54 - Distinct neuronal subtypes in fetal CSF characterized by single-cell transcriptomics

Friday, February 10, 2023

2:15 PM – 2:30 PM

Location: Room 3022, Moscone West, Level 3

Objective: Cells within the fetal cerebrospinal fluid (CSF) at 22-26 weeks’ gestation are only now being characterized. During fetoscopic myelomeningocele repair, fetal CSF is drawn from the lesion as part-and-parcel of the repair. We hypothesized most of these cells would be neuronal and neuroepithelial cell-types.

Study Design: We have generated a repository of paired amniotic fluid (AF) and fetal CSF specimens from >50 fetoscopic surgeries consenting for use. We used orthogonal methods to catalog cell populations in the fetal CSF during gestation. We performed flow cytometry to quantify CD34⁺hematopoietic stem cells, CD56⁺/CD45⁺NK cells, and CD133⁺/CD45⁺hematopoietic progenitor cells (N=4) and established a neurosphere culture system that could be maintained for 4 weeks (N=3). Lastly, we performed single-cell RNA-sequencing (scRNA-seq) as an unbiased approach to characterize paired fetal CSF and AF cell populations (N=9).

Results: By flow, neural precursor cells represented approximately 39% of the CSF cells, leaving many cell-types uncharacterized. Therefore, we utilized scRNA-seq resulting in 22,058 total cells clustered into 18 distinct cell-types (Fig 1a). We then focused on 3,742 transcriptomes of neuronal origin (Fig 1b), revealing fetal CSF neuronal cells were mainly composed of microglia, neuroblasts, and immature neurons (Fig 1c). A cluster of putative ARC capsids, which are endogenous 40nm virus-like capsids carrying neurotransmitter RNA, demonstrated that ARC was expressed in 261 CSF cells from 87% of the samples (Fig 2b). 73 transcriptomes putatively annotated as ARC capsids revealed RNA content payloads associated with cadmium ion response, cell cycle, and transcription regulation (GO pathways, f < 0.05 controlling for multiple comparisons).

Conclusion: Neuronal cell-types in the mid-gestation fetal CSF of ongoing pregnancies provide an unparalleled picture of neuronal development. Further analyses of the recently discovered ARC capsid neuronal signaling system will unveil the sophisticated differentiation trajectories of neurogenesis in utero and how neurons may be engineered or repaired.

Primary & Presenting Author(s)

Enrico R. Barrozo, PhD

Department of Obstetrics & Gynecology, Division of Maternal-Fetal Medicine, Baylor College of Medicine and Texas Children’s Hospital
Houston, Texas, United States

Slides

Coauthor(s)

AB

Angela P. H. Burgess, MD

Baylor College of Medicine and Texas Children’s Hospital, Department of Obstetrics & Gynecology, Division of Maternal-Fetal Medicine
Houston, TX, United States
AC

Alondra Carmona, BS

Baylor College of Medicine and Texas Children’s Hospital, Department of Obstetrics & Gynecology, Division of Maternal-Fetal Medicine
Houston, TX, United States
Maryshe S. Zietsman, BSA

Medical Student
Department of Obstetrics & Gynecology, Division of Maternal-Fetal Medicine, Baylor College of Medicine and Texas Children’s Hospital
Houston, Texas, United States
Maxim D. Seferovic, PhD

Assistant Professor
Department of Obstetrics & Gynecology, Division of Maternal-Fetal Medicine, Baylor College of Medicine and Texas Children’s Hospital
Houston, Texas, United States
Alireza A. Shamshirsaz, MD (he/him/his)

Director of the Maternal Fetal Care Center
Boston Children’s Hospital and Harvard Medical School
Newtown, Massachusetts, United States
Michael A. Belfort, MD,PhD (he/him/his)

Professor, Chairman
Baylor College of Medicine / Texas Children's Hospital
Houston, Texas, United States
Kjersti M. Aagaard, MD,PhD

Professor and Vice Chair of Research Department of Obstetrics and Gynecology, Division of MFM
Texas Children's and Baylor College of Medicine
Houston, Texas, United States