Think big and scale down: development of a multi-organ-on- the-chip-model for analysis of immune cell/skin interactions Christine Consentius 1,2 , Juliane Hübner 3,4 , Uwe Marx 3 , Hans-Dieter Volk 2,5 1 Charité Berlin, Berlin-Brandenburg School for Regenerative Therapies (BSRT), 2 Charité Berlin, Berlin-Brandenburg Center for Regenerative Therapies (BCRT), 3 TissUse GmbH, Berlin, 4 Intitute for Biotechnology, Medical Biotechnology, Technical University, Berlin, 5 Charité Berlin, Institute of Medical Immunology INTRODUCTION RESULTS CONCLUSION Our skin allograft on-the-chip model shows signs of rejection and might also be suitable to investigate pathogenetic mechanisms in chronic inflammatory diseases, e.g. psoriasis, or testing of novel therypeutic approaches with the advantage of reducing animal studies. mehr To date the interaction between skin and immune system in the context of chronic inflammatory skin diseases is poorly understood, complicating the development of targeted therapeutic strategies. So far, static 2D or 3D cultures, which are human but not systemic (A), or animal models, which are systemic but not human, can be used for testing. To be more close to the patient’s situation, we established a multi-organ-on-the-chip system (B) in which human skin and immune cells can be cultured together in a dynamic miniaturized human chip culture. This technology – for the first time ever – provides preclinical insight on a systemic level using human tissue. The system closely resembles the activity of multiple human organs in their true physiological context at the smallest possible biological scale. The suitability of multi-organ chips combining skin and immune cells was tested exemplary by establishing a “skin allograft on-the-chip model”. Human skin biopsies are cultured together with allogeneic Peripheral Blood Mononuclear Cells (PBMC) to induce skin rejection. IFNγ [pg/ml] IL2 [pg/ml] IL6 [pg/ml] IL17A [pg/ml] Co-culture skin/allogeneic PBMC Skin only Allogeneic PBMC only Enhanced IFNγ production in skin/immune cell co-culture Dynamic 3D cultures allow testing of disease models close to the patient‘s situation static 2D Culture static 3D Culture Patient dynamic 3D Culture Co-culture skin/allogeneic PBMC Allogeneic PBMC only T cells [%] NK cells [%] B cells [%] [A] [B] LDH [U/l] Glucose consumption [mg/l] [A] [A] [B] T cells preferentially survive and are more antigen-experienced in the co-culture PBMC can be tracked in the skin Fig. 2. [A] Glucose consumption as well as LDH production were analysed in medium. [B] H&E stain of skin freshly injected with allogeneic PBMC (day 0) and of single or co-culture at day 9. [C] Celltracker Green stained PBMC were injected into the skin, after 9 days nuclear counterstaining with Dapi was performed and fluorescence was recorded. 1/7,5s, 20x 1/60s, 20x merge Day 9 – skin only Day 9 – skin + allo PBMC Day 0 – skin + allo PBMC Day 9 – skin + allo PBMC (stained with Celltracker Green) [B] [C] Dapi Celltracker PBMC PBMC Fig. 3. [A] Expression of CD3, CD56 and CD19 on circulating PBMC was assessed by flow cytometry. [B] Flow cytometric analysis of CD3+ T cells for CD45RA and CCR7 expression at day 9. Gating for naive, effector memory (EM), central memory (CM) and terminally differentiated effector memory (EMRA) T cells is shown. Fig. 4. Concentrations of IFNγ, IL2, IL6 and IL17A were measured by Meso Scale Discovery. REJECTION Fig. 1. [A] Dynamic 3D cultures are as close to the patient as possible.. [B] Design of the TissUse Two- Organ-Chips, which are connected to control units. PBMC only Co-culture T naive T CM T EM T EMRA T naive T CM T EM T EMRA

Think big and scale down: development of a multi-organ-on ... Think big and scale down: development of a multi-organ-on-the-chip-model for analysis of immune cell/skin interactions

Download PDF Report

Upload
others
View
0
Download
0

Embed Size (px)

Citation preview

Page 1: Think big and scale down: development of a multi-organ-on ... Think big and scale down: development of a multi-organ-on-the-chip-model for analysis of immune cell/skin interactions

Think big and scale down: development of a multi-organ-on-the-chip-model for analysis of immune cell/skin interactions

Christine Consentius1,2, Juliane Hübner3,4, Uwe Marx3, Hans-Dieter Volk2,5

1 Charité Berlin, Berlin-Brandenburg School for Regenerative Therapies (BSRT), 2 Charité Berlin, Berlin-Brandenburg Center for Regenerative Therapies (BCRT), 3 TissUse GmbH, Berlin, 4 Intitute for Biotechnology, Medical Biotechnology, Technical

University, Berlin, 5 Charité Berlin, Institute of Medical Immunology

INTRODUCTION

RESULTS

CONCLUSIONOur skin allograft on-the-chip model shows signs of rejection and might also be suitable toinvestigate pathogenetic mechanisms in chronic inflammatory diseases, e.g. psoriasis, ortesting of novel therypeutic approaches with the advantage of reducing animal studies.

mehr

To date the interaction between skin and immune system in the context of chronic inflammatory skin diseases is poorly understood,complicating the development of targeted therapeutic strategies. So far, static 2D or 3D cultures, which are human but not systemic (A), oranimal models, which are systemic but not human, can be used for testing. To be more close to the patient’s situation, we established amulti-organ-on-the-chip system (B) in which human skin and immune cells can be cultured together in a dynamic miniaturized human chipculture. This technology – for the first time ever – provides preclinical insight on a systemic level using human tissue. The system closelyresembles the activity of multiple human organs in their true physiological context at the smallest possible biological scale. The suitabilityof multi-organ chips combining skin and immune cells was tested exemplary by establishing a “skin allograft on-the-chip model”. Humanskin biopsies are cultured together with allogeneic Peripheral Blood Mononuclear Cells (PBMC) to induce skin rejection.

IFNγ [pg/ml] IL2 [pg/ml] IL6 [pg/ml] IL17A [pg/ml]

Co-culture skin/allogeneic PBMC Skin only Allogeneic PBMC only

Enhanced IFNγ production in skin/immune cell co-culture

Dynamic 3D cultures allow testing of disease models close to the patient‘s situation

static2D Culture

static3D Culture

Patientdynamic3D Culture

Co-culture skin/allogeneic PBMC Allogeneic PBMC only

T cells [%] NK cells [%] B cells [%] [A] [B]

LDH [U/l]Glucose consumption [mg/l]

[A]

[A] [B]

T cells preferentially survive and are more antigen-experienced in the co-culture

PBMC can be tracked in the skin

Fig. 2. [A] Glucose consumption aswell as LDH production wereanalysed in medium. [B] H&E stainof skin freshly injected withallogeneic PBMC (day 0) and ofsingle or co-culture at day 9. [C]Celltracker Green stained PBMCwere injected into the skin, after 9days nuclear counterstaining withDapi was performed andfluorescence was recorded.

1/7,5s, 20x1/60s, 20x merge

Day 9 – skin only Day 9 – skin + allo PBMCDay 0 – skin + allo PBMC

Day 9 – skin + allo PBMC (stained with Celltracker Green)

[B]

[C]

Dapi Celltracker

PBMC

Fig. 3. [A] Expression of CD3, CD56and CD19 on circulating PBMC wasassessed by flow cytometry. [B]Flow cytometric analysis of CD3+ Tcells for CD45RA and CCR7expression at day 9. Gating fornaive, effector memory (EM),central memory (CM) andterminally differentiated effectormemory (EMRA) T cells is shown.

Fig. 4. Concentrations of IFNγ, IL2,IL6 and IL17A were measured byMeso Scale Discovery.

REJECTION

Fig. 1. [A] Dynamic 3D cultures areas close to the patient as possible..[B] Design of the TissUse Two-Organ-Chips, which are connectedto control units.

PBMC only Co-culture

TnaiveTCM

TEM TEMRA

TnaiveTCM

TEM TEMRA