University of Padova – Chair of Nephrology

Department of Nephrology Dialysis and Transplantation

International Renal Research Institute

St. Bortolo Hospital, Vicenza - Italy

Claudio Ronco

Renal Glomerular Filtration System

A size- (and charge-)selective blood-to-urine barrier

Glomerular capillary wall Glomerular filtration slit

S. Bowry

Dialysis Membranes: Classification

• Composition and structure (Biomaterial / Process)• Cellulosic vs Synthetic: Polymer Composition

• Pore size, Pore size distribution, thickness

• Performance characteristics:• Permeability / Efficiency / Flux / Cut-off / Adsorption

• Surface modification (functionalization):• Hydrophilic / hydrophobic

• Roughness

• Electrical charges

• Additives

New Spinning Technologies

Change of the spinning conditions:

• More homogeneous polymer solution mixing

• Refined precipitation conditions

• Improved design of spinning nozzles

Results in :

• Defined Pore structure of thin inner ‘skin’ region

• The fibre dimensions (ID + wall thickness)

• Optimised fibre structure (“wavy” fibres)

• Geometry & distribution of the pores of the innermost

surface controlled at nano-scale level

Conventional pores: ragged, tortuous below surface

Nanocontrolled pores: smooth, cylindrical: less resistance

Basis of the nanostructure of membrane pores

Outer membrane

Skin layer

Dead end pore

Stenotic pore

Old Synthetic Membranes

Standardized pore structure

Nanocontrolled Membrane

Improved Synthetic Membr

Mix of small and large pores

Membrane Surface

S. Bowry

Computer Image Analysis (CIA) of Transmission Electron Micrographs

1. Thin, separating layer.

2. High local porosity - and decreasing polymer concentration.

A B

C D

X 20.000

X 80.000

1 µm

100 µm 250 nm

X 20.000

X 80.000

1 µm

100 µm 250 nm

X 20.000

X 80.000

1 µm

100 µm 250 nm

X 20.000

X 80.000

1 µm

100 µm 250 nm

Surface Microdomains may limit Interactions with Proteins and Cells

IMPORTANCE OF HYDROPHILIC SURFACES OF PORES

Can pore size distribution

be precisely controlled ?

High Cut-Off (HCO) vs High-Flux Membranes

High Cut-Off

HighFlux

0

0,2

0,4

0,6

0,8

1

100 1000 10000 100000

Molecular Weight [Dalton]

Siev

ing

Co

effi

cien

t

Creatinine(113)

Vit. B12(1355)

Inulin(5200)

2-M(11.800)

Albumin(68.000)

Low-Flux

High-Flux S

Helixone

HPS

High-Flux LS

High Flux

Low Flux

High Flux

Mod Cell

Mod Surf

NCS Mod

High Cut off

Membranes and Therapies

Protein bound solutes

Mediators of inflammation

Erythropoiesis inhibitors

Smart Cut off

Figure 10

A B C

D

Blood In Blood Out

Dial. InUF or Effluent

Diffusable Solutes

membrane

Non diffusable solutes

Proteins

Cells

K = QF/S (S= [UF]/ [Pw]

Effluent equals K when S=1

S≠ [UF]/ [Pw] when

[UF] < [Pw]1 (membrane)

[Pw]2 < [Pw]1

Post Dilution

[Pw]1

Pre Dilution

> [Pw]2

Creatinine Sieving Coefficient over time

The Membrane as an active component

Endotoxins or proinflammatory mediators can be adsorbed

S. Bowry

Dialysis Membranes: Classification

• Composition and structure (Biomaterial + Process)– Cellulosic vs Synthetic: Polymer Composition

– Pore size, Pore size distribution, thickness

• Performance characteristics:– Permeability / Efficiency / Flux / Cut-off / Adsorption

• Surface modification (functionalization):– Hydrophilic / hydrophobic

– Roughness

– Electrical charges

– Additives

Effects of Surface Modification

Ultra-thin Layer

Biocompatibility, Permeability, Non-fouling effect

Highly Adsorbitive MembranesProteins

Polymer chain

Ionic

interactions

Van der Waals

forces

AdsorptionHydrophobic

domain

Hydrophobic

interactions

MatrixWeak

Medium

Strong

AN69 ST* and Heparin Adsorption

Polyethyleneimine

Hep.

Heparin

oXiris Membrane - Material

Free amine groups of PEI → endotoxin adsorption

CH2CH C

-

CH2

CH2

CH3

CN

SO3 Na-- --+

Bioactivity

AN69

N

NH

N

NH

NH2

NH NH

Polyethylene-imine

-

-

-

-

heparin

Available

active sites

for AT link

PS PMMA AN69

asymmetric structure

microporous membranes

symmetric structure

microporous membranes

symmetric structure

hydrogel membrane

Effective surface accessible for protein adsorptionProteins

Polymer chain

scanning electron microscopy

AN69 based membrane

Acrylonitrile Sodium Methallyl Sulfonate

Hydrophilic

Negative charge

−CH2−CH

CN n

CH2− C −

CH2

CH3

SO3 Na

m

- +

AN69 hydrogel membrane

(社内資料）

Hydrogel structure

TNF-α IL-1β IL-6 IL-8 IL-10 HMGB1

Cytokine clearance

(CHDF at 3 hour)

0

20

40

60

Clearance

(mL/min)

57.1±46.0

(n=15)

(n=19)

(n=32)

(n=23)

(n=25)

(n=13)

：AN69ST-CHDF trial

Data were used when blood cytokine levels before treatment were above the following cut-off values;

TNF-α: 10 pg/mL, IL-1β: 10pg/mL, IL-6: 100 pg/mL, IL-8：100 pg/mL, IL-10: 8 pg/mL, HMGB1: 10 ng/mL

amino group（positive）

sulfonate group

（negative）HH

HN

O

O

O

O O

O

O

OS

S

S

+-

Membrane nature allows ionic binding for targeted removal

Adsorptive β2-Microglobulin Removal

by AN69 vs Cellulose TriacetateClark et al, Kidney Int 1995

AN69

CTA

Β2M Adsorption Isotherms Clark et al, Kidney Int 1994

Porous AN69

Non-porous AN69

HMGB-1 (High Mobility Group 1 protein) is secreted by immune cells

(macrophages & monocytes) as a cytokine mediator of Inflammation.

Oxiris®: animal data

Oxiris®: animal data

- P. Aeruginosa porcine model of septic shock

- 2 × 10 pigs : 6 h of HF with oxiris versus 6 h of HF with a standard mb

- Arterial and Swan-Ganz catheters to assess hemodynamics

- MAP and PCWP maintained stable with crystalloids, colloids and epinephrine infusion

Endotoxin Removal – AN69 oXiris

Rimmele T et al., NDT 2009;24:354-357

Rimmelé, Peng, Kellum. unpublished data

oxiris ®

Hemoperfusion

Sham

Oxiris®: animal data (10 rats per group)

Cytokine Removal During Conventional CRRTDe Vriese et Al

Time (hours) T

NF

-α

IL-1

β

IL-6

TN

F-α

IL-1

β

IL-6

TN

F-α

IL-1

β

IL-6

TN

F-α

IL-1

β

IL-6

TN

F-α

IL-1

β

IL-6

TN

F-α

IL-1

β

IL-6

% t

ota

l re

moval 40

30

20

10

0

t=1 t=6 t=12 t=13 t=18 t=24

% AD% UF

*

**

*

*

*

*

Total amount of cytokine removed is expressed as a percentage of the amount in present in prefilter plasma

Hemofilter:AN69

In AKI, there is an imbalance between pro- and anti-oxidant factorsleading to potential tissue damage

In CRRT, blood membrane interaction may lead to production of reactive oxidant species contributing to damage

Vit E-coated membranes In addition to antioxidant therapy

Strategy

Direct scavenging on the membrane surface is preferable. Gastrointestinalabsorption of oral supplements can be affected by many factors, patients may notbe compliant, and oral supplements can be cleared by the dialysis process.

Taking advantage of new membrane manufacturing processes andmembrane surface modification/functionalization we can perform a

The effect on ROS is exerted in situ on blood cells without release of the vitamin E

This results in improved biocompatibility, with less activation of leukocytes and productionof inflammatory cytokines, as well as decreased endothelial cell activation due toscavenging of ROS products from vitamin E.

OxidativeSTRESS

The Role of Vitamin E- Coated Membrane

0

50.000

100.000

150.000

200.000

250.000

300.000

350.000

400.000

t0MPO t2MPO t24MPO

Filter E Filter NE

CRRT in Acute Kidney Injury Comparison activity MOPpg/m

l

Time from Tx beginning

n.16

MicroMacro Nano

mmcm nm

S. Bowry

Functionalization allows a new dimension in

dialysis membranes