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FJ
EXPRESS SUMMARY ARTICLE The Full-length version of this article is also available, published online April 7, 2005 as doi:10.1096/fj.04-3094fje. |
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,1
* The CBR Institute for Biomedical Research and
Department of Pathology, Harvard Medical School, Boston, Massachusetts, USA
1 Correspondence: The CBR Institute for Biomedical Research, 800 Huntington Ave., Boston, MA 02115, USA. E-mail: Wagner{at}cbr.med.harvard.edu
SPECIFIC AIMS
Soluble ICAM-1 (sICAM-1) is increased in many pathological conditions, including obesity. The aim of this study was to generate an animal model overexpressing sICAM-1 and to determine the long-term effects of elevated sICAM-1 on the animal. The emphasis was to investigate the immune responses in transgenic mice and to evaluate the effect of elevated sICAM-1 on adiposity.
PRINCIPAL FINDINGS
1. sICAM-1 transgenic mice produce up to 10-fold higher circulating ICAM-1
The highest sICAM-1 expression among the three founders identified by PCR genotyping generated a line that possessed at least 24 copies of the sICAM-1 (sICAM-1H). Another line with lower expression level (sICAM-1L) was kept for comparison; it had 6 copies of the transgene in the genome. Using immunoprecipitation, we detected a 90 kDa form of ICAM-1 in sICAM-1H mice plasma; the same size protein was detected in wild-type mice but to a lesser extent. The sICAM-1 expression level in the sICAM-1H line was 129 ± 3.5µg/mL, 
10-fold higher than that in wild-type mice.
2. Elevated sICAM-1 reduces neutrophil and monocyte/macrophage recruitment in thioglycollate-induced peritonitis
Thioglycollate induced recruitment of neutrophils to the peritoneal cavity in wild-type mice at 4 h (7.62±0.71x106). Reduction of neutrophil recruitment was observed in sICAM-1 transgenic mice. In the sICAM-1L line, the number of neutrophils in the peritoneal lavage was 4.98 ± 0.73 x 106, representing a 35% statistically significant decrease compared with wild-type (P<0.05). Recruitment was further reduced in sICAM-1H mice to 1.96 ± 0.57 x 106; compared with wild-type animals, the difference was highly significant (P<0.01). Reduction of neutrophil recruitment in the sICAM-1H mice was similar to that observed in the ICAM-1-deficient mice (P>0.05). Inhibition of macrophage recruitment to the peritoneum was observed in the sICAM-1H mice (4.99±0.20x106) 48 h after thioglycollate injection compared with wild-type mice (9.59±1.10x106), and this was statistically significant (P<0.001).
3. Contact hypersensitivity response was inhibited in sICAM-1 transgenic mice
To investigate contact hypersensitivity responses, animals were sensitized and the ear was elicited with 2,4-dinitro-1-fluorobenzene (DNFB). Ear swelling (Fig. 1
A) was significantly reduced in sICAM-1H transgenic mice compared with that in control mice (P<0.01). The response of the transgenic mice was comparable to the ICAM-1-deficient mice (P>0.05). Histological examination of ear skin sections revealed inhibition of inflammatory cell infiltration in the dermis in the sICAM-1H transgenic mice (Fig. 1B
). Accordingly, there was significant reduction of myeloperoxidase (MPO) activity in sICAM-1H mice compared with wild-type mice (Fig. 1C
, P<0.01). A similar reduction of MPO activity was present in the DNFB-treated ear in ICAM-1-deficient mice (P>0.05). Our results indicate that the presence of elevated levels of soluble ICAM-1 renders the mice immune deficient to an extent similar to that of ICAM-1-deficient mice in the contact hypersensitivity response.
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4. Draining lymph nodes from sICAM-1H mice were less responsive to DNFB application
The draining inguinal lymph nodes were examined 2 days after initial DNFB application. Notable lymph node hypertrophy was observed in wild-type mice; this enlargement was less obvious in sICAM-1H- and ICAM-1-deficient groups. The average size of inguinal lymph nodes was 6.56 ± 0.40 mg in wild-type mice and 5.01 ± 0.22 mg in sICAM-1H mice (P<0.01). Inguinal lymph nodes size in ICAM-1-deficient mice was comparable to that of the sICAM-1H mice group after DNFB application (P>0.05).
5. Elevated sICAM-1 levels increased adiposity of the transgenic mice
ICAM-1-deficient mice have increased adiposity in old age or on a high-fat diet compared with wild-type. To examine the long-term effect of elevated sICAM-1 on metabolism, animals were fed a Western-type diet. Both male and female sICAM-1H mice gained more body weight than wild-type groups (Fig. 2
A, B). The body weight of male sICAM-1H mice became significantly higher than wild-type 2 wk after administration of a Western-type diet (P<0.05). There were no statistical differences in food consumption in these mice (Fig. 2C, D
). The high-fat diet was withdrawn and a standard chow diet was provided for an additional 14 wk, allowing the mice to adjust and age. Body weight and BMI in male and female sICAM-1H mice were significantly higher than wild-type at age 35 wk. Their percent body fat was significantly increased over wild-type (19%:12%, P<0.03 for males and 18%:11%, P<0.008, for females), reminiscent of the phenotype of ICAM-1-deficient mice.
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CONCLUSIONS AND SIGNIFICANCE
Using transgenic mice, we have demonstrated that increased circulating sICAM-1 inhibits neutrophil and macrophage recruitment during inflammation. Stronger inhibition was observed in the sICAM-1H line, and this dose effect in the thioglycollate-induced inflammatory response supports the claim that the phenotype of transgenic mice results from elevated sICAM-1. Leukocyte transmigration is a multistep process. Firm adhesion of the leukocytes to the endothelium requires interaction of ICAM-1 with ß2 integrin; therefore, this interaction has become an important drug target in inflammatory diseases and ischemia/reperfusion injury. Our data demonstrate that sICAM-1 can act as a regulator during inflammatory processes. We speculate that excessive circulating sICAM-1 in the transgenic animals may bind to ß2 integrin on the leukocyte and thus decrease its availability for cell-cell interactions (Fig. 3
).
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A contact hypersensitivity response was inhibited in sICAM-1H mice, characterized by decreased ear swelling and less inflammatory cell infiltration after DNFB application. This finding indicates that increased levels of sICAM-1 have immuno-suppressant effects. The reduced immune reaction of the transgenic mice is similar in many respects to that of ICAM-1-deficient mice, which display impaired inflammation and immune responses. Moreover, we observed reduced MPO activity in DNFB-treated ear skin from sICAM-1H mice compared with wild-type animals.
The reduced contact hypersensitivity response in the sICAM-1H transgenic mice may also be explained by inhibition of the sensitization phase. Lymphocytes could be less activated in the sICAM-1H mice, as the draining lymph nodes sizes were smaller than wild-type and similar to those in ICAM-1-deficient mice. Given that LFA-1/ICAM-1 complexes are involved in the formation of immune synapses, which promote an efficient TCR-MHC interaction, sICAM-1 may interfere with T cell activation.
In addition to the defects in inflammatory and immune responses, sICAM-1H mice were more susceptible to weight gain (Fig. 2)
. Male and female sICAM-1H mice had significantly higher BMI in older age (8 months), with more body fat than wild-type mice. This suggests that elevation of sICAM-1 increases adiposity in the transgenic animals. Elevated sICAM-1 is reported in obese people. Levels of sICAM-1 positively correlate with BMI, and weight reduction decreases the soluble adhesion molecule levels. Thus, the high levels of sICAM-1 encountered in obesity may actually further worsen the condition. Earlier work from our laboratory demonstrated that ß2 integrin- or ICAM-1-deficient mice are susceptible to obesity. Recently we showed a severely diminished metabolic response to fasting in ICAM-1- and CD18-deficient mice. The study proposed that interaction between ICAM-1 and CD-18 is involved in regulating insulin metabolism and fat oxidation. Defects in these adhesion receptors or their inhibition with agents such as sICAM-1 thus may result in excessive body fat accumulation.
The transmembrane receptor ICAM-1 is traditionally considered an immune-regulating molecule; current observations broaden our view of the functional importance of this receptor by demonstrating biological activity of its cleaved form in plasma.
FOOTNOTES
To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.04-3094fje; doi: 10.1096/fj.04-3094fje
This article has been cited by other articles:
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  D. K. Brake, E. O. Smith, H. Mersmann, C. W. Smith, and R. L. Robker ICAM-1 expression in adipose tissue: effects of diet-induced obesity in mice Am J Physiol Cell Physiol, December 1, 2006; 291(6): C1232 - C1239. [Abstract] [Full Text] [PDF]
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