Leptin and Adiponectin Stimulate the Release of Proinflammatory Cytokines and Prostaglandins from Human Placenta and Maternal Adipose Tissue
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内分泌学杂志 2005年第8期
Department of Obstetrics and Gynecology (M.L., M.P.), University of Melbourne and Mercy Perinatal Research Centre, Mercy Hospital for Women, East Melbourne 3002, Victoria, Australia; and Translational Proteomics (G.E.R.), Baker Medical Research Institute, Baker Heart Research Institute, Melbourne 3004, Victoria, Australia
Address all correspondence and requests for reprints to: Martha Lappas, Department of Obstetrics and Gynaecology, University of Melbourne, Mercy Hospital for Women, Fourth Floor, 163 Studley Road, Heidelberg 3084, Victoria, Australia. E-mail: mlappas@unimelb.edu.au.
Abstract
Beyond their effects on central metabolic functions, leptin, resistin, and adiponectin have profound effects on a number of other physiologic processes, including immune function and inflammation. Although leptin, resistin, and adiponectin are produced in human placenta and adipose tissue, their immunoregulatory actions in these tissues are not known. Therefore, the aim of this study was to determine the effect of leptin, resistin, and adiponectin on the release of proinflammatory mediators in human placenta and sc adipose tissue. Samples were obtained from normal pregnancies at the time of cesarean section. Tissue explants (n = 5) were incubated in the absence (basal control) or presence of a leptin (1, 10, and 100 ng/ml), resistin (1, 10, and 100 ng/ml), and adiponectin (0.1 and 0.5 μg/ml). After 6 h incubation, the medium was collected, and the release of IL-1?, IL-6, TNF, prostaglandin (PG)F2 and PGE2 was quantified by ELISA. There was no effect of resistin on proinflammatory cytokine or prostaglandin release; however, leptin at 100 ng/ml and adiponectin at 0.1 and/or 0.5 μg/ml significantly increased the release of IL-1?, IL-6, TNF, and PGE2 from human placenta and adipose tissue. Although both leptin and adiponectin significantly increased PGF2 release from human placenta, there was no effect of these hormones on PGF2 release from adipose tissue. Furthermore, this leptin- and adiponectin-induced proinflammatory response could be abrogated by treatment with the antiinflammatory ERK1/2 MAPK inhibitor U0126, the peroxisomal proliferator-activated receptor- ligand troglitazone, and the nuclear factor-B inhibitor BAY 11-7082. Collectively, these data indicate that leptin and adiponectin activate proinflammatory cytokine release and phospholipid metabolism in human placenta and adipose tissue, and antiinflammatory agents can abrogate leptin- and adiponectin-induced inflammation.
Introduction
HUMAN PLACENTA AND adipose tissue have been reported to produce and secrete various proinflammatory factors including cytokine such as IL-1?, IL-6, TNF, and prostaglandins (PGs) such as PGE2 and PGF2. These inflammatory mediators have been demonstrated to play an important role in a number of normal and abnormal inflammatory processes, including the initiation and progression of human labor (reviewed in Refs. 1 and 2), obesity, and diabetes (reviewed in Refs. 3 and 4). We and others have demonstrated that the expression of these cytokines and prostaglandins in placenta and adipose tissue is regulated by a number of immune modulators, including lipopolysaccharide (LPS) and TNF (5). Recent evidence suggests that the hormones leptin, adiponectin, and resistin, which are released by both human placenta and adipose tissue, may also exhibit immunomodulatory functions (reviewed in Ref. 6) and therefore may be involved in many processes of normal and abnormal physiology. However, to date there is a paucity of data available on the regulatory role of leptin, resistin, and adiponectin on the release of proinflammatory cytokines and prostaglandins in human placenta and adipose tissue.
Leptin is a 16-kDa polypeptide hormone that is coded by the obese (ob) gene. In addition to regulating energy homeostasis, leptin is also a crucial hormone/cytokine for a number of diverse physiological processes such as inflammation, immune function, reproduction, and angiogenesis (reviewed in Ref. 6). The importance of leptin in immunity is evident in mice having a homozygous mutation in their leptin gene or receptor. These mice have impaired T-cell-mediated responses (7). In addition, leptin promotes helper T-cell (Th) activity in vitro, which coordinates most immune responses. In particular, leptin increases the proliferation of naive T cells, increases Th1 [interferon (IFN) and IL-2] and suppresses Th2 (IL-4) cytokine production (8). The biological actions of leptin are carried out through interaction with its specific surface receptor OB-R, resulting in the activation of divergent signal transduction pathways, such as the MAPK, nuclear factor-B (NF-B), and signal transducers and activators of transcription (STAT), which eventually induce expression of several genes involved in transcriptional regulation and signaling (9).
Adiponectin, in addition to its role in the modulation of glucose and lipid metabolism, has been reported to have potent antisuppressive properties due to its ability to induce the production of antiinflammatory cytokines, and inhibit the production of proinflammatory cytokines (10, 11). Circulating adiponectin exists in several isoforms, including trimers, hexamers, and high-molecular-weight multimers (12). Adiponectin contains three domains: an N-terminal signal sequence, a collagen domain, and a C-terminal globular domain. Proteolytic processing of adiponectin generates a truncated 16.5-kDa protein, gAcrp30, which corresponds to the entire C-terminal globular domain of adiponectin. gAcrp30 possesses unique pharmacological properties that are not shared with full-length adiponectin (13).
Resistin is a member of a family of tissue-specific signaling molecules called resistin-like molecules (reviewed in Ref. 14). Three physiological roles have been proposed for resistin: a mediator of regulation of metabolism, a regulator of adipogenesis, and a relationship to inflammation (reviewed in Ref. 14). The amino acid sequence of resistin is identical with the sequence of a family of proteins that are induced during lung inflammation, known as found in inflammatory zone (15). Although very little is known about found in inflammatory zone proteins, their pattern of expression and physiologic functions resemble that of other well-known proinflammatory cytokines such as IL-6 and TNF. Taken together, these data suggest that resistin may be involved in a number of inflammatory processes.
Leptin, resistin, and adiponectin, which are released by human placenta and adipose tissue, are cytokine-like peptide hormones that act as immune regulators; however, little is known about their immunomodulatory actions in placenta and adipose tissue. We therefore hypothesize that leptin, resistin, and adiponectin might exhibit cytokine-like properties in human placenta and adipose tissue. Therefore, the main purpose of this study was to investigate the effect of leptin, resistin, and adiponectin on proinflammatory cytokine release and phospholipid metabolism from human placenta and sc adipose tissue obtained from normal pregnant women. In addition, the role of the signaling pathways involving NF-B, peroxisomal proliferator-activated receptor (PPAR)-, and MAPK involved in this regulation was also investigated.
Materials and Methods
Reagents
All chemicals were purchased from BDH Chemicals Australia (Melbourne, Victoria, Australia) unless otherwise stated. RPMI 1640 (glucose free) was obtained from Life Technologies, Inc. Laboratories (Grand Island, NY). BSA (RIA grade), ?-NADH (disodium salt), 3,3',5,5'-tetramethylbenzidine, and pyruvic acid (dimer free) were supplied by Sigma (St. Louis, MO). The leptin ELISA kit was supplied by Biosource International (Camarillo, CA). The IL-1?, IL-6, TNF, and adiponectin kits were purchased from R & D Systems (Minneapolis, MN). The PGE2 and PGF2 enzyme immunoassay kits were supplied from Cayman Chemical Co. (Ann Arbor, MI). The resistin ELISA kit, the active fragment of adiponectin gAcrp30, human recombinant leptin, and human recombinant resistin were purchased from CytoLab (Rehovot, Israel). (E)-3-[4-Methylphenylsulfonyl]-2-propenenitrile (BAY 11-7082) was purchased from Sapphire Bioscience (Crows Nest, New South Wales, Australia). Troglitazone was generously provided by Sankyo (Japan, Tokyo), and U0126 was purchased from Tocris (Ellisville, MO).
Patients and sample collection
Human placenta and sc adipose tissue (from the anterior abdominal wall) were obtained from pregnant women who delivered healthy, singleton infants at term (37 wk gestation) undergoing elective cesarean section (indications for cesarean section were breech presentation and/or previous cesarean section). Approval for this study was obtained from the Mercy Hospital for Women’s Research and Ethics Committee, and informed consent was obtained from all participating subjects.
Tissue explants
Tissues were obtained within 10 min of delivery and dissected fragments were placed in ice-cold PBS as previously described (5, 16, 17). Briefly, tissues were dissected to remove visible connective tissue, vessels and/or calcium deposits. Tissue fragments were placed in RPMI 1640 (placenta) or DMEM (adipose tissue) containing 5 mM glucose at 37 C in a humidified atmosphere of 8% O2-5% CO2 for 1 h. Explants were blotted dry on sterile filter paper and transferred to 24-well tissue culture plates (100–200 mg wet weight per well). Placental explants (n = 5) were incubated, in duplicate, in 2 ml medium containing penicillin G (100 U/ml) and streptomycin (100 μg/ml), in the absence (basal release) or presence of leptin (1, 10, and 100 ng/ml), resistin (1, 10, and 100 ng/ml), and adiponectin (0.1 and 0.5 μg/ml). Due to the limited amount of tissue sample, only maximum concentrations of leptin (100 ng/ml), resistin (100 ng/ml), and adiponectin (0.5 μg/ml) were used for adipose tissue explants (n = 5). Placental explants (n = 3) were also cotreated with 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126. After 6 h incubation, tissues were collected and assayed for total protein, whereas the incubation medium was collected and assayed for TNF, IL-1?, IL-6, PGF2, PGE2, leptin, resistin, and adiponectin release by ELISA.
Validation of explant cultures and viability
To determine the effect of experimental treatment on cell membrane integrity, the release of the intracellular enzyme lactate dehydrogenase (LDH) into incubation medium was determined as described previously (5, 16, 17). LDH release was investigated over the 6 h time course of tissue explants (n = 3). In vitro incubation did not significantly affect LDH activity in the incubation medium, with none of these measurements exceeding 7% of total activity present in the tissue (data not shown). These data indicate that the concentrations of leptin, resistin, and adiponectin used in this study did not affect cell viability.
Experimental assays
The release of TNF, IL-1?, IL-6, PGE2, PGF2, leptin, resistin, and adiponectin into the explant incubation medium was performed by sandwich ELISA according to the manufacturers’ instructions. The intraassay and interassay coefficients of variation for the leptin ELISA was 3.4 and 4.5%, respectively, and the minimum detectable limit of the assay was 7.2 pg/ml. The intraassay and interassay coefficients of variation for the resistin ELISA was 3.4 and 4.5%, respectively, and the minimum detectable limit of the assay was 16 pg/ml. The intraassay and interassay coefficients of variation for the adiponectin ELISA was 4.2 and 4.7%, respectively, and the minimum detectable limit of the assay was 32 pg/ml. All tissue release data were corrected for total protein. The protein content of tissue homogenates was determined using BCA protein assay (Pierce, Rockford, IL), using BSA as a reference standard, as previously described (5, 16, 17).
Statistical analysis
Statistical analyses were performed using a commercially available statistical software package (Statgraphics, StatPoint, Inc., Herndon, VA). Homogeneity of data was assessed by Bartlett’s test, and when significant, data were logarithmically transformed before further analysis. The effect of experimental treatment on cytokine and prostaglandin release was analyzed by multivariate ANOVA. The method used to discriminate among the means is Fisher’s least significant difference procedure. Statistical difference was indicated by P < 0.05. Data are expressed as mean ± SEM.
Results
Effect of leptin, resistin, and adiponectin on leptin, resistin, and adiponectin release
Placental and adipose tissues were incubated in the presence of 1, 10, and/or 100 ng/ml of leptin; 1, 10, and/or 100 ng/ml resistin; and 0.1 and/or 0.5 μg/ml adiponectin and the release of leptin, resistin, and adiponectin measured by ELISA. Leptin, at all concentrations tested in this study, had no effect on resistin or adiponectin release from either placenta or adipose tissue (data not shown). Likewise, placental and adipose tissue release of leptin and adiponectin were unaffected by treatment with resistin (data not shown). Furthermore, addition of adiponectin to placenta and adipose tissue explants did not affect the release of leptin or resistin (data not shown).
Effect of leptin, resistin, and adiponectin on proinflammatory cytokine release
Compared with control, tissues incubated in the presence of 100 ng/ml leptin caused a significant increase in the release of IL-1?, IL-6, and TNF from human placenta (Fig. 1, A–C, respectively). Resistin, at all concentrations tested in this study, had no effect on proinflammatory cytokine release from placenta (data not shown). Adiponectin, at both 0.1 and 0.5 μg/ml, increased IL-1?, IL-6, and TNF release from human placenta (Fig. 2, A–C, respectively). Similarly in adipose tissue, IL-1? (Fig. 3), IL-6 (Fig. 4), and TNF (Fig. 5) release was significantly increased by treatment with 100 ng/ml leptin and 0.5 μg/ml adiponectin but not 100 ng/ml resistin.
FIG. 1. Effect of 1, 10, and 100 ng/ml leptin on the release of IL-1? (A), IL-6 (B), TNF (C), PGE2 (D), and PGF2 (E) from human placenta (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal cytokine release from placenta.
FIG. 2. Effect of 0.1 and 0.5 μg/ml adiponectin on the release of IL-1? (A), IL-6 (B), TNF (C), PGE2 (D), and PGF2 (E) from human placenta (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal cytokine release from placenta.
FIG. 3. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of IL-1? from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal IL-1? release from adipose tissue.
FIG. 4. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of IL-6 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal IL-6 release from adipose tissue.
FIG. 5. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of TNF from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal TNF release from adipose tissue.
Effect of leptin, resistin, and adiponectin on PG release
Placental tissues incubated in the presence of 100 ng/ml leptin significantly increased the release of PGE2 (Fig. 1D). Compared with control, incubation of placenta with 10 and 100 ng/ml leptin caused a significant increase in the release of PGF2 (Fig. 1E). Resistin, at all concentrations tested in this study, had no effect on PG release from placenta (data not shown). Adiponectin, at 0.5 μg/ml, significantly increased PGE2 (Fig. 2D) and PGF2 (Fig. 2E) release from human placenta. Both leptin and adiponectin significantly increased PGE2 release from adipose tissue (Fig. 6). Although leptin and adiponectin increased PGF2 release from adipose tissue, this failed to reach significance (Figs. 7). There was no effect of resistin on PGE2 (Fig. 6) and PGF2 release (Fig. 7).
FIG. 6. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of PGE2 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal PGE2 release from adipose tissue.
FIG. 7. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of PGF2 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal PGF2 release from adipose tissue.
Effect of U0126, BAY 11-7082, and troglitazone on leptin- and adiponectin-induced proinflammatory cytokine release
To determine the intracellular pathways involved in leptin- and adiponectin-induced proinflammatory cytokine release, placental explants were cotreated with the NF-B inhibitor BAY 11-7082, the PPAR ligand troglitazone, and the ERK1/2 MAPK inhibitor U0126. Treatment of placental explants with 50 μM BAY 11-7082, 30 μM troglitazone, or 10 μM U0126 significantly inhibited leptin (100 ng/ml), and adiponectin (0.5 μg/ml) induced IL-1? (Figs. 8A and 9A), IL-6 (Figs. 8B and 9B), and TNF (Figs. 8C and 9C) release.
FIG. 8. Effect of 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126 on 100 ng/ml leptin-stimulated IL-1? (A), IL-6 (B), and TNF (C) release from human placenta (n = 3). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal release; , P < 0.05 vs. leptin- stimulated release.
FIG. 9. Effect of 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126 on 0.5 μg/ml adiponectin-stimulated IL-1? (A), IL-6 (B), and TNF (C) release from human placenta (n = 3). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal release; , P < 0.05 vs. adiponectin-stimulated release.
Discussion
It has become increasingly apparent that leptin, resistin, and adiponectin have pleiotropic actions on various cell types beyond their effects on regulating energy homeostasis. In light of the involvement of leptin, resistin, and adiponectin in immune and inflammatory response, we employed a well-characterized in vitro human tissue explant system to examine their potential inflammatory effects in placenta and adipose tissue. Our data show that leptin and adiponectin, but not resistin, have proinflammatory actions, involving the stimulation of proinflammatory cytokines TNF, IL-1?, IL-6, and prostaglandins PGE2 and PGF2 from human placenta and adipose tissue. Our studies further aimed to investigate which intracellular signaling pathways are activated by leptin and adiponectin. We found that BAY 11-7082, troglitazone, and U0126 treatment suppressed leptin- and adiponectin-stimulated IL-1?, IL-6, and TNF release from human placenta, suggesting that NF-B, PPAR, and ERK 1/2 MAPK all appeared to be key transcription factors involved in leptin- and adiponectin-induced proinflammatory response.
Leptin directly regulates the production of several cytokines both in vivo and in vitro (reviewed in Refs. 18, 19, 20, 21). Leptin has been implicated in stimulating Th1 cytokines in rodents (8); increasing LPS-stimulated production of TNF, IL-6, and IL-12 in murine peritoneal macrophages and human monocytes (22, 23); and stimulating IL-6 secretion by human term trophoblast cells (24) and human placenta (25). Likewise, in this study, leptin increased the secretion of TNF, IL-1?, and IL-6 from human placenta and adipose tissue.
Leptin has also been shown to exert proinflammatory effects by stimulating the phospholipid metabolizing pathway. Leptin stimulates PGE2 and PGF2 production in neonatal rat hypothalamus (26); augments alveolar macrophage leukotriene synthesis by increasing phospholipase A2 (PLA2) activity and enhancing cytosolic PLA2 protein expression (27); up-regulates cytosolic PLA2 activity in human bone marrow stromal cells (28); and potentiates IFN-induced expression of cyclooxygenase-2 and PGE2 production from murine macrophages J774A.1 (29). In this study, we also demonstrated that treatment of placenta and adipose tissue with leptin resulted in an increased release of the proinflammatory prostaglandins PGE2 and PGF2.
Placenta and adipose tissue have both the long and the short isoforms of the leptin receptor. The Ob-Rb receptor, which is widely expressed in human placenta and adipose tissue, mediates leptin action by signaling via the Janus kinase/STAT pathway (30) including the downstream suppressor of cytokine signaling-3 feedback inhibitor and/or other common pathways such as the p38 MAPK, p42/p44 MAPK (28, 31, 32, 33, 34), and NF-B pathway (9, 31, 35). Cauzac et al. (36) demonstrated that transduction of leptin growth signals in placental cells is independent of Janus kinase/STAT activation; however, leptin activated the ERK 1/2 MAPK cascade. Likewise, in this study, we demonstrated that leptin-stimulated proinflammatory cytokine release from human placenta could be abrogated by treatment with the antiinflammatory ERK 1/2 MAPK inhibitor U0126, the PPAR ligand troglitazone, and the NF-B inhibitor BAY 11-7082. This is also in agreement with our previous studies, demonstrating the importance of NF-B and PPAR in the regulation of TNF and IL-6 release from human placenta (16, 17) and adipose tissue (37).
Although there are ample data available on the effect of proinflammatory mediators on the expression and release of resistin, to date, few data are available on the effect of resistin on proinflammatory cytokines and PGs. The available data do, however, suggest a proinflammatory role for resistin. In particular, treatment with 30 ng/ml resistin results in elevated secretion of IL-6 and TNF from human sc adipocytes (38). Furthermore, resistin promotes smooth muscle cell proliferation through activation of ERK 1/2 but not p38 MAPK (39). This is in contrast to the findings of this study, in which there was no effect of resistin (1–100 ng/ml) on the release of proinflammatory cytokines and PGs from human placenta and adipose tissue.
Adiponectin is postulated to be an antiinflammatory cytokine. It induces the antiinflammatory cytokines IL-10 in human leukocytes (10) and porcine macrophages (11), whereas adiponectin suppresses the production of the proinflammatory cytokine IFN in human macrophages (10); attenuates LPS-stimulated TNF and IL-6 release from porcine macrophages (11); and decreases LPS-induced hepatic TNF mRNA expression and release in KK-Ay obese mice (40). Furthermore, elevated TNF levels have been reported in the adiponectin knockout mouse, and the introduction of adiponectin by adenoviral infection normalized serum TNF in these mice (41). These antiinflammatory actions of adiponectin are mediated in part by suppression of NF-B signaling (11, 42) and ERK 1/2 MAPK activity (11).
In this study, adiponectin exerted proinflammatory effects by increasing the release of proinflammatory cytokines and PGs from human placenta and adipose tissue. This is in agreement with previous studies (43, 44) that demonstrated that adiponectin caused elevated expression of cyclooxygenase-2 and induced PGE2 release from stromal preadipocytes, and results of Schaffler et al. (45) demonstrated that adiponectin induced the release of IL-6 and matrix metalloproteinase-1 in synovial fibroblasts. Like leptin, the proinflammatory actions of adiponectin are mediated through a number of signaling pathways, including p38 MAPK (45) and NF-B (12). Similarly, in this study, adiponectin-induced proinflammatory response could be abrogated by treatment with the antiinflammatory ERK1/2 MAPK inhibitor U0126, the PPAR ligand troglitazone, and the NF-B inhibitor BAY 11-7082. Although the transcriptional regulation of proinflammatory cytokine gene expression in response to leptin and adiponectin treatment is not known, these findings are consistent with human IL-1?, IL-6, and TNF promoters containing response elements for a number of transcription factors, including NF-B, activator protein-1, and specificity protein-1 (reviewed in Ref. 46).
There are many possible reasons between the conflicting results between our studies and previous studies with respect to the response elicited upon resistin or adiponectin treatment. First, this may represent differences in tissue or cell-specific functions: (1) tissue explants are not comprised of just one cell type and paracrine mediators produced by, for example, nonadipocyte cells present in adipose tissue (such as immunocytes and vascular cells) may be necessary for adiponectin to stimulate proinflammatory cytokine release; (2) the explant system and the preservation of the extracellular matrix may provide a more natural environment that allows placental cells and adipocytes to respond to stimuli as they would in vivo; and (3) the conditions that are used to prepare and culture cells from may select for a subset of cells that respond to resistin and adiponectin in a different fashion than do mature fat cells or trophoblasts. Nevertheless, our studies are the first demonstration of such activities in placenta and adipose tissue. Second, the differences observed may also be due to different physiological functions of bacterially produced full-length adiponectin, the globular domain of adiponectin (gAcrp30), or the full-length adiponectin produced by mammalian cells. And third, it may also represent differences in the concentrations used. For example, in this study, adiponectin was used at 0.1 and 0.5 μg/ml, whereas others have used concentrations of adiponectin as high as 50 μg/ml (42).
The form of adiponectin used in this study was gAcrp30, which corresponds to the entire C-terminal globular domain of adiponectin; however, circulating adiponectin exists in several isoforms, including trimers, hexamers, and high-molecular-weight multimers (12). Therefore, it is uncertain whether the effects observed in human placenta and adipose tissue are purely pharmacological or whether they are reflective of a physiological aspect of adiponectin function. Future studies examining the effects of all forms of adiponectin will further contribute to our understanding of the role of this protein in regulating the inflammatory response.
In this study, leptin and adiponectin exerted proinflammatory actions in human placenta and adipose tissue, and this may therefore have important implications in a number of normal and abnormal physiological processes. Prostaglandins and proinflammatory cytokines that are produced within the intrauterine environment participate in the regulation of myometrial contractility, cervical ripening and rupture of membranes (reviewed in Refs. 1 and 2). Therefore, leptin and adiponectin, both of which are produced within the intrauterine environment, may play a role in the processes of human labor and delivery. Furthermore, these proinflammatory actions of leptin and adiponectin may also have significant implications in obesity and diabetes. The production of leptin and/or adiponectin within adipose tissue may contribute to the raised circulating levels of proinflammatory mediators that are evident in these disease states.
In summary, an in vitro tissue explant model system was used to investigate the inflammatory actions of leptin, resistin, and adiponectin on the release of proinflammatory mediators from placenta and adipose tissue. These data demonstrate that proinflammatory mediators in placenta and adipose tissue represent targets for up-regulation by leptin and adiponectin and provide mechanisms by which these hormones can promote inflammatory responses. These data have extended our knowledge of the factors involved in the regulation of proinflammatory cytokines and PGs.
Acknowledgments
The authors gratefully acknowledge the assistance of the clinical research midwives Val Bryant and Sarah Mitchell and the obstetrics and midwifery staff of the Mercy Hospital for Women for their cooperation.
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Address all correspondence and requests for reprints to: Martha Lappas, Department of Obstetrics and Gynaecology, University of Melbourne, Mercy Hospital for Women, Fourth Floor, 163 Studley Road, Heidelberg 3084, Victoria, Australia. E-mail: mlappas@unimelb.edu.au.
Abstract
Beyond their effects on central metabolic functions, leptin, resistin, and adiponectin have profound effects on a number of other physiologic processes, including immune function and inflammation. Although leptin, resistin, and adiponectin are produced in human placenta and adipose tissue, their immunoregulatory actions in these tissues are not known. Therefore, the aim of this study was to determine the effect of leptin, resistin, and adiponectin on the release of proinflammatory mediators in human placenta and sc adipose tissue. Samples were obtained from normal pregnancies at the time of cesarean section. Tissue explants (n = 5) were incubated in the absence (basal control) or presence of a leptin (1, 10, and 100 ng/ml), resistin (1, 10, and 100 ng/ml), and adiponectin (0.1 and 0.5 μg/ml). After 6 h incubation, the medium was collected, and the release of IL-1?, IL-6, TNF, prostaglandin (PG)F2 and PGE2 was quantified by ELISA. There was no effect of resistin on proinflammatory cytokine or prostaglandin release; however, leptin at 100 ng/ml and adiponectin at 0.1 and/or 0.5 μg/ml significantly increased the release of IL-1?, IL-6, TNF, and PGE2 from human placenta and adipose tissue. Although both leptin and adiponectin significantly increased PGF2 release from human placenta, there was no effect of these hormones on PGF2 release from adipose tissue. Furthermore, this leptin- and adiponectin-induced proinflammatory response could be abrogated by treatment with the antiinflammatory ERK1/2 MAPK inhibitor U0126, the peroxisomal proliferator-activated receptor- ligand troglitazone, and the nuclear factor-B inhibitor BAY 11-7082. Collectively, these data indicate that leptin and adiponectin activate proinflammatory cytokine release and phospholipid metabolism in human placenta and adipose tissue, and antiinflammatory agents can abrogate leptin- and adiponectin-induced inflammation.
Introduction
HUMAN PLACENTA AND adipose tissue have been reported to produce and secrete various proinflammatory factors including cytokine such as IL-1?, IL-6, TNF, and prostaglandins (PGs) such as PGE2 and PGF2. These inflammatory mediators have been demonstrated to play an important role in a number of normal and abnormal inflammatory processes, including the initiation and progression of human labor (reviewed in Refs. 1 and 2), obesity, and diabetes (reviewed in Refs. 3 and 4). We and others have demonstrated that the expression of these cytokines and prostaglandins in placenta and adipose tissue is regulated by a number of immune modulators, including lipopolysaccharide (LPS) and TNF (5). Recent evidence suggests that the hormones leptin, adiponectin, and resistin, which are released by both human placenta and adipose tissue, may also exhibit immunomodulatory functions (reviewed in Ref. 6) and therefore may be involved in many processes of normal and abnormal physiology. However, to date there is a paucity of data available on the regulatory role of leptin, resistin, and adiponectin on the release of proinflammatory cytokines and prostaglandins in human placenta and adipose tissue.
Leptin is a 16-kDa polypeptide hormone that is coded by the obese (ob) gene. In addition to regulating energy homeostasis, leptin is also a crucial hormone/cytokine for a number of diverse physiological processes such as inflammation, immune function, reproduction, and angiogenesis (reviewed in Ref. 6). The importance of leptin in immunity is evident in mice having a homozygous mutation in their leptin gene or receptor. These mice have impaired T-cell-mediated responses (7). In addition, leptin promotes helper T-cell (Th) activity in vitro, which coordinates most immune responses. In particular, leptin increases the proliferation of naive T cells, increases Th1 [interferon (IFN) and IL-2] and suppresses Th2 (IL-4) cytokine production (8). The biological actions of leptin are carried out through interaction with its specific surface receptor OB-R, resulting in the activation of divergent signal transduction pathways, such as the MAPK, nuclear factor-B (NF-B), and signal transducers and activators of transcription (STAT), which eventually induce expression of several genes involved in transcriptional regulation and signaling (9).
Adiponectin, in addition to its role in the modulation of glucose and lipid metabolism, has been reported to have potent antisuppressive properties due to its ability to induce the production of antiinflammatory cytokines, and inhibit the production of proinflammatory cytokines (10, 11). Circulating adiponectin exists in several isoforms, including trimers, hexamers, and high-molecular-weight multimers (12). Adiponectin contains three domains: an N-terminal signal sequence, a collagen domain, and a C-terminal globular domain. Proteolytic processing of adiponectin generates a truncated 16.5-kDa protein, gAcrp30, which corresponds to the entire C-terminal globular domain of adiponectin. gAcrp30 possesses unique pharmacological properties that are not shared with full-length adiponectin (13).
Resistin is a member of a family of tissue-specific signaling molecules called resistin-like molecules (reviewed in Ref. 14). Three physiological roles have been proposed for resistin: a mediator of regulation of metabolism, a regulator of adipogenesis, and a relationship to inflammation (reviewed in Ref. 14). The amino acid sequence of resistin is identical with the sequence of a family of proteins that are induced during lung inflammation, known as found in inflammatory zone (15). Although very little is known about found in inflammatory zone proteins, their pattern of expression and physiologic functions resemble that of other well-known proinflammatory cytokines such as IL-6 and TNF. Taken together, these data suggest that resistin may be involved in a number of inflammatory processes.
Leptin, resistin, and adiponectin, which are released by human placenta and adipose tissue, are cytokine-like peptide hormones that act as immune regulators; however, little is known about their immunomodulatory actions in placenta and adipose tissue. We therefore hypothesize that leptin, resistin, and adiponectin might exhibit cytokine-like properties in human placenta and adipose tissue. Therefore, the main purpose of this study was to investigate the effect of leptin, resistin, and adiponectin on proinflammatory cytokine release and phospholipid metabolism from human placenta and sc adipose tissue obtained from normal pregnant women. In addition, the role of the signaling pathways involving NF-B, peroxisomal proliferator-activated receptor (PPAR)-, and MAPK involved in this regulation was also investigated.
Materials and Methods
Reagents
All chemicals were purchased from BDH Chemicals Australia (Melbourne, Victoria, Australia) unless otherwise stated. RPMI 1640 (glucose free) was obtained from Life Technologies, Inc. Laboratories (Grand Island, NY). BSA (RIA grade), ?-NADH (disodium salt), 3,3',5,5'-tetramethylbenzidine, and pyruvic acid (dimer free) were supplied by Sigma (St. Louis, MO). The leptin ELISA kit was supplied by Biosource International (Camarillo, CA). The IL-1?, IL-6, TNF, and adiponectin kits were purchased from R & D Systems (Minneapolis, MN). The PGE2 and PGF2 enzyme immunoassay kits were supplied from Cayman Chemical Co. (Ann Arbor, MI). The resistin ELISA kit, the active fragment of adiponectin gAcrp30, human recombinant leptin, and human recombinant resistin were purchased from CytoLab (Rehovot, Israel). (E)-3-[4-Methylphenylsulfonyl]-2-propenenitrile (BAY 11-7082) was purchased from Sapphire Bioscience (Crows Nest, New South Wales, Australia). Troglitazone was generously provided by Sankyo (Japan, Tokyo), and U0126 was purchased from Tocris (Ellisville, MO).
Patients and sample collection
Human placenta and sc adipose tissue (from the anterior abdominal wall) were obtained from pregnant women who delivered healthy, singleton infants at term (37 wk gestation) undergoing elective cesarean section (indications for cesarean section were breech presentation and/or previous cesarean section). Approval for this study was obtained from the Mercy Hospital for Women’s Research and Ethics Committee, and informed consent was obtained from all participating subjects.
Tissue explants
Tissues were obtained within 10 min of delivery and dissected fragments were placed in ice-cold PBS as previously described (5, 16, 17). Briefly, tissues were dissected to remove visible connective tissue, vessels and/or calcium deposits. Tissue fragments were placed in RPMI 1640 (placenta) or DMEM (adipose tissue) containing 5 mM glucose at 37 C in a humidified atmosphere of 8% O2-5% CO2 for 1 h. Explants were blotted dry on sterile filter paper and transferred to 24-well tissue culture plates (100–200 mg wet weight per well). Placental explants (n = 5) were incubated, in duplicate, in 2 ml medium containing penicillin G (100 U/ml) and streptomycin (100 μg/ml), in the absence (basal release) or presence of leptin (1, 10, and 100 ng/ml), resistin (1, 10, and 100 ng/ml), and adiponectin (0.1 and 0.5 μg/ml). Due to the limited amount of tissue sample, only maximum concentrations of leptin (100 ng/ml), resistin (100 ng/ml), and adiponectin (0.5 μg/ml) were used for adipose tissue explants (n = 5). Placental explants (n = 3) were also cotreated with 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126. After 6 h incubation, tissues were collected and assayed for total protein, whereas the incubation medium was collected and assayed for TNF, IL-1?, IL-6, PGF2, PGE2, leptin, resistin, and adiponectin release by ELISA.
Validation of explant cultures and viability
To determine the effect of experimental treatment on cell membrane integrity, the release of the intracellular enzyme lactate dehydrogenase (LDH) into incubation medium was determined as described previously (5, 16, 17). LDH release was investigated over the 6 h time course of tissue explants (n = 3). In vitro incubation did not significantly affect LDH activity in the incubation medium, with none of these measurements exceeding 7% of total activity present in the tissue (data not shown). These data indicate that the concentrations of leptin, resistin, and adiponectin used in this study did not affect cell viability.
Experimental assays
The release of TNF, IL-1?, IL-6, PGE2, PGF2, leptin, resistin, and adiponectin into the explant incubation medium was performed by sandwich ELISA according to the manufacturers’ instructions. The intraassay and interassay coefficients of variation for the leptin ELISA was 3.4 and 4.5%, respectively, and the minimum detectable limit of the assay was 7.2 pg/ml. The intraassay and interassay coefficients of variation for the resistin ELISA was 3.4 and 4.5%, respectively, and the minimum detectable limit of the assay was 16 pg/ml. The intraassay and interassay coefficients of variation for the adiponectin ELISA was 4.2 and 4.7%, respectively, and the minimum detectable limit of the assay was 32 pg/ml. All tissue release data were corrected for total protein. The protein content of tissue homogenates was determined using BCA protein assay (Pierce, Rockford, IL), using BSA as a reference standard, as previously described (5, 16, 17).
Statistical analysis
Statistical analyses were performed using a commercially available statistical software package (Statgraphics, StatPoint, Inc., Herndon, VA). Homogeneity of data was assessed by Bartlett’s test, and when significant, data were logarithmically transformed before further analysis. The effect of experimental treatment on cytokine and prostaglandin release was analyzed by multivariate ANOVA. The method used to discriminate among the means is Fisher’s least significant difference procedure. Statistical difference was indicated by P < 0.05. Data are expressed as mean ± SEM.
Results
Effect of leptin, resistin, and adiponectin on leptin, resistin, and adiponectin release
Placental and adipose tissues were incubated in the presence of 1, 10, and/or 100 ng/ml of leptin; 1, 10, and/or 100 ng/ml resistin; and 0.1 and/or 0.5 μg/ml adiponectin and the release of leptin, resistin, and adiponectin measured by ELISA. Leptin, at all concentrations tested in this study, had no effect on resistin or adiponectin release from either placenta or adipose tissue (data not shown). Likewise, placental and adipose tissue release of leptin and adiponectin were unaffected by treatment with resistin (data not shown). Furthermore, addition of adiponectin to placenta and adipose tissue explants did not affect the release of leptin or resistin (data not shown).
Effect of leptin, resistin, and adiponectin on proinflammatory cytokine release
Compared with control, tissues incubated in the presence of 100 ng/ml leptin caused a significant increase in the release of IL-1?, IL-6, and TNF from human placenta (Fig. 1, A–C, respectively). Resistin, at all concentrations tested in this study, had no effect on proinflammatory cytokine release from placenta (data not shown). Adiponectin, at both 0.1 and 0.5 μg/ml, increased IL-1?, IL-6, and TNF release from human placenta (Fig. 2, A–C, respectively). Similarly in adipose tissue, IL-1? (Fig. 3), IL-6 (Fig. 4), and TNF (Fig. 5) release was significantly increased by treatment with 100 ng/ml leptin and 0.5 μg/ml adiponectin but not 100 ng/ml resistin.
FIG. 1. Effect of 1, 10, and 100 ng/ml leptin on the release of IL-1? (A), IL-6 (B), TNF (C), PGE2 (D), and PGF2 (E) from human placenta (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal cytokine release from placenta.
FIG. 2. Effect of 0.1 and 0.5 μg/ml adiponectin on the release of IL-1? (A), IL-6 (B), TNF (C), PGE2 (D), and PGF2 (E) from human placenta (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal cytokine release from placenta.
FIG. 3. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of IL-1? from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal IL-1? release from adipose tissue.
FIG. 4. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of IL-6 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal IL-6 release from adipose tissue.
FIG. 5. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of TNF from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal TNF release from adipose tissue.
Effect of leptin, resistin, and adiponectin on PG release
Placental tissues incubated in the presence of 100 ng/ml leptin significantly increased the release of PGE2 (Fig. 1D). Compared with control, incubation of placenta with 10 and 100 ng/ml leptin caused a significant increase in the release of PGF2 (Fig. 1E). Resistin, at all concentrations tested in this study, had no effect on PG release from placenta (data not shown). Adiponectin, at 0.5 μg/ml, significantly increased PGE2 (Fig. 2D) and PGF2 (Fig. 2E) release from human placenta. Both leptin and adiponectin significantly increased PGE2 release from adipose tissue (Fig. 6). Although leptin and adiponectin increased PGF2 release from adipose tissue, this failed to reach significance (Figs. 7). There was no effect of resistin on PGE2 (Fig. 6) and PGF2 release (Fig. 7).
FIG. 6. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of PGE2 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal PGE2 release from adipose tissue.
FIG. 7. Effect of 100 ng/ml leptin, 100 ng/ml resistin, and 0.5 μg/ml adiponectin on the release of PGF2 from human adipose tissue (n = 5). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal PGF2 release from adipose tissue.
Effect of U0126, BAY 11-7082, and troglitazone on leptin- and adiponectin-induced proinflammatory cytokine release
To determine the intracellular pathways involved in leptin- and adiponectin-induced proinflammatory cytokine release, placental explants were cotreated with the NF-B inhibitor BAY 11-7082, the PPAR ligand troglitazone, and the ERK1/2 MAPK inhibitor U0126. Treatment of placental explants with 50 μM BAY 11-7082, 30 μM troglitazone, or 10 μM U0126 significantly inhibited leptin (100 ng/ml), and adiponectin (0.5 μg/ml) induced IL-1? (Figs. 8A and 9A), IL-6 (Figs. 8B and 9B), and TNF (Figs. 8C and 9C) release.
FIG. 8. Effect of 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126 on 100 ng/ml leptin-stimulated IL-1? (A), IL-6 (B), and TNF (C) release from human placenta (n = 3). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal release; , P < 0.05 vs. leptin- stimulated release.
FIG. 9. Effect of 50 μM BAY 11-7082, 30 μM troglitazone, and 10 μM U0126 on 0.5 μg/ml adiponectin-stimulated IL-1? (A), IL-6 (B), and TNF (C) release from human placenta (n = 3). Each bar represents the mean ± SEM. *, P < 0.05 vs. basal release; , P < 0.05 vs. adiponectin-stimulated release.
Discussion
It has become increasingly apparent that leptin, resistin, and adiponectin have pleiotropic actions on various cell types beyond their effects on regulating energy homeostasis. In light of the involvement of leptin, resistin, and adiponectin in immune and inflammatory response, we employed a well-characterized in vitro human tissue explant system to examine their potential inflammatory effects in placenta and adipose tissue. Our data show that leptin and adiponectin, but not resistin, have proinflammatory actions, involving the stimulation of proinflammatory cytokines TNF, IL-1?, IL-6, and prostaglandins PGE2 and PGF2 from human placenta and adipose tissue. Our studies further aimed to investigate which intracellular signaling pathways are activated by leptin and adiponectin. We found that BAY 11-7082, troglitazone, and U0126 treatment suppressed leptin- and adiponectin-stimulated IL-1?, IL-6, and TNF release from human placenta, suggesting that NF-B, PPAR, and ERK 1/2 MAPK all appeared to be key transcription factors involved in leptin- and adiponectin-induced proinflammatory response.
Leptin directly regulates the production of several cytokines both in vivo and in vitro (reviewed in Refs. 18, 19, 20, 21). Leptin has been implicated in stimulating Th1 cytokines in rodents (8); increasing LPS-stimulated production of TNF, IL-6, and IL-12 in murine peritoneal macrophages and human monocytes (22, 23); and stimulating IL-6 secretion by human term trophoblast cells (24) and human placenta (25). Likewise, in this study, leptin increased the secretion of TNF, IL-1?, and IL-6 from human placenta and adipose tissue.
Leptin has also been shown to exert proinflammatory effects by stimulating the phospholipid metabolizing pathway. Leptin stimulates PGE2 and PGF2 production in neonatal rat hypothalamus (26); augments alveolar macrophage leukotriene synthesis by increasing phospholipase A2 (PLA2) activity and enhancing cytosolic PLA2 protein expression (27); up-regulates cytosolic PLA2 activity in human bone marrow stromal cells (28); and potentiates IFN-induced expression of cyclooxygenase-2 and PGE2 production from murine macrophages J774A.1 (29). In this study, we also demonstrated that treatment of placenta and adipose tissue with leptin resulted in an increased release of the proinflammatory prostaglandins PGE2 and PGF2.
Placenta and adipose tissue have both the long and the short isoforms of the leptin receptor. The Ob-Rb receptor, which is widely expressed in human placenta and adipose tissue, mediates leptin action by signaling via the Janus kinase/STAT pathway (30) including the downstream suppressor of cytokine signaling-3 feedback inhibitor and/or other common pathways such as the p38 MAPK, p42/p44 MAPK (28, 31, 32, 33, 34), and NF-B pathway (9, 31, 35). Cauzac et al. (36) demonstrated that transduction of leptin growth signals in placental cells is independent of Janus kinase/STAT activation; however, leptin activated the ERK 1/2 MAPK cascade. Likewise, in this study, we demonstrated that leptin-stimulated proinflammatory cytokine release from human placenta could be abrogated by treatment with the antiinflammatory ERK 1/2 MAPK inhibitor U0126, the PPAR ligand troglitazone, and the NF-B inhibitor BAY 11-7082. This is also in agreement with our previous studies, demonstrating the importance of NF-B and PPAR in the regulation of TNF and IL-6 release from human placenta (16, 17) and adipose tissue (37).
Although there are ample data available on the effect of proinflammatory mediators on the expression and release of resistin, to date, few data are available on the effect of resistin on proinflammatory cytokines and PGs. The available data do, however, suggest a proinflammatory role for resistin. In particular, treatment with 30 ng/ml resistin results in elevated secretion of IL-6 and TNF from human sc adipocytes (38). Furthermore, resistin promotes smooth muscle cell proliferation through activation of ERK 1/2 but not p38 MAPK (39). This is in contrast to the findings of this study, in which there was no effect of resistin (1–100 ng/ml) on the release of proinflammatory cytokines and PGs from human placenta and adipose tissue.
Adiponectin is postulated to be an antiinflammatory cytokine. It induces the antiinflammatory cytokines IL-10 in human leukocytes (10) and porcine macrophages (11), whereas adiponectin suppresses the production of the proinflammatory cytokine IFN in human macrophages (10); attenuates LPS-stimulated TNF and IL-6 release from porcine macrophages (11); and decreases LPS-induced hepatic TNF mRNA expression and release in KK-Ay obese mice (40). Furthermore, elevated TNF levels have been reported in the adiponectin knockout mouse, and the introduction of adiponectin by adenoviral infection normalized serum TNF in these mice (41). These antiinflammatory actions of adiponectin are mediated in part by suppression of NF-B signaling (11, 42) and ERK 1/2 MAPK activity (11).
In this study, adiponectin exerted proinflammatory effects by increasing the release of proinflammatory cytokines and PGs from human placenta and adipose tissue. This is in agreement with previous studies (43, 44) that demonstrated that adiponectin caused elevated expression of cyclooxygenase-2 and induced PGE2 release from stromal preadipocytes, and results of Schaffler et al. (45) demonstrated that adiponectin induced the release of IL-6 and matrix metalloproteinase-1 in synovial fibroblasts. Like leptin, the proinflammatory actions of adiponectin are mediated through a number of signaling pathways, including p38 MAPK (45) and NF-B (12). Similarly, in this study, adiponectin-induced proinflammatory response could be abrogated by treatment with the antiinflammatory ERK1/2 MAPK inhibitor U0126, the PPAR ligand troglitazone, and the NF-B inhibitor BAY 11-7082. Although the transcriptional regulation of proinflammatory cytokine gene expression in response to leptin and adiponectin treatment is not known, these findings are consistent with human IL-1?, IL-6, and TNF promoters containing response elements for a number of transcription factors, including NF-B, activator protein-1, and specificity protein-1 (reviewed in Ref. 46).
There are many possible reasons between the conflicting results between our studies and previous studies with respect to the response elicited upon resistin or adiponectin treatment. First, this may represent differences in tissue or cell-specific functions: (1) tissue explants are not comprised of just one cell type and paracrine mediators produced by, for example, nonadipocyte cells present in adipose tissue (such as immunocytes and vascular cells) may be necessary for adiponectin to stimulate proinflammatory cytokine release; (2) the explant system and the preservation of the extracellular matrix may provide a more natural environment that allows placental cells and adipocytes to respond to stimuli as they would in vivo; and (3) the conditions that are used to prepare and culture cells from may select for a subset of cells that respond to resistin and adiponectin in a different fashion than do mature fat cells or trophoblasts. Nevertheless, our studies are the first demonstration of such activities in placenta and adipose tissue. Second, the differences observed may also be due to different physiological functions of bacterially produced full-length adiponectin, the globular domain of adiponectin (gAcrp30), or the full-length adiponectin produced by mammalian cells. And third, it may also represent differences in the concentrations used. For example, in this study, adiponectin was used at 0.1 and 0.5 μg/ml, whereas others have used concentrations of adiponectin as high as 50 μg/ml (42).
The form of adiponectin used in this study was gAcrp30, which corresponds to the entire C-terminal globular domain of adiponectin; however, circulating adiponectin exists in several isoforms, including trimers, hexamers, and high-molecular-weight multimers (12). Therefore, it is uncertain whether the effects observed in human placenta and adipose tissue are purely pharmacological or whether they are reflective of a physiological aspect of adiponectin function. Future studies examining the effects of all forms of adiponectin will further contribute to our understanding of the role of this protein in regulating the inflammatory response.
In this study, leptin and adiponectin exerted proinflammatory actions in human placenta and adipose tissue, and this may therefore have important implications in a number of normal and abnormal physiological processes. Prostaglandins and proinflammatory cytokines that are produced within the intrauterine environment participate in the regulation of myometrial contractility, cervical ripening and rupture of membranes (reviewed in Refs. 1 and 2). Therefore, leptin and adiponectin, both of which are produced within the intrauterine environment, may play a role in the processes of human labor and delivery. Furthermore, these proinflammatory actions of leptin and adiponectin may also have significant implications in obesity and diabetes. The production of leptin and/or adiponectin within adipose tissue may contribute to the raised circulating levels of proinflammatory mediators that are evident in these disease states.
In summary, an in vitro tissue explant model system was used to investigate the inflammatory actions of leptin, resistin, and adiponectin on the release of proinflammatory mediators from placenta and adipose tissue. These data demonstrate that proinflammatory mediators in placenta and adipose tissue represent targets for up-regulation by leptin and adiponectin and provide mechanisms by which these hormones can promote inflammatory responses. These data have extended our knowledge of the factors involved in the regulation of proinflammatory cytokines and PGs.
Acknowledgments
The authors gratefully acknowledge the assistance of the clinical research midwives Val Bryant and Sarah Mitchell and the obstetrics and midwifery staff of the Mercy Hospital for Women for their cooperation.
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