Figure 1

Figure 1 Cbx7a/CBX7 is upregulated in different CCM disease models and lesion material of familial CCM patients.

(A) Volcano plot of microarray data of differentially-expressed genes (calculated with the limma R package (Ritchie et al, 2015) with moderated T-statistic, adjusted p value <0.05 and fold change >1.4) in zebrafish ccm2^m201 mutants (Renz et al, 2015). (B) Shown are expression fold-changes of Cbx7a/CBX7 mRNA in different CCM disease models. Elevated mRNA expression levels of zebrafish cbx7a in krit1^ty219c and ccm2^m201 mutants (RNA-sequencing of extracted hearts at 48 hours post fertilisation (hpf)), murine Cbx7 in Ccm1/Krit1^ECKO-derived brain microvascular endothelial cells (BMECs) (Koskimäki et al, 2019), and CBX7 in CCM1 and CCM2-deficient human iPSC-derived ECs (ECs) (RNA-sequencing). (C–E) Whole-mount in situ hybridization of zebrafish embryo at 56 hpf revealed low levels of cbx7a mRNA in wild-type (wt) embryos (C) and elevated levels in ccm2^m201 (D) and krit1^ty219c (E) mutants throughout the entire heart. The constriction of the atrioventricular canal (AVC) was lost in ccm2^m201 (D) and krit1^ty219c (E) mutants. (F) Quantifications of cbx7a mRNA levels by qRT-PCR from isolated tissue preparations of head, heart, or tail regions in ccm2^m201 and krit1^ty219c mutants as compared to wild-type (wt) (n = 3–4 biological replicates per group, each replicate contained 10–15 pooled tissue samples). Statistical testing is based on the student’s T-test (p values and s.e.m. bars indicated). (G, K) Immunohistochemistry stainings for CBX7 expression. CD34 marks ECs. Only low expression of CBX7 is detected in ECs of healthy brain (K, arrows). In lesions of CCM1 (G, H, J) and CCM2 (I) familial patients, CBX7 staining is markedly increased in ECs (arrows). Asterisks indicate vessel lumen. Ven ventricle, Atr atrium, OFT out flow tract. Experiments in zebrafish were done in three biological replicates. Scale bars are (C–E) 100 µm; (G–K) 200 µm. Source data are available online for this figure.