatrophy and compared the results with those for 15 samples from age- and body mass index-matched controls. The DNA and RNA analyses used PCR-based techniques, while proteins were quantified with enzyme-linked immunosorbent assay and measurement of activities with spectrophotometric assays. Depletion of mtDNA and mtDNA-encoded MT-CO2 mRNA was present, but normal levels of mtDNA-dependent activity (cytochrome c oxidase) and protein (MT-CO2p) showed that it was compensated for. An increase in nuclear-DNA-dependent mitochondrial activities (citrate synthase and malate dehydrogenase) and protein (COX4I1p), as well as transcriptional up-regulation of nuclear-DNA-encoded mitochondrial genes (COX4I1 and UCP2), demonstrated increased mitochondrial biogenesis. However, the expression of the known transcription factors of mitochondrial biogenesis (TFAM, NRF1, GABPA, PPARGC1A, PPARGC1B, and PPRC1) was normal or decreased. Increased amounts of activated caspase 3 and of DDIT3 mRNA showed the induction of apoptosis and oxidative stress, respectively. The mtDNA content did not correlate with any other mitochondrial parameter. In conclusion, mtDNA content does not appear to be an accurate biomarker of mitochondrial alteration in lipoatrophic adipose tissue. The preservation of mtDNA-dependent mitochondrial functions occurred despite severe mtDNA depletion. The presence of significant oxidative stress and apoptosis did not correlate with the mtDNA content.

Modified quantitative insulin sensitivity check index is better correlated to hyperinsulinemic glucose clamp than other fasting-based index of insulin sensitivity in different insulin-resistant states.

Fasting-based index estimates of insulin sensitivity were compared with euglycemic hyperinsulinemic clamp (IS clamp) measurements in 148 subjects: normal controls (n = 46), and obese (n = 12), polycystic ovary syndrome (n = 16), first-degree relatives of type 2 diabetic (n = 17), impaired glucose tolerance (n = 28), and type 2 diabetic (n = 29) patients. The fasting-based indexes tested included log homeostasis model assessment (HOMA), the quantitative insulin sensitivity check index (QUICKI), the revised QUICKI, and a new revised QUICKI using fasting plasma glycerol. In the population studied, at 40 mU/m(2).min (n = 30) revised QUICKI (r = 0.86; P < 0.0001) and QUICKI-glycerol (r = 0.87; P < 0.0001) gave higher correlations with the IS clamp than QUICKI and log HOMA (r = 0.78 and r = -0.78; P < 0.001). For subjects tested at 75 mU/m(2).min (n = 118), comparable correlations were found for all indexes (r > 0.80; P < 0.0001). When studied in subgroups, revised QUICKI and QUICKI-glycerol give significantly higher correlations with the IS clamp than other indexes for lean control subjects studied at 40mU/m(2).min and impaired glucose tolerance subjects. We confirmed, in a large patient population with a wide range of insulin sensitivities, that no single test is superior in all groups of patients. However, QUICKI and revised QUICKI are good indexes that offer correlations similar to or higher than values obtained with log HOMA. Such indexes are simple tools to estimate insulin sensitivity appropriate for epidemiological studies.

Altered fat differentiation and adipocytokine expression are inter-related and linked to morphological changes and insulin resistance in HIV-1-infected lipodystrophic patients.

OBJECTIVE: To achieve a better understand of the pathophysiology of HIV-related lipoatrophy, we compared the mRNA expression of adipocytokines in fat samples from patients and healthy HIV-seronegative controls together with fat morphology and we studied the relationship between changes in fat morphology, adipocytokine expression, markers of adipose tissue differentiation and whole body insulin sensitivity. DESIGN: Cross-sectional analytical study. SUBJECTS AND METHODS: The mRNA expression of adipocytokines and transcriptional factors in fat samples from 26 patients with peripheral lipoatrophy (all under anti-retroviral therapy associating protease inhibitor and nucleoside-analogue reverse transcriptase inhibitors) and from 16 non-HIV-infected controls was measured by real time quantitative RT-PCR. Fat morphology was assessed histologically on a subgroup of 10 patients and six controls: collagen fibres by Sirius Red staining, apoptosis by the TUNEL technique, vessels by smooth muscle alpha-actin staining and macrophages by CD68 staining. Insulin resistance was assessed by using the homeostasis model assessment. RESULTS: The patients' fat showed higher values of apoptosis (P=0.005), fibrosis (P<0.05), vessel density (P=0.001) and macrophage infiltration (P<0.05) than the controls' fat, together with lower adiponectin and leptin mRNA levels and higher interleukin (IL)-6 and tumour necrosis factor (TNF)alpha mRNA levels. TNFa and IL-6 expression correlated positively with the level of apoptosis (P=0.05 and P<0.05, respectively) and negatively with CCAAT-enhancer binding protein (C/EBP)alpha (P<0.001 and P<0.05, respectively). Apoptosis correlated negatively with the expression level of sterol-regulatory-element-binding-protein-1c (SREBP1c) (P=0.01) and C/EBPalpha (P=0.01) whilst the vessel density correlated negatively with SREBP1c (P<0.005), C/EBPalpha (P=0.001) and beta (P=0.001). Adiponectin and leptin expression correlated positively with each other, and also with adipogenic marker expression and overall insulin sensitivity. These relationships were also present when the patient group was studied separately. Finally, fat morphological abnormalities correlated positively with whole body insulin resistance. CONCLUSIONS: Adipose tissue from patients with HIV-1-related lipoatrophy shows increased apoptosis, together with decreased adipocyte differentiation. Increased TNFalpha and IL-6 expression could be a major phenomenon linking these alterations. Decreased adiponectin and leptin expression, which may result from decreased adipocyte differentiation, could be involved in the observed whole body insulin resistance.

Association between altered expression of adipogenic factor SREBP1 in lipoatrophic adipose tissue from HIV-1-infected patients and abnormal adipocyte differentiation and insulin resistance.

BACKGROUND: Lipodystrophy is a major side-effect of antiretroviral therapy but its pathophysiology remains elusive. In-vitro studies show that HIV-1-protease inhibitors affect adipocyte differentiation at an early step involving sterol-regulatory-element-binding-protein-1 (SREBP1), but in-vivo studies are lacking. METHODS: We compared fat morphology and mRNA and protein expression of major adipocyte differentiation markers and cytokines in subcutaneous abdominal adipose tissue from 26 HIV-1-infected patients who developed peripheral lipoatrophy while on protease inhibitors and from 18 HIV-1-seronegative healthy controls. FINDINGS: Patients' fat contained a higher proportion of small adipocytes than control fat, together with lower mRNA concentrations of the adipogenic differentiation factors CCAAT-enhancer binding protein (C/EBP) beta and alpha, peroxisome proliferator-activated receptor (PPAR) gamma, and the 1c isoform of SREBP1, with a median decrease of 93% in the latter. The SREBP1 protein concentration was increased 2.6-fold, whereas the PPARgamma protein concentration was decreased by 70%. The expression of adipocyte-specific markers, including leptin, was lower in fat from patients than in fat from controls, whereas expression of tumour necrosis factor (TNF) alpha was higher and correlated negatively with the expression of SREBP1c and downstream adipogenic factors. SREBP1c mRNA concentrations correlated negatively, and TNFalpha mRNA concentrations positively, with glycaemia and insulin resistance, but did not correlate with lipid variables. INTERPRETATION: The altered differentiation status of peripheral adipocytes in HIV-1-infected patients with antiretroviral-induced lipoatrophy is associated with greatly reduced SREBP1c expression. Since the differentiation factor SREBP1 is rapidly targeted by protease inhibitors in vitro, our results suggest that SREBP1c could be an important mediator of peripheral lipoatrophy in this setting, leading to metabolic alterations such as insulin resistance.