1. Volume 22, Issue 6, Pages 1522-1530 (February 2018)
Supraphysiologic Administration of GDF11 Induces Cachexia in Part by Upregulating GDF15 
Juli E. Jones, Samuel M. Cadena, Chenguang Gong, Xiaomei Wang, Zhiping Chen, Sharon X. Wang, Chad Vickers, Hong Chen, Estelle Lach-Trifilieff, John R. Hadcock, David J. Glass 
Cell Reports 
Volume 22, Issue 6, Pages (February 2018)
DOI: /j.celrep
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2. Cell Reports 2018 22, 1522-1530DOI: (10.1016/j.celrep.2018.01.044)
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3. Figure 1 GDF11 Induces a Reduction in Body Weight in Obese Mice
Obese mice fed ad libitum were administered empty vector (n = 9) or 3 μg (n = 10) or 10 μg (n = 9) of GDF11 DNA via HDI. Animals administered GDF11 DNA showed a dose-dependent decrease in body weight compared to empty vector controls. Food intake was suppressed to a similar degree as weight loss in each group (data not shown). Data are expressed as mean ± SEM of percent change from study day 0. ∗p < 0.05 versus vector by two-way ANOVA with Tukey’s post hoc test for multiple comparisons.
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4. Figure 2
GDF11 Overexpression Reduces Body Weight to a Greater Degree than Reduced Food Intake Alone
GDF11 was overexpressed following HDI DNA injection in diet-induced obesity (DIO) mice with access to high-fat diet (control vector, n = 9; GDF11-3 μg, n = 8, GDF11-pair-fed, n = 9, GDF11-10 μg, n = 8, GDF11-pair-fed, n = 9). Pair-feeding began 2 days following the HDI injection, and fresh diet was given on day 8, which consistently increases food intake for the subsequent 24 hr, but not for the GDF11-10 μg animals.
(A and B) Food intake (A) and body weight (B) significantly decreased in GDF11-treated mice from day 4 onward compared to control vector mice. Although animals received the same amount of food, animals administered GDF11 lost more weight than their pair-fed controls.
(C and D) End-of-study body composition in mass (C) or percent composition (D) indicates which compartment is contributing to the weight loss.
All data shown are means ± SEM. ∗p < 0.05 compared to control vector and #p < 0.05 compared to respective GDF11 DNA-treated mice. Statistical analysis was performed by two-way ANOVA followed by Bonferroni’s post hoc test (A and B) and Fisher’s least significant difference (LSD) post hoc test (C and D).
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5. Figure 3 GDF11 Induced Changes in GDF15
(A) Animals that received 10 μg of GDF11 (n = 8) or that are fed the same amount of food (n = 9) across the 11-day study had a significant increase in GDF15 gene expression in the gastrocnemius. Data are expressed as mean (relative expression to GAPDH) ∗ 100 ± SEM.
(B and C) Unlike muscle expression, plasma GDF15 levels only increased in animals with injection of GDF11 DNA (B), and the greater circulating GDF15 levels were associated with more weight loss (C). Data are expressed as mean plasma concentration (in picograms per milliliter) ± SEM.
∗p < 0.05 indicates significant difference from control vector by one-way ANOVA and Fisher’s LSD post hoc test.
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6. Figure 4
ActRIIA and ActRIIB Antagonism Blocks GDF11-Induced Body Weight Loss
(A) DIO mice injected, by HDI, with 3 μg of GDF11 DNA (n = 38) lost a significant amount of weight compared to control vector-injected mice (n = 36) five days following HDI injection. Five days post-HDI, animals (n = 9–10/group) were administered control antibody, anti-ActRIIA, anti-ActRIIB, or a combination of both antibodies. Only animals that received both antibodies showed a significant inhibition of body weight loss compared to animals receiving control antibody.
(B and C) Administration of a combination of anti-ActRIIA and anti-ActRIIB antibodies showed no change in fat mass (B), but led to a significant increase in lean body mass (C).
Data are expressed as mean ± SEM. ∗p < 0.05 versus control antibody by one-way ANOVA and Fisher’s LSD post hoc test.
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7. Figure 5
Suppression in Appetite Induced by GDF11 Is Blocked with an Anti-GDF15 Antibody
Animals injected with 3 μg of GDF11 (n = 20) lost significantly more weight than animals administered the vector control (n = 16) via HDI.
(A and B) Three days following administration of anti-GDF15 on day 11 post-HDI, animals in the GDF11 group (n = 10) began to consume more food (A), which elicited a regain of body weight (B).
(C and D) The weight that was gained in GDF11-treated animals was from an increase of fat mass (C) not lean mass (D).
Data are expressed as mean ± SEM. ∗p < 0.05 indicates significant difference from the vehicle Ab control group by one-way ANOVA and Fisher’s LSD post hoc test.
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8. Figure 6 GDF11 Induces SMAD2/3 to Bind to the GDF15 Promoter
(A) Spreading of SMAD2/3 binding motifs on DNA region upstream of GDF15 gene. Five pairs of primers were designed to flank each motif or motifs cluster on the GDF15 promoter; as a control, a previously determined SMAD2/3 binding motif on the ID1 promoter was used.
(B) Real-time PCR analysis of anti-SMAD2/3 ChIP on mouse primary myotubes treated with 300 ng/mL GDF11 for 24 hr.
Data are the result of three independent experiments and expressed as mean ± SEM. ∗p < 0.05 versus control by two-way ANOVA with Sidak’s post hoc test.
Cell Reports  , DOI: ( /j.celrep )
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