Muscle lipid storage, insulin sensitivity and oxidative capacity; a

microscopic approach

Sabine Daemen17-02-17Diabetes and Metabolism Research Groupdmrg.nl

Athletes Paradox

Pan et al. 1997 Diabetes

Goodpaster et al. 2004 Obes Res

Lipid droplets (LDs)

Morphology

Cellulardistribution

Coating proteins; PLIN5

Contact withmitochondria

Composition

Role for lipid droplets and their coat proteins in insulin sensitivity in skeletal muscle?

Aim

1. Morphology

2. Cellulardistribution

3. Coating proteins; PLIN5

4. Contact withmitochondria

5. Composition

Type 2 diabetes patients (T2D)

Endurance-trainedathletes

Methods morphology & localization

• Muscle biopsies: 7 μm thick sections• Staining:

- Bodipy for lipid droplets (green)- Laminin for cell membrane (blue)- MHC1 for fibertyping

• Confocal microscopy (Leica TCS SPE)• Deconvolution• Image Analysis (ImageJ & Matlab®)

1. Morphology & 2. Localization

Intramyocellular lipid (IMCL) storage is fiber type specific

All Type I Type II0.0

0.5

1.0

1.5

2.0

*

#

Athletes

T2D

IMC

L c

on

ten

t (%

lip

id a

rea f

racti

on

)

1. Morphology

Type I Type II0.0

0.1

0.2

0.3

0.4

0.5

* # Athletes

T2D

LD

siz

e (

m2)

Type I Type II0.00

0.02

0.04

0.06

0.08

Athletes

T2D

*

# #

LD

nu

mb

er

(# /

m2)

Increased subsarcolemmal lipid storage in T2D

2. Localization

SS IMF SS IMF0.0

0.5

1.0

1.5

2.0

2.5

Type I Type II

*

*

T2D

Athletes

IMC

L c

on

ten

t (%

lip

id a

rea f

racti

on

)

Type I Type II0

5

10

15

20

25

T2D

Athletes

*

*

Fra

cti

on

SS

LD

s (

%)

Subsarcolemmal (SS) lipid dropletsIntermyofibrillar (IMF) lipid droplets

Gemmink 2015 Diapedia

PLIN5 coating

3. PLIN5 coating

PLIN5 LDs Cel membrane

Insulin sensitivityIMCL

Bosma et al. 2013 Biochim Biophys Acta

PLIN5 coating does nog associate with improved insulin sensitivity

0.0 0.5 1.00

50

100

150 AthletesLeanObeseT2D

PLIN5+ Lipid Area Fraction (%)

GIR

(

mo

l/kg

le

an

ma

ss/m

in)

Ath

lete

s

Lean

Obes

eT2D

0.0

0.5

1.0

1.5

2.0

#*

* *

PL

IN5 p

rote

in (

AU

)

3. PLIN5 coating

Differential LD-mitochondrial contact in type 2 fibers

All Type I Type II0.0

0.1

0.2

0.3

*

# Athletes

T2D

Mit

o a

rea

fra

cti

on

(%

)

4. Contact with mitochondria

LD Mitochondria

LD composition by CARS microscopy

5. Composition

In collaboration with Dr. Sapun Parekh, Max Planck Institute for Polymer Research, Mainz

Billecke et al. 2015 Integr Biol

Working at the Max Planck Institute Mainz

• Technical Institute

• Partly home-build CARS microscope

5. Composition

Only subtle differences in CARS spectra

*

*

*** ***

***

5. Composition

Related to saturation and chain length

Differences especially in type 2 fibers

*

*

***

*** ***

5. Composition

Conclusions

• Differential lipid storage pattern between Athletes and T2D- Fiber type specific- Higher number of LDs in athletes <> larger LDs in T2D- More lipid storage in SS area in T2D

• PLIN5 coating of LDs does not directly explain the athletes paradox

• LD-mitochondrial contact differs in type 2 fibers between Athletes and T2D

• Only minor differences in lipid composition of the LD between Athletes and T2D

Acknowledgements

Department of Human Biology and Movement Sciences: • Anne Gemmink• Bram Brouwers• Gert Schaart• Joris Hoeks• Patrick Schrauwen• Matthijs K.C. Hesselink

Department of Molecular Spectroscopy, Max Planck Institute for Polymer Research Mainz

• Nils Billecke• Sapun Parekh

Graduate Programme

Diabetes and Metabolism Research Group: www.dmrg.nlfacebook.com/diabetesmaastricht