第 14回国際内分泌学会　京都　３月 28日

Journal ClubJournal Club

埼玉医科大学　総合医療センター　内分泌・糖尿病内科埼玉医科大学　総合医療センター　内分泌・糖尿病内科Department of Endocrinology and Diabetes, Department of Endocrinology and Diabetes,

Saitama Medical Center, Saitama Medical UniversitySaitama Medical Center, Saitama Medical University

松田　昌文松田　昌文Matsuda, MasafumiMatsuda, Masafumi

20102010年４月１日　年４月１日　 8:30-8:558:30-8:55８階　医局８階　医局

Sone H, Tanaka S, Iimuro S, Tanaka S, Oida K, Yamasaki Y, Oikawa S, Ishibashi S, Katayama S, Yamashita H, Ito H, Yoshimura Y, Ohashi Y, Akanuma Y, Yamada N; Japan Diabetes Complications Study Group.Long-term lifestyle intervention lowers the incidence of stroke in Japanese patients with type 2 diabetes: a nationwide multicentre randomised controlled trial (the Japan Diabetes Complications Study).Diabetologia. 2010 Mar;53(3):419-28.

Petersen KF, Dufour S, Hariri A, Nelson-Williams C, Foo JN, Zhang XM, Dziura J, Lifton RP, Shulman GI.Apolipoprotein C3 gene variants in nonalcoholic fatty liver disease.N Engl J Med. 2010 Mar 25;362(12):1082-9.

H. Sone : N. Yamada : Department of Internal Medicine, University of Tsukuba Institute of Clinical Medicine, Ibaraki, Japan S. Tanaka Laboratory of Biostatistics, Tokyo University of Science, Tokyo, Japan S. Iimuro : Y. Ohashi Department of Biostatistics, University of Tokyo School of Medicine, Tokyo, Japan S. Tanaka Translational Research Center, Kyoto University, Kyoto, Japan K. Oida Fukui Chuo Clinic, Fukui, Japan Y. Yamasaki Center for Advanced Science and Innovation, Osaka University, Osaka, Japan S. Oikawa Department of Medicine, Nippon Medical School, Tokyo, Japan S. Ishibashi Department of Endocrinology and Metabolism, Jichi Medical College, Tochigi, Japan S. Katayama The Fourth Department of Medicine, Saitama Medical School, Saitama, Japan H. Yamashita Department of Ophthalmology, Yamagata University School of Medicine, Yamagata, Japan

Aim

The aim of the study was to clarify whether a therapeutic intervention focused on lifestyle modification affected the incidence of vascular complications in patients with established diabetes.

MethodA total of 2,033 eligible Japanese men and women aged 40–70 years with type 2 diabetes from 59 institutes were randomised to a conventional treatment group (CON), which continued to receive the usual care, and a lifestyle intervention group (INT), which received education on lifestyle modification regarding dietary habits, physical activities and adherence to treatment by telephone counselling and at each outpatient clinic visit, in addition to the usual care. Randomisation and open-label allocation were done by a central computer system. Primary analysis regarding measurements of control status and occurrence of macro- and microvascular complications was based on 1,304 participants followed for an 8 year period.

Patients in the INT group also received 15 min telephone counselling sessions at least once every 2 weeks

previously diagnosed patients with type 2 diabetes aged 40–70 years whose HbA1c levels were ≥6.5%.

Initially registered from January 1995 to March 1996.

Goals were set for patients in the INT group and their physicians: i.e. HbA1c level <6.5%; BMI <22 kg/m2; BP <140/85 mmHg; serum cholesterol level <5.72 mmol/l (220 mg/dl) ; serum triacylglycerol level <1.65 mmol/l (150 mg/dl) ; serum HDLcholesterol >1.04 mmol/l (40 mg/dl); WHR <0.9 for men and <0.8 for women; smoking cessation; and abstinence from alcohol. Goals regarding BP and serum cholesterol levels were updated in accord with the revision of guidelines made by the Japan Diabetes Society, which were <130/80 mmHg and <5.17 mmol/l (200 mg/dl), respectively.

a duration of 10.9±7.2 years (both mean ± SD)

Chol 5.2 mmol/L (200 mg/d) TG 1.19 mmol/L (105 mg/dl) 1.09 ⇒ 95

Fig. 2 Kaplan–Meier curves for each complication. a Nephropathy, p=1.00. b Retinopathy, p=0.43. c CHD, p=0.40. d Stroke, p=0.02. p values by logrank test. Dotted curves, CON; solid curves, INT

Nephropathy Retinopathy

CHD Stroke 83% (n=75) were brain infarction, 9% (n=8) were brain haemorrhage and 8% (n=7) were transient ischaemic attack.

Δ ~－40%

All significant variables selected for the univariate analysis with the criterion of a p<0.1 were used in the multivariate analysis

Table 2 Risk factors for stroke analysed by Cox univariate and multivariate models

The mechanism of this apparent contradictory result is yet to be determined but it should be interpreted with care, especially since BP, which is a major risk factor for stroke, did not differ significantly between groups throughout the study period. Multifactorial or combined effects of lifestyle education/behaviours beyond individual factors [31] might have existed but can only be speculated upon. At the same time, the slight but significant differences in HbA1c in the first 3 years, which was reported previously [21], but that disappeared thereafter, could enhance the effects since past interventions to lower HbA1c reportedly have had a very long-term effect (i.e. ‘metabolic memory’ or ‘legacy effect’) [32, 33]. Other speculations for the apparent contradictory result include possible improvement in factors that were not determined in this study, such as postprandial glycaemia/ lipaemia, BP at home or psychological factors (stress, motivation or quality of life) [34], which could be ameliorated in the INT group rather than in the CON group. For example, Roumen et al. [35] recently reported that a lifestyle intervention successfully improved postprandial glucose levels in IGT patients. Changes in diet might also be effective, such as an increase in fruit intake, which is reportedly associated with reduced CVD mortality [36]. The reasons that only stroke, but not CHD or other complications, was found to be responsive to our intervention are speculated to include the following: (1) stroke is more frequent than CHD in Japan compared with other parts of the world, and (2) the independent risk factor for stroke was only systolic BP and lipoprotein(a), and so there would be room for other undetermined risk factors to work.

ResultsAlthough status of control of most classic cardiovascular risk factors, including body weight, glycaemia, serum lipids and BP, did not differ between groups during the study period, the incidence of stroke in the INT group (5.48/1,000 patient-years) was significantly lower than in the CON group (9.52/1,000 patient-years) by Kaplan– Meier analysis (p=0.02 by log rank test) and by multivariate Cox analysis (HR 0.62, 95% CI 0.39–0.98, p=0.04). The incidence of CHD, retinopathy and nephropathy did not differ significantly between groups. Lipoprotein(a) was another significant independent risk factor for stroke.

ConclusionThese findings suggest that lifestyle modification had limited effects on most typical control variables, but did have a significant effect on stroke incidence in patients with established type 2 diabetes.

First, our participants were hospital-based patients with diabetes of a relatively long duration. Therefore, we cannot make inferences beyond a similar group. Second, only Asian diabetic patients were involved and they are different from other ethnic groups in terms of degree of obesity. Third, we had a low follow-up rate, since the study was done mainly in large hospitals in urban areas where patients move quite frequently.

Limitations

Message

JDCSは生活介入で HbA1C(JDS)に差をつけるはずが，つかなかった。[ 最初の３年はついたのだが ]

ともあれ，理由は不明だが　血糖をよくしてやろうとすれば　脳卒中が減少した！？　で，この時に違ったのは　電話介入くらいだった。

N Engl J Med 2010;362:1082-9.

From the Departments of Internal Medicine (K.F.P., A.H., X.-M.Z., J.D., R.P.L., G.I.S.), Genetics (C.N.-W., J.N.F., R.P.L.), and Cellular and Molecular Physiology (G.I.S.), and the Howard Hughes Medical Institute (S.D., R.P.L., G.I.S.), Yale University School of Medicine, New Haven, CT.

BackgroundNonalcoholic fatty liver disease is associated with hepatic insulin resistance and type 2 diabetes mellitus. Whether this association has a genetic basis is unknown.

Asian Indian

MethodIn 95 healthy Asian Indian men, a group known to have a high prevalence of nonalcoholic fatty liver disease, we genotyped two single-nucleotide polymorphisms (SNPs) in the gene encoding apolipoprotein C3 (APOC3) that are known to be associated with hypertriglyceridemia (rs2854116 [T-455C] and rs2854117 [C-482T]). Plasma apolipoprotein C3 concentrations, insulin sensitivity, and hepatic triglyceride content were measured. We also measured plasma triglyceride concentrations and retinyl fatty acid ester absorption as well as plasma triglyceride clearance after oral and intravenous fat-tolerance tests. Liver triglyceride content and APOC3 genotypes were also assessed in a group of 163 healthy non–Asian Indian men.

VLDL

Genomic DNA was extracted from whole blood using the SDS/Proteinase-K, Phenol-chloroform method. Genotypes at SNP’s flanking the transcription start site of Apo C3 (position -455, rs2854116, and position -482, rs2854117) as well as Apo A5 (position -3, rs651821) were determined by amplifying the region encompassing the polymorphic sites followed by direct sequencing.

The segment of Apo C3 was amplified by polymerase chain reaction using the following primers: 5’GAAGGTGAACGAGAATCAGTCCTG3’ and 5’GCCTCGGGCCCATCTCAGCCTTTCACACTG 3’).

The segment of Apo A5 was amplified using the primers: 5’GACCTTTGCGAAGGGGTTAGAGCACCAC3’ and 5’CTCGCGAGCCATCTTCTGCTGATGGATC3’. Polymorphic SNPs in the PNPLA3 gene (GCCACTGTAGAAGGGG/CATGAAGCAGGAACAT), encompassing the I148M variant (rs738409) were amplified using primer set PNPLAV1F: 5’-GAGCCAACAA CCCTTGGTCC TGTCTG- and PNPLAV1R: 5'-GCTGCCCGG GTAGCCTGGA AATAG-3'. PCR fragments were sequenced with PNPLAV1F to determine the genotype.

Genotyping

Measurement of Hepatic Triglyceride ContentHepatic triglyceride content was measured with proton magnetic resonance spectroscopy on a BioSpec 4T system (Bruker Daltonics). Localized spectra of the liver were obtained with the use of a coil assembly composed of twin elliptical proton radiofrequency coils (13 cm by 9 cm) arranged in quadrature. Triglyceride content was measured by proton respiration-gated, stimulated echo acquisition mode spectroscopy in a voxel (15 mm by 15 mm by 15 mm) placed in three different locations within the liver to account for the heterogeneity of hepatic tissue.

(108 of whom were white, 26 Asian, 15 Hispanic, and 14 black)

due to decreased lipoprotein lipase activity

Figure 1. Scattergrams of Liver Triglyceride Content in Asian Indian and Non–Asian Indian Men. Carriers of APOC3 variant alleles (C-482T, T-455C, or both) were compared with APOC3 wild-type homozygotes (C-482 and T-455) in terms of liver triglyceride content. Results are shown for two groups of Asian Indian men (Panel A) and for two groups of non–Asian Indian men (Panel B). The boxes delineate subjects who had nonalcoholic fatty liver disease (hepatic lipid content >5.5%), all of whom were carriers of the variant allele. (insulin-sensitivity index, 2.0±2.0 [lower-third percentile for insulin sensitivity9], vs. 3.5±1.6; P<0.001).

Figure 2. Plasma Triglyceride and Retinyl Fatty Acid Ester Concentrations after an Oral Fat-Tolerance Test in Asian Indian Men. Panel A shows the area under the curve (AUC) for plasma triglyceride concentrations after an oral fat-tolerance test in 28 carriers of APOC3 variant alleles (C-482T, T-455C, or both) as compared with 10 APOC3 wild-type homozygotes. Data are means ±SE.

A liquid meal containing; 20.9 grams of protein, 76.2 grams of fat, 61 grams of carbohydrate and 60,000 IU/ m2 of Vitamin A was administered

Figure 2. Plasma Triglyceride and Retinyl Fatty Acid Ester Concentrations after an Oral Fat-Tolerance Test in Asian Indian Men. Panel B shows the AUC for plasma retinyl ester concentrations in the two groups. Data are means ±SE.

Retinyl fatty acid ester concentrations were measured in order to assess chylomicron and post chylomicron remnant metabolism

A bolus infusion of Liposyn (20%, 0.5 ml/kg, Abbott Laboratories, North Chicago, IL) per kg body weight was administered over 2 minutes

Individual SNPs and their respective effects on Apo C3 concentrations

Variant alleles = total # high risk alleles (T482 or C455 alleles) Variant haplotypes = total # haplotypes carrying at least one risk allele (C482-C455 or T482-C455; the T482-T455 haplotype was not observed in this cohort) Variant loci = # sites with the high risk allele present

Clinical characteristics of study participants by genotype classification

?

Effect of Weight Loss on Hepatic Steatosis and Insulin Resistance Seven of the subjects with hepatic steatosis and insulin resistance underwent a hypocaloric dietary intervention. Over a period of 3 to 6 months, their average weight decreased from 80.4±10.7 kg to 74.9±10.2 kg (P = 0.003). This weight loss was accompanied by a significant reduction in the liver triglyceride content (from 14.0 to 3.8%, P = 0.05). After weight loss, there was a marked improvement in insulin sensitivity on oral glucose-tolerance testing, with reductions in plasma glucose and plasma insulin concentrations and an increase in the insulin-sensitivity index (from 1.8±0.8 to 3.7±1.4, P<0.01).

The APOC3 variants C-482T and T-455C lead to in-creased plasma concentrations of apolipoprotein C3, which in turn inhibit lipoprotein lipase and triglyceride clearance, thus conferring a predisposition to both fasting? and postprandial hypertriglyceridemia due to an increase in chylomicronremnant particles. Increased concentrations of circulating chylomicron-remnant particles are then preferentially taken up by the liver by means of a receptor-mediated process, resulting in nonalcoholic fatty liver disease and hepatic insulin resistance.

DISCUSSION

ResultsCarriers of the APOC3 variant alleles (C-482T, T-455C, or both) had a 30% increase in the fasting plasma apolipoprotein C3 concentration, as compared with the wild type homozygotes. They also had a 60% increase in the fasting plasma triglyceride concentration, an increase by a factor of approximately two in the plasma triglyceride and retinyl fatty acid ester concentrations after an oral fat-tolerance test, and a 46% reduction in plasma triglyceride clearance. The prevalence of nonalcoholic fatty liver disease was 38% among variant-allele carriers and 0% among wild-type homozygotes (P<0.001). The subjects with nonalcoholic fatty liver disease had marked insulin resistance. A validation study involving non–Asian Indian men confirmed the association between APOC3 variant alleles and nonalcoholic fatty liver disease.

Conclusion

The polymorphisms C-482T and T-455C in APOC3 are associated with nonalcoholic fatty liver disease and insulin resistance.

Message高中性脂肪の患者で脂肪肝の場合にこの論文を思い出しましょう。 ( 「あなたの　 ApoC3　ちょっとおかしいもかも」と患者さんに言ってもよいかもしれません。 )

ともかく 5kgやせるだけで肝臓の TG含量が 14%から 3.8%に減っています！　インスリン感受性(Matsuda Index)が 1.8から 3.7に上昇します。