Extracellular Matrix Expression in Human Pancreatic Fat Cells of Patients with Normal Glucose Regulation, Prediabetes and Type 2 Diabetes.

Previously, we found that human pancreatic preadipocytes (PPAs) and islets influence each other and that the crosstalk with the fatty liver via the hepatokine fetuin-A/palmitate induces inflammatory responses. Here, we examined whether the mRNA-expression of pancreatic extracellular matrix (ECM)-forming and -degrading components differ in PPAs from individuals with normal glucose regulation (PPAs-NGR), prediabetes (PPAs-PD), and type 2 diabetes (PPAs-T2D), and whether fetuin-A/palmitate impacts ECM-formation/degradation and associated monocyte invasion. Human pancreatic resections were analyzed (immuno)histologically. PPAs were studied for mRNA expression by real-time PCR and protein secretion by Luminex analysis. Furthermore, co-cultures with human islets and monocyte migration assays in Transwell plates were conducted. We found that in comparison with NGR-PPAs, TIMP-2 mRNA levels were lower in PPAs-PD, and TGF-ß1 mRNA levels were higher in PPAs-T2D. Fetuin-A/palmitate reduced fibronectin, decorin, TIMP-1/-2 and TGF-ß1 mRNA levels. Only fibronectin was strongly downregulated by fetuin-A/palmitate independently of the glycemic status. Co-culturing of PPAs with islets increased TIMP-1 mRNA expression in islets. Fetuin-A/palmitate increased MMP-1, usherin and dermatopontin mRNA-levels in co-cultured islets. A transmigration assay showed increased monocyte migration towards PPAs, which was enhanced by fetuin-A/palmitate. This was more pronounced in PPAs-T2D. The expression of distinct ECM components differs in PPAs-PD and PPAs-T2D compared to PPAs-NGR, suggesting that ECM alterations can occur even in mild hyperglycemia. Fetuin-A/palmitate impacts on ECM formation/degradation in PPAs and co-cultured islets. Fetuin-A/palmitate also enhances monocyte migration, a process which might impact on matrix turnover.

Pancreatic fat cells of humans with type 2 diabetes display reduced adipogenic and lipolytic activity.

Obesity, especially visceral fat accumulation, increases the risk of type 2 diabetes (T2D). The purpose of this study was to investigate the impact of T2D on the pancreatic fat depot. Pancreatic fat pads from 17 partial pancreatectomized patients (PPP) were collected, pancreatic preadipocytes isolated, and in vitro differentiated. Patients were grouped using HbA1c into normal glucose tolerant (NGT), prediabetic (PD), and T2D. Transcriptome profiles of preadipocytes and adipocytes were assessed by RNAseq. Insulin sensitivity was estimated by quantifying AKT phosphorylation on Western blots. Lipogenic capacity was assessed with oil red O staining, lipolytic activity via fatty acid release. Secreted factors were measured using ELISA. Comparative transcriptome analysis of preadipocytes and adipocytes indicates defective upregulation of genes governing adipogenesis (NR1H3), lipogenesis (FASN, SCD, ELOVL6, and FADS1), and lipolysis (LIPE) during differentiation of cells from T2D-PPP. In addition, the ratio of leptin/adiponectin mRNA was higher in T2D than in NGT-PPP. Preadipocytes and adipocytes of NGT-PPP were more insulin sensitive than T2D-PPP cells in regard to AKT phosphorylation. Triglyceride accumulation was similar in NGT and T2D adipocytes. Despite a high expression of the receptors NPR1 and NPR2 in NGT and T2D adipocytes, lipolysis was stimulated by ANP 1.74-fold in NGT cells only. This stimulation was further increased by the PDE5 inhibitor dipyridamole (3.09-fold). Dipyridamole and forskolin increased lipolysis receptor independently 1.88-fold and 1.48-fold, respectively, solely in NGT cells. In conclusion, the metabolic status persistently affects differentiation and lipolysis of pancreatic adipocytes. These alterations could aggravate the development of T2D.

Metabolic crosstalk between fatty pancreas and fatty liver: effects on local inflammation and insulin secretion.

AIMS/HYPOTHESIS: Obesity-linked ectopic fat accumulation is associated with the development of type 2 diabetes. Whether pancreatic and liver steatosis impairs insulin secretion is controversial. We examined the crosstalk of human pancreatic fat cells with islets and the role of diabetogenic factors, i.e. palmitate and fetuin-A, a hepatokine released from fatty liver. METHODS: Human pancreatic resections were immunohistochemically stained for insulin, glucagon, somatostatin and the macrophage/monocyte marker CD68. Pancreatic adipocytes were identified by Oil Red O and adiponectin staining. Primary pancreatic pre-adipocytes and differentiated adipocytes were co-cultured with human islets isolated from organ donors and the metabolic crosstalk between fatty liver and fatty pancreas was mimicked by the addition of palmitate and fetuin-A. Insulin secretion was evaluated by ELISA and RIA. Cytokine expression and secretion were assessed by RT-PCR and multiplex assay, respectively. Subcellular distribution of proteins was examined by confocal microscopy and protein phosphorylation by western blotting. RESULTS: In human pancreatic parenchyma, highly differentiated adipocytes were detected in the proximity of islets with normal architecture and hormone distribution. Infiltration of adipocytes was associated with an increased number of CD68-positive cells within islets. In isolated primary pancreatic pre-adipocytes and differentiated adipocytes, palmitate and fetuin-A induced IL6, CXCL8 and CCL2 mRNA expression. Cytokine production was toll-like receptor 4 (TLR4)-dependent and further accentuated in pre-adipocytes when co-cultured with islets. In islets, IL6 and CXCL8 mRNA levels were also increased by fetuin-A and palmitate. Only in macrophages within the isolated islets, palmitate and fetuin-A stimulated the production of the cytotoxic cytokine IL-1ß. Palmitate, but not fetuin-A, exerted pro-apoptotic effects in islet cells. Instead, fetuin-A impaired glucose-induced insulin secretion in a TLR4-independent, but c-Jun N-terminal kinase- and Ca2+-dependent, manner. CONCLUSIONS/INTERPRETATION: These results provide the first evidence that fetuin-A-mediated metabolic crosstalk of fatty liver with islets may contribute to obesity-linked glucose blindness of beta cells, while fatty pancreas may exacerbate local inflammation.

Fetuin-A influences vascular cell growth and production of proinflammatory and angiogenic proteins by human perivascular fat cells.

AIMS/HYPOTHESIS: Fetuin-A (alpha2-Heremans-Schmid glycoprotein), a liver-derived circulating glycoprotein, contributes to lipid disorders, diabetes and cardiovascular diseases. In a previous study we found that perivascular fat cells (PVFCs) have a higher angiogenic potential than other fat cell types. The aim was to examine whether fetuin-A influences PVFC and vascular cell growth and the expression and secretion of proinflammatory and angiogenic proteins, and whether TLR4-independent pathways are involved. METHODS: Mono- and co-cultures of human PVFCs and endothelial cells were treated with fetuin-A and/or palmitate for 6-72 h. Proteins were quantified by ELISA and Luminex, mRNA expression by real-time PCR, and cell growth by BrDU-ELISA. Some PVFCs were preincubated with a nuclear factor κB NFκBp65 inhibitor, or the toll-like receptor 4 (TLR4) inhibitor CLI-095, or phosphoinositide 3-kinase (PI3K)/Akt inhibitors and/or stimulated with insulin. Intracellular forkhead box protein O1 (FoxO1), NFκBp65 and inhibitor of κB kinase ß (IKKß) localisation was visualised by immunostaining. RESULTS: PVFCs expressed and secreted IL-6, IL-8, plasminogen activator inhibitor 1 (PAI-1), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF)-BB, monocyte chemotactic protein-1 (MCP-1), vascular endothelial growth factor (VEGF), placental growth factor (PLGF) and hepatocyte growth factor (HGF). Fetuin-A upregulated IL-6 and IL-8, and this was potentiated by palmitate and blocked by CLI-095. Immunostaining and electrophoretic mobility shift assay (EMSA) showed partial NFκBp65 activation. MCP-1 was upregulated and blocked by CLI-095, but not by palmitate. However, HGF was downregulated, which was slightly potentiated by palmitate. This effect persisted after TLR4 pathway blockade. Stimulation of insulin-PI3K-Akt signalling by insulin resulted in nuclear FoxO1 extrusion and HGF upregulation. Fetuin-A counteracted these insulin effects. CONCLUSIONS/INTERPRETATION: Fetuin-A and/or palmitate influence the expression of proinflammatory and angiogenic proteins only partially via TLR4 signalling. HGF downregulation seems to be mediated by interference with the insulin-dependent receptor tyrosine kinase pathway. Fetuin-A may also influence angiogenic and proinflammatory proteins involved in atherosclerosis.

Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells.

Adeno-associated viral (AAV) vector suspensions produced in either human derived HEK cells or in Spodoptera frugiperda (Sf9) insect cells differ in terms of residual host cell components as well as species-specific post-translational modifications displayed on the AAV capsid proteins. Here we analysed the impact of these differences on the immunogenic properties of the vector. We stimulated human plasmacytoid dendritic cells with various lots of HEK cell-produced and Sf9 cell-produced AAV-CMV-eGFP vectors derived from different manufacturers. We found that AAV8-CMV-eGFP as well as AAV2-CMV-eGFP vectors induced lot-specific but not production platform-specific or manufacturer-specific inflammatory cytokine responses. These could be reduced or abolished by blocking toll-like receptor 9 signalling or by enzymatically reducing DNA in the vector lots using DNase. Successful HEK cell transduction by DNase-treated AAV lots and DNA analyses demonstrated that DNase did not affect the integrity of the vector but degraded extra-viral DNA. We conclude that both HEK- and Sf9-cell derived AAV preparations can contain immunogenic extra-viral DNA components which can trigger lot-specific inflammatory immune responses. This suggests that improved strategies to remove extra-viral DNA impurities may be instrumental in reducing the immunogenic properties of AAV vector preparations.

Synthetic Material Abdominal Swabs Reduce Activation of Platelets and Leukocytes Compared to Cotton Materials.

During surgical procedures, cotton abdominal swabs with their high absorptive capacity and malleability are used to retain organs and absorb blood or other body fluids. Such properties of the natural material cotton are advantageous for most operations, but in cardiopulmonary bypass (CPB) surgery, a high blood volume can accumulate in the thoracic cavity that is quickly retransfused via the heart-lung machine (HLM). This common practice is supposed to be safe due to the high anticoagulation. However, in vitro analyses showed that blood cells and plasma proteins were activated despite a high anticoagulation, which can propagate especially an inflammatory response in the patient. Thus, we investigated patients' blood during CPB surgery for inflammatory and coagulation-associated activation after contact to the HLM and either cotton or synthetic abdominal swabs. Contact with cotton significantly increased thrombocyte and neutrophil activation measured as ß-thromboglobulin and PMN-elastase secretion, respectively, compared to synthetic abdominal swabs. Both inflammatory cytokines, interleukin (IL) 1ß and IL6, were also significantly increased in the cotton over the synthetic patient group, while SDF-1α was significantly lower in the synthetic group. Our data show for the first time that cotton materials can activate platelets and leukocytes despite a high anticoagulation and that this activation is lower with synthetic materials. This additional activation due to the material on top of the activation exerted by the tissue contact that blood is exposed to during CPB surgery can propagate further reactions in patients after surgery, which poses a risk for this already vulnerable patient group.

[De-novo generation of vascularized tissue using different configurations of vascular pedicles in perforated and closed chambers]. / De-novo Generierung von vaskularisiertem Gewebe mittels unterschiedlicher Gefässstielkonfigurationen in perforierten und geschlossenen Wachstumskammern.

Growing three-dimensional tissue within a chamber requires vigorous angiogenesis initiated by, for example, an arteriovenous fistula or a ligated vascular pedicle. Growth may also be enhanced by contact with the external environment. In this study tissue growth in a rat model, vascularized via an arteriovenous loop (AV Loop) or ligated pedicle, was compared in chambers that were either closed or perforated. Chambers were harvested at 4 weeks and tissue volume and histology compared. In perforated chambers, more tissue were generated using the ligated pedicle (0.75 ml+/-0.04) than the AV Loop (0.59 ml+/-0.01). Perforated chambers generated larger volumes of tissue than closed chambers because they encouraged tissue ingrowth through the perforations. Both vessel configurations supported tissue growth but, interestingly, the ligated pedicle resulted in significantly more tissue in the perforated chambers.

Pancreatic Steatosis Associates With Impaired Insulin Secretion in Genetically Predisposed Individuals.

CONTEXT: Pancreatic steatosis leading to beta-cell failure might be involved in type 2 diabetes (T2D) pathogenesis. OBJECTIVE: We hypothesized that the genetic background modulates the effect of pancreatic fat on beta-cell function and investigated genotype × pancreatic fat interactions on insulin secretion. DESIGN: Two observational studies. SETTING: University hospital. PATIENTS OR PARTICIPANTS: A total of 360 nondiabetic individuals with elevated risk for T2D (Tuebingen Family Study [TUEF]), and 64 patients undergoing pancreatectomy (Pancreas Biobank [PB], HbA1c <9%, no insulin therapy). MAIN OUTCOME MEASURES: Insulin secretion calculated from 5-point oral glucose tolerance test (TUEF) and fasting blood collection before surgery (PB). A genome-wide polygenic score for T2D was computed from 484,788 genotyped variants. The interaction of magnetic resonance imaging-measured and histologically quantified pancreatic fat with the polygenic score was investigated. Partitioned risk scores using genome-wide significant variants were also computed to gain insight into potential mechanisms. RESULTS: Pancreatic steatosis interacted with genome-wide polygenic score on insulin secretion (P = 0.003), which was similar in the replication cohort with histological measurements (P = 0.03). There was a negative association between pancreatic fat and insulin secretion in participants with high genetic risk, whereas individuals with low genetic risk showed a positive correlation between pancreatic fat and insulin secretion. Consistent interactions were found with insulin resistance-specific and a liver/lipid-specific polygenic scores. CONCLUSIONS: The associations suggest that pancreatic steatosis only impairs beta-cell function in subjects at high genetic risk for diabetes. Genetically determined insulin resistance specifically renders pancreatic fat deleterious for insulin secretion.

Platelet derived bFGF mediates vascular integrative mechanisms of mesenchymal stem cells in vitro.

Patients with myocardial infarction reveal an altered number of circulating mesenchymal stem cells (MSCs). Recently, it was shown that MSCs are able to regenerate myocardial tissue and to differentiate into endothelial cells. The homing mechanisms of MSCs from the circulation into the target tissue, however, are not understood so far. In this study, we evaluated the impact of platelets on MSC recruitment, proliferation, migration and integration into the endothelium. MSCs expressing alpha(v)beta(3) integrin were recruited to human arterial endothelial cells exposed to isolated platelets or IL-1 beta under high shear conditions. Furthermore, induction of vascular injury in vivo resulted in increased recruitment of injected MSCs as assessed by intravital microscopy and depletion of platelets significantly reduced this adhesion. The interaction of platelets and MSCs was inhibited by pre-incubation with the mAb 7E3 or an RGD protein both blocking beta(3) integrin mediated adhesion. Platelets had a chemotactic effect on MSCs, promoted a migratory MSC phenotype and dose- and activation-dependently enhanced migration of MSCs, a process, which was mediated by basic fibroblast growth factor (bFGF). Similarly, platelet derived bFGF increased proliferation of MSCs. Coincubation of MSCs with platelets facilitated integration into an endothelial monolayer, which was significantly reduced by pre-incubation with a blocking mAb to bFGF. We conclude that platelets may play a critical part in the recruitment of MSCs to the endothelium, influence MSC function and promote integration of MSCs into the endothelium.

Platelet-derived stromal cell-derived factor-1 regulates adhesion and promotes differentiation of human CD34+ cells to endothelial progenitor cells.

BACKGROUND: Peripheral homing of progenitor cells in areas of diseased organs is critical for tissue regeneration. The chemokine stromal cell-derived factor-1 (SDF-1) regulates homing of CD34+ stem cells. We evaluated the role of platelet-derived SDF-1 in adhesion and differentiation of human CD34+ cells into endothelial progenitor cells. METHODS AND RESULTS: Adherent platelets express substantial amounts of SDF-1 and recruit CD34+ cells in vitro and in vivo. A monoclonal antibody to SDF-1 or to its counterreceptor, CXCR4, inhibits stem cell adhesion on adherent platelets under high arterial shear in vitro and after carotid ligation in mice, as determined by intravital fluorescence microscopy. Platelets that adhere to human arterial endothelial cells enhance the adhesion of CD34+ cells on endothelium under flow conditions, a process that is inhibited by anti-SDF-1. During intestinal ischemia/reperfusion in mice, anti-SDF-1 and anti-CXCR4, but not isotype control antibodies, abolish the recruitment of CD34+ cells in microcirculation. Moreover, platelet-derived SDF-1 binding to CXCR4 receptor promotes platelet-induced differentiation of CD34+ cells into endothelial progenitor cells, as verified by colony-forming assays in vitro. CONCLUSIONS: These findings imply that platelet-derived SDF-1 regulates adhesion of stem cells in vitro and in vivo and promotes differentiation of CD34+ cells to endothelial progenitor cells. Because tissue regeneration depends on recruitment of progenitor cells to peripheral vasculature and their subsequent differentiation, platelet-derived SDF-1 may contribute to vascular and myocardial regeneration.

Platelet lipoprotein interplay: trigger of foam cell formation and driver of atherosclerosis.

In the last decade, it was recognized that platelets and lipoproteins play a pivotal role in both early and late atherogenesis. Beside cellular interactions of platelets with other blood cells and vascular cells, interactions with lipoproteins seem to be quite important. Lipoproteins are fundamental 'players' in atherogenesis since they change the properties of different cells involved in atherosclerosis and thrombosis. Several studies have already shown that low density lipoproteins (LDL) are involved in the initiation of platelet signalling pathways. Platelets of hypercholesterolemic patients show hyperaggregability in vitro and enhanced activity in vivo. This review elucidates the major aspects concerning how native and modified lipoproteins influence the activation and metabolic behaviour of platelets, and shows a new way by which platelet-mediated lipoprotein transfer might contribute to foam cell formation. In hyperlipidaemia, circulating platelets are activated. This is accompanied by increased platelet aggregation, platelet-leukocyte aggregate formation, and platelet-induced superoxide anion production. Furthermore, oxidized LDL induces monocyte adhesion to the endothelium, migration and proliferation of smooth muscle cells, injures cells, interferes with nitric oxide release, and promotes procoagulant properties of vascular cells. New data about platelet-mediated lipoprotein transport and consequent foam cell formation, however, provide proof of how platelets might contribute to atheromatous lesion formation.

The immunoadhesin glycoprotein VI-Fc regulates arterial remodelling after mechanical injury in ApoE-/- mice.

AIMS: Rupture of advanced atherosclerotic plaques initiates platelet activation and aggregation as subendothelial collagen is exposed. Platelet collagen receptor glycoprotein VI (GPVI) was found to bind preferentially to the core region of human plaques. Consequently, platelets contribute to inflammatory processes and trigger atherosclerotic lesion progression. In this study, we examined binding of soluble platelet collagen receptor GPVI-Fc to atherosclerotic lesions and its effect on platelet-triggered athero-progression and neointima formation after wire-induced carotid injury. METHODS AND RESULTS: For binding studies after ligation-induced arterial injury, the left common carotid artery of C57BL/6J mice was ligated. For binding studies at spontaneously formed atherosclerotic lesion sites, Apolipoprotein E-deficient (ApoE(-/-)) mice were fed a 0.25% cholesterol diet over 16 weeks. Binding of [(124)I]GPVI-Fc was monitored by autoradiography 48 h after intravenous injection and by immunostaining. To study the effect of GPVI-Fc on neointima formation vs. control-Fc, a wire-induced injury of the left A. carotis communis of ApoE(-/-)-mice was performed. Mice were treated intraperitoneally with GPVI-Fc for 8 days and neointima formation was assessed 4 weeks after intervention. [(124)I]GPVI-Fc preferentially bound to injury sites after carotid ligation in C57BL/6J mice and to lipid-rich atherosclerotic lesions of the carotid artery and aortic arch in uninjured ApoE(-/-)-mice. Histological examinations of wire-injured carotid arteries showed that neointima formation was significantly reduced in GPVI-Fc-treated ApoE(-/-) mice compared to ApoE(-/-) mice receiving control-Fc (P < 0.05). CONCLUSION: GPVI-Fc preferentially bound to sites of vascular injury and was able to inhibit neointima formation after wire-induced vascular injury in ApoE(-/-) mice. Thus, soluble GPVI-Fc might be also a promising compound to attenuate lesion progression after plaque rupture.

What role do fat cells play in pancreatic tissue?

BACKGROUND: It is now generally accepted that obesity is a major risk factor for type 2 diabetes mellitus (T2DM). Hepatic steatosis in particular, as well as visceral and ectopic fat accumulation within tissues, is associated with the development of the disease. We recently presented the first study on isolated human pancreatic adipocytes and their interaction with islets [Gerst, F., Wagner, R., Kaiser, G., Panse, M., Heni, M., Machann, J., et al., 2017. Metabolic crosstalk between fatty pancreas and fatty liver: effects on local inflammation and insulin secretion. Diabetologia 60(11):2240-2251.]. The results indicate that the function of adipocytes depends on the overall metabolic status in humans which, in turn, differentially affects islet hormone release. SCOPE OF REVIEW: This review summarizes former and recent studies on factors derived from adipocytes and their effects on insulin-secreting ß-cells, with particular emphasis on the human pancreas. The adipocyte secretome is discussed with a special focus on its influence on insulin secretion, ß-cell survival and apoptotic ß-cell death. MAJOR CONCLUSIONS: Human pancreatic adipocytes store lipids and release adipokines, metabolites, and pro-inflammatory molecules in response to the overall metabolic, humoral, and neuronal status. The differentially regulated adipocyte secretome impacts on endocrine function, i.e., insulin secretion, ß-cell survival and death which interferes with glycemic control. This review attempts to explain why the extent of pancreatic steatosis is associated with reduced insulin secretion in some studies but not in others.

Platelets recruit human dendritic cells via Mac-1/JAM-C interaction and modulate dendritic cell function in vitro.

OBJECTIVE: Thrombotic events and immunoinflammatory processes take place next to each other during vascular remodeling in atherosclerotic lesions. In this study we investigated the interaction of platelets with dendritic cells (DCs). METHODS AND RESULTS: The rolling of DCs on platelets was mediated by PSGL-1. Firm adhesion of DCs was mediated through integrin alphaMbeta2 (Mac-1). In vivo, adhesion of DCs to injured carotid arteries in mice was mediated by platelets. Pretreatment with soluble GPVI, which inhibits platelet adhesion to collagen, substantially reduced recruitment of DCs to the injured vessel wall. In addition, preincubation of DCs with sJAM-C significantly reduced their adhesion to platelets. Coincubation of DCs with platelets induced maturation of DCs, as shown by enhanced expression of CD83. In the presence of platelets, DC-induced lymphocyte proliferation was significantly enhanced. Moreover, coincubation of DCs with platelets resulted in platelet phagocytosis by DCs, as verified by different cell phagocytosis assays. Finally, platelet/DC interaction resulted in apoptosis of DCs mediated by a JAM-C-dependent mechanism. CONCLUSIONS: Recruitment of DCs by platelets, which is mediated via CD11b/CD18 (Mac-1) and platelet JAM-C, leads to DC activation and platelet phagocytosis. This process may be of importance for progression of atherosclerotic lesions.

Periaortic Adipose Tissue Compared With Peribrachial Adipose Tissue Mass as Markers and Possible Modulators of Cardiometabolic Risk.

Increased perivascular fat mass contributes to cardiometabolic risk (CMR). High peribrachial adipose tissue (PBAT) associates with insulin resistance independently of established CMR parameters. It is unknown to what extent periaortic adipose tissue (PAAT) may have a similar impact. In 95 participants, precise quantification of total adipose tissue, PBAT, PAAT, visceral adipose tissue (VAT), and liver fat (LF) content was performed by whole-body magnetic resonance imaging. Insulin sensitivity was determined by oral glucose tolerance test and carotid intima-media thickness (cIMT) by high-resolution ultrasound. In univariate analyses, PAAT correlated with PBAT (ß = .65, P < .0001). A negative correlation of PAAT (ß = -.35, P = .0002) and PBAT (ß = -.43, P < .0001) with insulin sensitivity was observed. While in a stepwise forward regression analysis the relationship of PAAT with insulin sensitivity was no longer significant after adjustment for VAT, LF content, and other CMR factors ( P = 0.42), PBAT still correlated with insulin sensitivity ( r2 = .35, P = .01). The association between PAAT and cIMT (ß = .49, P < .0001) remained significant after adjustment for these variables ( r2 = .42, P = .0001). Although PAAT and PBAT strongly correlate, PAAT is not associated with insulin resistance, but with cIMT. Therefore, PAAT and PBAT may act differently as possible modulators of insulin resistance and subclinical atherosclerosis.

BMI-Independent Effects of Gestational Diabetes on Human Placenta.

Purpose: Recently, alterations in maternal lipid metabolism were associated with gestational diabetes mellitus (GDM). However, detailed plasma lipid profiles and their relevance for placental and fetal metabolism are currently not understood. Methods: Maternal and placental lipid profiles were characterized in women with GDM and women with normal glucose tolerance (NGT). Inflammatory gene expression was compared in placentas and primary term trophoblasts between the groups. In addition, trophoblasts were stimulated with nonesterified fatty acids (NEFAs), and effects on gene expression were quantified. Finally, placental macrophage content and cord blood concentrations of inflammatory parameters and NEFAs were compared between women with GDM and women with NGT with similar body mass index (BMI). Results: Palmitate and stearate levels were elevated in both maternal plasma and placental tissue of women with GDM. Placental GDM-associated elevations of IL6, IL8, and TLR2 expression were reflected in trophoblasts derived from women with GDM. Stimulation of primary trophoblasts with palmitate led to increased mRNA expression and protein release of the cytokine IL6 and the chemokine IL8. In line with this, elevated amounts of CD68-positive cells were quantified in the placental tissue of women with GDM. No GDM-associated elevations in a range of inflammatory parameters and NEFAs in cord blood of NGT vs GDM neonates was found. Conclusions: GDM, independently of BMI, altered maternal plasma NEFAs and the placental lipid profile. GDM was associated with trophoblast and whole-placenta lipoinflammation; however, this was not accompanied by elevated concentrations of inflammatory cytokines or NEFAs in neonatal cord blood.

Platelets induce differentiation of human CD34+ progenitor cells into foam cells and endothelial cells.

Recruitment of human CD34+ progenitor cells toward vascular lesions and differentiation into vascular cells has been regarded as a critical initial step in atherosclerosis. Previously we found that adherent platelets represent potential mediators of progenitor cell homing besides their role in thrombus formation. On the other hand, foam cell formation represents a key process in atherosclerotic plaque formation. To investigate whether platelets are involved in progenitor cell recruitment and differentiation into endothelial cells and foam cells, we examined the interactions of platelets and CD34+ progenitor cells. Cocultivation experiments showed that human platelets recruit CD34+ progenitor cells via the specific adhesion receptors P-selectin/PSGL-1 and beta1- and beta2-integrins. Furthermore, platelets were found to induce differentiation of CD34+ progenitor cells into mature foam cells and endothelial cells. Platelet-induced foam cell generation could be prevented partially by HMG coenzyme A reductase inhibitors via reduction of matrix metalloproteinase-9 (MMP-9) secretion. Finally, agonists of peroxisome proliferator-activated receptor-alpha and -gamma attenuated platelet-induced foam cell generation and production of MMP-9. The present study describes a potentially important mechanism of platelet-induced foam cell formation and generation of endothelium in atherogenesis and atheroprogression. The understanding and modulation of these mechanisms may offer new treatment strategies for patients at high risk for atherosclerotic diseases.

[Role of platelets in atherosclerosis and inflammation]. / Die Rolle von Thrombozyten bei Entzündungs- und Atheroskleroseprozessen.

BACKGROUND: Platelets are an important link between inflammation, thrombosis, and atherogenesis. Inflammation is characterized by interactions among platelets, leukocytes, and endothelial cells. These interactions trigger autocrine and paracrine activation processes that lead to leukocyte recruitment into the vascular wall. Platelet-induced chronic inflammatory processes at the vascular wall result in development of atherosclerotic lesions and atherothrombosis. METHODS: In vitro studies with co-incubated human endothelial cells and platelets after physical denudation or activation of the endothelium by cytokines provided important results for the understanding of adhesion mechanisms and the involvement of different adhesion receptors, like integrins and selectins. Furthermore, in in vivo mouse models platelet-endothelium interactions under dynamic flow conditions and increased shear stress could be analyzed. With the help of intravital microscopy, the availability of appropriate atherosclerotic animal models and first conclusive data obtained in humans, the important role of platelets in atheroprogression could be confirmed in vivo. CONCLUSION: This review highlights the molecular machinery and inflammatory pathways used by platelets to initiate and accelerate atherothrombosis. Understanding the specific requirements for platelets adhering to endothelium may lead to the development of novel therapeutic strategies.

Impact of fibroblast growth factor 21 on the secretome of human perivascular preadipocytes and adipocytes: a targeted proteomics approach.

CONTEXT: Perivascular adipose tissue (PVAT) is suggested to impact on vascular cells via humoral factors, possibly contributing to endothelial dysfunction and atherosclerosis. OBJECTIVE: To address whether the hepatokine fibroblast growth factor (FGF) 21 affects the PVAT secretome. METHODS: Human perivascular (pre)adipocytes were subjected to targeted proteomics and whole-genome gene expression analysis. RESULTS: Preadipocytes, as compared to adipocytes, secreted higher amounts of inflammatory cytokines and chemokines. Adipocytes released higher amounts of adipokines [e.g. adipisin, visfatin, dipeptidyl peptidase 4 (DPP4), leptin; p < 0.05, all]. In preadipocytes, omentin 1 release was 1.28-fold increased by FGF-21 (p < 0.05). In adipocytes, FGF-21 reduced chemerin release by 5% and enhanced DPP4 release by 1.15-fold (p < 0.05, both). FGF-21 altered the expression of four secretory genes in preadipocytes and of 18 in adipocytes (p < 0.01, all). CONCLUSION: The hepatokine FGF-21 exerts secretome-modulating effects in human perivascular (pre)adipocytes establishing a new liver-PVAT-blood vessel axis that possibly contributes to vascular inflammation and atherosclerosis.

Effects of local all-trans-retinoic acid delivery on experimental atherosclerosis in the rabbit carotid artery.

BACKGROUND: Retinoids regulate a variety of biological processes and play an important role in cell differentiation and proliferation. All-trans retinoid acid (atRA) is known to inhibit smooth muscle cell growth and thus is supposed to have favorable effects on the incidence of restenosis after percutaneous coronary interventions. The broad biological spectrum, however, leads to numerous severe side effects which limit the clinical use of a systemic application of atRA. In order to avoid systemic side effects, local delivery of atRA is preferable. The aim of this study was to evaluate the effects of atRA on the response to injury in a second-injury model of experimental balloon angioplasty. METHODS: After induction of a fibromuscular plaque in the right carotid artery of 40 New Zealand rabbits, 35 animals underwent balloon angioplasty of the preformed plaque formation. Subsequent local atRA delivery (10 ml, 10 microM) with the double-balloon catheter was performed in 15 animals. Five animals received vehicle only as sham controls, and five animals were solely electrostimulated, 15 animals served as control group with balloon angioplasty only. Vessels were excised 7 days (n=15) and 28 days (n=30) after intervention. Immunocytochemistry with antibodies against smooth muscle alpha-actin and myosin, bromodeoxyuridine, macrophages, collagen I and III and von Willebrand factor was performed. Quantitative analysis was done by computerized morphometry. RESULTS: After local atRA delivery in vivo, the extent of stenosis was markedly reduced with 21.7+/-8.3% (mean+/-S.D.) 4 weeks after intervention compared to 31.8+/-13.4% in balloon-dilated animals (P=0.0937). Both a reduced early neointimal proliferation (P=0.0002) and an increase in overall vessel diameter (4 weeks after intervention, P=0.0264) contributed to a limitation of restenosis in atRA-treated animals. Immunocytochemistry revealed a more intense alpha-actin staining pattern after local atRA therapy indicating redifferentiating effects of atRA on vascular smooth muscle cells. CONCLUSIONS: Local delivery of atRA led to limitation of restenosis formation in this animal model. The concept of a local atRA therapy might be a promising way to exploit the potential of atRA for vascular indications while minimizing the severe side effects of systemic retinoid therapy.