Published online January 4, 2006
PEDIATRICS
Vol. 117
No. 1
January 2006, pp.
220-223
(doi:10.1542/peds.2004-2854)
Obesity and Inflammation: Evidence for an Elementary Lesion
Andrea Sbarbati, MD, PhDa,
Francesco Osculati, MDa,
Davide Silvagni, MDb,
Donatella Benati, PhDa,
Mirco Galiè, PhDa,
Francesco Saverio Camoglio, MDc,
Gino Rigotti, MDd and
Claudio Maffeis, MDb
a Departments of Morphological-Biomedical Sciences, Human Anatomy and Histology Section
b Department of Mother and Child Biology-Genetics
c Pediatric Surgery Unit, University Hospital, University of Verona, Verona, Italy
d Plastic and Reconstructive Surgery (II Division), City Hospital, Verona, Italy
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ABSTRACT
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In obesity, an inflammatory process of the adipose tissue has been hypothesized; however, direct evidence for a tissue lesion is still lacking. Macrophage infiltration in the adipose tissue of obese individuals seems to be proven, but other alterations of the tissue have not been demonstrated.
Moreover, in humans it has not been clarified whether inflammation is an early characteristic of obesity, because no data from obese children are available. In the present study, we assessed the inflammatory involvement of the adipose tissue and identified the elementary "inflammatory" lesion in a group of obese children. The study of children gives us the chance to investigate adipose tissue during early phases of obesity. In all the obese subjects, ultramicroscopic analysis of the adipose tissue demonstrated inflammatory involvement, and the extent of the lesions seemed to depend on the SD score of body mass index. The elementary lesion is a microgranuloma, with fragments of adipocytes, that evolves to fibrosis. Macrophages (and less frequently, lymphocytes or granulocytes) were found in perivascular positions. The lesions were not found in nonobese children. Our study proved that an "inflammatory" process exists in the adipose tissue of obese children, confirming previous findings in animals and obese adults and demonstrating that it is an early alteration in humans. However, the accumulation of macrophages was just one of the components of the inflammatory lesion, which also involved adipocyte degeneration, fibrosis, and, to a lesser extent, granulocyte/lymphocyte accumulation. The finding of fragments of adipocytes in the elementary lesion suggests that, at the beginning of the process, adipocytes may degenerate and that the materials generated by this process can recruit macrophages and other leukocytes. These preliminary results suggest that additional studies should be designed to clarify the cause of adipocyte fragility in obese children.
Key Words: obesity • inflammation • children
Abbreviations: DXA, dual x-ray absorptiometry • SDS, SD score
Obesity is associated with a chronic low-grade inflammation,1,2 and inflammatory pathways could be critical in the mechanisms underlying obesity and its complications, although the factors triggering the inflammatory response are not known, especially in humans. The inflammatory process seems to originate and reside mainly in adipose tissue, which could regulate the process through cytokine production. Recent studies in mice documented leukocyte populations within adipose tissue, which are potentially involved in the development of the inflammatory status that is characteristic of obesity.3 Xu et al4 reported a significant infiltration of macrophages in adipose tissue, finding that inflammation and macrophage-specific genes are up-regulated in such tissue, which heralds an increase in circulating insulin levels. Genes typically expressed by macrophages correlated positively with fat mass in mice, and the percentage of cells expressing the macrophage marker F4/80 correlated positively with adipocyte size and body mass.5 Similar relationships were found in human subcutaneous adipose tissue stained for the macrophage antigen CD68.6
On the basis of evidence available mainly from animal models, macrophage infiltration in the adipose tissue of obese individuals seems to have been proven. However, it is not clear whether this is an isolated event or whether it is associated with other alterations of the tissue that have not been demonstrated by molecular biology or light-microscopy approaches. Moreover, it has not been clarified whether it is an early characteristic of obesity in humans, because no data are available from obese children.
This study was performed in a group of obese children who had not been exposed to overweight and its metabolic consequences for a long period and therefore constituted a model for the study of the early stages of the disease in humans. The aims of this study were to assess the inflammatory involvement of adipose tissue in children and identify the potential elementary inflammatory lesion.
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MATERIALS AND METHODS
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Nineteen prepubertal white children (11 males and 8 females), 11.0 ± 1 years old (range: 7.2–13.0 years), were recruited among the children who attended the obesity clinic of the Department of Pediatrics at University Hospital (Verona, Italy). Each child underwent a physical examination, anthropometric measures, and body densitometry (dual x-ray absorptiometry [DXA]) for body-composition analysis. The development of puberty was clinically assessed on the basis of Tanner stages.7 None of the children had any overt disease other than obesity, none of them were dieting at the time of the study, and none were on medication. Informed consent was obtained from the parents before the children took part in the study. Subcutaneous adipose tissue of nonobese children (4 males and 1 female; 7.4 ± 1.6 years old [range: 6.0–10.0 years]) was provided during surgical procedures and was used as a control. Height, weight, and body composition were measured for all the children. The protocol was approved by the University Hospital Ethical Committee, and informed consent was obtained from the parents of all children.
Anthropometric Measurements
Height and weight were measured in postabsorptive conditions, with the subject having an empty bladder. Height was measured to the nearest 0.5 cm on a standardized height board. Weight was determined to the nearest 0.1 kg on a standard physician's beam scale, with the subject dressed only in light underwear and no shoes. The body mass index (BMI) was calculated as weight (kg) divided by height squared (m2). The SD score (SDS) of BMI was calculated with the LMS method, using national reference values of BMI and LMS coefficients.8 Total-body DXA was performed to assess body composition. Fat mass and fat-free mass were measured by DXA using a Lunar DPX-L densitometer (Lunar, Madison, WI). The subjects were scanned in light clothing while they were lying flat on their backs. On the day of each test, the DPX-L was calibrated according to the procedures described by the manufacturer. Body fat mass was obtained by multiplying the percentage of body fat by body weight.
Experimental Design
After anthropometry and body-composition measurements were obtained, the children, in the postabsorptive state, underwent needle aspiration of abdominal subcutaneous adipose tissue 5 to 8 cm lateral to the umbilicus. Local anesthesia was achieved with 1% xylocaine, and 200 to 400 mg of subcutaneous adipose tissue were aspirated by using a 15-gauge needle.
Adipose-Tissue Evaluations
Adipocyte size was evaluated by light microscopy on whole mounts of unfixed tissue. The total number of adipose cells in the body was estimated by dividing total body fat by the mean of the adipocyte size. Specimens of subcutaneous fat obtained from the abdominal region were processed for light and electron microscopy. The specimens were fixed in 2.5% glutaraldehyde in Sorensen's buffer for 2 hours, postfixed in 1% osmium tetroxide in Sorensen's buffer for 1 hour, dehydrated in graded ethanols, embedded in Epon-Araldite, and cut with an Ultracut E ultramicrotome (Reichert-Jung, Wien, Austria). Semithin sections were stained by using toluidine blue. Four semithin sections for each sample were analyzed. Ultrathin sections were stained with lead citrate and uranyl acetate and observed under an EM10 electron microscope (Zeiss, Oberkochen, Germany).
Lesion quantification was performed on semithin sections. Briefly, the total sectional area of the adipose tissue and the lesioned area were selected and quantified by using Image-Pro Plus software (Media Cybernetics, San Diego, CA). The results were expressed as percentage ratio between the lesioned and the total sectional area. All values are expressed as mean ± SD. The statistical significance of the data were assessed by using the analysis of variance test. A P value of <.05 was accepted as statistically significant.
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RESULTS
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In the obese group, the SDS of the BMI was 2.60 ± 0.84 (range: 1.49–4.31), the body fat mass was 47% ± 8% (range: 35–60%), and the adipocyte diameter was 86 ± 22 mm (range: 58–144 mm). The total number of adipose cells in the body was 10.3 ± 7.4 x 1010 (range: 3.4–26.8 x 1010). In the nonobese group, the BMI was 16.5 ± 0.85 kg/m2 (range: 13.4–17.4 kg/m2), the body fat mass was 12.3% ± 1.9% (range: 10–15%), and the adipocyte diameter was 37.2 ± 1.9 mm (range: 35–40 mm).
In all the obese children, light and ultramicroscopic analysis of the adipose tissue demonstrated an inflammatory involvement of the tissue (Fig 1). At quantitative analysis, the lesion involved 6.8% ± 8.5% of the examined adipose tissue. Lesions appeared more evident in patients with a high BMI. In fact, within the obese group, the children with an SDS of BMI that was higher than the mean value (ie, 2.6) showed lesioned areas significantly larger (11.0% ± 11.5%, n = 8) than children with an SDS of BMI that was under the mean value (2.7 ± 2.6 [n = 11]; P < .05).
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FIGURE 1 Electron microscopic features of subcutaneous adipose tissue in obese children. A, A macrophage (m) is visible near a vessel (v) (original magnification, x4000). B, A granulocyte (g) near degenerating adipocytes (asterisks) is visible (original magnification, x3000). C, A lymphocyte (l) and degenerating adipocytes (asterisks) near a vessel (v) are visible (original magnification, x3500). D, A macrophage (m), lipid droplets (d), and degenerating adipocytes (asterisks) near a vessel (v) are visible (original magnification, x3000). The black lines mark thickened septa between adipocytes (a).
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At ultrastructural examination, the elementary lesion was a microgranuloma surrounding debris of adipocytes. One or more macrophages were often found in perivascular position (Fig 1A). Macrophages often showed lipid droplets. Granulocytes (Fig 1B) or lymphocytes (Fig 1C) were found also, but less frequently. The mature adipocytes showed normal morphology, being characterized by very thin cytoplasmic sheets, but adipocytes that had broken down were often visible in association with inflammatory cells (Fig 1D). The capillaries showed normal morphology. A thickening of the interadipose septa with a mild degree of fibrosis was found in some areas. The adipose tissue of nonobese children did not show any signs of inflammation.
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DISCUSSION
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The results of this study clearly demonstrate infiltration of blood elements in the adipose tissue of obese children. The novelty of these results exists in the identification of an elementary lesion by ultramicroscopic analysis in these young subjects. The study of children provides the opportunity to investigate the anatomy of adipose tissue during early phases of obesity. Children have had only short-term exposure to obesity or its complications, which potentially may affect the development or maintenance of inflammation in adipose tissue. We found that an inflammatory process exists in the adipose tissue of obese children, which confirms previous findings in animals and obese adults and suggests that this is an early alteration in human adipose tissue.
Furthermore, we demonstrated that the accumulation of macrophages in the adipose tissue of the obese is just one of the components of the inflammatory lesion, which also involves adipocyte degeneration, fibrosis, and, to a lesser extent, polymorphonucleates and lymphocyte accumulation. We did not find transitional elements between adipocytes and macrophages, and the latter were found mainly in perivascular positions. This suggests blood migration of macrophages rather than differentiation from preadipocytes, thereby confirming previous findings in adults.4,9
Although the cross-sectional design of this study did not allow us to establish a cause-and-effect relationship, it is likely that microgranuloma evolves to fibrosis, which, however, seems to be modest. The finding of fragments of adipocytes in the elementary lesion suggests that adipocytes may degenerate. The causes of this cell fragility are not known but could include deterioration of the cytoskeleton, cell membrane, or connective scaffold. A possible site of this weakness could be in the cytoplasmatic sheets of the adipocytes. The materials liberated by this process seem to recruit macrophages and, to a lesser extent, other leukocytes.
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CONCLUSIONS
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This study demonstrates that the elementary lesion in early obesity is a microgranuloma with lipodegenerative aspects. It is likely that it is the result of adipocyte fragility and a consequent recruitment of macrophages and that this process leads to a moderate fibrosis. Additional studies will clarify whether this lesion is merely an epiphenomenon accompanying obesity or whether it plays a pathogenic role in the maintenance of the inflammatory state that characterizes obesity.
Findings in adult humans and in animals suggest that the inflammatory status associated with the increase of fat mass may be involved in the pathogenetic pathways of obesity complications. The increase in adipose tissue contributes to a proinflammatory milieu, and inflammatory adipokines may promote insulin resistance, endothelial dysfunction, and finally atherosclerosis.2 A similar pathway can be hypothesized for the genesis of atheroma in obese children, because arterial stiffness and endothelial dysfunction have been shown to be related to obesity in childhood.10 The evidence of inflammatory involvement in the adipose tissue of obese children suggests an early onset of pathogenetic mechanisms that may favor the complications of obesity.
Identification of the elementary inflammatory lesion in the adipose tissue of obese subjects may suggest new potential targets for the treatment or prevention of the complications of obesity.
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ACKNOWLEDGMENTS
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We thank Dr Christine Harris for revising the manuscript.
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FOOTNOTES
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Accepted Apr 8, 2005.
Address correspondence to Andrea Sbarbati, MD, PhD, Human Anatomy and Histology Section, University of Verona, Medical Faculty, Strada Le Grazie 8, 37134 Verona, Italy. E-mail: andrea.sbarbati{at}univr.it
Dr Sbarbati had full access to all data in the study and had final responsibility for deciding what to submit for publication.
The authors have indicated they have no financial relationships relevant to this article to disclose.
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REFERENCES
|
|---|
- Yudkin JS. Adipose tissue, insulin action and vascular disease: inflammatory signals.
Int J Obes Relat Metab Disord. 2003;27
:S25
–S28
- Lyon CJ, Law RE, Hsueh WA. Minireview: adiposity, inflammation, and atherogenesis.
Endocrinology. 2003;144
:2195
–2200[Abstract/Free Full Text]
- Robker RL, Collins RG, Beaudet AL, Mersmann HJ, Smith CW. Leukocyte migration in adipose tissue of mice null for ICAM-1 and Mac-1 adhesion receptors.
Obes Res. 2004;12
:936
–940[ISI][Medline]
- Xu H, Barnes GT, Yang Q, et al. Chronic inflammation in fat plays a crucial role in the development of obesity-related insulin resistance.
J Clin Invest. 2003;112
:1821
–1830[CrossRef][ISI][Medline]
- Weisberg SP, McCann D, Desai M, Rosenbaum M, Leibel RL, Ferrante AW Jr. Obesity is associated with macrophage accumulation in adipose tissue.
J Clin Invest. 2003;112
:1796
–1808[CrossRef][ISI][Medline]
- Wellen KE, Hotamisligil GS. Obesity-induced inflammatory changes in adipose tissue.
J Clin Invest. 2003;112
:1785
–1788[CrossRef][ISI][Medline]
- Tanner JM.
Growth at Adolescence. 2nd ed. Oxford, United Kingdom: Blackwell; 1962
- Luciano A, Bressan F, Zoppi G. Body mass index reference curves for children aged 3–19 years from Verona, Italy.
Eur J Clin Nutr. 1997;51
:6
–10[CrossRef][ISI][Medline]
- Curat CA, Miranville A, Sengenes C, et al. From blood monocytes to adipose tissue-resident macrophages: induction of diapedesis by human mature adipocytes.
Diabetes. 2004;53
:1285
–1292[Abstract/Free Full Text]
- Tounian P, Aggoun Y, Dubern D, et al. Presence of increased stiffness of the common carotid artery and endothelial dysfunction in severely obese children: a prospective study.
Lancet. 2001;358
:1400
–1404[CrossRef][ISI][Medline]
PEDIATRICS (ISSN 1098-4275). ©2006 by the American Academy of Pediatrics
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ABSTRACT
MATERIALS AND METHODS
