Auraptene Enhances AMP-Activated Protein Kinase Phosphorylation and Thereby Inhibits the Proliferation, Migration and Expression of Androgen Receptors and Prostate-Specific Antigens in Prostate Cancer Cells.

Citrus hassaku extract reportedly activates AMPK. Because this extract contains an abundance of auraptene, we investigated whether pure auraptene activates AMPK and inhibits proliferation using prostate cancer cell lines. Indeed, auraptene inhibited the proliferation and migration of LNCaP cells and induced phosphorylation of AMPK or its downstream ACC in LNCaP, PC3, and HEK-293 cells, but not in DU145 cells not expressing LKB1. In addition, the mTOR-S6K pathway, located downstream from activated AMPK, was also markedly suppressed by auraptene treatment. Importantly, it was shown that auraptene reduced androgen receptor (AR) and prostate-specific antigen (PSA) expressions at both the protein and the mRNA level. This auraptene-induced downregulation of PSA was partially but significantly reversed by treatment with AMPK siRNA or the AMPK inhibitor compound C, suggesting AMPK activation to, at least partially, be causative. Finally, in DU145 cells lacking the LKB1 gene, exogenously induced LKB1 expression restored AMPK phosphorylation by auraptene, indicating the essential role of LKB1. In summary, auraptene is a potent AMPK activator that acts by elevating the AMP/ATP ratio, thereby potentially suppressing prostate cancer progression, via at least three molecular mechanisms, including suppression of the mTOR-S6K pathway, reduced lipid synthesis, and AR downregulation caused by AMPK activation.

Development of a simple high-performance liquid chromatography-ultraviolet method for sotorasib quantification in human plasma: Implications for therapeutic drug monitoring.

Sotorasib, an oral small-molecule inhibitor, reportedly exerts promising activity against Kirsten rat sarcoma viral oncogene homolog (KRAS)-mutant tumors. However, the currently administered dose may fail to represent the optimal dose based on the therapeutic efficacy. Herein, we developed a simple and sensitive method using high-performance liquid chromatography with ultraviolet (HPLC-UV) to measure the sotorasib concentration in human plasma. The sotorasib calibration curve exhibited linearity across the concentration range of 0.10-20.0 µg/mL (r2 = 0.9999). The coefficients of intra- and inter-day validation ranged between 0.79-9.75% and 3.01-6.13%, respectively. The assay accuracy ranged between -3.14 and 5.18%, with > 98.5% recovery. Subsequently, we applied the developed method to estimate sotorasib concentrations in a patient with KRAS G12C-mutated non-small cell lung cancer. We anticipate that our HPLC-UV method will be valuable for assessing the safety and efficacy of sotorasib in larger patient cohorts.

Hepatic Pin1 Expression, Particularly in Nuclei, Is Increased in NASH Patients in Accordance with Evidence of the Role of Pin1 in Lipid Accumulation Shown in Hepatoma Cell Lines.

Our previous studies using rodent models have suggested an essential role for Pin1 in the pathogenesis of non-alcoholic steatohepatitis (NASH). In addition, interestingly, serum Pin1 elevation has been reported in NASH patients. However, no studies have as yet examined the Pin1 expression level in human NASH livers. To clarify this issue, we investigated the expression level and subcellular distribution of Pin1 in liver specimens obtained using needle-biopsy samples from patients with NASH and healthy liver donors. Immunostaining using anti-Pin1 antibody revealed the Pin1 expression level to be significantly higher, particularly in nuclei, in the livers of NASH patients than those of healthy donors. In the samples from patients with NASH, the amount of nuclear Pin1 was revealed to be negatively related to serum alanine aminotransferase (ALT), while tendencies to be associated with other serum parameters such as aspartate aminotransferase (AST) and platelet number were noted but did not reach statistical significance. Such unclear results and the lack of a significant relationship might well be attributable to our small number of NASH liver samples (n = 8). Moreover, in vitro, it was shown that addition of free fatty acids to medium induced lipid accumulation in human hepatoma HepG2 and Huh7 cells, accompanied with marked increases in nuclear Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1), in accordance with the aforementioned observations in human NASH livers. In contrast, suppression of Pin1 gene expression using siRNAs attenuated the free fatty acid-induced lipid accumulation in Huh7 cells. Taken together, these observations strongly suggest that increased expression of Pin1, particularly in hepatic nuclei, contributes to the pathogenesis of NASH with lipid accumulation.

A pilot study of diabetic retinopathy in a porcine model of maturity onset diabetes of the young type 3 (MODY3).

Diabetic retinopathy (DR) is a leading cause of blindness in the working population. Because novel therapeutic intervention require testing, there is an urgent need for reliable animal models that faithfully replicate DR. Pig eyes have many similarities to human eyes anatomically and physiologically. Thus, attempts have been made to establish porcine models of DR by surgical, pharmaceutical or genetical induction of insulin deficiency, and dietary intervention. A previous study reported a transgenic pig model of maturity onset diabetes of the young type 3 (MODY3) developed signs of severe DR such as hemorrhage and proliferative tissue at the surface of the retina. However, the course of development of DR has not been studied in detail in this model. The purpose of this study was to investigate the early phase of DR in a MODY3. MODY3 and wild-type (WT) pigs underwent fundus photography and fluorescein angiogram (FA) before they developed cataracts. Animals were euthanized at age 1, 4, 7, and 10 months. Whole-mount retina and 10-µm thick paraffinized sections were stained with isolectin B4, and vessel density was determined by MATLAB software. At 4 and 7 months, retinal arterioles were immediately cannulated, and vasomotor action was measured by incubation with bradykinin and sodium nitroprusside. In the MODY3 pigs, fasting blood sugar levels gradually increased up to 500 mg/dL. Vascular tortuosity and yellowish spindle-shaped lesions were confirmed in MODY3 pigs at the age of 7 months; however, no microaneurysms were detected on FA. Compared with age-matched WT pigs, MODY3 pigs showed a significant decrease in blood vessel density in the intermediate and deep vascular plexus at 4 and 7 months of age and a slight decrease in capillary density in the superficial vascular plexus at 7 months of age. In MODY3 pigs, electron microscopy revealed thickening of the capillary basement membrane and leukostasis in the major blood vessels at 10 months of age. Bradykinin-induced dilation of retinal arterioles was diminished in MODY3 pigs as early as 7 months of age. Within 1 year after birth, MODY3 pigs show all typical early vascular lesions of diabetes except for microaneurysm formation. This pilot study suggests that the MODY3 pigs may serve as a suitable DR model to test effects of newly developed compounds on DR.

Synergistic deterioration of prognosis associated with decreased grip strength and hyporesponse to erythropoiesis-stimulating agents in patients undergoing hemodialysis.

INTRODUCTION: We examined the combined effect of erythropoietin (EPO) hyporesponsiveness and low handgrip strength (HGS) on the prognosis of patients undergoing hemodialysis (HD). METHODS: We recruited patients with chronic kidney disease (CKD) Stage 5, who were undergoing HD at our dialysis clinic between January 2015 and March 2015 (n = 182). Patients of ≥20 years of age and who had been undergoing HD for ≧3 months at enrollment were eligible for inclusion. Seven patients treated with epoetin-ß pegol were excluded. First, the erythropoietin resistance index (ERI) and HGS were measured. The patients were stratified by the ERI of 9.44 (U/kg/week/g/dL), and by the HGS of 28 kg for men and 18 kg for women. We then observed death and cardiovascular disease (CVD), composite endpoint (deaths or CVD) for a median of 2 years. RESULTS: A total of 175 patients (male, n = 122; female, n = 53; age, 34-92 years) were included in the analysis. During the observation period of 24 months, 57 events (14 deaths and 43 CVD) were observed. High ERI and low HGS were associated with a high incidence of endpoints compared to low ERI and high HGS. Among the four groups classified by ERI and HGS values, the highest risk group was the high ERI/low HGS group (HR: 4.20 95% CI 2.12-8.33). CONCLUSIONS: EPO hyporesponsiveness combined with low HGS were found to be significant predictors of a poor outcome, and the synergistic effects of the two factors had stronger predictive ability than either single factor.

Involvement of neuronal and muscular Trk-fused gene (TFG) defects in the development of neurodegenerative diseases.

Trk-fused gene (TFG) mutations have been identified in patients with several neurodegenerative diseases. In this study, we attempted to clarify the effects of TFG deletions in motor neurons and in muscle fibers, using tissue-specific TFG knockout (vMNTFG KO and MUSTFG KO) mice. vMNTFG KO, generated by crossing TFG floxed with VAChT-Cre, showed deterioration of motor function and muscle atrophy especially in slow-twitch soleus muscle, in line with the predominant Cre expression in slow-twitch fatigue-resistant (S) and fast-twitch fatigue-resistant (FR) motor neurons. Consistently, denervation of the neuromuscular junction (NMJ) was apparent in the soleus, but not in the extensor digitorum longus, muscle. Muscle TFG expressions were significantly downregulated in vMNTFG KO, presumably due to decreased muscle IGF-1 concentrations. However, interestingly, MUSTFG KO mice showed no apparent impairment of muscle movements, though a denervation marker, AChRγ, was elevated and Agrin-induced AChR clustering in C2C12 myotubes was inhibited. Our results clarify that loss of motor neuron TFG is sufficient for the occurrence of NMJ degeneration and muscle atrophy, though lack of muscle TFG may exert an additional effect. Reduced muscle TFG, also observed in aged mice, might be involved in age-related NMJ degeneration, and this issue merits further study.

The Effect of Sodium-Dependent Glucose Cotransporter 2 Inhibitor Tofogliflozin on Neurovascular Coupling in the Retina in Type 2 Diabetic Mice.

We investigated the effect of tofogliflozin, a sodium-dependent glucose cotransporter 2 inhibitor (SGLT2i), on retinal blood flow dysregulation, neural retinal dysfunction, and the impaired neurovascular coupling in type 2 diabetic mice. Tofogliflozin was added to mouse chow to deliver 5 mg/kg/day and 6-week-old mice were fed for 8 weeks. The longitudinal changes in the retinal neuronal function and blood flow responses to systemic hyperoxia and flicker stimulation were evaluated every 2 weeks in diabetic db/db mice that received tofogliflozin (n =6) or placebo (n = 6) from 8 to 14 weeks of age. We also evaluated glial activation and vascular endothelial growth factor (VEGF) expression by immunofluorescence. Tofogliflozin treatment caused a sustained decrease in blood glucose in db/db mice from 8 weeks of the treatment. In tofogliflozin-treated db/db mice, both responses improved from 8 to 14 weeks of age, compared with vehicle-treated diabetic mice. Subsequently, the electroretinography implicit time for the oscillatory potential was significantly improved in SGLT2i-treated db/db mice. The systemic tofogliflozin treatment prevented the activation of glial fibrillary acidic protein and VEGF protein expression, as detected by immunofluorescence. Our results suggest that glycemic control with tofogliflozin significantly improved the impaired retinal neurovascular coupling in type 2 diabetic mice with the inhibition of retinal glial activation.

Synergistic association of the copper/zinc ratio under inflammatory conditions with diabetic kidney disease in patients with type 2 diabetes: The Asahi Diabetes Complications Study.

AIMS/INTRODUCTION: We aimed to study the relationships among the copper (Cu)/zinc (Zn) ratio, inflammatory biomarkers, and the prevalence of diabetic kidney disease (DKD) in patients with type 2 diabetes. MATERIALS AND METHODS: A cross-sectional study was performed on 651 patients with type 2 diabetes. DKD was defined as a urinary albumin-to-creatinine ratio of ≥30 mg/g creatinine and/or an estimated glomerular filtration rate using cystatin C of < 60 mL/min/1.73 m2 . Areas under the curves (AUCs), cutoff values, and thresholds for detecting DKD were determined for the Cu/Zn ratio, soluble tumor necrosis factor-α receptor 1 (sTNFαR1), and high-sensitivity C-reactive protein (hsCRP). Patients were categorized by each cutoff value of sTNFαR1 and the Cu/Zn ratio. Odds ratios (ORs) and biological interactions for the prevalence of DKD were determined. RESULTS: DKD was identified in 220 patients. AUC/optimal cutoff values were 0.777/1300 pg/mL for sTNFαR1, 0.603/1.1648 for the Cu/Zn ratio, and 0.582/305 ng/mL for hsCRP. The ORs for DKD were higher, but not significantly, in the sTNFαR1 < 1300 and Cu/Zn ≥ 1.1648 group, significantly higher in the sTNFαR1 ≥ 1300 and Cu/Zn < 1.1648 group (P < 0.0001), and further synergistically elevated in the sTNFαR1 ≥ 1300 and Cu/Zn ≥ 1.1648 group (P < 0.0001) compared with the sTNFαR1 < 1300 and Cu/Zn < 1.1648 group after multivariable adjustment. Levels of sTNFαR1 were significantly higher in the sTNFαR1 ≥ 1300 and Cu/Zn ≥ 1.1648 group than in the sTNFαR1 ≥ 1300 and Cu/Zn < 1.1648 group (P = 0.0006). CONCLUSIONS: Under an inflammatory initiation signal of elevated serum sTNFαR1 levels, an increase in the Cu/Zn ratio may further exacerbate inflammation and is synergistically associated with a high prevalence of DKD in patients with type 2 diabetes.

Pathological Role of Pin1 in the Development of DSS-Induced Colitis.

Inflammatory bowel diseases (IBDs) are serious disorders of which the etiologies are not, as yet, fully understood. In this study, Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1) protein was shown to be dramatically upregulated in the colons of dextran sodium sulfate (DSS)-induced ulcerative colitis model mice. Interestingly, Pin1 knockout (KO) mice exhibited significant attenuation of DSS-induced colitis compared to wild-type (WT) mice, based on various parameters, including body weight, colon length, microscopic observation of the intestinal mucosa, inflammatory cytokine expression, and cleaved caspase-3. In addition, a role of Pin1 in inflammation was suggested because the percentage of M1-type macrophages in the colon was decreased in the Pin1 KO mice while that of M2-type macrophages was increased. Moreover, Pin1 KO mice showed downregulation of both Il17 and Il23a expression in the colon, both of which have been implicated in the development of colitis. Finally, oral administration of Pin1 inhibitor partially but significantly prevented DSS-induced colitis in mice, raising the possibility of Pin1 inhibitors serving as therapeutic agents for IBD.

Carnosic Acid and Carnosol Activate AMPK, Suppress Expressions of Gluconeogenic and Lipogenic Genes, and Inhibit Proliferation of HepG2 Cells.

Carnosic acid (CA), carnosol (CL) and rosmarinic acid (RA), components of the herb rosemary, reportedly exert favorable metabolic actions. This study showed that both CA and CL, but not RA, induce significant phosphorylation of AMP-dependent kinase (AMPK) and its downstream acetyl-CoA carboxylase 1 (ACC1) in HepG2 hepatoma cells. Glucose-6-phosphatase (G6PC) and phosphoenolpyruvate carboxykinase 1 (PCK1), rate-limiting enzymes of hepatic gluconeogenesis, are upregulated by forskolin stimulation, and this upregulation was suppressed when incubated with CA or CL. Similarly, a forskolin-induced increase in CRE transcriptional activity involved in G6PC and PCK1 regulations was also stymied when incubated with CA or CL. In addition, mRNA levels of ACC1, fatty acid synthase (FAS) and sterol regulatory element-binding protein 1c (SREBP-1c) were significantly reduced when incubated with CA or CL. Finally, it was shown that CA and CL suppressed cell proliferation and reduced cell viability, possibly as a result of AMPK activation. These findings raise the possibility that CA and CL exert a protective effect against diabetes and fatty liver disease, as well as subsequent cases of hepatoma.

Development of Pin1 Inhibitors and their Potential as Therapeutic Agents.

The prolyl isomerase Pin1 is a unique enzyme, which isomerizes the cis-trans conformation between pSer/pThr and proline and thereby regulates the function, stability and/or subcellular distribution of its target proteins. Such regulations by Pin1 are involved in numerous physiological functions as well as the pathogenic mechanisms underlying various diseases. Notably, Pin1 deficiency or inactivation is a potential cause of Alzheimer's disease, since Pin1 induces the degradation of Tau. In contrast, Pin1 overexpression is highly correlated with the degree of malignancy of cancers, as Pin1 controls a number of oncogenes and tumor suppressors. Accordingly, Pin1 inhibitors as anti-cancer drugs have been developed. Interestingly, recent intensive studies have demonstrated Pin1 to be responsible for the onset or development of nonalcoholic steatosis, obesity, atherosclerosis, lung fibrosis, heart failure and so on, all of which have been experimentally induced in Pin1 deficient mice. In this review, we discuss the possible applications of Pin1 inhibitors to a variety of diseases including malignant tumors and also introduce the recent advances in Pin1 inhibitor research, which have been reported.

Prolyl isomerase Pin1 in metabolic reprogramming of cancer cells.

Pin1 is one member of a group consisting of three prolyl isomerases. Pin1 interacts with the motif containing phospho-Ser/Thr-Pro of substrates and enhances cis-trans isomerization of peptide bonds, thereby controlling the functions of these substrates. Importantly, the Pin1 expression level is highly upregulated in most cancer cells and correlates with malignant properties, and thereby with poor outcomes. In addition, Pin1 was revealed to promote the functions of multiple oncogenes and to abrogate tumor suppressors. Accordingly, Pin1 is well recognized as a master regulator of malignant processes. Recent studies have shown that Pin1 also binds to a variety of metabolic regulators, such as AMP-activated protein kinase, acetyl CoA carboxylase and pyruvate kinase2, indicating Pin1 to have major impacts on lipid and glucose metabolism in cancer cells. In this review, we focus on the roles of Pin1 in metabolic reprogramming, such as "Warburg effects", of cancer cells. Our aim is to introduce these important roles of Pin1, as well as to present evidence supporting the possibility of Pin1 inhibition as a novel anti-cancer strategy.

Xanthine Oxidase Inhibitor Febuxostat Exerts an Anti-Inflammatory Action and Protects against Diabetic Nephropathy Development in KK-Ay Obese Diabetic Mice.

Hyperuricemia has been recognized as a risk factor for insulin resistance as well as one of the factors leading to diabetic kidney disease (DKD). Since DKD is the most common cause of end-stage renal disease, we investigated whether febuxostat, a xanthine oxidase (XO) inhibitor, exerts a protective effect against the development of DKD. We used KK-Ay mice, an established obese diabetic rodent model. Eight-week-old KK-Ay mice were provided drinking water with or without febuxostat (15 µg/mL) for 12 weeks and then subjected to experimentation. Urine albumin secretion and degrees of glomerular injury judged by microscopic observations were markedly higher in KK-Ay than in control lean mice. These elevations were significantly normalized by febuxostat treatment. On the other hand, body weights and high serum glucose concentrations and glycated albumin levels of KK-Ay mice were not affected by febuxostat treatment, despite glucose tolerance and insulin tolerance tests having revealed febuxostat significantly improved insulin sensitivity and glucose tolerance. Interestingly, the IL-1ß, IL-6, MCP-1, and ICAM-1 mRNA levels, which were increased in KK-Ay mouse kidneys as compared with normal controls, were suppressed by febuxostat administration. These data indicate a protective effect of XO inhibitors against the development of DKD, and the underlying mechanism likely involves inflammation suppression which is independent of hyperglycemia amelioration.

Possible involvement of normalized Pin1 expression level and AMPK activation in the molecular mechanisms underlying renal protective effects of SGLT2 inhibitors in mice.

BACKGROUND: Recently, clinical studies have shown the protective effects of sodium glucose co-transporter2 (SGLT2) inhibitors against progression of diabetic nephropathy, but the underlying molecular mechanisms remain unclear. METHODS: Diabetic mice were prepared by injecting nicotinamide and streptozotocin, followed by high-sucrose diet feeding (NA/STZ/Suc mice). The SGLT2 inhibitor canagliflozin was administered as a 0.03% (w/w) mixture in the diet for 4 weeks. Then, various parameters and effects of canagliflozin on diabetic nephropathy were investigated. RESULTS: Canagliflozin administration to NA/STZ/Suc mice normalized hyperglycemia as well as elevated renal mRNA of collagen 1a1, 1a2, CTGF, TNFα and MCP-1. Microscopic observation revealed reduced fibrotic deposition in the kidneys of canagliflozin-treated NA/STZ/Suc mice. Interestingly, the protein level of Pin1, reportedly involved in the inflammation and fibrosis affecting several tissues, was markedly increased in the NA/STZ/Suc mouse kidney, but this was normalized with canagliflozin treatment. The cells showing increased Pin1 expression in the kidney were mainly mesangial cells, along with podocytes, based on immunohistochemical analysis. Furthermore, it was revealed that canagliflozin induced AMP-activated kinase (AMPK) activation concentration-dependently in CRL1927 mesangial as well as THP-1 macrophage cell lines. AMPK activation was speculated to suppress mesangial cell proliferation and exert anti-inflammatory effects in hematopoietic cells. CONCLUSION: Therefore, we can reasonably suggest that normalized Pin1 expression and AMPK activation contribute to the molecular mechanisms underlying SGLT2 inhibitor-induced suppression of diabetic nephropathy, possibly at least in part by reducing inflammation and fibrotic change.

Prolyl isomerase Pin1 binds to and stabilizes acetyl CoA carboxylase 1 protein, thereby supporting cancer cell proliferation.

The prolyl isomerase Pin1 expression level is reportedly increased in most malignant tissues and correlates with poor outcomes. On the other hand, acetyl CoA carboxylase 1 (ACC1), the rate limiting enzyme of lipogenesis is also abundantly expressed in cancer cells, to satisfy the demand for the fatty acids (FAs) needed for rapid cell proliferation. We found Pin1 expression levels to correlate positively with ACC1 levels in human prostate cancers, and we focused on the relationship between Pin1 and ACC1. Notably, it was demonstrated that Pin1 associates with ACC1 but not with acetyl CoA carboxylase 2 (ACC2) in the overexpression system as well as endogenously in the prostate cancer cell line DU145. This association is mediated by the WW domain in the Pin1 and C-terminal domains of ACC1. Interestingly, Pin1 deficiency or treatment with Pin1 siRNA or the inhibitor juglone markedly reduced ACC1 protein expression without affecting its mRNA level, while Pin1 overexpression increased the ACC1 protein level. In addition, chloroquine treatment restored the levels of ACC1 protein reduced by Pin1 siRNA treatment, indicating that Pin1 suppressed ACC1 degradation through the lysosomal pathway. In brief, we have concluded that Pin1 leads to the stabilization of and increases in ACC1. Therefore, it is likely that the growth-enhancing effect of Pin1 in cancer cells is mediated at least partially by the stabilization of ACC1 protein, corresponding to the well-known potential of Pin1 inhibitors as anti-cancer drugs.

Alpha-Blockers As Colorectal Cancer Chemopreventive: Findings from a Case-Control Study, Human Cell Cultures, and In Vivo Preclinical Testing.

A retrospective case-controlled analysis was performed to identify drug candidates in the current use that may prevent colorectal cancer, outside of aspirin. A total of 37,510 patients aged ≥20 years were assessed to identify subjects who had been diagnosed with colorectal cancer by colonoscopy without a previous diagnosis of colorectal cancer, inflammatory bowel disease (IBD), or gastrointestinal symptoms; 1,560 patients were identified who were diagnosed with colorectal cancer by colonoscopy. The patients with colorectal cancer were matched with 1,560 age, gender, family history of colorectal cancer and comorbidity-matched control patients who were not diagnosed with colorectal cancer at colonoscopy. The medication histories were compared between the two groups. Next, candidate drugs that were more frequently used by the control patients were selected and their effects on human colorectal cancer cell lines in vitro and an inflammation-induced mouse model of colorectal cancer were tested. Putative colorectal cancer preventative agents were identified, including aspirin, vitamin D, vitamin B, vitamin C, vitamin E, xanthine oxidase inhibitor, alpha-blockers, angiotensin receptor blocker, nateglinide, probiotics, thienopyridine, folic acid, nitrovasodilators, bisphosphonates, calcium channel blockers, steroids, and statins (P < 0.05). Alpha-blockers and xanthine oxidase inhibitors were selected for further study because these agents have not been analyzed previously as factors that may affect colorectal cancer outcomes. In vitro doxazosin (alpha-blocker), but not febuxostat (xanthine oxidase inhibitor), suppressed the proliferation of human colorectal cancer cells. Doxazosin also decreased tumorigenesis in an AOM/DSS mouse colorectal cancer model. Alpha-blockers may prevent colorectal cancer.

Treatment of two mitochondrial disease patients with a combination of febuxostat and inosine that enhances cellular ATP.

Since mitochondria are energy-generating micro-organisms, most of the disorders in patients with mitochondrial diseases (mt-disease) are considered secondary to defects in ATP synthesis, although some other factors such as reactive oxygen species may be involved. A simultaneous oral administration of febuxostat and inosine was reported to elevate both hypoxanthine and ATP levels in peripheral blood. Based on those results, we attempted co-administration of febuxostat and inosine in two patients with mitochondrial disease: one patient with mitochondrial cardiomyopathy and the other patient with mitochondrial diabetes. In the former case, brain natriuretic peptide (BNP), which is a specific marker for heart failure, was decreased by 31%, and in the latter case, the insulinogenic index increased 3.1 times, suggesting the favorable action of the treatment. Considering that there is no effective treatment available for this disorder, the present therapy may be quite useful for the management of patients with mitochondrial diseases.

Roles of Gut-Derived Secretory Factors in the Pathogenesis of Non-Alcoholic Fatty Liver Disease and Their Possible Clinical Applications.

The rising prevalence of non-alcoholic fatty liver disease (NAFLD) parallels the global increase in the number of people diagnosed with obesity and metabolic syndrome. The gut-liver axis (GLA) plays an important role in the pathogenesis of NAFLD/non-alcoholic steatohepatitis (NASH). In this review, we discuss the clinical significance and underlying mechanisms of action of gut-derived secretory factors in NAFLD/NASH, focusing on recent human studies. Several studies have identified potential causal associations between gut-derived secretory factors and NAFLD/NASH, as well as the underlying mechanisms. The effects of gut-derived hormone-associated drugs, such as glucagon-like peptide-1 analog and recombinant variant of fibroblast growth factor 19, and other new treatment strategies for NAFLD/NASH have also been reported. A growing body of evidence highlights the role of GLA in the pathogenesis of NAFLD/NASH. Larger and longitudinal studies as well as translational research are expected to provide additional insights into the role of gut-derived secretory factors in the pathogenesis of NAFLD/NASH, possibly providing novel markers and therapeutic targets in patients with NAFLD/NASH.

Protective Effect of Sex Hormone-Binding Globulin against Metabolic Syndrome: In Vitro Evidence Showing Anti-Inflammatory and Lipolytic Effects on Adipocytes and Macrophages.

Sex hormone-binding globulin (SHBG) is a serum protein released mainly by the liver, and a low serum level correlates with a risk for metabolic syndrome including diabetes, obesity, and cardiovascular events. However, the underlying molecular mechanism(s) linking SHBG and metabolic syndrome remains unknown. In this study, using adipocytes and macrophages, we focused on the in vitro effects of SHBG on inflammation as well as lipid metabolism. Incubation with 20 nM SHBG markedly suppressed lipopolysaccharide- (LPS-) induced inflammatory cytokines, such as MCP-1, TNFα, and IL-6 in adipocytes and macrophages, along with phosphorylations of JNK and ERK. Anti-inflammatory effects were also observed in 3T3-L1 adipocytes cocultured with LPS-stimulated macrophages. In addition, SHBG treatment for 18 hrs or longer significantly induced the lipid degradation of differentiated 3T3-L1 cells, with alterations in its corresponding gene and protein levels. Notably, these effects of SHBG were not altered by coaddition of large amounts of testosterone or estradiol. In conclusion, SHBG suppresses inflammation and lipid accumulation in macrophages and adipocytes, which might be among the mechanisms underlying the protective effect of SHBG, that is, its actions which reduce the incidence of metabolic syndrome.

The SGLT2 Inhibitor Luseogliflozin Rapidly Normalizes Aortic mRNA Levels of Inflammation-Related but Not Lipid-Metabolism-Related Genes and Suppresses Atherosclerosis in Diabetic ApoE KO Mice.

Recent clinical studies have revealed the treatment of diabetic patients with sodium glucose co-transporter2 (SGLT2) inhibitors to reduce the incidence of cardiovascular events. Using nicotinamide and streptozotocin (NA/STZ) -treated ApoE KO mice, we investigated the effects of short-term (seven days) treatment with the SGLT2 inhibitor luseogliflozin on mRNA levels related to atherosclerosis in the aorta, as well as examining the long-term (six months) effects on atherosclerosis development. Eight-week-old ApoE KO mice were treated with NA/STZ to induce diabetes mellitus, and then divided into two groups, either untreated, or treated with luseogliflozin. Seven days after the initiation of luseogliflozin administration, atherosclerosis-related mRNA levels in the aorta were compared among four groups; i.e., wild type C57/BL6J, native ApoE KO, and NA/STZ-treated ApoE KO mice, with or without luseogliflozin. Short-term luseogliflozin treatment normalized the expression of inflammation-related genes such as F4/80, TNFα, IL-1ß, IL-6, ICAM-1, PECAM-1, MMP2 and MMP9 in the NA/STZ-treated ApoE KO mice, which showed marked elevations as compared with untreated ApoE KO mice. In contrast, lipid metabolism-related genes were generally unaffected by luseogliflozin treatment. Furthermore, after six-month treatment with luseogliflozin, in contrast to the severe and widely distributed atherosclerotic changes in the aortas of NA/STZ-treated ApoE KO mice, luseogliflozin treatment markedly attenuated the progression of atherosclerosis, without affecting serum lipid parameters such as high density lipoprotein, low density lipoprotein and triglyceride levels. Given that luseogliflozin normalized the aortic mRNA levels of inflammation-related, but not lipid-related, genes soon after the initiation of treatment, it is not unreasonable to speculate that the anti-atherosclerotic effect of this SGLT2 inhibitor emerges rapidly, possibly via the prevention of inflammation rather than of hyperlipidemia.