Inflammation in the Hypothalamus and Its Relevance to Obesity

Research in Obesity and Comorbidities

Licio A. Velloso University of Campinas Brazil www.ocrc.org.br

• 13 Principal investigators • 25 Associate investigators • 45 Postdoctoral fellows • 89 Graduate students www.ocrc.org.br

• Obesity • Diabetes • Metabolism • Endocrine physiology • Atherosclerosis • Hypertension • Nutrition • Physical activity www.ocrc.org.br

• USA • Spain – Harvard, Boston – University of Barcelona – Washington University, Seattle – Miguel Hernandes U, Elche – San Antonio U, Texas • Canada – Michigan U, Ann Arbor – Toronto University – U Nebraska, Lincoln • UK – Cambridge • Belgium – ULB, Brussels • Italy – Milan University www.ocrc.org.br

Development of biological compounds to accelerate wound healing in metabolic disease Diabetes wounds General objective

Define the mechanisms behind the development of obesity in order to develop more efficient preventive and therapeutic methods to tackle obesity and all the commonly associated disease

Brain

G Gut 3.0 x 1013

Sender PLoS Biol 2016 e1002533 3

3.0 x 1013 3.8 x 1013

Sender PLoS Biol 2016 e1002533 Karlsson Dabetes 2013 62:3341 Fraher Nat Rev Gastroenterol Hepatol 2012 9:312 Obesity

Manichanh Gut 2006 55:205 Turnbaugh Nature 2006 444:1027

Colon cancer

Zackular Mbio 2013 4:587 Dumas PNAS 2006 103:12511 Mafra Clin Kidney J 2015 8:332 Main features of the obese gut microbiome

1. Reduction of variability 2. Reduction of Firmicutes 3. Increase Proteobacteria

Turnbaugh Nature 2006 444:1027

Actually eats Improves caloric more calories harvesting in gut Improves caloric harvesting in the gut as compared with the microbiota of lean

Turnbaugh Nature 2006 444:1027 ?

Hypothalamic resistance to leptin and insulin - Obesity

Palmitate16:0 Citokines •TNFa Stearate18:0

Microglia •IL1b •IL6

Arachidate 20:0 TLR4,

What comes first in obesity

• Gut dysbiosis • Hypothalamic dysfunction Swiss outbred mice

HFD intake

25% lower 25% higher food intake food intake OR OP mice mice

Razolli et al Unpublished, 2017 P h y lu m Phylum 250000 Ac tin o b a c te r ia 200000 OR Chow 1 5 0 0 0 0 150000 OP Chow B a c te r o id e te s 100000 50000 C ya n o b a c te r ia

1 0 0 0 0 0 3000 F ir m ic u te s

Counts 2000 P r o te o b a c te r ia 1000 S p ir o c h a e te s 5 0 0 0 0 0 T e n e r ic u te s

U n c la s s ifie d Firmicutes Tenericutes Spirochaetes Unclassified O R C h o w O P C h o w ActinobacteriaBacteroidetesCyanobacteriaProteobacteria

P h y lu m Phylum 250000 Ac tin o b a c te r ia 200000 OR HFD B a c te r o id e te s 150000 OP HFD 1 0 0 0 0 0 100000 C ya n o b a c te r ia 50000 D e fe r r ib a c te r e s 1500

F ir m ic u te s Counts 1000 P r o te o b a c te r ia 5 0 0 0 0 500 S p ir o c h a e te s

T e n e r ic u te s 0

T h e r m o to g a e

U n c la s s ifie d l Firmicutes Tenericutes Bacteroidetes SpirochaetesThermotogaeUnclassified l O R H F D O P H F D ActinobacteriaCyanobacteriaDeferribacteresProteobacteria

Razolli et al Unpublished, 2017 C la s s Class

1 5 0 0 0 0 200000 B a c illi OR Chow 150000 OP Chow B a c te r o id ia 100000

C lo s tr id ia 50000 1 0 0 0 0 0 D e lta p r o te o b a c te r ia 4000

Counts 3000 E r ys ip e lo tr ic h i 2000 G a m m a p r o te o b a c te r ia 1000 5 0 0 0 0 0 M o llic u te s

S p h in g o b a c te r iia Bacilli Clostridia Mollicutes Bacteroidia U n c la s s ifie d Unclassified Erysipelotrichi Sphingobacteriia O R C h o w O P C h o w Deltaproteobacteria Gammaproteobacteria

C la s s Class 200000 OR HFD B a c illi 150000 1 0 0 0 0 0 OP HFD B a c te r o id ia 100000 50000 C lo s tr id ia 2000 D e lta p r o te o b a c te r ia Counts 1500 E r ys ip e lo tr ic h i 1000 5 0 0 0 0 500 G a m m a p r o te o b a c te r ia 0 M o llic u te s

Bacilli S p h in g o b a c te r iia Clostridia Mollicutes Bacteroidia Unclassified U n c la s s ifie d Erysipelotrichi Sphingobacteriia Deltaproteobacteria O R H F D O P H F D GammaproteobacteriaRazolli et al Unpublished, 2017 O r d e r Order 200000 1 5 0 0 0 0 OR Chow Ac h o le p la s m a ta le s 150000 OP Chow B a c te r o id a le s 100000

C lo s tr id ia le s 50000

D e s u lfo v ib r io n a le s 1 0 0 0 0 0 3000

Counts E r ys ip e lo tr ic h a le s 2000 L a c to b a c illa le s 1000 L e g io n e lla le s 5 0 0 0 0 0 N a tr a n a e r o b ia le s

S p h in g o b a c te r ia le s

Clostridiales U n c la s s ifie d Bacteroidales Legionellales Unclassified Lactobacillales Erysipelotrichales Natranaerobiales O R C h o w O P C h o w AcholeplasmatalesDesulfovibrionales Sphingobacteriales

O r d e r Order 200000 B a c te r o id a le s OR HFD 1 0 0 0 0 0 150000 OP HFD 100000 C lo s tr id ia le s 50000 D e s u lfo v ib r io n a le s 3000

E r ys ip e lo tr ic h a le s Counts 2000 5 0 0 0 0 L a c to b a c illa le s 1000

N a tr a n a e r o b ia le s 0

S p h in g o b a c te r ia le s

U n c la s s ifie d Clostridiales Bacteroidales Unclassified Lactobacillales ErysipelotrichalesNatranaerobiales O R H F D O P H F D Desulfovibrionales Sphingobacteriales Razolli et al Unpublished, 2017 Razolli et al Unpublished, 2017 POMC Processing

POMC

PC1/3

N-POMC/ACTH β-Lipotropin

PC1/3 PC2

Pro-γ-MSH ACTH γ-Lipotropin β-Endorphin

PC2 PC2 PC2/CPE

γ-MSH des-α-MSH β-MSH β-Endorphin

N-AT/PRCP

γ-Endorphin α-Endorphin α-MSH

Razolli et al Unpublished, 2017 Results – POMC Enzymes - WB

The infusion of palmitate led to increased hypothalamic α-MSH in OR as compared to CT. Enzymes involved on POMC processing were modulated between OP and OR mice after infusion of palmitic acid using osmotic pump.

ANOVA test; * p<0,05; n=5 Razolli et al Unpublished, 2017 Results – PC1/3 hypothalamic inhibition - Lentivirus OR mice transfected with PC1/3 lentivirus present body mass gain similar to OP mice transfected with scramble, followed by increased food intake.

Lentiviral clone selection: Five sequences from Sigma Aldrich - PCSK1 (NM_013628) ANOVA test; * p<0,05; n=5 Razolli et al Unpublished, 2017 Increase caloric intake Dysfunction of POMC neurons

Increase caloric harvesting Hypothalamus is the preferential target for treatment of obesity Thank you!

Grants Sao Paulo Research Foundation CNPq CAPES