Secretion Of Growth Differentiation Factor 15 Research Articles

Imperatorin ameliorates pulmonary fibrosis via GDF15 expression.

Background: Pulmonary fibrosis features in damaged pulmonary structure or over-produced extracellular matrix and impaired lung function, leading to respiratory failure and eventually death. Fibrotic lungs are characterized by the secretion of pro-fibrotic factors, transformation of fibroblasts to myofibroblasts, and accumulation of matrix proteins. Hypothesis/purpose: Imperatorin shows anti-inflammatory effects on alveolar macrophages against acute lung injury. We attempt to evaluate the properties of imperatorin on the basis of fibroblasts. Methods: In in vitro, zymosan was introduced to provoke pro-fibrotic responses in NIH/3T3 or MRC-5 pulmonary fibroblasts. Imperatorin was given for examining its effects against fibrosis. The mice were stimulated by bleomycin, and imperatorin was administered to evaluate the prophylactic potential in vivo. Results: The upregulated expression of connective tissue growth factor (CTGF), α-smooth muscle actin (α-SMA), and collagen protein due to zymosan introduction was decreased by imperatorin in fibroblasts. Zymosan induced the activity of transglutaminase 2 (TGase2) and lysyl oxidase (LOX), which was also inhibited by the administration of imperatorin. Imperatorin alone enhanced sirtuin 1 (SIRT1) activity and growth differentiation factor 15 (GDF15) secretion in fibroblasts via LKB1/AMPK/CREB pathways. In addition, GDF15 exerted a beneficial effect by reducing the protein expression of CTGF, α-SMA, and collagen and the activities of TGase and LOX. Moreover, orally administered imperatorin showed prophylactic effects on bleomycin-induced pulmonary fibrosis in mice. Conclusion: Imperatorin reduces fibrotic marker expression in fibroblasts and also increases GDF15 secretion via the LKB1/AMPK/CREB pathway, attenuating pro-fibrotic responses in vitro. Imperatorin also alleviates pulmonary fibrosis induced by bleomycin in vivo.

Hepatic glucuronyl C5-epimerase combats obesity by stabilising GDF15

Disturbed hepatic metabolism frequently results in excessive lipid accumulation in the adipose tissue. However, the specific role of the liver-adipose axis in maintaining lipid homeostasis, as well as the underlying mechanism, has not yet been fully elucidated. In this study, we investigated the role of hepatic glucuronyl C5-epimerase (Glce) in the progression of obesity. We determined the association between the expression of hepatic Glce and body mass index (BMI) in obese patients. Obesity models were established in hepatic Glce-knockout and wild-type mice fed a high-fat diet (HFD) to understand the effect of Glce on obesity development. The role of Glce in the progression of disrupted hepatokine secretion was examined via secretome analysis. Hepatic Glce expression was inversely correlated with BMI in obese patients. Moreover, Glce level was found to be decreased in the liver of a HFD murine model. Hepatic Glce deficiency led to impaired thermogenesis in adipose tissue and exacerbated HFD-induced obesity. Interestingly, decreased level of growth differentiation factor 15 (GDF15) was observed in the culture medium of Glce-knockout mouse hepatocytes. Treatment with recombinant GDF15 obstructed obesity progression derived from the absence of hepatic Glce, similar to the effect of Glce or its inactive mutant overexpressed both invitro and invivo. Furthermore, liver Glce deficiency led to diminished production and increased degradation of mature GDF15, resulting in reduced hepatic GDF15 secretion. Hepatic Glce deficiency facilitated obesity development, and decreased Glce expression further reduced hepatic secretion of GDF15, thereby perturbing lipid homeostasis invivo. Therefore, the novel Glce-GDF15 axis plays an important role in maintaining energy balance and may act as a potential target for combating obesity. Evidence suggests that GDF15 plays a key role in hepatic metabolism; however, the molecular mechanism for regulating its expression and secretion is largely unknown. Our work observes that hepatic Glce, as a key Golgi-localised epimerase, may work on the maturation and post-translational regulation of GDF15. Hepatic Glce deficiency reduces the production of mature GDF15 protein and facilitates its ubiquitination, resulting in the aggravation of obesity development. This study sheds light on the new function and mechanism of the Glce-GDF15 axis in lipid metabolism and provides a potential therapeutic target against obesity.

Abstract 5282: Mitotane induces GDF-15 expression in vitro and in vivo: A potential therapy-induced mechanism of immune escape in adrenocortical carcinoma

Abstract Context: Adrenocortical carcinoma (ACC) is a rare endocrine malignancy with poor prognosis and limited treatment options. The only approved therapy is mitotane, an adrenal toxicant, alone or in combination with cytotoxic chemotherapy. Response rate is only ~20%. Results of later line immune checkpoint inhibition (ICI) in ACC are disappointing, most likely due to an immunologically cold, immunosuppressive microenvironment. Among the factors induced in mitotane-treated ACC cell lines, we previously found growth/differentiation factor 15 (GDF-15), an anti-inflammatory cytokine physiologically expressed during pregnancy. Aim: As GDF-15 is an emerging mediator of tumor immune escape, we investigated changes in GDF-15 secretion upon mitotane treatment in ACC in vitro and in vivo, and explored a potential correlation with response to therapy and survival. Methods: Four different human ACC cell lines (NCI-H295R, CU-ACC1, CU-ACC2 and JIL-2266) were treated with mitotane. GDF-15 secretion and expression were determined by ELISA and immunoblot. GDF-15 levels in sera of 142 ACC patients (94 prior and 48 during mitotane) were analyzed by ELISA and correlated with survival from the time point of blood collection. GDF-15 serum levels were quantified in three responders and seven non-responders to ICI treatment. Results: In vitro, 24h treatment with 25 µM mitotane-induced GDF-15 secretion and expression in all cell lines by up to one order of magnitude. In 32 patient samples, GDF-15 serum concentrations increased from 0.6±1 ng/l prior mitotane to 2.1 ± 2.7 ng/l after mitotane initiation, p<0.0001.Importantly, patients with low serum GDF-15 levels before and during mitotane treatment had a significantly longer overall survival compared to patients with higher GDF-15 serum levels. This association retained statistical significance after adjustment for the known prognostic factors Ki67 and ENSAT tumour stage in the mitotane treated group (95% CI 1.15-.1.73, HR1.41, p=0.001). In an exploratory cohort of three responders and seven non-responders to ICI we found a trend for higher GDF-15 serum concentrations in the non-responders. Conclusion: Mitotane induces GDF-15 both in vivo and in vitro and potentially contributes to the poor response rates to ICI in ACC. Citation Format: Isabel Weigand, Tanja Anderlik, Jochen Schreiner, Hanna Remde, Otilia Kimpel, Laura-Sophie Landwehr, Eva Hoster, Florian Wedekink, Alexandra S. Triebig, Tanja Maier, Jörg Wischhusen, Martin Reincke, Martin Fassnacht, Matthias Kroiss. Mitotane induces GDF-15 expression in vitro and in vivo: A potential therapy-induced mechanism of immune escape in adrenocortical carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5282.

Berberine Ameliorates Obesity by Inducing GDF15 Secretion by Brown Adipocytes.

Berberine (BBR), which is a compound derived from the Chinese medicinal plant Coptis chinensis, promotes weight loss, but the molecular mechanisms are not well understood. Here, we show that BBR increases the serum level of growth differentiation factor 15 (GDF15), which is a stress response cytokine that can reduce food intake and lower body weight in diet-induced obese (DIO) mice. The body weight and food intake of DIO mice were decreased after BBR treatment, and the weight change was negatively correlated with the serum GDF15 level. Further studies show that BBR induced GDF15 mRNA expression and secretion in the brown adipose tissue (BAT) of DIO mice and primary mouse brown adipocytes. In addition, we found that BBR upregulates GDF15 mRNA expression and secretion by activating the integrated stress response (ISR) in primary mouse brown adipocytes. Overall, our findings show that BBR lowers body weight by inducing GDF15 secretion via the activation of the ISR in BAT.

Role of GDF15/MAPK14 Axis in Chondrocyte Senescence as a Novel Senomorphic Agent in Osteoarthritis.

Osteoarthritis (OA) is most prevalent in older individuals and exerts a heavy social and economic burden. However, an effective and noninvasive approach to OA treatment is currently not available. Chondrocyte senescence has recently been proposed as a key pathogenic mechanism in the etiology of OA. Furthermore, senescent chondrocytes (SnCCs) can release various proinflammatory cytokines, proteolytic enzymes, and other substances known as the senescence-associated secretory phenotype (SASP), allowing them to connect with surrounding cells and induce senesce. Studies have shown that the pharmacological elimination of SnCCs slows the progression of OA and promotes regeneration. Growth differentiation factor 15 (GDF15), a member of the tumor growth factor (TGF) superfamily, has recently been identified as a possible aging biomarker and has been linked to a variety of clinical conditions, including coronary artery disease, diabetes, and multiple cancer types. Thus, we obtained data from a publicly available single-cell sequencing RNA database and observed that GDF15, a critical protein in cellular senescence, is highly expressed in early OA. In addition, GDF15 is implicated in the senescence and modulation of MAPK14 in OA. Tissue and synovial fluid samples obtained from OA patients showed overexpression of GDF15. Next, we treated C20A4 cell lines with interleukin (IL)-1β with or without shGDF15 then removed the conditioned medium, and cultured C20A4 and HUVEC cell lines with the aforementioned media. We observed that C20A4 cells treated with IL-1β exhibited increased GDF15 secretion and that chondrocytes cultured with media derived from IL-1β–treated C20A4 exhibited senescence. HUVEC cell migration and tube formation were enhanced after culturing with IL-1β-treated chondrocyte media; however, decreased HUVEC cell migration and tube formation were noted in HUVEC cells cultured with GDF15-loss media. We tested the potential of inhibiting GDF15 by using a GDF15 neutralizing antibody, GDF15-nAb. GDF15-nAb exerted a similar effect, resulting in the molecular silencing of GDF15 in vivo and in vitro. Our results reveal that GDF15 is a driver of SnCCs and can contribute to OA progression by inducing angiogenesis.

Ionizing Radiation-Induced GDF15 Promotes Angiogenesis in Human Glioblastoma Models by Promoting VEGFA Expression Through p-MAPK1/SP1 Signaling.

Glioblastoma multiforme (GBM), the most aggressive cancer type that has a poor prognosis, is characterized by enhanced and aberrant angiogenesis. In addition to surgical resection and chemotherapy, radiotherapy is commonly used to treat GBM. However, radiation-induced angiogenesis in GBM remains unexplored. This study examined the role of radiation-induced growth/differentiation factor-15 (GDF15) in regulating tumor angiogenesis by promoting intercellular cross-talk between brain endothelial cells (ECs) and glioblastoma cells. Radiation promoted GDF15 secretion from human brain microvascular endothelial cells (HBMVECs). Subsequently, GDF15 activated the transcriptional promoter VEGFA in the human glioblastoma cell line U373 through p-MAPK1/SP1 signaling. Upregulation of vascular endothelial growth factor (VEGF) expression in U373 cells resulted in the activation of angiogenic activity in HBMVECs via KDR phosphorylation. Wound healing, tube formation, and invasion assay results revealed that the conditioned medium of recombinant human GDF15 (rhGDF15)-stimulated U373 cell cultures promoted the angiogenic activity of HBMVECs. In the HBMVEC-U373 cell co-culture, GDF15 knockdown mitigated radiation-induced VEGFA upregulation in U373 cells and enhanced angiogenic activity of HBMVECs. Moreover, injecting rhGDF15-stimulated U373 cells into orthotopic brain tumors in mice promoted angiogenesis in the tumors. Thus, radiation-induced GDF15 is essential for the cross-talk between ECs and GBM cells and promotes angiogenesis. These findings indicate that GDF15 is a putative therapeutic target for patients with GBM undergoing radio-chemotherapy.

Cytokines secreted by mesenchymal stem cells reduce demyelination in an animal model of Charcot-Marie-Tooth disease

IntroductionDemyelinating Charcot-Marie-Tooth disease (CMT) is caused by mutations in the genes that encode myelinating proteins or their transcription factors. Our study thus sought to assess the therapeutic effects of cytokines secreted from mesenchymal stem cells (MSCs) on this disease. MethodsThe therapeutic potential of Wharton's jelly MSCs (WJ-MSCs) and cytokines secreted by WJ-MSCs was evaluated on Schwann cells (SCs) exhibiting demyelination features, as well as a mouse model of demyelinating CMT. ResultsCo-culture with WJ-MSC protected PMP22-overexpressing SCs from apoptotic cell death. Using a cytokine array, the secretion of growth differentiation factor-15 (GDF-15) and amphiregulin (AREG) was found to be elevated in WJ-MSCs when co-incubated with the PMP22-overexpressing SCs. Administration of both cytokines into trembler-J (Tr-J) mice, an animal model of CMT, significantly enhanced motor nerve conduction velocity compared to the control group. More importantly, this treatment alleviated the demyelinating phenotype of Tr-J mice, as demonstrated by an improvement in the mean diameter and g-ratio of the myelinated axons. ConclusionsOur findings demonstrated that WJ-MSCs alleviate the demyelinating phenotype of CMT via the secretion of several cytokines. Further elucidation of the underlying mechanisms of GDF-15 and AREG in myelination might provide a robust basis for the development of effective therapies against demyelinating CMT.

Integrated stress response regulates GDF15 secretion from adipocytes, preferentially suppresses appetite for a high-fat diet and improves obesity

SummaryThe eIF2α phosphorylation-dependent integrated stress response (ISR) is a signaling pathway that maintains homeostasis in mammalian cells exposed to various stresses. Here, ISR activation in adipocytes improves obesity and diabetes by regulating appetite in a non-cell-autonomous manner. Adipocyte-specific ISR activation using transgenic mice decreases body weight and improves glucose tolerance and obesity induced by a high-fat diet (HFD) via preferential inhibition of HFD intake. The transcriptome analysis of ISR-activated adipose tissue reveals that growth differentiation factor 15 (GDF15) expression is induced by the ISR through the direct regulation of the transcription factors ATF4 and DDIT3. Deficiency in the GDF15 receptor GFRAL abolishes the adipocyte ISR-dependent preferential inhibition of HFD intake and the anti-obesity effects. Pharmacologically, 10(E), 12(Z)-octadecadienoic acid induces ISR-dependent GDF15 expression in adipocytes and decreases the intake of the HFD. Based on our findings the specific activation of the ISR in adipocytes controls the non-cell-autonomous regulation of appetite.

Abstract 993: GDF15 is a candidate circulating biomarker of eribulin response in breast cancer

Abstract Microtubule-targeting agents (MTAs) constitute a diverse group of chemical compounds that bind to microtubules and affect their structure and function. Disruption of microtubules induces various cellular responses, often leading to cell cycle arrest or cell death. Eribulin is a microtubule dynamic inhibitor approved for treatment in patients with metastatic breast cancer (MBC) who had previously been administered chemotherapy. Although MTAs are widely used in the clinic, there is a research gap in the identification of pharmacodynamic (PD) biomarkers that could be used as surrogates during the course of treatment and measured non-invasively. The proteomic interrogation of the cancer secretome has emerged as a promising strategy for biomarker and drug target discovery. Secretomes have much lower complexity than plasma and tissue homogenates and are likely to be enriched with tumour derived-proteins entering into circulation. Quantitative proteomics analysis of the cancer secretome could help monitor critical aspects of cancer progression and therapeutics. The working hypothesis for this study is that eribulin induces a specific secretome that can be measured by quantitative proteomics, and could be used to identify response biomarkers linked to the action of the drug. We used three cell line models, MDA-MB-231 and HS578T (representing triple-negative BC) and MCF7 (representing luminal BC), all of them sensitive to treatment with eribulin. After a 7-day treatment with an IC70 of eribulin, we identified a subpopulation of cells with a lower proliferation rate, a slow-cycling subpopulation of cells that survived the large cell death induced by the drug. This slow-cycling population is alive in the presence of eribulin but remains in a quiescent state. The secretome profiling of slow-cycling cells revealed an oversecretion of the growth differentiation factor 15 (GDF15) when compared to parental cells and cells that after continuous exposure to eribulin start proliferating and become drug-tolerant. Interestingly, drug-tolerant cells lose the secretion of GDF15, suggesting a role for GDF15 in the response to eribulin treatment. Currently, we are validating GDF15 as a candidate circulating biomarker for the response to eribulin in patient-derived xenograft (PDX) 3D-models derived from metastatic BC patients. Furthermore, we are conducting loss of GDF15 function experiments in slow-cycling cells to understand whether the secretion of GDF15 could also contribute to the establishment and/or maintenance of the slow-cycling state in cells responding to eribulin.We envision that our work will contribute to the validation of GDF15 as a circulating biomarker that could be used to monitor eribulin's activity in the clinical setting. Furthermore, these results could also help understand the initiation of acquired resistance to eribulin. Citation Format: Chiara Bellio, Marta Emperador, Laia Garrigós, Esther Zamora, Cristina Saura, Bruce A. Littlefield, Josep Villanueva. GDF15 is a candidate circulating biomarker of eribulin response in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 993.

GDF-15 as a Weight Watcher for Diabetic and Non-Diabetic People Treated With Metformin.

Weight gain and obesity are global health concerns contributing to morbidity with increased risks of cardiovascular disease, diabetes, liver steatohepatitis and cancer. Pharmacological therapies or bariatric surgery are often required for those who fail to adhere to diet and lifestyle modifications. Metformin, a widely used antidiabetic agent, seems to have a health benefit beyond its anti-hyperglycemic properties, with few side effects. Emerging evidence shows weight loss to be associated with metformin in both diabetic and non-diabetic individuals. Recently, the growth differentiation factor 15 (GDF-15), a member of the transforming growth factor beta superfamily, has been identified as a key mediator of metformin-induced weight loss. Metformin increases the secretion of GDF-15, which binds exclusively to glial cell-derived neurotrophic factor family receptor alpha-like (GFRAL). This gut-brain cytokine works as a prominent player in reducing food intake and body weight in health and disease, like anorexia nervosa and cancer. Herein, we critically review advances in the understanding of the weight-reducing effects of metformin via the GDF-15 pathway.

Potential associations of circulating growth differentiation factor-15 with sex hormones in male patients with coronary artery disease

This study aimed to explore the association between growth differentiation factor-15 (GDF-15), a stress-induced factor, and sex hormones in male patients with coronary artery disease (CAD). In this study, we recruited 253 male patients with CAD and 205 male controls. Patients were divided into three groups in accordance with GDF-15 tertiles. Serum levels of GDF-15, testosterone, estradiol and other biochemical variables were measured. Serum levels of GDF-15 were significantly increased and serum testosterone and testosterone/estradiol ratio (T/E2 ratio) were significantly decreased in CAD patients compared with controls. Patients with high GDF-15 levels had lower testosterone (203.97, 95% CI 154.67–328.30 vs. 303.98, 95% CI 246.93–345.66; P = 0.001) and T/E2 ratio (8.82, 95% CI 5.77–11.41 vs. 11.07, 95% CI 7.91–14.32; P = 0.013). Correlation analyses showed that serum GDF-15 levels inversely correlated with testosterone levels (r = −0.339) and T/E2 ratio (r = −0.365) (both P < 0.001). In multivariate regression analyses, the association between GDF-15 and T/E2 ratio was maintained (B=-0.442, 95% CI -99.568 to -6.991, P = 0.015). Furthermore, in vitro studies showed a synergistic effect of testosterone and estradiol on GDF-15 secretion, and demonstrated that testosterone association with estradiol decreased GDF-15 secretion through androgen receptor/estrogen receptor-mediated pathways. Together, these results suggest that upregulation of GDF-15 in the presence of low and imbalanced sex hormone levels may contribute to CAD. Thus, restoring the balance of testosterone and estradiol may inhibit the effects of GDF-15 and serve as a promising therapeutic strategy for the treatment of CAD.

Regulation of Systemic Glucose Homeostasis by T Helper Type 2 Cytokines.

Obesity results in an inflammatory microenvironment in adipose tissue, leading to the deterioration of tissue protective mechanisms. Although recent studies suggested the importance of type 2 immunity in an anti-inflammatory microenvironment in adipose tissue, the regulatory effects of T helper 2 (Th2) cytokines on systemic metabolic regulation are not fully understood. Recently, we identified the roles of the Th2 cytokine (interleukin 4 [IL-4] and IL-13)-induced adipokine, growth differentiation factor 15 (GDF15), in adipose tissue in regulating systemic glucose metabolism via signal transducer and activator of transcription 6 (STAT6) activation. Moreover, we showed that mitochondrial oxidative phosphorylation is required to maintain these macrophage-regulating autocrine and paracrine signaling pathways via Th2 cytokine-induced secretion of GDF15. In this review, we discuss how the type 2 immune response and Th2 cytokines regulate metabolism in adipose tissue. Specifically, we review the systemic regulatory roles of Th2 cytokines in metabolic disease and the role of mitochondria in maintenance of type 2 responses in adipose tissue homeostasis.

Effect of growth differentiation factor-15 secreted by human umbilical cord blood-derived mesenchymal stem cells on amyloid beta levels in in vitro and in vivo models of Alzheimer's disease

Alzheimer's disease (AD), which is the most common progressive neurodegenerative disease, causes learning and memory impairment. The pathological progress of AD can derive from imbalanced homeostasis of amyloid beta (Aβ) in the brain. In such cases, microglia play important roles in regulating the brain Aβ levels. In the present study, we found that human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) can increase, through paracrine action, the ability of microglial cells to clear Aβ. In order to identify the associated paracrine factors, a secretome of hUCB-MSCs co-cultured with Aβ-treated BV2 microglial cells was analyzed using a human cytokine protein array. As a result, growth differentiation factor-15 (GDF-15) was identified as a predominant candidate, and its association with Aβ clearance by microglial cells was investigated in vitro and in a 5XFAD mouse model. When Aβ-treated BV2 cells were treated with exogenous recombinant GDF-15, the Aβ levels in the culture medium decreased. Moreover, GDF-15 injection in the brain parenchyma of 5XFAD mice also led to decrease in Aβ plaques. In contrast, co-culture of BV2 cells and hUCB-MSCs treated with GDF-15-specific siRNA did not influence the Aβ levels in the culture medium. To elucidate how these phenomena are related, we confirmed that GDF-15 specifically increases insulin-degrading enzyme (IDE) expression in microglial cells through TGFβ receptor type II (TGFβRII), both in vitro and in vivo. These findings suggest that hUCB-MSCs promote the Aβ clearance ability of microglial cells through regulation of GDF-15 secretion, thus elucidating a therapeutic mechanism for AD.

Treatment-damaged hepatocellular carcinoma promotes activities of hepatic stellate cells and fibrosis through GDF15

The aim of this study was to investigate whether treatment-damaged hepatocellular carcinoma (HCC) would accelerate liver cirrhosis through promoting the activities of hepatic stellate cells (HSCs). HCC cells were exposed to chemotherapeutic agent or hypoxia to mimic the transarterial chemoembolization (TACE)-like treatment. Growth differentiation factor 15 (GDF15) expression was increased in cisplatin- or hypoxia-treated HCC cells. Treatment-induced GDF15 increase in HCC cells was mediated by p38MAPK, JNK, ERK1/2 activation. GDF15 from treatment-damaged HCC cells enhanced the proliferation and collagen synthesis of HSCs through ERK1/2- and Smad3-dependent pathways. Metformin significantly reduced the GDF15 production from treatment-damaged HCC cells by targeting JNK. The use of metformin could attenuate the in vivo fibrotic activities of HSCs promoted by treatment-damaged HCC cells and inhibit GDF15 expression. In conclusion, treatment-damaged HCC accelerates fibrosis by promoting the activities of HSCs via GDF15 secretion, which could be reversed by metformin. This provides a potential therapeutic target for alleviating TACE-aggravated liver cirrhosis.

Consolidation Chemotherapy Prevents Relapse by Indirectly Regulating Bone Marrow Adipogenesis in Patients with Acute Myeloid Leukemia

Background/Aims: Chemotherapy is still the main strategy used to prevent the relapse of acute myeloid leukaemia (AML). As the most abundant stromal component in bone marrow (BM), marrow adipocytes have been previously shown to promote leukaemogenesis. The present study was designed to further validate whether marrow adipocytes exert synergistic effects on strengthening chemotherapeutic efficacy and evaluate the underlying mechanism. Methods: A retrospective study of BM biopsies from 80 patients with AML in remission and 71 control subjects was applied to quantitatively analyse the marrow adipocyte volume. Toxicity tests were used to assess the effect of chemotherapy drugs on BM cells. The possible mechanisms by which chemotherapy regulated the reduced marrow adipocyte content were investigated using antibody neutralization experiments, with an emphasis on growth differentiation factor 15 (GDF15). Results: In our study, the marrow adipocyte content was obviously reduced in the AML- complete remission (CR) group compared with the control group (P<0.001). Moreover, patients with a reduced adipocyte content exhibited longer relapse-free survival (RFS) (P<0.001). We also confirmed that GDF15 was overexpressed in mononuclear cells (MNCs) after treatment with chemotherapy drugs and partially blocked mesenchymal stem cells (MSCs) adipogenesis. Intriguingly, this inhibitory effect on adipogenesis was rescued by treatment with a neutralizing anti-GDF15 antibody. Conclusion: Chemotherapy indirectly inhibited adipogenesis by promoting GDF15 secretion from BM MNCs, subsequently strengthening the efficacy of consolidation chemotherapy in patients with AML during CR.

Growth differentiation factor 15 is a myomitokine governing systemic energy homeostasis.

Reduced mitochondrial electron transport chain activity promotes longevity and improves energy homeostasis via cell-autonomous and -non-autonomous factors in multiple model systems. This mitohormetic effect is thought to involve the mitochondrial unfolded protein response (UPRmt), an adaptive stress-response pathway activated by mitochondrial proteotoxic stress. Using mice with skeletal muscle-specific deficiency of Crif1 (muscle-specific knockout [MKO]), an integral protein of the large mitoribosomal subunit (39S), we identified growth differentiation factor 15 (GDF15) as a UPRmt-associated cell-non-autonomous myomitokine that regulates systemic energy homeostasis. MKO mice were protected against obesity and sensitized to insulin, an effect associated with elevated GDF15 secretion after UPRmt activation. In ob/ob mice, administration of recombinant GDF15 decreased body weight and improved insulin sensitivity, which was attributed to elevated oxidative metabolism and lipid mobilization in the liver, muscle, and adipose tissue. Thus, GDF15 is a potent mitohormetic signal that safeguards against the onset of obesity and insulin resistance.

Morphological Effects on Expression of Growth Differentiation Factor 15 (GDF15), a Marker of Metastasis

Cancer cells typically demonstrate altered morphology during the various stages of disease progression as well as metastasis. While much is known about how altered cell morphology in cancer is a result of genetic regulation, less is known about how changes in cell morphology affect cell function by influencing gene expression. In this study, we altered cell morphology in different types of cancer cells by disrupting the actin cytoskeleton or by modulating attachment and observed a rapid up-regulation of growth differentiation factor 15 (GDF15), a member of the transforming growth factor-beta (TGF-β) super-family. Strikingly, this up-regulation was sustained as long as the cell morphology remained altered but was reversed upon allowing cell morphology to return to its typical configuration. The potential significance of these findings was examined in vivo using a mouse model: a small number of cancer cells grown in diffusion chambers that altered morphology increased mouse serum GDF15. Taken together, we propose that during the process of metastasis, cancer cells experience changes in cell morphology, resulting in the increased production and secretion of GDF15 into the surrounding environment. This indicates a possible relationship between serum GDF15 levels and circulating tumor cells may exist. Further investigation into the exact nature of this relationship is warranted.

Abstract 471: Triglyceride-rich Lipoprotein Lipolysis Products Induce Adipose Differentiation Related Protein Expression and Proatherogenic Cytokine Secretion in Monocytes

Elevated blood triglyceride levels are associated with an increased risk of atherosclerotic cardiovascular disease. Blood triglycerides are contained within very low density lipoproteins (VLDL) and chylomicrons, which are referred to as triglyceride-rich lipoproteins (TGRL). The enzyme lipoprotein lipase is responsible for the hydrolysis of these triglycerides, primarily into free fatty acids and glycerol. Our lab has shown that these lipolysis products induce lipid droplet formation in monocytes and can induce a proinflammatory state in multiple cell types. Other investigators have linked adipose differentiation related protein (ADRP, also known as perilipin 2 or adipophilin) to lipid droplet formation in monocyte-derived macrophage foam cells. The objectives of this study are to further understand the effects of TGRL lipolysis products on the formation of lipid droplets in monocytes and monocyte activation. We hypothesize that TGRL lipolysis products increase the expression of ADRP and the secretion of proatherogenic cytokines in THP-1 monocytes. In this study, we analyzed the global gene expression in THP-1 monocytes exposed to TGRL lipolysis products. ADRP was identified as a principal response lipid droplet associated gene induced by exposure to TGRL lipolysis products when compared to the control treatment. Induction of protein expression was analyzed by western blot. The TGRL lipolysis product treatment was also found to induce genes associated with atherosclerosis and monocyte adherence to endothelium including growth differentiation factor 15 (GDF15) and interleukin 8 (IL-8) when compared to the control treatment. We measured the secretion of GDF15 and IL-8 by ELISA. Our results indicate that mRNA and protein expression of ADRP and secretion of proatherogenic cytokines (GDF15 and IL-8) were increased in monocytes after exposure to TGRL lipolysis products. In conclusion, our study demonstrates that TGRL lipolysis products induce the expression of ADRP and the secretion of proatherogenic cytokines in monocytes. These results may indicate a role for TGRL lipolysis products in the increased risk of atherosclerotic cardiovascular disease for individuals with elevated blood triglyceride levels.

Discovery of Growth Differentiation Factor 15 as an Erythroblast-Secreted Regulator of Hepcidin with Very High Level Expression in Patients with Thalassemia.

Iron overload causes considerable morbidity associated with thalassemia due to inappropriate suppression of the iron regulator, hepcidin. One possible explanation for this phenomenon is that protein(s) that are normally secreted into the marrow microenvironment by erythroblasts become endocrine signals in patients with thalassemia due to the myeloproliferative nature of the disease. To test this hypothesis, progenitor and precursor cell transcriptional profiles were generated with Affymetrix GeneChip Human Genome U133 Plus 2.0 Arrays using primary erythroblasts cultured from 15 healthy human donors. For this study, informatic analyses were focused upon 54 members of the TGF–B/BMP superfamily. Among the subset of genes identified by this screen, evidence for high-level expression of a gene named growth differentiation factor 15 (GDF15) was discovered in the precursor cell profiles. Quantitative PCR, Western, and ELISA analyses confirmed expression and secretion of GDF15 during erythroblast maturation. GDF15 is an apoptosis-associated protein expressed primarily by the placenta. To determine whether GDF15 serves as a regulator of hepcidin expression, hepcidin expression assays were performed using a human hepatoma cell line (HuH-7). BMP2 and BMP4 were studied for comparison. Addition of BMP2 and BMP4 (range 10,000–500,000 pg/ml) resulted in dosed increases in hepcidin mRNA (5–30 fold). At the concentrations of GDF15 normally found in human blood (500 pg/ml), a 2-fold increase in the expression of hepcidin was measured compared to matched cultures containing no supplemental GDF15. However, GDF15 dosed to levels above 5,000 pg/ml resulted in a significant reduction in hepcidin expression. Next, plasma levels of GDF15 were measured in the peripheral blood of 162 donors (donor groups included healthy controls, sickle-cell syndromes, thalassemia syndromes, and other causes of anemia) to determine whether plasma GDF15 levels are dysregulated in thalassemia. Plasma samples from 21 hereditary hemochromatosis donors provided evidence that significantly elevated GDF15 expression is not associated with iron overload in the absence of erythroid pathology. Compared with mean plasma GDF15 levels of 536±222 pg/ml among the control samples, only patients with thalassemia intermedia, thalassemia major, and HbE-beta thalassemia showed significantly elevated plasma levels of GDF15 (9 donors; mean GDF15 24,600 pg/ml; range 8,980–75,100 pg/ml; p<0.01). Elevated ferritin levels (>370 ug/L) were noted in each of those patients regardless of their transfusion or chelation history. These novel findings suggest that GDF15 is secreted from human erythroblasts, released into the circulation at extremely high levels in thalassemia patients, and contributes to iron overloading in those patients by suppressing hepcidin expression.