With the increasing prevalence of diabetes in recent years, diabetic nephropathy (DN) has become a severe disease that greatly threatens human health. DN not only is a common complication of diabetes, but also takes an important place in kidney disease. To this end, the present study was designed to explore the effects of Forkhead box protein O1 (FoxO1) on reactive oxygen species (ROS) production in DN mice. DN mice were treated with recombinant protein of FoxO1. Afterward, inflammation ELISA kits were used to measure the levels of TNF-α, IL-1β, IL-6, and IL-18. The levels of MDA, SOD, GSH, and GSH-PX were measured using kits according to the manufacturer's instructions. In addition, the production of ROS was assessed. Interestingly, the expression of FoxO1 was down-regulated in DN mice. The treatment of FoxO1 recombinant protein ameliorated MDA levels, increased the levels of SOD, GSH, and GSH-PX, and induced both mRNA and protein expression of hepatic serine protease inhibitor B1 (serpinB1) in ND mice. Similarly, FoxO1 reduced MDA levels and ROS production, increased the levels of SOD, GSH, and GSH-PXs, and induced the mRNA and protein expression of serpinB1 in in vitro model of DN. The inhibition of serpinB1 attenuated the effects of FoxO1 on ROS production-induced oxidative stress in in vitro model of DN. Overall, FoxO1/SERPINB1 ameliorated ROS production-induced oxidative stress in DN.

Transcription factors forkhead box protein O1 (FOXO1) and paired box 3 (PAX3) have been reported to play important roles in various cancers. However, their role in epithelial ovarian cancer (EOC) has not been elucidated yet. Therefore, we evaluated the expression and clinical significance of FOXO1 and PAX3 in EOC.

Calorie restriction (CR) reportedly prevents atherosclerotic diseases. Furthermore, CR induces forkhead box protein-O1 (FOXO-1) expression in the skeletal muscle, altering the character of the skeletal muscle. We previously reported that the change in skeletal muscle character, induced by the overexpression of peroxisome proliferator-activated receptor γ coactivator-1α, suppresses atherosclerotic progression in an atherosclerotic apolipoprotein E-knockout (ApoE-KO) mouse model. Thus, we hypothesized that skeletal muscle alternation induced by FOXO-1 may also have an anti-atherosclerotic effect in ApoE-KO mice. In this study, we investigated whether skeletal muscle-specific FOXO-1 overexpression suppresses the progression of atherosclerosis in ApoE-KO mice. We generated ApoE-KO/FOXO-1 mice, in which an ApoE-KO mouse was crossbred with a mouse presenting skeletal muscle-specific FOXO-1 overexpression (FOXO-1Tg). The mice were sacrificed at 20 weeks of age, and atherosclerotic plaque area and protein expression in the plaque were measured. Additionally, we measured the tumor necrosis factor α (TNFα)- induced mRNA expression in human umbilical vein endothelial cells (HUVECs), using serum collected from the FOXO-1Tg mice. Accordingly, ApoE-KO/FOXO-1 mice showed a 65% reduced atherosclerotic plaque area when compared with the ApoE-KO mice, with concomitantly reduced vascular cell adhesion molecule-1 (VCAM-1) and macrophage infiltration. As compared to serum from wild-type mice, the serum collected from the FOXO-1Tg mice significantly suppressed the mRNA expression of VCAM-1, an atherosclerosis initiation factor, in TNFα-treated HUVECs. Therefore, these data suggest that skeletal muscle-specific FOXO-1 overexpression suppresses the progression of atherosclerosis in ApoE-KO mice. In part, the CR-induced anti-atherosclerotic effect could be attributed to FOXO-1 upregulation in the skeletal muscle.

Forkhead box O (FoxO) transcription factors and E3 ubiquitin ligases such as Muscle RING finger 1 (MuRF1) are believed to participate in the regulation of skeletal muscle mass. The function of FoxO transcription factors is regulated by post-translational modifications such as phosphorylation and acetylation. In the present study FoxO1 protein expression, phosphorylation and acetylation as well as MuRF1 protein expression, were examined in atrophic and hypertrophic denervated skeletal muscle.

Porphyromonas gingivalis (P. gingivalis) is a pathogen involved in periodontal disease. Recently, periodontal disease has been demonstrated to increase the risk of developing diabetes mellitus, although the molecular mechanism is not fully understood. Forkhead box protein O1 (FoxO1) is a transcriptional factor that regulates gluconeogenesis in the liver. Gluconeogenesis is a key process in the induction of diabetes mellitus; however, little is known regarding the relationship between periodontal disease and gluconeogenesis. In this study, to investigate whether periodontal disease influences hepatic gluconeogenesis, we examined the effects of P. gingivalis on the phosphorylation and translocation of FoxO1 in insulin-induced human hepatocytes.

The invariant NKT (iNKT) cells recognize glycolipid antigens presented by the non-classical MHC like molecule CD1d. They represent an innate T-cell lineage with the ability to rapidly produce a variety of cytokines in response to agonist stimulation to bridge innate and adaptive immunity. In thymus, most iNKT cells complete their maturation and differentiate to multiple effector lineages such as iNKT-1, iNKT-2, and iNKT-17 cells that possess the capability to produce IFNγ, IL-4, and IL-17A, respectively, and play distinct roles in immune responses and diseases. Mechanisms that control iNKT lineage fate decisions are still not well understood. Evidence has revealed critical roles of Foxo1 of the forkhead box O1 subfamily of transcription factors in the immune system. However, its role in iNKT cells has been unknown. In this report, we demonstrate that deletion of Foxo1 causes severe decreases of iNKT cell total numbers due to impairment of late but not early iNKT cell development. Deficiency of Foxo1 results in decreases of iNKT-1 but increases of iNKT-17 cells. Our data reveal that Foxo1 controls iNKT effector lineage fate decision by promoting iNKT-1 but suppressing iNKT-17 lineages.

MicroRNA (miR)-421 has been reported to serve various important roles in numerous types of cancer, including neuroblastoma and gastric cancer. However, to the best of our knowledge, few reports have determined the role of miR-421 in lung cancer. The aim of the current study was to analyze the expression levels of miR-421 in A549 lung cancer cells, to determine the target gene of miR-421, and to investigate the function and mechanism of miR-421 in cellular cytotoxicity. miR-421 expression levels were analyzed in A549 lung cancer cells using reverse transcription-quantitative PCR, a MTT assay was performed to determine the effect of miR-421 on A549 cell cytotoxicity and the protein expression levels of forkhead box O1 (FOXO1) were determined via western blotting. The target gene of miR-421 was predicted and verified using TargetScan and a dual-luciferase reporter assay, respectively. The results revealed that miR-421 expression levels were significantly upregulated in A549 lung cancer cell lines compared with the normal cells (P<0.01). Additionally, it was discovered that miR-421 promoted A549 cell viability (P<0.01) compared with A549 transfected with negative control. miR-421 was also identified to bind to the 3'-untranslated region of FOXO1. In A549 cells transfected with miR-421-mimics, the expression levels of phosphorylated (p)-AKT, p-glycogen synthase kinase-3β, p-retinoblastoma and cyclin D1 were significantly upregulated (P<0.01), whereas the expression levels of FOXO1 and p21 were significantly downregulated (P<0.01) compared with the control group. In conclusion, the results of the present study suggested that miR-421 may promote the viability of A549 lung cancer cells by targeting FOXO1 and modulating cell cycle, indicating that targeting miR-421 and FOXO1 may represent future therapeutic strategies for the treatment of patients with lung cancer.

Insulin resistance and compensatory hyperinsulinemia are characteristic features of obesity and polycystic ovary syndrome, and both are associated with reduced fertility and implantation. There is little knowledge about the effect of insulin on the decidualization process and previous findings are contradictory. We investigated the effect of insulin on the regulation of forkhead box protein O1 (FOXO1), one of the most important transcription factors during decidualization. Endometrial stromal cells were isolated from six healthy, regularly menstruating women and decidualized in vitro. Gene expression levels of six putative FOXO1 target genes (including insulin-like growth factor binding protein-1 (IGFBP1) and prolactin (PRL)) were measured with Real-Time PCR following FOXO1 inhibition or insulin treatment. PI3K inhibition was used to identify the possible mechanism behind regulation. Subcellular localization of FOXO1 was analyzed with immunofluorescence. All the genes (IGFBP1, CTGF, INSR, DCN, LEFTY2), except prolactin, were evaluated as FOXO1 target genes in decidualizing stromal cells. Insulin caused a significant dose-dependent inhibition of the verified FOXO1 target genes. It was also demonstrated that insulin regulated FOXO1 target genes by transcriptional inactivation and nuclear export of FOXO1 via PI3K pathway. However, insulin did not inhibit the morphological transformation of endometrial stromal cells via transcriptional inactivation of FOXO1. This study provides new insights on the action of insulin on the endometrium via regulation of FOXO1. It is suggested that hyperinsulinemia results in dysregulation of a high number of FOXO1 controlled genes that may contribute to endometrial dysfunction and reproductive failure. Our findings may illuminate possible reasons for unexplained infertility.

As the major cause of female anovulatory infertility, polycystic ovary syndrome (PCOS) affects a great proportion of women at childbearing age. Although glucagon-like peptide 1 receptor agonists (GLP-IRAs) show therapeutic effects for PCOS, its target and underlying mechanism remains elusive. In the present study, we identified that, both in vivo and in vitro, GLP-1 functioned as the regulator of proliferation and antiapoptosis of MGCs of follicle in PCOS mouse ovary. Furthermore, forkhead box protein O1 (FoxO1) plays an important role in the courses. Regarding the importance of granulosa cells (GCs) in oocyte development and function, the results from the current study could provide a more detailed illustration on the already known beneficial effects of GLP-1RAs on PCOS and support the future efforts to develop more efficient GLP-1RAs for PCOS treatment.

Forkhead box O1 (foxo1) is a transcription factor and plays important roles in glucose metabolism. In the present study, foxo1 in turbot Scophthalmus maximus was cloned and characterized. The siRNA of foxo1 was used to investigate the functions of foxo1 in turbot hepatocytes glucose metabolism. After that, a 10-week feeding trial with two different dietary carbohydrate levels (15% and 21%, respectively) was conducted to analyze the function of foxo1 in glucose metabolism in vivo. Results showed that the foxo1 was identified as 2176 bp (base pair) with a 2025 bp open reading frame, which encoded 675 amino acids. Sequence analysis showed that foxo1 of turbot was highly homologous to most of fishes. Tissue distribution analysis revealed that the highest expression of foxo1 was in liver. After in vitro analysis, foxo1-specific small interfering RNA (sifoxo1) treatment significantly decreased the expressions of cytosolic phosphoenolpyruvate carboxykinase (cpepck) and glucose-6-phosphatase1(g6pase1) in primary hepatocytes. Expression of mitochondrial phosphoenolpyruvate carboxykinase (mpepck) was not significantly inhibited. In contrast, the expression of glucose-6-phosphatase2 (g6pase2) increased significantly. After the in vivo study (feeding trial), with the decreased expression of foxo1 in turbot due to high dietary carbohydrate level (21%), the expression of g6pase2 was significantly upregulated. However, the expression of glucokinase (gk) was not changed significantly. These increased the level of blood glucose and hepatic glycogen. In conclusion, data from both in vitro (primary hepatocytes) and in vivo (feeding trial) showed that downregulated foxo1 in turbot could not result in significant depression of gluconeogenesis and activation of glycolysis. This could be one of the reasons why intake of high level of carbohydrate resulted in prolonged hyperglycemia in turbot.

Type 2 diabetes mellitus (T2DM) is a chronic disease manifested by hyperglycemia. It is essential to effectively control hyperglycemia to prevent complications of T2DM. Here, we hypothesize that repression of transcriptional activity of forkhead box O1 (FoxO1) via histone deacetylase inhibitors (HDACi) ameliorates hyperglycemia in T2DM rats.

The recessive disease arterial calcification due to deficiency of CD73 (ACDC) presents with extensive nonatherosclerotic medial layer calcification in lower extremity arteries. Lack of CD73 induces a concomitant increase in TNAP (tissue nonspecific alkaline phosphatase; ALPL), a key enzyme in ectopic mineralization. Our aim was to investigate how loss of CD73 activity leads to increased ALPL expression and calcification in CD73-deficient patients and assess whether this mechanism may apply to peripheral artery disease calcification. Approach and Results: We previously developed a patient-specific disease model using ACDC primary dermal fibroblasts that recapitulates the calcification phenotype in vitro. We found that lack of CD73-mediated adenosine signaling reduced cAMP production and resulted in increased activation of AKT. The AKT/mTOR (mammalian target of rapamycin) axis blocks autophagy and inducing autophagy prevented calcification; however, we did not observe autophagy defects in ACDC cells. In silico analysis identified a putative FOXO1 (forkhead box O1 protein) binding site in the human ALPL promoter. Exogenous AMP induced FOXO1 nuclear localization in ACDC but not in control cells, and this was prevented with a cAMP analogue or activation of A2a/2b adenosine receptors. Inhibiting FOXO1 reduced ALPL expression and TNAP activity and prevented calcification. Mutating the FOXO1 binding site reduced ALPL promoter activation. Importantly, we provide evidence that non-ACDC calcified femoropopliteal arteries exhibit decreased CD73 and increased FOXO1 levels compared with control arteries.

BACKGROUND Gegen qinlian decoction (GGQLD) is a form of traditional Chinese medicine used for hundreds of years for its efficacy in treating diabetes. However, the mechanisms underlying the therapeutic effects of GGQLD on diabetes are still not clear. We aimed to evaluate the effect of GGQLD on hepatic insulin resistance (IR) through silent information regulator1 (SIRT1)/forkhead box O1 (FOXO1) in an IR mouse model. MATERIAL AND METHODS A high-fat diet (HFD) mouse model was established and GGQLD was administrated by oral gavage. Metabolic parameters were detected, including body weights, triglyceride, fasting glucose, fasting insulin and HOMA-IR index, glucose intolerance, and insulin resistance. HE-stained sections were used to observe the histopathology of liver tissue. For in vitro study, GGQLD-medicated serum was used to treat palmitic acid-stimulated HepG2 cells. The glycogen synthesis and downstream SIRT1/FOXO1 signaling pathways were examined. Specific siRNAs were used to knock down SIRT1 in HepG2 cells. RESULTS GGQLD administration significantly decreased body weights, triglyceride level, fasting glucose level, fasting insulin level, and HOMA-IR index, and improved IR in HFD mice. GGQLD enhanced SIRT1 expression and suppressed the expression of Ac-FOXO1 in liver tissues. Further, GGQLD-medicated serum promoted SIRT1 upregulation and suppressed Ac-FOXO1 levels in palmitate-stimulated HepG2 cells. GGQLD-medicated serum also increased the protein expression of PPARγ and reduced the expression of FABP4 in palmitate-stimulated HepG2 cells. CONCLUSIONS We found that GGQLD alleviates insulin resistance through SIRT1-dependent deacetylation of FOXO1.

This study aims to explore the impact of Rehmannioside D (RD) on ovarian functions of rats with diminished ovarian reserve (DOR) and its underlying mechanisms of action. A single injection of cyclophosphamide was performed to establish a DOR rat model, and fourteen days after the injection, the rats were intragastrically administrated with RD for two weeks. Rat estrus cycles were tested using vaginal smears. Ovarian tissues were histologically evaluated, the number of primordial, mature, and atretic follicles was calculated, and the apoptotic rate of granulosa cells. Follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) levels were determined by ELISA assays. Protein levels of Forkhead Box O1 (FOXO1), KLOTHO, Bcl-2, and Bax were investigated in ovarian tissues of DOR rats. The binding between FOXO1 and KLOTHO was verified by ChIP assay. High-dose administration of RD into DOR rats improved their estrus cycles, increased ovarian index, enhanced the number of primordial and mature follicles, reduced the number of atretic follicle number, and ovarian granulosa cell apoptosis in addition to inhibiting FSH and LH levels and upregulating E2 expression. FOXO1 and KLOTHO were significantly suppressed in DOR rats. FOXO1 knockdown partially suppressed the protective effects of RD on DOR rats, and KLOTHO overexpression could restore RD-induced blockade of DOR development despite knocking down FOXO1. FOXO1 antibody enriched KLOTHO promoter, and the binding between them was reduced in DOR group compared to that in sham group. RD improved ovarian functions in DOR rats and diminished granulosa cell apoptosis via the FOXO1/KLOTHO axis.

Ovarian cancer is a prominent disease that demonstrates high incidence rates in women and often presents multidrug resistance. Propofol has been demonstrated to suppress the malignancy of various types of human cancer; however, the underlying molecular mechanisms of propofol in ovarian cancer remain largely unknown. The present study aimed to investigate whether and how propofol inhibits proliferation and cisplatin (DDP) resistance in ovarian cancer cells. Ovarian cancer cell viability was assessed by the Cell Counting kit‑8 assay; apoptosis and cell cycle progression were determined by flow cytometry; the relative expression levels of microRNA (miR)‑374a and forkhead box O1 (FOXO1) were analyzed using reverse transcription‑quantitative PCR; the binding ability of miR‑374a to FOXO1 was assessed by the dual‑luciferase reporter assay; cellular sensitivity to DDP was detected using the MTT assay; and finally, the protein expression levels of FOXO1, p27, and Bcl‑2‑like‑protein 11 (Bim) were analyzed by western blotting. Propofol reduced viability, promoted apoptosis and decreased miR‑374a expression levels in A2780 cells. In addition, the viability of A2780/DDP cells in the propofol + DDP treatment group was significantly inhibited, and the apoptotic rate was increased. In addition, miR‑374a overexpression increased cell viability and the proportion of cells in the S phase, and decreased the proportion of cells in the G0/G1 phase. Conversely, genetic knockdown of miR‑374a exerted the opposite effects on cell viability and cell cycle progression. Moreover, miR‑374a was demonstrated to bind to FOXO1. Propofol promoted the expression of FOXO1, p27 and Bim, induced cell cycle arrest and decreased ovarian cancer cell viability. In addition, treatment with propofol and DDP regulated FOXO1 and increased apoptosis of ovarian cancer cells. In conclusion, propofol downregulated miR‑374a and modulated the FOXO1 pathway to reduce proliferation and DDP resistance in ovarian cancer cells.

Decidualization is a prerequisite for successful implantation and the establishment of pregnancy. Krüppel-like factor 12 (KLF12) is a negative regulator of endometrial decidualization in vitro. We investigated whether KLF12 was associated with impaired decidualization under conditions of repeated implantation failure (RIF).

Hypertension is an independent risk factor for the progression of chronic renal failure, and oxidative stress plays a critical role in hypertensive renal damage. Forkbox O1(FoxO1) signaling protects cells against oxidative stress and may be a useful target for treating oxidative stress-induced hypertension. Tongxinluo is a traditional Chinese medicine with cardioprotective and renoprotective functions. Therefore, this study aimed to determine the effects of Tongxinluo in hypertensive renal damage in spontaneously hypertensive rats(SHRs)and elucidate the possible involvement of oxidative stress and FoxO1 signaling in its molecular mechanisms. SHRs treated with Tongxinluo for 12 weeks showed a reduction in systolic blood pressure. In addition to increasing creatinine clearance, Tongxinluo decreased urinary albumin excretion, oxidative stress injury markers including malondialdehyde and protein carbonyls, and expression of nicotinamide adenine dinucleotide phosphate oxidase subunits and its activity in SHR kidneys. While decreasing phosphorylation of FoxO1, Tongxinluo also inhibited the phosphorylation of extracellular signal-regulated kinase1/2 and p38 and enhanced manganese superoxide dismutase and catalase activities in SHR kidneys. Furthermore, histology revealed attenuation of glomerulosclerosis and renal podocyte injury, while Tongxinluo decreased the expression of α-smooth muscle actin, extracellular matrixprotein, transforming growth factor β1 and small mothers against decapentaplegic homolog 3,and improved tubulointerstitial fibrosis in SHR kidneys. Finally, Tongxinluo inhibited inflammatory cell infiltration as well as expression of tumor necrosis factor-α and interleukin-6. In conclusion, Tongxinluo protected SHRs against hypertension-induced renal injury by exerting antioxidant, antifibrotic, and anti-inflammatory activities. Moreover, the underlying mechanisms of these effects may involve inhibition of oxidative stress and functional activation of FoxO1 signaling.

Forkhead box protein O1 (FoxO1) is a transcription factor which promotes hepatic glucose production (HGP) by up-regulating the transcription of gluconeogenic enzymes in monogastric species. The activity of FoxO1 is inhibited by insulin-induced phosphorylation. The aims of the present study were to find associations between FoxO1 expression and variables associated with HGP as affected by feeding regimen in dairy cows during the transition period. Twenty one healthy German Holstein cows were allocated to four groups (LC-CON, HC-CON, LC-NA with 5 cows/group and HC-NA with 6 cows/group, respectively). Cows received 0 (LC-CON and HC-CON) or 24 (LC-NA and HC-NA) g/d nicotinic acid with high (HC) or low (LC) concentrate proportion from -42 days (-41.8 + 4.8; mean + standard deviation) relative to expected calving date (d-42) to d24. Liver biopsy was taken at d-42, 1, 21, and 100. The total protein expression of FoxO1 (tFoxO1) and the extent of phosphorylation of FoxO1 at serine 256 (pFoxO1) were analysed semiquantitatively by Western Blotting. The expression of hepatic mRNA of FoxO1 and seven genes associated with HGP was measured by real-time RT-PCR. Mixed model and Pearson's correlation were used for statistical evaluation with the level of significance at P<0.05. No dietary effect was observed either on feed intake, energy balance, or on the concentration of blood metabolites. Neither time nor diet affected the expression of FoxO1 total protein and mRNA. A NA × concentrate interaction was found in pFoxO1. However, no corresponding dietary effect was found in the mRNA expression of investigated genes. Different patterns of correlations between FoxO1-related variables and investigated indicators for HGP were found at d21 and 100. The results indicated that the regulation of HGP did not take place on the levels of mRNA and protein expression and the phosphorylation of FoxO1 in dairy cows in early lactation.

Phosphatidylinositol Transfer Protein, Membrane-Associated 3 (PITPNM3) often bind with chemokine (C-C motif) ligand 18 (CCL18) to promote tumor progression. However, the role of PITPNM3 in intrahepatic cholangiocarcinoma (ICC) is unclear. We first searched GEPIA database and detected the PITPNM3 expression using immunohistochemistry and real-time quantitative PCR. The results showed that PITPNM3 is high expression in ICC tissues and cells. Then we investigated the cell function of CLL18 and PITPNM3 through cell clone formation assay and transwell assay. The results indicated that CCL18 treatment promoted the proliferation, migration, and invasion of ICC cells. Silence of PITPNM3 reversed the effect of CCL18 on cell function. Simultaneously, we detected key protein expression of forkhead box O1 (FOXO1) and nuclear factor kappa B (NF-KB) through western blotting and found that CCL18 activated NF-KB pathway while inhibited FOXO1 pathway, the effect of which were attenuated by silence of PITPNM3. Finally, we confirmed which pathway affected the cell function using inhibitor of FOXO1 (AS1842856) and activator of NF-KB (Asatone). The results showed that AS1842856, not Asatone, relieved the inhibitory effect of si-PITPNM3 on the cell function of CCL18. In short, CCL18 treatment activated PITPNM3 to promote the proliferation, migration, and invasion of ICC via FOXO1 signaling pathway. These results provided a new insight for the diagnosis and therapy of ICC.

Myocardial infarction occurs due to insufficient (ischemia) blood supply to heart for long time; plasmacytoma variant translocation 1 (PVT1) is a long non-coding RNAs (lncRNAs) involved in the pathogenesis of various diseases, including heart disease; However, few studies have explored its role. The present study evaluated the effects of lncRNA PVT1 on hypoxic rat H9c2 cells.