1. To establish optimized approaches for the isolation and characterization of BOECs, CD34+-derived MKs, iPSC- ECs and iPSC-MKs.2. Dissect the cellular mechanisms that control endothelial and platelet (secretory) function, thereby understanding the…
ID
Source
Brief title
Condition
- Other condition
- Coagulopathies and bleeding diatheses (excl thrombocytopenic)
- Blood and lymphatic system disorders congenital
Synonym
Health condition
inherited platelet function disorders
Research involving
Sponsors and support
Intervention
Outcome measures
Primary outcome
A maximum of seventy milliliters of blood will be drawn for the isolation of
PBMCs, platelets, plasma and DNA. Several parameters will be measured in plasma
to characterize the hemostatic and angiogenic profile. BOECs, iPSC-ECs/MKs and
CD34+-derived primary MKs will be characterized in vitro. Protein expression
profiles will be determined by whole proteome mass spectrometry. DNA will be
genotyped for SNPs and mutations in VWF and VWF associated genes.
Secondary outcome
n/a
Background summary
Rationale:
Hemostasis is critically dependent on Von Willebrand factor (VWF), a multimeric
adhesive plasma protein that is crucial for mediating platelet adhesion to
sites of vascular damage and that acts as a chaperone for coagulation factor
VIII (FVIII) in plasma. The bulk of plasma VWF is synthesized by endothelial
cells (ECs) where it is stored in and released from Weibel-Palade bodies
(WPBs). Platelets also store and secrete VWF from alpha-granules after
activation, which contributes to thrombus formation. The molecular mechanisms
that control secretion of VWF from ECs and platelets are poorly understood.
Abnormalities in the (secretory) function of platelets and endothelial cells
can lead to bleeding, such as observed in platelet storage pool disorders
(SPD), Von Willebrand disease (VWD) or in individuals with *low VWF*.
Problem definition:
In ~30% of type 1 VWD patients and individuals with low VWF (defined as VWF
levels <50 IU/dL) no VWF mutations are found. Another problem is that in around
50% of bleeding patients referred to tertiary centers the underlying mechanisms
that cause clinically relevant bleeding problems cannot yet be identified,
so-called bleeding of unknown cause (BUC). Desmopressin is administered
therapeutically to improve platelet function and raise VWF in plasma by
inducing WPB exocytosis from endothelial cells. In ~25% of patients
desmopressin fails to trigger (sufficient) release of VWF/FVIII for reasons
largely unknown.
Research hypothesis:
The main hypothesis of this study is that defects in components of the
secretory machinery of VWF are causative for reduced VWF levels and lack of
desmopressin response. Also, defects in this machinery may be causative in
patients with BUC. To identify new determinants of VWF levels and vascular
health we will study secretory mechanisms in ECs and platelets of individuals
with abnormalities in hemostatic and/or secretory function. For this we will
establish ex vivo (patient-derived) cellular model systems of endothelial and
platelet secretion using blood outgrowth endothelial cells (BOECs), platelets,
CD34+-derived megakaryocytes and induced pluripotent stem cell (iPSC)-derived
endothelial cells (iPSC-ECs) and megakaryocytes (iPSC-MKs).
Study objective
1. To establish optimized approaches for the isolation and characterization of
BOECs, CD34+-derived MKs, iPSC- ECs and iPSC-MKs.
2. Dissect the cellular mechanisms that control endothelial and platelet
(secretory) function, thereby understanding the pathophysiology of bleeding
disorders and the genetic control of VWF secretion using BOECs, platelets,
CD34+-derived MKs, iPSC- ECs and iPSC-MKs.
3. To develop in vitro methods to introduce or correct (disease-causing)
mutations in regulators of endothelial or platelet function in BOECs or
iPSC-derived ECs/MK using CRISPR-mediated genome engineering.
Study design
The study is a basic science, experimental in vitro study that requires the
sampling of venous peripheral blood of the participants.
Study burden and risks
The objectives of this study can only be reached by studying patients and
individuals with hemostatic and/or secretory abnormalities, their family
members and healthy controls. The participants do not directly benefit from
this research, however the burden is minimal as it is restricted to a
venepuncture of negligible risk. Participants may be requested for repeated
sampling, however this will be limited to three times.
Dr. Molewaterplein 40
Rotterdam 3015 GD
NL
Dr. Molewaterplein 40
Rotterdam 3015 GD
NL
Listed location countries
Age
Inclusion criteria
In order to be eligible to participate in this study, a subject must meet all
of the following criteria:
Patients
• Prior diagnosis of VWD, low VWF, inherited platelet function defects,
bleeding of unknown cause or genetic disorders in which dysfunction of the
secretory pathway has been implicated in disease pathogenesis.
• For patients >=16 years old; written informed consent
• For patients 12-15 years old; written informed consent from both the patient
and their parent(s)/legal guardian(s)
• For patients <12 years old; written informed consent from their
parent(s)/legal guardian(s)
Family members
• Age 18 years or older
• Family member of a patient with VWD, low VWF, inherited platelet function
defects, bleeding of unknown cause or genetic disorders in which dysfunction of
the secretory pathway has been implicated in disease pathogenesis.
• Family members may or may not be affected.
Healthy controls
• Age 18 years or older
• Not previously diagnosed with VWD, low VWF, inherited platelet function
defects, bleeding of unknown cause or genetic disorders in which dysfunction of
the secretory pathway has been implicated in disease pathogenesis.
Exclusion criteria
A potential subject who meets any of the following criteria will be excluded
from participation in this study:
• Unable to give written informed consent.
• Use of medication that can compromise platelet function or hemostasis.
Design
Recruitment
Followed up by the following (possibly more current) registration
No registrations found.
Other (possibly less up-to-date) registrations in this register
No registrations found.
In other registers
| Register | ID |
|---|---|
| CCMO | NL72564.078.20 |