Immune Phenotype of patients long-term after allogeneic Hematopoietic Stem Cell Transplantation (HSCT)

Francesca RossiOncology-Pathology dep.,Huddinge HospitalSupervisor: Dr. Michael UhlinCo-supervisor: Arwen Stikvoort

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Hematopoietic Stem Cell Transplantation (HSCT)

transplantation of multipotent hematopoietic stem cells (HSC) Multipotent: they can become one of several types of cells within a given organ Hematopoietic: they are located in the bone marrow and give rise to all other blood cells

through haematopoiesis

An HSCT can be: Autologous: when the patient’s own HSCs are used for the transplantation Allogeneic: when the stem cells come from another individual

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Procedure

1. Conditioning: administration of chemotherapeutic drugs

2. Transplant: HSC get infused into the recipient’s vein

3. Engraftment: the cells reach the sinuses in the bone barrow

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http://www.mdlingo.com/article/reduced-intensity-conditioning-improves-outcome-of-bone-marrow-engraftment

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Possible reactions after HSCT

GVL: immune-mediated response of remission of the hematological malignancy

Graft failure: when host’s immune cell reject the donor’s cells or when part of the HSCT does not work

GVHD: when donor’s cells attack the host’s tissues

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The closest the compatibility is between donor and recipient,the lower is the risk that complications arise

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AIM

Determine whether the donor and patient’s immune system differed after 10 years since transplantation

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HSC

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DON: > 10 years

REC: > 10 years

DrugsDiseaseLifestyle…

DON REC

HSCT

DON/REC

HSC

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Material & Methods (1)

Groups:1. 7 donors and 7 recipients Unpaired

2. 5 donors and 5 recipients Paired (donors and patients are siblings)

HSCTs took place between 1987 and 2003

Samples collected at least 10 years ago

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Material & Methods (2)

Samples: PBMC “ring” removed from the blood and frozen (-180°C)

BD FACS Canto Flow cytometer (42 different markers used)

FlowJo analysing software

Statistics: IBM – SPSS Statistics, version 23 GraphPad Prism, version 5.00 P values <0.05

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Unpaired: Mann-Whitney (box plots) Paired: Wilcoxon (paired scatter plots)

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Results (1)

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In a healthy person:

CD4+ > CD8+

Previous reports: shortly after transplantation:

CD8+ > CD4+

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Results (2)

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CD95+ recipients > donors

CD95+ ➡️ CD8+ > CD4+ T cells

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Results (3)

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CD95+ ➡️ CD8+ > CD4+ T cells

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CCR7 – CD45RO

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CD45RO+

CCR7+

TN TCM

TEMTTD

TN: naïve T cells express CCR7 marker (lymph nodes)

TCM: cells that guard against infections

TEM: T cells entered in contact with antigens and are ready to enter the bloodstream to defend the body

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Results (4)

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TEMs ➡️ patients > donors in T cells and CD4+ T cells (A and B) Transplantation increases immune system reaction: mature memory T cells

TNs ➡️ donors > patients in CD4+ T cells (C) Less T naïve cells in patients after HSCT

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Results (5)

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CD45RO+ memory Treg ➡️ patients > donors.

Patients after transplantation: lower levels of naïve T cells

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Results (6)

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PD-1: exhaustion marker ➡️ patients > donors

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To summarize…

CD4+/CD8+ shifted

Higher levels of CD95+ in patients (in both groups)

Higher levels of TEM cells in patients and higher levels of TN in donors

Increased levels of CD45RO+ in Treg memory cells in patients

Higher amounts of PD-1 exhaustion marker in patients’ senescent cells

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Considerations & Conclusions

Aspects that may influence immune system cell’s reconstitution: Quality/quantity of mononuclear cells transplanted Genetic disparity/compatibility between donor and recipient Type of transplantation

Immune system’s cells after HSCT The immune system cells in the patient are its donor’s cells Even after many years in the patient some cell populations seem to show relevant differences

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Thank you!

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Antibodies panel – Flow Cytometry

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Medical data of donors and patients included in the study

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