Omental Macrophages Research Articles

Abstract 6810: Macrophage secreted CCL9/CCL5 induces CCR1-mediated ovarian cancer metastasis to the omentum in the absence of CCL6

Abstract The tumor-microenvironment (TME) of the omentum plays an important role in providing a pre-metastatic niche for high-grade serous ovarian cancer (HGSC-OvCa) progression. Omentum, a layer of fatty adipose tissue within the peritoneal cavity, is the preferred primary metastatic site for OvCa. Omentum is a well-vascularized fatty tissue containing wide-spread immune clusters (milky spots). Milky spots in the omentum are primarily composed of B cells, T cells, macrophages, NK cells, and dendritic cells. Previous studies from our laboratory have highlighted the importance of CCL6 secreted by omental macrophages as a key chemokine in providing a pre-metastatic niche by activating the CCR1 axis on the cancer cells required for OvCa invasion. We engineered and characterized a CCL6-depleted in vivo mouse model using CRISPR/Cas9 to study the effect on OvCa progression and macrophage biology. We demonstrate that deletion of CCL6 in mice alters characteristics of omental tissue-resident (OM-Macφ) and bone marrow-derived macrophages (BMDM) derived from circulating monocytes. RNA sequencing of OM-Macφ and microarray study of BMDM revealed that CCL6 deletion produces enhanced TGFβ1+CSF-1+CCL5+ expression in OM-Macφ and an increase in TGFβ1+CSF-1+CCL9+ in BMDM. This acts as a rescue mechanism to maintain activation of their shared CCR1 receptor with a pro-M2 stimulation profile. Deep multiplexed imaging of the omentum tissue by CODEX showed that CCL6 deletion alters the architecture of milky spots, with enriched macrophage content and reduced B-cell and T-cell-rich neighborhoods. While our previous studies indicated that CCL6 deletion could potentially have an anti-tumoral effect, we found that it does not significantly affect murine ID8 ovarian cancer tumor burden in both short-term (7 days) and long-term in vivo studies and provides no added survival advantage. We report that CCL9+F4/80+ macrophages accumulate in the omentum post-tumor infiltration in the CCL6-KO model, and neutralizing CCL9 in the CCL6-KO BMDM macrophage-derived conditioned media significantly reduces migration of ID8 cancer cells in vitro. Furthermore, we demonstrate that murine CCL9 and its analog, human CCL15, promote the epithelial-mesenchymal transition (EMT) of mouse ID8 and human A2780 OvCa cells. Moreover, our findings indicate that human CCL15 is a prognostic biomarker for reduced survival in OvCa patient cohorts (Stages I&II and Stage III&IV). Further classification of HGSC-OV patients (n=375) from TCGA-OV based on CCL15 and CCR1-high expression correlated with a higher expression of TGFβ1, CCL5, and a T-cell exhaustion signature in the TME. Taken together, given the apparent redundancy of the chemokine network controlling CCR1 signaling, our findings support the strategy of therapeutically blocking the CCR1 receptor to potentially produce a favorable anti-tumor response in HGSC-OvCa patients. Citation Format: Supreeti Tallapragada, Justine Chan, Venkatesh Krishnan, Oliver Dorigo. Macrophage secreted CCL9/CCL5 induces CCR1-mediated ovarian cancer metastasis to the omentum in the absence of CCL6 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6810.

Laminins in tumor-derived exosomes upregulated by ETS1 reprogram omental macrophages to promote omental metastasis of ovarian cancer

Tumor-derived exosomes participate in omental metastatic colonization of ovarian cancer by inducing an adaptive response in the tumor microenvironment. However, cell–cell communication via exosomes between primary tumor cells and the microenvironment of distant omentum and the mechanism of pre-metastatic niche formation are poorly understood. Here, we demonstrated that ETS1-overexpressing ovarian cancer cells secreted larger exosomes with higher laminin levels. In addition, ovarian cancer exosomes could be taken up by omental macrophages through integrin and laminin interaction. Compared with control exosomes, exosomes derived from ETS1-overexpressing ovarian cancer cells (LV-ETS1 Exos) stimulated the polarization of more macrophages toward the M2 phenotype (CD163 marker), as well as the production of more CXCL5 and CCL2 in macrophages, via integrin αvβ5/AKT/Sp1 signaling. In vivo experiments showed that LV-ETS1 Exos promoted omental metastasis of ovarian cancer by mediating the tumor-promoting effect of macrophages, which could be neutralized by integrin ανβ5 inhibitor cilengitide. These results indicated that ETS1 could drive ovarian cancer cells to release exosomes with higher laminin levels, thereby accelerating the exosome-mediated pro-metastatic effects of omental macrophages via the integrin αvβ5/AKT/Sp1 signaling pathway, and the integrin ανβ5 inhibitor cilengitide could inhibit omental metastasis of ovarian cancer driven by tumor-derived exosomes.

Cell origin and niche availability dictate the capacity of peritoneal macrophages to colonize the cavity and omentum.

The relationship between macrophages of the peritoneal cavity and the adjacent omentum remains poorly understood. Here, we describe two populations of omental macrophages distinguished by CD102 expression and use an adoptive cell transfer approach to investigate whether these arise from peritoneal macrophages, and whether this depends upon inflammatory status, the origin of peritoneal macrophages and availability of the omental niches. We show that whereas established resident peritoneal macrophages largely fail to migrate to the omentum, monocyte-derived resident cells readily migrate and form a substantial component of omental CD102+ macrophages in the months following resolution of peritoneal inflammation. In contrast, both populations had the capacity to migrate to the omentum in the absence of endogenous peritoneal and omental macrophages. However, inflammatory macrophages expanded more effectively and more efficiently repopulated both CD102+ and CD102− omental populations, whereas established resident macrophages partially reconstituted the omental niche via recruitment of monocytes. Hence, cell origin determines the migration of peritoneal macrophages to the omentum and predisposes established resident macrophages to drive infiltration of monocyte-derived cells.

IL-1β Impacts Vascular Integrity and Lymphatic Function in the Embryonic Omentum.

The chromatin-remodeling enzyme BRG1 (brahma-related gene 1) regulates gene expression in a variety of rapidly differentiating cells during embryonic development. However, the critical genes that BRG1 regulates during lymphatic vascular development are unknown. We used genetic and imaging techniques to define the role of BRG1 in murine embryonic lymphatic development, although this approach inadvertently expanded our study to multiple interacting cell types. We found that omental macrophages fine-tune an unexpected developmental process by which erythrocytes escaping from naturally discontinuous omental blood vessels are collected by nearby lymphatic vessels. Our data indicate that circulating fibrin(ogen) leaking from gaps in omental blood vessels can trigger inflammasome-mediated IL-1β (interleukin-1β) production and secretion from nearby macrophages. IL-1β destabilizes adherens junctions in omental blood and lymphatic vessels, contributing to both extravasation of erythrocytes and their uptake by lymphatics. BRG1 regulates IL-1β production in omental macrophages by transcriptionally suppressing the inflammasome trigger RIPK3 (receptor interacting protein kinase 3). Genetic deletion of Brg1 in embryonic macrophages leads to excessive IL-1β production, erythrocyte leakage from blood vessels, and blood-filled lymphatics in the developing omentum. Altogether, these results highlight a novel context for epigenetically regulated crosstalk between macrophages, blood vessels, and lymphatics.

LYVE1+ macrophages of murine peritoneal mesothelium promote omentum-independent ovarian tumor growth.

Two resident macrophage subsets reside in peritoneal fluid. Macrophages also reside within mesothelial membranes lining the peritoneal cavity, but they remain poorly characterized. Here, we identified two macrophage populations (LYVE1hi MHC IIlo-hi CX3CR1gfplo/- and LYVE1lo/- MHC IIhi CX3CR1gfphi subsets) in the mesenteric and parietal mesothelial linings of the peritoneum. These macrophages resembled LYVE1+ macrophages within surface membranes of numerous organs. Fate-mapping approaches and analysis of newborn mice showed that LYVE1hi macrophages predominantly originated from embryonic-derived progenitors and were controlled by CSF1 made by Wt1+ stromal cells. Their gene expression profile closely overlapped with ovarian tumor-associated macrophages previously described in the omentum. Indeed, syngeneic epithelial ovarian tumor growth was strongly reduced following in vivo ablation of LYVE1hi macrophages, including in mice that received omentectomy to dissociate the role from omental macrophages. These data reveal that the peritoneal compartment contains at least four resident macrophage populations and that LYVE1hi mesothelial macrophages drive tumor growth independently of the omentum.

Omental macrophages secrete chemokine ligands that promote ovarian cancer colonization of the omentum via CCR1

The omentum is the most common site of ovarian cancer metastasis. Immune cell clusters called milky spots are found throughout the omentum. It is however unknown if these immune cells contribute to ovarian cancer metastasis. Here we report that omental macrophages promote the migration and colonization of ovarian cancer cells to the omentum through the secretion of chemokine ligands that interact with chemokine receptor 1 (CCR1). We found that depletion of macrophages reduces ovarian cancer colonization of the omentum. RNA-sequencing of macrophages isolated from mouse omentum and mesenteric adipose tissue revealed a specific enrichment of chemokine ligand CCL6 in omental macrophages. CCL6 and the human homolog CCL23 were both necessary and sufficient to promote ovarian cancer migration by activating ERK1/2 and PI3K pathways. Importantly, inhibition of CCR1 reduced ovarian cancer colonization. These findings demonstrate a critical mechanism of omental macrophage induced colonization by ovarian cancer cells via CCR1 signaling.

Tissue-resident macrophages in omentum promote metastatic spread of ovarian cancer.

Experimental and clinical evidence suggests that tumor-associated macrophages (TAMs) play important roles in cancer progression. Here, we have characterized the ontogeny and function of TAM subsets in a mouse model of metastatic ovarian cancer that is representative for visceral peritoneal metastasis. We show that the omentum is a critical premetastatic niche for development of invasive disease in this model and define a unique subset of CD163+ Tim4+ resident omental macrophages responsible for metastatic spread of ovarian cancer cells. Transcriptomic analysis showed that resident CD163+ Tim4+ omental macrophages were phenotypically distinct and maintained their resident identity during tumor growth. Selective depletion of CD163+ Tim4+ macrophages in omentum using genetic and pharmacological tools prevented tumor progression and metastatic spread of disease. These studies describe a specific role for tissue-resident macrophages in the invasive progression of metastatic ovarian cancer. The molecular pathways of cross-talk between tissue-resident macrophages and disseminated cancer cells may represent new targets to prevent metastasis and disease recurrence.

Abstract TMIM-076: OMENTAL MACROPHAGES REGULATE OVARIAN CANCER METASTATIC COLONIZATION THROUGH THE CCL6-CCR1 SIGNALING AXIS

Abstract BACKGROUND: Metastatic ovarian cancer remains an urgent clinical problem. The homing and invasion of cancer cells into the omental adipose tissue, which is the preferred site of ovarian cancer metastasis, is a critical step in disease progression. Omental adipose tissue contains adipocytes, blood vessels, and clusters of leukocytes known as milky spots. We have previously demonstrated in several mouse models that intraperitoneally injected ovarian cancer cells localize to the omentum and specifically to the milky spots. However, we found that B, T, or NK cells were not required for omental colonization. We hypothesize that macrophages within the milky spots play a crucial role in promoting ovarian cancer progression. METHODS AND RESULTS: Depleting macrophages disrupts cancer cells' localization to the omentum. This result supports our hypothesis that macrophages play an essential role in attracting ovarian cancer cells to the omental milky spots. Correspondingly, factors secreted by omental macrophages promote ovarian cancer migration in vitro, in contrast to factors secreted by macrophages isolated from other peritoneal adipose tissues. In order to understand the specific mechanism(s) that regulate metastatic colonization of the omentum, we performed RNA-Seq of CD45+CD11b+F4/80+ flow-sorted macrophages from the omentum and the mesenteric fat of naïve and cancer-bearing mice. We found that omental macrophages have a distinct and dynamic gene expression pattern even when compared to a very similar cell type. The chemokine CCL6 is one of the most highly upregulated genes during metastatic colonization. The human homolog of CCL6 is CCL23 and is also expressed by human omental macrophages. Genomic inactivation of CCR1, the gene encoding receptor for CCL6 in mice and CCL23 in humans, in mouse ovarian cancer cells abolishes their ability to home to omental milky spots. Our findings establish a role for chemokine signaling in the early colonization of the omentum by ovarian cancer. CONCLUSION: We demonstrate that omental milky spots are the preferential sites for cancer colonization of peritoneal adipose. Further, we have identified omental macrophages as the critical mediators of colonization via the CCL6/CCR1 chemokine axis. Future studies are focused on understanding the cancer cell-macrophage interactions that can be targeted therapeutically to disrupt metastatic growth and extend disease-free survival. Citation Format: Venkatesh Krishnan, Supreeti Tallapragada, Bruce Schaar, Anita Chanana, Carrie Rinker-Schaeffer, Oliver Dorigo. OMENTAL MACROPHAGES REGULATE OVARIAN CANCER METASTATIC COLONIZATION THROUGH THE CCL6-CCR1 SIGNALING AXIS [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr TMIM-076.

Whole Body Vibration-Induced Omental Macrophage Polarization and Fecal Microbiome Modification in a Murine Model.

Human nutrient metabolism, developed millions of years ago, is anachronistic. Adaptive features that offered survival advantages are now great liabilities. The current dietary pattern, coupled with massively reduced physical activities, causes an epidemic of obesity and chronic metabolic diseases, such as type 2 diabetes mellitus. Chronic inflammation is a major contributing factor to the initiation and progression of most metabolic and cardiovascular diseases. Among all components of an innate immune system, due to their dual roles as phagocytic as well as antigen-presenting cells, macrophages play an important role in the regulation of inflammatory responses, affecting the body’s microenvironment and homeostasis. Earlier studies have established the beneficial, anti-inflammatory effects of whole body vibration (WBV) as a partial exercise mimetic, including reversing the effects of glucose intolerance and hepatic steatosis. Here for the first time, we describe potential mechanisms by which WBV may improve metabolic status and ameliorate the adverse consequences through macrophage polarization and altering the fecal microbiome.

Abstract TMEM-028: OMENTAL MACROPHAGES REGULATE OVARIAN CANCER METASTATIC COLONIZATION

Abstract BACKGROUND: Metastatic ovarian cancer remains an urgent clinical problem. The homing and invasion of cancer cells into the omental adipose tissue, which is the preferred site of ovarian cancer metastasis, is a critical step in disease progression. Improving patient outcomes requires a mechanistic understanding of how cancer cells colonize the omentum and ultimately give rise to widespread peritoneal metastases. Unlike other peritoneal adipose tissues, omental adipose contains immune aggregates known as milky spots, which contain macrophages, B, T, natural killer (NK), and stromal cells within capillary nests. Our data shows that milky spots play an active role in ovarian cancer metastatic colonization to the omentum. We hypothesize that macrophages within the milky spots play a crucial role in promoting ovarian cancer progression. METHODS AND RESULTS: Immunocompetent mice were injected with syngeneic murine ID8 and immunocompromised mice were injected with human (SKOV3ip.1, CaOV3, HeyA8), ovarian cancer cells. After seven days post-injection, mice were sacrificed and analyzed for the distribution of cancer cells. We found metastases in omental milky spots and not in adipose lacking milky spots. Use of genetic models (C57BL/6, Athymic Nude, Beige Nude, Rag1-/-, Igh6-/- mice) ruled out a requirement for B, T, and NK cells in ovarian cancer cell homing to milky spots, suggesting a critical role for macrophages in this process. Our recent data shows that depletion of omental adipose tissue macrophages abrogates cancer cell colonization of milky spots. Furthermore, factors secreted by omental macrophages have an enhanced ability to promote ovarian cancer migration in vitro, compared to factors secreted by other peritoneal adipose tissue macrophages. Screening omental adipose conditioned medium with a cytokine array revealed higher concentration of macrophage-secreted cytokines (e.g., MCSF-1, IL10) than control medium conditioned by adipose lacking milky spots. Examination of naïve human omentum confirmed the presence of macrophages in immune clusters. Clinical samples from patients with high-grade serous cancer showed the incidence of microscopic metastases within omental immune clusters. Importantly, RNA-Seq of CD45+CD11b+F4/80+ flow-sorted macrophages from omentum and mesentery of naïve and cancer-bearing mice revealed that omental macrophages have a very distinct gene expression signature. CONCLUSION: We demonstrate that omental milky spots are the preferential sites for cancer colonization of peritoneal adipose. Further, we have identified omental macrophages as the key mediators of colonization. Future studies are focused on understanding the cancer cell-macrophage interactions that can be targeted therapeutically to disrupt metastatic growth and extend disease-free survival. Citation Format: Venkatesh Krishnan, Bruce Schaar, Carrie Rinker-Schaeffer, Oliver Dorigo. OMENTAL MACROPHAGES REGULATE OVARIAN CANCER METASTATIC COLONIZATION [abstract]. In: Proceedings of the 11th Biennial Ovarian Cancer Research Symposium; Sep 12-13, 2016; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(11 Suppl):Abstract nr TMEM-028.

Abstract 4853: Omental milky-spots serve as a niche for ovarian cancer metastatic colonization

Abstract Metastatic ovarian cancer remains an urgent clinical problem. The homing and invasion of cancer cells into the omental adipose tissue, the preferred site of ovarian cancer metastasis, is a rate-limiting step in disease progression. Improving patient outcomes requires a mechanistic understanding of how cancer cells colonize the omentum and ultimately give rise to widespread peritoneal metastases. Unlike other peritoneal adipose, omental adipose contains functional immune aggregates known as milky spots, which contain macrophages, B, T, natural killer (NK), and stromal cells within capillary nests. In addition to their role in peritoneal homeostasis and immune defense, our data shows that milky spots play an active role in ovarian cancer metastatic colonization. We hypothesize that omental microenvironment plays a crucial role in instructing the ovarian cancer cells to achieve an aggressive phenotype that enables wide spread metastases. Well-established in vitro migration assays and in vivo experimental metastasis assays were used to answer the following questions: Does cancer cell localization depend upon the immune composition of the milky spots? Do cancer cells utilize the native milky spot microenvironment, or does it undergo remodeling during colonization? Answers to these questions will be the foundation for mechanism-based studies aimed at identifying ovarian cancer-omental interactions that can be targeted therapeutically. Human (SKOV3ip.1, CaOV3, HeyA8), and murine (ID8) ovarian cancer cells rapidly localized to omental milky spots and not adipose lacking milky spots. Use of genetic models (C57BL/6, Athymic Nude, Beige Nude, Rag1-/-, Igh6-/- mice) ruled out a requirement of B, T, and NK cells in ovarian cancer cell homing to milky spots suggesting a critical role for macrophages in this process. Quantitative data found no difference in the incidence of cancer cell foci in the omentum across these strains. Our recent data shows that depletion of omental adipose tissue macrophages (ATM) abrogates cancer cell colonization of milky spots. In vitro assays have found that omental adipose-conditioned media causes 75% more cell migration and contains a higher concentration of macrophage-secreted cytokines than media conditioned by adipose lacking milky spots. Importantly, our data shows that factors secreted by omental ATMs have an enhanced ability to promote ovarian cancer migration compared to factors secreted by other peritoneal ATMs. We have shown that omental milky spots are required for cancer colonization of peritoneal adipose. Further, we have identified omental macrophages as the key mediators of colonization. Future studies are focused on understanding the cancer cell-macrophage interactions that can be targeted therapeutically to disrupt metastatic growth and extend disease-free survival. Acknowledgement: The DOD (W81XWH-09-1-0127), NCI/NIH (2RO1CA089569) and Marsha Rivkin Center Pilot Study Award. Note: This abstract was not presented at the meeting. Citation Format: Venkatesh Krishnan, Robert Clark, Carrie Rinker-Schaeffer. Omental milky-spots serve as a niche for ovarian cancer metastatic colonization. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4853. doi:10.1158/1538-7445.AM2014-4853

Macrophage infiltration and stress-signaling in omental and subcutaneous adipose tissue in diabetic pregnancies

Objective: To examine if, as in obesity, pregnancies complicated by gestational diabetes mellitus (GDM) exhibit increased macrophage infiltration and activated MAP-kinases in omental adipose tissue.Methods: Paired omental (OM) and abdominal subcutaneous (SC) fat samples were collected from 11 GDM and 20 normal pregnancies during cesarean delivery. Tissues were stained to detect macrophages, and analyzed to assess MAP-kinases.Results: OM had higher macrophage counts than SC in GDM (6.10 ± 2.20 versus 2.53 ± 1.45, p = 0.04), but not in normal pregnancies (p = 0.346). GDM pregnancies had more macrophages than normal pregnancies in OM (6.10 ± 2.20 versus 1.29 ± 0.55, p = 0.01), while only a trend was observed in SC fat (p = 0.08). Significant correlation (R = 0.619, p = 0.005) was observed between OM-macrophage infiltration and insulin resistance. Using multivariate analysis, only obesity independently associated with GDM. Expression of total p38MAP-kinase was higher in OM versus SC in both normal and GDM pregnancies, without significant differences between these groups. However, expression of activated p-p38MAP-kinase, and its upstream kinase MKK4, was comparable between fat depots.Conclusion: GDM pregnancies demonstrate increased macrophage infiltration to OM fat, correlating with higher insulin resistance. As in non-pregnant-patients obesity and OM macrophage infiltration may be on the same causal pathway, leading to GDM. Yet, this occurs without activation of p38MAP-kinase signaling.

FTO mRNA Expression in Extremely Obese and Type 2 Diabetic Human Omental and Subcutaneous Adipose Tissues

Fat mass and obesity-associated protein (FTO) gene expression is known to correlate with obesity. Our aim was to investigate the FTO gene expression in paired omental and subcutaneous human adipose tissues from morbid and obese patients. To understand the role of CD68-positive macrophages in adipose tissues, the correlation with adiposity parameters such as adipocyte diameter and adipocyte radius was also measured. Drug and adiposity correlations were also analyzed. Paired omental and subcutaneous adipose tissue were excised during elective surgery from morbidly obese (n = 9) and obese (n = 5) patients. FTO expressions were determined by quantitative PCR. Tissue sections were analyzed for their CD68 protein expressions by immunuhistochemistry. Omental and subcutaneous adipose tissue FTO gene expression levels were not found to differ significantly among morbidly obese and obese study groups. Serum aspartate aminotransferase e and alanine transaminase levels were found to be in negative correlation with subcutaneous fat tissue FTO expression rate. Antidiabetic drug use was found to be in correlation with adiposity. Both subcutaneous and omental fat cell diameters were found to have correlation with antidiabetic drug use. Omental fat cell diameter was found to enlarge together with omental CD68 protein expression. Subcutaneous macrophage number decreased while omental fat cell radius increased. Omental macrophage number was found in correlation with subcutaneous macrophage number. Antidiabetic therapy was found to increase adiposity in omental and subcutaneous fat. Further research is needed with larger samples to explore the exact role of FTO in obesity.

240: A significant higher rate of omental macrophages infiltration in gestational diabetes mellitus pregnancies

240: A significant higher rate of omental macrophages infiltration in gestational diabetes mellitus pregnancies

Chemerin Correlates with Markers for Fatty Liver in Morbidly Obese Patients and Strongly Decreases after Weight Loss Induced by Bariatric Surgery

Chemerin is a new adipokine involved in in vitro adipogenesis and insulin resistance and associates with body mass index (BMI) in vivo. We investigated the role of chemerin in morbid obesity, associated metabolic diseases (insulin resistance, hepatic diseases), and postsurgery-induced weight loss. This was a prospective study performed at a university hospital. Subjects included 60 obese female patients (BMI 50.0 +/- 1.0 kg/m(-2)) being candidates for gastric bypass. Patients were examined before and 3, 6, and 12 months after surgery. In 27 patients, chemerin was measured after 2 yr. Outcomes included chemerin, anthropometric parameters, homeostasis model assessment for insulin resistance index (HOMA-IR), cholesterol, high-density lipoprotein, triglycerides, C-reactive protein, adipokines at all time points; and liver histology and macrophage content in fat at baseline. Chemerin was substantially elevated in obese patients compared with nonobese persons (353.8 +/- 18.0 vs. 191 +/- 14 ng/ml, P < 0.001). Preoperatively, chemerin concentrations correlated positively with BMI, C-reactive protein, IL-6, HOMA-IR, and the amount of omental macrophages and negatively with high-density lipoprotein levels. Baseline chemerin was elevated in patients with a significant activity score for nonalcoholic fatty liver disease, portal inflammation, fibrosis, and fibroinflammation. After surgery, chemerin decreased significantly to 253.0 +/- 14.9 ng/ml after 1 yr and pursued its decrease in patients studied for 2 yr. After surgery, chemerin concentrations positively correlated with triglycerides. The strong decrease of chemerin in the 3 months after surgery was associated with the decrease in HOMA-IR and blood glucose. Chemerin concentrations are elevated in morbidly obese patients and correlated with insulin resistance and markers of liver pathology. Chemerin plasma concentrations decreased after bariatric surgery. This study suggests that chemerin might mediate metabolic alterations in obesity, drastically improving after gastric bypass.

Association between omental adipose tissue macrophages and liver histopathology in morbid obesity: Influence of glycemic status

Recently we showed that macrophage accumulation in omental adipose tissue is associated with liver fibro-inflammation in morbidly obese subjects. Here, we evaluated the influence of glycemic status and extended the analysis to the spectrum of obesity-linked liver damage. Liver biopsies, subcutaneous and omental adipose tissue were collected in 132 obese subjects during gastric bypass surgery. HAM56+ adipose tissue macrophages were counted in subjects classified by liver histopathology and by their degree of insulin resistance. In the whole population, the number of omental macrophages increased with the score of steatosis, the non-alcoholic fatty liver disease activity score, the stage of fibrosis and with fibro-inflammation index. None of these relationships were significant with subcutaneous macrophage count. In insulin-sensitive participants, omental macrophages accumulation was higher in subjects with high indexes of fibro-inflammation (p=0.012 vs. low indexes). In insulin-resistant including type 2 diabetic participants, omental macrophage count was higher both in subjects with high scores of steatosis and in subjects with high indexes of fibro-inflammation (p<0.05 vs. low scores). Macrophage accumulation in omental adipose tissue is associated with aggravated steatosis and fibro-inflammation in insulin-resistant obese subjects independently of altered glycemic status.

Insufficient ability of omental milky spots to prevent peritoneal tumor outgrowth supports omentectomy in minimal residual disease.

The greater omentum is frequently involved in the course of gastrointestinal and ovarian tumors. Therefore, common practice in surgical treatment for especially gastric and ovarian cancer includes removal of the greater omentum. Paradoxically, many immune cells, such as macrophages that accumulate in so-called milky spots, reside within the omentum and are cytotoxic against tumor cells ex vivo. Consequently, omental macrophages might play an important role in killing tumor cells, and may hereby prevent development into local peritoneal recurrences. In the present study, we therefore evaluated the role of the omentum and the clinical relevance of omentectomy in minimal residual disease (MRD). Tumor cell dissemination patterns on the omentum in a rat model were examined using DiI-labelled CC531s tumor cells. Additionally, intra peritoneal (i.p.) tumor load was investigated in rats that underwent omentectomy or sham laparotomy followed by i.p. injection of CC531s cells on day 21, which represented MRD. At 4 h post injection, tumor cells predominantly adhered on milky spots. Number of cells thereafter declined rapidly suggesting initial tumor killing functions in these specific immune aggregates. Despite initial reduction observed in milky spots, numbers of tumor cells however increased at fatty tissue stripes that border the omentum. This indicated proliferation at these locations, which corresponded to macroscopic observations of the omenta on day 21 after tumor cell injection. Omentectomy resulted in reduced intra-abdominal tumor load, which was completely attributable to the absence of the omentum, as tumor development did not differ on other sites. Even in the MRD group microscopic clusters of tumor cells located in the omentum eventually developed into macroscopic nodules. Since the ability of omental milky spots is, even in MRD, insufficient to prevent intra abdominal tumor outgrowth, omentectomy, which reduces tumor load, is recommended in surgical treatment of intra abdominal tumors that are prone to disseminate intraperitoneally.

Elimination of various subpopulations of macrophages and the development of multiple-organ dysfunction syndrome in mice.

To evaluate the role of specific macrophage subpopulations in the development of zymosan-induced multiple-organ dysfunction syndrome by selective elimination of liver, splenic, alveolar, and peritoneal macrophages. Randomized animal trial. Central animal laboratory at the University Hospital Nijmegen, Nijmegen, the Netherlands. Male C57Bl/6 mice. Elimination of macrophages was accomplished by administration of multilamellar liposomes that contained dichloromethylene bisphosphonate (Cl2MBP). Intravenous, intratracheal, and intraperitoneal administrations induced an elimination of liver and splenic, alveolar, and peritoneal and omental macrophages, respectively. Zymosan (1 mg/g) was injected intraperitoneally at day 0. The liposomes that contained Cl2MBP were administered before and after zymosan challenge. At day 12, all surviving mice were killed. The body weights, temperatures, and mortality rates of the mice were monitored daily. Relative organ weights (ROWs) were calculated from the lungs, liver, spleen, and kidneys after the mice were killed. The liposomes that contained Cl2MBP, administered intravenously before or after zymosan challenge, did not induce significant changes in the body weight, temperature, or mortality rate. The ROW of the liver was significantly decreased in both treatment groups. Elimination of liver and splenic macrophages after zymosan challenge induced an increased ROW of the lung and a decreased ROW of the liver. The liposomes that contained Cl2MBP, administered intratracheally before zymosan challenge, completely prevented deaths. The body weights, temperatures, and ROWs of the mice were not changed. The liposomes that contained Cl2MBP, administered intraperitoneally, did not change the body weight, temperature, or ROW. The liposomes that contained Cl2MBP, administered intraperitoneally before zymosan challenge, increased the mortality from 50% to 90%. These data show that the elimination of specific macrophage subpopulations and the elimination on specific time points in this model had differential effects, indicating a differential role of specific macrophage subpopulations, either protective or detrimental, in the development of multiple-organ dysfunction syndrome.

Macrophage differentiation and expression of macrophage colony-stimulating factor in murine milky spots and omentum after macrophage elimination.

To elucidate the differentiation mechanisms of macrophages in the murine omentum, we studied the repopulation of these cells and the expression of macrophage colony-stimulating factor (M-CSF) in the milky spots and omental tissues in mice depleted of macrophages following administration of liposome-encapsulated dichloromethylene diphosphonate (clodronate). The macrophages in the omentum were spindle or dendritic in shape, expressed several macrophage-specific antigens and Ia antigen, and phagocytized intraperitoneally injected carbon particles. In the milky spots, macrophages and macrophage precursors were detected, and the number of precursors increased after elimination of macrophages by intraperitoneal injection of liposome-encapsulated clodronate. Macrophage precursors in the milky spots proliferated, moved to the omentum, and transformed into dendritic-shaped macrophages. Expression of M-CSF mRNA extracted from the milky spots was markedly enhanced at 2 and 3 days after macrophage depletion. Localization of M-CSF protein and mRNA was observed in the stromal cells of the milky spots. In osteopetrosis (op/op) mutant mice that are defective in the production of functional M-CSF omental macrophages were absent. These results indicate that M-CSF locally produced in the milky spots plays an important role in providing a microenvironment for development and differentiation of omental macrophages.

Effect of intraperitoneal administration of granulocyte/macrophage-colony-stimulating factor in rats on omental milky-spot composition and tumoricidal activity in vivo and in vitro.

Milky spots in the greater omentum are small accumulations of leucocytes that consist mainly of macrophages and have recently shown to be a selective dissemination site of intraperitoneal (i.p.) inoculated tumour cells. However, milky-spot macrophages show tumoricidal activity and may, therefore, be an excellent source of effector cells suited for local immunotherapy. In the present study we first examined whether granulocyte/macrophage-colony-stimulating factor (GM-CSF) treatment of isolated milky-spot macrophages affects the cytotoxicity against syngeneic colon carcinoma cells (CC531) in vitro. Secondly, we studied the influence of intraperitoneal GM-CSF administration on the number and antitumour activity of milky-spot and peritoneal macrophages. All studies were performed in Wag/Rij rats in which a syngeneic colon carcinoma cell line (CC531) is available. The results of the in vitro study showed that GM-CSF treatment of the omental macrophages led to an increased cytotoxicity against the tumour cell line. Intraperitoneal administration of 1000 U GM-CSF daily for 7 consecutive days demonstrated both an enhanced antitumour activity of the milky-spot macrophages and an increase in the milky-spot macrophage population. An increase in the proliferative capacity, according to bromodeoxyuridine incorporation, was shown in the milky-spot macrophages. Taking into account both the enhanced macrophage number and their enhanced activity upon i.p. GM-CSF treatment, the milky-spot macrophages may provide a rationale for local intraperitoneal immunotherapy in the prevention of intra-abdominal tumour growth.