Purification and Biochemical Characterization of a Novel Fibrinolytic

M ET AB O LI S M CL IN I CA L A N D E XP E RI ME N TAL 6 4 ( 2 01 5 ) 3 8 0–39 0

Available online at www.sciencedirect.com

Metabolism www.metabolismjournal.com

The role of fibroblast growth factor 21 in the pathogenesis of non-alcoholic fatty liver disease and implications for therapy☆ Jia Liu, Yuan Xu, Yanjin Hu, Guang Wang⁎ Department of Endocrinology, Beijing Chao-yang Hospital, Capital Medical University, No. 8, Gongti South Road, Chaoyang District, Beijing 100020, China

A R T I C LE I N FO

AB S T R A C T

Article history:

Non-alcoholic fatty liver disease (NAFLD) includes a cluster of liver disorders ranging from

Received 30 June 2014

simple fatty liver to non-alcoholic steatohepatitis (NASH) and cirrhosis. Due to its liver and

Accepted 25 November 2014

vascular complications, NAFLD has become a public health problem with high morbidity and

Keywords:

lipotoxicity, oxidative stress, endoplasmic reticulum stress, a chronic inflammatory state

Non-alcoholic fatty liver disease

and mitochondrial dysfunction. Fibroblast growth factor 21 (FGF21) is a member of the

Fibroblast growth factor 21

fibroblast growth factor family with multiple metabolic functions. FGF21 directly

Insulin resistance

regulates lipid metabolism and reduces hepatic lipid accumulation in an insulin-

Lipid metabolism

independent manner. Several studies have shown that FGF21 can ameliorate the

mortality. The pathogenesis of NAFLD is considered a “multi-hit hypothesis” that involves

“multi-hits” in the pathogenesis of NAFLD. The administration of FGF21 reverses hepatic steatosis, counteracts obesity and alleviates insulin resistance in rodents and nonhuman primates. Using several strategies, we show that the reversal of simple fatty liver and NASH is mediated by activation of the FGF21 signaling pathway. In this review, we describe the molecular mechanisms involved in the onset and/or progression of NAFLD, and review the current literature to highlight the therapeutic procedures associated with the FGF21 signaling pathway for simple fatty liver and NASH, which are the two most important types of NAFLD. © 2015 Elsevier Inc. All rights reserved.

Abbreviations: NAFLD, non-alcoholic fatty liver disease; NASH, non-alcoholic steatohepatitis; FGF21, fibroblast growth factor 21; SREBP-1c, sterol regulatory element-binding protein-1c; ACC, acetyl-CoA carboxylase; FAS, fatty acid synthase; SCD1, stearoyl-CoA desaturase 1; ChREBP, carbohydrate response element binding protein; PKC, protein kinase C; ERK, extracellular regulated protein kinases; ROS, reactive oxygen species; IL-6, interleukin-6; TNF-α, tumor necrosis factor-α; CRP, C-reactive protein; JNK, c-jun N-terminal kinase; IκB, inhibitor of nuclear factor-κB; NF-κB, nuclear factor-κB; CHOP, CCAAT enhancer binding protein homologous protein; FGFR, fibroblast growth factor receptor; PPARα, peroxisome proliferator activated receptor α; AMPK, AMP-activated protein kinase; SIRT1, sirtuin 1; PPARγ, peroxisome proliferators-activated receptor γ; UCP-1, uncoupling protein 1; UCP-2, uncoupling protein 2; PI3K, phosphatidylinosistol 3-kinase; ATF4, activating transcription factor 4; OLETF, Otsuka Long Evans Tokushima Fatty; GLP-1, glucagon-like peptide-1; MCAD, medium-chain acyl-CoA dehydrogenase. ☆ Disclosure statement: The authors have nothing to disclose. ⁎ Corresponding author at: Department of Endocrinology; Beijing Chao-yang Hospital, Capital Medical University, No. 8, Gongti South Road, Chaoyang District, Beijing 100020, China. Tel./fax: +86 10 85231710. E-mail address: [email protected] (G. Wang). http://dx.doi.org/10.1016/j.metabol.2014.11.009 0026-0495/© 2015 Elsevier Inc. All rights reserved.

M E TAB O LI S M CL IN I CA L A N D EX PE RI ME N TA L 6 4 ( 2 01 5 ) 3 8 0–3 90

1.

Introduction

Non-alcoholic fatty liver disease (NAFLD) includes a cluster of liver disorders ranging from simple fatty liver to nonalcoholic steatohepatitis (NASH) to cirrhosis [1]. A sedentary lifestyle along with excessive energy intake contributes to obesity, insulin resistance and NAFLD [1]. NAFLD is a public health problem with high morbidity and mortality due to its liver and vascular complications [1]. The current therapies are limited to reducing weight and improving insulin sensitivity with drugs or lifestyle interventions, such as dietary changes and physical activity, which have uncertain therapeutic effects [1]. Fibroblast growth factor 21 (FGF21) is a member of the fibroblast growth factor family with multiple metabolic functions [2]. The administration of FGF21 reverses hepatic steatosis, counteracts obesity, and alleviates insulin resistance and dyslipidemia in both rodents and nonhuman primates. These findings support its development as a novel therapy for the treatment of NAFLD and other metabolic disorders [3–9]. In this review article, we describe the molecular mechanisms involved in the onset and/or progression of NAFLD. Furthermore, we review the current literature to highlight the therapeutic procedures associated with the FGF21 signaling pathway for simple fatty liver and NASH, which are the two most important types of NAFLD.

2.

The Pathogenesis of NAFLD

The pathogenesis of NAFLD has been modified to the “multihit hypothesis” from the “two-hit hypothesis” [10]. The “firsthit” is known as hepatic lipid accumulation/steatosis, which is the initial histological characteristic of NAFLD [11]. Insulin resistance causes an imbalance of fatty acid metabolism of hepatocytes and further contributes to hepatic steatosis [10]. Under physiological conditions, insulin suppresses the lipolysis of white adipose tissue and hepatic gluconeogenesis and also promotes hepatic lipogenesis by stimulating the transcription of sterol regulatory element-binding protein-1c (SREBP-1c). SREBP-1c is a master regulator of lipogenesis that regulates the transcription of acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1) [12,13]. However, in some insulin-resistant states, including obesity, type 2 diabetes and metabolic syndrome, lipolysis and hepatic gluconeogenesis are not inhibited by insulin. However, insulin retains the ability to effectively promote hepatic lipogenesis [14]. Elevated lipolysis of white adipose tissue increases plasma free fatty acids and enhances gluconeogenesis. These changes result in hyperglycemia and further increase de novo lipogenesis by stimulating carbohydrate response element binding protein (ChREBP) [14]. In NAFLD patients, approximately 60% of hepatic lipid accumulation is derived from the re-esterification of plasma free fatty acids. 26% occurs by de novo lipogenesis and 14% is derived from dietary fatty acids [15]. Additional lipids are deposited in the liver and simple fatty liver histologically manifests as steatosis with more than 5% hepatic lipid accumulation [11].

381

The exposure and overload of fatty acid harm hepatocytes by intracellular accumulation of lipid intermediates, such as diglycerides and ceramides, which is defined as lipotoxicity [16]. The lipids and intermediates induce endoplasmic reticulum stress and mitochondrial dysfunction directly or via activation of Toll-like receptors 2 and 4 [16]. Ceramides activate protein kinase C (PKC), inhibit Akt and increase protein phosphatase 2A levels. The increase in protein phosphatase 2A levels contributes to mitochondrial dysfunction and endoplasmic reticulum stress [17]. In addition, the accumulation of lipids and intermediates activates p38 mitogen-activated protein kinase, extracellular regulated protein kinases (ERK) and c-Jun. The activation of these pathways further exacerbates insulin resistance [18]. Elevated fatty acid β-oxidation increases reactive oxygen species (ROS) and activates oxidative stress [16]. The hepatic lipid accumulation and intracellular stresses activate the transcription and release of pro-inflammatory factors, such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and C-reactive protein (CRP) [19,20]. A sedentary lifestyle in conjunction excessive energy intake promotes obesity and dysfunction of white adipose tissue. White adipose tissue secretes more TNF-α and IL-6 and reduces the secretion of adiponectin [20]. The elevated circulating levels of pro-inflammatory cytokines and reduced antiinflammatory factors cause a chronic low-grade inflammatory state that is recognized as an important pathogenic mechanism of NAFLD [20,21]. These “hits” further deteriorate insulin resistance and initiate inflammatory signaling pathways and apoptosis cascades [10]. TNF-α activates c-Jun N-terminal kinase (JNK), PKC and inhibitor of nuclear factor-κB (IκB), which inhibits insulin signal and triggers inflammatory pathways [20]. TNF-α also activates de novo ceramide synthesis by stimulating sphingomyelinase [22]. IL-6 stimulates Janus kinase activation and induces the phosphorylation of signal transducters and activators of transcription to increase the transcription of the suppressor of cytokine signaling and promote insulin resistance [23]. An increase of ROS levels and oxidative stress triggers the ROS–PKC– nuclear factor-κB (NF-κB) pathway and further activates inflammatory signaling pathways that stimulate the infiltration of inflammatory cells [10,20]. Endoplasmic reticulum stress also stimulates cell apoptosis through both CCAAT enhancer binding protein homologous protein (CHOP)induced and JNK-mediated mitochondria-dependent apoptosis pathways [24,25]. Hepatocytes with excess lipid accumulation are susceptible to subsequent “multi-hits”, including oxidative stress, endoplasmic reticulum stress, a chronic inflammatory state and mitochondrial dysfunction. These changes lead to the infiltration of inflammatory cells, activation of inflammatory signaling pathways and hepatocytes apoptosis. Additionally, these changes promote the progression from simple fatty liver to NASH [10] (Fig. 1). Previous studies have shown more than 25% of NASH patients will develop cirrhosis within 10 years, and a few patients will develop end-stage liver disease and hepatocellular carcinoma [26]. Thus, NASH is a turning point from benign to irreversible lesions including cirrhosis and hepatocellular carcinoma [26].

382


Fig. 1 – The role of FGF21 in the pathogenesis of NAFLD. FGF21 directly regulates lipid metabolism and reduces hepatic lipid accumulation in an insulin-independent manner. Moreover, FGF21 also has the ability to ameliorate the “multi-hits” of the pathogenesis of NAFLD, including oxidative stress, endoplasmic reticulum stress, chronic inflammatory state and mitochondrial dysfunction, which were mainly mediated by stimulating the FGF21 signaling pathway in liver, skeletal muscle and adipose tissue.

3.

FGF21 and NAFLD

FGF21 is predominantly released from hepatocytes and to a lesser extent from other tissues, including white adipose tissue, skeletal muscle and pancreatic β cells [2]. FGF21 binds to β-klotho and fibroblast growth factor receptor (FGFR) and induces the dimerization and autophosphorylation of FGFR. Activated FGFR then initiates its biological functions [2]. Recent studies have shown that FGF21 is a metabolic regulator of energy homeostasis, glucose-lipid metabolism and insulin sensitivity [2]. FGF21 reverses hepatic steatosis and prevents diet-induced obesity in both rodents and nonhuman primates [3–9]. There are several underlying mechanisms responsible for the above-mentioned beneficial effect of FGF21 (Fig. 1). FGF21 directly regulates lipid metabolism and reduces hepatic lipid accumulation in an insulin-independent manner [12,27,28]. Additionally, FGF21 inhibits the lipolysis of white adipose tissue and further decreases circulating free fatty acids levels [29]. The over-expression of FGF21 reverses the up-regulated expression of SREBP-1c and FAS, the two key enzymes for lipid synthesis, in fatty acids-treated human liver-derived HepG2 cells [12]. In addition to suppressing lipid synthesis by lowering the hepatic influx of fatty acids and reducing the expression of lipid synthesis genes, FGF21 also promotes fatty acid β-oxidation mediated by the peroxisome proliferator activated receptor α (PPARα)-FGF21 transcriptional regulatory axis [28,30,31]. Moreover, FGF21 activates AMPactivated protein kinase (AMPK) and sirtuin 1 (SIRT1). The activation of these proteins up-regulates gene expression

required for mitochondrial biogenesis and fatty acids βoxidation [32]. A recent study showed that FGF21 also had the ability to ameliorate endoplasmic reticulum stress-mediated apoptosis in ApoE-/- mice [33]. FGF21 phosphorylates Akt and ameliorates insulin resistance in peripheral tissues [5]. The elevated insulin sensitivity decreased ChREBP-mediated de novo lipogenesis by promoting glucose utilization in the liver and skeletal muscle and inhibiting hepatic gluconeogenesis [14,17]. Moreover, the reduced insulin resistance also decreased the accumulation of lipid and its intermediates and lowered PKC activation in liver and skeletal muscle. Collectively, the result is the inhibition of inflammatory signaling pathways and apoptosis [24]. White adipose tissue highly expresses FGFR1 and β-klotho, and is considered to be the predominant site of FGF21 action [34,35]. FGF21 stimulated glucose uptake by adipocytes is mediated through the activation of ERK1/ERK2 and the induction of glucose transporter-1 expression in an insulinindependent manner [27]. FGF21 inhibits lipolysis stimulated by growth hormone and catecholamine in adipocytes [29]. In white adipocytes, FGF21 also enhances mitochondrial oxidative capacity and potentiates peroxisome proliferator activated receptor γ (PPARγ) activity [36]. FGF21 is expressed in human brown adipose tissue and has a significant positive correlation with uncoupling protein 1 (UCP-1) expression [37]. FGF21 signaling elicits thermogenic activation and stimulates browning of white adipose tissue by inducing the expression of UCP-1 and ACC2, and then leading to an increase of energy expenditure [38]. FGF21 has been identified as a key modulator for adiponectin secretion in white adipose tissue [3]. FGF21 also counteracts the negative effects of TNF-α on adiponectin secretion [39]. Adiponectin alleviates steatosis, suppresses the NF-κB signaling pathway and improves insulin sensitivity by promoting fatty acid β-oxidation, reducing de novo lipogenesis and inhibiting the expression of pro-inflammatory cytokines [40]. Moreover, adiponectin also stimulates deacetylation of ceramides and decreases lipotoxicity [39]. Adiponectin has been reported to ameliorate hepatic mitochondrial dysfunction by stimulating uncoupling protein 2 (UCP-2) expression, which is a mitochondrial inner membrane transporter [41]. FGF21 administration significantly decreases the lipid content and improves the insulin sensitivity of skeletal muscle [4]. In human skeletal muscle myotubes, FGF21 prevents palmitate-induced insulin resistance by inhibiting the activation of stress-related kinases including JNK, NF-κB and PKC [42]. FGF21 has been proven as a myokine expressed and released by skeletal muscle cells, and its expression is regulated by phosphatidylinosistol 3-kinase (PI3K)/Akt signaling pathway [43,44]. Exercise activates the Akt1 signaling pathway in muscle, which ameliorates metabolic disorders partially by promoting the production and secretion of FGF21 [44,45]. Consistent with this hypothesis, the manifestation of muscle Akt1 activation is similar to FGF21 administration in mice [44,46]. Hojman et al. demonstrated that there was no significant correlation between muscle FGF21 mRNA and plasma FGF21 levels during chronic hyperinsulinemia, which might suggest that muscle tissue is not the main source of circulating FGF21 and its release from muscle cells may have a paracrine role in the muscle tissue [47].


4.

FGF21 Resistance

There are several stimulators that regulate the expression and release of FGF21. Acute, hour-long stimulation of overnutrition manifests as an inhibitory effect on FGF21 release [48]. However, a 3-day overfeeding significantly raises circulating FGF21 in healthy humans [49]. An increased fatty acid influx has been shown to induce hepatic FGF21 production via activating PPARα [50], and glucose activates FGF21 mRNA expression in a ChREBP-dependent way in rat hepatocytes [12]. Both oxidative stress and endoplasmic reticulum stress are related to increased expression of FGF21 [51,52]. A recent study demonstrated that FGF21 is the target gene of activating transcription factor 4 (ATF4) and CHOP [53]. Endoplasmic reticulum stress induced by triglycerides can directly regulate FGF21 expression through an ATF and CHOP dependent mechanism [53]. Moreover, intraperitoneal injection of the endoplasmic reticulum stressor – tunicamycin – can stimulate hepatic FGF21 expression and increase circulating FGF21 levels [52]. In vivo, insulin increases FGF21 levels in patients with insulin resistance during a euglycemic clamp test [50]. The circulating FGF21 levels are positively correlated with plasma insulin levels in patients with impaired glucose tolerance and type 2 diabetes [54]. Moreover, mitochondrial disorders in patients with NAFLD/NASH are related to elevated FGF21 expression [41].

383

Circulating FGF21 levels are significantly elevated in patients with simple fatty liver and NASH. The plasma levels and hepatic mRNA expression of FGF21 increase with the degree of steatosis (Table 1) [55–64]. There were similar results found in animal studies [65,66]. The induction of ERK1/2 phosphorylation by FGF21 is impaired in diet-induced obese mice [67]. Furthermore, the ability of FGF21 to promote adiponectin production is decreased significantly in mice with obesity and type 2 diabetes. The ability of FGF21 to decrease plasma levels of fatty acids, triglycerides and glucose is also restricted in diet-induced obese mice [6,68], and more notably, a higher dose of intravenous FGF21 still reverses hepatic steatosis, improves insulin sensitivity and decreases fasting glucose in NAFLD animal models [6,7,9]. These findings might suggest an FGF21-resistant state in animal models and patients with NAFLD/NASH. The increased FGF21 levels may be a protective response against disorders of glucose-lipid metabolism. In animal models of NAFLD, the expressions of FGFR1 and β-klotho in white adipose tissue are markedly lower. Furthermore, the expressions of FGFR2, FGFR substrates 2 and β-klotho in liver are also reduced [67,69]. The activation of inflammatory signaling pathways represses β-klotho expression by the JNK1 pathway in adipose tissue [68]. Additionally, in vitro studies have shown high glucose directly inhibits the expression of β-klotho [70]. Thus, the mechanisms responsible for FGF21 resistance may involve the inhibition of

Table 1 – Main characteristics and findings of clinical studies assessing FGF21 in human NAFLD. Year

Country (ethnicity)

Cross-sectional studies 2013 China (Asian)

Study design (diagnostic method)

Major findings

Ref.

253 subjects (hepatic ultrasonography)

NAFLD patients showed significantly higher serum FGF21 than those without NAFLD. NASH patients have higher serum FGF21 than those without NAFLD. NASH patients have higher serum FGF21 than controls. Serum FGF21 levels were positively correlated with hepatic fat content especially in patients with mild/moderate hepatic steatosis. Fasting plasma FGF21 levels were significantly higher in patients with NASH. Serum FGF21 in newly diagnosed type 2 diabetic patients with NAFLD group was significantly increased. NAFLD patients have significantly higher serum FGF21 levels than healthy controls; serum FGF21 levels were an independent predictor of hepatic steatosis scores in NAFLD patients. Serum FGF21 levels of NAFLD patients were significantly higher than control subjects; in human liver tissues, FGF21 mRNA expression increased with the degree of steatosis. Serum FGF21 of NASH patients was increased.

[55]

Baseline FGF21 level was an independent predictor of NAFLD.

[64]

2012

Turkey (European)

2012

China Hong Kong (Asian)

2011

China (Asian)

2011

USA (Caucasian)

2011

China (Asian)

2010

Turkey (European)

2010

China (Asian)

348 subjects NAFLD (n = 224)/control subjects (n = 124) (hepatic ultrasonography); 17 subjects with different degrees of steatosis (biopsy)

2010

Spain (Caucasian)

30 subjects overweight or obese nondiabetic women (n = 10)/healthy, lean subjects (n = 20) (biopsy)

Longitudinal studies 2012 China (Asian)

136 subjects NASH (n = 60)/healthy controls (n = 76) (biopsy) 220 subjects NAFLD (n = 146)/healthy controls (n = 74) (biopsy) 138 subjects with ultrasonography diagnosed fatty liver (hepatic fat contents were measured by proton magnetic resonance spectroscopy) 74 subjects NASH (n = 36)/healthy controls (n = 38) (biopsy) 99 subjects patients with newly diagnosed type 2 diabetes (n = 69)/healthy controls (n = 30) (hepatic ultrasonography) 159 subjects NAFLD (n = 82)/healthy controls (n = 77) (biopsy)

808 subjects (follow-up 3 years) (ultrasonography)

[56] [57] [58]

[59] [60]

[61]

[62]

[63]

384


FGF21 downstream signaling pathways induced by multiple pathologic conditions.

5.

FGF21 and the Therapy of NAFLD

As a novel metabolic regulator, FGF21 has beneficial effects on simple fatty liver and NASH, which are the two most important types of NAFLD [6,7,9]. Several strategies have shown the ability to reverse simple fatty liver and NASH is potentially mediated by activation of the FGF21 signaling pathway (Tables 2 and 3) (Fig. 2). Lifestyle modifications, including a calorie-restricted diet and regular physical exercise, are widely believed as the firstline treatment of simple fatty liver and NASH [1]. Previous population studies have shown that both exercise and caloric restriction significantly decrease liver enzyme levels and reverse hepatic steatosis [81]. Uebanso et al. demonstrated that a hypocaloric high-protein diet promotes hepatic lipolysis and lipid utilization and further leads to a significant improvement of simple fatty liver through a FGF21dependent pathway [78]. Physical activity for two weeks has been shown to significantly increase circulating FGF21 levels in sedentary young healthy women [82]. However, both exercise and caloric restriction prevent obesity and NAFLD development of Otsuka Long Evans Tokushima Fatty (OLETF) rats by reducing circulating FGF21 levels and hepatic FGF21 mRNA expression [69]. The interesting finding is the up-

regulation of hepatic FGF21 downstream effectors including FGFR2, FGF21 receptor substrate 2 and β-klotho in this model [69]. These data might indicate chronic exercise and caloric restriction alleviate FGF21 resistance. Therefore, it is likely that the acute effect of lifestyle modification is due to the stimulation of FGF21 expression, whereas chronic actions are attributable to improving hepatic FGF21 resistance. Another study showed that moderate weight loss by exercise and caloric restriction did not cause the changes of FGF21 levels in humans [73]. This controversy might be related to different duration of treatment, excise intensity and the degree of body weight loss. In addition to the improvement of lipid profiles, PPARα agonists significantly reverse hepatic steatosis, necroinflammation and collagen deposition in animal models with simple fatty liver and NASH [83,84]. The FGF21 gene has a PPARα response element and is considered as an important downstream target factor of PPARα [30]. Increasing evidence suggests that fenofibrate, by activating PPARα, shows the therapeutic benefits on NAFLD by increasing fatty acid βoxidation, improving lipid profiles, decreasing hepatic insulin resistance and inhibiting the expression of inflammatory mediators [83]. However, the above-mentioned effects of fenofibrate are impaired in the FGF21 knockout mice [30]. Therefore, these data suggest the effects of PPARα agonists are mediated by activating the FGF21 signaling pathway. Metformin treatment alleviates hepatic inflammation, steatosis and fibrosis in simple fatty liver and NASH patients

Table 2 – Clinical trials assessing the effect of pharmacologic and non-pharmacologic interventions on FGF21 levels. Year Country (ethnicity) Study design diagnostic method) 2012

Germany (Caucasian)

2011

China (Asian)

2011

USA (Caucasian)

2011 2009

Germany (Caucasian) China (Asian)

2009

Czech (European)

2008

Sweden (Caucasian)

Obese children (n = 60)/ normal-weight children (n = 40)

Interventions (duration)

Major findings

Exercise, behavior, and Compared with the normal-weight nutrition therapy (12 months) children, obese children had significantly increased FGF21 levels; a decrease of BMI was associated with a significant reduction of FGF21 levels. Patients with newly diagnosed 30 patients with newly Plasma FGF21 levels were higher in the type 2 diabetes (n = 68)/controls diagnosed type 2 diabetes newly diagnosed type 2 diabetes (n = 52) with FBG ≥14.0 mmol/L were mellitus group than controls; fasting treated with CSII (2 weeks) plasma FGF21 levels were significantly decreased after CSII treatment. Type 2 diabetic patients (n = 24) Pioglitazone (45 mg/d) or Pioglitazone treatment reduced on diet and/or metformin pioglitazone (45 mg/d) + hepatic fat, but plasma FGF21 levels (magnetic resonance spectroscopy) exenatide (10 μg twice daily) did not change; combined treatment (12 months) with pioglitazone and exenatide decreased FGF21 levels and hepatic fat. Obese subjects (n = 30) A weight reduction program Moderate weight loss did not induce (6 months) changes of FGF21 levels in humans. Patients with newly diagnosed type Type 2 diabetic patients with Plasma FGF21 levels were higher in 2 diabetes (n = 30)/type 2 diabetic poor glycemic control were type 2 diabetic patients than in the patients with poor glycemic control added rosiglitazone (4 mg/day) controls; rosiglitazone treatment after the treatment with metformin (12 weeks) significantly decreased plasma FGF21 (n = 34)/healthy controls (n = 30) levels. Obese patients (n = 26)/type 2 Very low calorie diet Both 3 weeks of VLCD and 3 months diabetic patients (n = 11)/healthy (3 weeks) or fenofibrate of fenofibrate treatment significantly controls (n = 32) (200 mg/d) (3 months) increased FGF21 levels. Normal-weight, nondiabetic Placebo or fenofibrate Fenofibrate treatment significantly patients with primary (200 mg/d) (3 weeks) increased FGF21 levels. hypertriglyceridemia (n = 19)

Ref. [71]

[72]

[66]

[73] [74]

[75]

[76]


385

Table 3 – Experimental studies investigating implicating FGF21 in the treatment of NAFLD. Year

Study design

FGF21-related agents 2014 Liver-specific SIRT1 knockout (SIRT1 LKO) mice

2013

Diet-induced obese mice were infused with FGF21 (1 mg/kg/d) or vehicle (1 week)

2013

Adiponectin knockout mice were injected intraperitoneally with either rmFGF21 (2 mg/kg/d) or vehicle

2012

Db/db mice were treated with PEGylation of human FGF21 (0.25 mg/kg/d) (12 days)

2009

Diet-induced obese mice were randomly assigned to intraperitoneally injection with FGF21 twice daily (0.1, 1, and 10 mg/kg/d) or vehicle (6 weeks)

2008

Diet-induced obese mice and ob/ob mice were treated with FGF21(0.1, 0.3, and 1 mg/kg/d) or vehicle (2 weeks) Male diabetic rhesus monkeys were administered FGF21 (1.0 mg/kg/d) (6 weeks)

2007

Strategies potentially affecting FGF21 signaling pathway 2014 C57BL/6J mice fed high-fat diet were treated with exendin-4 intraperitoneal injection (1 nmol/kg/d) (10 weeks) 2012 High-fat diet (HFD) fed ApoE(-/-) mice with adiponectin (Acrp30) knockdown were administrated liraglutide(1 mg/kg/d)(8 weeks)

2011

Diet-induced obese mice were treated with exendin-4 (4 weeks)

2012

OLETF rats were grouped into ad libitum fed, sedentary (OLETF-SED), voluntary wheel running exercise (OLETF-EX), or caloric restriction (CR) (OLETF-CR; 70% of SED) (36 weeks)

2009

High-sucrose induced obese rats were assigned randomly to one of two energy restricted diets: a carbohydrate-based control diet (CD) or a highprotein diet (HPD) (4 weeks) Diet-induced fatty liver mice and db/db mice were given molecular hydrogen-water (12 weeks)

2011

2007

Wild-type mice, PPARα deficient mice and ob/ob mice were treated with the PPARα agonist Wy-14,643

2013

C57/B6J mice were randomly divided into three groups: high-fat diet, artemisia santolinaefolia (SANT) and artemisia scoparia (SCO) (4 weeks)

Effect and potential mechanism

Ref.

Hepatic overexpression of FGF21 increased the gene expressions of fatty acid β-oxidation, decreased fasting-induced steatosis, reduced obesity and promoted browning of white adipose tissue. In diet-induced obese mice, FGF21 infusion improved insulin responsiveness, decreased hepatocellular and myocellular diacylglycerol content and reduced protein kinase C activation in liver and skeletal. The effects of FGF21 on ameliorating insulin resistance of liver and skeletal muscle were abrogated in adiponectin knockout mice, whereas FGF21-mediated activation of ERK1/ERK2 in adipose tissues remained unaffected. PEGylation of human FGF21 reduced blood glucose, lipid profiles and hepatic lipid content, and enhanced islet number and glucose-dependent insulin secretion. FGF21 significantly reversed hepatic steatosis, reduced body weight, blood glucose, insulin and lipid levels, and improved hepatic and peripheral insulin sensitivity; the reduction of hepatic triglyceride levels was associated with FGF21-dependent inhibition of nuclear SREBP-1 expression. In diet-induced obese and ob/ob mice, administration of FGF21 lowered body weight, increased energy expenditure, reduced hepatic steatosis and ameliorated hyperglycemia. FGF21 administration significantly reduced plasma levels of glucose, triglyceride and insulin, improved lipid profiles and caused a small but significant weight loss.

[77]

Exendin-4 could improve fatty liver by increasing SIRT1-mediated FGF21 expression.

[28]

In HFD fed ApoE(-/-) mice with adiponectin knockdown, FGF21 mRNA expressions in both liver and adipose tissues were significantly increased, while FGFR1 and β-klotho mRNA levels in adipose tissue, as well as FGFR1-3 and β-klotho mRNA levels in liver were significantly decreased; liraglutide improved insulin resistance and increased FGF21 expression in liver and FGFR3 and β-klotho in adipose tissue, as well as FGFR1-3, β-klotho levels in liver. Exendin-4 significantly reduced hepatic triglyceride content and FGF21 expression, and enhanced phosphorylation of hepatic AMPK and ACC, but no significant difference in weight and body fat was observed. Both daily EX and CR prevented obesity and NAFLD development, accompanied by significantly lower serum FGF21 and hepatic FGF21 expression; Hepatic FGFR2, hepatic FGF21 receptor substrate 2 and β-klotho were elevated in the OLETF-EX and OLETF-CR animals compared with the OLETF-SED animals. HPD diet caused greater improvement in fatty liver and hypertriglyceridemia than the CD diet; Expression of FGF21 was significantly increased in obese rats fed the HPD.

[65]

Drinking molecular hydrogen-water significantly enhanced the expression of FGF21, reduced hepatic oxidative stress, and alleviated fatty liver in db/db mice as well as diet-induced fatty liver mice. Fasting or treatment of mice with the PPARα agonist induced FGF21 mRNA expression; in PPARα deficient mice, FGF21 expression was lower than wild-type mice, and fasting or treatment with PPARα agonist did not induce FGF21. Fewer lipid droplets were observed in the SCO group; SCO significantly increased hepatic insulin sensitivity and AMPK activity, decreased the expressions of lipid synthesis genes. Neither SANT nor SCO significantly altered plasma FGF21 levels.

[79]

[4]

[3]

[5]

[6]

[7]

[8]

[66]

[69]

[78]

[30]

[80]

386


Fig. 2 – The potential pharmacologic and non-pharmacologic agents affecting FGF21 in NAFLD. Some strategies reverse simple fatty liver and NASH potentially mediated by stimulating the expression and release of FGF21 or up-regulating the expression of FGF21 downstream effectors.

[85]. These beneficial effects are caused by its ability to decrease hepatic gluconeogenesis, stimulate glucose uptake by skeletal muscle and increase fatty acid β-oxidation of adipose tissue and liver by activating AMPK [85]. Metformin stimulates the FGF21 expression of rat and human hepatocytes in vitro in a dose-dependent manner, and this can be blocked by the AMPK inhibitor Compound C [86]. Therefore, FGF21 may be involved in the AMPK signaling pathway and further contributes to the therapeutic effects of metformin on NAFLD. However, in human clinical trials, metformin is not beneficial for NAFLD patients [87]. This controversial result can be explained because these studies were shortterm and have inconsistent outcomes due to the different duration and dose. Further larger randomized controlled trials with sufficient duration and histological endpoints are urgently needed to assess the effectiveness of metformin on NAFLD. PPARγ agonist, another insulin sensitizer, has two representative medicine—pioglitazone and rosiglitazone. The PPARγ agonists ameliorate insulin resistance, reverse hepatic steatosis and inhibit liver inflammation and ballooning necrosis [36]. In vitro, PPARγ agonists induce FGF21 expression of murine and human adipocytes [88]. Animal studies have shown that FGF21 is an autocrine factor that regulates the PPARγ activity of adipose tissues [36]. In FGF21-/- mice, rosiglitazone fails to exert its beneficial effects [36]. This might suggest that FGF21 is required for the therapeutic effects of PPARγ agonists in NAFLD patients. Moreover, treatment with rosiglitazone alleviates metabolic disorders by reducing circulating FGF21 levels in type 2 diabetic patients [74]. Several in vitro studies have shown that rosiglitazone reverses the inhibition on β-klotho caused by high glucose [70]. It suggests PPARγ agonists alleviate FGF21 resistance,

which might contribute to the beneficial effects of PPARγ agonists in reversing the progression of NAFLD. The glucagon-like peptide-1 (GLP-1) receptor agonist is a novel agent approved for treating type 2 diabetes [89]. Its therapeutic effects on simple fatty liver and NASH have been demonstrated by several human and animal studies [90–92]. Dipeptidyl peptidase IV inhibitors, which enhance endogenous GLP-1 levels by inhibiting degradation of GLP-1, ameliorate hepatocyte ballooning and reduce liver enzyme levels in type 2 diabetic patients with NASH [93]. Liraglutide, a longacting GLP-1 receptor agonist, significantly increases FGF21 expression in animal models of NAFLD [65]. In high fat dietinduced obese mice, exendin-4 (GLP-1 analog) injections increased hepatic FGF21 expression [28]. Exendin-4 promotes fatty acid β-oxidation by increasing expression of PPARα and medium-chain acyl-CoA dehydrogenase (MCAD). However, the inhibition of FGF21 by siRNA attenuated the effect on PPARα and MCAD of exendin-4 [28]. Therefore, the beneficial effect of exendin-4 treatment may be due to the activation of the FGF21 signaling pathway. Interestingly, GLP-1 treatment for 16 weeks significantly decreases plasma levels and hepatic mRNA expression of FGF21 [94]. A combination of exenatide and pioglitazone treatment is associated with a significant decrease in plasma FGF21 levels and hepatic fat content [94]. Moreover, exendin-4 administration for 4 weeks significantly reduces hepatic triglyceride content and hepatic FGF21 expression in diet-induced obese mice, despite the lack of any significant change in weight or body mass index [94]. In high-fat diet fed ApoE-/- mice with adiponectin knockdown, liraglutide upregulated β-klotho expression in adipose tissue, and FGFR1-3 and β-klotho levels in the liver [65]. The intraperitoneal injection of exendin-4 for 10 weeks increases the expression of FGFR in diet-induced obese mice [28].


Moreover, in diet-induced obese mice, exendin-4 treatment increases hepatic AMPK phosphorylation and circulating adiponectin levels, and reduces hepatic expression and plasma levels of FGF21 [66,94]. The elevated expression of hepatic FGF21 downstream effectors as well as the raised ability of FGF21 to promote AMPK phosphorylation and adiponectin expression suggests the amelioration of FGF21 resistance [66,94]. Thus, the therapeutic effects of GLP-1 receptor agonists on simple fatty liver and NASH may be partly mediated by its direct effect on FGF21 activity. There are several emerging drugs with the ability to treat simple fatty liver and NASH by regulating the FGF21 pathway [77,79,80]. Resveratrol, a natural SIRT1 activator, is extracted from grapes and other plants [95]. Animal studies showed that resveratrol prevents hepatic steatosis and hyperlipidemia in diabetic and diet-induced obese mice [95,96]. In vitro studies showed that SIRT1 promotes fatty acid β-oxidation by stimulating FGF21 expression [77]. The FGF21 signaling pathway may be involved in the therapeutic effects of resveratrol for NAFLD. Molecular hydrogen has been reported to improve oxidative stress, reduce body weight and plasma glucose levels and improve insulin sensitivity [79]. Drinking molecular hydrogen-water significantly alleviates fatty liver in db/db mice and diet-induced fatty liver in wild-type mice [79]. A recent study demonstrated that molecular hydrogen improves metabolism dysfunction by inducing hepatic FGF21 in db/db mice [79]. More and more researchers want to investigate whether FGF21 is a potential candidate for the treatment of NAFLD. In diabetic rhesus monkeys, FGF21 administration for 6 weeks improves insulin sensitivity and decreases fasting plasma glucose [8]. Significant improvements in lipid profiles are observed in FGF21-treated diabetic rhesus monkeys [8]. The subcutaneous administration of LY2405319, a novel FGF21 variant, reduced plasma glucose and body weight in ob/ob and diet-induced obese mice [97]. However, a short circulating half-life limits the use of FGF21 in clinical application. The site-specific PEGylation of human FGF21 shows a dramatically prolonged half-life and enhanced efficacy in db/db mice [5]. The twice-weekly dosing of human PEGylated FGF21 reduced the hepatic lipid content and improved plasma lipid profiles, and exerts some beneficial effects on glucose metabolism [5]. There is still an urgent need for more animal studies and further clinical research studies to assess the safety and effectiveness of FGF21-related biological agents.

6.

Summary

FGF21 is a novel metabolic regulator with beneficial effects on the regulation of energy homeostasis, glucose-lipid metabolism and insulin sensitivity. Several strategies can reverse simple fatty liver and NASH by activating FGF21 signaling pathway. Many researchers are interested in investigating whether FGF21 is a potential candidate for the treatment of NAFLD. Although FGF21 administration has some therapeutic effects for NAFLD in animal studies, there is still an urgent need for more animal studies and further clinical research to assess the safety and effectiveness of FGF21-related biological agents.

387

Author Contributions G. W. planned the review. J. L. reviewed papers and wrote the manuscript. Y. X. and Y. J. H. provided advice on planning the review.

Acknowledgments This work was supported by grants from the Major National Basic Research Program of P. R. China (2011CB503904) and the Chinese National Natural Science Foundation (81270369; 81070244) to Guang Wang and the Capital Clinical Research Foundation of Beijing Municipal Commission of Science and Technology (Z131107002213024) to Yuan Xu.

REFERENCES

[1] Chalasani N, Younossi Z, Lavine JE, Diehl AM, Brunt EM, Cusi K, et al. The diagnosis and management of non-alcoholic fatty liver disease: practice guideline by the American Gastroenterological Association, American Association for the Study of Liver Diseases, and American College of Gastroenterology. Gastroenterology 2012;142:1592–609. [2] Li H, Zhang J, Jia W. Fibroblast growth factor 21: a novel metabolic regulator from pharmacology to physiology. Front Med 2013;7:25–30. [3] Lin Z, Tian H, Lam KS, Lin S, Hoo RC, Konishi M, et al. Adiponectin mediates the metabolic effects of FGF21 on glucose homeostasis and insulin sensitivity in mice. Cell Metab 2013;17:779–89. [4] Camporez JP, Jornayvaz FR, Petersen MC, Pesta D, Guigni BA, Serr J, et al. Cellular mechanisms by which FGF21 improves insulin sensitivity in male mice. Endocrinology 2013;154:3099–109. [5] Mu J, Pinkstaff J, Li Z, Skidmore L, Li H, Myler H, et al. FGF21 analogs of sustained action enabled by orthogonal biosynthesis demonstrate enhanced antidiabetic pharmacology in rodents. Diabetes 2012;61:505–12. [6] Xu J, Lloyd DJ, Hale C, Stanislaus S, Chen M, Sivits G, et al. Fibroblast growth factor 21 reverses hepatic steatosis, increases energy expenditure, and improves insulin sensitivity in diet-induced obese mice. Diabetes 2009;58:250–9. [7] Coskun T, Bina HA, Schneider MA, Dunbar JD, Hu CC, Chen Y, et al. Fibroblast growth factor 21 corrects obesity in mice. Endocrinology 2008;149:6018–27. [8] Kharitonenkov A, Wroblewski VJ, Koester A, Chen YF, Clutinger CK, Tigno XT, et al. The metabolic state of diabetic monkeys is regulated by fibroblast growth factor-21. Endocrinology 2007;148:774–81. [9] Kim HW, Lee JE, Cha JJ, Hyun YY, Kim JE, Lee MH, et al. Fibroblast growth factor 21 improves insulin resistance and ameliorates renal injury in db/db mice. Endocrinology 2013; 154:3366–76. [10] Takaki A, Kawai D, Yamamoto K. Multiple hits, including oxidative stress, as pathogenesis and treatment target in non-alcoholic steatohepatitis (NASH). Int J Mol Sci 2013;14: 20704–28. [11] Straub BK, Schirmacher P. Pathology and biopsy assessment of non-alcoholic fatty liver disease. Dig Dis 2010;28:197–202. [12] Zhang Y, Lei T, Huang JF, Wang SB, Zhou LL, Yang ZQ, et al. The link between fibroblast growth factor 21 and sterol regulatory element binding protein 1c during lipogenesis in hepatocytes. Mol Cell Endocrinol 2011;342:41–7.

388


[13] Kohan AB, Qing Y, Cyphert HA, Tso P, Salati LM. Chylomicron remnants and nonesterified fatty acids differ in their ability to inhibit genes involved in lipogenesis in rats. J Nutr 2011; 141:171–6. [14] Kawano Y, Cohen DE. Mechanisms of hepatic triglyceride accumulation in non-alcoholic fatty liver disease. J Gastroenterol 2013;48:434–41. [15] Puri P, Baillie RA, Wiest MM, Mirshahi F, Choudhury J, Cheung O, et al. A lipidomic analysis of nonalcoholic fatty liver disease. Hepatology 2007;46:1081–90. [16] Gentile CL, Pagliassotti MJ. The role of fatty acids in the development and progression of nonalcoholic fatty liver disease. J Nutr Biochem 2008;19:567–76. [17] Gariani K, Philippe J, Jornayvaz FR. Non-alcoholic fatty liver disease and insulin resistance: from bench to bedside. Diabetes Metab 2013;39:16–26. [18] Weickert MO, Pfeiffer AF. Signalling mechanisms linking hepatic glucose and lipid metabolism. Diabetologia 2006;49: 1732–41. [19] Pirgon O, Bilgin H, Cekmez F, Kurku H, Dundar BN. Association between insulin resistance and oxidative stress parameters in obese adolescents with non-alcoholic fatty liver disease. J Clin Res Pediatr Endocrinol 2013;5:33–9. [20] Asrih M, Jornayvaz FR. Inflammation as a potential link between nonalcoholic fatty liver disease and insulin resistance. J Endocrinol 2013;218:R25–36. [21] Tarantino G, Savastano S, Colao A. Hepatic steatosis, low-grade chronic inflammation and hormone/growth factor/adipokine imbalance. World J Gastroenterol 2010;16: 4773–83. [22] Meyer SG, de Groot H. Cycloserine and threodihydrosphingosine inhibit TNF-alpha-induced cytotoxicity: evidence for the importance of de novo ceramide synthesis in TNF-alpha signaling. Biochim Biophys Acta 2003; 1643:1–4. [23] Ueki K, Kondo T, Kahn CR. Suppressor of cytokine signaling 1 (SOCS-1) and SOCS-3 cause insulin resistance through inhibition of tyrosine phosphorylation of insulin receptor substrate proteins by discrete mechanisms. Mol Cell Biol 2004;24:5434–46. [24] Greene MW, Burrington CM, Ruhoff MS, Johnson AK, Chongkrairatanakul T, Kangwanpornsiri A. PKC{delta} is activated in a dietary model of steatohepatitis and regulates endoplasmic reticulum stress and cell death. J Biol Chem 2010;285:42115–29. [25] Xiong Y, Zhang J, Liu M, An M, Lei L, Guo W. Human leptin protein activates the growth of HepG2 cells by inhibiting PERK-mediated ER stress and apoptosis. Mol Med Rep 2014; 10:1649–55. [26] Caldwell S, Argo C. The natural history of non-alcoholic fatty liver disease. Dig Dis 2010;28:162–8. [27] Li K, Li L, Yang M, Liu H, Boden G, Yang G. The effects of fibroblast growth factor-21 knockdown and over-expression on its signaling pathway and glucose-lipid metabolism in vitro. Mol Cell Endocrinol 2012;348:21–6. [28] Lee J, Hong SW, Park SE, Rhee EJ, Park CY, Oh KW, et al. Exendin-4 regulates lipid metabolism and fibroblast growth factor 21 in hepatic steatosis. Metabolism 2014;63:1041–8. [29] Chen W, Hoo RL, Konishi M, Itoh N, Lee PC, Ye HY, et al. Growth hormone induces hepatic production of fibroblast growth factor 21 through a mechanism dependent on lipolysis in adipocytes. J Biol Chem 2011;286:34559–66. [30] Lundasen T, Hunt MC, Nilsson LM, Sanyal S, Angelin B, Alexson SE, et al. PPARalpha is a key regulator of hepatic FGF21. Biochem Biophys Res Commun 2007;360:437–40. [31] Christodoulides C, Dyson P, Sprecher D, Tsintzas K, Karpe F. Circulating fibroblast growth factor 21 is induced by peroxisome proliferator-activated receptor agonists but not ketosis in man. J Clin Endocrinol Metab 2009;94:3594–601.

[32] Chau MD, Gao J, Yang Q, Wu Z, Gromada J. Fibroblast growth factor 21 regulates energy metabolism by activating the AMPK-SIRT1-PGC-1alpha pathway. Proc Natl Acad Sci U S A 2010;107:12553–8. [33] Wu X, Qi YF, Chang JR, Lu WW, Zhang JS, Wang SP, et al. Possible role of fibroblast growth factor 21 on atherosclerosis via amelioration of endoplasmic reticulum stress-mediated apoptosis in apoE mice. Heart Vessels 2014. http://dx.doi.org/ 10.1007/s00380-014-0557-9. [34] Veniant MM, Hale C, Helmering J, Chen MM, Stanislaus S, Busby J, et al. FGF21 promotes metabolic homeostasis via white adipose and leptin in mice. PLoS One 2012;7: e40164. [35] Adams AC, Yang C, Coskun T, Cheng CC, Gimeno RE, Luo Y, et al. The breadth of FGF21's metabolic actions are governed by FGFR1 in adipose tissue. Mol Metab 2012;2:31–7. [36] Dutchak PA, Katafuchi T, Bookout AL, Choi JH, Yu RT, Mangelsdorf DJ, et al. Fibroblast growth factor-21 regulates PPARgamma activity and the antidiabetic actions of thiazolidinediones. Cell 2012;148:556–67. [37] Hondares E, Gallego-Escuredo JM, Flachs P, Frontini A, Cereijo R, Goday A, et al. Fibroblast growth factor-21 is expressed in neonatal and pheochromocytoma-induced adult human brown adipose tissue. Metabolism 2014;63:312–7. [38] Lee P, Brychta RJ, Linderman J, Smith S, Chen KY, Celi FS. Mild cold exposure modulates fibroblast growth factor 21 (FGF21) diurnal rhythm in humans: relationship between FGF21 levels, lipolysis, and cold-induced thermogenesis. J Clin Endocrinol Metab 2013;98:E98-102. [39] Holland WL, Miller RA, Wang ZV, Sun K, Barth BM, Bui HH, et al. Receptor-mediated activation of ceramidase activity initiates the pleiotropic actions of adiponectin. Nat Med 2011; 17:55–63. [40] Kadowaki T, Yamauchi T, Kubota N, Hara K, Ueki K, Tobe K. Adiponectin and adiponectin receptors in insulin resistance, diabetes, and the metabolic syndrome. J Clin Invest 2006;116: 1784–92. [41] Zhou M, Xu A, Tam PK, Lam KS, Chan L, Hoo RL, et al. Mitochondrial dysfunction contributes to the increased vulnerabilities of adiponectin knockout mice to liver injury. Hepatology 2008;48:1087–96. [42] Lee MS, Choi SE, Ha ES, An SY, Kim TH, Han SJ, et al. Fibroblast growth factor-21 protects human skeletal muscle myotubes from palmitate-induced insulin resistance by inhibiting stress kinase and NF-kappaB. Metabolism 2012;61: 1142–51. [43] Keipert S, Ost M, Johann K, Imber F, Jastroch M, van Schothorst EM, et al. Skeletal muscle mitochondrial uncoupling drives endocrine cross-talk through the induction of FGF21 as a myokine. Am J Physiol Endocrinol Metab 2014;306:E469–82. [44] Izumiya Y, Bina HA, Ouchi N, Akasaki Y, Kharitonenkov A, Walsh K. FGF21 is an Akt-regulated myokine. FEBS Lett 2008; 582:3805–10. [45] Nader GA, Esser KA. Intracellular signaling specificity in skeletal muscle in response to different modes of exercise. J Appl Physiol (1985) 2001;90:1936–42. [46] Cheung BM, Deng HB. Fibroblast growth factor 21: a promising therapeutic target in obesity-related diseases. Expert Rev Cardiovasc Ther 2014;12:659–66. [47] Hojman P, Pedersen M, Nielsen AR, Krogh-Madsen R, Yfanti C, Akerstrom T, et al. Fibroblast growth factor-21 is induced in human skeletal muscles by hyperinsulinemia. Diabetes 2009;58:2797–801. [48] Matikainen N, Taskinen MR, Stennabb S, Lundbom N, Hakkarainen A, Vaaralahti K, et al. Decrease in circulating fibroblast growth factor 21 after an oral fat load is related to postprandial triglyceride-rich lipoproteins and liver fat. Eur J Endocrinol 2012;166:487–92.


[49] Heilbronn LK, Campbell LV, Xu A, Samocha-Bonet D. Metabolically protective cytokines adiponectin and fibroblast growth factor-21 are increased by acute overfeeding in healthy humans. PLoS One 2013;8:e78864. [50] Mai K, Andres J, Biedasek K, Weicht J, Bobbert T, Sabath M, et al. Free fatty acids link metabolism and regulation of the insulin-sensitizing fibroblast growth factor-21. Diabetes 2009; 58:1532–8. [51] Liu Y, Wang C, Wang Y, Ma Z, Xiao J, McClain C, et al. Cobalt chloride decreases fibroblast growth factor-21 expression dependent on oxidative stress but not hypoxia-inducible factor in Caco-2 cells. Toxicol Appl Pharmacol 2012;264: 212–21. [52] Schaap FG, Kremer AE, Lamers WH, Jansen PL, Gaemers IC. Fibroblast growth factor 21 is induced by endoplasmic reticulum stress. Biochimie 2013;95:692–9. [53] Wan XS, Lu XH, Xiao YC, Lin Y, Zhu H, Ding T, et al. ATF4- and CHOP-dependent induction of FGF21 through endoplasmic reticulum stress. Biomed Res Int 2014;2014:807874. [54] Chavez AO, Molina-Carrion M, Abdul-Ghani MA, Folli F, Defronzo RA, Tripathy D. Circulating fibroblast growth factor21 is elevated in impaired glucose tolerance and type 2 diabetes and correlates with muscle and hepatic insulin resistance. Diabetes Care 2009;32:1542–6. [55] Shen Y, Ma X, Zhou J, Pan X, Hao Y, Zhou M, et al. Additive relationship between serum fibroblast growth factor 21 level and coronary artery disease. Cardiovasc Diabetol 2013;12:124. [56] Yilmaz Y, Eren F. Identification of a support vector machine-based biomarker panel with high sensitivity and specificity for nonalcoholic steatohepatitis. Clin Chim Acta 2012;414:154–7. [57] Shen J, Chan HL, Wong GL, Choi PC, Chan AW, Chan HY, et al. Non-invasive diagnosis of non-alcoholic steatohepatitis by combined serum biomarkers. J Hepatol 2012;56:1363–70. [58] Yan H, Xia M, Chang X, Xu Q, Bian H, Zeng M, et al. Circulating fibroblast growth factor 21 levels are closely associated with hepatic fat content: a cross-sectional study. PLoS One 2011;6: e24895. [59] Dasarathy S, Yang Y, McCullough AJ, Marczewski S, Bennett C, Kalhan SC. Elevated hepatic fatty acid oxidation, high plasma fibroblast growth factor 21, and fasting bile acids in nonalcoholic steatohepatitis. Eur J Gastroenterol Hepatol 2011; 23:382–8. [60] Li X, Fan X, Ren F, Zhang Y, Shen C, Ren G, et al. Serum FGF21 levels are increased in newly diagnosed type 2 diabetes with nonalcoholic fatty liver disease and associated with hsCRP levels independently. Diabetes Res Clin Pract 2011;93:10–6. [61] Yilmaz Y, Eren F, Yonal O, Kurt R, Aktas B, Celikel CA, et al. Increased serum FGF21 levels in patients with nonalcoholic fatty liver disease. Eur J Clin Invest 2010;40:887–92. [62] Li H, Fang Q, Gao F, Fan J, Zhou J, Wang X, et al. Fibroblast growth factor 21 levels are increased in nonalcoholic fatty liver disease patients and are correlated with hepatic triglyceride. J Hepatol 2010;53:934–40. [63] Dushay J, Chui PC, Gopalakrishnan GS, Varela-Rey M, Crawley M, Fisher FM, et al. Increased fibroblast growth factor 21 in obesity and nonalcoholic fatty liver disease. Gastroenterology 2010;139:456–63. [64] Li H, Dong K, Fang Q, Hou X, Zhou M, Bao Y, et al. High serum level of fibroblast growth factor 21 is an independent predictor of non-alcoholic fatty liver disease: a 3-year prospective study in China. J Hepatol 2013;58:557–63. [65] Yang M, Zhang L, Wang C, Liu H, Boden G, Yang G, et al. Liraglutide increases FGF-21 activity and insulin sensitivity in high fat diet and adiponectin knockdown induced insulin resistance. PLoS One 2012;7:e48392. [66] Samson SL, Sathyanarayana P, Jogi M, Gonzalez EV, Gutierrez A, Krishnamurthy R, et al. Exenatide decreases hepatic fibroblast growth factor 21 resistance in non-alcoholic fatty

[67]

[68]

[69]

[70]

[71]

[72]

[73]

[74]

[75]

[76]

[77]

[78]

[79]

[80]

[81]

[82]

389

liver disease in a mouse model of obesity and in a randomised controlled trial. Diabetologia 2011;54:3093–100. Fisher FM, Chui PC, Antonellis PJ, Bina HA, Kharitonenkov A, Flier JS, et al. Obesity is a fibroblast growth factor 21 (FGF21)resistant state. Diabetes 2010;59:2781–9. Diaz-Delfin J, Hondares E, Iglesias R, Giralt M, Caelles C, Villarroya F. TNF-alpha represses beta-Klotho expression and impairs FGF21 action in adipose cells: involvement of JNK1 in the FGF21 pathway. Endocrinology 2012;153:4238–45. Fletcher JA, Meers GM, Laughlin GM, Ibdah JA, Thyfault JP, Rector RS. Modulating fibroblast growth factor 21 in hyperphagic OLETF rats with daily exercise and caloric restriction. Appl Physiol Nutr Metab 2012;37:1054–62. So WY, Cheng Q, Chen L, Evans-Molina C, Xu A, Lam KS, et al. High glucose represses beta-klotho expression and impairs fibroblast growth factor 21 action in mouse pancreatic islets: involvement of peroxisome proliferator-activated receptor gamma signaling. Diabetes 2013;62:3751–9. Reinehr T, Woelfle J, Wunsch R, Roth CL. Fibroblast growth factor 21 (FGF-21) and its relation to obesity, metabolic syndrome, and nonalcoholic fatty liver in children: a longitudinal analysis. J Clin Endocrinol Metab 2012;97: 2143–50. Yang M, Dong J, Liu H, Li L, Yang G. Effects of short-term continuous subcutaneous insulin infusion on fasting plasma fibroblast growth factor-21 levels in patients with newly diagnosed type 2 diabetes mellitus. PLoS One 2011;6: e26359. Mai K, Schwarz F, Bobbert T, Andres J, Assmann A, Pfeiffer AF, et al. Relation between fibroblast growth factor-21, adiposity, metabolism, and weight reduction. Metabolism 2011;60: 306–11. Li K, Li L, Yang M, Zong H, Liu H, Yang G. Effects of rosiglitazone on fasting plasma fibroblast growth factor-21 levels in patients with type 2 diabetes mellitus. Eur J Endocrinol 2009;161:391–5. Mraz M, Bartlova M, Lacinova Z, Michalsky D, Kasalicky M, Haluzikova D, et al. Serum concentrations and tissue expression of a novel endocrine regulator fibroblast growth factor-21 in patients with type 2 diabetes and obesity. Clin Endocrinol (Oxf) 2009;71:369–75. Galman C, Lundasen T, Kharitonenkov A, Bina HA, Eriksson M, Hafstrom I, et al. The circulating metabolic regulator FGF21 is induced by prolonged fasting and PPARalpha activation in man. Cell Metab 2008;8:169–74. Li Y, Wong K, Giles A, Jiang J, Lee JW, Adams AC, et al. Hepatic SIRT1 attenuates hepatic steatosis and controls energy balance in mice by inducing fibroblast growth factor 21. Gastroenterology 2014;146:539-49 e7. Uebanso T, Taketani Y, Fukaya M, Sato K, Takei Y, Sato T, et al. Hypocaloric high-protein diet improves fatty liver and hypertriglyceridemia in sucrose-fed obese rats via two pathways. Am J Physiol Endocrinol Metab 2009;297:E76–84. Kamimura N, Nishimaki K, Ohsawa I, Ohta S. Molecular hydrogen improves obesity and diabetes by inducing hepatic FGF21 and stimulating energy metabolism in db/db mice. Obesity (Silver Spring) 2011;19:1396–403. Wang ZQ, Zhang XH, Yu Y, Tipton RC, Raskin I, Ribnicky D, et al. Artemisia scoparia extract attenuates non-alcoholic fatty liver disease in diet-induced obesity mice by enhancing hepatic insulin and AMPK signaling independently of FGF21 pathway. Metabolism 2013;62:1239–49. Petersen KF, Dufour S, Befroy D, Lehrke M, Hendler RE, Shulman GI. Reversal of nonalcoholic hepatic steatosis, hepatic insulin resistance, and hyperglycemia by moderate weight reduction in patients with type 2 diabetes. Diabetes 2005;54:603–8. Cuevas-Ramos D, Almeda-Valdes P, Meza-Arana CE, BritoCordova G, Gomez-Perez FJ, Mehta R, et al. Exercise increases

390

[83]

[84]

[85]

[86]

[87]

[88]

[89]

[90]


serum fibroblast growth factor 21 (FGF21) levels. PLoS One 2012;7:e38022. Hong XZ, Li LD, Wu LM. Effects of fenofibrate and xuezhikang on high-fat diet-induced non-alcoholic fatty liver disease. Clin Exp Pharmacol Physiol 2007;34:27–35. Shiri-Sverdlov R, Wouters K, van Gorp PJ, Gijbels MJ, Noel B, Buffat L, et al. Early diet-induced non-alcoholic steatohepatitis in APOE2 knock-in mice and its prevention by fibrates. J Hepatol 2006;44:732–41. Doycheva I, Loomba R. Effect of metformin on ballooning degeneration in nonalcoholic steatohepatitis (NASH): when to use metformin in nonalcoholic fatty liver disease (NAFLD). Adv Ther 2014;31:30–43. Nygaard EB, Vienberg SG, Orskov C, Hansen HS, Andersen B. Metformin stimulates FGF21 expression in primary hepatocytes. Exp Diabetes Res 2012;2012:465282. Mazza A, Fruci B, Garinis GA, Giuliano S, Malaguarnera R, Belfiore A. The role of metformin in the management of NAFLD. Exp Diabetes Res 2012;2012:716404. Zhang X, Yeung DC, Karpisek M, Stejskal D, Zhou ZG, Liu F, et al. Serum FGF21 levels are increased in obesity and are independently associated with the metabolic syndrome in humans. Diabetes 2008;57:1246–53. Giorda CB, Nada E, Tartaglino B. Pharmacokinetics, safety, and efficacy of DPP-4 inhibitors and GLP-1 receptor agonists in patients with type 2 diabetes mellitus and renal or hepatic impairment. A systematic review of the literature. Endocrine 2014. http://dx.doi.org/10.1007/s12020-014-0179-0. Trevaskis JL, Griffin PS, Wittmer C, Neuschwander-Tetri BA, Brunt EM, Dolman CS, et al. Glucagon-like peptide-1 receptor agonism improves metabolic, biochemical, and

[91]

[92]

[93]

[94]

[95]

[96]

[97]

histopathological indices of nonalcoholic steatohepatitis in mice. Am J Physiol Gastrointest Liver Physiol 2012;302: G762–72. Cuthbertson DJ, Irwin A, Gardner CJ, Daousi C, Purewal T, Furlong N, et al. Improved glycaemia correlates with liver fat reduction in obese, type 2 diabetes, patients given glucagonlike peptide-1 (GLP-1) receptor agonists. PLoS One 2012;7: e50117. Klonoff DC, Buse JB, Nielsen LL, Guan X, Bowlus CL, Holcombe JH, et al. Exenatide effects on diabetes, obesity, cardiovascular risk factors and hepatic biomarkers in patients with type 2 diabetes treated for at least 3 years. Curr Med Res Opin 2008; 24:275–86. Yilmaz Y, Yonal O, Deyneli O, Celikel CA, Kalayci C, Duman DG. Effects of sitagliptin in diabetic patients with nonalcoholic steatohepatitis. Acta Gastroenterol Belg 2012;75:240–4. Samson SL, Gonzalez EV, Yechoor V, Bajaj M, Oka K, Chan L. Gene therapy for diabetes: metabolic effects of helperdependent adenoviral exendin 4 expression in a diet-induced obesity mouse model. Mol Ther 2008;16:1805–12. Heeboll S, Thomsen KL, Pedersen SB, Vilstrup H, George J, Gronbaek H. Effects of resveratrol in experimental and clinical non-alcoholic fatty liver disease. World J Hepatol 2014;6:188–98. Cho SJ, Jung UJ, Choi MS. Differential effects of low-dose resveratrol on adiposity and hepatic steatosis in diet-induced obese mice. Br J Nutr 2012;108:2166–75. Kharitonenkov A, Beals JM, Micanovic R, Strifler BA, Rathnachalam R, Wroblewski VJ, et al. Rational design of a fibroblast growth factor 21-based clinical candidate, LY2405319. PLoS One 2013;8:e58575.

Purification and Biochemical Characterization of a Novel Fibrinolytic

Recommend Documents