Pregnancies with extreme fetal-maternal HLA incompatibility
Description
Pregnancy requires local immune tolerization. Oocyte donation (OD) pregnancies, with extensive fetal-maternal HLA mismatching, are at higher risk of pre-eclampsia. We hypothesize that immune adaptations are needed at the fetal-maternal interface to maintain healthy pregnancy despite high HLA dissimilarity. This study is by X. Tian et al. The data sets concern raw HLA genotyping data, imaging mass cytometry data and RNA sequencing data generated from decidua basalis of fully-allogeneic healthy pregnancies, semi-allogeneic healthy pregnancies, and fully-allogeneic pregnancies with pre-eclampsia.
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HLA genotype: HLA typing was applied on maternal and fetal DNA using AlloSeq Tx methodology (CareDx, Brisbane CA, USA). Results of HLA-A, -B, -C, -DRB1, -DRB3/4/5, -DQB1, -DQA1, -DPB1, -DPA1 were used to calculate fetal-maternal HLA mismatches at allele level (4-digit). Software HLA epitope mismatch algorithm (HLA-EMMA) was applied. Imaging mass cytometry: placental slides were cut in 4-μm thick slides and stained with an IMC antibody panel of 42 markers. After deparaffinization and heat-induced epitope retrieval using citrate buffer the slides were incubated for 30 min with 200 μL of Superblock solution (Thermo Fisher Scientific). Next, 100 μL of CD33 and CD4 antibodies diluted in staining buffer (PBS/1%BSA) was added to the slides and incubated overnight at 4oC. The slides were washed three times for 5 min with washing buffer (PBS/1%BSA/0.05% Tween) and then incubated with 100 μL of QDot800-labeled anti-mouse (1:20) and 145Nd-labeled anti-rabbit (1:100) secondary antibodies for 1 hour at room temperature (RT). After washing again for 3x5min, the slides were incubated with 100 μL of metal-labeled antibody mix for 5 hours at RT. The second antibody mix was added to the slides after 3x5min washing and the slides were incubated overnight at 4oC in a humid chamber. After washing again for 3x5min, the slides were incubated for 5 min with 100 μL of Intercalator Iridium (1.25 μM, Fluidigm). The slides were washed 2x5min with washing buffer and 1x5min with demineralized water, and then dried under an airflow. RNA sequencing: to specifically isolate decidua basalis from frozen placental sections, 40-μm thick tissue sections were placed on a glass slide to localize the basalis under a stereomicroscope and swiftly remove it from the placental villous part with a pipette tip. Then, the decidua basalis tissue was immediately placed in ML lysis buffer for RNA extraction. Sequencing was performed on 250 ng of RNA on the Illumina NovaSeq system by GenomeScan (Leiden, The Netherlands). A ribosomal RNA depletion step was incorporated and coverage was 30 million paired end reads (150 bp) per sample. RNAseq data were normalized between samples by count-per-million (cpm) approach.
Institutions
- Leids Universitair Medisch Centrum