Am J Physiol Endocrinol Metab 287: E182–E187, 2004;
10.1152/ajpendo.00189.2003.
Report
Lipid and carbohydrate metabolism in mice with a targeted
mutation in the IL-6 gene: absence of development
of age-related obesity
Gina B. Di Gregorio,1 Lori Hensley,1 Tong Lu,1 Gouri Ranganathan,1 and Philip A. Kern2
1
Division of Endocrinology, Department of Medicine, University of Arkansas for Medical Sciences;
and 2The Central Arkansas Veterans HealthCare System, Little Rock, Arkansas 72205
Submitted 25 April 2003; accepted in final form 6 February 2004
INSULIN RESISTANCE IS AN EARLY FEATURE in the development of
type 2 diabetes and is closely associated with obesity (7). Most
whole body glucose uptake occurs in skeletal muscle (8), and
obesity is associated with increased lipid accumulation in both
adipose tissue and muscle, possibly resulting in lipotoxicity
and decreased insulin action (29). The link between insulin
resistance and adipose tissue has been tightened considerably
with the description of cytokine expression by adipocytes (14).
Leptin-deficient mice become obese, insulin resistant, and
diabetic, and recent studies suggest that leptin may sensitize
skeletal muscle to catecholamine-mediated increases in lipid
oxidation (21), resulting in a decrease in muscle lipid accumulation. Another important cytokine in the obesity-insulin resistance syndrome is tumor necrosis factor-␣ (TNF-␣). Obese
rodents express higher levels of TNF-␣ in adipose tissue, and
the infusion of such animals with anti-TNF-␣-binding proteins
reverses the insulin resistance (12). Although the mechanism
of TNF-␣-mediated insulin resistance is not clear, TNF-␣ or
TNF-␣ receptor knockout (KO) mice demonstrate less insulin
resistance with high-fat feeding (30).
IL-6 is another important component of obesity-related insulin resistance. IL-6 is expressed by many cells and tissues,
including adipose tissue, and circulates at fairly high levels in
plasma. IL-6 is expressed at higher levels by omental adipose
tissue (10) and is released following a meal (24). IL-6, TNF-␣,
and perhaps other adipokines may interact with each other. In
both 3T3-L1 adipocytes and mice, TNF-␣ caused an increase
in IL-6 expression, suggesting a possible role of IL-6 in
TNF-␣-mediated alterations in carbohydrate and lipid metabolism (4, 11). Plasma IL-6 is higher in subjects with hyperinsulinemia and obesity (3) and was significantly associated with
insulin resistance independently of obesity (16). Together, the
association between IL-6 and insulin resistance in humans is
compelling and leads to the hypothesis that the suppression of
IL-6 might have favorable effects on insulin sensitivity.
IL-6 KO (IL-6⫺/⫺) mice have been available for some time.
These mice are viable, and earlier studies did not evaluate these
mice for a phenotype involving carbohydrate metabolism (2,
25, 27). In a recent study, IL-6⫺/⫺ mice developed maturity
onset obesity, with disturbed carbohydrate and lipid metabolism, and increased leptin levels (32). These results were
surprising, because high IL-6 is associated with insulin resistance in humans, and therefore one would have expected
IL-6⫺/⫺ mice to be more insulin sensitive than wild-type mice.
In this study, we examined IL-6⫺/⫺ mice, paying particular
attention to phenotypes associated with obesity and insulin
resistance. We did not observe an increase in obesity or in total
body fat with aging, even when the mice were fed a high-fat
diet. Although there were some subtle differences in carbohydrate and lipid metabolism between IL-6⫺/⫺ and IL-6⫹/⫹ mice
when they were fed a high-fat diet, there were no consistent
changes suggestive of insulin resistance. In addition, adipose
tissue secretions of leptin and TNF-␣ were similar. Therefore,
our results show that IL-6⫺/⫺ mice do not develop any obvious
phenotype related to maturity onset obesity and diabetes.
Address for reprint requests and other correspondence: P. A. Kern, Research, 151LR, Central Arkansas Healthcare System, 4300 W. 7th St., Little
Rock, AR 72205 (E-mail: [email protected]).
The costs of publication of this article were defrayed in part by the payment
of page charges. The article must therefore be hereby marked “advertisement”
in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
interleukin-6; adipose tissue
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Di Gregorio, Gina B., Lori Hensley, Tong Lu, Gouri Ranganathan, and Philip A. Kern. Lipid and carbohydrate metabolism in
mice with a targeted mutation in the IL-6 gene: absence of development of age-related obesity. Am J Physiol Endocrinol Metab 287:
E182–E187, 2004; 10.1152/ajpendo.00189.2003.—Obesity-related
insulin resistance may be caused by adipokines such as IL-6, which is
known to be elevated with the insulin resistance syndrome. A previous
study reported that IL-6 knockout mice (IL-6⫺/⫺) developed maturity
onset obesity, with disturbed carbohydrate and lipid metabolism, and
increased leptin levels. Because IL-6 is associated with insulin resistance, one might have expected IL-6⫺/⫺ mice to be more insulin
sensitive. We examined body weights of growing and older IL-6⫺/⫺
mice and found them to be similar to wild-type (IL-6⫹/⫹) mice.
Dual-energy X-ray absorptiometry analysis at 3 and 14 mo revealed
no differences in body composition. There were no differences in
fasting blood insulin and glucose or in triglycerides. To further
characterize these mice, we fed 11-mo-old IL-6⫺/⫺ and IL-6⫹/⫹ mice
a high- (HF)- or low-fat diet for 14 wk, followed by insulin (ITT) and
glucose tolerance tests (GTT). An ITT showed insulin resistance in
the HF animals but no difference due to genotype. In the GTT,
IL-6⫺/⫺ mice demonstrated elevated postinjection glucose levels by
60% compared with IL-6⫹/⫹ but only in the HF group. Although
IL-6⫺/⫺ mice gained weight and white adipose tissue (WAT) with the
HF diet, they gained less weight than the IL-6⫹/⫹ mice. Total
lipoprotein lipase activity in WAT, muscle, and postheparin plasma
was unchanged in the IL-6 ⫺/⫺ mice compared with IL-6⫹/⫹ mice.
There were no differences in plasma leptin or TNF-␣ due to genotype.
Plasma adiponectin was ⬃53% higher (71.7 ⫾ 14.1 ␮g/ml) in IL6⫺/⫺ mice than in IL-6⫹/⫹ mice but only in the HF group. Thus these
data show that IL-6⫺/⫺ mice do not demonstrate obesity, fasting
hyperglycemia, or abnormal lipid metabolism, although HF IL-6⫺/⫺
mice demonstrate elevated glucose after a GTT.
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IL-6 KNOCKOUT MICE
METHODS
⫺/⫺
tm1Kopf
Fig. 1. Effect of age and IL-6 deficiency on body weights of wild-type
(IL-6⫹/⫹) and IL-6 knockout (IL-6⫺/⫺) mice at 3 and 8 mo of age. Bar graph
shows the average weight in grams of male mice at both ages (n ⫽ 7 mice per
group). A similar trend in body weight was seen for female mice at 9 and 11
mo of age (IL-6⫹/⫹: 27.6 ⫾ 1.6 g; IL-6⫺/⫺: 24.4 ⫾ 2.0 g). Values represent
means ⫾ SE.
AJP-Endocrinol Metab • VOL
BMI, g/cm2
Age,
mo
5
14
Percent Body Fat
Diet
IL-6⫹/⫹
IL-6⫺/⫺
IL-6⫹/⫹
IL-6⫺/⫺
Low fat
High fat
Low fat
High fat
0.31⫾0.01
0.38⫾0.02
NA
NA
0.30⫾0.01
0.39⫾0.03
NA
NA
19.2⫾1.2
39.3⫾3.6*
27.9⫾6.9
NA
21.1⫾2.1
44.3⫾2.2*
29.2⫾7.2
NA
Data represent means ⫾ SE. DEXA, dual-energy X-ray absorptiometry;
BMI, body mass index; NA, not applicable. *Significant difference (P ⬍
0.001) from the low-fat diet group with similar genotype.
incubated with 500 ␮l of DMEM containing 20 mM HEPES for 4 h.
The samples were centrifuged at 1,500 g, and the medium was
collected and frozen at ⫺20°C for assay at a later date.
Hormone and metabolic assays. Blood was collected from mice at
the time of death, and the serum was assayed for leptin, adiponectin,
and TNF-␣. TNF-␣ was measured with a sensitive enzyme-linked
immunosorbent assay (R&D Systems) that has interassay and intrassay variations of 5.1 and 3.1 pg/ml, respectively. Leptin was measured
by a nonequilibrium radioimmunoassay that has a sensitivity of 0.2
ng/ml (Linco, St. Charles, MO). Adiponectin was measured by equilibrium radioimmunoassay that has a sensitivity of 1 ng/ml (Linco).
Serum triglycerides were measured using Infinity Triglycerides Reagent (Sigma-Aldrich, St. Louis, MO). All samples were assayed
together in one batch.
Measurement of lipoprotein lipase activity. Lipoprotein lipase
(LPL) catalytic activity was measured as total LPL after a detergent
extraction, and LPL mRNA was measured by Northern blotting, as
described previously (23). LPL activity was measured using a
[3H]triolein-containing substrate emulsified with lecithin, 3% bovine
serum albumin (BSA) as a stabilizer, and fetal bovine serum as a
source of apolipoprotein C-II (22). After the samples were incubated
Fig. 2. Effect of diet on body weight of IL-6⫺/⫺ mice. IL-6⫺/⫺ and IL-6⫹/⫹
mice were fed a chow diet until 11 mo of age (week 0). At that time, mice were
randomized to receive either the low-fat (LF) or high-fat (HF) diet (see
METHODS). Body weights were recorded at 0, 4, 8, and 14 wk. Data are
expressed as %baseline, and both male and female mice were included.
Baseline is the mean body weight for each group at week 0 and is designated
100%. Body weights, which were similar among the groups at week 0, were
28.5 ⫾ 2.7 g for IL-6⫹/⫹ mice and 27.9 ⫾ 4.6 g for IL-6⫺/⫺ mice. HF feeding
significantly increased the body weights of mice (P ⬍ 0.001), and mean body
weights of HF-fed IL-6⫹/⫹ (⫹/⫹/HF) mice were significantly greater than
those of HF-fed IL-6⫺/⫺ (⫺/⫺/HF) mice (P ⬍ 0.001). Values represent
means ⫾ SE.
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Animals. IL-6
mice breeder pairs (C57BL/6J-Il6
) were
purchased from Jackson Labs (Bar Harbor, ME), and a colony of
IL-6⫺/⫺ mice was established. This is the same strain of IL-6⫺/⫺ mice
reported previously (32). A breeder pair of normal C57BL/6 mice was
also purchased from Jackson Labs, and their littermate mice were used
as wild-type controls for the experiment. All experiments were conducted from this colony, and no backcrosses to other strains were
made. The breeding strategy used by Jackson Labs to maintain the
IL-6 KO mouse strain was similar to that used by others (32). Animals
were maintained on mouse chow and water ad libitum according to
guidelines provided by the Animal Care and Use Committee, in
accordance with accepted standards of humane animal care as outlined in The Endocrine Society’s Ethical Guidelines for Research. For
the high-fat feeding experiment, 3-mo and 11-mo-old mice were
randomly assigned to receive either mouse chow (6% fat; Harlan
Teklad, Madison, WI, www.harlan.com) or a high-fat diet (59% fat;
BioServ, Frenchtown, NJ, www.bio-serv.com). The precise composition of these diets can be found on the company websites. In brief, the
low-fat diet contained 20% protein, 6% fat, and 4% fiber, and the
high-fat diet contained 16% protein, 59% fat, and 0.1% fiber.
Verification of IL-6⫺/⫺ mice. Polymerase chain reaction was used
to identify mice containing an insertion of the neor cassette in exon 2
of the IL-6 gene. Primers were designed to lie within exon 2 on either
side of the neor cassette such that a 174-bp product was generated for
wild-type mice and a 1314-bp product was generated for KO mice.
The primer sequence for the forward primer was 5⬘-TTCCATCCAGTTGCCTTCTTGG-3⬘, and the sequence for the reverse primer
was 5⬘-TTCTCA-TTTCCACGATTTCCCAG-3⬘. The presence of the
neor cassette was confirmed in IL-6⫺/⫺ mice.
Measurement of serum IL-6 was used in combination with genotyping to distinguish IL-6⫺/⫺ mice from IL-6⫹/⫹ mice. IL-6 was
measured with a sensitive enzyme-linked immunosorbent assay (R&D
Systems, Minneapolis, MN) that has a sensitivity of 3.1 pg/ml. Mean
(⫾SE) serum levels of IL-6 were 23.7 ⫾ 7.6 pg/ml in IL-6⫹/⫹ mice
and were undetectable in IL-6⫺/⫺ mice. All samples were measured in
duplicate in one assay. The blood levels of IL-6 confirmed the
genotyping results and verified that the mice did not express IL-6.
Medium secretion. White adipose tissue (WAT) was collected from
mice at the time of death, and 100 mg of WAT from each animal were
Table 1. Percent body fat in young (5-mo-old) and old (14mo-old) mice as determined by DEXA and BMI
Report
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IL-6 KNOCKOUT MICE
Table 2. Effects of age and genotype on blood glucose levels
Fasting Glucose
Nadir Glucose During ITT
Peak Glucose During GTT
Age,
mo
IL-6⫹/⫹
IL-6⫺/⫺
IL-6⫹/⫹
IL-6⫺/⫺
IL-6⫹/⫹
IL-6⫺/⫺
3
8
208.3⫾13.4
192.9⫾12.5
218.6⫾10.7
193.3⫾8.5
68⫾5.4
72.0⫾5.9
64.8⫾5.3
86.9⫾11.9
554.6⫾20.6
395.0⫾30.9
564.9⫾18.1
288.3⫾36.9
Data represent means ⫾ SE in mg/dl. ITT and GTT, insulin and glucose tolerance tests.
RESULTS
Effects of genotype and diet on body weight and composition. IL-6⫺/⫺ and wild-type mice grew and developed normally, and no evidence of obesity was observed. Figure 1
shows body weight at 3 and 8 mo of age. IL-6⫺/⫺ mice
weighed 10% less than IL-6⫹/⫹ males at 3 mo of age (P ⫽
0.002), and body weights of the two groups were similar at 8
mo (Fig. 1). To examine both weight and body composition,
the body mass index (BMI) was calculated as weight (g)/length
(cm)2, as described previously (20), and percent body fat was
assessed using dual-energy X-ray absorptiometry. As shown in
Table 1, a high-fat diet resulted in a predictable increase in
BMI and percent body fat; however, there were no differences
between IL-6⫺/⫺ and IL-6⫹/⫹ mice. Older mice (14 mo) had a
higher body fat than the younger mice (5 mo), but there was no
difference due to genotype. In a previous study, the obesity/
diabetes phenotype of IL-6⫺/⫺ mice was more pronounced
with aging (32). To further characterize body weight with
aging and diet in these mice, mice were fed a chow diet until
11 mo of age and were then randomized to either a low-fat or
high-fat diet for 14 wk. Both male and female IL-6⫺/⫺ and
IL-6⫹/⫹ mice gained weight on the high-fat diet, but IL-6⫺/⫺
mice gained slightly less weight compared with IL-6⫹/⫹ mice
(P ⬍ 0.001; Fig. 2).
Effects of genotype and diet on carbohydrate and lipid
metabolism. IL-6⫹/⫹ and IL-6⫺/⫺ mice had similar fasting
glucose levels at 3 and 8 mo of age (Table 2). At 8 mo of age,
after ad libidum consumption of normal chow (low-fat diet),
carbohydrate metabolism was assessed with a GTT and ITT, as
described in METHODS. When the IL-6⫺/⫺ mice were compared
with wild-type mice, peak glucose levels during a GTT at 8 mo
of age were not significantly different, nor were nadir glucose
levels during an ITT (Table 2). Hence, these studies provided
no evidence for insulin resistance at this age. Additional
studies were performed in older mice after high-fat feeding. In
response to high-fat feeding, both IL-6⫺/⫺ and IL-6⫹/⫹ mice
demonstrated evidence of insulin resistance, with higher blood
glucose values in response to a GTT, and elevated fasting
serum insulin and a higher value for homeostasis model assessment of insulin resistance (HOMA-IR; Table 3). During
the GTT, IL-6⫺/⫺ mice fed a high fat diet had 1.6-fold higher
glucose levels compared with IL-6⫹/⫹ mice (Table 3). Such an
effect would imply that the IL-6⫺/⫺ mice were more insulin
resistant than the IL-6⫹/⫹ mice. However, these findings were
not confirmed by the ITT. During the ITT, both genotypes of
mice demonstrated less insulin-mediated glucose suppression
in response to high-fat feeding (P ⬍ 0.001; Fig. 3), and overall
glucose levels were twofold higher compared with low-fat-fed
mice. However, there were no significant differences between
the IL-6⫺/⫺ and IL-6⫹/⫹ mice in the ITT. Therefore, there
were no consistent changes in the IL-6⫺/⫺ mice to suggest
genotype-mediated insulin resistance in these mice.
Effects of genotype and diet on LPL activity. We measured
total LPL activity in WAT, muscle, and postheparin plasma
(PHP) of male and female mice and found that LPL activity
was similar in IL-6⫺/⫺ mice and IL-6⫹/⫹ mice (Table 4). This
absence of effect of genotype was noted in both young (5 mo)
and older (14 mo) mice. In addition, no effect of gender was
observed. After young mice consumed a high-fat diet, there
was an approximately twofold increase in PHP LPL activity.
Although the increase in PHP activity was slightly greater in
the IL-6⫹/⫹ mice, the difference between the IL-6⫹/⫹ and
Table 3. Effects of genotype and high-fat diet on fasting blood glucose and insulin levels in 14-mo-old mice
Fasting Glucose, mg/dl
Genotype
⫹/⫹
IL-6
IL-6⫺/⫺
Peak Glucose During GTT, mg/dl
Fasting Serum Insulin, ng/ml
HOMA-IR
Low fat
High fat
Low fat
High fat
Low fat
High fat
Low fat
High fat
195.0⫾11.7
209.7⫾8.4
213.0⫾20.6
168.7⫾25.3
611.7⫾51.7
471.7⫾40.4
678.3⫾20.9
1090⫾158.3†
1.7⫾0.4
1.1⫾0.2
7.6⫾1.9*
8.0⫾1.5*
0.8⫾0.3
0.58⫾0.06
3.8⫾0.7*
3.4⫾1.1*
Data represent means ⫾ SE. HOMA-IR, homeostasis model assessment of insulin resistance. *P ⬍ 0.05, significant difference from low-fat-fed mice; †P ⬍
0.05 from high-fat-fed IL-6⫹/⫹ mice.
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with substrate for 1 h at 37°C, liberated 3H-labeled free fatty acids
(3H-FFA) were separated and quantitated by liquid scintillation.
Adipose LPL activity was expressed as nanomoles of FFA released
per milligram of protein per hour, and postheparin plasma (PHP) LPL
activity was expressed as nanomoles of FFA released per milliliter per
hour.
Glucose and insulin tolerance tests. Mice were fasted for 4 h before
the glucose tolerance test (GTT). GTT and insulin tolerance tests
(ITT) were performed ⬃1 wk apart. Animals were given 2 g of 50%
D-glucose/kg body wt intraperitoneally for GTT and human regular
insulin (0.75 U/kg; Eli Lilly, Indianapolis, IN) intraperitoneally for
ITT, as described previously (1). Mice were not anesthetized during
this procedure, and a 75-␮l aliquot of blood was taken retroorbitally
from mice after they received an eye drop of proparacaine (Bausch
and Lomb Pharmaceuticals, Tampa, FL) at 0, 15, 30, 60, and 120 min.
Blood samples were assayed for glucose.
Statistics. Data were tested for normality and homogeneity of
variance. Differences among treatment groups for GTT and ITT tests
were determined using one-way analysis of variance, and comparisons
were made using Tukey’s test. A two-tailed t-test was used to identify
differences among groups for body weights and WAT weights, total
LPL activity, and quantities of cytokines in the blood and secreted by
adipose tissue.
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IL-6 KNOCKOUT MICE
Table 5. Effects of genotype and high-fat diet in 14-mo-old
mice on blood cytokines and their secretion by WAT
Adiponectin, ␮g/ml
Leptin, ng/ml
Genotype
⫹/⫹
IL-6
IL-6⫺/⫺
Diet
Serum
Adipose
Serum
Adipose
Low fat
High fat
Low fat
High fat
12.3⫾2.6
52.6⫾6.2†
9.1⫾1.1
52.9⫾14.3†
9.9⫾1.7
25.9⫾1.6†
7.2⫾0.7
17.3⫾3.6†
31.7⫾5.7
33.0⫾5.9
37.2⫾5.2
71.7⫾14.1‡
0.5⫾0.05
0.4⫾0.1
0.9⫾0.1*
0.6⫾0.1
Data represent means ⫾ SE. White adipose tissue (WAT, 100 mg) was
incubated for 4 h in 500 ␮l of serum-free medium. Serum and medium leptin
and adiponectin were measured as described in METHODS. †P ⬍ 0.05,
significant difference from low-fat-fed mice; ‡P ⬍ 0.001 and *P ⫽ 0.012,
significant difference from all other treatment group means.
IL-6⫺/⫺ mice was not statistically significant (Table 4). Northern blots were performed on adipose tissue, and no differences
in mRNA levels were observed between IL-6⫹/⫹ and IL-6⫺/⫺
mice (data not shown). We also examined the effects of diet
and genotype on serum triglyceride levels. There were no
significant differences in triglyceride levels between IL-6⫹/⫹
and IL-6⫺/⫺ mice. In young mice, fasting triglyceride levels
were 93 ⫾ 18.9 and 78.6 ⫾ 4.8 mg/dl, respectively, in IL-6⫹/⫹
and IL-6⫺/⫺ mice [P ⫽ not significant (NS)]. In a similar
fashion, there were no significant gender-related differences in
triglyceride in older mice or in high-fat-fed mice.
Effects of genotype and diet on adipokines. Diet and genotype did not affect serum TNF-␣ levels or secretion of TNF-␣
by adipose tissue in any of the treatment groups (data not
shown). In contrast, serum adiponectin levels were increased
nearly twofold in high-fat-fed IL-6⫺/⫺ mice compared with
other treatment groups (P ⫽ 0.006; Table 5). Secretion of
adiponectin by adipose tissue was also higher in IL-6⫺/⫺ mice,
but only in the low-fat-fed group. As expected, serum leptin
Table 4. Effects of genotype and high-fat diet on total LPL
activity and serum triglycerides in young and old mice
Young Mice
DISCUSSION
IL-6 is expressed by many tissues and has been extensively
studied with regard to numerous physiological processes and
disease states. IL-6 is expressed and secreted by adipose tissue
and therefore may represent one of a number of obesity-related
inflammatory cytokines that cause insulin resistance. In a
number of human studies, there was a significant correlation
between plasma IL-6 and insulin sensitivity. IL-6 was associated with insulin resistance in obese and lean humans (3, 16),
and this relationship between plasma IL-6 and insulin sensitivity was independent of obesity. The injection of IL-6 into
healthy human volunteers increased blood glucagon and increased fasting glucose (28). Finally, several studies in humans
have suggested that common polymorphisms in the IL-6 gene,
which may alter expression, result in altered insulin sensitivity
(9, 18).
As with any knockout mouse, one would not necessarily
expect a phenotype that mimics a human disease entity. In
addition to the species difference, the IL-6⫺/⫺ mouse lacked
IL-6 during embryogenesis and development, and other systems likely substituted for the functions of this rather ubiquitous cytokine. Nonetheless, we hypothesized that the absence
of IL-6 would result in a more insulin-sensitive animal. To test
our hypothesis, we studied IL-6⫺/⫺ mice and have summarized
our findings on the growth and metabolic characteristics for
IL-6⫺/⫺ mice from these studies in Table 6. IL-6⫺/⫺ mice had
Table 6. Phenotype of IL-6⫺/⫺ mice
Old Mice
Parameter
Genotype
Diet
Adipose
Muscle
PHP
Adipose
IL-6⫹/⫹
Low fat
High fat
Low fat
High fat
326⫾38
437⫾83
282⫾49
287⫾47
3.9⫾0.4
3.4⫾0.2
4.9⫾0.9
4.7⫾0.5
4,651⫾386
10,476⫾846†
4,688⫾164
7,459⫾1240
302⫾76
350⫾83
393⫾69
313⫾102
IL-6⫺/⫺
Body weight
Data represent means ⫾ SE. Tissue lipoprotein lipase (LPL) activity is expressed as nmol䡠h⫺1䡠mg protein⫺1. Postheparin plasma (PHP) LPL activity is
expressed as nmol䡠h⫺1䡠ml⫺1. Old mice were 14 mo old and had been on a high-fat
diet for 12 wk. Young mice were 5 mo old and had been fed a high fat diet for 8
wk. †P ⬍ 0.05, significant difference from low-fat-fed IL-6⫹/⫹ mice.
AJP-Endocrinol Metab • VOL
Body composition
LPL activity
Adipose TNF
Adipose leptin
Adipose adiponectin
Serum triglycerides
Insulin sensitivity
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IL-6⫺/⫺ vs. IL-6⫹/⫹
Low-fat diet: less at 3 mo
High-fat diet: less weight gain
No difference
No difference
No difference
No difference
High-fat diet: increased
No difference
GTT: elevated glucose, high-fat diet only
ITT: no difference
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Fig. 3. Effect of diet on insulin tolerance in IL-6⫺/⫺ mice. Blood glucose
levels during an insulin tolerance test (ITT) in 14-mo-old male and female
mice fed either an LF- or HF diet. Data are expressed as %control. Control is
the mean glucose values for each treatment group at time 0 of the ITT and is
designated 100%. Glucose levels, which were similar for each treatment group
at time 0, were ⫹/⫹/LF: 194.1 ⫾ 10.7 mg/dl; ⫹/⫹/HF: 217.4 ⫾ 10 mg/dl;
⫺/⫺/LF: 181.7 ⫾ 10.1 mg/dl; and ⫺/⫺/HF: 223 ⫾ 13 mg/dl. *Significant
difference from contemporary genotype-matched LF-fed mice. Values represent means ⫾ SE.
levels were markedly higher in high-fat-fed mice (P ⫽ 0.002),
and there were no differences due to genotype (Table 5). A
similar trend was observed for secretion of leptin by adipose
tissue, which was markedly elevated in response to high-fat
feeding and was not influenced by genotype (P ⬍ 0.001).
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pancreatic insulin-producing cells, with the resultant phenotype of near-euglycemia due to hyperinsulinemia (19). It is
possible that ␤-cell hyperplasia and the resultant hyperinsulinemia are dependent on IL-6, since our IL-6⫺/⫺ mice became
hyperglycemic on a high-fat diet. As with this study, Wallenius
et. al. (32) studied IL-6⫺/⫺ mice on a C57BL/6 background.
Therefore, it is unclear why we obtained different results,
unless there are subtle differences in strains related to either
breeding strategy, or important differences in environment or
diet.
Removal of endogenous IL-6 in mice did not influence
adipose tissue secretion of TNF-␣, adiponectin, or leptin.
High-fat feeding resulted in an increase in adiponectin only in
IL-6⫺/⫺ mice and not in IL-6⫹/⫹ mice. Circulating levels of
adiponectin are reduced in several mouse models of insulin
resistance (26). Others found decreased levels of adiponectin in
obese and diabetic subjects (13), and there is a strong positive
association between plasma adiponectin and insulin sensitivity
(15). These data suggest that the increase in adiponectin
secretion in high-fat-fed IL-6⫺/⫺ mice may be uniquely associated with the IL-6⫺/⫺ genotype and perhaps is due to a
regulatory interaction between IL-6 and adiponectin.
In summary, IL-6 ⫺/⫺ mice did not manifest maturity onset
obesity. Although several subtle phenotypes were observed,
such as a higher glucose following a GTT in fat-fed mice, and
some changes in adiponectin, these mice were generally characterized by a lack of overt diabetes and obesity compared with
wild-type mice.
ACKNOWLEDGMENTS
We thank Dr. Charles O’Brien for providing the primer sequences for the
PCR to genotype the IL-6⫺/⫺ mice. We also thank Brian Wells, Dale Paulson,
Ginger Brown, and Margaret McIntire for technical assistance.
GRANTS
This study was funded in part by a Career Development Award from the
American Diabetes Association, Grant DK-39176 from the National Institute
of Diabetes and Digestive and Kidney Diseases, and a Merit Review Grant
from the Veterans Administration.
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