Fibroblast growth factor 21 attenuates the progression of hyperuricemic nephropathy through inhibiting inflammation, fibrosis and oxidative stress.

Elevated levels of circulating fibroblast growth factor 21 (FGF21) have been reported in patients with hyperuricemia. However, the effect of FGF21 in hyperuricemic nephropathy (HN) remains unexplored. Here, we investigated the effect and mechanism of action of FGF21 on HN. HN model was induced with adenine and potassium oxysalt in wild-type C57BL/6 mice and FGF21-/- mice. For in vitro studies, human renal tubular epithelial (HK-2) cells were exposed to uric acid with/without FGF21 or ß-Klotho-siRNA. Here, we reported aggravated renal dysfunction and structural damage in the FGF21-/- mice compared to the wild-type mice. These were evident in the upsurge of inflammatory factors IL-1ß, TNF-α, IL-6, and IL-18; fibrotic markers Collagen I and α-SMA; and oxidation products ROS and MDA. However, exogenous administration of FGF21 to wild-type HN mice significantly reversed these negative effects. In terms of mechanism, FGF21 significantly inhibited NF-κB/NLRP3 and TGF-ß1/Smad3 pathways and promoted nuclear translocation of Nrf2 both in vivo and in vitro. Furthermore, the silencing of ß-Klotho was marked by the attenuation of the improved effect of FGF21 on cell damage. In conclusion, our studies revealed that exogenous FGF21 treatment significantly improved HN, which was achieved by the inhibition of inflammation, fibrosis, and oxidative stress.

Fibroblast growth factor-21 as a novel metabolic factor for regulating thrombotic homeostasis.

Fibroblast growth factor-21 (FGF-21) performs a wide range of biological functions in organisms. Here, we report for the first time that FGF-21 suppresses thrombus formation with no notable risk of bleeding. Prophylactic and therapeutic administration of FGF-21 significantly improved the degree of vascular stenosis and reduced the thrombus area, volume and burden. We determined the antithrombotic mechanism of FGF-21, demonstrating that FGF-21 exhibits an anticoagulant effect by inhibiting the expression and activity of factor VII (FVII). FGF-21 exerts an antiplatelet effect by inhibiting platelet activation. FGF-21 enhances fibrinolysis by promoting tissue plasminogen activator (tPA) expression and activation, while inhibiting plasminogen activator inhibitor 1 (PAI-1) expression and activation. We further found that FGF-21 mediated the expression and activation of tPA and PAI-1 by regulating the ERK1/2 and TGF-ß/Smad2 pathways, respectively. In addition, we found that FGF-21 inhibits the expression of inflammatory factors in thrombosis by regulating the NF-κB pathway.

FGF21 ameliorates hepatic fibrosis by multiple mechanisms.

BACKGROUND: Previous study reports that fibroblast growth factor 21 (FGF21) could ameliorate hepatic fibrosis, but its mechanisms have not been fully investigated. METHODS AND RESULTS: In this study, three models were used to investigate the mechanism by which FGF21 alleviates liver fibrosis. Hepatic fibrosis animal models were respectively induced by CCL4 and dimethylnitrosamine. Our results demonstrated that liver index and liver function were deteriorated in both models. Hematoxylin and eosin and Masson's staining showed that the damaged tissue architectonics were observed in the mice of both models. Treatment with FGF21 significantly ameliorated these changes. ELISA analysis showed that the serum levels of IL-1ß, IL-6 and TNF-α were significantly elevated in both models. However, administration of FGF21 significantly reduced these inflammatory cytokines. Real-time PCR and Western blot analysis showed that treatment with FGF21 significantly decreased mRNA and protein expressions of collagenI, α-SMA and TGF-ß. Platelet-derived growth factor-BB (PDGF-BB) stimulant was used to establish the experimental cell model in hepatic stellate cells (HSCs). Real-time PCR and Western blot analysis demonstrated that the expression of collagenI and α-SMA were significantly upregulated by this stimulant in model group. Interestingly, our results showed that mRNA and protein expressions of leptin were also significantly induced in PDGF-BB treated HSCs. Administration of FGF21 significantly reduced leptin expression in a dose dependent manner and these effects were reversed in siRNA (against ß-klotho) transfected HSCs. Furthermore, the leptin signaling pathways related protein p-ERK/t-ERK, p-STAT3/STAT3 and TGF-ß were significantly downregulated by FGF21 treatment in a dose dependent manner. The expressions of SOCS3 and Nrf-2 were enhanced by treatment with FGF21. The underlying mechanism may be that FGF21 regulates leptin-STAT3 axis via Nrf-2 and SOCS3 pathway in activated HSCs. CONCLUSIONS: FGF21 ameliorates hepatic fibrosis by multiple mechanisms.

FGF21 alleviates chronic inflammatory injury in the aging process through modulating polarization of macrophages.

Previous studies reported that FGF21 prolongs life span and delays the body senescence, but the mechanism is not clear. The present study was designed to investigate the effects of FGF21 on hepatic senescence in aging mice and further research the mechanism. The 14-month-old male mice were administered with PBS, FGF21 or metformin once daily for 6 months. Results showed that FGF21 alleviated liver injury and inhibited accumulation of senescence markers SASP, P53 and P16 in the livers of aging mice. Subsequently we found that the aging mice treated by FGF21 showed transition of type 1 macrophages (M1) to type 2 macrophages (M2) in the livers. Next, we used THP-1 macrophages triggered by LPS to study effects of FGF21 on macrophages. Macrophages triggered by LPS exhibited features of M1, but the addition of FGF21 decreased the expression of M1 markers, and promoted the macrophages to exhibit features of M2. Results showed that the effects of FGF21 on macrophages were associated with the AMPK pathway. After adding AMPK inhibitor, the effects of FGF21 were inhibited, which was associated with the NF-κB signaling pathway. Finally, co-culturing differentiated macrophages and hepatocytes, we found that the large amount of pro-inflammatory factors such as IL-6 promoted hepatocyte senescence, which exhibited enhanced P53, P16 and ß-galactosidase. This was contrary to hepatocytes co-cultured with macrophages treated by FGF21. These results indicate that FGF21 alleviates hepatic senescence injury by modulating the polarization of macrophages through the AMPK /NF-κB signaling pathway.

Fibroblast growth factor 21: a novel long-acting hypoglycemic drug for canine diabetes.

Currently, insulin is commonly used in the clinical management of canine diabetes. However, it must be injected preprandially causing much inconvenience to the owners. Therefore, the development of long-acting hypoglycemic agents has attracted much attention in the scientific community. This study aimed to investigate the long-acting hypoglycemic effect of canine fibroblast growth factor 21 (cFGF-21) in diabetic dogs. Diabetic dogs were administered with cFGF-21, polyethylene glycol-modified cFGF-21 (PEG-cFGF-21), or insulin once a day, once every 2, 3, or 4 days subcutaneously. The results showed that cFGF-21 and PEG-cFGF-21 maintained blood glucose comparable to normal levels for 2 and 3 days respectively while insulin maintained the blood glucose for only 2 h after a single injection. After treatment with cFGF-21, oral glucose tolerance test (OGTT) was significantly improved with glycosylated hemoglobin (HbA1c) close to the normal levels. In addition, cFGF-21 significantly repaired islet ß cells, increased insulin content, and protected the pancreas from streptozotocin-induced injury. Furthermore, cFGF-21 exhibited both antioxidant and anti-inflammatory properties in the pancreas. We conclude, therefore, that cFGF-21 and PEG-cFGF-21 can maintain blood glucose comparable to normal levels for 2 and 3 days respectively after a single dose. The long-acting efficacy of cFGF-21 can be attributed to improvement in oxidative stress and the reduction of inflammation in the pancreas.

Ultraviolet light activates PMK-1/p38 MAPK signaling via MOM-4 and JKK-1 in Caenorhabditis elegans.

P38 mitogen-activated protein kinase (p38 MAPK) plays an important role in innate immunity and is activated by ultraviolet (UV) radiation. However, the molecular mechanism underlying UV stress remains unclear. In this study, we reported that UV activated PMK-1/p38 MAPK signaling via JKK-1 and MOM-4 in Caenorhabditis elegans. In C. elegans, different UV radiation doses resulted in PMK-1 phosphorylation. However, pmk-1 mutants failed to demonstrate an altered survival time in response to UV when compared with wild-type worms. Further analysis showed that JKK-1, but not SEK-1 mutants, displayed impaired PMK-1 activation following UV irradiation, suggesting that JKK-1 is the upstream MAP2K for the activation of PMK-1 in C. elegans under UV stimulation. UV-induced activation of PMK-1 was markedly reduced in MOM-4, but not in NSY-1 and DLK-1 mutant worms, suggesting that MOM-4 is the upstream MAP3K regulator of PMK-1 activation in response to UV stress in C. elegans. Additionally, daf-16 mutants displayed a shorter lifespan under UV stress, but UV-induced activation of PMK-1 was not markedly reduced in daf-16 and age-1 mutant worms. Our results revealed the signaling pathway involved in PMK-1 activation in C. elegans in response to UV radiation.

Fibroblast Growth Factor-21 ameliorates hepatic encephalopathy by activating the STAT3-SOCS3 pathway to inhibit activated hepatic stellate cells.

Neurological dysfunction, one of the consequences of acute liver failure (ALF), and also referred to as hepatic encephalopathy (HE), contributes to mortality posing challenges for clinical management. FGF21 has been implicated in the inhibition of cognitive decline and fibrogenesis. However, the effects of FGF21 on the clinical and molecular presentations of HE has not been elucidated. HE was induced by fulminant hepatic failure using thioacetamide (TAA) in male C57BL/6J mice while controls were injected with saline. For two consecutive weeks, mice were treated intraperitoneally with FGF21 (3 mg/kg) while controls were treated with saline. Cognitive, neurological, and activity function scores were recorded. Serum, liver, and brain samples were taken for analysis of CCL5 and GABA by ELISA, and RT qPCR was used to measure the expressions of fibrotic and pro-inflammatory markers. We report significant improvement in both cognitive and neurological scores by FGF21 treatment after impairment by TAA. GABA and CCL5, key factors in the progression of HE were also significantly reduced in the treatment group. Furthermore, the expression of fibrotic markers such as TGFß and Col1 were also significantly downregulated after FGF21 treatment. TNFα and IL-6 were significantly reduced in the liver while in the brain, TNFα and IL-1 were downregulated. However, both in the liver and the brain, IL-10 was significantly upregulated. FGF21 inhibits CXCR4/CCL5 activation and upregulates the production of IL-10 in the damaged liver stimulating the production pro-inflammatory cytokines and apoptosis of hepatic stellate cells through the STAT3-SOCS3 pathway terminating the underlying fibrosis in HE.

The composite biological adjuvants enhance immune response of porcine circovirus type2 vaccine.

The porcine circovirus (PCV) is one of the most economically important infection diseases of pigs. Granulocyte-macrophage colony-stimulating factor (GM-CSF) or Flic as an immune adjuvant has been shown to enhance the immunogenicity of vaccines in previous study. However, the composite biological adjuvants stimulate more effective immunological response. In this study, the porcine GM-CSF (pGM-CSF) and FliC protein were expressed by pSUMO in E.coli Rosetta (DE3) and purified by Ni-NTA Sepharose, respectively. The immunogenicity of PCV vaccine with pGM-CSF and FliC was firstly evaluated to identify the immunoenhancement in mice. The results indicated that mice immunized with vaccine + pGM-CSF + FliC enhanced immune responses ability significantly and quickly. Then, the immune response level of PCV vaccine with pGM-CSF and FliC was assessed in piglets. The results indicated that pigs immunized with vaccine + pGM-CSF + FliC showed significantly higher PCV antibody level than those immunized with vaccine and single adjuvant or vaccine alone. Furthermore, pigs in the vaccine + pGM-CSF + FliC group elicited stronger CD4+ and CD8 + T cells proliferative responses than those in all other groups and showed the effectively up-regulated transcriptional level of IL-1, IL-8 and IL-17 stimulating the immune system. We demonstrated that GM-CSF and FliC as composite biological adjuvants of the vaccine showed a stronger immune response and it is promising application for vaccines to against PCV.

Efficacy of a combination of high and low dosage of PEGylated FGF-21 in treatment of diabetes in db/db mice.

The aim of this study is to explore a new method-high dose starting and low dose maintaining for PEGylated Fibroblast growth factor 21 (pFGF-21) treatment. Db/db mice were initially treated with pFGF-21 of high dose, then treated with pFGF-21 of low doses. The mice were treated with pFGF-21 at initial dose of 1.0mg/kg for 14days, then treated with pFGF-21 at maintenance doses of 0.125/0.250/0.375/0.500mg/kg for 30days. The hypoglycemic and hypolipidemic effects of pFGF-21 of different maintenance doses were compared. The pharmacological efficacy of the maintenance doses of pFGF-21 was evaluated by blood glucose levels, oral glucose tolerance test, glycosylated hemoglobin levels, insulin levels, body weight, lipid profile parameters, the mRNA expressions of glycolysis-related genes, the mRNA expressions of gluconeogenesis-related genes and the mRNA expressions of lipid metabolism-related genes. Results showed that in comparison to the mice treated only with initial dose, the treatment with pFGF-21 at maintenance doses of 0.125/0.250/0.375/0.500mg/kg exhibited favorable efficacy in lowering blood glucose levels and glycosylated hemoglobin levels, thus alleviating insulin resistance and improving dyslipidemia. However, among all of the maintenance doses, the dose of 0.125mg/kg was less effective than the other maintenance doses. These results suggest that using the treatment method of high dose of PEGylated FGF-21 in the start and low dose maintaining results in favorable control of the glycolipid metabolic balance. This study provides a new method for PEGylated FGF-21 treatment which is beneficial to promote the clinical application of PEGylated FGF-21.

Canine Fibroblast Growth Factor 21 Ameliorates Hyperglycemia Associated with Inhibiting Hepatic Gluconeogenesis and Improving Pancreatic Beta-Cell Survival in Diabetic Mice and Dogs.

Diabetes mellitus is a common endocrinopathy in dog. Fibroblast growth factor 21 (FGF-21) is a secreted protein, which is involved in glucose homeostasis. We speculate that the recombinant canine FGF-21 (cFGF-21) has the potential to become a powerful therapeutics to treat canine diabetes. The cFGF-21 gene was cloned and expressed in E. coli Rosetta (DE3). After purification, a cFGF-21 protein with the purity exceeding 95% was obtained. Mouse 3T3-L1 adipocytes and type 1 diabetic mice/dogs induced by STZ were used to examine the biological activity of cFGF-21 in vitro and in vivo, respectively. Results showed that cFGF-21 stimulated glucose uptake in adipocytes significantly in a dose-dependent manner, and reduced plasma glucose significantly in diabetic mice/dogs. After treatment with cFGF-21, the serum insulin level, glycosylated hemoglobin (HbA1c) level and the expressions of the hepatic gluconeogenesis genes (glucose-6-phosphatase, G6Pase and phosphoenolpyruvate carboxykinase, PCK) of the diabetic mice/dogs were attenuated significantly. In the mouse experiment, we also found that the phosphorylation of signal transducer and activator of transcription 3 (STAT3) and the expression of suppressor of cytokine signaling 3 (SOCS3) were up-regulated significantly in the livers after treatment. Histopathological and immunohistochemical results showed that treatment with cFGF-21 promoted recovery of pancreatic islets from STZ-induced apoptosis. Besides, we also found that treatment with cFGF-21 protected liver against STZ or hyperglycemia induced damage and the mechanism of this action associated with inhibiting oxidative stress. In conclusion, cFGF-21 represents a promising candidate for canine diabetes therapeutics. The mechanism of cFGF-21 ameliorates hyperglycemia associated with inhibiting hepatic gluconeogenesis by regulation of STAT3 signal pathway and improving pancreatic beta-cell survival.

A robust H.264/AVC video watermarking scheme with drift compensation.

A robust H.264/AVC video watermarking scheme for copyright protection with self-adaptive drift compensation is proposed. In our scheme, motion vector residuals of macroblocks with the smallest partition size are selected to hide copyright information in order to hold visual impact and distortion drift to a minimum. Drift compensation is also implemented to reduce the influence of watermark to the most extent. Besides, discrete cosine transform (DCT) with energy compact property is applied to the motion vector residual group, which can ensure robustness against intentional attacks. According to the experimental results, this scheme gains excellent imperceptibility and low bit-rate increase. Malicious attacks with different quantization parameters (QPs) or motion estimation algorithms can be resisted efficiently, with 80% accuracy on average after lossy compression.