Hypoglycemia measured by flash glucose monitoring system predicts liver-related events in chronic liver disease patients

Impaired glucose tolerance, glucose fluctuations, and hypoglycemia have been observed in patients with chronic liver disease (CLD). The flash glucose monitoring (FGM) system, which recognises continuous and dynamic glucose changes in real time, is used in daily clinical practice. This study aimed to examine the association between glucose fluctuations and hypoglycemia, as measured by the FGM system, and liver-related events. Seventy-two patients with CLD and type 2 DM who had their blood glucose measured using Freestyle Libre Pro between April 2017 and July 2018 at our institution were enrolled in this retrospective study. We assessed the results of the FGM system measurements and liver-related events, as defined by gastrointestinal bleeding, infection, ascites, encephalopathy, and liver-related death. The standard deviation (SD) of mean glucose as measured by the FGM system was 41.55 mg/dl, and hypoglycemia was observed in 48.6% (35/72) of the patients. Liver-related event-free survival was not significant when stratified based on SD; however, the event-free survival was significantly lower when stratified by hypoglycemia (p = 0.007). In a multivariate analysis using the Cox proportional hazards model, Child–Pugh class B [Hazards ratio (HR) 2.347 (95% confidence interval (CI): 1.042–5.283), p = 0.039] and hypoglycemia [HR 2.279 (95% CI: 1.064–4.881), p = 0.034] were identified as factors contributing to event-free survival. Hypoglycemia, as determined by the FGM system, was identified as a significant factor that was closely associated with liver-related events. In addition to measuring glucose levels, the FGM system is useful in predicting the occurrence of liver-related events.

Statistical analysis.Continuous variables (age, body mass index, duration of DM, platelet count, alanine transferase, FPG, immunoreactive insulin, HbA1c, glycoalbumin, SD of mean glucose, CV, and MAGE) were dichotomized with respect to the median or clinically meaningful values in a multivariate analysis.Spearman's rank correlation coefficient was used for statistical analysis, and the Mann-Whitney U test was used for comparison between the two groups.The Kaplan-Meier method and log-rank test were used to estimate the eventfree survival rate.Cox proportional hazards regression analysis was performed to evaluate the risk factors for event-free survival.Statistical significance was set at p < 0.05.Data analysis was performed using SPSS ver.22.0 (SPSS, Chicago, IL, USA).

Results
Patient characteristics.The baseline characteristics of the 72 patients included in this study are summarized in Table 1.The median duration of diabetes was 9.0 years.All patients received glucose-lowering therapy and 25.0% used insulin.Overall, 88% of the patients had cirrhosis and 48.6% had a history of liver cancer.The median observation period was 35.0 months.www.nature.com/scientificreports/Glucose parameters and FGM parameters.The results of glucose metabolism using peripheral blood measurements and the FGM system are shown in Table 2.The SD of mean glucose level as measured by the FGM system was 41.55 mg/dl and median CV was 29.6%, and hypoglycemia was observed in 48.6% (35/72) of the patients.The median percentage of time below range was 1.0% overall and 4.5% for the hypoglycemia group only.In addition, 55.0% of the measured hypoglycemic events were detected in the nocturnal period (0000-0559 h).There was a correlation between classical glycemic variability parameters and SD values (Fig. 1).The correlation between HbA1c and glycoalbumin was weak (Fig. 1A and B), while MAGE showed a strong correlation (Fig. 1C).However, no significant difference was observed between hypoglycemia and SD values (p = 0.241, Fig. 1D).
Incidence rates of liver-related events.During the observation period, five patients underwent liver transplantation.There were 28 liver-related events and the event-free survival rate at 1 year was 74.6%.Liverrelated events included encephalopathy in nine cases, ascites in eight cases, gastrointestinal bleeding in seven cases, and severe infection in four cases.Liver-related event-free survival was compared between the two groups according to mean sensor glucose, SD of mean glucose, and CV values, but no significant difference was found (Fig. 2A, B, C).Contrarily, liver-related event-free survival according to glucose levels showed significantly lower event-free survival in the hypoglycemia group (p = 0.007, Fig. 2D).www.nature.com/scientificreports/Multivariate analysis of factors contributing to liver-related event-free survival.Table 3 shows the multivariate analysis of factors contributing to event-free survival using the Cox proportional hazards model.In univariate analysis, branched-chain amino acids (BCAA), Child-Pugh class, HbA1c, and hypoglycemia were identified as factors contributing to event-free survival.In multivariate analysis, Child-Pugh class B (hazard's ratio (HR) 2.347 [95% confidence interval (CI): 1.042-5.283],p = 0.039) and hypoglycemia (HR 2.279 [95% CI: 1.064-4.881),p = 0.034) were identified as factors contributing to event-free survival.

Discussion
Hepatic glycogen storage is decreased in patients with CLD 10 .Postprandial glucose uptake from the blood by the liver is delayed, resulting in hyperglycemia.In addition, patients with CLD have a reduced storage capacity for hepatic glycogen, resulting in inadequate glucose release from the liver into the blood during fasting, and impaired gluconeogenesis, leading to hypoglycemia; therefore, the management of diabetes in patients with liver disease can be difficult 11 .The metabolic state of patients with CLD after an overnight fast is similar to that In the analysis focusing on glucose fluctuation, there was a weak correlation with DM parameters such as HbA1c and GA, and a strong correlation with MAGE, a classical fluctuation parameter (Fig. 1).The SD of mean glucose level as measured by FGM can be considered a surrogate index for MAGE.The CV was also examined as a measure of fluctuation; however, SD had a stronger correlation with MAGE.In our previous study, the SD was 24.1 mg/dl 6 , and it was reported to be approximately 14 in healthy subjects 12 .In the present study, the SD value of blood glucose was 41.5 mg/dl, which was as high as expected and considered to be affected by CLD.There was no significant relationship between glucose fluctuations and liver-related events.
The CGM system has been reported to be useful in detecting hidden abnormalities in blood glucose fluctuations in patients with type 2 DM and CLD 13 .Abnormal blood glucose fluctuations have also been reported to be a risk factor for sleep disturbance and decreased quality of life in patients with LC 6 .However, glucose fluctuation does not predict liver-related events such as encephalopathy, infection, and liver failure, which are more severe in patients with CLD.Similar results indicating that hypoglycemia is more important than blood glucose fluctuations have been reported, although not in patients with liver disease 14 .
In this study, hypoglycemia, as determined by the FGM system, was identified as a significant factor closely associated with liver-related events.Hypoglycemia plays an important role in inflammation, thrombotic events, and endothelial dysfunction by inducing oxidative stress 15 .Previous reports have shown that hypoglycemia is an important prognostic factor for short-term mortality in patients with cirrhosis 16 .In addition, hypoglycemia has been reported to be associated with nutritional deficiencies, infections, and poor glucogenesis 17,18 .The presence of DM itself is associated with infections, variceal hemorrhage, and encephalopathy 19,20 .Previous reports have consistently shown that hypoglycemia is an important factor in liver-related events.Therefore, it is reasonable to accept hypoglycemia as a risk factor for liver-related event-free survival in our study.
Hypoglycemia measured using the FGM system showed a higher frequency in the total patient population (48.6%, 35/72).Previous studies of continuous blood glucose measurement using iPro2 have shown that hypoglycemia is infrequent (16.3%) 6 .Compared to iPro2, FGM has a long measurement period of up to 14 days, www.nature.com/scientificreports/making it easier to detect hypoglycemia.It has been reported that CGM systems using FreeStyle Libre Pro can detect hypoglycemia better than point-of-care capillary glucose testing 21 .In addition, glucose measurements using this CGM system have been reported to be slightly lower than blood glucose levels 21 .Furthermore, 55.0% of the measured hypoglycemic events were detected during the nocturnal period, indicating that FGM is superior in detecting latent hypoglycemia and has a higher hypoglycemic frequency than previously reported.Glycated hemoglobin A1c (HbA1c) and glycoalbumin are the gold standard indicators of glycemic control in diabetes.However, HbA1c cannot adequately represent the glycemic control status in patients with CLD because of the short lifespan of erythrocytes caused by hypersplenism.Glycoalbumin is affected by impaired albumin metabolism; in patients with CLD, the half-life of serum albumin is prolonged owing to decreased albumin synthesis 22 .Therefore, it is difficult to accurately monitor the glycemic control status in patients with CLD.The FGM system has enabled the identification of hypoglycemia in patients with CLD at a high risk of liver-related events.It is an excellent system for detecting latent hypoglycemia during a routine examination in a population with apparently good glycemic control, including in those with low HbA1c and glycoalbumin levels.
Late evening snack (LES) with BCAA supplementation is considered to be effective in improving proteinenergy nutrition 23 and avoiding nocturnal hypoglycemia in patients with CLD.In this study, BCAA and LES with BCAA were administered to 31.4 and 4.2% of the patients, respectively.Since all patients had type 2 DM, the possibility that they avoided calorie intake was considered, and there is a potential opportunity for intervention in the future.
A limitation of this study is that it was a single-centre retrospective analysis and there were no treatment interventions based on the FGM system measurements.Whether an intervention for hypoglycemia with FGM leads to a reduction in liver-related events is unknown 24 and is a subject for future research.Another limitation is that the FGM system measurements were performed under health insurance and patients with CLD without therapeutic intervention for DM were not monitored by the FGM system.Despite these limitations, this is the first report to describe the relationship between hypoglycemia identified using the FGM system and liver disease-related events.These results suggest that the FGM system, in addition to measuring glucose levels, is useful for predicting the occurrence of liver-related events.

Figure 1 .
Figure 1.Association of the standard deviation of mean glucose with classical DM control index and hypoglycemia.The standard deviation of mean glucose correlated with HbA1c (A), Glycoalbumin (B), and MAGE (C).(D) Standard deviation of mean glucose did not differ between patients with and without hypoglycemia.

Figure 2 .
Figure 2. Liver-related event-free survival.No significant difference was seen (A, B, C).Liver-related eventfree survival based on glucose levels showed significantly lower event-free survival in the hypoglycaemia group (p = 0.007, D).

Table 2 .
Laboratory characteristics associated with flash glucose monitoring.Data are given as the medians with interquartile ranges or numbers.MAGE mean amplitude of glycemic excursions.

Table 3 .
Multivariate analysis of factors contributing to event-free survival using the Cox proportional hazards model.BMI body mass index, ALT alanine aminotransferase, FPG fasting plasma glucose, IRI immunoreactive insulin, HbA1c hemoglobin A1c, SD standard deviation, MAGE mean amplitude of glycemic excursions.