Fibroblast Growth Factor mRNA Research Articles

Treatment with dapagliflozin increases FGF-21 gene expression and reduces triglycerides content in myocardial tissue of genetically obese mice.

The association between obesity and some cardiovascular complications such as heart failure (HF) is well established, and drugs affecting adiposity are supposed to be promising treatments for these conditions. The sodium-glucose cotransporter-2 inhibitors (SGLT2i) are antidiabetic drugs showing benefits in patients with HF, despite the underlying mechanisms have not been completely understood yet. SGLT2i are supposed to promote systemic effects, such as triglycerides mobilization, through the enhancement of fibroblast growth factor-21 (FGF-21) activity. So, in this study, weevaluated the effects of dapagliflozin treatment on FGF-21 and related receptors (FGF-Rs) gene expression and on lipid content in myocardial tissue in an animal model of genetically induced obesity to unravel possible metabolic mechanisms accounting for the cardioprotection of SGLT2i. Six-week-old C57BL/6J wild-type mice and B6.V-LEP (ob/ob) mice were randomly assigned to the control or treatment group (14 animals/group). Treatment was based on the administration of dapagliflozin 0.15mg/kg/day for 4weeks. The gene expression of FGF-21 and related receptors (FGF-R1, FGF-R3, FGF-R4, and β-klotho co-receptor) was assessed at baseline and after treatment by real-time PCR. Similarly, cardiac triglycerides concentration was measured in the control group and treated animals. At baseline, FGF-21 mRNA expression in the heart did not differ between lean and obese ob/ob mice. Dapagliflozin administration significantly increased heart FGF-21 gene expression, but only in ob/ob mice (p < 0.005). Consistently, when measuring the amount of triglycerides in the cardiac tissue, SGLT2i treatment reduced the lipid content in obese ob/ob mice, while no significant effects were observed in treated lean animals (p < 0.001). The overall expression of the FGF-21 receptors was only minimally affected by dapagliflozin treatment both in obese ob/ob mice and in lean controls. Dapagliflozin administrationincreases FGF-21gene expression and reduces triglyceride content in myocardial tissue of ob/ob mice, while no significant effect was observed in lean controls. These results might help understand the cardiometabolic effects of SGLT2i inducing increased FGF-21 synthesis while reducing lipid content in cardiomyocytes as a possible expression of the switch to different energy substrates. This mechanism could represent a potential target of SGLT2i in obesity-related heart diseases.

Fibroblast growth factor 13-mediated regulation of medium spiny neuron excitability and cocaine self-administration.

Cocaine use disorder (CUD) is a prevalent neuropsychiatric disorder with few existing treatments. Thus, there is an unmet need for the identification of new pharmacological targets for CUD. Previous studies using environmental enrichment versus isolation paradigms have found that the latter induces increased cocaine self-administration with correlative increases in the excitability of medium spiny neurons (MSN) of the nucleus accumbens shell (NAcSh). Expanding upon these findings, we sought in the present investigation to elucidate molecular determinants of these phenomena. To that end, we first employed a secondary transcriptomic analysis and found that cocaine self-administration differentially regulates mRNA for fibroblast growth factor 13 (FGF13), which codes for a prominent auxiliary protein of the voltage-gated Na+ (Nav) channel, in the NAcSh of environmentally enriched rats (i.e., resilient behavioral phenotype) compared to environmentally isolated rats (susceptible phenotype). Based upon this finding, we used in vivo genetic silencing to study the causal functional and behavioral consequences of knocking down FGF13 in the NAcSh. Functional studies revealed that knockdown of FGF13 in the NAcSh augmented excitability of MSNs by increasing the activity of Nav channels. These electrophysiological changes were concomitant with a decrease in cocaine demand elasticity (i.e., susceptible phenotype). Taken together, these data support FGF13 as being protective against cocaine self-administration, which positions it well as a pharmacological target for CUD.

FGF2 promotes the chemotherapy resistance in colon cancer cells through activating PI3K/Akt signaling pathway

Abstract Background To investigate the role of fibroblast growth factor 2 (FGF2) in chemotherapy resistance of colon cancer. Methods An HCT116/5-fluorouracil (5-FU)–resistant cell line was established, and FGF2 levels were detected in a sensitive cell group (HCT116) and a resistant cell group (HCT1116-R) using different methods. Fibroblast growth factor 2 levels in the medium were determined by enzyme-linked immunoassay. The protein expressions of FGF2, fibroblast growth factor receptor 1 (FGFR1), and phospho-FGFR1 were assessed by Western blotting, and FGF2 mRNA levels were detected by quantitative real-time polymerase chain reaction. Fibroblast growth factor 2 recombinant protein was added to sensitive cells, and FGFR inhibitor AZD4547 was added to resistant cells, and the cell survival rate was determined using the cell counting kit-8 method and the protein expressions of PI3K (phosphatidylinositol 3 kinase), p-PI3K (phospho-PI3K), Akt (protein kinase B), p-Akt (phospho-Akt), mammalian target of rapamycin (mTOR), p-mTOR (phospho-mTOR), Bad (Bcl-xL/Bcl-2–associated death promoter), NF-κB (nuclear factor κB), GSK-3 (glycogen synthase kinase-3), FKHR (forkhead box protein O1), and PTEN (phosphatase and tensin homolog deleted on chromosome ten) were detected by Western blotting. Results Fibroblast growth factor 2 protein and mRNA expression levels in the HCT116-R group were significantly higher than those in the HCT116 group. Fibroblast growth factor 2 increased the survival rate of HCT116 cells; improved tolerance to 5-FU; upregulated p-PI3K, p-Akt, and p-mTOR; and downregulated Bad. The FGFR inhibitor AZD4547 decreased cell survival rate and tolerance to 5-FU; downregulated p-PI3K, p-Akt, and p-mTOR expression; and upregulated Bad. Conclusions Fibroblast growth factor 2 promotes chemotherapy tolerance in colon cancer cells by activating the Akt/mTOR and Akt/Bad signaling pathways downstream of PI3K.

The Effect of Altered Mucin1, FGF2, and HBEGF Gene Expression at The Ectopic Implantation Site and Endometrial Tissues in The Tubal Pregnancy Pathogenesis: A Case-Control Study.

Ectopic pregnancy (EP) is defined as implantation and development of an embryo outside of the uterine tissue. Women undergoing assisted reproductive technologies (ART), particularly frozen embryo transfer (FET), are in high-risk populations for EP. Mucin1 (MUC1), fibroblast growth factor-2 (FGF2), and Heparin-binding epidermal growth factor (HBEGF) genes are involved in the endometrial receptivity pathway, leading to normal eutopic implantation; Although, their relevance in the tubal pregnancy after FET is unknown. We aimed evaluation of Mucin1, FGF2, and HBEGF expression fold as endometrial receptive markers in the EP patients following the FET cycle. A case-control study was conducted on ten patients (five EP patients and five women in the pseudo-pregnancy group, as the control samples). Pseudo-pregnancy group was established in women who were candidates for hysterectomy for benign diseases. Fallopian tube biopsies and corresponding endometrial tissues from these patients were taken during the hysterectomy. However, the fallopian tube and endometrial tissues of EP patients were obtained during salpingectomy. The mRNA expressions of Mucin1, FGF2, and HBEGF genes in the fallopian tube and endometrial tissues were measured by real-time polymerase chain reaction (PCR) assay. MUC1 mRNA expression level in the endometrium of the case group was higher than in the control group (P=0.04); however, its mRNA expression in the fallopian samples of the case group in comparison with the control group was significantly decreased (P=0.001). The HBEGF mRNA expression level was not significantly different between the case and control endometrium, whereas its expression was significantly increased in the case fallopian samples compared with the control ones (P=0.001). The same pattern was observed for FGF2 mRNA expression level in the fallopian samples of the case group but was significantly reduced in the endometrial samples in comparison with the control samples (P=0.03). Mucin1, FGF2, and HBEGF gene mRNA expression changes may explain the embryo rejection from the uterus and the establishment of a receptive phenotype in fallopian cells.

Investigation of FGF21 mRNA levels and relative mitochondrial DNA copy number levels and their relation in nonalcoholic fatty liver disease: a case-control study.

Background: Although the exact mechanisms of nonalcoholic fatty liver disease (NAFLD) are not fully understood, numerous pieces of evidence show that the variations in mitochondrial DNA (mtDNA) level and hepatic Fibroblast growth factor 21 (FGF21) expression may be related to NAFLD susceptibility. Objectives: The main objective of this study was to determine relative levels of mtDNA copy number and hepatic FGF21 expression in a cohort of Iranian NAFLD patients and evaluate the possible relationship. Methods: This study included 27 NAFLD patients (10 with nonalcoholic fatty liver (NAFL) and 17 with non-alcoholic steatohepatitis (NASH)) and ten healthy subjects. Total RNA and genomic DNA were extracted from liver tissue samples, and then mtDNA copy number and FGF21 expression levels were assessed by quantitative real-time PCR. Results: The relative level of hepatic mtDNA copy number was 3.9-fold higher in patients than in controls (p < 0.0001). NAFLD patients showed a 2.9-fold increase in hepatic FGF21 expression compared to controls (p < 0.013). Results showed that hepatic FGF21 expression was positively correlated with BMI, serum ALT, and AST levels (p < 0.05). The level of mitochondrial copy number and hepatic FGF21 expression was not significantly associated with stages of change in hepatic steatosis. Finally, there was a significant correlation between FGF21 expression and mitochondrial copy number in NAFLD patients (p = 0.027). Conclusion: Our findings suggest a considerable rise of hepatic FGF21 mRNA levels and mtDNA-CN and show a positive correlation between them in the liver tissue of NAFLD patients.

Vaginal Birth Alters Gut Transcriptome of Neonatal Pigs

Background: Women at risk of preterm birth are routinely administered prenatal doses of glucocorticoids to facilitate lung maturation and improve pulmonary outcomes in preterm infants. Additionally, preterm infants are often administered low doses of glucocorticoids postnatally for prevention of bronchopulmonary dysplasia. However, there is limited understanding about the impact of perinatal glucocorticoid exposure on the gut and liver in preterm infants. In pigs, there is a spike in plasma cortisol in newborns during parturition. We previously showed in pigs that plasma cortisol levels positively correlate with plasma fibroblast growth factor 19 (FGF19), a gut hormone that negatively regulates hepatic bile acid synthesis. FGF19 is secreted by epithelial cells predominantly in the distal ileum in response to bile acids. However, in vaginally-born pigs, FGF19 mRNA levels were higher in the proximal jejunum. This led us to hypothesize that exposure to glucocorticoids in the perinatal period has a profound impact on the transcription of genes involved in the development of gut and liver. Objective: The objective of this study was to characterize the impact of spontaneous vaginal birth and the exposure to elevated cortisol on the transcriptome in gut and liver tissue of neonatal pigs Methods: Two sows underwent a cesarean section on gestation day 114 (“Term”; term=115 day), two sows underwent a cesarean section on gestation day 105 (“Preterm”) and two additional sows were allowed to farrow at term gestational age (“Vaginal”). Small intestine tissue and plasma was collected from newborn pigs (n= 10 Preterm; n=8 Term; n=6 Vaginal). RNA-seq analysis was performed on intestinal samples mapped to the latest Sus scrofa Sscrofa11.1 reference genome (NCBI). Results: Cortisol levels were statistically highest in the vaginally-born pigs (139 ng/mL), followed by the Term (101 ng/mL) and lowest in the Preterm (61 ng/mL) pigs. Gene Ontology (GO) analysis showed significant enrichment in pathways of upregulated genes involved in response to toxic substances, stress, detoxification and inorganic substances in the jejunum of Vaginal pigs compared to Preterm and Term pigs. KEGG analysis of the proximal jejunum indicated enrichment in pathways of upregulated genes in steroid hormone biosynthesis, bile secretion, TNF signaling and IL-17 Vaginal, compared to Term and Preterm pigs. The same pattern was not the same in the distal ileum where GO analysis showed pathway enrichment of upregulated genes in oxidation-reduction processes, and transition metal ion binding in Vaginal compared to Term and Preterm pigs. KEGG analysis of the distal ileum identified pathway enrichment of upregulated genes in complement and coagulation, bile acid synthesis and secretion, fatty acid metabolism, and amino acid metabolism in Vaginal pigs. Conclusion: Genes in metabolic and cellular signaling pathways associated with cortisol are upregulated in vaginally-born pigs. This work was supported in part by federal funds from the USDA, Agricultural Research Service under Cooperative Agreement Number 3092-51000-060-01, and grants from the National Institutes of Health Grant DK-094616 (D.G.B), and the Texas Medical Center Digestive Diseases Center (NIH Grant P30 DK-56338). Caitlin Vonderohe was supported by T32 DK007664 and Greg Guthrie was supported by K01 DK129408. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Modulation of GCN2/eIF2α/ATF4 Pathway in the Liver and Induction of FGF21 in Young Goats Fed a Protein- and/or Phosphorus-Reduced Diet.

Mammals respond to amino acid (AA) deficiency by initiating an AA response pathway (AAR) that involves the activation of general control nonderepressible 2 (GCN2), phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), and activation of transcription factor 4 (ATF4). In this study, the effects of protein (N) and/or phosphorus (P) restriction on the GCN2/eIF2α/ATF4 pathway in the liver and the induction of fibroblast growth factor 21 (FGF21) in young goats were investigated. An N-reduced diet resulted in a decrease in circulating essential AA (EAA) and an increase in non-essential AA (NEAA), as well as an increase in hepatic mRNA expression of GCN2 and ATF4 and protein expression of GCN2. Dietary N restriction robustly increased both hepatic FGF21 mRNA expression and circulating FGF21 levels. Accordingly, numerous significant correlations demonstrated the effects of the AA profile on the AAR pathway and confirmed an association. Furthermore, activation of the AAR pathway depended on the sufficient availability of P. When dietary P was restricted, the GCN2/eIF2α/ATF4 pathway was not initiated, and no increase in FGF21 was observed. These results illustrate how the AAR pathway responds to N- and/or P-reduced diets in ruminants, thus demonstrating the complexity of dietary component changes.

Fibroblast growth factor 21 is expressed and secreted from skeletal muscle following electrical stimulation via extracellular ATP activation of the PI3K/Akt/mTOR signaling pathway.

Fibroblast growth factor 21 (FGF21) is a hormone involved in the regulation of lipid, glucose, and energy metabolism. Although it is released mainly from the liver, in recent years it has been shown that it is a "myokine", synthesized in skeletal muscles after exercise and stress conditions through an Akt-dependent pathway and secreted for mediating autocrine and endocrine roles. To date, the molecular mechanism for the pathophysiological regulation of FGF21 production in skeletal muscle is not totally understood. We have previously demonstrated that muscle membrane depolarization controls gene expression through extracellular ATP (eATP) signaling, by a mechanism defined as "Excitation-Transcription coupling". eATP signaling regulates the expression and secretion of interleukin 6, a well-defined myokine, and activates the Akt/mTOR signaling pathway. This work aimed to study the effect of electrical stimulation in the regulation of both production and secretion of skeletal muscle FGF21, through eATP signaling and PI3K/Akt pathway. Our results show that electrical stimulation increases both mRNA and protein (intracellular and secreted) levels of FGF21, dependent on an extracellular ATP signaling mechanism in skeletal muscle. Using pharmacological inhibitors, we demonstrated that FGF21 production and secretion from muscle requires the activation of the P2YR/PI3K/Akt/mTOR signaling pathway. These results confirm skeletal muscle as a source of FGF21 in physiological conditions and unveil a new molecular mechanism for regulating FGF21 production in this tissue. Our results will allow to identify new molecular targets to understand the regulation of FGF21 both in physiological and pathological conditions, such as exercise, aging, insulin resistance, and Duchenne muscular dystrophy, all characterized by an alteration in both FGF21 levels and ATP signaling components. These data reinforce that eATP signaling is a relevant mechanism for myokine expression in skeletal muscle.

Role of Thyrotropin-Releasing Hormone in Regulating Fibroblast Growth Factor 21 in Mouse Pancreatic β Cells.

Background: Thyrotropin-releasing hormone (TRH) is primarily produced in the hypothalamus and regulates the thyrotropin secretion from the pituitary. TRH is distributed ubiquitously in the extrahypothalamic region, especially in pancreatic islets, while its physiological role remains nebulous. We have previously established a TRH-deficient mouse model, and showed impaired glucose tolerance and downregulated expression of fibroblast growth factor 21 (FGF21) in islets. Recent studies have demonstrated the physiological roles of pancreatic FGF21. Therefore, in this study, we elucidate the direct functions of TRH in pancreatic islets via the regulation of FGF21. Methods: To explore the functions of TRH in pancreatic islets, a microarray analysis using isolated islets from TRH-knockout mice was conducted. The regulatory mechanism of TRH in pancreatic FGF21 was investigated using islet cell lines; reverse transcription-quantitative polymerase chain reaction and Western blotting were used to determine the mRNA and protein expression levels of FGF21 in pancreatic islets and islet cell lines. Induction of FGF21 expression by TRH treatment was examined in vitro. To identify the transcription factors binding to the region responsible for TRH-induced stimulation of the FGF21 promoter, electromobility shift assays were conducted. Results: Among the detected and considerably changed genes in microarray, FGF21 was the most consistently downregulated in TRH-deficient mice islets. FGF21 was strongly co-expressed with insulin in mouse islets, and TRH stimulated endogenous Fgf21 mRNA expression in the islet cell line βHC9. The E-box site in the FGF21 promoter was responsible for TRH-induced stimulation via the extracellular signal-regulated kinase (ERK)1/2 signaling pathway. The transcription factor upstream stimulatory factor 1 (USF1) could specifically bind to the E-box site. Overexpression of USF1 increased FGF21 promoter activity. Conclusion: FGF21 was transcriptionally upregulated by TRH through the ERK1/2 and USF1 pathways in pancreatic β cells.

Sodium Propionate or Sodium Butyrate Promotes Fatty Acid Oxidation in HepG2 Cells Under Oxidative Stress.

The beneficial effects of sodium butyrate (NaB) and sodium propionate (NaP) on fatty acid oxidation (FAO) genes and production of proinflammatory cytokines related to nonalcoholic fatty liver disease (NAFLD) were evaluated using HepG2 human liver hepatocellular carcinoma cells exposed to palmitate/oleate or lipopolysaccharides (LPSs) as a model. The results showed that NaP or NaB was able to promote FAO, regulate lipolysis, and reduce reactive oxygen species production by significantly increasing the mRNA expression of peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α), peroxisome proliferator-activated receptor alpha (PPARα), adipose triglyceride lipase (ATGL), carnitine palmitoyltransferase 1 alpha (CPT1α), fibroblast growth factor 21 (FGF21), and uncoupling protein 2 (UCP2) in HepG2 cells. Together, NaP and NaB may produce greater effects by increasing CPT1α, PPARα, and UCP2 mRNA expression in LPS-treated HepG2 cells and by increasing CPT1α and ATGL mRNA expression in palmitate-/oleate-treated HepG2 cells. Only NaP treatment significantly increased FGF21 mRNA expression in palmitate-/oleate-treated HepG2 cells. The enzyme-linked immunosorbent assay results revealed that only pretreatment with LPSs and not palmitate/oleate significantly increased tumor necrosis factor alpha (TNF-α) expression in HepG2 cells. NaP alone or in combination with NaB significantly decreased TNF-α expression in LPS-induced HepG2 cells. The expression of interleukin-8 in both models showed no significant differences in all treatments. NaP and NaB show potential for in vivo studies on NAFLD.

PSUN311 JTV-519 Alleviates the Impairment of FGF21 Axis Signaling after Spinal Cord Contusion in Mice

Abstract Spinal cord injury (SCI) causes adverse changes in body composition and metabolic profile due to dramatic loss of skeletal muscle mass and an extremely sedentary lifestyle. These abrupt and permanent changes increase the risk for cardiovascular disease, liver disorders, and type 2 diabetes compared to the general population. We have reported that metabolic dysregulation following SCI due to transection is associated with impaired fibroblast growth factor 21 (FGF21) and adiponectin (ADPN) signaling [1], two important regulators of metabolism [2, 3]. JTV519, which stabilizes ryanodine receptors and blocks calcium uptake via SERCA [4], has been reported to have a strong cardioprotective effect via stabilization of ryanodine receptor 2. We examined the effects of SCI caused by contusion (a more common cause of SCI), with or without JTV519 administration, on FGF21 and ADPN signaling in tissues collected from mice at 56 days after SCI. SCI reduced serum and hepatic mRNA levels of FGF21 by 75% and 45%, respectively, compared to sham mice. SCI also significantly diminished FGF21 receptor (FGF-R1) and coreceptor (beta-klotho, KLB) mRNA expression in adipose tissue. In addition, SCI decreased serum ADPN levels, particularly its high molecular weight (HMW) isoform which is the most potent form of the protein. These inhibitory effects were associated with decreased hepatic mRNA expression for PPARalpha (55%), type-2 adiponectin receptor (AdipoR2; 70%), and Glut4 (75%). In addition, SCI reduced mRNA expression of leptin (by 85%) and UCP1 (by 95%) in adipose tissue. In skeletal muscle (gastrocnemius), SCI lowered mRNA expression of PGC1alpha (by 38%), ADPN (by 80%), FGF-R1 (by 35%) and AdipoR1 (by 20%) compared to sham mice. Treating sham-mice with JTV519 upregulated FGF21 and ADPN serum and mRNA levels as well as the expression of the associated metabolic genes of interest (mentioned above). Likewise, SCI-mice treated with JTV519 also demonstrated upregulation of serum levels of FGF21, the HMW isoform of ADPN, and hepatic FGF21 mRNA expression compared to vehicle-treated SCI-mice. Decreased hepatic mRNA expression of PPARalpha, AdipoR2, Glut4 and FGF-R1 and KLB mRNA in adipose tissue were also reversed after JTV519 administration, with levels approximately equivalent to those observed in sham-vehicle treated mice but lower than sham-JTV519 treated mice. Moreover, JTV519 treatment protected against SCI-induced decreases of PGC1alpha, ADPN and FGF-R1 mRNA expression in the gastrocnemius. These data imply that JTV-519 is able to alleviate SCI-induced impairments of FGF21/ADPN signaling. Therefore, JTV519 could be a potential pharmacological agent to treat metabolic dysfunction after SCI.

Alternative polyadenylation writer CSTF2 forms a positive loop with FGF2 to promote tubular epithelial-mesenchymal transition and renal fibrosis

Effective therapies for renal fibrosis, the common endpoint for most kidney diseases, are lacking. We previously reported that alternative polyadenylation (APA) drives transition from acute kidney injury to chronic kidney disease, suggesting a potential role for APA in renal fibrogenesis. Here, we found that among canonical APA writers, CSTF2 expression was upregulated in tubular epithelial cells (TEC) of fibrotic kidneys. CSTF2 was also identified as a TGF-β-inducible pro-fibrotic gene. Further analysis revealed that CSTF2 promoted epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) overproduction in TEC by inducing 3’UTR shortening and upregulation of the expression of basic fibroblast growth factor 2 (FGF2). Additionally, 3’UTR shortening stabilised FGF2 mRNA through miRNA evasion. Interestingly, FGF2 enhanced CSTF2 expression, leading to the forming of a CSTF2-FGF2 positive loop in TEC. Furthermore, CSTF2 knockdown alleviated unilateral ureteral obstruction-induced renal fibrosis in vivo. Finally, we developed a CSTF2-targeted antisense oligonucleotide (ASO) and validated its effectiveness in vitro. These results indicate that the expression of the APA writer, CSTF2, is upregulated by TGF-β and CSTF2 facilitates TGF-β-induced FGF2 overexpression, forming a TGF-β-CSTF2-FGF2 pro-fibrotic axis in TEC. CSTF2 is a potentially promising target for renal fibrosis that does not directly disrupt TGF-β.

The regulation mechanism of different hair types in inner Mongolia cashmere goat based on PI3K-AKT pathway and FGF21.

There is genetic diversity of hair types in the Inner Mongolia cashmere goat population. Previous studies have found that fibroblast growth factor 21 (FGF21) and PI3K-AKT signal pathways may be related to different hair types in Inner Mongolia cashmere goats. Therefore, the purpose of this study was to explore the effects of the PI3K-AKT signal pathway on different hair types, the expression of mRNA and protein expression sites of FGF21 in the hair follicles of cashmere goats with different hair types, so as to lay a foundation for understanding the molecular mechanism of different hair types and the role of skin hair follicle development. In this experiment, the skin tissues of long hair type (LHG) and short hair type (SHG) of Inner Mongolia cashmere goat were collected in three key periods of secondary hair follicle growth, namely, anagen (September), catagen (December), and telogen (March). The relative expression of FGF21 and PI3K-AKT signal pathway candidate gene mRNA in different periods and different hair types was detected by real-time fluorescence quantitative technique (qRT-PCR), and the expression site of FGF21 protein was located by immunohistochemical technique. Through qRT-PCR, it was found that the relative expression of FGF21, FGFR1, AKT3, BRCA1, PKN3, SPP1, and GNG4 was significantly different between LHG and SHG. The expression of FGF21 in the skin of LHG was significantly higher than that of SHG in the three periods. Through immunohistochemical test, it was found that FGF21 protein was mainly expressed in primary hair follicle connective tissue sheath, primary hair follicle outer root sheath, secondary hair follicle outer root sheath, and sebaceous glands. It was also found that the expression of LHG skin tissue in the outer root sheath of primary hair follicles was higher than that of SHG in three periods. In summary, it is suggested that the PI3K-AKT signal pathway may play an important role in the formation of different hair types in Inner Mongolia cashmere goats.

Central FGF21 production regulates memory but not peripheral metabolism.

SUMMARYFibroblast growth factor 21 (FGF21) is a liver-derived endocrine hormone that functions to regulate energy homeostasis and macronutrient intake. Recently, FGF21 was reported to be produced and secreted from hypothalamic tanycytes, to regulate peripheral lipid metabolism; however, rigorous investigation of FGF21 expression in the brain has yet to be accomplished. Using a mouse model that drives CRE recombinase in FGF21-expressing cells, we demonstrate that FGF21 is not expressed in the hypothalamus, but instead is produced from the retrosplenial cortex (RSC), an essential brain region for spatial learning and memory. Furthermore, we find that central FGF21 produced in the RSC enhances spatial memory but does not regulate energy homeostasis or sugar intake. Finally, our data demonstrate that administration of FGF21 prolongs the duration of long-term potentiation in the hippocampus and enhances activation of hippocampal neurons. Thus, endogenous and pharmacological FGF21 appear to function in the hippocampus to enhance spatial memory.

MiR‑491‑3p functions as a tumor suppressor in non‑small cell lung cancer by targeting fibroblast growth factor 5

The present study aimed to identify the function of miR-491-3p in regulating non-small cell lung cancer (NSCLC). Tumor tissues and adjacent normal tissues were collected from 43 patients with NSCLC. A549 and H1299 cells were transfected with microRNA (miR)-491-3p mimic, mimic negative control (NC), miR-491-3p inhibitor, inhibitor NC, pcDNA3.1-FGF5 vector and control vector. Cell counting kit-8 assay and Edu experiments were performed to assess cell viability and proliferation. Matrigel experiment, wound healing assay and flow cytometric analysis were performed to explore cell invasion, migration and apoptosis, respectively. A dual-luciferase reporter experiment was performed to identify the relationship between miR-491-3p and fibroblast growth factor 5 (FGF5). In vivo study was conducted by using nude mice. The miR-491-3p and FGF5 protein expression levels were investigated using reverse transcription-quantitative polymerase chain reaction and western blot analysis. In NSCLC tumor tissues, miR-491-3p was downregulated and FGF5 was upregulated (P<0.01). Low miR-491-3p expression and high FGF5 mRNA expression was associated with poor outcomes in patients, including advanced TNM stage and lymph node metastasis (P<0.05). upregulation of miR-491-3p suppressed viability, proliferation, invasion and migration of NSCLC cells; however, it promoted apoptosis (P<0.01). FGF5 was a target gene for miR-491-3p. miR-491-3p directly inhibited FGF5 expression. upregulation of FGF5 significantly reversed the inhibitory effects of miR-491-3p on malignant phenotypes of NSCLC cells (P<0.01). miR-491-3p overexpression suppressed the in vivo growth of NSCLC. Thus, it was identified that miR-491-3p functions as a tumor suppressor in NSCLC by directly targeting FGF5.

The Association Between FGF21 and Diabetic Erectile Dysfunction: Evidence from Clinical and Animal Studies.

Erectile dysfunction (ED), a complication of diabetes mellitus (DM), affects 50–75% of men with diabetes. Fibroblast growth factor 21 (FGF21) is a liver-derived metabolic regulator which plays a role in insulin-independent glucose uptake in adipocytes. We designed a clinical study and an animal experiment to investigate the relationship between FGF21 and DM-induced ED. The clinical study enrolled 93 participants aged > 18 years (61 patients with type 2 DM and 32 healthy controls) from Taian City Central Hospital (TCCH) in Shandong Province, China, amongst whom the association between serum FGF21 and diabetic ED was analyzed. To further validate this association, we developed animal model of diabetic ED using Sprague-Dawley (SD) rats. Serum FGF21 concentration and FGF21 mRNA expression in penile samples of the rats were determined with Western blotting and quantitative real-time PCR. Among the 93 participants, the level of serum FGF21 was negatively correlated with the IIEF-5 score (r = -0.74, P < 0.001). The analysis on the performance of FGF21 for ED diagnosis showed that the area under the receiver operating characteristic (ROC) curve was 0.875 (95% confidence interval [CI]: 0.803 to 0.946). In the animal experiment, the levels of serum FGF21, 2-Δ Δ Ct values of FGF21 mRNA expression, and relative levels of FGF21 in penile samples were higher in the ED group compared to the DM and control groups. Our findings demonstrated an association between the FGF21 level and diabetic ED, indicating the potential of this cytokine in predicting diabetic ED.

Subcutaneous delivery of FGF21 mRNA therapy reverses obesity, insulin resistance, and hepatic steatosis in diet-induced obese mice

Fibroblast growth factor 21 (FGF21) is a promising therapeutic agent for treatment of type 2 diabetes (T2D) and non-alcoholic steatohepatitis (NASH). We show that therapeutic levels of FGF21 were achieved following subcutaneous (s.c.) administration of mRNA encoding human FGF21 proteins. The efficacy of mRNA was assessed following 2-weeks repeated s.c. dosing in diet-induced obese (DIO), mice which resulted in marked decreases in body weight, plasma insulin levels, and hepatic steatosis. Pharmacokinetic/pharmacodynamic (PK/PD) modelling of several studies in both lean and DIO mice showed that mRNA encoding human proteins provided improved therapeutic coverage over recombinant dosed proteins in vivo. This study is the first example of s.c. mRNA therapy showing pre-clinical efficacy in a disease-relevant model, thus, showing the potential for this modality in the treatment of chronic diseases, including T2D and NASH.

Next-Generation Sequencing of Circular RNAs Reveals the Molecular Mechanisms of Chondrogenic Differentiation in Human Adipose-derived Stem Cells.

Adipose-derived stem cells are one of the potential sources of cells for the treatment of cartilage defects. This study aimed to investigate the molecular mechanisms that account for the chondrogenic differentiation of human adipose-derived stem cells (hADSCs). We employed integrin β1 (ITGB1) overexpression to induce chondrogenic differentiation of hADSCs. Next-generation sequencing was used to determine the mRNAs and circular RNAs (circRNAs) expression profiles in ITGB1-overexpresing and negative control cells. The potential functions of differentially expressed mRNAs were analyzed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses. Moreover, differentially expressed circRNAs with the greatest fold change were validated by polymerase chain reaction (PCR), Sanger sequencing, and quantitative real-time PCR (qRT-PCR). These three circRNAs and their downstream microRNAs and mRNAs were used to construct a circRNA-microRNA-mRNA interaction network. The results showed that we identified 713 differentially expressed circRNAs (150 upregulated and 563 downregulated in ITGB1-overexpressing hADSCs versus negative control cells, respectively). Meanwhile, 2383 mRNAs were differentially expressed between two groups (1672 upregulated and 711 downregulated in ITGB1-overexpressing cells compared with the negative control cells). The GO and KEGG analysis results showed that the differentially expressed mRNAs were enriched in biological processes, cellular components, and molecular functions, especially in the phosphatidylinositol 3-kinase (PI3K)-AKT and mitogen-activated protein kinase signaling pathways. Three differentially expressed circRNAs, including hsa_circ_0071127, hsa_circ_0008637, and hsa_circ_0020028, were validated by qRT-PCR. Moreover, the circRNA-microRNA-mRNA network predicted that fibroblast growth factor 2 (FGF2) was a common node regulated by these three circRNAs through several microRNAs, including miR-195-3p, miR-205-3p, and miR-152-3p. We further found that the knockdown of hsa_circ_0020028, but not the two other circRNAs, significantly reduced FGF2 mRNA expression in hADSCs. Furthermore, the knockdown of hsa_circ_0020028 significantly inhibited the protein expression of FGF2, chondrogenic differentiation markers (COL II, aggrecan, and SOX9), and PI3K/AKT signaling in ITGB1-overexpressing hADSCs. This study uncovered the differentially expressed mRNA and circRNA profiles in the chondrogenic differentiation of hADSCs induced by ITGB1 overexpression. Our findings demonstrate that hsa_circ_0020028 regulates the ITGB1 overexpression-mediated chondrogenic differentiation of hADSCs through regulation of FGF2-related signaling pathways.

Transmembrane G protein-coupled receptor 5 signaling stimulates fibroblast growth factor 21 expression concomitant with up-regulation of the transcription factor nuclear receptor Nr4a1

Fibroblast growth factor 21 (FGF21) acts as an endocrine factor, playing important roles in the regulation of energy homeostasis, glucose and lipid metabolism. It is induced by diverse metabolic and cellular stresses, such as starvation and cold challenge, which in turn facilitate adaptation to the stress environment. The pharmacological action of FGF21 has received much attention, because the administration of FGF21 or its analogs has been shown to have an anti-obesity effect in rodent models. In the present study, we found that 3-O-acetyloleanolic acid, an active constituent isolated from the fruits of Forsythia suspensa, stimulated FGF21 production concomitant with the up-regulation of a transcription factor, nuclear receptor Nr4a1, in C2C12 myotubes. Additionally, significant increases in mFgf21 promoter activity were observed in C2C12 cells overexpressing TGR5 receptor in response to 3-O-acetyloleanolic acid treatment. Treatment with the p38 MAPK inhibitor SB203580 was effective at suppressing these stimulatory effects of 3-O-acetyloleanolic acid. Pretreatment with SB203580 also significantly repressed FGF21 mRNA abundance and FGF21 secretion in C2C12 myotubes after 3-O-acetyloleanolic acid stimulation, suggesting that p38 activation is required for the induction of FGF21 by ligand-activated TGR5 in C2C12 myotubes. These findings collectively indicated that TGR5 receptor signaling drives FGF21 expression via p38 activation, at least partly, by mediating Nr4a1 expression. Thus, the novel biological function of 3-O-acetyloleanolic acid as an agent having anti-obesity effects is likely to be mediated through the activation of TGR5 receptors.

Upregulation of fibroblast growth factor 2 contributes to endometriosis through SPRYs/DUSP6/ERK signaling pathway

Previous studies report that fibroblast growth factor 2 (FGF2) modulates Sproutys (SPRYs)/dual specificity phosphatase 6 (DUSP6)/extracellular signal-regulated kinase (ERK) signaling pathway in endometrial glandular epithelial cells. However, its role in endometriosis remains unclear. The expression patterns and localization of related proteins in endometrium patients’ samples were determined using quantitative reverse transcription PCR, Western blotting, and immunohistochemistry, respectively. Human endometrial stromal cells (HESCs) were isolated and transfected with small interfering RNA (siRNA) targeting FGF2 (FGF2-siRNA). Cell viability was determined using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. It was found that FGF2 mRNA and protein levels were increased in the ectopic endometrium, whilst the mRNA and protein levels of SPRYs/DUSP6/ERK signaling pathway related-genes were dysregulated. Spearman’s rank correlation analysis revealed a negative correlation between FGF2 and SPRYs/DUSP6 signaling pathway-related proteins. In vitro study demonstrated that FGF2 silencing suppressed cell proliferation. Our results suggest that FGF2 upregulation might contribute to endometriosis via the regulation of the SPRYs/DUSP6/ERK signaling pathway.