transport and metabolism Dr Rohan Lewis - - PowerPoint PPT Presentation

Placental amino acid transport and metabolism

Dr Rohan Lewis rohan.lewis@southampton.ac.uk

Note: for non-commercial purposes only

Why do we need to understand placental amino acid transfer?

• Amino acid transfer….
• Is required for fetal growth
• Is reduced in fetal growth restriction
• Is reduced prior to fetal growth restriction
• Interventions could optimise amino acid

transfer and fetal growth

• But interventions are more likely to succeed if we

understand the mechanisms

There are multiple determinants of placental function

Transporters MVM BM Blood flow Maternal Fetal Structure Diffusion distance Surface area Metabolism Consumption Inter-conversion

Which of these are most likely to be rate limiting for fetal growth?

Placental blood flow

Amino acids must be delivered before they can be transported

Blood flow and placental transfer

• Oxygen transfer is flow limited
• Rapid diffusion down concentration gradients
• Maternal and fetal blood flow maintain the gradients
• Is amino acid transfer flow limited?
• Placental uptake occurs against the gradient so is

unlikely to be flow dependent

• But placental transfer to the fetus occurs down

gradients so could be flow dependent

• 1. Maternal artery:
• Perfuse 14C-phenylalanine
• Control flow rate 10, 14, 18 ml/min
• 2. Fetal artery:
• Control flow rate 3, 6, 9 ml/min
• 3. Fetal vein:
• Measure 14C-phenylalanine

transfer

Flow effects in the isolated perfused human placenta

Study design: flow rates the perfused human placenta

2 4 6 8 10 12 14 16 18 20 50 100 150 200 250 300 Flow rate ml/min Time from start of tracer perfusion maternal flow fetal flow

Time (minutes) Flow rate (ml/min) Maternal flow rate 10, 14, 18 ml/min Fetal flow rate 3, 6, 9 ml/min

Increasing maternal flow increases amino acid uptake

5 10 15 20 25 30 35 10M3F 10M6F 10M9F 14M3F 14M6F 14M9F 18M3F 18M6F 18M9F Plaxental uptake (umol/min)

Data is mean and SEM 2 way ANOVA Maternal flow P = 0.011, Fetal flow P = 0.41, Interaction P = 0.96 Flow ml/min Maternal 10 10 10 14 14 14 18 18 18 Fetal 3 6 9 3 6 9 3 6 9 Placental uptake (pmol/min)

Increasing fetal flow decreases fetal vein amino acid concentration

0,3 0,6 0,9 1,2 1,5 1,8 Fetal vein Phe concentration (umol/ml)

Data is mean and SEM 2 way ANOVA Maternal flow P = 0.54, Fetal flow P < 0.001,Interaction P = 0.22 Flow ml/min Maternal 10 10 10 14 14 14 18 18 18 Fetal 3 6 9 3 6 9 3 6 9 Fetal vein concentration (pmol/l)

But neither maternal or fetal flow affect placental transfer

1 2 3 4 5 Placental Phe transfer (umol/min)

Data is mean and SEM 2 way ANOVA Maternal flow P = 0.71, Fetal flow P = 0.79,Interaction P = 0.39 Flow ml/min Maternal 10 10 10 14 14 14 18 18 18 Fetal 3 6 9 3 6 9 3 6 9 Placental transfer (pmol/min)

Phenylalanine transfer is not flow limited

• Placental uptake
• Was associated with maternal but not fetal flow
• Placental transfer was independent of

flow

• Higher uptake did not mean more transfer!
• So while flow is essential for amino acid

transfer it is not rate limiting

• Even at half physiological flow rates

Membrane transport

20 amino acids 20+ transport proteins

Amino acid uptake

Maternal blood Placental syncytiotrophoblast Fetal blood

Amino acids = aa aa1 aa2 aa1 aa3 aa2 aa2 aa1 aa1

BM MVM

aa3 aa3 aa2 aa3 aa1 aa1

Accumulative Exchanger

Amino acid efflux

Cleal et al (2011) J. Physiol 589:987-989 Amino acids = aa aa1 aa1 aa2

BM MVM

aa3 aa2

Accumulative Exchanger Facilitated

aa3

Exchanger

aa3 aa3 aa3

Maternal blood Placental syncytiotrophoblast Fetal blood

aa3 aa3 aa3

Amino acid transfer requires complex interactions

• Amino acid transfer requires interactions
• Between different classes of transporter
• Between the MVM and the BM
• Its more complex than shown
• 20 amino acids and more than 20 transporters
• Can we predict the effect of changing the

level of one transporter or substrate?

• Computational modelling…

Does metabolism facilitate amino acid transport?

Amino acid consumption Amino acid inter-conversion

Placental amino acid metabolism

AA2 AA1 AA1 CONSUMPTION Reduces the size of amino acid pool available for transfer to fetus INTERCONVERSION Alters the composition of the amino acid pool available for transfer to the fetus AA1 AA2 Mother Placenta Fetus AA = amino acid

Human placental glutamate metabolism

Day et al 2013 Placenta 34, 1223-1231

Glutamate glutamine proline aspartate α-ketoglutarate Intermediary metabolites glutamine proline aspartate leucine alanine Nitrogen Carbon NH4

+

ATP isoleucine valine

Placental amino acid metabolism

Day et al 2013 Placenta 34, 1223-1231

glutamine glutamate glutamate CONSUMPTION Intermediary metabolites and energy INTERCONVERSION glutamate glutamine Mother Placenta Fetus

glutamine

≈45% ≈45% ≈ 10% ≈ 90%

4. Maternal blood Placental syncytiotrophoblast Fetal blood

Accumulative transporter

5 mmol/l glutamate glutamine

Exchanger Exchanger Facilitated BM MVM

Glutamine synthesis may drive uptake of maternal amino acids by exchange

Lower abundance essential amino acids Cycling of abundant amino acid Delivery of lower abundance essential amino acids to the fetus 1. 2. 3. 4.

Metabolism summary

• Glutamine synthesis may occur in order to

drive amino acid uptake by exchange

• Makes the system less dependent on accumulative

transporters such as system A

• The question we need to model is whether

metabolism is a rate limiting factor

Placental structure

Is this important for amino acid transfer?

Placental structure

• We don’t know how placental structure

may relate to amino acid transfer

• Factors like surface area and diffusion

distance will be important

• But not as important as they are for oxygen
• Modern imaging techniques may help us

to investigate these questions

microCT can image large scale vascular structure

Key Red: GLUT1 Green: Capillaries

Light sheet microscopy can image up to 1 cm3

(this image 0.2 cm3)

Lat2

Transporters may be localised to specific regions on the villi

Key Blue: LAT2 Green: MVM

Computational Modelling

An integrated approach to placental transfer

Placental function determines fetal development

• We often assume that changes in placental

characteristics correlate with function

• e.g. more transporter = more transfer & vice versa
• But placental function is complex and this

assumption may not be justified

• Computational modelling allows us to study

complex interactions and make predictions

The simple model

3 amino acids, 3 compartments, one transporter of each type

Syncytiotrophoblast aa1, aa2, aa3, 425 µM each Maternal artery aa1, aa2, aa3, 100 µM each Fetal artery aa1, aa2, aa3, 150 µM each Flow Flow

Exchanger aa1, aa2, aa3

Facilitated

diffusion aa3 Accumulative aa1

All KM values 100 μM

Exchanger aa1, aa2, aa3

What can the simplified model tell us?

• How changing individual factors are likely

to affect placental function

• Plasma amino acid concentrations
• Transporter activity
• Blood flow

Varying maternal amino acids

(± 10 fold)

10 x 1.7 x 10 x

• 3.6 x

Increasing one amino acid alone decreases transfer of the others

Accumulative transporter Vmax

The V max of all other transporters A F X X

MVM BM

MVM Exchanger Vmax

A F X X

MVM BM

The V max of all other transporters

BM facilitated transporter Vmax

The V max of all other transporters A F X X

MVM BM

Maternal and fetal flow

Maternal flow* Fetal flow*

*For a given level of transporter activity

Conclusions

• Placental amino acid transfer requires

multiple mechanisims

• But many of these necessary processes may

never become rate limiting

• e.g. flow is essential but even at rates well below

physiological is not rate limiting

• Interventions to optimise fetal growth need

to target rate limiting steps

• Modelling will help us to identify these

Acknowledgements

Southampton - Biologists

• Dr Jane Cleal
• Emma Lofthouse
• Prof Alan Jackson
• Prof Mark Hanson
• Dr Suzanne Brookes
• Dr Priscilla Day

Southampton – Modellers

• Dr Bram Sengers
• Nont Panitchob
• Simone Perazzollo

Oxford - Maths

• Prof Colin Please

Manchester

• Prof Colin Sibley
• Dr Ed Johnstone
• Dr Ian Crocker
• Dr Jo Glazier
• Dr Kate Widdows

Gerald Kerkut Charitable Trust