2013 List_The role of GH in adipose tissue-LiGHRKO mice

The Role of GH in Adipose Tissue: Lessons fromAdipose-Specific GH Receptor Gene-Disrupted Mice

Edward O. List, Darlene E. Berryman, Kevin Funk, Elahu S. Gosney, Adam Jara,Bruce Kelder, Xinyue Wang, Laura Kutz, Katie Troike, Nicholas Lozier,Vincent Mikula, Ellen R. Lubbers, Han Zhang, Clare Vesel, Riia K. Junnila,Stuart J. Frank, Michal M. Masternak, Andrzej Bartke, and John J. Kopchick

Edison Biotechnology Institute (E.O.L., D.E.B., K.F., E.S.G., A.J., B.K., X.W., L.K, K.T., N.L., V.M., E.R.L.,H.Z., C.V, R.K.J., J.J.K.), Department of Specialty Medicine (E.O.L.), School of Applied Health Sciencesand Wellness (D.E.B.), Department of Biomedical Sciences (D.E.B., A.J., J.J.K.), Department of Pediatrics(B.K.), Heritage College of Osteopathic Medicine, Ohio University, Athens, Ohio 45701; Department ofMedicine (S.J.F.), Division of Endocrinology, Diabetes and Metabolism, University of Alabama atBirmingham, Birmingham, Alabama; College of Medicine (M.M.M.), Burnett School of BiomedicalSciences, University of Central Florida, Orlando, Florida 32827; Institute of Human Genetics (M.M.M.),Polish Academy of Sciences, Strzeszynska 32, 60–479 Poznan, Poland; and Department of InternalMedicine (A.B.), Geriatrics Research, Southern Illinois University School of Medicine, Springfield, Illinois62702

GH receptor (GHR) gene-disrupted mice (GHR�/�) have provided countless discoveries as to thenumerous actions of GH. Many of these discoveries highlight the importance of GH in adiposetissue. For example GHR�/� mice are insulin sensitive yet obese with preferential enlargement ofthe sc adipose depot. GHR�/� mice also have elevated levels of leptin, resistin, and adiponectin,compared with controls leading some to suggest that GH may negatively regulate certain adipo-kines. To help clarify the role that GH exerts specifically on adipose tissue in vivo, we selectivelydisrupted GHR in adipose tissue to produce Fat GHR Knockout (FaGHRKO) mice. Surprisingly,FaGHRKOs shared only a few characteristics with global GHR�/� mice. Like the GHR�/� mice,FaGHRKO mice are obese with increased total body fat and increased adipocyte size. However,FaGHRKO mice have increases in all adipose depots with no improvements in measures of glucosehomeostasis. Furthermore, resistin and adiponectin levels in FaGHRKO mice are similar to controls(or slightly decreased) unlike the increased levels found in GHR�/� mice, suggesting that GH doesnot regulate these adipokines directly in adipose tissue in vivo. Other features of FaGHRKO miceinclude decreased levels of adipsin, a near-normal GH/IGF-1 axis, and minimal changes to a largeassortment of circulating factors that were measured such as IGF-binding proteins. In conclusion,specific removal of GHR in adipose tissue is sufficient to increase adipose tissue and decreasecirculating adipsin. However, removal of GHR in adipose tissue alone is not sufficient to increaselevels of resistin or adiponectin and does not alter glucose metabolism. (Molecular Endocrinology27: 0000–0000, 2013)

The GH/IGF-1 axis has important roles in growth, me-tabolism, and lifespan. Because GH signaling requires

the binding of GH to its cognate receptor (R), disruptionor deletion of either the GH or GHR gene ablates GH

action. In this regard, GHR-disrupted or knockout mice(GHR�/� mice), which were generated in our laboratoryapproximately 15 yr ago, have produced many novel andclinically relevant discoveries that have played an impor-

ISSN Print 0888-8809 ISSN Online 1944-9917Printed in U.S.A.Copyright © 2013 by The Endocrine Societydoi: 10.1210/me.2012-1330 Received October 11, 2012. Accepted December 10, 2012.

Abbreviations: BAT, brown adipose tissue; CRP, C-reactive protein; ECL, enhanced chemi-luminescence; FAS, fatty acid synthase; GHR, GH receptor; GHRKO, GHR knockout; GTT,glucose tolerance test; HMW, high molecular weight; IGFBP, IGF-binding protein; ITT,insulin tolerance test; KOMP, Knockout Mouse Project; LS, Laron syndrome; PPAR, per-oxisome proliferator-activated receptor; sTNFR, soluble TNF receptor; sVEGFR, solublevascular endothelial growth factor receptor.

O R I G I N A L R E S E A R C H

Mol Endocrinol, March 2013, 27(3):0000–0000 mend.endojournals.org 1

Molecular Endocrinology. First published ahead of print January 24, 2013 as doi:10.1210/me.2012-1330

Copyright (C) 2013 by The Endocrine Society

tant role in our understanding of GH function (1).GHR�/� mice are dwarf with low levels of IGF-1 andincreased GH; thus, they are GH resistant (2). They arealso extremely insulin sensitive (3), presumably due to theabsence of the anti-insulin effects of GH. Remarkably,they exhibit up to a 50% increase in lifespan that cannotbe further extended with calorie restriction (4, 5). Theextended lifespan in GHR�/� mice is associated withlower morbidity and disease-related mortality, with al-most half of the long-lived mice dying without obviouslethal pathological lesions as compared with 10% of theirwild-type littermate controls (6). Importantly, the uniquephenotype of GHR�/� mice has notable similarities witha population of Ecuadorian Laron Syndrome (LS) indi-viduals (7). This population has a reduction in IGF-1 lev-els, an elevation in GH levels, and enhanced insulin sen-sitivity. Thus far, these individuals do not appear toexperience life extension, but they are protected from di-abetes and fatal neoplasms. The increased insulin sensi-tivity in LS individuals and GHR�/� mice is particularlyinteresting considering that both are obese, a characteris-tic not typically associated with improved glucose homeo-stasis or lifespan extension.

The adiposity of GHR�/� mice has been extensivelystudied. GHR�/� mice have a significantly higher per-cent body fat throughout their lifespan (8) with a dispro-portionate amount of fat deposition in the sc white adi-pose tissue (WAT) depot (8). In terms of adipokineexpression, leptin levels (9) are elevated in GHR�/�mice, which is consistent with their increased obesity.Interestingly, adiponectin levels, which are usually nega-tively correlated with obesity, are elevated in GHR�/�mice (10). Adiponectin is an important adipokine withbeneficial effects on inflammation and insulin sensitivityand is positively correlated with increased longevity in an-imals and humans (11–13). Furthermore, adiponectin hasbeen reported to be negatively regulated by GH in multi-ple systems (14, 15). As stated above, GHR�/� mice areobese throughout life and have high adiponectin levels.This suggests that the lack of GH action, either directlyvia negative regulation by GH or indirectly through otherphysiological alterations in these mice, overrides the pres-ence of obesity. Furthermore, the increased adiponectinmay also contribute to the improved glucose metabolismof GHR�/� mice. Although less thoroughly studied, LSindividuals have increased serum levels of adiponectinand leptin as well as notable enlargements in WAT (7,16). Taken together, LS individuals and GHR�/� miceprovide a means to better understand how adipose tissuemass can be enlarged without notable deleterious effectson health and lifespan. Because GHR signaling is dis-rupted in all tissues of LS patients and GHR�/� mice, it

would be of value to disrupt GH signaling selectively inadipose tissue to better understand the impact of this tis-sue on whole-body metabolism and physiology.

Multiple groups have utilized the Cre-LoxP system toevaluate GHR disruption in various tissues and cell types.Liver-specific deletion of GHR results in no majorchanges in adiposity although these mice are reported tohave marked insulin resistance and severe hepatic steato-sis (17). Muscle-specific deletion of GHR has been re-ported by two groups with different results. Mavalli et al(18) report peripheral adiposity, insulin resistance, andglucose intolerance using the Mef-2c promoter/enhancerwhereas mice generated using the muscle creatine kinasepromoter/enhancer have reduced adiposity and overallimprovement in glucose metabolism (19). Disruption ofGHR in �-cells impairs insulin secretion that is exacer-bated by a high-fat diet (20). Collectively, these resultsdemonstrate that disruption of GHR in specific tissuescan dramatically influence glucose homeostasis and adi-posity. To date, no studies have assessed GHR gene dis-ruption in adipose tissue. Because GH’s action on adiposetissue plays an essential metabolic role in terms of whole-body physiology, we set out to generate and characterizeadipose tissue GHR gene-disrupted mice. In this study, weused the Cre-LoxP system to disrupt the GHR�/� gene inadipose tissue to produce Fat GHR Knockout(FaGHRKO) mice. We hypothesized that disruption ofthe GHR in adipose tissue will 1) increase adiposity, 2)increase leptin and adiponectin levels, and 3) improveglucose homeostasis, all of which occur in globalGHR�/� mice. Here, we report initial characterizationof these mice including adiposity, adipokine levels, andglucose metabolic results as well as effects on morpho-metric, endocrine, and physiological parameters. We alsodiscuss these results in comparison with previous reportscharacterizing the global GHR�/� mice to provide noveland more specific insight into the specific role of GH inadipose tissue.

Materials and Methods

FaGHRKO mouse productionThe mouse strain carrying the conditional GHR floxed allele

(GHRflox/flox) was generated by the Knockout Mouse Project(KOMP) as previously described (21). Adipose tissue-specificGHR�/� mice (FFCx) and floxed littermate controls (FFxx)were generated by breeding conditional GHRflox/flox mice toB6.Cg-Tg(Fabp4-cre)1Rev/J mice purchased from The JacksonLaboratory (Bar Harbor, Maine). B6.Cg-Tg(Fabp4-cre)1Rev/Jmice have been crossed to C57BL/6 mice for nine generations atThe Jackson Laboratory.

2 List et al Disruption of the GHR Gene in Adipose Tissue Mol Endocrinol, March 2013, 27(3):0000–0000

In the current study, 146 mice divided into two main cohortswere used. The first cohort of male and female FaGHRKO andlittermate controls (63 total, n � 15–16 per group) were used forall measurements except body composition over time. The sec-ond cohort of FaGHRKO and littermate controls of both sexes(83 total, n � 16–25 per group) were used only to collect lon-gitudinal body composition. Mice were housed three to four percage and given ad libitum access to water and standard labora-tory rodent chow (ProLab RMH 3000). The cages were main-tained in a temperature- and humidity-controlled room and ex-posed to a 14-hour light, 10-hour dark cycle. All procedureswere approved by the Ohio University Institutional AnimalCare and Use Committee.

Quantitative real-time PCRWhole frozen tissue was homogenized using the Precellys

24-Dual (Bertin Technologies, Montigny-le-Bretonneux, France).The homogenization conditions were optimized for each tissue.RNA was purified using Qiagen RNeasy Mini Kit (QIAGEN,Chatsworth, Calfornia), and the concentration and integrity ofthe mRNA was verified by the Thermo NanoDrop 2000c andAgilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, Cal-ifornia), respectively. Qiagen QuantiTect Reverse TranscriptionKit was used for cDNA synthesis. For real-time data collection,Qiagen QuantiTect SYBR Green PCR Kit was used with a BIO-RAD iCycler Thermal Cycler (Bio-Rad Laboratories, Inc, Her-cules, California). Qbase Plus from Biogazelle (Zwijnaarde, Bel-gium) was used to analyze the qPCR results. Each tissue wastested with a pair of primers for the GHR gene as well as sevenreference genes (EEF2, RPS3, B2M, ACTB, HPRT, EIF3F, andRPL38). For all other qPCR, the reactions and calculations wereperformed as previously described (22).

Western blot analysis of GHRFrozen tissue samples were homogenized in lysis buffer (1%

vol/vol Triton X-100, 150 mM NaCl, 10% vol/vol glycerol, 50mM Tris-HCl [pH 8.0], 100 mM NaF, 2 mM EDTA, 1 mMphenylmethylsulfonyl fluoride, 1 mM sodium orthovanadate,10 �g/mL aprotinin) and proteins resolved by 8% SDS-PAGE.After electrophoresis, proteins were transferred to an Amer-sham Hybond-enhanced chemiluminescence (ECL) membrane(GE Healthcare, Pittsburgh, Pennsylvania), and the membraneincubated in Tris buffered saline (TBS) containing 2% gelatinand 0.05% Tween 20. The membrane was then rinsed briefly inTBS/Tween and incubated with rabbit anti-GHR primary anti-body (AL47) in 1% gelatin-TBS/Tween overnight (23). The fol-lowing day, the membranes were washed and incubated withsecondary antibody for 2 h (NA934, ECL Antirabbit IgG HRPLinked, GE Healthcare, Piscataway, New Jersey) in 1% gelatin-TBS/Tween 20. The membrane was then washed and incubatedfor 5 minutes with ECL Plus Western Blotting Detection System(RPN2132, GE Healthcare) and exposed to Kodak BiomaxXAR film (Eastman Kodak, Rochester, New York).

Body composition measurementsBody composition was measured in two cohorts of mice. In

the first cohort, FaGHRKO and littermate controls (n � 15–16per group per sex) were measured at 5 months of age beforedissection and subsequent tissue analysis at 6 mo. In the secondcohort, FaGHRKO and littermate controls of both sexes (n �

16–25 per group per sex) were measured over time starting at 2months until 12 months of age. Body composition was mea-sured using a Bruker Minispec NMR (Bruker Corp, The Wood-lands, Texas) as previously described (8, 24).

Fasting blood glucose, glucose tolerance test (GTT),and insulin tolerance test (ITT) measurements

Fasting blood glucose was determined at 5 months of ageusing OneTouch Ultra test strips and glucometers (Lifescan, Inc,Milpitas, California). Blood samples were obtained by cuttingapproximately 1 mm from the tip of the tail and collecting thefirst drop of blood. Fasting blood glucose measurements oc-curred starting at 9:00 AM after a 12-hour overnight fast. GTTswere performed at 5 months and 1 week of age. Mice werefasted for 12 hours before commencement of the experiment at9:00 AM. Each mouse received an ip injection of 10% glucose ata dose of 1 g/kg body weight. Blood glucose measurements weremonitored before the glucose injection and at 15, 30, 45, 60, and90 minuntes after injection. ITTs were performed at 5 monthsand 2 weeks of age in a fed state at approximately 3:00 PM.Recombinant human insulin (Humulin-R; Eli Lilly & Co, Indi-anapolis, Indiana) was prepared by diluting Humulin-R (100U/ml) to 0.075 U/mL in sterile 0.9% NaCl. Each mouse receivedan ip injection of the 0.075 U/ml insulin solution at a dose of0.75U/kg body weight. Blood glucose measurements were per-formed before the insulin injection and at 15, 30, 45, and 60minutes after injection.

Serum measurementsSerum measurements were performed at 6 mo of age. Serum

was collected starting at approximately 9:00 AM after a 12-hfast. IGF-1 levels (total IGF-1) were measured using IGF-1(mouse, rat) ELISA kits (Catalog no. 22-IG1MS-E01; ALPCODiagnostics, Salem, New Hampshire). High molecular weight(HMW) and total adiponectin levels were measured usingELISA kits (47-ADPMS-E01) from ALPCO Diagnostics (Salem,New Hampshire). Insulin, c-peptide, leptin, resistin, and gastricinhibitory polypeptide were measured using a Mouse MetabolicPanel (catalog no. MMHMAG-44K; Millipore Corp., Billerica,Massachusetts). IGF binding proteins-1, -2, -3, -5, -6, and -7were measured using the Mouse IGF Binding Protein MAG-NETIC Bead Panel (catalog no. MIGFBPMAG-43; MilliporeCorp.). Adipsin, AGP, �-2-macroglobulin, C-reactive protein(CRP), and haptoglobin were measured using the MILLIPLEXMAP Mouse Acute Phase Magnetic Bead Panel 2 (catalog no.MAP2MAG-76K; Millipore Corp.). Soluble receptors (sCD30,sgp130, sIL-1RI, sIL-1RII, sIL-2Ra, sIL-4R, sIL6R, sTNFRI,sTNFRII, soluble vascular endothelial growth factor receptor(sVEGFR)1, sVEGFR2, sVEGFR3) were measured using theMILLIPLEX MAP Mouse Soluble Cytokine Receptor Panel(catalog no. MSCR-42K, Millipore Corp.). Lipocalin-2 andpentraxin-3 were measured using the MILLIPLEX MAP MouseAcute Phase Magnetic Bead Panel-1 (catalog no. MAP1MAG-76K; Millipore Corp.). All MILLIPEX kits were analyzed usinga Milliplex 200 Analyzer (Millipore Corp.). All of the aboveprocedures were performed according to the manufacturer’sinstructions.


Tissue collection, liver, and WAT analysisAll tissue was collected from 6-mo-old mice. Mice were

killed starting at 9:00 AM following a 12-hour overnight fast.Mice were first placed in a CO2 chamber until unconscious,after which blood was quickly collected from the orbital sinus.After blood collection, the mice were killed by cervical disloca-tion. Kidney, heart, lung, spleen, brain, skeletal muscle (gastroc-nemius, soleus, and quadriceps), and interscapular BAT werecollected, weighed, and flash frozen in liquid nitrogen andstored at �80ºC.

Liver tissue was collected and weighed, and a portion wasflash frozen in liquid nitrogen and stored at �80ºC until pro-cessing for determining triglyceride content, while another por-tion was fixed in 10% formalin, embedded in paraffin, andprocessed for histology. For determining triglyceride content,liver tissues were thawed and used for extraction and measure-ment of triacylglycerol levels as described previously (25).

Subcutaneous, retroperitoneal, mesenteric, and perigonadalWAT were collected and weighed. A portion of WAT was flashfrozen in liquid nitrogen and stored at �80ºC. For subcutaneousand perigonadal WAT, a portion of the sample was processedfor histology by fixing in 10% formalin and embedding in par-

affin. Adipocyte cell size and number weredetermined as previously described (26).

Statistical analysisAll values are given as means � SEMs.

Statistics were performed using SPSS ver-sion 14.0 (Chicago, Illinois). The two-tailed unpaired Student’s t test was used toassess the significance of difference be-tween two sets of data. Differences wereconsidered to be statistically significantwhen P � .05.

Results

FaGHRKO miceFaGHRKO mice (FFCx) and floxed

littermate controls (FFxx) were gener-ated by breeding conditional floxedGHRflox/flox mice to B6.Cg-Tg(Fabp4-cre)1Rev/J mice (Figure 1, A and B). Ab-sence of GHR protein in WAT ofFaGHRKO mice was shown by Westernblot analysis (Figure 1C). Adipose tissue-specific deletion of GHR was quantifiedusing qPCR in FaGHRKO mice (FFCx)and floxed littermate controls (FFxx)(Figure 1D). In the FaGHRKO mice,GHR mRNA expression is significantlydecreased 89%, 77%, 85%, 60%, and92% in epididymal, mesenteric, retro-peritoneal, and sc WAT, and in brownadipose tissue (BAT), respectively, vscontrols. No change in GHR gene ex-

pression was observed for liver, lung, kidney, brain, skeletalmuscle, or heart.

Body weight and body compositionFive-month mean body weights of FaGHRKO mice

that were also used for dissection were significantlygreater than littermate controls with 14% and 23% in-creases for male and female, respectively (Figure 2A).Body composition analysis showed a 96% increase intotal body fat mass in FaGHRKO mice compared withcontrols (Figure 2B). Total lean body mass was signifi-cantly increased in female (8%) but not male FaGHRKOmice compared with controls (Figure 2C). Additionally,total body fluid was significantly increased in both sexesof FaGHRKO mice (Figure 2D).

Body composition measurements over time showedthat the increase in body weight became significant forFaGHRKO males by 3 mo of age and for FaGHRKO

Figure 1. Generation and characterization of FaGHRKO mice. A, FaGHRKO mice weregenerated by crossing mice with a “floxed” exon 4 of the GHR to transgenic mice thatexpress Cre recombinase under the control of the aP2 promoter/enhancer (aP2-Cre). Solidarrowheads depict the LoxP sites. B, PCR analysis detected the presence of the LoxP sites andthe aP2-Cre transgene. C, Western blot analysis of GHR protein from WAT in FaGHRKO(FFCx) vs controls (FFxx). D, GHR mRNA expression level in various tissues from FaGHRKO mice(n � 6; black bars) compared with controls (n � 6; white bars). GHR expression wassignificantly decreased in both WAT and BAT. No changes to GHR gene expression wereobserved for liver, lung, kidney, brain, skeletal muscle (SM), or heart. Epi, epididymal; Mes,mesenteric; Retro, retroperitoneal; Std, standard; SubQ, subcutaneous.


females by 5 months of age (Figure 2, E and H). Thisdifference continued to increase with age. Fat mass wassignificantly increased at 2 months in male FaGHRKOmice (Figure 2F). Although exhibiting a similar trend,female FaGHRKO mice had a delayed increase in fat masswith a significant difference not seen until 4 months of age(Figure 2I). Lean mass for FaGHRKO males increasedonly at 10 mo of age (Figure 2G), whereas FaGHRKOfemales had a significant increase from 5–10 months ofage (Figure 2J).

Adipose tissue depot mass and adipocyte sizeand number

WAT mass was significantly increased in all four de-pots for female FaGHRKO mice compared with controls.In males, all depots but the perigonadal fat pad wereincreased (Figure 3). The sc depot in FaGHRKO mice wasthe most impacted by removal of the GHR because itshowed the largest increase in males (204% increase) andfemales (237% increase). In male FaGHRKO mice, theretroperitoneal depot was increased by 74% and the mes-enteric depot was increased by 111%, whereas the 20%increase in the perigonadal depot did not reach statistical

significance. In female FaGHRKO mice, the retroperito-neal depot was increased by 188%, the mesenteric depotwas increased by 124%, and the perigonadal depot wasincreased by 116%. The mass of BAT collected fromFaGHRKO mice was significantly increased comparedwith littermate controls with 87% and 93% increases formale and female, respectively.

Subcutaneous and perigonadal WAT depots were an-alyzed for cell size and number. Adipocyte cell size wassignificantly increased in the sc depot (Figure 3F) in bothmale FaGHRKO (157% compared with controls) andfemale FaGHRKO mice (135% compared with controls).Adipocytes from the perigonadal depot (Figure 3G) weremodestly increased in FaGHRKO mice. Adipocyte cellnumbers did not differ in sc or perigonadal adipose depots(data not shown).

Organ weight and liver triglyceride contentNo change was observed in the absolute weights of

liver, heart, lung, brain, gastrocnemius muscle, and quad-riceps between FaGHRKO vs control mice of males (Fig-ure 4A) or females (Figure 4B). However, several sex-specific differences were observed in other organs. For

Figure 2. GHR deletion in adipose tissue increases total body fat mass. Two cohorts of mice were studied, including 63 mice (n � 15–16 animalsper group) that were used to measure body composition at 6 months of age before a 6-month dissection (A–D) and 83 mice (n � 16–25 animalsper group) that were used to measure body composition over time (E–J). For the 63 mice used for dissections, body weight (A), fat mass (B), leanmass (C), and fluid mass (D) are shown for FaGHRKO (FFCx; black bars) and controls (FFxx; white bars) in both males and females. Body weight (Eand H), fat mass (F and I), and lean mass (G and J) are shown for males and females over time. FaGHRKO (FFCx; black boxes) and controls (FFxx;white circles) are indicated. Values in A–J are represented as mean � SEM (n � 15–25 per group). *P � .05; **P � .01; ***P � .001, FaGHRKO(FFCx) vs control (FFxx). BW, body weight.


example, kidneys and soleus muscle from FaGHRKOmice were significantly larger than controls in females butnot in males, whereas the spleens from FaGHRKO micewere smaller than controls in males but normal in females.Liver triglyceride levels did not differ in FaGHRKO mice com-pared with controls (Figure 4C).

AdipokinesMean values for circulating leptin were only signifi-

cantly increased in female FaGHRKO mice (Table 1).Resistin levels were unchanged in FaGHRKO mice com-pared with controls. Mean values for total and HMWadiponectin were decreased slightly in both male and fe-male FaGHRKO mice with only male total adiponectinvalues reaching statistical significance (P � .047). Circu-lating levels of adipsin were significantly decreased inboth male and female FaGHRKO mice compared withcontrols.

Glucose metabolismFasting blood glucose, serum insulin, and C-peptide

did not differ between FaGHRKO and controls (Figure5). GTTs and ITTs also showed no significant differencesbetween FaGHRKO mice and controls. No changes wereseen in RNA levels encoding intracellular signaling mol-ecules that are known to affect glucose metabolism and

other processes in WAT from FaGHRKO mice including:peroxisome proliferator-activated receptor (PPAR)� co-activator 1, sirtuin1 (SIRT1), PPAR�, PPAR�, PPAR�,4-1BB, sterol regulatory element binding protein(SREBP), fatty acid synthase (FAS), and glucose trans-porter (GLUT)4 (Figure 6).

GH, IGF-1, and IGF-binding proteins (IGFBPs)GH levels were not significantly different in FaGHRKO

mice relative to controls. Serum IGF-1 levels were ele-vated in FaGHRKO compared with controls for bothmales (22%) and females (8%); however, only in malesdid this reach statistical significance (Table 2). No signif-icant changes were observed for any of the IGFBPs exceptfor IGFBP-5, which was significantly increased (19%) infemale FaGHRKO mice (P � .033) but did not quitereach statistical significance in males (P � .06).

Other blood parametersCirculating levels of IL-6 were significantly elevated in

female FaGHRKO compared with controls (P � .005),whereas no change was seen in males (Table 3). MCP-1did not differ between FaGHRKO and controls for eithersex. Circulating levels of acute phase peptides includinglipocalin-2, pentraxin-3, AGP, CRP, �-2-macroglobulin,and haptoglobin did not differ between FaGHRKO and

Figure 3. FaGHRKO mice have increased adipose tissue depot weight and increased adipocyte cell size. Adipose tissue was collected at 6 mo ofage. Subcutaneous (A), perigonadal (B), retroperitoneal (C), and mesenteric (D) WAT depots as well as BAT (E) are shown for male and femaleFaGHRKO (FFCx; black bars) and controls (FFxx; white bars). Mean adipocyte cell size is shown for sc (F) and perigonadal (G) depots. Hematoxylinand eosin (H&E) staining of sc (top four images) and perigonadal (bottom four images) adipose tissue depots are shown. Values in A–G arerepresented as mean � SEM (n � 15–16 per group). *P � .05; **P � .01; ***P � 0.001, FaGHRKO (FFCx) vs control (FFxx). Mes, mesenteric; Peri,perigonadal; Retro, retroperitoneal; SubQ, subcutaneous.


controls regardless of sex. A large number of soluble se-rum receptors including sgp130, sIL-1RI, sIL-1RII, sIL-2Ra, sIL6R, sTNFRI, sTNFRII, sVEGFR1, sVEGFR2,and sVEGFR3 were similar between FaGHRKO and con-trols regardless of sex. However, sCD30 (decreased infemale FaGHRKO) and sIL-4R (increased in maleFaGHRKO) differed between FaGHRKO and controls.

Discussion

The GHR�/� mouse was generated inour laboratory nearly 15 yr ago (2).These mice are dwarf with low IGF-1and high GH levels. GHR�/� mice areobese with the sc WAT depot beingpreferentially increased (8). Despite theobese phenotype, the mice are insulinsensitive with very low levels of seruminsulin (4). Interestingly, these mice arelong-lived (4) with decreased rates ofcancer (6). Another interesting findingis that GHR�/� mice have elevatedlevels of adiponectin despite beingobese (10). In an effort to determine thetissues responsible for the above men-tioned phenotypes, attempts are beingmade to disrupt the GHR gene in a tis-sue-specific manner. Others have al-ready reported on muscle-, liver-, andpancreas-specific GHRKO mice (17–19). Here, we describe the fat GHRknockout (FaGHRKO) mouse.

To delete the GHR in WAT,GHRflox/flox mice were crossed withtransgenic aP2-cre mice. This cre pro-moter/enhancer has been used by manyfor disruption of a variety of genes inadipose tissue (27–30). Whereas aP2expression is induced in nonadipogenictissues during early development, al-beit in cells that are of an analogouscell lineage (31), studies that have ana-lyzed adult tissues show a specific lo-

calization to WAT and BAT (29, 30). Our results supportthe expression of aP2 specifically in adult adipose tissuebecause the levels of GHR mRNA were reduced in fourWAT depots and interscapular BAT but not significantlyaltered in any other tissue tested. It has been previouslyclaimed that aP2 expression is induced in activated mac-

Figure 4. Organ weights and liver tryglyceride content. A total of 63 mice were dissected at6 mo of age. A and B, Liver, kidney, spleen, lung, heart, skeletal muscle (gastrocnemius,soleus, and quadriceps), and brain, are shown for FaGHRKO (FFCx; black bars) and controls(FFxx; white bars) for males (A) and females (B). C, Liver triglyceride is shown for FaGHRKOmice and controls for both sexes. Values in A–C are represented as mean � SEM (n � 15–16per group). *P � .05, FaGHRKO (FFCx) vs control (FFxx). Gast, gastrocnemius; Quad,quadriceps; Sol, soleus.

Table 1. Serum Adipokine Levels of FaGHRKO and Control Male and Female Mice

Fat GHR�/� (Male) Fat GHR�/� (Female)

FFxx FFCx P Value FFxx FFCx P ValueTotal adiponectin, n � 9–10 (pg/ml) 22 302 � 1227 18 782 � 1106a .047 43 736 � 7135 39 642 � 12 224 .372HMW adiponectin, n � 9–10 (pg/ml) 3923 � 1412 3405 � 864 .335 11 214 � 3429 10 922 � 6816 .905Leptin, n � 15–16 (pg/ml) 3850 � 736 5451 � 835 .160 3183 � 739 6267 � 1057a .023Resistin, n � 15–16 (pg/ml) 11 347 � 857 13 318 � 1575 .272 11 427 � 1216 11 779 � 907 .818Adipsin, n � 9–10 (pg/ml) 1816 � 88 1117 � 100a 8.E-05 2020 � 187 1248 � 153a .005

Abbreviations: FFxx, controls; FFCx, FaGHRKO. Adipokine values of mice at 6 months of age. Values are represented as mean � SEM (n � 9–16per group). a Indicates significance with P values given to the right.


rophages (32). However, more recent reports show thatthe efficiency of Cre recombination in macrophages ismuch less than that in adipocytes (27, 28). Because wecannot rule out macrophage expression or embryonic ex-pression, these possibilities need to be taken into consid-eration when interpreting our results.

The body weights of male and female FaGHRKO micewere significantly increased. This weight gain was mainlyattributed to a near doubling of fat mass. The increase infat mass was expected because GH possesses lipolytic andantilipogenic effects on adipose tissue, and removal of thisaction should result in increased fat mass (33, 34). More-over, GHR�/� mice have repeatedly been shown to beobese relative to littermate controls throughout life (8).This is also true for FaGHRKO mice up to 1 year of age;thus, our first hypothesis that FaGHRKO mice wouldhave increased adiposity is supported. However, there areimportant differences in the adiposity of FaGHRKO micerelative to GHR�/� mice. In this study, all adipose depots(four WAT depots and interscapular BAT) analyzed with

the exception of the perigonadal (epidid-ymal) fat pad in males were significantlyenlarged in the FaGHRKO mice com-pared with controls. The fact thatperigonadal was the lone exception inmales is not surprising because previousstudies have shown this particular depotto be the least responsive to GH treat-ment (24). In contrast to FaGHRKOmice, our laboratory and others haveshown that GHR�/� mice in a similarC57BL/6 genetic background have apreferential enlargement of the sc depotand occasionally the retroperitoneal de-pot with other fat pads being propor-tional to their dwarf size (8, 35). It hasbeen proposed that the obesity inGHR�/� mice may represent a form of“healthy” obesity because of the prefer-ential accumulation of excess of sc adi-pose tissue (10). Although the sc WATmass did show the largest increase inmass compared with other depots, all fatpads (except for perigonadal in males) inthe FaGHRKO mice were enlarged. Theenlargement of most depots, as seen inFaGHRKO mice, may not provide thesame benefit to lifespan and health asseen in the GHR�/� mice. Ongoing lon-gevity studies will provide important in-formation about the long-term outcomeof this alternative fat deposition.

Other than leptin, circulating adipokine levels in theFaGHRKO mice are distinct from what has previouslybeen reported for GHR�/� mice. Leptin levels in maleGHR�/� mice are consistently elevated although femalemice are less thoroughly studied (12). Likewise, leptinlevels are increased in female FaGHRKO mice, whereasthe increase in males did not reach statistical significance.The elevation of leptin in both GHR�/� and femaleFaGHRKO mice is not surprising considering that thesemice are obese, and this hormone has been shown to beconsistently and positively correlated with an increase infat mass. However, the physiological consequences of el-evated leptin in these mice have not been thoroughly stud-ied. Possible connections between elevated leptin and al-terations in adaptive and innate immunity (36) as well asorgan function and disease states (37) are worthy of fur-ther exploration. However, it should also be noted thatobese states are typically associated with leptin resistance(38); thus, the increased leptin in these mice may not be

Figure 5. GHR deletion in adipose tissue does not alter glucose homeostasis. Fasting bloodglucose (A), fasting serum insulin (B), and c-peptide (C) are shown for FaGHRKO (FFCx; blackbars) and controls (FFxx; white bars). D and E, GTTs were performed at 5 months and 1 weekof age after a 12-hour fast by ip injection of a 10% glucose solution at 0.01 ml/g bodyweight for FaGHRKO (FFCx; black boxes) and controls (FFxx; white circles) in males (D) andfemales (E). F and G, ITTs were performed in males (F) and females (G) at 5 months and 2weeks of age in nonfasted mice via ip injection of 0.075 U/ml insulin solution at 0.01 ml/gbody weight. Values in A–G are represented as mean � SEM (n � 15–16 per group).*P � .05, FaGHRKO (FFCx) vs. control (FFxx).


accompanied by significant changes in leptin action at thecellular level.

Unlike leptin, adiponectin levels have been shown todecrease as fat mass increases (39). In contrast, GHR�/�mice are obese yet have elevated levels of adiponectin.This observation in GHR�/� mice suggests that GH maynegatively regulate adiponectin. Cell culture studies havealso shown this to be the case because GH treatment ofdifferentiated 3T3-L1 adipocytes results in a decrease inadiponectin levels (40). Furthermore, studies on GH-transgenic and GH-deficient rodents suggest that GHsuppresses adiponectin secretion (12). Based on the aboveobservations, our second hypothesis was that disruptionof the GHR in adipose tissue will increase adiponectinlevels similar to what is seen in GHR�/� mice. Sur-prisingly, we found that FaGHRKO mice did not haveelevated levels of adiponectin, but rather had no changein females or decreased levels in males. Thus, our sec-

ond hypothesis did not hold true. This suggests that anynegative regulatory activity of GH on adipose tissue, asobserved in global GHR�/� mice, is dependent on theconsequences of disrupting GHR in other tissues orthat GHR is not important for regulating adiponectinsecretion.

We also measured resistin and adipsin, two additionaladipokines, in FaGHRKO mice. Resistin levels remainedunchanged and adipsin levels were significantly decreasedin both sexes. Recently, it has been shown that resistin isincreased in the GHR�/� mice (41). Thus, it appearsthat, like adiponectin, circulating resistin also is differen-tially regulated when GHR is selectively disrupted fromadipose tissue as opposed to global disruption. Adipsinlevels have not been assessed in GHR�/� mice. Becauseadipsin is thought to function primarily in the alternativepathway of the complement system, it is possible that theFaGHRKO mice have alterations to immune function,which was not assessed in this study. However, recentevidence suggests that the complement system in adiposetissue may play an important role in fat storage and insu-lin sensitivity (42). Thus, further investigation into theeffects of adipsin reduction in FaGHRKO and GHR�/�mice would also be of interest.

Various measures of glucose homeostasis did not differbetween FaGHRKO mice and controls including fastingglucose, fasting insulin, glucose tolerance, and insulin tol-erance. We expected adipose tissue to dispose of glucosemore efficiently than control mice because the diabeto-genic action of GH action was lacking in this tissue. How-ever, it appears that removal of GHR in adipose tissuedoes not produce a measurable effect on whole-bodyreadouts of glucose metabolism. Because the largest pro-portion of glucose disposal occurs in skeletal muscle (43),it is likely that adipose tissue’s contribution to whole-body glucose disposal is negligible. Moreover, most other

Figure 6. GHR gene disruption in adipose tissue does not alter RNAlevels encoding intracellular signaling molecules that are known toaffect glucose metabolism in WAT. Retroperitoneal WAT from femaleFaGHRKO mice was collected at 8 months of age. Relative mRNA levelsof PGC1�, SIRT1, PPAR�, PPAR�, PPAR�, 4-1BB, SREBP, FAS, andGLUT4, are shown for FaGHRKO (FFCx; black bars) and controls (FFxx;white bars). Values are presented relative to control values � SEM(n � 6–7 per group). FaGHRKO (FFCx) vs control (FFxx).

Table 2. IGF-1 and IGFBP Levels of FaGHRKO and Control Male and Female Mice


FFxx FFCx P Value FFxx FFCx P ValueGH and IGF-1

GH, n � 9–10 (ng/ml) 7.9 � 3.2 12.1 � 4.5 .466 7.6 � 1.6 7.9 � 2.1 .915IGF-1, n � 9–10 (ng/ml) 503 � 19 613 � 30a .006 655 � 31 705 � 32 .275

IGFBPsIGFBP-1, n � 9–10 (pg/ml) 7.4 � 2.0 11 � 1.7 .147 22 � 6.4 38 � 13 .285IGFBP-2, n � 9–10 (pg/ml) 160 � 8.5 172 � 8.0 .329 199 � 10 204 � 16 .789IGFBP-3, n � 9–10 (pg/ml) 189 � 19 229 � 14 .109 248 � 13 241 � 11 .711IGFBP-5, n � 9–10 (pg/ml) 5.8 � 0.3 7 � 0.5 .062 7 � 0.4 9 � 0.8a .033IGFBP-6, n � 9–10 (pg/ml) 133 � 12 142 � 11 .586 155 � 8.8 134 � 14 .228IGFBP-7, n � 9–10 (pg/ml) 13 � 1.0 13 � 1.2 .850 13 � 1.4 15 � 1.5 .428

Abbreviations: FFxx, controls; FFCx, FaGHRKO. Circulating peptide values of male and female mice at 6 months of age. Values are represented asmean � SEM (n � 9–10 per group). a Significance with P values given to the right.


adipose tissue-specific mouse models show similar lack ofeffect on whole-body glucose metabolism. A partial ex-ception can be seen with adipose-specific overexpressionof GLUT4 in isolated adipocytes ex vivo, where a 2- to3-fold increase in glucose disposal is reported; however,no difference in insulin-stimulated glucose disposal can bedetected in vivo (44). When adipose tissue is selectivelymade insulin resistant by fat-specific removal of insulinreceptor (seen in the FIRKO mouse), no changes in glu-cose or insulin tolerance occur at a young age (2 mo), butthese parameters do change in older mice (10 mo); thus, itis also possible that we may see changes at more advancedages (45). Alternatively, adiponectin is considered a po-tent insulin sensitizer, and high levels of this adipokinecould be important for the increased insulin sensitivity inGHR�/� mice (12). However, adiponectin levels are notincreased in FaGHRKO mice, which may partially ex-plain why no improvements were seen in glucose homeo-stasis. Furthermore, the difference in glucose metabolismmay also be partially affected by location of fat storage.As discussed earlier, GHR�/� mice have increased adi-posity primarily due to increased sc adipose tissue (8).FaGHRKO mice have increases in all WAT depots includ-ing mesenteric, which is thought to have a negative effecton glucose homeostasis (10). Analysis of genes involved in

glucose metabolism by quantitative real time RT-PCR inadipose tissue of FaGHRKO mice revealed similar resultsto that of whole-body analysis of glucose metabolism asno changes were seen. We hypothesized that theFaGHRKO mice would have improved glucose homeo-stasis; however, this was not the case.

Interestingly, FaGHRKO mice are quite different fromglobal GHR�/� mice. We expect that the differencesbetween the global GHR�/� and FaGHRKO mice withregard to nonadipose parameters (such as body size) aredue to the fact that GHR is disrupted in all tissues inGHR�/� mice whereas FaGHRKO mice have normallevels of GHR in nonadipose tissues. In contrast, differ-ences in adipose tissue parameters (such as adiponectinproduction and depot differences) are difficult to explainbecause the GHR is disrupted in adipose tissue in both mouselines;however,wespeculate that thesedifferencesareduetotheaction of GH in tissues other than adipose, and these othertissues, in turn, are able to influence adipose tissue physiologyvia endocrine/paracrine mechanisms.

Because this is the first description of the FaGHRKOmice, many additional studies will be performed. For ex-ample, analyzing gene expression and/or protein produc-tion in tissues such as adipose, liver, and muscle will beneeded to investigate the potential of tissue cross talk in

Table 3. Circulating Cytokines, Acute Phase Proteins, and Soluble Receptors of FaGHRKO and Control Male andFemale Mice


FFxx FFCx P Value FFxx FFCx P ValueCytokines

IL-6, n � 15–16 (pg/ml) 51 � 13 58 � 13 .675 30 � 4 56 � 7a .005MCP-1, n � 15–16 (pg/ml) 112 � 27 121 � 27 .819 402 � 221 135 � 21 .247

Acute phase 1Lipocalin-2, n � 9–10 (pg/ml) 59 � 4 106 � 22 .061 37 � 2 49 � 9 .218Pentraxin-3, n � 9–10 (pg/ml) 15 � 1 21 � 2a .008 18 � 2 18 � 1 .726

Acute phase 2AGP, n � 9–10 (pg/ml) 176 � 14 223 � 21 .083 160 � 9 214 � 57 .382�-2-Macroglobulin, n � 9–10 (ng/ml) 2490 � 117 2381 � 105 .499 2242 � 98 2139 � 201 .662CRP, n � 9–10 (ng/ml) 14 � 1 17 � 1 .058 15 � 1 16 � 2 .956Haptoglobin, n � 9–10 (ng/ml) 32 � 11 69 � 27 .232 12 � 2 47 � 37 .383

Soluble receptorsCD30, n � 9–10 (pg/ml) 65 � 19 129 � 69 .383 145 � 36 47 � 17a .029sgp130, n � 9–10 (pg/ml) 653 � 54 973 � 295 .326 944 � 362 3142 � 1777 .266sIL-1RI, n � 9–10 (pg/ml) 810 � 94 1084 � 205 .248 415 � 67 549 � 78 .214sIL-1RII, n � 9–10 (pg/ml) 4601 � 114 4445 � 87 .287 3581 � 370 3193 � 116 .310sIL-2Ra, n � 9–10 (pg/ml) 414 � 34 368 � 25 .289 394 � 59 345 � 22 .430sIL-4R, n � 9–10 (pg/ml) 1385 � 105 2305 � 287a .010 1734 � 138 1872 � 198 .585sIL-6R, n � 9–10 (pg/ml) 9010 � 301 8628 � 362 .434 10 430 � 956 9888 � 367 .608sTNFRI, n � 9–10 (pg/ml) 1457 � 85 1663 � 114 .174 1525 � 146 1515 � 87 .954sTNFRII, n � 9–10 (pg/ml) 3398 � 382 4478 � 533 .125 2838 � 396 3235 � 395 .489sVEGFR1, n � 9–10 (pg/ml) 2739 � 115 2826 � 202 .724 2187 � 438 1940 � 110 .574sVEGFR2, n � 9–10 (ng/ml) 29 � 1 28 � 1 .635 30 � 1 29 � 2 .768sVEGFR3, n � 9–10 (ng/ml) 32 � 1 31 � 1 .542 32 � 1 32 � 1 .566

Abbreviations: FFxx, controls; FFCx, FaGHRKO. Circulating peptide values of male and female mice at 6 mo of age. Values are represented asmean � SEM (n � 9–16 per group). a Significance with P values given to the right.


FaGHRKO vs GHR�/� mice. It also would be of interestto determine whether local IGF-1 expression is altered invarious tissues of these mice because local IGF-1 may bedecreased in tissues where GHR is absent. Additionally,we would like to quantify expression of various lipogenic/lypolytic enzymes (such as lipoprotein lipase, adrenergicreceptors, lipid droplet proteins, and intracellular lipases)in adipose tissue depots of these mice. Because lipoproteinlipase has been shown previously to respond to GH dif-ferently in different adipose tissue depots (33), such stud-ies may help explain the differences observed in the cur-rent study. Finally, we now have the capability to crossvarious tissue-specific GHRKO lines (such as muscle- andliver-specific GHRKO mice) with the FaGHRKO line inorder to determine which tissue(s) require GHR disrup-tion in concert with adipose to achieve a similar pheno-type as global GHR�/� mice.

In conclusion, FaGHRKO mice share few characteris-tics with global GHR�/� mice. FaGHRKO mice areobese with increased total body fat, increased adipocytecell size, and increased circulating leptin. However, unlikeglobal GHR�/� mice, these mice show no improvementsin measures of glucose homeostasis, have normal levels ofresistin, and normal/decreased levels of adiponectin.Thus, it appears that the increases in adipokines seen inglobal GHR�/� mice are probably due to the removal ofGH’s action in all tissues and not a result of deletion of theGHR in adipose tissue alone.

Acknowledgments

Address all correspondence and requests for reprints to:John, J. Kopchick, PhD, 101 Konneker Research Laborato-ries, Ohio University, Athens, Ohio 45701. E-mail:[email protected].

This work was supported by the State of Ohio’s EminentScholar Program that includes a gift from Milton and LawrenceGoll; by National Institutes of Health (NIH) GrantsP01AG031736, AG032290, DK58259, and DK083729; by theAMVETS; by the Diabetes Institute at Ohio University; and byPolish Ministry of Science and Higher Education GrantNN401042638. The floxed GHR mouse strain used for thisresearch project was generated by the trans-NIH KOMP andobtained from the KOMP Repository (www.komp.org). NIHgrants to Velocigene at Regeneron, Inc (U01HG004085) andthe CHORI-Sanger-UCDavis Consortium (U01HG004080)funded the generation of gene-targeted embryonic stem cells for8500 genes in the KOMP Program and archived and distributedby the KOMP Repository at University of California Davis andChildren’s Hospital Oakland Research Institute (CHORI)(U42RR024244).

Disclosure Summary: The authors have nothing to disclose.

References

1. List EO, Sackmann-Sala L, Berryman DE, et al. Endocrine param-eters and phenotypes of the growth hormone receptor gene dis-rupted (GHR�/�) mouse. Endocr Rev. 2011;32:356–386.

2. Zhou Y, Xu BC, Maheshwari HG, et al. A mammalian model forLaron syndrome produced by targeted disruption of the mousegrowth hormone receptor/binding protein gene (the Laron mouse).Proc Natl Acad Sci USA. 1997;94:13215–13220.

3. Dominici FP, Arostegui Diaz G, Bartke A, Kopchick JJ, Turyn D.Compensatory alterations of insulin signal transduction in liver ofgrowth hormone receptor knockout mice. J Endocrinol. 2000;166:579–590.

4. Coschigano KT, Holland AN, Riders ME, List EO, Flyvbjerg A,Kopchick JJ. Deletion, but not antagonism, of the mouse growthhormone receptor results in severely decreased body weights, insu-lin and IGF-1 levels and increased lifespan. Endocrinology. 2003;144:3799–3810.

5. Bonkowski MS, Dominici FP, Arum O, et al. Disruption of growthhormone receptor prevents calorie restriction from improving insu-lin action and longevity. PLoS ONE. 2009;4:e4567.

6. Ikeno Y, Hubbard GB, Lee S, et al. Reduced incidence and delayedoccurrence of fatal neoplastic diseases in growth hormone receptor/binding protein knockout mice. J Gerontol A Biol Sci Med Sci.2009;64:522–529.

7. Guevara-Aguirre J, Balasubramanian P, Guevara-Aguirre M, et al.Growth hormone receptor deficiency is associated with a majorreduction in pro-aging signaling, cancer, and diabetes in humans.Sci Transl Med. 2011;3:70ra13.

8. Berryman DE, List EO, Palmer AJ, et al. Two-year body composi-tion analyses of long-lived GHR null mice. J Gerontol A Biol SciMed Sci. 2010;65:31–40.

9. Egecioglu E, Bjursell M, Ljungberg A, et al. Growth hormone re-ceptor deficiency results in blunted ghrelin feeding response, obe-sity, and hypolipidemia in mice. Am J Physiol Endocrinol Metab.2006;290:E317–E325.

10. Berryman DE, List EO, Sackmann-Sala L, Lubbers E, Munn R,Kopchick JJ. Growth hormone and adipose tissue: beyond the adi-pocyte. Growth Horm IGF Res. 2011;21:113–123.

11. Kloting N, Fasshauer M, Dietrich A, et al. Insulin-sensitive obesity.Am J Physiol Endocrinol Metab. 299:E506–E515.

12. Berryman DE, List EO, Coschigano KT, Behar K, Kim JK, Kop-chick JJ. Comparing adiposity profiles in three mouse models withaltered GH signaling. Growth Horm IGF Res. 2004;14:309–318.

13. Bik W, Baranowska B. Adiponectin—a predictor of higher mortal-ity in cardiovascular disease or a factor contributing to longer life?Neuro Endocrinol Lett. 2009;30:180–184.

14. Ibanez L, Lopez-Bermejo A, Diaz M, Jaramillo A, Marin S, deZegher F. Growth hormone therapy in short children born small forgestational age: effects on abdominal fat partitioning and circulat-ing follistatin and high-molecular-weight adiponectin. J Clin Endo-crinol Metab. 2010;95:2234–2239.

15. Nilsson L, Binart N, Bohlooly YM, et al. Prolactin and growthhormone regulate adiponectin secretion and receptor expression inadipose tissue. Biochem Biophys Res Commun. 2005;331:1120–1126.

16. Laron Z, Ginsberg S, Lilos P, Arbiv M, Vaisman N. Body compo-sition in untreated adult patients with Laron syndrome (primaryGH insensitivity). Clin Endocrinol (Oxf). 2006;65:114–117.

17. Fan Y, Menon RK, Cohen P, et al. Liver-specific deletion of thegrowth hormone receptor reveals essential role of GH signaling inhepatic lipid metabolism. J Biol Chem. 2009;284(30):19937–19944.

18. Mavalli MD, DiGirolamo DJ, Fan Y, et al. Distinct growth hor-mone receptor signaling modes regulate skeletal muscle develop-ment and insulin sensitivity in mice. J Clin Invest. 2010;120:4007–4020.


www.komp.org

19. Vijayakumar A, Wu Y, Sun H, et al. Targeted loss of GHR signalingin mouse skeletal muscle protects against high-fat diet-induced met-abolic deterioration. Diabetes. 2012;61:94–103.

20. Wu Y, Liu C, Sun H, et al. Growth hormone receptor regulates �

cell hyperplasia and glucose-stimulated insulin secretion in obesemice. J Clin Invest. 2011;121:2422–2426.

21. Skarnes WC, Rosen B, West AP, et al. A conditional knockoutresource for the genome-wide study of mouse gene function. Na-ture. 2011;474:337–342.

22. Masternak MM, Al-Regaiey KA, Del Rosario Lim MM, et al. Ca-loric restriction results in decreased expression of peroxisome pro-liferator-activated receptor superfamily in muscle of normal andlong-lived growth hormone receptor/binding protein knockoutmice. J Gerontol A Biol Sci Med Sci. 2005;60:1238–1245.

23. Zhang Y, Guan R, Jiang J, et al. Growth hormone (GH)-induceddimerization inhibits phorbol ester-stimulated GH receptor prote-olysis. J Biol Chem. 2001;276:24565–24573.

24. List EO, Palmer AJ, Berryman DE, Bower B, Kelder B, Kopchick JJ.Growth hormone improves body composition, fasting blood glu-cose, glucose tolerance and liver triacylglycerol in a mouse model ofdiet-induced obesity and type 2 diabetes. Diabetologia. 2009;52:1647–1655.

25. Salmon DM, Flatt JP. Effect of dietary fat content on the incidenceof obesity among ad libitum fed mice. Int J Obes. 1985;9:443–449.

26. Tchoukalova YD, Koutsari C, Votruba SB, et al. Sex- and depot-dependent differences in adipogenesis in normal-weight humans.Obesity (Silver Spring). 2010;18:1875–1880.

27. Wueest S, Rapold RA, Schumann DM, et al. Deletion of Fas inadipocytes relieves adipose tissue inflammation and hepatic mani-festations of obesity in mice. J Clin Invest. 2010;120:191–202.

28. Kumar A, Lawrence JC Jr, Jung DY, et al. Fat cell-specific ablationof rictor in mice impairs insulin-regulated fat cell and whole-bodyglucose and lipid metabolism. Diabetes. 2010;59:1397–1406.

29. He W, Barak Y, Hevener A, et al. Adipose-specific peroxisomeproliferator-activated receptor � knockout causes insulin resistancein fat and liver but not in muscle. Proc Natl Acad Sci USA. 2003;100:15712–15717.

30. Abel ED, Peroni O, Kim JK, et al. Adipose-selective targeting of theGLUT4 gene impairs insulin action in muscle and liver. Nature.2001;409:729–733.

31. Urs S, Harrington A, Liaw L, Small D. Selective expression of anaP2/fatty acid binding protein 4-Cre transgene in non-adipogenictissues during embryonic development. Transgenic Res. 2006;15:647–653.

32. Makowski L, Boord JB, Maeda K, et al. Lack of macrophage fatty-acid-binding protein aP2 protects mice deficient in apolipoprotein Eagainst atherosclerosis. Nat Med. 2001;7:699–705.

33. Richelsen B, Pedersen SB, Borglum JD, Moller-Pedersen T, Jor-gensen J, Jorgensen JO. Growth hormone treatment of obesewomen for 5 wk: effect on body composition and adipose tissueLPL activity. Am J Physiol. 1994;266:E211–E216.

34. Etherton TD. Porcine growth hormone: a central metabolic hor-mone involved in the regulation of adipose tissue growth. Nutri-tion. 2001;17:789–792.

35. Liu JL, Coschigano KT, Robertson K, et al. Disruption of growthhormone receptor gene causes diminished pancreatic islet size andincreased insulin sensitivity in mice. Am J Physiol EndocrinolMetab. 2004;287:E405–E413.

36. Moraes-Vieira PM, Bassi EJ, Araujo RC, Camara NO. Leptin as alink between the immune system and kidney-related diseases: lead-ing actor or just a coadjuvant? Obes Rev. 2012;13:733–743.

37. Mantzoros CS, Magkos F, Brinkoetter M, et al. Leptin in humanphysiology and pathophysiology. Am J Physiol Endocrinol Metab.2011;301:E567–E584.

38. Moon HS, Matarese G, Brennan AM, et al. Efficacy of metreleptinin obese patients with type 2 diabetes: cellular and molecular path-ways underlying leptin tolerance. Diabetes. 2011;60:1647–1656.

39. Arita Y, Kihara S, Ouchi N, et al. Paradoxical decrease of an adi-pose-specific protein, adiponectin, in obesity. Biochem Biophys ResCommun. 1999;257:79–83.

40. Wolfing B, Neumeier M, Buechler C, Aslanidis C, Scholmerich J,Schaffler A. Interfering effects of insulin, growth hormone and glu-cose on adipokine secretion. Exp Clin Endocrinol Diabetes. 2008;116:47–52.

41. Vijeyta F. Effects of Growth Hormone on Circulating Resistin Lev-els in Mice. [masters’ thesis] Ohio University, Athens, Ohio: 2012;1–141.

42. Schaffler A, Scholmerich J. Innate immunity and adipose tissuebiology. Trends Immunol. 2010;31:228–235.

43. DeFronzo RA. Pathogenesis of type 2 diabetes: metabolic and mo-lecular implications for identifying diabetes genes. Diabetes Rev.1997;5:177–269.

44. Carvalho E, Kotani K, Peroni OD, Kahn BB. Adipose-specific over-expression of GLUT4 reverses insulin resistance and diabetes inmice lacking GLUT4 selectively in muscle. Am J Physiol EndocrinolMetab. 2005;289:E551–E561.

45. Bluher M, Michael MD, Peroni OD, et al. Adipose tissue selectiveinsulin receptor knockout protects against obesity and obesity-re-lated glucose intolerance. Dev Cell. 2002;3:25–38.


2013 List_The role of GH in adipose tissue-LiGHRKO mice

Documents

Transcript of 2013 List_The role of GH in adipose tissue-LiGHRKO mice