Semicarbazide-Sensitive Amine Oxidase/Vascular Adhesion Protein-1 Deficiency Reduces Leukocyte Infiltration into Adipose Tissue and Favors Fat Deposition
2009; Elsevier BV; Volume: 174; Issue: 3 Linguagem: Inglês
10.2353/ajpath.2009.080612
ISSN1525-2191
AutoresSandy Bour, Sylvie Caspar‐Bauguil, Zsuzsa Iffiú-Soltész, Maryse Nibbelink, Béatrice Cousin, Mari Miiluniemi, Marko Salmi, Craig Stolen, Sirpa Jalkanen, Louis Casteilla, Luc Pénicaud, Philippe Valet, Christian Carpéné,
Tópico(s)Body Contouring and Surgery
ResumoObesity is associated with low-grade inflammation and leukocyte infiltration in white adipose tissue (WAT) and is linked to diabetic complications. Semicarbazide-sensitive amine oxidase, also known as vascular adhesion protein-1 (SSAO/VAP-1), is a membrane protein that is highly expressed in adipocytes and is also present on the endothelial cell surface where it is involved in leukocyte extravasation. We studied fat deposition and leukocyte infiltration in WAT of mice with a null mutation in the amine oxidase copper-containing-3 (AOC3) gene encoding SSAO/VAP-1. Both epididymal and inguinal WATs were larger in 6-month-old AOC3-KO males than in age-matched wild-type controls. However, WAT from AOC3-KO mice contained lower CD45 mRNA levels and fewer CD45+ leukocytes. Subpopulation analyses revealed a diminished infiltration of WAT by T cells, macrophages, natural killer, and natural killer T cells. A decrease in leukocyte content in WAT was also detected in female AOC3-KO mice as early as 2 months of age, whereas increased fat mass was evident by 6 months of age. Reduced CD45+ populations in WAT of AOC3-KO mice was not rescued by human SSAO/VAP-1 expression on adipocytes under the control of aP2, suggesting the importance of vascular AOC3 in leukocyte entrance into fat. Our results indicate that SSAO/VAP-1 is instrumental for the presence of leukocytes in WAT. Therefore, AOC3-KO mice present a unique model of mild obesity, characterized by increased WAT devoid of low-grade inflammation. Obesity is associated with low-grade inflammation and leukocyte infiltration in white adipose tissue (WAT) and is linked to diabetic complications. Semicarbazide-sensitive amine oxidase, also known as vascular adhesion protein-1 (SSAO/VAP-1), is a membrane protein that is highly expressed in adipocytes and is also present on the endothelial cell surface where it is involved in leukocyte extravasation. We studied fat deposition and leukocyte infiltration in WAT of mice with a null mutation in the amine oxidase copper-containing-3 (AOC3) gene encoding SSAO/VAP-1. Both epididymal and inguinal WATs were larger in 6-month-old AOC3-KO males than in age-matched wild-type controls. However, WAT from AOC3-KO mice contained lower CD45 mRNA levels and fewer CD45+ leukocytes. Subpopulation analyses revealed a diminished infiltration of WAT by T cells, macrophages, natural killer, and natural killer T cells. A decrease in leukocyte content in WAT was also detected in female AOC3-KO mice as early as 2 months of age, whereas increased fat mass was evident by 6 months of age. Reduced CD45+ populations in WAT of AOC3-KO mice was not rescued by human SSAO/VAP-1 expression on adipocytes under the control of aP2, suggesting the importance of vascular AOC3 in leukocyte entrance into fat. Our results indicate that SSAO/VAP-1 is instrumental for the presence of leukocytes in WAT. Therefore, AOC3-KO mice present a unique model of mild obesity, characterized by increased WAT devoid of low-grade inflammation. Anatomical and functional relationships exist between the immune system and adipose tissue. 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The SVF contains all of the cell types found in adipose tissue (endothelial cells, fibroblasts, preadipocytes, immune cells) except the lipid-laden adipocytes. 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Furthermore, an increase of T cells was reported in the fatter WAT of diet-induced obese mice.15Caspar-Bauguil S Cousin B André M Nibbelink M Galinier A Periquet B Casteilla L Pénicaud L Weight-dependent changes of immune system in adipose tissue: importance of leptin.Exp Cell Res. 2006; 312: 2195-2202Crossref PubMed Scopus (36) Google Scholar, 16Wu H Ghosh S Perrard X Feng L Garcia G Perrard J Sweeney J Peterson L Chan L Smith C Ballantyne C T-cell accumulation and regulated on activation, normal T cell expressed and secreted upregulation in adipose tissue in obesity.Circulation. 2007; 115: 1029-1038Crossref PubMed Scopus (528) Google Scholar SSAO/VAP-1 (semicarbazide-sensitive amine oxidase/vascular adhesion protein-1) is abundantly present on adipocytes.17Morin N Lizcano JM Fontana E Marti L Smih F Rouet P Prévot D Zorzano A Unzeta M Carpéné C Semicarbazide-sensitive amine oxidase substrates stimulate glucose transport and inhibit lipolysis in human adipocytes.J Pharmacol Exp Ther. 2001; 297: 563-572PubMed Google Scholar This membrane protein is encoded by the AOC3 gene (amine oxidase copper containing-3) and is also expressed on pericytes and vascular endothelium, where it is involved in leukocyte extravasation to inflamed tissues in different models.18Salmi M Jalkanen S Cell-surface enzymes in control of leukocyte trafficking.Nat Rev Immunol. 2005; 5: 760-771Crossref PubMed Scopus (228) Google Scholar SSAO/VAP-1 protein located on the endothelial cell surface is implicated in rolling, firm adhesion, and transmigration of different types of leukocytes through the endothelial cell barrier to penetrate into tissues.19Stolen CM Marttila-Ichihara F Koskinen K Yegutkin GG Turja R Bono P Skurnik M Hanninen A Jalkanen S Salmi M Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo.Immunity. 2005; 22: 105-115Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar Mice were recently created in which the AOC3 gene was genetically invalidated. They are devoid of VAP-1 protein, SSAO activity, and leukocyte extravasation to diverse sites of experimental inflammation is impaired.19Stolen CM Marttila-Ichihara F Koskinen K Yegutkin GG Turja R Bono P Skurnik M Hanninen A Jalkanen S Salmi M Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo.Immunity. 2005; 22: 105-115Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar Here we asked whether SSAO/VAP-1 could be involved in leukocyte infiltration in WAT since we recently observed that AOC3-KO mice are moderately overweight and have enlarged adipose tissues when compared with their age-matched wild-type controls.20Bour S Prévot D Guigné C Stolen C Jalkanen S Valet P Carpéné C Semicarbazide-sensitive amine oxidase substrates fail to induce insulin-like effects in fat cells from AOC3 knockout mice.J Neural Transm. 2007; 114: 829-833Crossref PubMed Scopus (15) Google Scholar We used wild-type, AOC3-deficient and newly generated fat cell-specific AOC-3 transgenic mice to study the infiltration of leukocytes into WAT. The following results indicate that SSAO/VAP-1 contributes to leukocyte extravasation at the level of adipose tissue because there were fewer immune cells in the WAT of mice invalidated for SSAO/VAP-1, despite their moderate overweight. Therefore, AOC3-deficient mice may serve as a new model of mild obesity, in which fat accumulation is not associated with increased inflammatory cell infiltration. Moreover, they may be valuable tools for future studies on the obesity-related development of diabetes. Mice deficient in SSAO/VAP-1 were produced on a pure 129 background by replacing a portion of the first exon of the mouse AOC3 gene with a neomycin-resistance cassette.19Stolen CM Marttila-Ichihara F Koskinen K Yegutkin GG Turja R Bono P Skurnik M Hanninen A Jalkanen S Salmi M Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo.Immunity. 2005; 22: 105-115Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar AOC3-KO mice, homozygous for the null mutation, were backcrossed onto C57BL/6 for eight generations and compared with wild-type mice with a C57BL/6 background. The aP2hVAP-1TG/KO line was created by crossing the aP2hVAP-1 TG (FBV/n mice expressing human VAP-1 on adipocytes)21Stolen CM Yegutkin GG Kurkijarvi R Bono P Alitalo K Jalkanen S Origins of serum semicarbazide-sensitive amine oxidase.Circ Res. 2004; 95: 50-57Crossref PubMed Scopus (119) Google Scholar to AOC3-KO mice on C57BL/6 background for eight generations. The aP2hVAP-1 transgene, mouse VAP-1 mutant allele (null), and endogenous mouse VAP-1 allele (wt) were all identified by polymerase chain reaction (PCR) screening of purified genomic DNA with specific primers and verified immunohistochemically with human and mouse SSAO/VAP-1 antibodies and enzymatic assays (Amplex Red-based fluorometric detection) as previously described.19Stolen CM Marttila-Ichihara F Koskinen K Yegutkin GG Turja R Bono P Skurnik M Hanninen A Jalkanen S Salmi M Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo.Immunity. 2005; 22: 105-115Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar All of the mice used for this study were handled in accordance with the European Communities Council Directives for experimental animal care. They were housed under specific pathogen-free conditions with constant temperature (20 to 22°C) and with a 12-hour light-dark cycle. All mice had free access to food and water. Glucose tolerance tests were performed on 5-hour-fasted conscious male mice (20 to 25 weeks of age) and glucose was measured on arterio-venous blood from tail vessels using an Accu-Chek glucometer from Roche Diagnostics (Meylan, France). To measure total body fat and lean mass by nuclear magnetic resonance, living female mice were placed into a Plexiglas tubular holder inserted into an Echo MRI NMR machine (100TM3; Echo Medical Systems, Houston, TX) and measured during a period of less than 1 minute. All other comparisons between genotypes were made on mice sacrificed after overnight fasting, at the age of 6 months, unless otherwise stated. Blood samples were immediately used for cell count analyses in an automated analyzer (MICROS-60; Horiba-ABX, Montpellier, France). Epididymal (EPI) and inguinal (ING) WATs were dissected and visible blood vessels were removed. LNs were carefully isolated from ING adipose tissue and used for further analysis. Adipose tissues and LNs were immediately processed for cell isolation or frozen in liquid nitrogen and stored at −80°C until RNA extraction. Adipose tissues were digested in Dulbecco's modified Eagle's medium/F12 (1:1) medium (Invitrogen, Cergy-Pontoise, France) containing 2% bovine serum albumin and 2 mg/ml collagenase (type II collagenase; Sigma-Aldrich, St. Quentin Fallavier, France) for 30 minutes at 37°C. Digested tissues were filtered through a 25-μm mesh nylon membrane. After centrifugation (630 × g for 10 minutes), the pellet from adipose tissues contained a heterogeneous cell population called SVF. Buoyant adipocytes were discarded because their large cell size and fragility and were incompatible with cell sorter analyzer performances. Pelleted cells were washed in Dulbecco's modified Eagle's medium/F12 (1:1) containing 10% newborn calf serum (Sigma-Aldrich). Digested LNs were passed twice through a needle (0.8 mm) before washing. Isolated LNs and cells forming SVF were suspended in Dulbecco's modified Eagle's medium F12 10% newborn calf serum. Red cells were lysed in a buffer containing 155 mmol/L ammonium chloride, 20 mmol/L Tris, pH 7.6, for 5 minutes. Cells were then centrifuged (560 × g for 5 minutes) and resuspended in supernatant from hybridoma-secreting murine monoclonal CD16/32 antibodies (clone 2-4G-2) to block nonspecific Fc receptors before counting on a Coulter counter (Beckman-Coulter, Roissy, France). Staining of lymphocytes and macrophages were performed as previously described.5Caspar-Bauguil S Cousin B Galinier A Segafredo C Nibbelink M André M Casteilla L Pénicaud L Adipose tissues as ancestral immune organ: site-specific change in obesity.FEBS Lett. 2005; 579: 3487-3492Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar, 22Prunet-Marcassus B Cousin B Caton D Andé M Pénicaud L Casteilla L From heterogeneity to plasticity in adipose tissues: site-specific differences.Exp Cell Res. 2006; 312: 727-736Crossref PubMed Scopus (233) Google Scholar Briefly, cells were incubated with conjugated anti-mouse monoclonal antibodies for 20 minutes at room temperature in the dark. Fluorescein isothiocyanate-conjugated anti-CD3 antibodies, phycoerythrin-conjugated NK1.1 and CD115 antibodies, peridinin-chlorophyll-protein complex-conjugated anti-CD45 antibodies, and allophycocyanin-conjugated anti-CD11b monoclonal antibodies were purchased from Becton Dickinson (Le Pont-de-Claix, France). Phycoerythrin-conjugated anti-CD19 monoclonal antibody was obtained from Immunotech (Marseille, France). Cells were then washed in phosphate-buffered saline and analyzed on a fluorescence-activated cell sorter (FACS Calibur, Becton Dickinson). Subpopulations and total lymphocytes were identified by appropriate marker combinations and light scatter properties. Briefly, the combination of phycoerythrin-conjugated anti-NK1.1 and fluorescein isothiocyanate-conjugated anti-CD3 allowed differentiation of three cell populations: NK1.1− CD3+, T cells; NK1.1+CD3−, NK cells; and NK1.1+CD3+, NKT cells. Anti-CD19 was used to sort B cells and CD11b, CD115 to differentiate subsets of macrophages. Data acquisition and analysis were performed with Cell Quest software (Becton Dickinson). Four hundred mg of ING WAT were homogenized in 2 ml of a highly denaturing guanidine-thiocyanate-containing buffer supplied by the RNeasy mini kit (Qiagen, Courtaboeuf, France) plus 20 μl of β-mercaptoethanol. After lipid elimination by chloroform extraction, total RNA was extracted according to the supplier's instructions. Then, 0.5 μg of total RNA was reverse-transcribed using random hexamers and Superscript II reverse transcriptase (Invitrogen, Cergy Pontoise, France). A reaction was performed in parallel without reverse transcriptase (RT−) to estimate genomic DNA contamination. Real-time PCR was performed starting with 6.25 ng of cDNA and both sense and antisense oligonucleotides in a final volume of 20 μl using the SYBR Green TaqMan Universal PCR Mastermix (Eurogentec, Angers, France). Fluorescence was monitored in GeneAmp 7500 detection system instrument (Applied Biosystems, Foster City, CA). Oligonucleotide primers were designed using Primer Express (Perkin-Elmer Life Sciences, Courtaboeuf, France) and verified on Blast Nucleotide software. Primer specificity was checked by the occurrence of a single peak of expected size during dissociation experiments. Analysis of the 18S ribosomal RNA was performed using the Ribosomal RNA Control TaqMan assay kit (Applied Biosystems) to normalize gene expression. Results were expressed as arbitrary units relative to 18S expression, as already reported23Bour S Daviaud D Gres S Lefort C Prévot D Zorzano A Wabitsch M Saulnier-Blache JS Valet P Carpéné C Adipogenesis-related increase of semicarbazide-sensitive amine oxidase and monoamine oxidase in human adipocytes.Biochimie. 2007; 89: 916-925Crossref PubMed Scopus (55) Google Scholar using the following equation: 2(Ct18S − Ct gene "RT+") × [1 − 1/(2{Ct gene "RT-" − Ct gene "RT+"})], where Ct corresponds to the number of cycles needed to generate a fluorescence threshold. For immunohistological analysis, WAT was fixed in zinc buffer and paraffin-embedded sections were prepared. The sections were first incubated with 1 μg/ml of rat anti-mouse CD45, rat anti-VAP-1 (TK10-79 mAb recognizing both human and mouse VAP-1) or class-matched negative control antibody followed by biotinylated anti-rat IgG (Vector Laboratories, Peterborough, UK) in 5% normal mouse serum and streptavidin horseradish peroxidase (Vector Laboratories). 3,3′-Diaminobenzidine was used as a chromogen. The sections were counterstained with hematoxylin. For immunofluorescence stainings the frozen sections were fixed with acetone and stained with monoclonal antibodies recognizing mouse VAP-1 (clone 7-106),24Merinen M Irjala H Salmi M Jaakkola I Hänninen A Jalkanen S Vascular adhesion protein-1 is involved in both acute and chronic inflammation in the mouse.Am J Pathol. 2005; 166: 793-800Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar human VAP-1 (JG2.10),21Stolen CM Yegutkin GG Kurkijarvi R Bono P Alitalo K Jalkanen S Origins of serum semicarbazide-sensitive amine oxidase.Circ Res. 2004; 95: 50-57Crossref PubMed Scopus (119) Google Scholar and a negative control antibody followed by fluorescein isothiocyanate anti-rat IgG (Sigma, St. Louis, MO) containing 5% normal mouse serum. Statistical significance was assessed by the use of nonparametric Kruskal-Wallis one-way analysis for the nonnormal data of flow cytometry, which are reported as mean ± SD. For all other parameters, means are given ± SEM and Student's t-test was used to compare genotypes. A P value of less than 0.05 was considered significant for all tests; n denotes the number of cases. The body weights of 28-week-old males were significantly higher for AOC3-KO mice than that for wild-type C57BL/6 mice (KO: 37.4 ± 1.2 g, n = 15; and wild-type: 32.4 ± 0.8 g, n = 12; P < 0.001). An increase in epididymal (EPI) and inguinal (ING) WAT weight was found in male AOC3-KO mice when compared with wild-type mice (Table 1). In ING, the enlarged tissue was characterized by a higher level of lipid content (KO: 728 ± 12 mg lipid/g tissue versus wild-type: 661 ± 18 mg lipid/g tissue, n = 8; P < 0.01) and a reduced protein content (KO: 13.1 ± 1.6 mg protein/g tissue versus wild-type: 19.3 ± 1.6 mg protein/g tissue, n = 8; P < 0.05), as generally observed in most forms of murine obesities.25Valet P Tavernier G Castan-Laurell I Saulnier-Blache JS Langin D Understanding adipose tissue development from transgenic animal models.J Lipid Res. 2002; 43: 835-860Abstract Full Text Full Text PDF PubMed Google Scholar As a consequence, the lipid-to-protein ratio was significantly increased in ING depot of AOC3-KO mice (58.3 ± 5.0 versus 35.9 ± 2.9 for KO and wild-type mice, respectively; n = 8, P < 0.02). Although fatter, the AOC3-KO mice were normoglycemic (fasting blood glucose was 108 ± 5 mg/dl, ie, not significantly different from wild type: 98 ± 5 mg/dl, n = 12) and did not exhibit alteration of glucose tolerance (area under the curve of the hyperglycemic response to glucose challenge (1 g/kg, i.p.) was 4379 ± 575 and 4920 ± 157 arbitrary units in KO and wild type, respectively; n = 12). Accordingly, we have previously reported that the dose-response curves to insulin stimulation of glucose transport are superimposable in adipocytes isolated from KO and wild-type mice.20Bour S Prévot D Guigné C Stolen C Jalkanen S Valet P Carpéné C Semicarbazide-sensitive amine oxidase substrates fail to induce insulin-like effects in fat cells from AOC3 knockout mice.J Neural Transm. 2007; 114: 829-833Crossref PubMed Scopus (15) Google ScholarTable 1Adipose Depot Weight and CD45-Positive Cells in Stroma-Vascular Fraction of Wild-Type (WT) or AOC3-KO Male MiceWTAOC3-KOWeight (g)CD45+ (cells/g)Weight (g)CD45+ (cells/g)EPI WAT1.04 ± 0.12416,182 ± 105,5391.44 ± 0.08*P < 0.05 and79,166 ± 11,074*P < 0.05 andING WAT1.05 ± 0.12566,346 ± 124,6361.85 ± 0.20†P < 0.001 for the difference between WT and AOC3-KO mice.183,490 ± 26,915*P < 0.05 andThe number of CD45+ cells was expressed per g of tissue. Means ± SEM of 5 to 15 control (WT) and 4 to 12 AOC3-KO mice.* P < 0.05 and† P < 0.001 for the difference between WT and AOC3-KO mice. Open table in a new tab The number of CD45+ cells was expressed per g of tissue. Means ± SEM of 5 to 15 control (WT) and 4 to 12 AOC3-KO mice. Among the cells that constitute the SVF of WAT (endothelial cells, fibroblasts, leukocytes, preadipocytes), we focused on leukocytes (CD45-positive cells) and compared them to the cell population (mainly lymphocytes) of the inguinal LN as a reference immune organ. CD45-positive cells (CD45+) were analyzed by flow cytometry according to their size and granulosity. As shown in Figure 1A, CD45+ cells present in SVF exhibited heterogeneous size and granulosity whereas CD45+ cells from LN were more homogeneous. In control mice, cells expressing CD45 represented 39 ± 6% and 56 ± 7% of the SVF cells counted in EPI and ING fat tissues, respectively (Figure 1B). These proportions were strongly reduced in AOC3-KO male mice because CD45+cells represented only 9 ± 1% and 16 ± 2% of SVF cells in EPI and ING. By contrast, in both genotypes, the leukocytes accounted for more than 96% of the cells found in LN (Figure 1B). The reduction of the abundance of CD45+ cells in WAT from AOC3-KO mice was also marked when results were expressed as number of cells per g of tissue, as shown in Table 1. Among SVF cells, two CD45-positive populations with different fluorescence intensities for CD45 could be distinguished (Figure 1C). This allowed us to define two regions, R1 and R2, corresponding, respectively, to CD45-high and CD45-low populations. As previously described, gate R1 was defined according to size and granulosity of cells present in LN and mainly contains lymphocytes.5Caspar-Bauguil S Cousin B Galinier A Segafredo C Nibbelink M André M Casteilla L Pénicaud L Adipose tissues as ancestral immune organ: site-specific change in obesity.FEBS Lett. 2005; 579: 3487-3492Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar Macrophages are present in the gate R2.22Prunet-Marcassus B Cousin B Caton D Andé M Pénicaud L Casteilla L From heterogeneity to plasticity in adipose tissues: site-specific differences.Exp Cell Res. 2006; 312: 727-736Crossref PubMed Scopus (233) Google Scholar The two populations were found in all adipose depots from wild-type or AOC3-KO mice. Noteworthy, the proportions of both lymphocytes (R1) and macrophages (R2) were strongly reduced in the SVF of ING and EPI WAT of AOC3-KO male mice (Figure 1C). In whole blood, the white blood cell number remained unchanged between wild-type and AOC3-KO (6.4 ± 0.6 and 5.8 ± 0.8 103 cells/μl, respectively; n = 8). A small but significant weight increase also occurred in AOC3-KO female mice because the body mass was 21.6 ± 0.6 g and 24.6 ± 1.2 g in 6-month-old wild-type and AOC3-KO, respectively (n = 11 and 9, P < 0.05). All of the dissected AOC3-KO fat depots tended to be heavier although significance was not reached in any, eg, 0.25 ± 0.05 g versus 0.59 ± 0.15 g for parametrial WAT or 0.29 ± 0.04 g versus 0.61 ± 0.18 g for ING (n = 11 and 9). Body composition measurements using nuclear magnetic resonance indicated that total fat mass increased from 8.9 ± 1.0% in wild-type to 11.4 ± 0.7% in AOC3-KO when expressed as percentage of body mass (n = 11 and 17, P < 0.05). A reduction of the abundance of CD45+ cells also occurred in WAT from SSAO/VAP-1-deficient female mice, decreasing from 42 ± 2% in wild-type to 15 ± 4% in AOC3-KO. A decrease of lymphocyte abundance (R1) was detected in ING of 9-week-old AOC3-KO females (12 ± 1% versus 39 ± 2% in age-matched control; n = 6, P < 0.001). Taken together, these observations indicated that a defect in leukocyte presence occurred in WAT of AOC3-KO mice independent of age or gender. A complete lack of AOC3 expression was found in the ING WAT by real-time PCR experiments (not shown), therefore confirming the total disruption of the gene encoding SSAO/VAP-1. A reduction of CD45 expression was observed in the mRNAs prepared from ING adipose tissue of 28-week-old male AOC3-KO mice (Figure 2A). This was in accordance with the reduction of leukocyte number in adipose depot SVF reported above. The expression of the chemoattractant protein MCP-1 was reduced by 51% in the subcutaneous adipose tissue of AOC3-KO mice, when compared with wild-type mice (Figure 2B). The expression of leptin, an adipokine known to possess pro-inflammatory action exhibited a clear increase in ING WAT of AOC3-KO mice (Figure 2C). This increased leptin expression is in agreement with the enhanced fat deposition found in these mice. The fact that AOC3-KO mice demonstrated reduced leukocyte infiltration suggests that the adhesion defects in the absence of SSAO/VAP-1 dominate over the recognized pro-inflammatory functions of leptin. The reduction of leukocyte presen
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