KLF9 drives intermittent hypoxia-induced NAFLD by suppressing the NR4A1-p38 MAPK hepatic metabolic axis.

PurposeGrowing evidence has revealed that nonalcoholic fatty liver disease (NAFLD) is associated with type 2 diabetes. This study aimed to assess the association between glycometabolism and NAFLD in patients with obstructive sleep apnea (OSA).MethodsPatients with suspected OSA were enrolled consecutively and then underwent polysomnography, liver ultrasound, and biochemical measurements. Logistic regressions were used to identify factors associated with NAFLD.ResultsIn total, 415 patients were included. The prevalence of NAFLD in the non-OSA, mild OSA, moderate OSA, and severe OSA groups was 37.21%, 69.09%, 68.34%, and 78.08%, respectively. Stepwise logistic regression suggested that percentage of total sleep time spent with oxygen saturation of < 90% (TS90), lowest oxygen saturation (LaSO2), and homeostatic model assessment of insulin resistance (HOMA-IR) were independently associated with NAFLD in all subjects, after adjusting for confounders (odds ratio [OR] = 1.037, p = 0.014; OR = 1.056, p = 0.004; OR = 0.732, p = 0.009; respectively). TS90, LaSO2, and HOMA-IR were also independent predictors for NAFLD in patients with mild and moderate OSA, whereas TS90, LaSO2, and ODI were independent predictors for NAFLD in patients with severe OSA.ConclusionsThere is a relationship between OSA and NAFLD, and the combination of disordered glycometabolism and intermittent hypoxia may act as a “two hit” mechanism to promote the development of NAFLD. Furthermore, intermittent hypoxia alone was an independent predictor for NAFLD in severe OSA patients.

Obstructive sleep apnoea and nonalcoholic fatty liver disease: risk factor or just coincidence?

Recent studies have demonstrated that obstructive sleep apnea (OSA) is associated with the development and evolution of nonalcoholic fatty liver disease (NAFLD), independent of obesity or other shared risk factors. Like OSA, NAFLD is a prevalent disorder associated with major adverse health outcomes: Patients with NAFLD may develop cirrhosis, liver failure, and hepatocellular carcinoma. One major finding that has emerged from these studies is that the OSA-NAFLD association is related to the degree of nocturnal hypoxemia in OSA. Animal models have therefore largely focused on intermittent hypoxia, a key manifestation of OSA, to shed light on the mechanisms by which OSA may give rise to the complex metabolic disturbances that are seen in NAFLD. Intermittent hypoxia leads to tissue hypoxia and can result in oxidative stress, mitochondrial dysfunction, inflammation, and overactivation of the sympathetic nervous system, among many other maladaptive effects. In such models, intermittent hypoxia has been shown to cause insulin resistance, dysfunction of key steps in hepatic lipid metabolism, atherosclerosis, and hepatic steatosis and fibrosis, each of which is pertinent to the development and/or progression of NAFLD. However, many intriguing questions remain unanswered: Principally, how aggressively should the clinician screen for NAFLD in patients with OSA, and vice versa? In this review, we attempt to apply the best evidence from animal and human studies to highlight the relationship between these two disorders and to advocate for further trials aimed at defining these relationships more precisely.

the phenotypic characterization of obstructive sleep apnea (OSA) continues to unfold. Recent studies provide evidence that this syndrome has the potential to disturb cardiovascular, neurological, and endocrine homeostatic processes ([1][1], [6][2], [8][3], [9][4]). Li et al. ([5][5]) present

Fibroblast growth factor 21 as a biomarker for NAFLD: Integrating pathobiology into clinical practice

Intestinal FGF19 has emerged as a novel endocrine regulator of hepatic bile salt and lipid metabolism. In patients with nonalcoholic fatty liver disease (NAFLD) hepatic lipid metabolism is deranged. A possible role of FGF19 in NAFLD has not been reported yet. In this study, we assessed intestinal FGF19 production and the hepatic response to FGF19 in NAFLD patients with and without insulin resistance [homeostasis model of assessment (HOMA) score > or =2.5 (n = 12) and HOMA score <2.5 (n = 8), respectively]. To this end, NAFLD patients received a standardized oral fat challenge. Postprandial excursions of triglycerides, bile salts, and FGF19 were monitored, and plasma levels of a marker for bile salt synthesis (7alpha-hydroxy-4-cholesten-3-one) were determined. Fasted FGF19 levels were comparable in a control group of healthy volunteers (n = 15) and in NAFLD patients (0.26 +/- 0.28 vs. 0.18 +/- 0.09 ng/ml, respectively, P = 0.94). Postprandial FGF19 levels in both controls and NAFLD patients peaked between 3-4 h and were three times higher than baseline levels. The areas under the postprandial FGF19 curve were similar in controls and in the HOMA score-based NAFLD subgroups. In NAFLD patients with HOMA score <2.5, the postprandial increase in plasma FGF19 was accompanied by a lowering of plasma levels of 7alpha-hydroxy-4-cholesten-3-one (-30%, P = 0.015). This anticipated decline was not observed in insulin-resistant NAFLD patients (+10%, P = 0.22). In conclusion, patients with NAFLD show an unimpaired intestinal FGF19 production. However, the hepatic response to FGF19 is impaired in NAFLD patients with insulin resistance (HOMA score > or =2.5). This impaired hepatic response to FGF19 may contribute to the dysregulation of lipid homeostasis in NAFLD.

Obstructive sleep apnea (OSA) is associated with metabolic dysfunction, including progression of nonalcoholic fatty liver disease (NAFLD). Chronic intermittent hypoxia (IH) as a model of OSA worsens hepatic steatosis and fibrosis in rodents with diet induced obesity. However, IH also causes weight loss, thus complicating attempts to co-model OSA and NAFLD. We sought to determine the effect of various durations of IH exposure on metabolic and liver-related outcomes in a murine NAFLD model. We hypothesized that longer IH duration would worsen the NAFLD phenotype. Male C57BL/6J mice (n = 32) were fed a high trans-fat diet for 24 weeks, to induce NAFLD with severe steatohepatitis. Mice were exposed to an IH profile modeling severe OSA, for variable durations (0, 6, 12, or 18 weeks). Intraperitoneal glucose tolerance test was measured at baseline and at six-week intervals. Liver triglycerides, collagen and other markers of NAFLD were measured at sacrifice. Mice exposed to IH for 12 weeks gained less weight (p = 0.023), and had lower liver weight (p = 0.008) relative to room air controls. These effects were not observed in the other IH groups. IH of longer duration transiently worsened glucose tolerance, but this effect was not seen in the groups exposed to shorter durations of IH. IH exposure for 12 or 18 weeks exacerbated liver fibrosis, with the largest increase in hepatic collagen observed in mice exposed to IH for 12 weeks. Duration of IH significantly impacts clinically relevant outcomes in a NAFLD model, including body weight, fasting glucose, glucose tolerance, and liver fibrosis.

Obstructive sleep apnea is the most common sleep-related breathing disorder worldwide and remains underdiagnosed. Its multiple associated comorbidities contribute to a decreased quality of life and work performance as well as an increased risk of death. Standard treatment seems to have limited effects on cardiovascular and metabolic aspects of the disease, emphasising the need for early diagnosis and additional therapeutic approaches. Recent evidence suggests that the dysregulation of circadian rhythms, processes with endogenous rhythmicity that are adjusted to the environment through various cues, is involved in the pathogenesis of comorbidities. In patients with obstructive sleep apnea, altered circadian gene expression patterns have been demonstrated. Obstructive respiratory events may promote circadian dysregulation through the effects of sleep disturbance and intermittent hypoxia, with subsequent inflammation and disruption of neural and hormonal homeostasis. In this review, current knowledge on obstructive sleep apnea, circadian rhythm regulation, and circadian rhythm sleep disorders is summarised. Studies that connect obstructive sleep apnea to circadian rhythm abnormalities are critically evaluated. Furthermore, pathogenetic mechanisms that may underlie this association, most notably hypoxia signalling, are presented. A bidirectional relationship between obstructive sleep apnea and circadian rhythm dysregulation is proposed. Approaching obstructive sleep apnea as a circadian rhythm disorder may prove beneficial for the development of new, personalised diagnostic, therapeutic and prognostic tools. However, further studies are needed before the clinical approach to obstructive sleep apnea includes targeting the circadian system.

Nocturnal intermittent hypoxia (IH) featuring obstructive sleep apnea (OSA) dysregulates hepatic lipid metabolism and might contribute to the development of non-alcoholic fatty liver disease (NAFLD) observed in OSA patients. However, further research is required to better understanding the molecular mechanisms underlying IH-induced hepatic lipid accumulation. Therefore, the aim of the present study was to determine the effects of OSA on hepatic CD36 expression and the impact of IH by using a mouse model of OSA. Histological analysis, lipid content and CD36 expression were assessed in livers from subjects who underwent liver biopsy and polygraphic study during sleep, and in livers from mice submitted to chronic IH mimicking OSA. Among those who presented OSA features, NAFLD were significantly more frequent than in control subjects with normal respiratory function (77.8 vs. 36.4%, respectively), and showed more severe liver disease. Interestingly, CD36 expression was significantly overexpressed within the liver of OSA patients with respect to controls, and a significant positive correlation was observed between hepatic levels of CD36 and the values of two well-known respiratory parameters that characterized OSA: apnea/hypopnea index (AHI) and oxygen desaturation index (ODI). Moreover, hepatic lipid accumulation as well as induction of hepatic lipogenic genes, and CD36 mRNA and protein expression were significantly higher in livers from mice exposed to IH conditions for 8 weeks than in their corresponding littermates. This study provides novel evidence that IH featuring OSA could contribute to NAFLD setup partly by upregulating hepatic CD36 expression.

Introduction: Nonalcoholic fatty liver disease (NAFLD) is strongly linked to insulin resistance. As a result, type 2 diabetes mellitus (T2DM) is commonly comorbid in NAFLD and can increase morbidity and reduce weight loss treatment efficacy. We studied weight loss in adult NAFLD patients without and with T2DM (NAFLD-DM) receiving care in the Yale Fatty Liver Disease Program (YFLDP), which focuses on the integration of medical weight management treatments and hepatology care. Methods: We analyzed retrospective data for adult patients with diagnostic codes for NAFLD and T2DM with complete data on body weight and at least one clinic visit between 5/23/2018 and 5/02/2022. Patients were categorized as NAFLD or NAFLD-DM. Total body weight loss (WL) was quantified using 0%, 5%, and 10% thresholds. Between NAFLD and NAFLD-DM groups, we compared total body weight loss (WL) and across WL threshold groups, we also compared initial weights, follow-up time, medical interventions received, and results of non-invasive liver fibrosis tests. Results: There were 933 patients with NAFLD, of whom 463 (49.6%) had NAFLD-DM. After a median follow-up of 7.7 months, 632 (67.7%) of all patients achieved any weight loss. Significantly more patients with NAFLD-DM lost weight compared to NAFLD alone (73.9 vs 61.7%, p< 0.0001). Weight loss between NAFLD and NAFLD-DM groups varied, respectively: 0-5% WL: 34.7% vs. 34.8% (p=0.98); 5-10% WL: 13.2% vs. 19.2% (p=0.01), >10% WL: 13.8% vs. 19.9% (p=0.01). Compared to those not losing weight, NAFLD and NAFLD-DM patients with >10% WL had significantly greater proportion of participation in a complete meal replacement program (Optifast®) or treatment with GLP-1 agonist semaglutide. For NAFLD and NAFLD-DM groups, there was no significant difference between initial Fibroscan stiffness or FIB-4 index and subsequent WL percentages (Table). Conclusion: The majority of NAFLD and NAFLD-DM patients receiving care in a specialty clinic incorporating medical weight management with liver care successfully achieved weight loss in a relatively short period. Patients with NAFLD-DM did not have significantly worse outcomes with weight loss, and NAFLD-T2DM patients lost more weight than NAFLD patients. These findings suggest that an integrative care liver clinic with concurrent weight management care can help patients improve metabolic diseases associated with NAFLD, particularly T2DM, and in turn improve liver-related outcomes.Figure 1.: Comparison of Clinic Interventions between No Weight Loss (WL) versus >10% WL patients with NAFLD (top panel, a) and NAFLD-DM (bottom panel, b). *Statistically significant p-values using chi-square analysis, which can be found in Table Table 1. - Characteristics of and Clinical Interventions for NAFLD and NAFLD-DM Patients in the Yale Fatty Liver Disease Program (YFLDP). There was a significantly lower proportion of patients with NAFLD compared to NAFLD-DM who were able to achieve > 10% Weight Loss (13.8 vs. 19.9, p=0.014), and significantly more NAFLD patients compared to NAFLD-DM who did not achieve any weight loss (38.3 vs. 26.1, p<0.001) NAFLD (n=470 patients) NAFLD-DM (n=463 patients) Patient Categories n (%) No WL180 (38.3) >10% WL65 (13.8) p No WL121 (26.1) >10% WL92 (19.9) p Clinic Interventions n (%) Nutrition Consultation 80 (44.4) 31 (47.7) 0.652 58 (47.9) 40 (43.5) 0.518 OPTIFAST® Program 4 (2.2) 5 (7.7) 0.045 2 (1.7) 8 (8.7) 0.016 Naltrexone/Buproprion 6 (3.3) 5 (7.7) 0.146 16 (13.2) 6 (6.5) 0.111 Semaglutide 15 (8.3) 17 (26.2) < 0.001 30 (24.8) 37 (40.2) 0.016 Liraglutide 11 (6.1) 7 (10.8) 0.217 21 (17.4) 7 (7.6) 0.037 Phentermine 9 (5.0) 10 (15.4) 0.007 15 (12.4) 17 (18.5) 0.219 Bariatric Surgery Referral 13 (7.2) 5 (7.7) 0.901 21 (17.4) 31 (33.7) 0.006 Health Outcomes mean (SD) Follow-Up Time, days 256 (276.5) 422 (296.8) < 0.001 271 (280.2) 403 (297.3) 0.001 Initial FibroScan® Stiffness, kPa 8.63 (6.72) 13.18 (14.01) 0.663 13.18 (14.01) 12.48 (12.49) 0.701 Initial FIB-4 Score 1.32 (1.13) 1.70 (1.98) 0.068 1.70 (1.98) 1.83 (2.11) 0.648 Initial Weight, kg 93.82 (25.45) 102.20 (24.90) 0.015 102.20 (24.90) 109.61 (25.07) 0.056 *Denotes statistical significance p<0.05. WL = Weight Loss.

We aimed to elucidate the frequency of polymorphic genotypes and alleles of patatin-like phospholipase domain containing 3 rs738409 polymorphism and its possible associations with non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis in a cohort from Turkey.We enrolled 200 patients diagnosed with NAFLD and genotyped for rs738409 I148M polymorphism by real-time polymerase chain reaction, particularly by melting curve analysis. SPSS analysis software was used for statistical significance. Continuous variable values were expressed as mean ± standard deviation. Significant statistical level was chosen as p = 0.05.Our results demonstrate in a cohort from Turkey that rs738409 C > G polymorphism (I148M) of patatin-like phospholipase domain containing 3 gene is significantly able to affect individuals to have NAFLD in unadjusted regression model.Consistent with the previous studies in other populations, our study group showed a significantly higher risk of having NAFLD in unadjusted regression model but not in the adjusted model indicating that non-genetic factors such as age and sex may be responsible for the association. However, independent studies need to validate our findings with a larger group of NAFLD patients, as well as in different ethnic cohorts.

The association of obstructive sleep apnea (OSA) with nonalcoholic fatty liver disease (NAFLD) has only been studied in selected subgroups such as the morbidly obese. We aimed to determine the prevalence and effect of OSA on NAFLD and vice versa in unselected patients attending the outpatient department. OSA was diagnosed by polysomnography, done in patients having symptoms of OSA, in patients with NAFLD attending the liver clinic. Polysomnography-proven patients with OSA attending the chest clinic were evaluated for NAFLD by ultrasonography. Anthropometry, liver function tests, metabolic syndrome evaluation and transient elastography were performed in all patients. Three (3%; 95% CI 1.03-8.45%) out of 100 patients with NAFLD (mean age 41 ± 11 years) had symptomatic OSA. Of 23 patients with OSA (mean age 46 ± 12 years,), 3 (13%) had mild, 5 (22%) moderate and 15 (65%) severe OSA. Twenty-one (91.3%; 95% CI 73.2-97.6%) patients with OSA had NAFLD, while raised hepatic transaminase levels were seen in seven (30.4%; 95% CI 15.6-50.9%). Body mass index (OR 1.21, 95% CI 1.02-1.44) and male gender (OR 4.79, 95% CI 1.12-20.48) were significant independent predictors of OSA in NAFLD. The apnea-hypopnea index (OR 1.084, 95% CI 1.002-1.172), a marker of OSA severity, was the only significant independent predictor of significant fibrosis in patients with NAFLD. Prevalence of symptomatic OSA in patients with NAFLD is low and is predicted by male gender and obesity. Prevalence of NAFLD in patients with OSA is very high. Significant hepatic fibrosis in patients with NAFLD is predicted by OSA independent of obesity and metabolic syndrome.

Reduction of Hepatosteatosis and Lipid Levels by an Adipose Differentiation-Related Protein Antisense Oligonucleotide

The biological factors that promote inflammation or nonalcoholic steatohepatitis (NASH) in the setting of nonalcoholic fatty liver disease remain incompletely understood. Clinical studies have demonstrated an association between obstructive sleep apnea (OSA) and both inflammation and fibrosis in NASH, but the mechanism has not been identified. In this study, we use in vitro modeling to examine the impact of intermittent hypoxia on the liver. Hepatocyte, stellate cell, and macrophage cell lines were exposed to intermittent or sustained hypoxia. Candidate genes associated with inflammation, fibrosis, and lipogenesis were analyzed. Circulating cytokines were assessed in human serum of patients with nonalcoholic fatty liver disease. Intermittent hypoxia results in significant induction of interleukin (IL)‐6 expression in both hepatocytes and macrophages. The increase in IL‐6 expression was independent of hypoxia inducible factor 1 induction but appeared to be in part related to antioxidant response element and nuclear factor kappa B activation. Mature microRNA 365 (miR‐365) has been demonstrated to regulate IL‐6 expression, and we found that miR‐365 expression was decreased in the setting of intermittent hypoxia. Furthermore, macrophage cell lines showed polarization to an M1 but not M2 phenotype. Finally, we found a trend toward higher circulating levels of IL‐6 in patients with OSA and NASH. Conclusion: Intermittent hypoxia acts as a potent proinflammatory stimulus, resulting in IL‐6 induction and M1 macrophage polarization. Increased IL‐6 expression may be due to both induction of antioxidant response element and nuclear factor kappa B as well as inhibition of miR‐365 expression. Higher levels of IL‐6 were observed in human samples of patients with OSA and NASH. These findings provide biological insight into mechanisms by which obstructive sleep apnea potentiates inflammation and fibrosis in patients with fatty liver disease. (Hepatology Communications 2017;1:326–337)

589 Background: The proportion of non-alcoholic fatty liver disease (NAFLD) has been increasing as a cause of hepatocellular carcinoma (HCC) worldwide. Natural killer (NK) cells are involved in the first line of immune defense against cancer. NK cell function is regulated by activating and inhibitory NK cell receptors. However, the role of NK cells in the pathogenesis of NAFLD and NAFLD-HCC is still largely unknown. In this study, we aimed to clarify the phenotypic and functional features of NK cells in NAFLD and NAFLD-HCC patients. Methods: We performed mass cytometry (CyTOF) and flow cytometry analysis of NK cells in 33 NAFLD patients (22 chronic hepatitis (CH), 8 liver cirrhosis (LC), 3 with HCC (HCC)) and 9 healthy donors (HDs). We compared surface markers on NK cells in cancerous and non-cancerous intrahepatic lymphocytes (IHLs). We also measured NK cell function in the presence of IL-12 and IL-18. Results: The frequency of NK cells was lower in NAFLD patients compared to HDs. PD-1, CD57, ILT2 were highly expressed, and Siglec-7, NKp30, NKp46 were downregulated on NK cells from NAFLD patients, compared with those of HDs. In NAFLD patients, Siglec-7 levels on NK cells were negatively correlated with PD-1 and CD57, and positively correlated with NKp30 and NKp46. The other inhibitory markers (NKG2A, KIR3DL1 and KIRDL2/L3), activating markers (CD69 and NKG2D) and checkpoint markers (Tim-3 and TIGIT) were comparable between NAFLD patients and HDs. PD-1 and CD57 expression levels on NK cells were also significantly upregulated in NAFLD-HCC patients than those in HDs. CD57 was rarely expressed on NK cells in non-cancerous IHLs, on the other hand, highly expressed in cancerous IHLs. The IFNγ production and CD107a expression on NK cells were also decreased in NAFLD patients. PD-1+CD57+Siglec-7− NK cells were observed in NAFLD patients, rarely in HDs. PD-1+CD57+Siglec-7− NK cells were functionally impaired compared to other NK subsets. Conclusions: In patients with NAFLD, NK cells are reduced and functionally impaired, the reason of which may be the increased proportion of dysfunctional PD-1+CD57+Siglec-7− NK subset, and dysfunctional NK cells might be related to impairment of surveillance for HCC.