Insulin level, cognition, liver disease and CR

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Hi All,

There are three publications that are pdf-available below and the first (1)

introduces how insulin levels, which are measured using plasma C peptide levels

as

an index, are associated with reduced cognition, the second (2) is the paper

introduced in (1) and the third (3) is a paper that correlates insulin levels

(along

with other health-related factors) with a fairly common liver disease,

non-alcoholic

steatohepatitis, which is shown in the paper to respond to CR, but to respond

better

with CR in combination with metformin treatment.

Plasma C peptide is defined as: " a byproduct of normal insulin production by the

beta cells in the pancreas. Normal values are 0.5 to 3.0 ng/ml. Normal levels

indicate that the body is still producing its own insulin. Low levels indicate

that

the pancreas is producing little or no insulin. "

Plasma C peptide, or C peptide, is better if lower for mental functions and

CRers

seem to have healthier levels. The below pdf-available papers (1, 2) may be

more

significant considering that it is not in diabetics.

The definition of steatosis is: " Fatty degeneration. " Therefore, non-alcoholic

steatohepatitis seems to be degeneration of the liver tissues, with lipid tissue

pathology.

(1) Arch Intern Med Table of Contents for July 25, 2005; Vol. 165, No. 14

In This Issue of Archives of Internal Medicine

Arch Intern Med. 2005;165:1569.

Plasma C Peptide Level and Cognitive Function Among Older Women Without Diabetes

Mellitus

The impact of high insulin levels on cognition, in the absence of diabetes

mellitus,

has not been well studied. In this investigation, Okereke et al evaluated the

relation between mid-life plasma C peptide level (representing insulin

secretion)

and later-life cognition among 718 nondiabetic women. Higher levels of insulin

secretion were consistently associated with worse performance on tests of global

cognition and verbal memory. Compared with those in the lowest quartile, women

in

the highest quartile of C peptide level were 3 times as likely to have impaired

performance on both of these measures. The authors estimated that the impact of

high

C peptide level was cognitively equivalent to aging by 5 to 6 years.

(2) Plasma C Peptide Level and Cognitive Function Among Older Women Without

Diabetes

Mellitus

Olivia Okereke; E. Hankinson; B. Hu; Francine Grodstein

Arch Intern Med. 2005;165:1651-1656.

ABSTRACT

Background Growing evidence suggests that type 2 diabetes mellitus and

hyperinsulinemia may be related to diminished cognition. To help differentiate

between the effects of diabetes and insulin, we examined the relation of insulin

to

cognitive function among nondiabetic participants of the Nurses’ Health Study.

Methods We measured the C peptide level, representing insulin secretion, in

blood

samples provided by 718 women from June 14, 1989, to October 4, 1990, when they

were

aged 61 to 69 years. We administered telephone interviews an average of 10 years

after blood collection, testing general cognition, verbal memory, category

fluency,

and attention; second cognitive assessments were conducted 2 years later. The

primary outcomes were global cognitive function across all tests and verbal

memory.

We used regression models to estimate multivariable-adjusted mean differences in

cognitive function and cognitive decline, and odds of cognitive impairment,

across C

peptide levels.

Results Cognitive function was worse among women in the fourth C peptide

quartile

compared with those in the first quartile (eg, on the global score combining all

cognitive tests, the multivariable-adjusted mean difference was –1.7 standard

units

[95% confidence interval, –2.9 to –0.6 standard units]; P = .002); the odds of

cognitive impairment (defined as the worst 10% of the distribution) were 3-fold

higher among women in the fourth vs first quartile (95% confidence interval,

1.3-7.8). On verbal memory, women in the fourth quartile scored significantly

worse

than those in the first quartile; the odds of impairment were 2.8-fold higher

(95%

confidence interval, 1.1-7.0). Consistent findings were observed for cognitive

decline.

Conclusion Higher insulin secretion may be related to worse cognition, even

among

those without diabetes.

INTRODUCTION

Decreasing Alzheimer disease is a public health priority. Identifying mutable

factors in the early stages of preclinical dementia may be critical to effective

prevention.1-2 A small impairment in cognition among healthy older individuals

strongly predicts dementia development,3-5 and may be considered a marker of

preclinical dementia.

Epidemiologic studies6 have identified type 2 diabetes mellitus as an important

possible risk factor for diminished cognition. ... Type 2 diabetes is initially

characterized by elevated insulin levels, and insulin receptors are concentrated

in

the hippocampus.8 In animal and human models,9-11 an elevated insulin level

increases the amyloid (A) level; A accumulation is implicated in the

pathogenesis

of Alzheimer disease.12

.... METHODS

....

COGNITIVE FUNCTION ASSESSMENT

From February 1995 to July 2001, Nurses’ Health Study subjects 70 years and

older,

free of diagnosed stroke, participated in a telephone cognitive assessment.

.... ASCERTAINMENT OF INSULIN SECRETION

C peptide is cleaved in a 1:1 ratio in the conversion of proinsulin to insulin

and

provides an accurate representation of insulin secretion.19-21 Although insulin

and

C peptide are secreted in equimolar amounts, C peptide is not excreted by the

liver

and its half-life in the circulation is 2 to 5 times longer than insulin22;

therefore, C peptide is a more stable indicator of insulin secretion.

.... RESULTS

There was a broad distribution of C peptide levels among our subjects (Table 1);

the

median C peptide level in the fourth quartile was more than 4 times greater than

that in the first quartile. Characteristics, including age and educational

attainment, were generally similar across C peptide quartiles. However, the

prevalence of hypertension increased with increasing C peptide level, and women

in

the third and fourth quartiles of C peptide used hormone therapy less often than

those in the lower quartiles. Compared with women in the first quartile of C

peptide, women in the fourth quartile had worse mean performance on all our

cognitive tests.

After adjustment for age and educational attainment (Table 2), we found

statistically significantly worse performance on the verbal memory and global

scores

in the second through fourth quartiles of C peptide, compared with the first

quartile. On the TICS, findings were slightly weaker, but mean scores among

women in

the fourth quartile of C peptide were significantly lower than among women in

the

first quartile. Further adjustment for various potential confounding factors had

relatively little impact on results. For example, on the global score, after

multivariable adjustment, women in the fourth C peptide quartile scored 1.7

standard

units lower than those in the first quartile (P = .002). To help interpret these

mean differences, we calculated the effect of age on cognitive performance. In

our

subjects, women 6 years apart in age had a mean difference of approximately 1.5

standard units on the global score; thus, being in the highest quartile of C

peptide

seemed cognitively equivalent to aging by 6 years.

Furthermore, multivariable-adjusted analyses of C peptide as a continuous

variable

also indicated significant trends of worse cognition with increasing C peptide

level. Each 1-SD increase in C peptide was associated with a mean difference of

–0.4

standard units (P = .03 for trend) on the global score, or the approximate

equivalent of aging by 2 years.

Table 2. Mean Differences in Cognitive Function, According to Plasma C Peptide

Quartile*

.........................................

Cognitive Test † Per SD Increase in C Peptide ‡----C Peptide Quartile

------------------------------------------1 2 3 4

.........................................

Verbal memory (n = 574)

Age and education adjusted & #8722;0.3 ( & #8722;0.5 to & #8722;0.1) 0.0 & #8722;0.9

( & #8722;1.6 to & #8722;0.3) & #8722;0.7 ( & #8722;1.3 to 0.0) & #8722;1.1 ( & #8722;1.8

to

& #8722;0.4)

Multivariable adjusted ** & #8722;0.3 ( & #8722;0.5 to 0.0) 0.0 & #8722;0.9

( & #8722;1.6

to & #8722;0.3) & #8722;0.6 ( & #8722;1.3 to 0.0) & #8722;1.0 ( & #8722;1.7 to

& #8722;0.3)

P value for trend .02 NA NA NA NA

TICS (n = 718)

Age and education adjusted & #8722;0.2 ( & #8722;0.4 to 0.0) 0.0 & #8722;0.5

( & #8722;1.0

to 0.1) & #8722;0.4 ( & #8722;0.9 to 0.2) & #8722;0.6 ( & #8722;1.2 to & #8722;0.1)

Multivariable adjusted ** & #8722;0.2 ( & #8722;0.4 to 0.0) 0.0 & #8722;0.5

( & #8722;1.1

to 0.1) & #8722;0.3 ( & #8722;0.9 to 0.3) & #8722;0.6 ( & #8722;1.2 to 0.0)

P value for trend .05 NA NA NA NA

Global score (n = 574)

Age and education adjusted & #8722;0.5 ( & #8722;0.9 to & #8722;0.1) 0.0 & #8722;1.9

( & #8722;2.9 to & #8722;0.8) & #8722;1.3 ( & #8722;2.3 to & #8722;0.2) & #8722;1.9

( & #8722;3.0 to & #8722;0.9)

Multivariable adjusted ** & #8722;0.4 ( & #8722;0.8 to 0.0) 0.0 & #8722;1.9

( & #8722;3.0

to & #8722;0.9) & #8722;1.2 ( & #8722;2.2 to & #8722;0.1) & #8722;1.7 ( & #8722;2.9 to

& #8722;0.6)

P value for trend .03 NA NA NA NA

...........................................

Abbreviations: NA, data not applicable; SD, standard deviation; TICS, Telephone

Interview for Cognitive Status.

*Data are given as mean difference (95% confidence interval) unless otherwise

indicated.

†The verbal memory score combines the results of immediate and delayed

recalls of

the East Boston Memory Test and the 10-word list; and the global score

combines the results of the TICS, the category fluency test, digit span

backward,

and immediate and delayed recalls of the East Boston Memory Test and the

10-word list.

‡The average SD for C peptide is 1.32 ng/mL (0.44 nmol/L).

**Adjusted for age (in years), education (associate’s degree, bachelor’s

degree, or

master’s or doctoral degree), high blood pressure (yes or no),

postmenopausal hormone therapy (current, past, or never), vitamin E use (yes or

no),

cigarette smoking (current, past, or never), use of antidepressants (yes or

no), and alcohol intake (in tertiles).

Analyses of the odds of cognitive impairment (Table 3) suggested the potential

clinical importance of these differences in cognitive performance. Women in the

fourth C peptide quartile had statistically significant, roughly 3-fold greater

odds

of impairment on verbal memory and the global score, compared with women in the

first quartile. In analyses of C peptide as a continuous measure, we found

significant trends of 30% to 50% increased odds of cognitive impairment with

each

1-SD increase in C peptide level on the verbal memory and global scores.

Table 3. Risk of Cognitive Impairment, According to Plasma C Peptide Quartile

............................

Cognitive Test* Per SD Increase in C Peptide †----C Peptide Quartile

-----------------------------------------1 2 3 4

.............................

Verbal memory (n = 574)

No. of cases of impairment NA 8 14 14 19

Multivariable-adjusted OR (95% CI) ‡ 1.3 (1.1-1.7) 1.0 1.9 (0.7-4.8) 1.8

(0.7-4.6)

2.8 (1.1-7.0)

P value for trend .02 NA NA NA NA

TICS (n = 718)

No. of cases of impairment NA 13 21 14 17

Multivariable-adjusted OR (95% CI) ‡ 1.1 (0.9-1.4) 1.0 1.7 (0.8-3.5) 1.0

(0.4-2.2)

1.2 (0.5-2.7)

P value for trend .40 NA NA NA NA

Global score (n = 574)

No. of cases of impairment NA 8 17 11 22

Multivariable-adjusted OR (95% CI) ‡ 1.5 (1.2-1.8) 1.0 2.3 (0.9-5.6) 1.3

(0.5-3.4)

3.2 (1.3-7.8)

P value for trend .001 NA NA NA NA

.......................................

Abbreviations: CI, confidence interval; NA, data not applicable; OR, odds ratio;

SD,

standard deviation; TICS, Telephone Interview for Cognitive Status.

*The verbal memory score combines the results of the immediate and delayed

recalls

of the East Boston Memory Test and the 10-word list; and the global

score combines the results of the TICS, the category fluency test, digit span

backward, and immediate and delayed recalls of the East Boston Memory Test and

the

10-word list.

†The average SD for C peptide is 1.32 ng/mL (0.44 nmol/L).

‡Adjusted for age (in years), education (associate’s degree, bachelor’s degree,

or

master’s or doctoral degree), high blood pressure (yes or no),

postmenopausal hormone therapy (current, past, or never), vitamin E use (yes or

no),

cigarette smoking (current, past, or never), use of antidepressants (yes or

no), and alcohol intake (in tertiles).

In a secondary analysis, we evaluated whether eventual development of diabetes

might

explain our findings. We excluded all women who developed type 2 diabetes (an

additional 35 women) between blood draw and the initial cognitive assessment,

although in fact most such women were excluded in the primary analyses because

they

would have been cases in the nested case-control study of diabetes; results

remained

identical. In a separate analysis, we further adjusted for fasting status: C

peptide

measures are equally valid in fasting and nonfasting samples, but the absolute

values of nonfasting samples may be slightly higher.27 However, these results

were

similar to primary findings, as were results excluding nonfasting samples. In

analyses adjusted for body mass index, results were attenuated, as expected;

nevertheless, there were significant differences in cognitive performance

between

those in the fourth vs first C peptide quartile (eg, on the global score, the

multivariable-adjusted mean difference was –1.3 standard units [95% confidence

interval, –2.4 to –0.1 standard units]). In addition, in a model further

adjusting

for depression using continuous scores from the mental health index, results

were

again identical. Finally, in models using covariates updated through the initial

cognitive interview, findings were unchanged.

We had a short period for measuring cognitive decline (2 years), compared with

an

average 10 years between blood collection and initial cognitive testing. Thus,

we

likely underestimate the relation of C peptide level to cognitive decline.

However,

there was a suggestion of greater decline in performance with increasing C

peptide

level, generally supporting our findings from the initial cognitive testing. On

the

global score, after multivariable adjustment, there was a mean difference in

decline

of –0.5 standard units (95% confidence interval, –1.4 to 0.4 standard units)

comparing the fourth with the first C peptide quartile, with a borderline

significant trend of worse rates of decline with increasing C peptide quartile

(P =

..1) (data not shown).

COMMENT

Among women without diabetes, we found significantly worse cognitive function

for

those with higher compared with lower levels of C peptide. These findings

persisted

after adjustment for covariates, at blood draw and subsequent to blood draw, and

persisted after exclusion of women who developed diabetes during the average 10

years between blood draw and cognitive testing. Specifically, being in the

highest C

peptide quartile seemed cognitively equivalent to aging by 6 years, and was

associated with up to a 3-fold increased odds of cognitive impairment.

There are several possible explanations for these findings. An elevated insulin

level and diminished cognition may share some common underlying cause, rather

than

having a direct relation to each other. Alternatively, higher insulin secretion,

even before the development of diabetes, may lead to vascular damage and, thus,

be

an indirect source of cognitive impairment. However, accumulating evidence

suggests

a direct link between insulin level and cognition. A high insulin level may

inhibit

neuron firing28 and decrease the activity of choline acetyltransferase,29 an

enzyme

involved in forming neurotransmitters’ regulating memory and learning. In vitro

studies11 indicate insulin causes a 3- to 4-fold increase in extracellular A

levels.

The insulin-degrading enzyme provides a further possible link between insulin

and A

levels. The insulin-degrading enzyme is primarily responsible for insulin

degradation,30 but also degrades A.31-32 In mice, insulin-degrading enzyme

deficiency was associated with greater than 50% reduction in A degradation, and

cerebral A accumulation was increased by up to 64% (the mice also developed a

diabetic phenotype).10 Insulin binds more readily to insulin-degrading enzyme30

than

other substrates, and is a competitive inhibitor of A degradation; a recent

investigation of 16 healthy older adults9 found that, on infusion of insulin,

cerebrospinal fluid A levels increased by 15% (P = .02).

Although relatively few large-scale epidemiologic studies have addressed this

issue,

existing data are consistent with our finding of a relation of elevated insulin

level to cognition in nondiabetic persons.33-38 In a prospective cohort, 297

prediabetic women (fasting glucose level, >110 but <126 mg/dL [>6.1 but <7.0

mmol/L]) had a significantly elevated risk of developing cognitive impairment

(odds

ratio, 1.64; 95% confidence interval, 1.03-2.61) compared with women with a

normal

glucose level.38 Among 386 nondiabetic men,33 those in the highest quartile of

fasting insulin level had 25% more errors on the Mini-Mental State Examination

compared with those in the lowest quartile (95% confidence interval, 4%-50%; P =

..02

for trend across quartiles). Stolk and colleagues34 observed a steady and

significant decrease in mean Mini-Mental State Examination score with increasing

postload serum insulin level among 3278 women without dementia; results remained

similar after excluding diabetic persons. Among 1897 older women without

diabetes,

there was a 30% increased risk of developing cognitive impairment with each 1%

increase in glycosylated hemoglobin level,35 a measure of glucose control.

.... Overall, increasing evidence suggests that insulin level may have a direct

effect on cognitive function. Further research is clearly necessary to confirm

findings in our study and others, and to establish whether the apparent effects

of

insulin on cognition are indeed direct. Such research could have a large

influence

on public health, especially given the growing epidemic of obesity, which is

often

accompanied by insulin resistance and increased insulin levels.

(3) Uygun A, Kadayifci A, Isik AT, Ozgurtas T, Deveci S, Tuzun A, Yesilova Z,

Gulsen

M, Dagalp K.

Metformin in the treatment of patients with non-alcoholic steatohepatitis.

Aliment Pharmacol Ther. 2004 Mar 1;19(5):537-44.

PMID: 14987322

http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve & db=pubmed & dopt=Abstra\

ct & list_uids=14987322 & query_hl=16

INTRODUCTION

Non-alcoholic steatohepatitis is a common disease that

may progress to end-stage liver disease. It is now

considered to be a widespread liver disease in Western

countries.1 Despite its common occurrence, there is no

proven pharmaceutical therapy for patients. Without

doubt, weight loss is the therapy of choice for those who

are overweight, but this requires a dramatic change in

dietary habits and lifestyle.1, 2 Therefore, it can be

achieved or maintained by only a limited number of

patients.

.... Patient selection

.... all patients underwent a detailed clinical and

dietary treatment alone and the second group of

patients was given metformin 850 mg b.d. plus dietary

treatment, for 6 months. For dietary treatment, a

dietician was consulted by all patients to restrict their

intake of lipids and non-complex carbohydrates. All

obese and overweight patients were advised to lose

weight with a restriction of daily calorie intake to

1600–1800 calories per day.

.... RESULTS

Subject characteristics

Thirty-four patients completed the study protocol suc-cessfully.

One patient in the group given dietary

treatment alone did not come to control visits regularly,

and one patient in the metformin group was excluded

from the study because of a suspicion of autoimmune

hepatitis that appeared at the third month. The

demographic characteristics of the groups are summar-ized

in Table 1. In both groups, nearly one-half of

patients reported good compliance with their dietary

recommendations, whereas the compliance was mod-erate

or poor in the others. No significant difference in

dietary compliance was observed between the two

groups. The co-morbidities associated with non-alcoholic

steatohepatitis in the group given dietary

treatment alone were as follows: obesity in five,

Table 1. Demographic characteristics of the patients

........................................

Variable DT group MDT group P

Number (n)1717

.........................................

Gender (male/female) 10/7 11/6 1

Age (years, range) 41.5±9.1 (23–61) 39.8±10.6 (22–64) 0.12

Body mass index (kg/m2) 28.4±3.9 30.1±3.4 0.09

..........................................

DT group, group given dietary treatment alone; MDT group, group

given metformin and dietary treatment.

hyperlipidaemia in three, obesity and hyperlipidaemia

in seven and undefined in two. In the metformin group,

the associated co-morbidities were as follows: obesity in

six, hyperlipidaemia in two, obesity and hyperlipidae-mia

in eight and undefined in one.

Treatment response

The results of the mean serum parameters in both

groups during the study period are shown in Tables 2

and 3. A significant decrease was noted in the mean

serum alanine aminotransferase, aspartate amino-transferase,

insulin and C-peptide levels and in the

index of insulin resistance during and at the end of

treatment in the metformin group. The rate of decrease

was found to be most prominent during the first month

of treatment, and continued to decrease during the

study period. Comparison of the mean serum alanine

aminotransferase and aspartate aminotransferase levels

and the index of insulin resistance before and after

treatment showed a strong statistical difference in

the metformin group (P <0.0001). A gradual mild

decrease was observed in the mean serum glucose, total

cholesterol and triglyceride levels during treatment

without reaching statistical significance (P > 0.05).

The mean serum alanine aminotransferase and aspar-tate

aminotransferase levels also decreased in the group

given dietary treatment alone (P=0.001 and

P=0.06, respectively), but was not as significant as

in the metformin group. The other serum parameters,

including insulin and C-peptide levels and the index of

insulin resistance, did not change significantly in the

dietary treatment group compared with the mean levels

before treatment.

For both groups, the mean (± s.d.) change in the

serum parameters, index of insulin resistance and body

mass index between time zero and the sixth month of

treatment and the percentage rate are shown in

Table 4. At the end of the sixth month, the serum

alanine aminotransferase level had reached the normal

range in 10 of 17 patients (59%) and the aspartate

aminotransferase level in 12 of 16 patients (75%) in the

Table 2. Results of biochemical parameters in the group given dietary treatment

alone (mean±s.d.)

........................................

Parameter 0 month 1 month 3 months 6 months P*

........................................

Alanine aminotransferase (U/L) 72.8±31.2 70.6±28.5 62.7±24.5 55.4±16.3 0.001

Aspartate aminotransferase (U/L) 48.1±26.3 46.3±25.8 45.5±17.8 41.3±13.5 0.06

Glucose (mg/dL) 96.4±19.2 97.3±18.8 95.2±17.3 92.7±15.7 0.04

Body mass index (kg/m2) 28.4±3.9 28.1±4.1 26.2±3.2 26.5±3.7 0.01

Insulin (lIU/mL) 13.7±4.7 13.2±4.9 13.2±5.1 11.8±4.4 0.05

C-peptide (ng/mL) 4.95±5.6 5.1±4.9 4.87±5.2 4.86±5.8 0.09

Index of insulin resistance (%) 1.83±0.74 1.82±0.81 1.85±0.72 1.81±0.67 0.18

Cholesterol (mg/dL) 197.8±39.9 191±40.1 180.1±35.6 173.4±31.7 0.01

Triglyceride (mg/dL) 203±68.5 188±54.2 184.4±49.7 185.3±51.6 0.13

...........................................

* P value shows dual comparisons between 0 and 6 months.

Table 3. Results of biochemical parameters in the group given metformin and

dietary

treatment (mean±s.d.)

..............................................

Parameter 0 month 1 month 3 months 6 months P*

..............................................

Alanine aminotransferase (U/L) 83.5±24.6 70.2±25.3 61.6±35.9 46.4±23.3 0.0001

Aspartate aminotransferase (U/L) 57.9±17.3 49.1±14.6 39.8±12.8 35.8±10.5 0.0001

Glucose (mg/dL) 87±14.2 86.5±13 84.8±8.1 80.7±6.7 0.033

Body mass index (kg/m2) 30.1±3.4 29.3±3 28.4±2.7 27.7±2.5 0.001

Insulin (lIU/mL) 12±5.1 10.4±4.6 9.06±4.3 7.1±4 0.001

C-peptide (ng/mL) 4.18±1.3 3.7±1.4 3.5±1.3 2.8±1.3 0.003

Index of insulin resistance (%) 2.53±0.98 2.2±0.9 1.86±0.76 1.38±0.71 0.0001

Cholesterol (mg/dL) 189±39.2 184±31 182±24.7 171±50 0.05

Triglyceride (mg/dL) 178±70.5 163±74.1 160±70 172±65.8 0.68

Lactic acid (mmol/L) 1.78±0.4 2.07±0.48 1.98±0.0.41 2.11±0.52 0.21

...............................................

* P value shows dual comparisons between 0 and 6 months.

metformin group, whereas the corresponding values

were six of 16 patients (37.5%) and four of 15 patients

(26%), respectively, in the group given dietary treat-ment

alone. The rate of decrease of the mean serum

insulin and C-peptide levels and the index of insulin

resistance was also more significant in the metformin

group than in the group given dietary treatment alone

(P <0.05). The body mass index decreased by more

than 1 point in 10 patients in the metformin group and

in seven patients in the dietary treatment group. The

mean body mass index had decreased significantly in

both groups by the end of the sixth month, but the

results were more prominent in the metformin group.

Thus, metformin plus dietary treatment was observed to

be more effective for the achievement of weight loss

than dietary treatment alone. The basal level of serum

lactic acid increased by more than 0.5 mmol/L in six

patients and was elevated above the upper limit in three

patients in the metformin group during treatment, but

no signs or symptoms of lactic acidosis were detected.

The mean increase in the serum lactic acid level was not

statistically significant at the end of treatment (Table 4).

Fourteen patients in the metformin group (82.5%) and

13 patients in the dietary treatment group (76.5%) had

three of the five components of the metabolic syndrome

according to ATPIII criteria. No significant difference

was observed in the response to treatment in patients

with or without metabolic syndrome in both groups. As

the number of cases without metabolic syndrome was

insufficient, no statistical comparison was performed.

Four patients in the metformin group and seven

patients in the dietary treatment group refused the

control biopsy even though written consent had been

obtained at the beginning of the study. For this reason,

control liver biopsies were performed in 13 patients in

the metformin group and in 10 patients in the dietary

treatment group at the end of treatment. The mean

histopathological scores of necro-inflammatory activity

and fibrosis before and after treatment are summarized

in Table 5. There was a slight decrease in the mean

grade of necro-inflammatory activity in the metformin

group, but this result was not statistically significant.

No change in the mean grade of necro-inflammatory

activity was detected in the group given dietary

treatment alone. The grade of necro-inflammatory

activity was unchanged in five patients, improved in

six and had progressed in two in the metformin group,

whereas it was unchanged in seven, improved in none

and had progressed in three in the dietary treatment

group. The fibrosis score was unchanged in 10 patients,

Table 4. Mean (± s.d.) changes in param-eters and percentage rate at the end of

treatment in the two groups

................................................

Parameter DT group Mean±s.d. (%) MDT group Mean±s.d. (%) P

..................................................

Alanine aminotransferase (U/L) 17.4±14.1 (24) 37.1±22.2 (44.4) 0.003

Aspartate aminotransferase (U/L) 6.8±5.9 (14) 22.1±14.3 (38) 0.0001

Glucose (mg/dL) 3.7±7.1 (3.8) 6.3±11.2 (7.2) 0.38

Body mass index (kg/m2) 1.9±2.1 (6.7) 2.4±1.9 (7.9) 0.01

Insulin (lIU/mL) 1.9±0.9 (13.8) 4.9±3.7 (40.8) 0.002

C-peptide (ng/mL) 0.09±0.2 (1.8) 1.4±1.3 (33) 0.002

Index of insulin resistance (%) 0.02±0.03 (1.1) 1.15±0.82 (45) 0.001

Cholesterol (mg/dL) 24.4±41.1 (12.3) 18.4±37.6 (9.7) 0.9

Triglyceride (mg/dL) 17.7±39.4 (8.7) 6.3±48.4 (3.5) 0.8

Lactic acid (mmol/L) 0.33±0.2 (18.5)

.....................................................

DT group, group given dietary treatment alone; MDT group, group given metformin

and

dietary treatment.

Table 5. Mean histopathological necro-inflammatory activity grade and fibrosis

score

in the two groups before and after treatment

.....................................................

------DT group MDT group

------0 month (n=17) 6 months (n=10) P 0 month (n=17) 6 months (n=13) P

......................................................

Necro-inflammatory activity 1.41±0.6 1.3±0.48 0.62 1.41±0.61 1.15±0.68 0.31

Fibrosis 1.05±1.1 1.12±1.1 0.91 0.94±1.02 0.92±1.03 0.96

.....................................................

DT group, group given dietary treatment alone; MDT group, group given metformin

and

dietary treatment.

improved in none and had progressed in three in the

metformin group, whereas it was unchanged in eight,

improved in none and had progressed in two in the

dietary treatment group. The frequency of improvement

in the necro-inflammatory activity was greater in the

metformin group than in the group given dietary

treatment alone (46% vs. 10%), but the result was not

statistically significant (P=0.17 by Fisher’s exact test).

The mean grade of steatosis by upper abdominal

ultrasonography decreased from 1.53±0.73 to

1.28±0.63 in the dietary treatment group and from

1.62±0.51 to 0.98±0.43 in the metformin group at

the end of the sixth month (P=0.17 and P=0.038,

respectively). The sonography grade showed improve-ment

in nine patients (53%) and was evaluated as

normal in five of these patients (29%) at the end of

therapy in the metformin group. A decrease in sono-graphy

grade was only detected in three patients

(17.6%) and was unchanged in the others in the

dietary treatment group.

None of the patients discontinued metformin because

of intolerability during treatment. No patient reported

symptoms of hypoglycaemia. Four patients complained

of gas and bloating and two patients of mild to moderate

abdominal pain in the first month. However, these

complaints did not require cessation of the drug.

A 6-month follow-up period in 15 patients and a 1-year

follow-up period in 11 patients were completed after

metformin treatment. The mean serum alanine amino-transferase

levels at 6 and 12 months of follow-up were

45.1±19.9 U/L and 44.2±16.7 U/L, respectively.

The mean aspartate aminotransferase levels were

37.8±13.9 U/L and 39.4±12.8 U/L, respectively.

The body mass indices were 27.1±3.1 kg/m2 and

26.7±3.9 kg/m2 at 6 and 12 months of follow-up,

respectively. No significant change in the mean alanine

aminotransferase and aspartate aminotransferase levels

or body mass index was detected during the follow-up

period. Serum alanine aminotransferase and aspartate

aminotransferase levels were elevated above the upper

limits in only one patient at follow-up, but normalized in

two other patients.

DISCUSSION

Metformin increases insulin-mediated glucose utiliza-tion

in peripheral tissues and has an anti-lipolytic effect

that lowers serum free fatty acid concentrations.17 In

this study, metformin was found to cause a decrease in

serum insulin and C-peptide concentrations by produ-cing

a significant improvement in insulin action. This

improvement probably led to a significant reduction in

the accumulation of free fatty acids in hepatocytes and

suppressed the oxidation of fatty acids contributing to

cell injury and inflammation. A significant and greater

decrease in the mean serum alanine aminotransferase

and aspartate aminotransferase levels in the metformin

group than in the group given dietary treatment alone

confirmed that the drug probably restricted the damage

to hepatocytes. The biochemical response to treatment

continued during the study period, suggesting a sus-tained

effect of the drug. In this study, the effect of

metformin treatment on hepatic histopathology was

evaluated for the first time in patients with non-alcoholic

steatohepatitis. More patients in the metfor-min

group than in the dietary treatment group showed

improved hepatic necro-inflammatory activity, with

decreased steatosis, ballooning of hepatocytes and

acinar/portal inflammation, but the difference between

the groups was not significant. No significant change

was detected in the fibrosis score of the liver at the end

of treatment in both groups.

et al. showed that metformin reversed the

insulin resistance induced by tumour necrosis factor-a

in liver cells.18 Lin et al. also suggested that the

potential mechanisms of metformin with regard to the

elimination of fat from the liver in animal models were

probably related to the inhibition of hepatic tumour

necrosis factor-a and of several tumour necrosis factor-inducible

responses promoting hepatic steatosis and

inflammation.11 In a recent study, Zhou et al. showed

that metformin regulated the adenosine monophos-phate-

activated protein kinase in hepatocytes, which is

a major cellular regulator of lipid and glucose meta-bolism.19

These authors suggested that the activation of

the adenosine monophosphate-activated protein kinase

provided a unified explanation for the beneficial effect of

metformin. All of these studies have attempted to

explain the mechanism of metformin in the liver and

have reached a consensus that metformin may prevent,

restrict or reverse hepatic steatosis and inflammation in

non-alcoholic fatty liver disease.

Older age, obesity and the presence of diabetes mellitus

have been reported to be independent predictors of more

advanced disease in patients with non-alcoholic steato-hepatitis

by Angulo et al.20 In addition, Marchesini et al.

have shown that the frequency of metabolic syndrome

based on ATPIII criteria is high (88%) in patients with

non-alcoholic steatohepatitis, and these patients carry a

higher risk of potentially progressive liver disease.

21 Using the same criteria, we also found a high rate of

metabolic syndrome in patients with non-alcoholic

steatohepatitis. This is an extremely important point,

as this sub-group of patients would be expected to derive

more benefit from metformin treatment.

Lactic acidosis is the most important potential risk

during metformin treatment. The US Food and Drug

Administration reported about five cases of lactic

acidosis in 100 000 metformin-treated patients in a

year.22 In a recent study, Lalau and Race searched the

medical reports for a link between lactic acidosis and

metformin treatment.

23 They found no true metformin-associated

lactic acidosis or mortality due to metformin

alone. It seems that lactic acidosis is always associated

with other contributing factors. In this study, the

appearance of autoimmune hepatitis in one patient

during the third month of therapy raised the suspicion

of an adverse event of metformin. However, we do not

have sufficient data to make a clear comment.

The small number of patients, the unblind nature of

the study and the lack of a placebo group were major

drawbacks of this investigation. In addition, post-treatment

liver biopsy could not be performed in a

certain proportion of patients. Moreover, the treatment

period was short and a longer course may be necessary

to observe more significant changes in hepatic hist-ology.

The baseline fibrosis score was low in the study

because some patients without fibrosis, but with

moderate ballooning and chronic inflammation in

biopsies, were included. This might have limited the

overall impact of the study, as the patients most likely to

benefit are those with more advanced stages of fibrosis.

In conclusion, 6 months of therapy with metformin

was well tolerated by patients with non-alcoholic

steatohepatitis and led to a greater improvement in

insulin resistance and liver enzymes than dietary

treatment alone. Although metformin seemed to

improve the severity of steatohepatitis in some patients,

no effect on fibrosis was seen. The results of this pilot

study suggest that metformin should be evaluated in

larger controlled trials with extended follow-up and liver

histology as the end-point.

Al Pater, PhD; email: old542000@...

__________________________________________________