See the published version of this preprint at Fluids and Barriers of the CNS.
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Study protocol
Leandro Castaneyra-Ruiz
whashington University in Saint Louis
Corresponding Author
License:
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Background:
Severe intraventricular hemorrhage (IVH) is one of the most devastating neurological complications in preterm infants, with the majority suffering long-term neurological morbidity and up to 50 percent developing post hemorrhagic hydrocephalus (PHH). Despite the importance of this disease, its cytopathological mechanisms are not well known. An in vitro model of IVH is required to investigate the effects of blood and its components on the developing ventricular zone (VZ) and its stem cell niche. To address this need, we developed a new in vitro model to mimic the cytopathological conditions of IVH in the preterm infant.
Methods:
Maturing neuroepithelial cells from the VZ were harvested from the entire lateral ventricles of wild type C57BL/6 mice at 1-4 days of age and expanded in proliferation media for 3-5 days. At confluence, cells were re-plated onto 24-well plates in differentiation media to generate ependymal cells (EC). At approximately 3-5 days, which corresponded to the onset of ependymal cell differentiation based on the appearance of multiciliated cells , phosphate-buffered saline for controls or syngeneic whole blood for IVH was added to the ependymal cell surface. The cells were examined for the expression of EC markers of differentiation and maturation to qualitatively and quantitatively assess the effect of blood exposure on VZ transition from neuroepithelial cells to EC.
Discussion:
This model will allow investigators to test cytopathological mechanisms contributing to the pathology of IVH with high temporal resolution and query the impact of injury to the maturation of the VZ. This technique recapitulates features of normal maturation of the VZ in vitro, offering the capacity to investigate the developmental features of VZ biogenesis.
Keywords
Cell Culture, Neural Stem Cells, Ependyma, Ventricular Zone, Intraventricular Hemorrhage, Post- hemorrhagic Hydrocephalus, Preterm, Premature, Neonate
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CURRENT STATUS: Published
View the published article at Fluids and Barriers of the CNS.
No community comments so far
Editorial decision: Accept
On 10 Jul, 2020
Editor assigned
On 08 Jul, 2020
Submission checks complete
On 07 Jul, 2020
Editor invited
On 07 Jul, 2020
No comments provided
Editorial decision: Minor revision
On 19 Jun, 2020
Review #3 received
Received 17 Jun, 2020
Review #2 received
Received 29 May, 2020
Review #1 received
Received 29 May, 2020
Reviewer #3 agreed
On 28 May, 2020
Reviewers invited
Invitations sent on 19 May, 2020
Reviewer #1 agreed
On 19 May, 2020
Reviewer #2 agreed
On 19 May, 2020
Editor assigned
On 16 May, 2020
Submission checks complete
On 15 May, 2020
Editor invited
On 15 May, 2020
Version 1
Posted 12 Mar, 2020
No comments provided
Editorial decision: Major revision
On 23 Mar, 2020
Review #2 received
Received 12 Mar, 2020
Review #1 received
Received 12 Mar, 2020
Review #3 received
Received 12 Mar, 2020
Reviewer #3 agreed
On 09 Mar, 2020
Reviewers invited
Invitations sent on 09 Mar, 2020
Reviewer #1 agreed
On 09 Mar, 2020
Reviewer #2 agreed
On 09 Mar, 2020
Editor assigned
On 06 Mar, 2020
First submitted
On 05 Mar, 2020
Submission checks complete
On 05 Mar, 2020
Editor invited
On 05 Mar, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Cellular & Molecular Neuroscience
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A new preprint design is coming!
We have improved readability, clarity, accessibility, and opportunities for engagement.
See the published version of this preprint at Fluids and Barriers of the CNS.
This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Study protocol
Leandro Castaneyra-Ruiz
whashington University in Saint Louis
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Published
View the published article at Fluids and Barriers of the CNS.
No community comments so far
Editorial decision: Accept
On 10 Jul, 2020
Editor assigned
On 08 Jul, 2020
Submission checks complete
On 07 Jul, 2020
Editor invited
On 07 Jul, 2020
No comments provided
Editorial decision: Minor revision
On 19 Jun, 2020
Review #3 received
Received 17 Jun, 2020
Review #2 received
Received 29 May, 2020
Review #1 received
Received 29 May, 2020
Reviewer #3 agreed
On 28 May, 2020
Reviewers invited
Invitations sent on 19 May, 2020
Reviewer #1 agreed
On 19 May, 2020
Reviewer #2 agreed
On 19 May, 2020
Editor assigned
On 16 May, 2020
Submission checks complete
On 15 May, 2020
Editor invited
On 15 May, 2020
Version 1
Posted 12 Mar, 2020
No comments provided
Editorial decision: Major revision
On 23 Mar, 2020
Review #2 received
Received 12 Mar, 2020
Review #1 received
Received 12 Mar, 2020
Review #3 received
Received 12 Mar, 2020
Reviewer #3 agreed
On 09 Mar, 2020
Reviewers invited
Invitations sent on 09 Mar, 2020
Reviewer #1 agreed
On 09 Mar, 2020
Reviewer #2 agreed
On 09 Mar, 2020
Editor assigned
On 06 Mar, 2020
First submitted
On 05 Mar, 2020
Submission checks complete
On 05 Mar, 2020
Editor invited
On 05 Mar, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Cellular & Molecular Neuroscience
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Fluids and Barriers of the CNS.
Background:
Severe intraventricular hemorrhage (IVH) is one of the most devastating neurological complications in preterm infants, with the majority suffering long-term neurological morbidity and up to 50 percent developing post hemorrhagic hydrocephalus (PHH). Despite the importance of this disease, its cytopathological mechanisms are not well known. An in vitro model of IVH is required to investigate the effects of blood and its components on the developing ventricular zone (VZ) and its stem cell niche. To address this need, we developed a new in vitro model to mimic the cytopathological conditions of IVH in the preterm infant.
Methods:
Maturing neuroepithelial cells from the VZ were harvested from the entire lateral ventricles of wild type C57BL/6 mice at 1-4 days of age and expanded in proliferation media for 3-5 days. At confluence, cells were re-plated onto 24-well plates in differentiation media to generate ependymal cells (EC). At approximately 3-5 days, which corresponded to the onset of ependymal cell differentiation based on the appearance of multiciliated cells , phosphate-buffered saline for controls or syngeneic whole blood for IVH was added to the ependymal cell surface. The cells were examined for the expression of EC markers of differentiation and maturation to qualitatively and quantitatively assess the effect of blood exposure on VZ transition from neuroepithelial cells to EC.
Discussion:
This model will allow investigators to test cytopathological mechanisms contributing to the pathology of IVH with high temporal resolution and query the impact of injury to the maturation of the VZ. This technique recapitulates features of normal maturation of the VZ in vitro, offering the capacity to investigate the developmental features of VZ biogenesis.
Figure 1
Figure 2
Figure 3
The full text of this article is available to read as a PDF.
This is a list of supplementary files associated with this preprint. Click to download.
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