Sage Journals: Discover world-class research

Abstract

Background

We previously reported that the indicator of glycaemic control, glycated albumin (GA) levels, are low in relation to glycaemia in patients with high alanine aminotransferase (ALT) levels in non-alcoholic fatty liver disease because of chronic inflammation, and that the GA/glycated haemoglobin ratio (G/H ratio) is inversely correlated with hepatic function in patients with chronic liver disease. The severity of liver fibrosis is known to be a good indicator for surveillance, and for determining the prognosis and optimal treatment of non-alcoholic steatohepatitis (NASH). In this study, we aimed to investigate the clinical usefulness of measuring the G/H ratio for predicting the severity of liver fibrosis in patients with NASH.

Methods

The study subjects were 36 patients with histologically diagnosed NASH (19 men, 17 women; mean age54.8 ± 12.2 years, body mass index 28.3 ± 5.0 kg/m²). The relationships of the G/H ratio to hepatic function tests and fibrosis stage in the liver were investigated.

Results

The G/H ratio in patients with NASH was inversely correlated with ALT (P < 0.001) and platelet count (P < 0.0001). Furthermore, the G/H ratio was positively correlated with the fibrosis stage in liver (P = 0.003).

Conclusions

These results suggest that the G/H ratio increases along with the fibrosis stage in patients with NASH.

Introduction

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease, affecting up to 20% of the population in Western countries, and 70–80% of obese individuals.^1,2 The incidence of NAFLD is increasing due to the ongoing epidemic of its two major risk factors, obesity and type 2 diabetes mellitus, related to a sedentary lifestyle and inadequate dietary choices. It encompasses a spectrum of distinct histological entities with different natural histories and outcomes, ranging from simple fat accumulation in hepatocytes to liver steatosis accompanied by a microinflammatory component that may have associated fibrosis. In contrast to simple steatosis, which is defined as a benign form of NAFLD with minimal risk of progression, non-alcoholic steatohepatitis (NASH) can progress to liver cirrhosis in up to 20% of patients and subsequently lead to hepatic failure or hepatocellular carcinoma.^3,4

Diabetic patients are known to show greater glycation of various proteins than non-diabetic subjects and it has been suggested that some of these glycated proteins are involved in the onset and development of the chronic complications of diabetes.⁵ One of these glycated proteins, glycated haemoglobin (HbA_1c), is often used in clinical settings as an indicator of glycaemic control.^6,7 Since the lifespan of erythrocytes is approximately 120 days, HbA_1c reflects the glycaemic control status of the previous 2–3 months. However, measurement of HbA_1c is affected in diseases in which the erythrocyte lifespan is shortened, such as haemolytic anaemia and renal anaemia, and with variant haemoglobins, and thus HbA_1c does not accurately reflect the glycaemic control status in such cases.^8,9

Glycated albumin (GA) is used as another glycaemic control indicator.⁶ Since the half-life of serum albumin is shorter than that of erythrocytes, at 14 days, GA reflects plasma glucose concentrations over a shorter period (approximately two weeks). Moreover, GA has a benefit in that its measurements are not affected by haemoglobin disorders.¹⁰ However, measurements of GA are affected in patients with disorders of albumin metabolism, such as in nephrotic syndrome or thyroid dysfunction, and do not correctly reflect the glycaemic control status in such cases.¹¹

It has been recently demonstrated that GA is low relative to plasma glucose concentrations in obese patients with diabetes.¹² We observed a significant negative correlation between body mass index and GA even in non-diabetic subjects.¹³ We also demonstrated a significant negative correlation between highly sensitive C-reactive protein (hs-CRP) and GA, and that hs-CRP was a significant negative explanatory variable for GA in a multivariate analysis.¹³ From the finding that hs-CRP is elevated as a result of accelerated release of various cytokines in obesity, we hypothesized that the negative regulatory mechanism of GA resulting from obesity is based on accelerated albumin catabolism accompanied by chronic microinflammation.¹³ We have also reported low levels of GA by similar mechanisms in smoking,¹⁴ hyperuricaemia¹⁵ and hypertriglyceridaemia,¹⁶ which are representative conditions causing elevated hs-CRP. High levels of hs-CRP have also been reported in NAFLD patients with high alanine aminotransferase (ALT).^17,18 We reported GA concentrations to be low in relation to glycaemia in patients with high ALT without a drinking habit.¹⁹

It has been demonstrated that patients with chronic liver disease (CLD) have apparently low HbA_1c ^20,21 due to a shortened lifespan of erythrocytes that is caused by hypersplenism. Furthermore, since the ability of albumin synthesis is impaired and the half-life of serum albumin is prolonged in patients with CLD,²² their GA concentrations appear to be high relative to glycaemia.²³ Therefore, the GA/HbA_1c ratio (G/H ratio) in patients with CLD was shown to be correlated with hepatic function irrespective to glycaemia.²⁴

The severity of liver fibrosis is known to be a good indicator for surveillance, and for determining the prognosis and optimal treatment of NASH. In this study, we aimed to investigate the clinical usefulness of measuring the G/H ratio for predicting the severity of liver fibrosis in patients with NASH.

Methods

Study patients

GA and HbA_1c were simultaneously measured in 36 patients with NASH (Table 1) undergoing treatment at Fukui-ken Saiseikai Hospital or Hyogo College of Medicine. Among the 36 patients, 26 patients (72%) displayed complications of type 2 diabetes mellitus. Patients with unstable glycaemic control were excluded. The diagnosis of NASH was made histologically, and the fibrosis stage was classified as S1–S4 according to the method of Brunt et al. ²⁵This study was approved by the ethics committee at Fukui-ken Saiseikai Hospital and Hyogo College of Medicine. The purpose of the study was explained to all patients, and all patients provided their written informed consent.

Table 1

Clinical characteristics of the study patients classified by the stage of fibrosis*

	S1	S2	S3	S4	P
n (M/F)	7 (5/2)	17 (8/9)	6 (3/3)	6 (3/3)	0.546
Age (y)	53.0 ± 10.3	50.2 ± 3.1	63.5 ± 4.0	60.8 ± 3.6	0.051
BMI (kg/m²)	28.7 ± 3.6	28.9 ± 5.7	26.9 ± 4.8	27.5 ± 5.3	0.099
DM (%)	3 (43)	12 (71)	5 (83)	5 (83)	0.252
Serum ALT (U/L)	80.9 ± 40.0	58.1 ± 30.3	51.2 ± 25.0	42.2 ± 16.9	0.026
Platelet (×10³/μL)	20.9 ± 3.1	25.5 ± 5.1	18.6 ± 5.7	11.8 ± 1.4	0.001
Hb (g/dL)	15.2 ± 1.7	14.5 ± 1.0	13.5 ± 1.6	12.6 ± 1.6	0.001
HbA_1c (%)	6.4 ± 0.7	6.3 ± 0.7	7.6 ± 1.3	7.0 ± 1.6	0.077
GA (%)	15.5 ± 2.5	14.8 ± 2.4	20.0 ± 3.4	19.5 ± 5.4	0.004
G/H ratio	2.43 ± 0.25	2.37 ± 0.28	2.62 ± 0.13	2.79 ± 0.27	0.003

BMI, body mass index; DM, diabetes mellitus; ALT, alanine aminotransferase; Hb, haemoglobin; HbA_1c, glycated haemoglobin; GA, glycated albumin; G/H, GA/HbA_1c

*The fibrosis stage was classified as S1–S4 according to the method of Brunt et al. ²⁵

Laboratory measurements

HbA_1c was measured by high-performance liquid chromatography, with calibration using Japan Diabetes Society (JDS) Lot 2.²⁶ The value for HbA_1c (%) was estimated as a National Glycohemoglobin Standardization Program (NGSP) equivalent value (%) calculated using the formula HbA_1c (%) = HbA_1c (Japan Diabetes Society: JDS) (%) + 0.4%, considering the relational expression of HbA_1c (JDS) (%) measured using previous Japanese standard substances and measurement methods and HbA_1c (NGSP).²⁷ Serum GA was determined by enzymatic methods using albumin-specific protease, ketoamine oxidase and albumin assay reagent (Lucica GA-L; Asahi Kasei Pharma, Tokyo, Japan).²⁸ GA was hydrolysed to amino acids by an albumin-specific proteinase and then oxidized by ketoamine oxidase to produce hydrogen peroxide, which was measured quantitatively.

Statistical analysis

All data are shown as mean ± SD. The significance of the differences between the two groups was calculated using Student's t-test. To evaluate the relationship between the G/H ratio and other variables, Spearman's correlation coefficient was performed. The relationship between the stages of fibrosis on histology and the G/H ratio was tested by analysis of variance with StatView (Version 5.0 for Windows; Abacus Concepts, Berkeley, CA, USA) software. A P value <0.05 was considered significant.

Results

The clinical characteristics of the study patients classified by the stage of fibrosis are shown in Table 1. Serum ALT, platelet count and haemoglobin decreased with the progression of fibrosis. On the other hand, higher GA, but not HbA_1c, was seen with the progression of fibrosis. Higher G/H ratios were also seen with the progression of fibrosis (Table 1). A significant inverse correlation was seen between the G/H ratio and serum ALT in patients with NASH (R = −0.572, P < 0.001) (Figure 1a). The G/H ratio was also inversely correlated with platelet count (R = −0.663, P < 0.0001) (Figure 1b).

Figure 1

Correlation between the GA/HbA_1c (G/H) ratio and serum ALT (a) and platelet count (b) in patients with NASH. HbA_1c, glycated haemoglobin; GA, glycated albumin; NASH, non-alcoholic steatohepatitis; ALT, alanine aminotransferase

Discussion

A significant inverse correlation was seen between the G/H ratio and serum ALT in patients with NASH. We previously reported that serum ALT showed significant inverse correlations with both GA and the G/H ratio in patients with NAFLD.¹⁹ In those patients, hs-CRP had a significant positive correlation with serum ALT and a significant inverse correlation with GA,¹⁹ indicating the possibility that GA values declined as a result of accelerated albumin metabolism due to chronic inflammation²⁹ in patients with NAFLD with high serum ALT. This suggests that GA concentrations are set lower relative to plasma glucose concentrations because of hepatic inflammation in the early stages of NASH, resulting in lower G/H ratios.

The G/H ratio in patients with NASH showed a significant inverse correlation with platelet count, which reflects hepatic function. A significant increase in the G/H ratio was also seen with progressing fibrosis stage histologically. While there was an apparent decrease in HbA_1c and an apparent increase in GA in patients with CLD, we previously showed that these indicators diverged markedly with the state of glycaemic control as hepatic function decreased.²³ We also showed that the G/H ratio was related to hepatic function irrespective to glycaemia in patients with CLD.²⁴ Therefore, it is thought that in patients with NASH, hepatic function declines with the progression of histological findings including fibrosis, finally reaching a state of cirrhosis, while the G/H ratio becomes higher.

The above results are thought to indicate that the G/H ratio in patients with NASH declines in the early stage of the disease, but increases with the progression of the disease, and becomes high in the final stage. These results suggest that the G/H ratio is associated with the progression of fibrosis in patients with NASH. Since other fibrosis markers (hyaluronic acid and type IV collagen) were measured only in a small number of the patients in this study, statistical analysis was difficult to compare the G/H ratio with these markers. The relation between the G/H ratio and such fibrosis markers will need to be studied in future.

DECLARATIONS

Competing interests: None.

Funding: No funding was required in this research.

Ethical approval: The ethics committee of Fukui-ken Saiseikai Hospital and Hyogo College of Medicine approved this study.

Guarantors: KaN and S-hN.

Contributions: MK wrote the first draft of the manuscript. YB, DT, KaN, NT, S-hN, KeN, HN and SK edited and contributed to the final manuscript. HK, KA, NT and HE looked after the patients. All authors reviewed and edited the manuscript and approved its final version.

Acknowledgements: None.

References

Chitturi

, Farrell

, Hashimoto

, Saibara

, Lau

, Sollano

. Non-alcoholic fatty liver disease in the Asia-Pacific region: definitions and overview of proposed guidelines. J Gastroenterol Hepatol 2007;22:778–87

Amarapurkar

, Hashimoto

, Lesmana

, Sollano

, Chen

, Goh

. How common is non-alcoholic fatty liver disease in the Asia-Pacific region and are there local differences? J Gastroenterol Hepatol 2007;22:788–93

Matteoni

, Younossi

, Gramlich

, Boparai

, Liu

, McCullough

. Nonalcoholic fatty liver disease: a spectrum of clinical and pathological severity. Gastroenterology 1999;116:1413–9

Farrell

, Larter

. Nonalcoholic fatty liver disease: from steatosis to cirrhosis. Hepatology 2006;43 (Suppl 1):S 99–112

Cohen

. Nonenzymatic glycation: a central mechanism in diabetic microvasculopathy? J Diabet Complications 1998;2:214–7

Koenig

, Peterson

, Jones

, Saudek

, Lehrman

, Cerami

. Correlation of glucose regulation and hemoglobin A_Ic in diabetes mellitus. N Engl J Med 1976;295:417–20

Bunn

, Gabbay

, Gallop

. The glycosylation of hemoglobin: relevance to diabetes mellitus. Science 1978;20:21–7

Jeffcoate

. Diabetes control and complications: the role of glycated haemoglobin, 25 years on. Diabetic Med 2004;21:657–65

Bry

, Chen

, Sacks

. Effects of hemoglobin variants and chemically modified derivatives on assays for glycohemoglobin. Clin Chem 2001;47:153–63

10.

Guthrow

, Morris

, Day

, Thorpe

, Baynes

. Enhanced nonenzymatic glucosylation of human serum albumin in diabetes mellitus. Proc Natl Acad Sci USA 1979;76:4258–61

11.

Koga

, Kasayama

. Clinical impact of glycated albumin as an another glycemic control marker. Endocr J 2010;57:751–62

12.

Koga

, Matsumoto

, Saito

, Kasayama

. Body mass index negatively influences glycated albumin, but not glycated hemoglobin, in diabetic patients. Endocr J 2006;53:387–91

13.

Koga

, Otsuki

, Matsumoto

, Saito

, Mukai

, Kasayama

. Negative association of obesity and its related chronic inflammation with serum glycated albumin but not glycated hemoglobin levels. Clin Chim Acta 2007;378:48–52

14.

Koga

, Saito

, Mukai

, Otsuki

, Kasayama

. Serum glycated albumin levels are influenced by smoking status, independent of plasma glucose levels. Acta Diabetol 2009;46:141–4

15.

Koga

, Murai

, Saito

, Mukai

, Kasayama

. Serum glycated albumin, but not glycated hemoglobin, is low in relation to glycemia in hyperuricemic men. Acta Diabetol 2010;47:173–7

16.

Koga

, Murai

, Saito

, Mukai

, Kasayama

. Serum glycated albumin, but not glycated hemoglobin, is low in relation to glycemia in men with hypertriglyceridemia. J Diabetes Invest 2010;1:202–7

17.

Hanley

, Williams

, Festa

, Wagenknecht

, D'Agostino

Jr , Haffner

. Liver markers and development of the metabolic syndrome: the insulin resistance atherosclerosis study. Diabetes 2005;54:3140–7

18.

, Cho

, Kang

, The association between increased alanine aminotransferase activity and metabolic factors in nonalcoholic fatty liver disease. Metabolism 2006;55:1604–9

19.

Koga

, Murai

, Saito

, Mukai

, Kasayama

. Serum glycated albumin levels, but not glycated hemoglobin, is low in relation to glycemia in non-diabetic men with nonalcoholic fatty liver disease with high alanine aminotransferase levels. Clin Biochem 2010;43:1023–5

20.

Cacciatore

, Cozzolino

, Giardina

, Abnormalities of glucose metabolism induced by liver cirrhosis and glycosylated hemoglobin levels in chronic liver disease. Diabetes Res 1988;7:185–8

21.

Nomura

, Nanjo

, Miyano

, Hemoglobin A1 in cirrhosis of the liver. Diabetes Res 1989;11:177–80

22.

Stefanovic

, Bosnjakovic

, Stajnfl

, Vujnic

, Petric

, Ristic

. Characteristics of albumin turnover in different stages of cirrhosis of liver evaluated by means of radioisotope technique and analog computer analysis. Radiobio Radiother 1972;13:105–11

23.

Koga

, Kasayama

, Kanehara

, Bando

. CLD (chronic liver disease)-HbA_1c as a novel indicator for estimation of mean plasma glucose in the patients with chronic liver disease. Diabetes Res Clin Pract 2008;81:258–62

24.

Bando

, Kanehara

, Toya

, Tanaka

, Kasayama

, Koga

. Association of serum glycated albumin to glycated haemoglobin A_1c ratio with hepatic function tests in patients with chronic liver disease. Ann Clin Biochem 2009;46:368–72

25.

Brunt

, Janney

, Di Biscegli

, Neuschwander-Tetri

, Bacon

. Nonalcoholic steatohepatitis: a proposal for grading and staging the histological lesions. Am J Gastroenterol 1999;94:2467–74

26.

Tominaga

, Makino

, Yoshino

, Japanese standard reference material for JDS Lot 2 haemoglobin A_1c. I: comparison of Japan Diabetes Society-assigned values to those obtained by the Japanese and USA domestic standardization programmes and by the International Federation of Clinical Chemistry reference laboratories. Ann Clin Biochem 2005;42:41–6

27.

The Committee of Japan Diabetes Society on the diagnostic criteria of diabetes mellitus. Report of the Committee on the classification and diagnostic criteria of diabetes mellitus. J Diabetes Invest 2010;1:212–28

28.

Kouzuma

, Usami

, Yamakoshi

, Takahashi

, Imamura

. An enzymatic method for the measurement of glycated albumin in biological samples. Clin Chim Acta 2004;324:61–71

29.

Das

. Raised C-reactive protein levels in serum from smokers. Clin Chem Acta 1985;153:9–13

The glycated albumin to glycated haemoglobin ratio increases along with the fibrosis stage in non-alcoholic steatohepatitis

Abstract

Background

Methods

Results

Conclusions

Introduction

Methods

Study patients

Laboratory measurements

Statistical analysis

Results

Discussion

DECLARATIONS

References