11ß-HSD1 inhibition in men mitigates prednisolone-induced adverse effects in a proof-of-concept randomised double-blind placebo-controlled trial.

Glucocorticoids prescribed to limit inflammation, have significant adverse effects. As 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) regenerates active glucocorticoid, we investigated whether 11ß-HSD1 inhibition with AZD4017 could mitigate adverse glucocorticoid effects without compromising their anti-inflammatory actions. We conducted a proof-of-concept, randomized, double-blind, placebo-controlled study at Research Unit, Churchill Hospital, Oxford, UK (NCT03111810). 32 healthy male volunteers were randomized to AZD4017 or placebo, alongside prednisolone treatment. Although the primary endpoint of the study (change in glucose disposal during a two-step hyperinsulinemic, normoglycemic clamp) wasn't met, hepatic insulin sensitivity worsened in the placebo-treated but not in the AZD4017-treated group. Protective effects of AZD4017 on markers of lipid metabolism and bone turnover were observed. Night-time blood pressure was higher in the placebo-treated but not in the AZD4017-treated group. Urinary (5aTHF+THF)/THE ratio was lower in the AZD4017-treated but remained the same in the placebo-treated group. Most anti-inflammatory actions of prednisolone persisted with AZD4017 co-treatment. Four adverse events were reported with AZD4017 and no serious adverse events. Here we show that co-administration of AZD4017 with prednisolone in men is a potential strategy to limit adverse glucocorticoid effects.

AKR1D1 knockout mice develop a sex-dependent metabolic phenotype.

Steroid 5ß-reductase (AKR1D1) plays important role in hepatic bile acid synthesis and glucocorticoid clearance. Bile acids and glucocorticoids are potent metabolic regulators, but whether AKR1D1 controls metabolic phenotype in vivo is unknown. Akr1d1-/- mice were generated on a C57BL/6 background. Liquid chromatography/mass spectrometry, metabolomic and transcriptomic approaches were used to determine effects on glucocorticoid and bile acid homeostasis. Metabolic phenotypes including body weight and composition, lipid homeostasis, glucose tolerance and insulin tolerance were evaluated. Molecular changes were assessed by RNA-Seq and Western blotting. Male Akr1d1-/- mice were challenged with a high fat diet (60% kcal from fat) for 20 weeks. Akr1d1-/- mice had a sex-specific metabolic phenotype. At 30 weeks of age, male, but not female, Akr1d1-/- mice were more insulin tolerant and had reduced lipid accumulation in the liver and adipose tissue yet had hypertriglyceridemia and increased intramuscular triacylglycerol. This phenotype was associated with sexually dimorphic changes in bile acid metabolism and composition but without overt effects on circulating glucocorticoid levels or glucocorticoid-regulated gene expression in the liver. Male Akr1d1-/- mice were not protected against diet-induced obesity and insulin resistance. In conclusion, this study shows that AKR1D1 controls bile acid homeostasis in vivo and that altering its activity can affect insulin tolerance and lipid homeostasis in a sex-dependent manner.

Acute intermittent hypoxia drives hepatic de novo lipogenesis in humans and rodents.

Background and aims: Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver condition. It is tightly associated with an adverse metabolic phenotype (including obesity and type 2 diabetes) as well as with obstructive sleep apnoea (OSA) of which intermittent hypoxia is a critical component. Hepatic de novo lipogenesis (DNL) is a significant contributor to hepatic lipid content and the pathogenesis of NAFLD and has been proposed as a key pathway to target in the development of pharmacotherapies to treat NAFLD. Our aim is to use experimental models to investigate the impact of hypoxia on hepatic lipid metabolism independent of obesity and metabolic disease. Methods: Human and rodent studies incorporating stable isotopes and hyperinsulinaemic euglycaemic clamp studies were performed to assess the regulation of DNL and broader metabolic phenotype by intermittent hypoxia. Cell-based studies, including pharmacological and genetic manipulation of hypoxia-inducible factors (HIF), were used to examine the underlying mechanisms. Results: Hepatic DNL increased in response to acute intermittent hypoxia in humans, without alteration in glucose production or disposal. These observations were endorsed in a prolonged model of intermittent hypoxia in rodents using stable isotopic assessment of lipid metabolism. Changes in DNL were paralleled by increases in hepatic gene expression of acetyl CoA carboxylase 1 and fatty acid synthase. In human hepatoma cell lines, hypoxia increased both DNL and fatty acid uptake through HIF-1α and -2α dependent mechanisms. Conclusions: These studies provide robust evidence linking intermittent hypoxia and the regulation of DNL in both acute and sustained in vivo models of intermittent hypoxia, providing an important mechanistic link between hypoxia and NAFLD.

Metformin maintains intrahepatic triglyceride content through increased hepatic de novo lipogenesis.

OBJECTIVE: Metformin is a first-line pharmacotherapy in the treatment of type 2 diabetes, a condition closely associated with non-alcoholic fatty liver disease (NAFLD). Although metformin promotes weight loss and improves insulin sensitivity, its effect on intrahepatic triglyceride (IHTG) remains unclear. We investigated the effect of metformin on IHTG, hepatic de novo lipogenesis (DNL), and fatty acid (FA) oxidation in vivo in humans. DESIGN AND METHODS: Metabolic investigations, using stable-isotope tracers, were performed in ten insulin-resistant, overweight/obese human participants with NAFLD who were treatment naïve before and after 12 weeks of metformin treatment. The effect of metformin on markers of s.c. adipose tissue FA metabolism and function, along with the plasma metabolome, was investigated. RESULTS: Twelve weeks of treatment with metformin resulted in a significant reduction in body weight and improved insulin sensitivity, but IHTG content and FA oxidation remained unchanged. Metformin treatment was associated with a significant decrease in VLDL-triglyceride (TG) concentrations and a significant increase in the relative contribution of DNL-derived FAs to VLDL-TG. There were subtle and relatively few changes in s.c. adipose tissue FA metabolism and the plasma metabolome with metformin treatment. CONCLUSIONS: We demonstrate the mechanisms of action of metformin whereby it improves insulin sensitivity and promotes weight loss, without improvement in IHTG; these observations are partly explained through increased hepatic DNL and a lack of change in FA oxidation.

Sodium-glucose cotransporter 2 inhibition does not reduce hepatic steatosis in overweight, insulin-resistant patients without type 2 diabetes.

BACKGROUND AND AIM: Non-alcoholic fatty liver disease (NAFLD) is rapidly becoming the leading indication for liver transplant and is associated with increased cardiovascular and liver mortality, yet there are no licensed therapies. Sodium-glucose cotransporter 2 (SGLT2) inhibitors are widely used for their glucose-lowering effects in patients with type 2 diabetes (T2D). Preclinical models have suggested a beneficial impact on NAFLD, but clinical data are limited, and there are currently no data on patients without T2D. We aimed to investigate the impact of SGLT2 inhibition on NAFLD in overweight, nondiabetic patients and establish the effect these agents may have on the processes that regulate hepatic steatosis in vivo. METHODS: We conducted an open-label, experimental medicine pilot study on insulin-resistant overweight/obese individuals (n = 10) using gold-standard noninvasive assessments of NAFLD phenotype, including magnetic resonance spectroscopy, two-step hyperinsulinemic euglycemic clamps, and stable isotope tracers to assess lipid and glucose metabolism. Investigations were performed before and after a 12-week treatment with the SGLT2 inhibitor, dapagliflozin. RESULTS: Despite a body weight reduction of 4.4 kg, hepatic steatosis was unchanged following treatment. Hepatic glucose production increased, and there was impairment of glucose disposal during the low-dose insulin infusion. Although circulating, nonesterified, fatty acid levels did not change, the ability of insulin to suppress lipolysis was reduced. CONCLUSIONS: SGLT2 inhibition for 12 weeks does not improve hepatic steatosis in patients without T2D. Additional studies in patients with established T2D or impairments of fasting or postprandial glucose homeostasis are needed to determine whether SGLT2 inhibition represents a viable therapeutic strategy for NAFLD. (http://clinicaltrials.gov Number NCT02696941).

Co-administration of 5α-reductase Inhibitors Worsens the Adverse Metabolic Effects of Prescribed Glucocorticoids.

CONTEXT: Glucocorticoids (GCs) are commonly prescribed, but their use is associated with adverse metabolic effects. 5α-reductase inhibitors (5α-RI) are also frequently prescribed, mainly to inhibit testosterone conversion to dihydrotestosterone. However, they also prevent the inactivation of GCs. OBJECTIVE: We hypothesized that 5α-RI may worsen the adverse effects of GCs. DESIGN: Prospective, randomized study. PATIENTS: A total of 19 healthy male volunteers (age 45 ±â€…2 years; body mass index 27.1 ±â€…0.7kg/m2). INTERVENTIONS: Participants underwent metabolic assessments; 2-step hyperinsulinemic, euglycemic clamp incorporating stable isotopes, adipose tissue microdialysis, and biopsy. Participants were then randomized to either prednisolone (10 mg daily) or prednisolone (10 mg daily) plus a 5α-RI (finasteride 5 mg daily or dutasteride 0.5 mg daily) for 7 days; metabolic assessments were then repeated. MAIN OUTCOME MEASURES: Ra glucose, glucose utilization (M-value), glucose oxidation, and nonesterified fatty acids (NEFA) levels. RESULTS: Co-administration of prednisolone with a 5α-RI increased circulating prednisolone levels (482 ±â€…96 vs 761 ±â€…57 nmol/L, P = 0.029). Prednisolone alone did not alter Ra glucose (2.55 ±â€…0.34 vs 2.62 ±â€…0.19 mg/kg/minute, P = 0.86), M-value (3.2 ±â€…0.5 vs 2.7 ±â€…0.7 mg/kg/minute, P = 0.37), or glucose oxidation (0.042 ±â€…0.007 vs 0.040 ±â€…0.004 mmol/hr/kg/minute, P = 0.79). However, co-administration with a 5α-RI increased Ra glucose (2.67 ±â€…0.16 vs 3.05 ±â€…0.18 mg/kg/minute, P < 0.05) and decreased M-value (4.0 ±â€…0.5 vs 2.6 ±â€…0.4 mg/kg/minute, P < 0.05), and oxidation (0.043 ±â€…0.003 vs 0.036 ±â€…0.002 mmol/hr/kg, P < 0.01). Similarly, prednisolone did not impair insulin-mediated suppression of circulating NEFA (43.1 ±â€…28.9 vs 36.8 ±â€…14.3 µmol/L, P = 0.81), unless co-administered with a 5α-RI (49.8 ±â€…8.6 vs 88.5 ±â€…13.5 µmol/L, P < 0.01). CONCLUSIONS: We have demonstrated that 5α-RIs exacerbate the adverse effects of prednisolone. This study has significant translational implications, including the need to consider GC dose adjustments, but also the necessity for increased vigilance for the development of adverse effects.

Accurate non-invasive diagnosis and staging of non-alcoholic fatty liver disease using the urinary steroid metabolome.

BACKGROUND: The development of accurate, non-invasive markers to diagnose and stage non-alcoholic fatty liver disease (NAFLD) is critical to reduce the need for an invasive liver biopsy and to identify patients who are at the highest risk of hepatic and cardio-metabolic complications. Disruption of steroid hormone metabolic pathways has been described in patients with NAFLD. AIM(S): To assess the hypothesis that assessment of the urinary steroid metabolome may provide a novel, non-invasive biomarker strategy to stage NAFLD. METHODS: We analysed the urinary steroid metabolome in 275 subjects (121 with biopsy-proven NAFLD, 48 with alcohol-related cirrhosis and 106 controls), using gas chromatography-mass spectrometry (GC-MS) coupled with machine learning-based Generalised Matrix Learning Vector Quantisation (GMLVQ) analysis. RESULTS: Generalised Matrix Learning Vector Quantisation analysis achieved excellent separation of early (F0-F2) from advanced (F3-F4) fibrosis (AUC receiver operating characteristics [ROC]: 0.92 [0.91-0.94]). Furthermore, there was near perfect separation of controls from patients with advanced fibrotic NAFLD (AUC ROC = 0.99 [0.98-0.99]) and from those with NAFLD cirrhosis (AUC ROC = 1.0 [1.0-1.0]). This approach was also able to distinguish patients with NAFLD cirrhosis from those with alcohol-related cirrhosis (AUC ROC = 0.83 [0.81-0.85]). CONCLUSIONS: Unbiased GMLVQ analysis of the urinary steroid metabolome offers excellent potential as a non-invasive biomarker approach to stage NAFLD fibrosis as well as to screen for NAFLD. A highly sensitive and specific urinary biomarker is likely to have clinical utility both in secondary care and in the broader general population within primary care and could significantly decrease the need for liver biopsy.

Nonalcoholic Fatty Liver Disease in Adults: Current Concepts in Etiology, Outcomes, and Management.

Nonalcoholic fatty liver disease (NAFLD) is a spectrum of disease, extending from simple steatosis to inflammation and fibrosis with a significant risk for the development of cirrhosis. It is highly prevalent and is associated with significant adverse outcomes both through liver-specific morbidity and mortality but, perhaps more important, through adverse cardiovascular and metabolic outcomes. It is closely associated with type 2 diabetes and obesity, and both of these conditions drive progressive disease toward the more advanced stages. The mechanisms that govern hepatic lipid accumulation and the predisposition to inflammation and fibrosis are still not fully understood but reflect a complex interplay between metabolic target tissues including adipose and skeletal muscle, and immune and inflammatory cells. The ability to make an accurate assessment of disease stage (that relates to clinical outcome) can also be challenging. While liver biopsy is still regarded as the gold-standard investigative tool, there is an extensive literature on the search for novel noninvasive biomarkers and imaging modalities that aim to accurately reflect the stage of underlying disease. Finally, although no therapies are currently licensed for the treatment of NAFLD, there are interventions that appear to have proven efficacy in randomized controlled trials as well as an extensive emerging therapeutic landscape of new agents that target many of the fundamental pathophysiological processes that drive NAFLD. It is highly likely that over the next few years, new treatments with a specific license for the treatment of NAFLD will become available.

A multidisciplinary approach to the management of NAFLD is associated with improvement in markers of liver and cardio-metabolic health.

Non-alcoholic fatty liver disease (NAFLD) is a globally prevalent health problem, associated in its more severe forms with increased liver-related and cardiovascular-related morbidity and mortality. We established a multidisciplinary metabolic hepatology clinic in 2014 and have analysed the clinical data to evaluate the effectiveness of this service. Patients with NAFLD (n=165) who had attended two or more appointments were included. Prespecified clinical data were collected prospectively at clinic appointments and analysed retrospectively. Interventions offered included lifestyle advice, signposting to weight loss services and pharmacological treatment of diabetes and cardiovascular risk factors. Median follow-up was 13 months (range: 2-34). 59% (n=97) of patients had type 2 diabetes mellitus (T2DM). 53% (n=87) underwent liver biopsy of whom 18% (n=16) had cirrhosis. Median alanine aminotransferase (ALT) reduced by 11 IU/L (p<0.0001), median weight reduced by 3.3 kg (p=0.0005). There were significant reductions in HbA1c, total cholesterol and liver stiffness. Specifically, in patients with T2DM, HbA1c decreased by 4 mmol/mol (p=0.01) with significant reductions in ALT, weight and total cholesterol. Relative cardiovascular risk assessed by the QRISK3 score reduced in the whole cohort and in those with T2DM. Health economic modelling suggested the clinic intervention among those patients with poorly controlled T2DM was cost-effective. In conclusion, a multidisciplinary approach to the management of patients with NAFLD in this observational cohort study was associated with improvements in liver-related and cardio-metabolic related health parameters and with evidence of cost-effectiveness in patients with poorly controlled T2DM.

Mapping tissue water T1 in the liver using the MOLLI T1 method in the presence of fat, iron and B0 inhomogeneity.

Modified Look-Locker inversion recovery (MOLLI) T1 mapping sequences can be useful in cardiac and liver tissue characterization, but determining underlying water T1 is confounded by iron, fat and frequency offsets. This article proposes an algorithm that provides an independent water MOLLI T1 (referred to as on-resonance water T1 ) that would have been measured if a subject had no fat and normal iron, and imaging had been done on resonance. Fifteen NiCl2 -doped agar phantoms with different peanut oil concentrations and 30 adults with various liver diseases, nineteen (63.3%) with liver steatosis, were scanned at 3 T using the shortened MOLLI (shMOLLI) T1 mapping, multiple-echo spoiled gradient-recalled echo and 1 H MR spectroscopy sequences. An algorithm based on Bloch equations was built in MATLAB, and water shMOLLI T1 values of both phantoms and human participants were determined. The quality of the algorithm's result was assessed by Pearson's correlation coefficient between shMOLLI T1 values and spectroscopically determined T1 values of the water, and by linear regression analysis. Correlation between shMOLLI and spectroscopy-based T1 values increased, from r = 0.910 (P < 0.001) to r = 0.998 (P < 0.001) in phantoms and from r = 0.493 (for iron-only correction; P = 0.005) to r = 0.771 (for iron, fat and off-resonance correction; P < 0.001) in patients. Linear regression analysis revealed that the determined water shMOLLI T1 values in patients were independent of fat and iron. It can be concluded that determination of on-resonance water (sh)MOLLI T1 independent of fat, iron and macroscopic field inhomogeneities was possible in phantoms and human subjects.

Liver biochemical abnormalities in Turner syndrome: A comprehensive characterization of an adult population.

OBJECTIVE: Abnormal liver function tests (LFTs) are frequent in Turner syndrome (TS). The causes and clinical significance are unclear. AIMS: To investigate the prevalence of elevated LFTs in adult TS; secondly, to analyse the associations between elevated LFTs, TS-karyotypes and TS-related conditions; and thirdly, to evaluate liver stiffness and histological assessment. METHODS: A total of 125 TS women were retrospectively studied. Karyotypes, clinical and biochemical details and aortic measurements were recorded. Fibroscan and liver biopsy results were noted. RESULTS: Elevated LFTs were found in 49.6%: gamma-glutamyltransferase (GGT) in 88.7%, ALK in 45.2%, ALT in 40.3% and AST in 29%. A FIB-4 index >1.3 was found in 11.8%. Women with isochromosome of the X long arm, iso[X](q), had a higher prevalence of elevated LFTs. A lower prevalence of abnormal GGT was found in patients with 45,X/46,XX, 45,X/47,XXX or 45,X/46,XX/47,XXX. Subjects with raised GGT were older, shorter and more likely to have higher triglyceride levels. There was no association with HRT duration after adjusting for age. Among patients with elevated aminotransferases, no differences were noted, except for higher HDL-cholesterol levels. The sinuses and ascending aorta diameter were greater in the elevated LFTs group. Fibroscan was suggestive of significant liver fibrosis in 38.1%. Among 11 biopsies, liver architectural changes were reported in 45.4%, including two with cirrhosis. CONCLUSIONS: Elevated LFTs in TS are common and important to detect given the possible progression towards severe liver disease. An association between raised LFTs and karyotype iso[X]q was demonstrated. We have also shown a new association between abnormal LFTs and aortic dilatation.

Non-invasive Markers of Liver Fibrosis: Adjuncts or Alternatives to Liver Biopsy?

Liver fibrosis reflects sustained liver injury often from multiple, simultaneous factors. Whilst the presence of mild fibrosis on biopsy can be a reassuring finding, the identification of advanced fibrosis is critical to the management of patients with chronic liver disease. This necessity has lead to a reliance on liver biopsy which itself is an imperfect test and poorly accepted by patients. The development of robust tools to non-invasively assess liver fibrosis has dramatically enhanced clinical decision making in patients with chronic liver disease, allowing a rapid and informed judgment of disease stage and prognosis. Should a liver biopsy be required, the appropriateness is clearer and the diagnostic yield is greater with the use of these adjuncts. While a number of non-invasive liver fibrosis markers are now used in routine practice, a steady stream of innovative approaches exists. With improvement in the reliability, reproducibility and feasibility of these markers, their potential role in disease management is increasing. Moreover, their adoption into clinical trials as outcome measures reflects their validity and dynamic nature. This review will summarize and appraise the current and novel non-invasive markers of liver fibrosis, both blood and imaging based, and look at their prospective application in everyday clinical care.