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CELLULAR AND MOLECULAR
Third Department of Internal Medicine, Miyazaki Medical College, Miyazaki University, Kiyotake, Miyazaki, Japan
Received December 29, 2003; accepted March 1, 2004.
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Abstract |
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1–7, 9, 10, 
1–4, 
, and 
subunit mRNAs are expressed in adipocytes. Immunocytochemical experiments also suggested the presence of 7 and 2 subunits. The receptor binding assay revealed a binding site for nicotine (Kd = 39.2 x 10-9 M) on adipocytes. Adipocytes incubated with nicotine for 12 and 36 h released tumor necrosis factor- (TNF-), adiponectin, and free fatty acid (FFA) into the medium in a dose-dependent manner with increasing nicotine concentration from 6 x 10-8 to 6 x 10-4 M. However, TNF- protein levels in adipocytes incubated for 12 and 36 h decreased in a dose-dependent manner with increasing nicotine concentration from 6 x 10-8 to 6 x 10-4 M. These results show that adipocytes have functional nAChRs and suggest that nicotine reduces TNF- protein production in adipocytes through the activation of nAChRs. Nicotine may temporarily lower insulin sensitivity by stimulating the secretion of TNF- and FFA, whereas long-term direct stimulation of nAChRs by nicotine in addition to autonomic nervous system stimulation may contribute to better insulin sensitivity in vivo through a modulated secretion of adipocytokines.
). nAChRs are divided into muscle (1, 1, 
/, and ) and neuronal AChRs (2–10 and 2–4), and neuronal AChRs are further subdivided into those that form homomeric receptors when expressed in heterologous systems (7–10) and those that form heteromeric receptors (2–6 and 2–4 in different combinations) (Gotti et al., 2000). The 8 nAChR subunit has not been studied since it is only expressed in the chicken (Rubboli et al., 1994). The muscle receptor subtypes are comprised of 1, 1, , and (embryonal) or 1, 1, , and (adult) subunits (Tassonyi et al., 2002). ACh and its receptors are among the best characterized neurotransmitter/receptor systems (Albuquerque et al., 1997). Cholinergic neurotransmission is used in a variety of neuronal systems and in a broad range of animals ranging from invertebrates to mammals (Lindstrom et al., 1995). The main function of this receptor family is to transmit signals through the neurotransmitter acetylcholine at neuromuscular junctions and in the central and peripheral nervous systems (Marubio and Changeux, 2000). In addition, non-neuronal cells may express a functional nAChR. Human bronchial epithelial cells and aortic endothelial cells express a functional 3 subtype nAChR, which modulates cell shape and affects cell-to-cell contact (Maus et al., 1998). Human skin keratinocytes express functional nAChRs of different subtypes, including 3, 9, and possibly 7 subunits (Nguyen et al., 2000). Specific receptors for nicotine are present on lymphocytes in humans (Hoss et al., 1986) and rats (Maslinski et al., 1992); however, whether functional nAChRs exist in adipocytes is unclear.
Excess adipose tissue leads to insulin resistance, thereby increasing the risk of type 2 diabetes mellitus and cardiovascular disease (Saltiel and Kahn, 2001). Adipocytes release cytokines that influence energy expenditure, insulin sensitivity, vasomotor tone, and fibrinolysis, and obesity perturbs the regulation of these cytokines. Physiologically active substances produced in adipose tissues, called adipocytokines and free fatty acid (FFA), play a role in the progression of insulin resistance in obesity (Matsuzawa et al., 1999). Adipocytes play a role in systemic energy homeostasis by producing molecules such as leptin, plasminogen activator inhibitor-1, and several cytokines including tumor necrosis factor- (TNF-) and interleukin-6 that influence key metabolic pathways (Mohamed-Ali et al., 1998). TNF- is a mediator of lipid metabolism, adipocyte differentiation, and in vivo insulin sensitivity (Hotamisligil et al., 1993). TNF- is expressed in macrophages and adipocytes and is substantially elevated in obesity in rodents (Hotamisligil et al., 1993) and humans (Hofmann et al., 1994).
In contrast to other adipocytokines, adiponectin is proposed to play a role in the regulation of energy homeostasis and insulin sensitivity (Hu et al., 1996). Adiponectin levels are depressed in obesity and associated comorbidities such as type 2 diabetes. Decreased expression of adiponectin correlates with insulin resistance. Adiponectin is a hormone secreted from adipocytes and has antidiabetic and antiatherogenic effects (Yamauchi et al., 2003). The mechanism of regulation of adiponectin secretion remains to be clarified.
In a previous study, we reported that oral nicotine administration reduces insulin resistance in obese diabetic rats possibly through decreased expression of TNF- in visceral fat tissues and reduced hepatic glucose release (Liu et al., 2001, 2003). To clarify whether nicotine has a direct effect on the function of adipocytes, we evaluated nAChR expression in adipocytes and the direct effects of nicotine on the production of adipocytokines (TNF- and adiponectin).
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Materials and Methods |
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Conditioned Media. Preadipocytes seeded at 2 x 104 cells/cm2 into 24-well plates (Falcon; BD Biosciences) were cultured to confluence. Under the culture conditions, preadipocytes differentiated into matured adipocytes. Adipocytes were incubated in medium for 12 and 36 h with nicotine (nicotine tartrate dihydrate dissolved in Dulbecco's modified Eagle's medium) with a concentration ranging between 6 x 10-8 and 6 x 10-4 M. The conditioned medium was then removed, centrifuged for 5 min at 4°C at 1000 rpm, and the medium and cells were separately stored at -80°C.
Assays for TNF-, Adiponectin, and FFA. Using a specific antibody, medium TNF- and adiponectin were measured by enzyme-linked immunosorbent assay using kits obtained from Bio-Source International, Inc. (Camarillo, CA) and Otsuka Pharmaceutical Co., Ltd (Tokyo, Japan), respectively. The medium FFA was measured by spectrophotometric assays using a commercially available kit (Wako Bioproducts, Richmond, VA).
Adipocyte Number and Form. Adipocytes were counted by a hemocytometer, and viable and dead cells were counted by 0.4% trypan blue staining. Micrographs were taken at 40 to 200x magnification.
RNA Extraction and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR). Total RNA was isolated from the adipocytes using a QIAamp RNA mini kit (QIAGEN, Tokyo, Japan). Before RT-PCR experiments, total RNA was quantified by measuring the absorbance at 260 nm (U-1100 spectrophotometer; Hitachi Software Engineering, Yokohama, Japan). The RNA samples had A260/A280 ratios ranging from 1.8 to 2.0. All RNA samples were treated with amplification grade DNase I according to the manufacturer's instructions (Invitrogen, Carlsbad, CA) to eliminate residual DNA. cDNA synthesis and PCR were performed in a single tube using gene-specific primers and total RNA by SuperScript One-Step RT-PCR with a Platinum Taq kit (Invitrogen). For the conversion of total RNA (0.5 µg) to cDNA, a 50-µl single-tube reaction mixture was prepared from a master mix containing 25 µl of 2x reaction mix, 0.5 µg of template RNA, and 1 µl of RT/Platinum Taq mix. Then 10 µM of each gene-specific primer pair was added to the tubes. Primer sequences were selected from the unique cytoplasmic domain region of each nAChR subunit (Table 1) (LaPolla et al., 1984; Buonanno et al., 1989; Witzemann et al., 1990; Rohwedel et al., 1995; Liu et al., 1998; Tseng et al., 2001). Glyceraldehyde-3-phosphate dehydrogenase (G3PDH) (Toyobo Engineering) was used as an internal control to verify the quality of each RNA sample and its subsequent RT-PCR analysis. The RT-PCR cycling profiles using a Thermal Cycler (GeneAmp PCR System 9600; PerkinElmer Life and Analytical Sciences, Boston, MA) were as follows: 1 cycle at 50°C for 30 min, 1 cycle at 94°C for 2 min, 35 cycles at 94°C for 1 min (46–57°C, respectively, in Table 1), 72°C for 1 min, and a final cycle at 72°C for 7 min. A 12-µl aliquot of each sample was electrophoresed on a 2.4% agarose gel containing 0.6 mg/ml ethidium bromide.
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Immunofluorescence Microscopy of 7 and 2 Subunits in Cultured Adipocytes. Adipocytes grown on Labtek 4-well slides were cooled on ice and washed in ice-cold PBS. The cells were fixed with 4% paraformaldehyde in PBS for 20 min and permeabilized with 0.1% Triton X-100 in PBS for 20 min at room temperature. The slides were then blocked for 20 min with 5% normal goat serum in PBS, after which they were incubated for 1 h in mouse anti-7 antibody (1:200 dilution in PBS, mAb 306; Sigma-Aldrich, St. Louis, MO) or rabbit anti-2 antibody (1:200 dilution in PBS; Santa Cruz Biotechnology Inc., Santa Cruz, CA) at room temperature, respectively. The cells were then washed three times for 5 min with ice-cold PBS and incubated for 1 h at room temperature with fluorescein isothiocyanate-conjugated goat anti-mouse IgG (1:200 dilution in PBS; Jackson ImmunoResearch Laboratories Inc., West Grove, PA) or fluorescein isothiocyanate-conjugated goat anti-rabbit IgG (1:200 dilution in PBS, Santa Cruz Biotechnology Inc.), respectively. They were then washed three times with PBS for 5 min each, mounted in PermaFluor mounting medium (Thermo Shandon, Pittsburgh, PA), and observed with a FluoView FV500 confocal laser scanning microscope (Olympus Optical Co., Ltd., Tokyo, Japan).
[3H](-)-Nicotine Binding Assays. The analysis of binding data were performed as previously described (Didier et al., 1995). Briefly, a 0.8-ml cell suspension (50 mM NaCl, 5 mM KCl, 10 mM CaCl2 · 2H2O, 2 mM EDTA, 50 mM Tris base, 50 mM HEPES, 10 mM D-glucose, 1% bovine serum albumin, pH 8.0) (cell count was 4 x 107 cells/ml) was added to tubes containing unlabeled (-)-nicotine (0, 1, 10, 100, and 1000 x 10-6 M) and 5 x 10-9 M[3H](-)-nicotine [(-)-N-methyl-[3H]nicotine (81 Ci/mmol), PerkinElmer Life and Analytical Sciences, Inc.]. A final cell suspension of 1 ml was incubated at 37°C for 30 min. The reaction was terminated by adding 2 ml of cold buffer, and the samples were then filtered through Whatman GF/C glass filters (Whatman International Ltd., Kent, England) presoaked with 0.3% polyethylenimine solution for 5 h. The filters were washed three times with the same ice-cold buffer, and the radioactivity retained on the filter was measured by liquid scintillation spectrometry. For Scatchard analysis, the particulate fractions were incubated with various concentrations of [3H](-)-nicotine (0 to 100 x 10-9 M) under the same conditions as described above. The dissociation constant (Kd) and maximal binding sites (Bmax) for adipocyte [3H]nicotine binding were estimated by Scatchard analysis. The GraphPad Prism program was used for data manipulations, and all values including Bmax and Kd values were determined using this program.
Western Blot Analysis. Harvested adipocytes were homogenized in ice-cold buffer (50 mM HEPES, 150 mM sodium chloride, 1% Triton X-100, pH 7.8, 0.2 mM phenylmethylsulfonyl fluoride, 0.5 mg/ml pepstatin, 1 mg/ml leupeptin, and 2 mg/ml aprotinin), and the resulting homogenate was centrifuged for 10 min at 4°C at 12,000 rpm. Samples containing 50 µg of protein were resolved by electrophoresis in a 12% SDS-polyacrylamide gel. Rat TNF- (PeproTech, Inc., Rocky Hill, NJ) and Rainbow markers (Amersham Biosciences Inc., Piscataway, NJ) were used as molecular markers. Proteins were transferred to polyvinylidene fluoride membranes (Millipore Corporation, Bedford, MA). Blots were incubated with anti-mouse TNF- polyclonal antibody (Pierce Endogen, Rockford, IL) at 0.65 µg/ml in a low-fat milk solution overnight at 4°C. After the membranes were incubated for 1 h at room temperature with horseradish peroxidase-conjugated donkey anti-rabbit IgG [0.2 mg/ml (1:5000), Amersham Biosciences Inc.)] in Tris-buffered saline/Tween 20-bovine serum albumin, they were incubated with enhanced chemiluminescence (ECL) reagents (Amersham Biosciences Inc.) and exposed to X-ray film for 3 min. Densitometric analysis of immunoblots was performed using Adobe PhotoShop software (Adobe Systems, Inc., Mountain, CA) for Apple Macintosh computers (Apple Computer, Inc., Cupertino, CA).
Statistical Analysis. The unpaired Student's t test was used to determine significance, and values were represented as the means ± S.E.M. of the number of experiments stated. StatView 5.0 software was used for all statistical calculations. A p value of less than 0.05 was accepted as being statistically significant.
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Results |
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, , , , and nAChR subunits in cultured adipocytes by RT-PCR using specific primers for the (1–7, 9, and 10), (1–4), , , and nAChR subunits (Table 1). Ethidium bromide staining of the gel shows the presence of nAChRs subunits (1–7, 9, 10, 1–4, , and ), G3PDH, and molecular weight markers (M) obtained by RT-PCR as seen in Fig. 1. The data show that rat adipocytes express (1–7, 9, and 10), (1–4), , and subunits. Amplification yielded PCR products of expected sizes: 288 bp for 1, 300 bp for 2–7 and 9, 209 bp for 10, 355 bp for 1, 507 bp for 2, and 300 bp for 3 and 4. Amplification of the G3PDH gene product (452 bp) was used as an internal control to verify the quality of each RNA sample and its subsequent RT-PCR. However, in the present study gene-specific primers for rat and subunits (Table 1) amplified larger products in size (, 395 bp; , 402 bp) than expected from previous findings in skeletal muscle (, 235/291 bp; , 222/340 bp) (Fig. 1). The nAChR subunit was not expressed (Fig. 1).
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Immunocytochemical Studies on Adipocytes Using Specific Antibodies Against 7 and 2 Subunits. To further ensure the expression of nAChR subunits in adipocytes, we performed an immunocytochemical analysis using specific antibody against 7 or 2 subunits. The presence of antibodies bound to the cells was revealed by the binding of fluorescein isothiocyanate-labeled goat anti-rabbit IgG antibody. A representative field is shown in Fig. 2, A and B. 7 and 2 subunit immunoreactivity was observed on the membrane and in the cytoplasm of the adipocytes, whereas no immunoreactivity was detected in the nuclei.
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[3H](-)-Nicotine Binding to Adipocytes. Specific binding of [3H](-)-nicotine (5 x 10-9 M) to adipocytes decreased in a dose-dependent manner with increasing (-)-nicotine concentration from 0 to 1 x 10-3 M and was significant for unlabeled (-)-nicotine concentrations from 1 x 10-4 to 1 x 10-3 M (Fig. 3).
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Saturation Analysis of [3H](-)-Nicotine. Saturation studies on adipocytes using concentrations of [3H](-)-nicotine from 1 x 10-10 M to 1 x 10-7 M revealed the presence of saturable binding sites. Nonlinear regression analysis of nicotine binding yielded a Kd value of 39.15 ± 2.67 x 10-9 M and a Bmax of 43,236 ± 1152 sites/cell for the affinity site by Scatchard analysis as seen by the curved appearance of the graph (Fig. 4).
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Adipocyte Number and Histological Examination of Culture Adipocytes. There was no significant difference in the counts of viable cells between the nicotine and control groups from 6 x 10-8 to 6 x 10-4 M for 12 and 36 h (Table 2). The histological forms and size of adipocytes were not significantly different in both groups from 6 x 10-8 to 6 x 10-4 M for 12 and 36 h (not shown).
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Adiponectin Concentration in the Medium. Adipocytes incubated with (-)-nicotine for 12 and 36 h released more adiponectin than the control into the culture medium. The release of adiponectin was augmented in a dose-dependent manner with increasing (-)-nicotine concentration from 6 x 10-8 to 6 x 10-4 M. Moreover, nicotine-stimulated adiponectin secretion for 36 h was significantly higher than that for 12 h at the same nicotine-stimulated concentration (Fig. 5).
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TNF- and FFA Concentration in the Medium. Adipocytes stimulated with (-)-nicotine released more TNF- and FFA than the control for 12 and 36 h into the culture medium. The release of TNF- and FFA was augmented in a dose-dependent manner with increasing (-)-nicotine concentration from 6 x 10-8 to 6 x 10-4 M. Nicotine-stimulated TNF- release for 36 h was significantly lower than that for 12 h at the same nicotine concentration (Fig. 6A); however, there was no significant difference in FFA release between 12 and 36 h at the same nicotine concentration (Fig. 6B).
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Expression of TNF- Protein. The results of Western blot studies performed using anti-mouse TNF- polyclonal antibody are shown in Fig. 7A. The percentage ratio to the corresponding standard TNF was calculated as a TNF- protein relative intensity and is shown in Fig. 7B. Western blot analysis revealed that TNF- protein in adipocytes in the nicotine group was significantly lower than in the control group for 12 and 36 h (p < 0.001). Moreover, the TNF- protein levels decreased in a dose-dependent manner with increasing (-)-nicotine concentration from 6 x 10-8 to 6 x 10-4 M in adipocytes (Fig. 7, A and B).
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Discussion |
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TopAbstractMaterials and MethodsResultsDiscussionReferences |
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1–7, 9, 10, 1–4, , and subunit mRNAs are expressed in adipocytes. Immunocytochemical experiments also suggested the presence of 7 and 2 subunits. Therefore, the subunits that characterize neuronal and muscle (1 and 1) nicotinic receptors (Witzemann et al., 1990; Liu et al., 1998; Tseng et al., 2001) are present in rat adipocytes. PCR could not amplify the subunit transcript because the subunit was replaced by an subunit to become the adult-type receptor (Saito et al., 2002). However, the present findings of the size of and mRNA expression in the adipocytes differ from that expressed in muscle (LaPolla et al., 1984; Buonanno et al., 1989). The differences in the size of the and subunit mRNAs between muscle and fat tissues can be explained by tissue-specific splicing. Further studies are needed to clarify this.
The saturation curve was analyzed by a nonlinear regression model, and the Kd and Bmax values were subsequently determined. The curved appearance of the Scatchard plot supports this interpretation. The Kd values for (-)-nicotine binding to the high-affinity sites is 39.15 ± 2.67 x 10-9 M. This is similar to that for nicotine binding sites found in brain and peripheral blood cells, which have Kd for nicotine between 2 and 43 x 10-9 M (Wonnacott, 1987; Lebargy et al., 1996).
[3H](-)-nicotine binding to adipocytes decreased in a dose-dependent manner with increasing (-)-nicotine concentration from 0 to 1 x 10-3 M and was significant for unlabeled (-)-nicotine concentrations from 1 x 10-4 to 1 x 10-3 M. Chronic administration of nicotine to animals up-regulates nAChR in the central nervous system when examined by [3H]nicotine radiolabeled ligand for nAChRs (Ke et al., 1998). In addition, several investigations reported that the up-regulation of the receptors is due to increased numbers of 4, 2, and 7 nAChRs subtypes in neurons and non-neuronal cells (Bencherif et al., 1995). Although the nAChRs subtypes up-regulated in rat adipocytes used in this study remain to be clarified, the present findings suggest that rat adipocytes express functional nAChRs.
To investigate the direct effect of nicotine exposure on cytokine secretion from adipocytes, we examined TNF-, adiponectin, and FFA levels in the culture cell medium with nicotine stimulation for 12 and 36 h. These results showed that TNF-, adiponectin, and FFA was released into the medium in a dose-dependent manner with increasing nicotine concentration (Figs. 5 and 6, A and B). However, after stimulation with nicotine at the same concentration for 36 h, adiponectin release was significantly higher, TNF- release was significantly lower, and FFA release did not change compared with the results for 12 h. On the other hand, TNF- protein levels in adipocytes incubated for 12 and 36 h decreased in a dose-dependent manner with increasing nicotine concentration (Fig. 7, A and B).
Several studies have been carried out on nicotine inhibition of cytokine synthesis. Nicotine exerts immunosuppressive activity through T cell-dependent and -independent mechanisms (Mabley et al., 2002). It also modulates the production of various cytokines (Yoshida et al., 1998). Nicotine inhibits the production of IL-2 and TNF- from human mononuclear cells (Madretsma et al., 1996). Recently, Wang et al. (2003) reported that the nAChR 7 subunit is required for acetylcholine inhibition of macrophage TNF release. Electrical stimulation of the vagus nerve inhibits TNF synthesis in wild-type mice but fails to inhibit TNF synthesis in 7-deficient mice (Wang et al., 2003). The present study revealed that the amounts of TNF- protein in adipocytes is also significantly reduced by nicotine. The mechanism responsible for this remains to be clarified.
TNF- negatively regulates adiponectin production (Fasshauer et al., 2002). Kern et al. (2003) reported that TNF- and adiponectin may be antagonists of each other or that one cytokine may control the expression of the other cytokines. Several agents such as TNF- mediate their effects on insulin metabolism by modulating adiponectin secretion from adipocytes (Ukkola and Santaniemi, 2002). TNF- expression was higher in adiponectin knockout mice, and the administration of adiponectin in these mice resulted in an improvement in insulin resistance along with a decrease in TNF expression (Maeda et al., 2002).
We showed that short-term exposure to nicotine stimulates the secretion of TNF-, adiponectin, and FFA into the culture medium. Also, long-term exposure reduces the expression of TNF- protein in adipocytes but increases the secretion of adiponectin, possibly in part, through decreased TNF- protein production. Our previous studies showed that nicotine reduces insulin resistance in vivo through decreased production of TNF- protein in visceral tissues and reduces hepatic glucose release (Liu et al., 2001, 2003). Together, these results suggest that adipocytes have functional nAChRs and that nicotine reduces TNF- production in adipocytes although continuing to increase the secretion of adiponectin through the activation of nAChRs. Adipose tissues are under sympathetic and parasympathetic control (Kreier et al., 2002). The present study suggests that nicotine temporarily reduces insulin sensitivity by stimulating the secretion of TNF- and FFA, whereas the long-term direct stimulation of nAChRs by nicotine, in addition to autonomic nervous stimulation, contributes to better insulin sensitivity in vivo through the modulatory secretion of adipocytokines.
Although nicotine administration by smoking is unlikely to be a preventative therapy for diabetes due to deleterious effects on other body systems, the discovery of the presence of nAChRs in adipocytes may lead to the development of a specific agonist for adipocytes. This may prove to be an effective therapy for increasing insulin sensitivity as described in our previous paper (Liu et al., 2001).
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Footnotes |
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, tumor necrosis factor-; FFA, free fatty-acid; RT-PCR, reverse transcriptase-polymerase chain reaction; PBS, phosphate-buffered saline; G3PDH, glyceraldehyde-3-phosphate dehydrogenase; bp, base pair(s).
1 Current address: Kishiwada City Hospital, 1001 Gakuhara, Kishiwada, Osaka 596-8501, Japan. 
Address correspondence to: Dr. Masanari Mizuta, Third Department of Internal Medicine, Miyazaki Medical College, Miyazaki University, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan. E-mail: mmizuta{at}fc.miyazaki-med.ac.jp
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References |
|---|
|
TopAbstractMaterials and MethodsResultsDiscussionReferences |
|---|
Albuquerque EX, Alkondon M, Pereira EF, Castro NG, Schrattenholz A, Barbosa CT, Bonfante-Cabarcas R, Aracava Y, Eisenberg HM, and Maelicke A (1997) Properties of neuronal nicotinic acetylcholine receptors: pharmacological characterization and modulation of synaptic function. J Pharmacol Exp Ther 280: 1117-1136.
Bencherif M, Fowler K, Lukas RJ, and Lippiello PM (1995) Mechanisms of up-regulation of neuronal nicotinic acetylcholine receptors in clonal cell lines and primary cultures of fetal rat brain. J Pharmacol Exp Ther 275: 987-994.
Buonanno A, Mudd J, and Merlie JP (1989) Isolation and characterization of the beta and epsilon subunit genes of mouse muscle acetylcholine receptor. J Biol Chem 264: 7611-7616.
Didier M, Berman SA, Lindstrom J, and Bursztajn S (1995) Characterization of nicotinic acetylcholine receptors expressed in primary cultures of cerebellar granule cells. Brain Res Mol Brain Res 30: 17-28.[Medline]
Fasshauer M, Klein J, Neumann S, Eszlinger M, and Paschke R (2002) Hormonal regulation of adiponectin gene expression in 3T3–L1 adipocytes. Biochem Biophys Res Commun 290: 1084-1089.[CrossRef][Medline]
Gotti C, Carbonnelle E, Moretti M, Zwart R, and Clementi F (2000) Drugs selective for nicotinic receptor subtypes: a real possibility or a dream? Behav Brain Res 113: 183-192.[CrossRef][Medline]
Hofmann C, Lorenz K, Braithwaite SS, Colca JR, Palazuk BJ, Hotamisligil GS, and Spiegelman BM (1994) Altered gene expression for tumor necrosis factor-alpha and its receptors during drug and dietary modulation of insulin resistance. Endocrinology 134: 264-270.
Hoss W, Lin JP, Matchett S, and Davies BD (1986) Characterization of noncholinergic nicotine receptors on human granulocytes. Biochem Pharmacol 35: 2367-2372.[CrossRef][Medline]
Hotamisligil SG, Shargill NS, and Spiegelman BM (1993) Adipose expression of tumor necrosis factor-alpha: direct role in obesity-linked insulin resistance. Science (Wash DC) 259: 87-97.
Hu E, Liang P, and Spiegelman BM (1996) AdipoQ is a novel adipose-specific gene dysregulated in obesity. J Biol Chem 271: 10697-10703.
Ke L, Eisenhour CM, Bencherif M, and Lukas RJ (1998) Effects of chronic nicotine treatment on expression of diverse nicotinic acetylcholine receptor subtypes. I. Dose- and time-dependent effects of nicotine treatment. J Pharmacol Exp Ther 286: 825-840.
Kern PA, Di Gregorio GB, Lu T, Rassouli N, and Ranganathan G (2003) Adiponectin expression from human adipose tissue: relation to obesity, insulin resistance and tumor necrosis factor-alpha expression. Diabetes 52: 1779-1785.
Kreier F, Fliers E, Voshol PJ, Van Eden CG, Havekes LM, Kalsbeek A, Van Heijningen CL, Sluiter AA, Mettenleiter TC, Romijn JA, et al. (2002) Selective parasympathetic innervation of subcutaneous and intra-abdominal fat—functional implications. J Clin Investig 110: 1243-1250.[CrossRef][Medline]
LaPolla RJ, Mayne KM, and Davidson N (1984) Isolation and characterization of a cDNA clone for the complete protein coding region of the delta subunit of the mouse acetylcholine receptor. Proc Natl Acad Sci USA 81: 7970-7974.
Lebargy F, Benhammou K, Morin D, Zini R, Urien S, Bree F, Bignon J, Branellec A, and Largue G (1996) Tobacco smoking induces expression of very-high-affinity nicotine binding sites on blood polymorphonuclear cells. Am J Respir Crit Care Med 153: 1056-1063.[Abstract]
Lindstrom J, Anand R, Peng X, Gerzanich V, Wang F, and Li Y (1995) Neuronal nicotinic receptor subtypes. Ann NY Acad Sci 757: 100-116.[Medline]
Liu L, Chang GQ, Jiao YQ, and Simon SA (1998) Neuronal nicotinic acetylcholine receptors in rat trigeminal ganglia. Brain Res 809: 238-245.[CrossRef][Medline]
Liu RH, Kurose T, and Matsukura S (2001) Oral nicotine administration decreases tumor necrosis factor-alpha expression in fat tissues in obese rats. Metabolism 50: 79-85.[CrossRef][Medline]
Liu RH, Mizuta M, and Matsukura S (2003) Long-term oral nicotine administration reduces insulin resistance in obese rats. Eur J Pharmacol 458: 227-234.[CrossRef][Medline]
Mabley JG, Pacher P, Southan GJ, Salzman AL, and Szabo C (2002) Nicotine reduces the incidence of type I diabetes in mice. J Pharmacol Exp Ther 300: 876-881.
Madretsma GS, Donze GJ, van Dijk AP, Tak CJ, Wilson JH, and Zijlstra FJ (1996) Nicotine inhibits the in vitro production of interleukin 2 and tumour necrosis factor-alpha by human mononuclear cells. Immunopharmacology 35: 47-51.[CrossRef][Medline]
Maeda N, Shimomura I, Kishida K, Nishizawa H, Matsuda M, Nagaretani H, Furuyama N, Kondo H, Takahashi M, Arita Y, et al. (2002) Diet-induced insulin resistance in mice lacking adiponectin/ACRP30. Nat Med 8: 731-737.[CrossRef][Medline]
Marubio LM and Changeux J-P (2000) Nicotinic acetylcholine receptor knockout mice as animal models for studying receptor function. Eur J Pharmacol 393: 113-121.[CrossRef][Medline]
Maslinski W, Laskowska-Bozek H, and Ryzewski J (1992) Nicotinic receptors of rat lymphocytes during adjuvant polyarthritis. J Neurosci Res 31: 336-340.[CrossRef][Medline]
Matsuzawa Y, Funahashi T, and Nakamura T (1999) Molecular mechanism of metabolic syndrome X: contribution of adipocytokines adipocyte-derived bioactive substances. Ann NY Acad Sci 892: 146-154.[CrossRef][Medline]
Maus ADJ, Pereira EFR, Karachunski PI, Horton RM, Navaneetham D, Macklin KD, Cortes WS, Albuquerque EX, and Conti-Fine BM (1998) Human and rodent epithelial cells express functional nicotinic acetylcholine receptors. Mol Pharmacol 54: 779-788.
Mohamed-Ali V, Pinkney JH, and Coppack SW (1998) Adipose tissue as an endocrine and paracrine organ. Int J Obes Relat Metab Disord 22: 1145-1158.[CrossRef][Medline]
Nguyen VT, Ndoye A, and Grando SA (2000) Novel human 9 acetylcholine receptor regulating keratinocyte adhesion is targeted by pemphigus vulgaris autoimmunity. Am J Pathol 157: 1377-1391.
Rohwedel J, Horak V, Hebrok M, Fuchtbauer EM, and Wobus AM (1995) M-twist expression inhibits mouse embryonic stem cell-derived myogenic differentiation in vitro. Exp Cell Res 220: 92-100.[CrossRef][Medline]
Rubboli F, Court JA, Sala C, Morris C, Chini B, Perry E, and Clementi F (1994) Distribution of nicotinic receptors in the human hippocampus and thalamus. Eur J Neurosci 6: 1596-1604.[CrossRef][Medline]
Saito T, Ohnuki Y, Saeki Y, Nakagawa Y, Ishibashi K, Yanagisawa K, and Yamane A (2002) Postnatal changes in the nicotinic acetylcholine receptor subunits in rat masseter muscle. Arch Oral Biol 47: 417-421.[CrossRef][Medline]
Saltiel AR and Kahn CR (2001) Insulin signalling and the regulation of glucose and lipid metabolism. Nature (Lond) 414: 799-806.[CrossRef][Medline]
Tassonyi E, Charpantier E, Muller D, Dumont L, and Bertrand D (2002) The role of nicotinic acetylcholine receptors in the mechanisms of anesthesia. Brain Res Bull 57: 133-150.[CrossRef][Medline]
Tseng J, Kwitek-Black AE, Erbe CB, Popper P, Jacob HJ, and Wackym PA (2001) Radiation hybrid mapping of 11 alpha and beta nicotinic acetylcholine receptor genes in Rattus norvegicus. Brain Res Mol Brain Res 91: 169-173.[Medline]
Ukkola O and Santaniemi M (2002) Adiponectin: a link between excess adiposity and associated comorbidities? J Mol Med 80: 696-702.[CrossRef][Medline]
Wang H, Yu M, Ochani M, Amella CA, Tanovic M, Susarla S, Li JH, Wang H, Yang H, Ulloa L, et al. (2003) Nicotinic acetylcholine receptor alpha7 subunit is an essential regulator of inflammation. Nature (Lond) 421: 384-388.[CrossRef][Medline]
Witzemann V, Stein E, Barg B, Konno T, Koenen M, Kues W, Criado M, Hofmann M, and Sakmann B (1990) Primary structure and functional expression of the alpha-, beta-, gamma-, delta- and epsilon-subunits of the acetylcholine receptor from rat muscle. Eur J Biochem 194: 437-448.[Medline]
Wonnacott S (1987) Brain nicotine binding sites. Hum Toxicol 6: 343-353.[Medline]
Yamauchi T, Kamon J, Waki H, Imai Y, Shimozawa N, Hioki K, Uchida S, Ito Y, Takakuwa K, Matsui J, et al. (2003) Globular adiponectin protected ob/ob mice from diabetes and apoE-deficient mice from atherosclerosis. J Biol Chem 278: 2461-2468.
Yoshida H, Sakagami H, Yamanaka Y, Amano Y, Yamaguchi M, Yamamura M, Fukuchi K, Gomi K, Ohata H, Momose K, et al. (1998) Induction of DNA fragmentation by nicotine in human myelogenous leukemic cell lines. Anticancer Res 18: 2507-2511.[Medline]
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, p < 0.01 versus 1 x 10-6 M group.
