Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: A rare germline BMP15 missense mutation causes hereditary ovarian immature teratoma in human.

doi: 10.1073/pnas.2310409121

Figure Lengend Snippet: Fig. 1. Pathogenic mutant in familial ovarian immature teratoma. (A) Cases from two pedigree of OIT and their corresponding genotypes of BMP15. (B) Pathological diagnosis of corresponding patients. Blue dash-circled areas and arrowheads indicate two to three germ layer-derived tissues in the tumor tissue; neural components and neuroglia in the ectoderm; and the neuroepithelium is connected into pieces in a daisy doughnut or irregular glandular shape. (C) Sanger sequencing results of corresponding patients. (D) Conservation of Arg 88 across various species. (E) Hydrophilicity of Arg 88 across various species. (F) Predicted structural change of c.C262T (MT) mutated BMP15.

Article Snippet: Mouse Model. C57BL/6 mice harboring BMP15 R86C mutation (equivalent to BMP15 R88C in humans) were generated by Cas9/CRISPR- mediated genome editing (Cyagen Biosciences).

Techniques: Mutagenesis, Biomarker Discovery, Derivative Assay, Sequencing

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: A rare germline BMP15 missense mutation causes hereditary ovarian immature teratoma in human.

doi: 10.1073/pnas.2310409121

Figure Lengend Snippet: Fig. 3. C262T mutant significantly stimulates parthenogenetic activation of oocytes. Oocytes were collected from genetically engineered mice harboring the orthologous of BMP15 wild type or C262T variants. Superovulation was induced at 4 wk of age mice. Ten ovaries were assessed from each genotype. The parthenogenetic activation in oocyte was assessed by morphological changes (panel A–C) or histological stain (panel D and E). (A) Two pronucleus were seen in oocyte with spontaneous parthenogenetic activation (yellow arrowhead); (B) two pronucleus and two polar bodies were seen in oocyte with spontaneous parthenogenetic activation (yellow and blue arrowheads, respectively); (C) Two-cell (green arrowheads) was seen with spontaneous parthenogenetic activation; (D) The oocytes appear to have two pronucleus in the HE section (yellow arrowhead); (E) Two polar bodies and one pronucleus appear in the oocyte in the HE section (yellow and blue arrowheads, respectively); (F) Two-cell (green arrowheads) appeared in HE sections; (G) The percentage of parthenogenetically activated oocytes in different BMP15 variants was counted under the Leica DMI3000B inverted biological microscope.

Article Snippet: Mouse Model. C57BL/6 mice harboring BMP15 R86C mutation (equivalent to BMP15 R88C in humans) were generated by Cas9/CRISPR- mediated genome editing (Cyagen Biosciences).

Techniques: Mutagenesis, Activation Assay, Staining, Microscopy

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: A rare germline BMP15 missense mutation causes hereditary ovarian immature teratoma in human.

doi: 10.1073/pnas.2310409121

Figure Lengend Snippet: Fig. 2. C262T mutant affects the protein expression of BMP15. (A) Wild type and C262T mutated BMP15 were overexpressed in HEK293 cells. Amounts of secreted BMP15 (s-BMP15) were compared by western blot. Signal intensity was semi-quantified using ImageJ with the arbitrary unit (P < 0.05). (B) The general BMP15 protein level in a healthy ovarian, BMP15 was highly expressed in both granulosa cells (red arrow) and oocytes (blue arrow) of healthy ovaries; (C) Wild-type BMP15 in sporadic ovarian immature teratoma, BMP15 is highly expressed in the follicular epithelium of tumors (black arrow). (D) C262T variant in familial ovarian immature teratoma. The expression level of the mature BMP15 protein was significantly decreased in immature teratomas carrying the C262T mutation (blue arrow).

Article Snippet: Mouse Model. C57BL/6 mice harboring BMP15 R86C mutation (equivalent to BMP15 R88C in humans) were generated by Cas9/CRISPR- mediated genome editing (Cyagen Biosciences).

Techniques: Mutagenesis, Expressing, Western Blot, Variant Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: A rare germline BMP15 missense mutation causes hereditary ovarian immature teratoma in human.

doi: 10.1073/pnas.2310409121

Figure Lengend Snippet: Fig. 4. H-Ras activated MAPK stimulate spontaneous activation of the oocytes and overgrown of the ovary. (A) Venn diagram showing the enriching strategy of oocytes specific RNA sequencing results. (B) Heat map showing the distinct diverse pattern of differential expressed genes in oocytes with wild-type or mutated BMP15. (C) H-Ras and MEK1 RNA levels in CC, CT, or TT type oocytes. (D) IHC stain showing the BMP15, H-Ras, MEK1, and p-MEK1 protein levels in CC, CT, or TT type ovarian tissues at week 4. (E) (i) Hematoxylin and eosin stain of ovarian tissues at the longest latitude section; blue arrowheads show follicles; (ii) the actual ovary size of CC and CT type at month 6; (iii) average counts of ovarian follicles per slicing section at month 6. PF, primary follicle; SF, secondary follicle; MF, mature follicle; AF, atretic follicle; *** indicates P-value < 0.001. (F) An ovarian teratoma in the CT type ovary. Blue arrowhead shows a digestive tract structure composed of layers of epithelial cells. (G) IHC stain showing the BMP15, H-Ras, MEK1, and p-MEK1 protein levels in a CC type healthy ovary and the CT type ovary with the teratoma at month 6.

Article Snippet: Mouse Model. C57BL/6 mice harboring BMP15 R86C mutation (equivalent to BMP15 R88C in humans) were generated by Cas9/CRISPR- mediated genome editing (Cyagen Biosciences).

Techniques: Activation Assay, RNA Sequencing, Staining, H&E Stain