Associations between glucose metabolism disorders and prognosis in patients with acute-on- chronic liver failure

Han Hu A liated Hospital of Zunyi Medical University Xinxin Hu A liated Hospital of Zunyi Medical University Caiyun Tian A liated Hospital of Zunyi Medical University Yanping Zhu A liated Hospital of Zunyi Medical University Yujuan Liu A liated Hospital of Zunyi Medical University Qijiao Cheng A liated Hospital of Zunyi Medical University Fangwan Yang A liated Hospital of Zunyi Medical University Jun Liu School of public health, Zunyi Medical University Ying Li A liated Hospital of Zunyi Medical University Shi De Lin (  linshide6@hotmail.com ) A liated Hospital of Zunyi Medical University https://orcid.org/0000-0001-8803-4069


Introduction
The liver plays an important role in glucose metabolism. Excessive liver damage disturbs glucose homeostasis, thus resulting in hypoglycemia, impaired glucose tolerance (IGT) and diabetes in patients with severe liver diseases [1]. Diabetes, second to severely impaired liver function, is known as hepatogenous diabetes (HD), although the World Health Organization does not classify it as an independent disease [2]. Hyperglycemia can also occur in patients with severe acute diseases such as sepsis [3], coronavirus disease 2019 [4], and myocardial infarction [5] and is de ned as stress-induced hyperglycemia [6].
Acute-on-chronic liver failure (ACLF) is a severe liver injury accompanied by reduced liver function, organ failure and systemic in ammation in patients with chronic liver diseases [7]. In China, ACLF occurs most frequently in patients with chronic hepatitis B or hepatitis B virus (HBV)-related cirrhosis. Patients with HBV-related ACLF have different degrees of liver brosis and severe acute systemic in ammation [8]. Previous studies found that HD is a common complication in patients with liver cirrhosis. The HD prevalence in patients with liver cirrhosis depends on the degree of liver brosis, the etiology of the chronic liver disease and the severity of the liver injury [9,10]. Hypoglycemia and temporal hyperglycemia can also occur in patients with severe acute hepatitis and acute liver failure (ALF) [11]. Therefore, glucose metabolism disorders in patients with ACLF may differ from those in patients with liver cirrhosis. In patients with ACLF, glucose metabolism disorders may result from both the loss of liver function and the stress response to acute systemic in ammation. However, the clinical characteristics of glucose metabolism disorders in patients with ACLF remain unclear.
Both hypoglycemia and hyperglycemia have been closely correlated with high mortality in patients with severe diseases [3,12]. Previous studies have found that diabetes was associated with a poor long-term prognosis and high risk of complications such as bacterial infections, esophageal variceal bleeding, hepatic encephalopathy, and hepatocellular carcinoma in patients with liver cirrhosis [13][14][15][16]. However, the diagnostic methods used to test for diabetes and the etiologies of the patients varied widely in these studies. Moreover, the effects of diabetes on short-term mortality in patients with liver cirrhosis remain controversial, and the effects of glucose metabolism disorders on the prognosis of patients with ACLF are unknown [17,18]. Here, we evaluated the association between glucose metabolism disorders and 90day mortality in patients with HBV-related ACLF.

Study participants
We retrospectively analyzed 206 patients with ACLF associated with chronic HBV infection who were hospitalized in the Department of Infectious Diseases at the A liated Hospital of Zunyi Medical University (Zunyi, China) from October 2017 to December 2020. Forty-six patients who did not undergo an oral glucose tolerance test (OGTT) on admission because they were complicated with esophageal or gastric variceal bleeding, hepatic encephalopathy or hepatorenal syndrome were excluded. Fifty-one patients were excluded for coexistence of other liver diseases: 18 had alcoholic liver disease, 13 had druginduced hepatitis, 9 had hepatocellular carcinoma, 6 had a previous diagnosis of type 2 diabetes, and 5 were previously or presently taking steroids. Six patients who were lost to follow-up within 90 days and 7 with incomplete clinical data were also excluded. Finally, 96 patients were enrolled in the study.
Diagnostic criteria for ACLF, liver cirrhosis and bacterial infections ACLF was diagnosed as the recent development of jaundice (total bilirubin [TBil] ≥5×upper limits of normal [ULN])and coagulopathy (prothrombin activity [PTA] <40% or international normalized ratio [INR] ≥1.5), complicated within 4 weeks by ascites and/or encephalopathy in patients with previously diagnosed or undiagnosed chronic liver disease [19].
Liver cirrhosis was diagnosed based on previous liver biopsy ndings, ultrasonography, computed tomography, liver stiffness measurement or magnetic resonance imaging ndings. Patients with irregular and nodular livers, small and shrunken livers, splenomegaly and hypersplenism, or evidence of portosystemic collaterals together with impaired liver synthetic function were diagnosed with liver cirrhosis [20].
Clinical and laboratory assessment Patient demographics, clinical and laboratory variables, and imaging ndings were collected within 24 h before and after admission.
Fasting plasma glucose (FPG), fasting insulin (FINS) and fasting C-peptide (FCP) were detected within 24 h after admission. OGTTs were performed as described by the World Health Organization with an oral anhydroglucose load of 75 g, and glycemia was measured at 0 and 2 hours. Glucose abnormalities were diagnosed based on the American Diabetes Association criteria [22]. Hypoglycemia was de ned as FPG <3.8 mmol/L; diabetes was established as FPG ≥7 mmol/L or OGTT 2-h plasma glucose (OGTT 2h-PG) ≥11.1 mmol/L. IGT included patients with impaired fasting glucose (FPG of ≥5.6 mmol/L but <7 mmol/L) and/or impaired glucose tolerance (OGTT 2h-PG of ≥7.8 mmol/L and <11.1 mmol/L). Because glycated hemoglobin (HbA1c) levels may be inappropriately normal in patients with severe liver cirrhosis owing to reduced erythrocyte lifespans [2], we excluded HbA1c from this study. The homeostasis model of assessment 2-insulin resistance (HOMA2-IR), HOMA2-insulin sensitivity (HOMA2-IS), and HOMA2-β-cell function (HOMA2-β) indices were estimated with the HOMA2 calculator, version 2.2 released by the Diabetes Trials Unit at the Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford [23]. Liver disease severity was assessed using the model for end-stage liver disease (MELD) score, which was calculated using the following formula: MELD score = 3.78 × ln[TBil (mg/dL)] + 11.2 × ln[INR] + 9.57 × ln[Cr (mg/dL)] + 6.43 × (constant for liver disease etiology = 0 if cholestatic or alcoholic, otherwise = 1).

Therapeutic method and follow-up
For patients with diabetes, oral hypoglycemic agents or insulin were initiated if dietary therapy was insu cient to obtain good glycemic control. For patients with IGT, no speci c treatment was administered. All patients were followed for up to 90 days after admission. In 22 patients with diabetes or IGT without oral hypoglycemic agents or insulin treatment, an OGTT was repeated to evaluate the regression of their glucose metabolism disorders. Based on their OGTT results at baseline and follow-up, patients were retrospectively assigned to one of two outcomes groups: (1) regression: from diabetes or IGT to normal glucose tolerance (NGT) or from diabetes to IGT or (2) non-regression: no change in their glucose metabolism disorders or progression from IGT or NGT to diabetes or from NGT to IGT.
Statistical analysis SPSS, version 19.0 (IBM Corp., Armonk, NY, USA) was used for statistical processing. Differences between groups were assessed using unpaired t-tests, paired t-tests, Mann Whitney U-tests, or one-way analysis of variance (ANOVA), followed by the Bonferroni test for post hoc multiple comparisons. The chisquare test was used for categorical data. Univariate logistic regression was used to determine the risk factors associated with regression of glucose metabolism disorders. A multivariate Cox proportional hazards model was used to determine independent predictive factors of mortality. Cumulative survival was analyzed using the Kaplan-Meier method, and the curves were compared using the log-rank test. P<0.05 was considered statistically signi cant. Patients with diabetes had signi cantly lower HOMA2-β levels than did patients with NGT (Table 1). No other clinical or laboratory parameters differed signi cantly among patients with NGT, IGT and diabetes. Patients with hypoglycemia had signi cantly higher HOMA2-IS, HOMA2-β, alanine aminotransferase, aspartate aminotransferase, and TBil levels and MELD scores and signi cantly lower HOMA2-IR, FINS and FCP levels than did patients without hypoglycemia (Supplementary Table 1).  Table 2). Eight patients had regression of their glucose metabolism disorders (2 with diabetes and 1 with IGT regressed to NGT, 5 with diabetes regressed to IGT). Fourteen patients did not regress (one progressed from IGT to diabetes; the others remained unchanged; Fig. 1). Patients whose glucose metabolism disorders regressed had signi cantly higher platelet (PLT) and albumin (ALB) levels and a signi cantly lower prevalence of Child-Pugh class C liver cirrhosis than did patients without regression when the OGTT was repeated (   and diabetes (OR = 3.601, 95% CI: 1.342-9.661) were the independent risk factors associated with death in patients with chronic HBV-related ACLF. Patients with both chronic HBV-related ACLF and diabetes had signi cantly higher mortality than did those with only HBV-related ACLF (Fig. 2).

Discussion
In patients with ACLF, acute and chronic liver dysfunction may result in HD, and severe liver and systemic in ammation may result in stress-induced hyperglycemia. Hypoglycemia occurs frequently in both patients with HD and patients with acute stress. Clinically, HD is di cult to distinguish from stressinduced hyperglycemia, and HD-related hypoglycemia is di cult to distinguish from hypoglycemia induced by acute stress. In this study, we excluded patients with pre-existing diabetes. Glucose metabolism disorders in the patients in this study were mostly due to severe liver injury and severe systemic in ammation. Most patients with diabetes had normal FPG, and most patients with hypoglycemia had diabetes or IGT, demonstrating that various glucose metabolism disorders existed in patients with ACLF.
Previous studies have found that both pancreatic β-cell dysfunction and increased insulin resistance play central roles in HD development in patients with liver cirrhosis [24]. Stress-induced hyperglycemia is also considered a result of pancreatic failure in compensating for increased insulin production and release [25]. In the current study, patients with diabetes had signi cantly lower HOMA2-β levels than did patients with NGT, demonstrating that an impaired pancreatic β-cell response to acute liver in ammation and damage plays a determining role in the development of diabetes in patients with ACLF.
In patients with HD or stress-induced hyperglycemia, managing the hyperglycemia remains a clinical challenge. Hyperglycemia increased the mortality rates of patients with liver cirrhosis and critically ill patients. However, strictly controlling patients' hyperglycemia increased the prevalence of hypoglycemia, which was also associated with high mortality [26]. Most patients with ACLF have severe digestive symptoms, making it impossible to perform the OGTT, and managing diabetes in patients with ACLF is especially di cult. Therefore, the dynamic changes in glucose metabolism disorders in patients with ACLF must be better understood. In the current study, eight of 22 patients (36.4%) with ACLF exhibited regressed glucose metabolism disorders and improved liver function during the 32.8 ± 28.8-day follow-up.
PLT levels were positively associated with the regression. Our results indicated that in patients with ACLF and sustained low PLTs, OGTTs should be repeated to evaluate the regression of the glucose metabolism disorders.
One major nding in this study was that diabetes was an independent risk factor for 90-day mortality in patients with ACLF. Previous studies showed that hyperglycemia accelerated the progression of liver brosis and provoked liver in ammation [27]. Several clinical studies demonstrated that HD or preexisting diabetes carried high risks for poor long-term outcomes and complications in patients with liver cirrhosis [16, 28]. One study evaluated the association of HD and 30-day survival in 78 patients with decompensated liver cirrhosis and found that HD and IGT better predicted a poor 30-day survival than did NGT [18]. However, in another study, Bianchi et al. found no association between diabetes and short-term mortality in patients with liver cirrhosis [17]. The reason for the con icting results in these studies is unknown. Bianchi et al. did not exclude pre-existing diabetes or perform OGTTs in their study; therefore, the diabetes prevalence might have been underestimated. In this study, we excluded pre-existing diabetes and performed OGTTs in 96 patients with ACLF. We found that diabetes resulted from severe liver injury and liver in ammation as one independent risk factor of 90-day mortality in patients with ACLF.
Hypoglycemia was not a risk factor for 90-day mortality in this study. Patients with ALF are generally considered to be at high risk for hypoglycemia [10,29]; however, to our knowledge, the correlation between hypoglycemia and mortality in patients with ACLF remains unknown. One retrospective study found that hypoglycemia was a risk factor for poor prognosis in patients with decompensated liver cirrhosis [30]; however, no OGTT was performed in that study. Because the diagnosis of HD in patients with chronic liver diseases depends on OGTT results, and patients with HD are prone to hypoglycemia. Whether the effects of hypoglycemia found in that study actually resulted from the effects of HD requires further analysis. In the current study, the prevalence of hypoglycemia was much lower than that of diabetes, and most patients with hypoglycemia had diabetes. Diabetes was only the risk factor associated with death in patients with ACLF, indicating that diabetes had a worse effect on patients' prognosis than did hypoglycemia in patients with ACLF.
Our study had several limitations. First, although it included the largest cohort to date to undergo OGTTs to analyze the effects of glucose metabolism disorders on the prognosis of patients with ACLF, it was a single-center study, and we could not perform OGTTs on some severe patients with ACLF. Thus, the number of patients included was relatively small, and the results should be veri ed in multicenter studies with more patients. Second, we evaluated only the association between baseline hyperglycemia and mortality, and whether the changes in glucose metabolism disorders after admission and the different methods for managing hyperglycemia in uence patient outcomes remains to be clari ed.
In conclusion, this was the rst study to evaluate the effects of glucose metabolism disorders on the prognosis of patients with chronic HBV-related ACLF. The major nding in this study was that diabetes, but not hypoglycemia, is associated with high mortality in patients with ACLF. Dynamic changes in glucose metabolism disorders in patients with acute-on-chronic liver failure NGT, normal glucose tolerance; IGT, impaired glucose tolerance Cumulative survival rates of patients with acute-on-chronic liver failure with and without diabetes

Supplementary Files
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