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High Plasma Level of Interleukin-18 in HIV-Infected Subjects With Lipodystrophy
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JAIDS Journal of Acquired Immune Deficiency Syndromes: Volume 36(1) 1 May 2004
Lindegaard, Birgitte MD*†; Hansen, Ann-Brit Eg MD*; Gerstoft, Jan MD, DrSci*; Pedersen, Bente Klarlund MD, DrSci*†
From *Department of Infectious Diseases, Rigshospitalet, University of Copenhagen, Denmark, and †the Copenhagen Muscle Research Centre, Rigshospitalet, University of Copenhagen, Denmark.
SUMMARY:
The study examined men with fat accumulation (mixed group) (n = 12) and one without fat accumulation (lipoatrophic group) (n = 15). Controls were HIV-positive men without LD (n = 15) and HIV-negative, age-matched men (n = 12). The study found that the lipoatrophic group had the highest IL-18.
The main finding of this study was that the level of circulating IL-18 is elevated in HIV-infected patients with lipoatrophy (both with and without central fat accumulation) compared with HIV-infected patients without LD.
High levels of IL-18 were found especially in patients who had received antiretroviral therapy for a long period. This could reflect lipoatrophy or that IL-18 was related to a direct effect of >=1 antiretroviral treatment.
A positive correlation was found between the total duration of NRTI use and IL-18 in HIV subjects without the mixed group as well as in the lipoatrophic group. The present study was not designed to study the specific effect of stavudine or other treatments. Patients currently receiving stavudine (n = 11) had significantly higher levels of plasma IL-18 compared with patients currently on regimens without stavudine. Patients who had stavudine switched to other antiretroviral drugs did not have a decrease in the level of IL-18 We found that IL-18 was correlated with TNF-a, thus supporting its role as a true proinflammatory cytokine. Given that IL-18 induces tissue destruction, it may be involved in adipocyte apoptosis occurring in patients with LD
IL-18 is a cytokine with proinflammatory features and is involved in tissue destruction. Thus, it may play a role in the host defense against infectious diseases and tumor growth, but it seems also to play a pathogenetic role in inflammatory conditions.
IL-18 induces interferon-g production and IL-6. In addition, IL-18 induces tumor necrosis factor-a (TNF-a) and TNF-a also stimulates IL-18 production. Thus, the two cytokines IL-18 and TNF-a seem to work in synergy.
The level of interleukin-18 (IL-18) is elevated in patients with HIV infection as well as in people with insulin resistance (IR). The level of IL-18 is elevated in patients with LD and closely linked to limb atrophy, whereas it is not associated with cholesterol or IR.
Elevated levels of IL-18 have recently been demonstrated in HIV-infected patients and especially in those with immunodeficiency and advanced clinical disease and to decline with HAART for 24 months. Recently, the levels of IL-18 were found to be elevated in persons with obesity, type 2 diabetes, and arteriosclerosis and to decline with weight loss. Given that these conditions share common features with HIV-associated LD, we hypothesized that IL-18 would be elevated in these patients. Furthermore, we specifically evaluated whether IL-18 was associated with fat mass, fat distribution, and NRTI treatment and whether IL-18 was associated with other cytokines such as TNF-a, IL-6, and adiponectin.
Abstract:
The level of interleukin-18 (IL-18) is elevated in patients with HIV infection as well as in people with insulin resistance (IR). As HIV-associated lipodystrophy (LD) shares metabolic characteristics with the metabolic syndrome, it was hypothesized that IL-18 would be elevated in patients with LD.
Two groups of HIV-infected men with LD, one with fat accumulation (mixed group) (n = 12) and one without fat accumulation (lipoatrophic group) (n = 15) were included. Controls were HIV-positive men without LD (n = 15) and HIV-negative, age-matched men (n = 12).
The levels of plasma IL-18 were elevated in all 3 HIV groups compared with HIV-negative controls (P <0.01). In the HIV groups the lipoatrophic group had the highest IL-18, followed by the mixed group and the HIV-positive controls. Only the differences between the lipoatrophic group and the HIV-positive controls were significant (P <0.01).
Plasma IL-18 correlated with tumor necrosis factor-a (P <0.05), but not IL-6, adiponectin, or HOMA-IR (homeostasis model of insulin resistance). In contrast to the HIV-negative controls, IL-18 did not correlate with total or low-density cholesterol in either of the HIV groups. An inverse correlation was observed between IL-18 and limb fat (P <0.05).
In conclusion, the level of IL-18 is elevated in patients with LD and closely linked to limb atrophy, whereas it is not associated with cholesterol or IR.
INTRODUCTION
A syndrome of lipodystrophy (LD), including peripheral subcutaneous fat loss, lipoatrophy, and central fat accumulation has been described in HIV-infected patients receiving highly active antiretroviral therapy (HAART). It is still not established whether loss of subcutaneous adipose tissue on the one hand and central fat accumulation on the other constitutes one pathogenetic entity. However, in many studies most patients presented a mixture of peripheral fat loss and central fat accumulation, named mixed LD. As other types of lipodystrophies (congenital as well as acquired) are characterized by selective loss of subcutaneous fat tissue only, some authors recognize HIV-associated LD as being primarily due to loss of peripheral fat, with fat accumulation as a secondary phenomenon. In support of this, a prospective study has recently demonstrated that HAART may induce a selective loss of limb fat. The development of LD was first observed in patients treated with a combination of protease inhibitors (PIs) and nucleoside reverse transcriptase inhibitors (NRTIs). Recently, LD has also been recognized in patients only receiving NRTIs. While the NRTI stavudine has especially been associated with a high risk for the development of lipoatrophy, the pathogenesis of HIV-associated LD is not known.
The HIV-associated LD syndrome has characteristics in common with the metabolic syndrome: hypertriglyceridemia, hypercholesterolemia, increased lipolysis, and insulin resistance, but the metabolic abnormalities are not only a result of the fat distribution in HIV. The metabolic alterations may also result from a direct effect from the PIs.
IL-18 is a cytokine with proinflammatory features and is involved in tissue destruction. Thus, it may play a role in the host defense against infectious diseases and tumor growth, but it seems also to play a pathogenetic role in inflammatory conditions. IL-18 is synthesized as an inactive precursor and converted to its biologic form by the cysteine protease caspase-1. IL-18 induces interferon-g production and IL-6. In addition, IL-18 induces tumor necrosis factor-a (TNF-a) and TNF-a also stimulates IL-18 production. Thus, the two cytokines IL-18 and TNF-a seem to work in synergy. The cytokine adiponectin is associated with both diabetes and LD, but its possible relationship to IL-18 has not been evaluated.
Elevated levels of IL-18 have recently been demonstrated in HIV-infected patients and especially in those with immunodeficiency and advanced clinical disease and to decline with HAART for 24 months. IL-18 has been reported both to inhibit and to induce HIV-1 expression in monocytes. Recently, the levels of IL-18 were found to be elevated in persons with obesity, type 2 diabetes, and arteriosclerosis and to decline with weight loss. Given that these conditions share common features with HIV-associated LD, we hypothesized that IL-18 would be elevated in these patients. Furthermore, we specifically evaluated whether IL-18 was associated with fat mass, fat distribution, and NRTI treatment and whether IL-18 was associated with other cytokines such as TNF-a, IL-6, and adiponectin.
RESULTS
The HIV groups did not differ with regard to duration of HIV infection, CD4 cell count, or HIV RNA. The lipoatrophic group had received therapy for a longer period than the HIV controls and had a lower body mass index compared with the other groups. The DXA scans supported the clinical diagnosis of LD and revealed 3 distinct different HIV groups with regard to phenotype.
The lipoatrophic group was characterized by reduced limb fat mass compared with the mixed group, the HIV controls, and the healthy controls. All groups had the same lean body mass. All HIV-infected patients had reduced limb fat mass compared with the healthy controls. The mixed group had increased truncal fat mass compared with the lipoatrophic group and the HIV controls but not compared with the healthy controls.
The levels of plasma IL-18 were elevated in all 3 HIV groups compared with HIV-negative controls (P <0.01). In the HIV groups, the lipoatrophic group had the highest IL-18, followed by the mixed group, and the HIV controls. Only the differences between the lipoatrophic group and the HIV controls were significant: 345.9 (264.3-427.3 pg/mL) vs. 212.6 (151.0-259.5 pg/mL), P <0.01.
A positive correlation between IL-18 and TNF-a was found in all HIV-infected patients. IL-18 did not correlate with IL-6, adiponectin, or HOMA-IR in any of the groups. In the healthy group, IL-18 correlated with the total cholesterol, LDL, and triglycerides. However, such correlations did not exist in either of the HIV groups, although triglycerides correlated with IL-18 within HIV subjects without the mixed group. Within all subjects a negative correlation between IL-18 and limb fat mass was found, but no correlation within the HIV subgroups except for HIV subjects with exclusion of the mixed group. In contrast to the HIV-infected patients, IL-18 correlated positively to the total fat mass and to the truncal fat mass in the healthy controls.
Moreover, a positive correlation was found between the total duration of NRTI use and IL-18 in HIV subjects without the mixed group as well as in the lipoatrophic group.
A negative correlation was also found between total duration of NRTI use and limb fat mass within the HIV groups (HIV subjects without the mixed group: rs = -0.497, n = 26, P <0.02; all HIV subjects: rs = -0.484, n = 35, P <0.001). There was no correlation between the duration of NRTI use and trunk fat mass in either of the HIV groups (HIV subjects without the mixed group: rs = -0.270, n = 26, P = NS; All HIV subjects: rs =-0.188, n = 35, P = NS) (data not shown).
Patients currently receiving stavudine (n = 11) had significantly higher levels of plasma IL-18 compared with patients currently on regimens without stavudine (n = 28, 369.4 [323.3-409.5] vs. 221.5 [170.2-351.49] pg/mL, P <0.01).
Plasma IL-18 did not differ between HIV-infected patients with HIV-1 RNA >200 copies/mL (n = 8) and patients with HIV-1 RNA <200 copies/mL (n = 31). Furthermore, IL-18 did not correlate to CD4 cell count in any of the HIV groups.
DISCUSSION by authors
The main finding of this study was that the level of circulating IL-18 is elevated in HIV-infected patients with lipoatrophy (both with and without central fat accumulation) compared with HIV-infected patients without LD.
Elevated levels of IL-18 have recently been demonstrated in HIV-infected patients and especially in those with immunodeficiency and advanced clinical disease. This could indicate that IL-18 was related to high viremia. However, we did not find that viral load had any influence on the level of IL-18, presumably because all patients were on stable and successful HAART. In addition, the level of IL-18 was not related to the CD4 cell count. Chronic inflammation is well documented in HIV disease, even in patients who are successfully treated with HAART. The elevated level of IL-18 correlated well with circulating levels of TNF-a and thus is likely to reflect such inflammation. In addition, inflammation may be a cause of HAART-induced metabolic changes. Although longitudinal prospective studies are required to fully answer this question, we found indications for a relationship between inflammation and altered metabolism.
The finding that IL-18 was elevated in both the mixed and the lipoatrophic groups indicates that IL-18 is linked with the peripheral fat atrophy. In support, IL-18 correlated inversely with limb fat mass. The fact that the levels of IL-18 did not differ between patients in the mixed group and the HIV control group would be in accordance with the possibility that IL-18 was mechanistically linked to the low total limb fat mass, as the DXA scans revealed that the HIV controls also had low limb fat mass, despite their phenotype. This DXA scan finding is in agreement with recent observations in Fat Redistribution and Metabolic Change (FRAM) Study. IL-18 did not correlate to LDL and trunk fat in HIV patients as opposed to the healthy controls, indicating that IL-18 is differently regulated in patients with and patients without HIV, a suggestion further supported by the fact that IL-18 is positively correlated to the total fat mass in the healthy; but when all the HIV-infected subjects are included in the analysis the correlation is negative. A strong correlation exists between IL-18 and triglycerides in healthy subjects. Such a correlation is also present in HIV subjects if patients with fat accumulation are excluded.
We found an association between IL-18 and the duration of NRTI therapy. In a previous study the level of IL-18 declined following HAART for 24 months. However, in the present study, all patients received HAART, and high levels of IL-18 were found especially in patients who had received antiretroviral therapy for a long period. This could reflect the lipoatrophy phenotype or that IL-18 was related to a direct effect of >=1 antiretroviral treatment.
Stavudine has been associated with the development of lipoatrophy. The present study was not designed to study the specific effect of stavudine or other treatments. However, patients on current treatment with stavudine had higher levels of IL-18 than other patients. Such a possible relationship is not likely to be due to an acute effect of stavudine as patients who had stavudine switched to other antiretroviral drugs did not have a decrease in the level of IL-18 (median and 25 and 75% quartiles, 247 (186-366) vs. 273 (166-399) pg/mL, before and after, respectively stavudine switching, unpublished data). We found that IL-18 was correlated with TNF-a, thus supporting its role as a true proinflammatory cytokine. Given that IL-18 induces tissue destruction, it may be involved in adipocyte apoptosis occurring in patients with LD. Thus, IL-18 could represent a link between NRTI therapy and fat atrophy.
In conclusion, the level of IL-18 is elevated in patients with LD (both with and without fat accumulation) and closely linked to limb atrophy, whereas it is not associated with cholesterol or IR.
Patients and Methods
Forty-two HIV-infected men were recruited from the outpatient clinic of the Department of Infectious Disease, Rigshospitalet in Copenhagen. Informed consent was obtained from all patients according to declaration of the local ethical committee. LD was defined clinically by physical examination of peripheral lipoatrophy (fat loss from face, arms, buttocks, or legs) with or without central accumulation (abdomen, dorsocervical fat pad). Age-matched patients were enrolled into the following 3 groups according to clinical examination: patients with lipoatrophy with truncal fat accumulation (the mixed group) (n = 12); patients with lipoatrophy without truncal fat accumulation (the lipoatrophic group) (n =15); and patients without LD (HIV controls) (n=15). All patients were on a stable and effective nucleoside analogue-based HAART with no changes in antiretroviral therapy during the preceding 8 weeks. The combinations were as follows: 10 patients were receiving PIs and 4 nonnucleoside analogues in the mixed group; 10 patients were receiving PIs and 4 nonnucleoside analogues in the lipoatrophic group; in the HIV control group, 9 patients were receiving PIs and 5 nonnucleoside analogues; and 12 age-matched HIV-negative healthy men served as controls.
None of the HIV-infected patients had signs of ongoing infections or fasting glucose >7 mM.
Peripheral blood samples were obtained after an overnight fasting. Measurements of total cholesterol (mM), high-density lipoprotein (HDL) cholesterol (mM), low-density lipoprotein (LDL) cholesterol (mM), triglycerides (mM), plasma glucose (mM), and insulin (pM) were determined immediately using routine methods.
CD4 cell counts were calculated by flow cytometry and HIV RNA copies were measured by the Amplicor HIV Monitor (Roche Molecular Systems, Branchburg, NJ) (lower limit of detection: 20 copies/mL).
Cytokines IL-18, IL-6, and TNF-a were measured in plasma.
Adiponectin was determined by a radioimmunoassay kit (LINCO Research, Inc., St. Charles, MO). Threshold of detection was 1 ng/mL. Insulin resistance was assessed from fasting plasma insulin and glucose by using the homeostasis model (HOMA-IR) as described previously.
Body Composition Analysis
Fat and fat-free tissue masses for the whole body, trunk, and extremities were measured using dual-energy x-ray absorptiometry (DXA) scanner, Norland XR 36 (version 3.94; Norland Corp., Fort Atkinson, WI). All measurements were done in a single laboratory. Whole-body and regional fat measurements (trunk and extremity) were determined.
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