<em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>), a metabolic hormone predominantly produced by the liver, is also expressed in adipocytes and the pancreas. It regulates glucose and lipid metabolism through pleiotropic actions in these tissues and the brain. In mice, fasting leads to increased PPAR-α mediated expression of FGF<em>21</em> in the liver where it stimulates gluconeogenesis, fatty acid oxidation, and ketogenesis, as an adaptive response to fasting and starvation. In the fed state, FGF<em>21</em> acts as an autocrine <em>factor</em> in adipocytes, regulating the activity of PPAR-γ through a feed-forward loop mechanism. Administration of recombinant FGF<em>21</em> has been shown to confer multiple metabolic benefits on insulin sensitivity, blood glucose, lipid profile and body weight in obese mice and diabetic monkeys, without mitogenic or other side effects. Such findings highlight the potential role of FGF<em>21</em> as a therapeutic agent for obesity-related medical conditions. However, in human studies, high circulating FGF<em>21</em> levels are found in obesity and its related cardiometabolic disorders including the metabolic syndrome, type 2 diabetes, non-alcoholic fatty liver disease and coronary artery disease. These findings may indicate the presence of FGF<em>21</em> resistance or compensatory responses to the underlying metabolic stress, and imply the need for supraphysiological doses of FGF<em>21</em> to achieve therapeutic efficacy. On the other hand, serum FGF<em>21</em> has been implicated as a potential biomarker for the early detection of these cardiometabolic disorders. This review summarizes recent developments in the understanding of FGF<em>21</em>, from physiological and clinical perspectives.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>) is active in murine adipocytes and has beneficial metabolic effects in animal models of type 2 diabetes mellitus. We assessed whether FGF<em>21</em> influences lipolysis in human adipocytes and 3T3-L1 cells. FGF<em>21</em> had no short-time effect (h) while a 3-day incubation with FGF<em>21</em> attenuated hormone-stimulated lipolysis. FGF<em>21</em> did not influence the mRNA expression of genes involved in regulating lipolysis, but significantly reduced the expression of the lipid droplet-associated phosphoprotein perilipin without affecting differentiation. Via reduced release of fatty acids into the circulation, the anti-lipolytic effect could be a mechanism through which FGF<em>21</em> promotes insulin sensitivity in man.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> (FGF)-<em>21</em> is a novel regulator of insulin-independent glucose transport in 3T3-L1 adipocytes and has glucose and triglyceride lowering effects in rodent models of diabetes. The precise mechanisms whereby FGF-<em>21</em> regulates metabolism remain to be determined. Here we describe the early signaling events triggered by FGF-<em>21</em> treatment of 3T3-L1 adipocytes and reveal a functional interplay between FGF-<em>21</em> and peroxisome proliferator-activated receptor gamma (PPARgamma) pathways that leads to a marked stimulation of glucose transport. While the early actions of FGF-<em>21</em> on 3T3-L1 adipocytes involve rapid accumulation of intracellular calcium and phosphorylation of Akt, GSK-3, p70(S6K), SHP-2, MEK1/2, and Stat3, continuous treatment for 72 h induces an increase in PPARgamma protein expression. Moreover, chronic activation of the PPARgamma pathway in 3T3-L1 adipocytes with the PPARgamma agonist and anti-diabetic agent, rosiglitazone (BRL 49653), enhances FGF-<em>21</em> action to induce tyrosine phosphorylation of FGF receptor-2. Strikingly, treatment of cells with FGF-<em>21</em> and rosiglitazone in combination leads to a pronounced increase in expression of the GLUT1 glucose transporter and a marked synergy in stimulation of glucose transport. Together these results reveal a novel synergy between two regulators of glucose homeostasis, FGF-<em>21</em> and PPARgamma, and further define FGF-<em>21</em> mechanism of action.

BACKGROUND

Macronutrient intake varies substantially between individuals, and there is evidence that this variation is partly accounted for by genetic variants.

OBJECTIVE

The objective of the study was to identify common genetic variants that are associated with macronutrient intake.

METHODS

We performed 2-stage genome-wide association (GWA) meta-analysis of macronutrient intake in populations of European descent. Macronutrients were assessed by using food-frequency questionnaires and analyzed as percentages of total energy consumption from total fat, protein, and carbohydrate. From the discovery GWA (n = 38,360), 35 independent loci associated with macronutrient intake at P < 5 × 10(-6) were identified and taken forward to replication in 3 additional cohorts (n = 33,533) from the DietGen Consortium. For one locus, fat mass obesity-associated protein (FTO), cohorts with Illumina MetaboChip genotype data (n = 7724) provided additional replication data.

RESULTS

A variant in the chromosome 19 locus (rs838145) was associated with higher carbohydrate (β ± SE: 0.25 ± 0.04%; P = 1.68 × 10(-8)) and lower fat (β ± SE: -0.<em>21</em> ± 0.04%; P = 1.57 × 10(-9)) consumption. A candidate gene in this region, <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>), encodes a <em>fibroblast</em> <em>growth</em> <em>factor</em> involved in glucose and lipid metabolism. The variants in this locus were associated with circulating FGF<em>21</em> protein concentrations (P < 0.05) but not mRNA concentrations in blood or brain. The body mass index (BMI)-increasing allele of the FTO variant (rs14<em>21</em>085) was associated with higher protein intake (β ± SE: 0.10 ± 0.02%; P = 9.96 × 10(-10)), independent of BMI (after adjustment for BMI, β ± SE: 0.08 ± 0.02%; P = 3.15 × 10(-7)).

CONCLUSIONS

Our results indicate that variants in genes involved in nutrient metabolism and obesity are associated with macronutrient consumption in humans. Trials related to this study were registered at clinicaltrials.gov as NCT00005131 (Atherosclerosis Risk in Communities), NCT00005133 (Cardiovascular Health Study), NCT00005136 (Family Heart Study), NCT000051<em>21</em> (Framingham Heart Study), NCT00083369 (Genetic and Environmental Determinants of Triglycerides), NCT01331512 (InCHIANTI Study), and NCT00005487 (Multi-Ethnic Study of Atherosclerosis).

BACKGROUND

An unmet medical need exists for patients with metastatic renal cell carcinoma who have progressed on VEGF-targeted and mTOR-inhibitor therapies. Fibroblast growth factor (FGF) pathway activation has been proposed as a mechanism of escape from VEGF-targeted therapies. Dovitinib is an oral tyrosine-kinase inhibitor that inhibits VEGF and FGF receptors. We therefore compared dovitinib with sorafenib as third-line targeted therapies in patients with metastatic renal cell carcinoma.

METHODS

In this multicentre phase 3 study, patients with clear cell metastatic renal cell carcinoma who received one previous VEGF-targeted therapy and one previous mTOR inhibitor were randomly assigned through an interactive voice and web response system to receive open-label dovitinib (500 mg orally according to a 5-days-on and 2-days-off schedule) or sorafenib (400 mg orally twice daily) in a 1:1 ratio. Randomisation was stratified by risk group and region. The primary endpoint was progression-free survival (PFS) assessed by masked central review. Efficacy was assessed in all patients who were randomly assigned and safety was assessed in patients who received at least one dose of study drug. This study is registered with ClinicalTrials.gov, number NCT01223027.

RESULTS

284 patients were randomly assigned to the dovitinib group and 286 to the sorafenib group. Median follow-up was 11·3 months (IQR 7·9-14·6). Median PFS was 3·7 months (95% CI 3·5-3·9) in the dovitinib group and 3·6 months (3·5-3·7) in the sorafenib group (hazard ratio 0·86, 95% CI 0·72-1·04; one-sided p=0·063). 280 patients in the dovitinib group and 284 in the sorafenib group received at least one dose of study drug. Common grade 3 or 4 adverse events included hypertriglyceridaemia (38 [14%]), fatigue (28 [10%]), hypertension (22 [8%]), and diarrhoea (20 [7%]) in the dovitinib group, and hypertension (47 [17%]), fatigue (24 [8%]), dyspnoea (21 [7%]), and palmar-plantar erythrodysaesthesia (18 [6%]) in the sorafenib group. The most common serious adverse event was dyspnoea (16 [6%] and 15 [5%] in the dovitinib and sorafenib groups, respectively).

CONCLUSIONS

Dovitinib showed activity, but this was no better than that of sorafenib in patients with renal cell carcinoma who had progressed on previous VEGF-targeted therapies and mTOR inhibitors. This trial provides reference outcome data for future studies of targeted inhibitors in the third-line setting.

BACKGROUND

Novartis Pharmaceuticals Corporation.

Overexpression and hyperactivation of the type I insulin-like <em>growth</em> <em>factor</em> receptor (IGF-IR) has been observed in human breast tumor biopsies. In addition, in vitro studies indicate that overexpression of IGF-IR is sufficient to transform cells such as mouse embryo <em>fibroblasts</em> and this receptor promotes proliferation and survival in breast cancer cell lines. To fully understand the function of the IGF-IR in tumor initiation and progression, transgenic mice containing human IGF-IR under a doxycycline-inducible MMTV promoter system were generated. Administration of 2 mg/ml doxycycline in the animals' water supply beginning at <em>21</em> days of age resulted in elevated levels of IGF-IR in mammary epithelial cells as detected by Western blotting and immunohistochemistry. Whole mount analysis of 55-day-old mouse mammary glands revealed that IGF-IR overexpression significantly impaired ductal elongation. Moreover, histological analyses revealed multiple hyperplasic lesions in the mammary glands of these 55-day-old mice. The formation of palpable mammary tumors was evident at approximately 2 months of age and was associated with increased levels of IGF-IR signaling molecules including phosphorylated Akt, Erk1/Erk2 and STAT3. Therefore, these transgenic mice provide evidence that IGF-IR overexpression is sufficient to induce mammary epithelial hyperplasia and tumor formation in vivo and provide a model to further understand the function of IGF-IR in mammary epithelial transformation.

<em>Fibroblast</em> <em>growth</em> <em>factors</em> (FGFs) 15/19 and <em>21</em> belong to a subfamily of FGFs that function as hormones. Produced in response to specific nutritional cues, they act on overlapping sets of cell surface receptors composed of classic FGF receptors in complex with βKlotho, and regulate metabolism and related processes during periods of fluctuating energy availability. Pharmacologically, both FGF15/19 and FGF<em>21</em> cause weight loss and improve both insulin-sensitivity and lipid parameters in rodent and primate models of metabolic disease. Recently, FGF<em>21</em> was shown to have similar effects in obese patients with type 2 diabetes. We discuss here emerging concepts in FGF15/19 and FGF<em>21</em> tissue-specific actions and critically assess their putative role as candidate targets for treating metabolic disease.

Brown adipose tissue (BAT) is the main site of adaptive thermogenesis and experimental studies have associated BAT activity with protection against obesity and metabolic diseases, such as type 2 diabetes mellitus and dyslipidaemia. Active BAT is present in adult humans and its activity is impaired in patients with obesity. The ability of BAT to protect against chronic metabolic disease has traditionally been attributed to its capacity to utilize glucose and lipids for thermogenesis. However, BAT might also have a secretory role, which could contribute to the systemic consequences of BAT activity. Several BAT-derived molecules that act in a paracrine or autocrine manner have been identified. Most of these <em>factors</em> promote hypertrophy and hyperplasia of BAT, vascularization, innervation and blood flow, processes that are all associated with BAT recruitment when thermogenic activity is enhanced. Additionally, BAT can release regulatory molecules that act on other tissues and organs. This secretory capacity of BAT is thought to be involved in the beneficial effects of BAT transplantation in rodents. <em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em>, IL-6 and neuregulin 4 are among the first BAT-derived endocrine <em>factors</em> to be identified. In this Review, we discuss the current understanding of the regulatory molecules (the so-called brown adipokines or batokines) that are released by BAT that influence systemic metabolism and convey the beneficial metabolic effects of BAT activation. The identification of such adipokines might also direct drug discovery approaches for managing obesity and its associated chronic metabolic diseases.

The binding of biologically active, 125I-labeled basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) to baby hamster kidney-derived cell line cells (BHK-<em>21</em>) was studied at 4 degrees C. Unlabeled FGF displaced cell surface bound 125I-FGF, but platelet-derived <em>growth</em> <em>factor</em>, epidermal <em>growth</em> <em>factor</em>, insulin, or transferrin did not. Binding was saturable both as a function of time and as a function of increasing 125I-FGF concentrations. Scatchard analysis of the binding data revealed the presence of about 1.2 X 10(5) binding sites/cell with an apparent KD of 270 pM. The number of the binding sites was down-regulated following preincubation of the cells with FGF. The density of binding sites/cell also decreased as an inverse function of cell density. When 125I-FGF binding was studied in a BHK-<em>21</em> cell membrane preparation, it was found that the membranal binding site displayed a lower KD of <em>21</em> pM. 125I-FGF was covalently cross-linked to its cell surface receptor on intact BHK-<em>21</em> cells using the homobifunctional agent disuccinimidyl suberate. Two macromolecular species with an apparent molecular weight of 145,000 and 125,000, respectively, were labeled under both reducing and nonreducing conditions. Unlabeled FGF competed with 125I-FGF for binding to both macromolecular species. The labeling of the macromolecules was also inhibited by heparin. No labeling was observed in the absence of the cross-linkers or when heat-inactivated 125I-FGF was used instead of radiolabeled, biologically active FGF.

Nutrient deprivation or starvation frequently correlates with amino acid limitation. Amino acid starvation initiates a signal transduction cascade starting with the activation of the kinase GCN2 (general control non-derepressible 2) phosphorylation of eIF2 (eukaryotic initiation <em>factor</em> 2), global protein synthesis reduction and increased ATF4 (activating transcription <em>factor</em> 4). ATF4 modulates a wide spectrum of genes involved in the adaptation to dietary stress. The hormone FGF<em>21</em> (<em>fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em>) is induced during fasting in liver and its expression induces a metabolic state that mimics long-term fasting. Thus FGF<em>21</em> is critical for the induction of hepatic fat oxidation, ketogenesis and gluconeogenesis, metabolic processes which are essential for the adaptive metabolic response to starvation. In the present study, we have shown that FGF<em>21</em> is induced by amino acid deprivation in both mouse liver and cultured HepG2 cells. We have identified the human FGF<em>21</em> gene as a target gene for ATF4 and we have localized two conserved ATF4-binding sequences in the 5' regulatory region of the human FGF<em>21</em> gene, which are responsible for the ATF4-dependent transcriptional activation of this gene. These results add FGF<em>21</em> gene induction to the transcriptional programme initiated by increased levels of ATF4 and offer a new mechanism for the induction of the FGF<em>21</em> gene expression under nutrient deprivation.

We isolated and expanded BMSCs from human alveolar/jaw bone at a high success rate (70%). These cells had potent osteogenic potential in vitro and in vivo, although their chondrogenic and adipogenic potential was less than that of iliac cells.

BACKGROUND

Human bone marrow stromal cells (BMSCs) have osteogenic, chondrogenic, and adipogenic potential, but marrow aspiration from iliac crest is an invasive procedure. Alveolar BMSCs may be more useful for regenerative medicine, because the marrow can be aspirated from alveolar bone with minimal pain.

METHODS

In this study, alveolar bone marrow samples were obtained from 41 patients, 6-66 years of age, during the course of oral surgery. BMSCs were seeded and maintained in culture with 10% FBS and basic fibroblast growth factor. In addition, BMSCs were induced to differentiate into osteoblasts, chondrocytes, or adipocytes in appropriate medium.

CONCLUSIONS

From a small volume (0.1-3 ml) of aspirates, alveolar BMSCs expanded at a success ratio of 29/41 (70%). The success rate decreased with increasing donor age, perhaps because of age-dependent decreases in the number and proliferative capacity of BMSCs. The expanded BMSCs differentiated into osteoblasts under osteogenic conditions in 21-28 days: the mRNA levels of osteocalcin, osteopontin, and bone sialoprotein, along with the calcium level, in alveolar BMSC cultures were similar to those in iliac cultures. However, unlike iliac BMSC, alveolar BMSC showed poor chondrogenic or adipogenic potential, and similar differences were observed between canine alveolar and iliac BMSCs. Subsequently, human alveolar BMSCs attached to beta-tricalcium phosphate were transplanted into immunodeficient mice. In transplants, new bone formed with osteoblasts and osteocytes that expressed human vimentin, human osteocalcin, and human GAPDH. These findings suggest that BMSCs have distinctive features depending on their in vivo location and that alveolar BMSCs will be useful in cell therapy for bone diseases.

BACKGROUND

Recent studies suggest human neck brown adipose tissue (BAT) to consist of 'brown adipocyte (BA)-like' or beige adipocytes. However, little is known about their thermogenic function. Within the beige adipocyte transcriptome, <em>fibroblast</em> <em>growth</em> <em>factor</em>-<em>21</em> (FGF<em>21</em>) is a gene whose protein product acts as an adipokine, regulating cold-induced thermogenesis in animals. Here, we explored (i) the adipogenic potential, thermogenic function and FGF<em>21</em> secretory capacity of beige adipocytes derived from human neck fat and (ii) the role of FGF<em>21</em> in modulating adipose bioenergetics.

METHODS

Progenitors isolated from human cervical fat were differentiated into adipocytes with either a BA-like or white adipocyte (WA) phenotype. FGF<em>21</em> secretion was measured by enzyme-linked immuosorbent assay. Real-time PCR/western blotting was used to determine cellular mRNA/protein levels. Extracellular flux bioanalyzer was used to quantify adipocyte oxygen consumption and fatty acid oxidation. Adipocyte heat production was measured by infrared thermography.

RESULTS

Under hormonal manipulation, primary human neck pre-adipocytes differentiated into adipocytes with either BA-like or WA phenotypes, on gene/protein and functional levels. BA-like cells expressed beige but not classic BA markers. During BA differentiation, FGF<em>21</em> gene expression and secretion were increased, and were augmented following norepinephrine exposure (a cold mimic in vitro). Differentiated WA expressed β-klotho, a critical co-<em>factor</em> mediating FGF<em>21</em> action. Treatment of WA with FGF<em>21</em>-induced UCP1 expression and increased oxygen consumption, respiratory uncoupling, norepinephrine-mediated thermogenesis, fatty acid oxidation and heat production, thus recapitulating the association between cold-induced FGF<em>21</em> secretion and cold-induced thermogenesis in vivo.

CONCLUSIONS

Beige adipocytes are thermogenic in humans. FGF<em>21</em> is a beige adipokine capable of promoting a brown fat-like thermogenic program in WAs.

CONCLUSIONS

This study provides first evidence of inducible functional thermogenic beige adipogenesis in human neck fat. FGF<em>21</em> holds promise as a cold-induced beige adipokine with metabolic benefits of therapeutic relevance through browning of white adipose tissue.

OBJECTIVE

To improve the success of culturing olfactory neurons from human nasal mucosa by investigating the intranasal distribution of the olfactory epithelium and devising new techniques for growing human olfactory epithelium in vitro.

METHODS

Ninety-seven biopsy specimens were obtained from 33 individuals, aged 21 to 74 years, collected from 6 regions of the nasal cavity. Each biopsy specimen was bisected, and 1 piece was processed for immunohistochemistry or electron microscopy while the other piece was dissected further for explant culture. Four culture techniques were performed, including whole explants and explanted biopsy slices. Five days after plating, neuronal differentiation was induced by means of a medium that contained basic fibroblast growth factor. After another 5 days, cultures were processed for immunocytochemical analysis.

RESULTS

The probability of finding olfactory epithelium in a biopsy specimen ranged from 30% to 76%, depending on its location. The dorsoposterior regions of the nasal septum and the superior turbinate provided the highest probability, but, surprisingly, olfactory epithelium was also found anteriorly and ventrally on both septum and turbinates. A new method of culturing the olfactory epithelium was devised. This slice culture technique improved the success rate for generating olfactory neurons from 10% to 90%.

CONCLUSIONS

This study explains and overcomes most of the variability in the success in observing neurogenesis in cultures of adult human olfactory epithelium. The techniques presented here make the human olfactory epithelium a useful model for clinical research into certain olfactory dysfunctions and a model for the causes of neurodevelopmental and neurodegenerative diseases.

Preventing reproduction during nutritional deprivation is an adaptive process that is conserved and essential for the survival of species. In mammals, the mechanisms that inhibit fertility during starvation are complex and incompletely understood. Here we show that exposure of female mice to <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>), a fasting-induced hepatokine, mimics infertility secondary to starvation. Mechanistically, FGF<em>21</em> acts on the suprachiasmatic nucleus (SCN) in the hypothalamus to suppress the vasopressin-kisspeptin signaling cascade, thereby inhibiting the proestrus surge in luteinizing hormone. Mice lacking the FGF<em>21</em> co-receptor, β-Klotho, in the SCN are refractory to the inhibitory effect of FGF<em>21</em> on female fertility. Thus, FGF<em>21</em> defines an important liver-neuroendocrine axis that modulates female reproduction in response to nutritional challenge.

OBJECTIVE

The hepatocyte-derived hormone <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>) is a hormone-like regulator of metabolism. The nicotinamide adenine dinucleotide-dependent deacetylase SIRT1 regulates fatty acid metabolism through multiple nutrient sensors. Hepatic overexpression of SIRT1 reduces steatosis and glucose intolerance in obese mice. We investigated mechanisms by which SIRT1 controls hepatic steatosis in mice.

METHODS

Liver-specific SIRT1 knockout (SIRT1 LKO) mice and their wild-type littermates (controls) were divided into groups that were placed on a normal chow diet, fasted for 24 hours, or fasted for 24 hours and then fed for 6 hours. Liver tissues were collected and analyzed by histologic examination, gene expression profiling, and real-time polymerase chain reaction assays. Human HepG2 cells were incubated with pharmacologic activators of SIRT1 (resveratrol or SRT1720) and mitochondrion oxidation consumption rate and immunoblot analyses were performed. FGF<em>21</em> was overexpressed in SIRT1 LKO mice using an adenoviral vector. Energy expenditure was assessed by indirect calorimetry.

RESULTS

Prolonged fasting induced lipid deposition in livers of control mice, but severe hepatic steatosis in SIRT1 LKO mice. Gene expression analysis showed that fasting up-regulated FGF<em>21</em> in livers of control mice but not in SIRT1 LKO mice. Decreased hepatic and circulating levels of FGF<em>21</em> in fasted SIRT1 LKO mice were associated with reduced hepatic expression of genes involved in fatty acid oxidation and ketogenesis, and increased expression of genes that control lipogenesis, compared with fasted control mice. Resveratrol or SRT1720 each increased the transcriptional activity of the FGF<em>21</em> promoter (-2070/+117) and levels of FGF<em>21</em> messenger RNA and protein in HepG2 cells. Surprisingly, SIRT1 LKO mice developed late-onset obesity with impaired whole-body energy expenditure. Hepatic overexpression of FGF<em>21</em> in SIRT1 LKO mice increased the expression of genes that regulate fatty acid oxidation, decreased fasting-induced steatosis, reduced obesity, increased energy expenditure, and promoted browning of white adipose tissue.

CONCLUSIONS

SIRT1-mediated activation of FGF<em>21</em> prevents liver steatosis caused by fasting. This hepatocyte-derived endocrine signaling appears to regulate expression of genes that control a brown fat-like program in white adipose tissue, energy expenditure, and adiposity. Strategies to activate SIRT1 or FGF<em>21</em> could be used to treat fatty liver disease and obesity.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>) is a distinctive member of the FGF family with potent beneficial effects on lipid, body weight, and glucose metabolism and has attracted considerable interest as a potential therapeutic for treating diabetes and obesity. As an alternative to native FGF<em>21</em>, we have developed a monoclonal antibody, mimAb1, that binds to βKlotho with high affinity and specifically activates signaling from the βKlotho/FGFR1c (FGF receptor 1c) receptor complex. In obese cynomolgus monkeys, injection of mimAb1 led to FGF<em>21</em>-like metabolic effects, including decreases in body weight, plasma insulin, triglycerides, and glucose during tolerance testing. Mice with adipose-selective FGFR1 knockout were refractory to FGF<em>21</em>-induced improvements in glucose metabolism and body weight. These results in obese monkeys (with mimAb1) and in FGFR1 knockout mice (with FGF<em>21</em>) demonstrated the essential role of FGFR1c in FGF<em>21</em> function and suggest fat as a critical target tissue for the cytokine and antibody. Because mimAb1 depends on βKlotho to activate FGFR1c, it is not expected to induce side effects caused by activating FGFR1c alone. The unexpected finding of an antibody that can activate FGF<em>21</em>-like signaling through cell surface receptors provided preclinical validation for an innovative therapeutic approach to diabetes and obesity.

Atrial fibrosis is important for the pathogenesis of atrial fibrillation (AF) but the underlying signal transduction is incompletely understood. We therefore studied the role of microRNA-<em>21</em> (miR-<em>21</em>) and its downstream target Sprouty 1 (Spry1) during atrial fibrillation. Left atria (LA) from patients with AF showed a 2.5-fold increased expression of miR-<em>21</em> compared to matched LA of patients in sinus rhythm. Increased miR-<em>21</em> expression correlated positively with atrial collagen content and was associated with a reduced protein expression of Spry1 and increased expression of connective tissue <em>growth</em> <em>factor</em> (CTGF), lysyl oxidase and Rac1-GTPase. Neonatal cardiac <em>fibroblasts</em> treated with angiotensin II (AngII) or CTGF showed an increased miR-<em>21</em> and decreased Spry1 expression. Pretreatment with an inhibitor of Rac1 GTPase, NSC23766, reduced the AngII-induced upregulation of miR-<em>21</em>. A small molecule inhibitor of lysyl oxidase, BAPN, prevented the AngII as well as the CTGF-induced miR-<em>21</em> expression. Transgenic mice with cardiac overexpression of Rac1, which develop spontaneous AF and atrial fibrosis with increasing age, showed upregulation of miR-<em>21</em> expression associated with reduced Spry1 expression. miR-<em>21</em> expression and signalling in vivo were prevented by long-term treatment of the mice with statins. Direct inhibition of miR-<em>21</em> by antagomir-<em>21</em> prevented fibrosis of the atrial myocardium post-myocardial infarction. Left atria of patients with atrial fibrillation are characterized by upregulation of miR-<em>21</em> und reduced expression of Spry1. Activation of Rac1 by angiotensin II leads to a CTGF- and lysyl oxidase-mediated increase of miR-<em>21</em> expression contributing to structural remodelling of the atrial myocardium.

Previous studies suggest that Fgf8 has a key role in regulating vertebrate development. In the rostral head of the embryonic chicken, there are increasing numbers of separate Fgf8 domains; these are present in tissues that appear to have previously expressed Otx2. As Fgf8 expression becomes established, Otx2 expression weakens, but remains in cells abutting the Fgf8 expression domain. These Fgf8 expression domains are closely associated with tissues expressing Bmp4 and Shh. Based on analogy with the embryonic limb, we suggest that Fgf8, Bmp4 and Shh function together in patterning regions of the embryonic head. Gene expression changes are particularly prominent in 14-<em>21</em> somite stage embryos in the rostral forebrain, during early morphogenesis of the telencephalic and optic vesicles, when several new interfaces of Fgf8, Bmp4 and Shh are generated. To gain insights into the functions of <em>fibroblast</em> <em>growth</em> <em>factor</em> 8 (FGF8) in the embryonic forebrain, we studied the effects of implanting beads containing this protein in the dorsal prosencephalon of embryonic day 2 chicken embryos. Ectopic FGF8 had profound effects on morphogenesis of the telencephalic and optic vesicles. It disrupted formation of the optic stalk and caused a transformation of the pigment epithelium into neural retina. Within the telencephalon, FGF8 beads frequently induced a sulcus that had features of an ectopic rostral midline. The sulcus separated the telencephalon into rostral and caudal vesicles. Furthermore, we present evidence that FGF8 can regulate regionalization of the prosencephalon through inhibition of Otx2 and Emx2 expression. Thus, these experiments provide evidence that FGF8 can regulate both morphogenesis and patterning of the rostral prosencephalon (telencephalic and optic vesicles). FGF8 beads can induce midline properties (e.g. a sulcus) and can modulate the specification and differentiation of adjacent tissues. We suggest that some of these effects are through regulating the expression of homeobox genes (Otx2 and Emx2) that are known to participate in forebrain patterning.

Nonalcoholic fatty liver disease (NAFLD) is the most prevalent liver disease worldwide, and there is no approved pharmacotherapy. The efficacy of vitamin E and pioglitazone has been established in nonalcoholic steatohepatitis (NASH), a progressive form of NAFLD. GLP-1RA and SGLT2 inhibitors, which are currently approved for use in diabetes, have shown early efficacy in NASH, and also have beneficial cardiovascular or renal effects. Innovative NASH therapies include four main pathways. The first approach is targeting hepatic fat accumulation. Medications in this approach include modulation of peroxisome proliferator-activator receptors (e.g., pemafibrate, elafibranor), medications targeting farnesoid X receptor axis [obeticholic acid; OCA)], inhibitors of de novo lipogenesis (aramchol, ACC inhibitor), and <em>fibroblast</em> <em>growth</em> <em>factor</em>-<em>21</em> analogues. A second target is oxidative stress, inflammation, and apoptosis. This class of drug includes apoptosis signaling kinase 1 (ASK1) inhibitor and emricasan (an irreversible caspase inhibitor). A third target is intestinal microbiomes and metabolic endotoxemia. Several agents are in ongoing trials, including IMMe124, TLR4 antagonist, and solithromycin (macrolide antibiotics). The final target is hepatic fibrosis, which is strongly associated with all-cause or liver-related mortality in NASH. Antifibrotic agents are a cysteine-cysteine motif chemokine receptor-2/5 antagonist (cenicriviroc; CVC) and galectin 3 antagonist. Among a variety of medications in development, four agents such as OCA, elafibranor, ASK1 inhibitor, and CVC are currently being evaluated in an international phase 3 trial for the treatment of NASH. Within the next few years, the availability of therapeutic options for NASH will hopefully curb the rising trend of NASH-related diseases.

Despite quantitative differences, the activity of basic and acidic <em>fibroblast</em> <em>growth</em> <em>factors</em> (FGF) on a wide variety of normal diploid cells derived from neuroectoderm and mesoderm is intrinsically similar. This suggests that they bind to the same cell surface receptors. This was investigated using a baby hamster kidney cell line (BHK-<em>21</em>) as a model. BHK-<em>21</em> cell membrane components that interact with basic and acidic FGF have been identified by covalent cross-linking to their respective 125I-labeled ligands. Under appropriate conditions, basic and acidic 125I-FGF were cross-linked, using disuccinimidyl suberate, to two receptor species with apparent molecular masses of 145,000 and 125,000 daltons, respectively. The labeling of those receptors is inhibited when either native basic or acidic FGF are present in excess during incubation of cells with either acidic or basic 125I-FGF. Competition of basic 125I-FGF with increasing concentrations of native acidic FGF results in a preferential decrease in the labeling of the 125,000-dalton species, whereas competition of acidic 125I-FGF with increasing concentrations of native basic FGF leads to a preferential decrease in the labeling of the 145,000-dalton species. The data suggest that qualitatively both mitogens interact with the same 145,000- and 125,000-dalton receptor species. The different affinities displayed by acidic and basic FGF toward their common receptor molecules could explain why acidic FGF, depending on the cell type considered, is 20-100-fold less potent than basic FGF.

BACKGROUND

Autologous bone marrow-derived or cardiac-derived stem cell therapy for heart disease has demonstrated safety and efficacy in clinical trials, but functional improvements have been limited. Finding the optimal stem cell type best suited for cardiac regeneration is the key toward improving clinical outcomes.

OBJECTIVE

To determine the mechanism by which novel bone-derived stem cells support the injured heart.

RESULTS

Cortical bone-derived stem cells (CBSCs) and cardiac-derived stem cells were isolated from enhanced green fluorescent protein (EGFP+) transgenic mice and were shown to express c-kit and Sca-1 as well as 8 paracrine <em>factors</em> involved in cardioprotection, angiogenesis, and stem cell function. Wild-type C57BL/6 mice underwent sham operation (n=<em>21</em>) or myocardial infarction with injection of CBSCs (n=67), cardiac-derived stem cells (n=36), or saline (n=60). Cardiac function was monitored using echocardiography. Only 2/8 paracrine <em>factors</em> were detected in EGFP+ CBSCs in vivo (basic <em>fibroblast</em> <em>growth</em> <em>factor</em> and vascular endothelial <em>growth</em> <em>factor</em>), and this expression was associated with increased neovascularization of the infarct border zone. CBSC therapy improved survival, cardiac function, regional strain, attenuated remodeling, and decreased infarct size relative to cardiac-derived stem cells- or saline-treated myocardial infarction controls. By 6 weeks, EGFP+ cardiomyocytes, vascular smooth muscle, and endothelial cells could be identified in CBSC-treated, but not in cardiac-derived stem cells-treated, animals. EGFP+ CBSC-derived isolated myocytes were smaller and more frequently mononucleated, but were functionally indistinguishable from EGFP- myocytes.

CONCLUSIONS

CBSCs improve survival, cardiac function, and attenuate remodeling through the following 2 mechanisms: (1) secretion of proangiogenic factors that stimulate endogenous neovascularization, and (2) differentiation into functional adult myocytes and vascular cells.

Hepatic gluconeogenesis is a main source of blood glucose during prolonged fasting and is orchestrated by endocrine and neural pathways. Here we show that the hepatocyte-secreted hormone <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>) induces fasting gluconeogenesis via the brain-liver axis. Prolonged fasting induces activation of the transcription <em>factor</em> peroxisome proliferator-activated receptor α (PPARα) in the liver and subsequent hepatic production of FGF<em>21</em>, which enters into the brain to activate the hypothalamic-pituitary-adrenal (HPA) axis for release of corticosterone, thereby stimulating hepatic gluconeogenesis. Fasted FGF<em>21</em> knockout (KO) mice exhibit severe hypoglycemia and defective hepatic gluconeogenesis due to impaired activation of the HPA axis and blunted release of corticosterone, a phenotype similar to that observed in PPARα KO mice. By contrast, intracerebroventricular injection of FGF<em>21</em> reverses fasting hypoglycemia and impairment in hepatic gluconeogenesis by restoring corticosterone production in both FGF<em>21</em> KO and PPARα KO mice, whereas all these central effects of FGF<em>21</em> were abrogated by blockage of hypothalamic FGF receptor-1. FGF<em>21</em> acts directly on the hypothalamic neurons to activate the mitogen-activated protein kinase extracellular signal-related kinase 1/2 (ERK1/2), thereby stimulating the expression of corticotropin-releasing hormone by activation of the transcription <em>factor</em> cAMP response element binding protein. Therefore, FGF<em>21</em> maintains glucose homeostasis during prolonged fasting by fine tuning the interorgan cross talk between liver and brain.

OBJECTIVE

The oncogenic drivers of triple-negative (TN) and basal-like breast cancers are largely unknown. Substantial evidence now links aberrant signaling by the fibroblast growth factor receptors (FGFR) to the development of multiple cancer types. Here, we examined the role of FGFR signaling in TN breast cancer.

METHODS

We examined the sensitivity of a panel of 31 breast cancer cell lines to the selective FGFR inhibitor PD173074 and investigated the potential mechanisms underlying sensitivity.

RESULTS

TN breast cancer cell lines were more sensitive to PD173074 than comparator cell lines (P = 0.011), with 47% (7/15) of TN cell lines showing significantly reduced growth. The majority of TN cell lines showed only modest sensitivity to FGFR inhibition in two-dimensional growth but were highly sensitive in anchorage-independent conditions. PD173074 inhibited downstream mitogen-activated protein kinase and PI3K-AKT signaling and induced cell-cycle arrest and apoptosis. Basal-like breast cancer cell lines were found to express FGF2 ligand (11/21 positive) and, similarly, 62% of basal-like breast cancers expressed FGF2, as assessed by immunohistochemistry compared with 5% of nonbasal breast cancers (P < 0.0001). RNA interference targeting of FGF2 in basal-like cell lines significantly reduced growth in vitro and reduced down stream signaling, suggesting an autocrine FGF2 signaling loop. Treatment with PD173074 significantly reduced the growth of CAL51 basal-like breast cancer cell line xenografts in vivo.

CONCLUSIONS

Basal-like breast cancer cell lines, and breast cancers, express autocrine FGF2 and show sensitivity to FGFR inhibitors, identifying a potential novel therapeutic approach for these cancers.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>21</em> (FGF<em>21</em>) plays an important role in the regulation of energy homeostasis during starvation and has an excellent therapeutic potential for the treatment of type 2 diabetes in rodents and monkeys. Acute exercise affects glucose and lipid metabolism by increasing glucose uptake and lipolysis. However, it is not known whether acute exercise affects FGF<em>21</em> expression. Here, we showed that serum FGF<em>21</em> level is increased in mice after a single bout of acute exercise, and that this is accompanied by increased serum levels of free fatty acid, glycerol and ketone body. FGF<em>21</em> gene expression was induced in the liver but not in skeletal muscle and white adipose tissue of mice after acute exercise, and further, the gene expression levels of hepatic peroxisome proliferator-activated receptor α (PPARα) and activating transcription <em>factor</em> 4 (ATF4) were also increased. In addition, we observed increased FGF<em>21</em> level in serum of healthy male volunteers performing a treadmill run at 50 or 80% VO2max. These results suggest that FGF<em>21</em> may also be associated with exercise-induced lipolysis in addition to increased catecholamines and reduced insulin.