Journal: EMBO Reports

Article Title: Cyst-independent oocyte phagocytosis builds the female reproductive reserve in mice

doi: 10.1038/s44319-025-00663-7

Figure Lengend Snippet: ( A ) A representative time-lapse image showing the oocyte developmental dynamics in live ovaries in vitro over a period of 162 h, from c-17.5 dpc to c-PD4. Scale bar: 20 μm. ( B ) Tracing the developmental dynamics of oocytes in the time-lapse image. All oocytes were inverted to black/white (b/w), and a survival oocyte was colored cyan, while a sacrificed oocyte was colored orange to highlight their developmental dynamics. Scale bar: 20 μm. All images of the surviving oocyte development are shown in Appendix Fig. . ( C ) Representative images showing the typical behaviors of oocytes during development, including growth (arrows in lane 1), shrinkage (arrows in lane 2), degradation and deformation. Scale bar: 10 μm. ( D ) Representative images showing the filopodia formation (arrows) of oocytes during development. Green, oocytes; Red, somatic cells. Scale bar: 10 μm. ( E ) 3D static images showing that oocytes are connected to cyst-like structures at any detected time points. Green, oocyte; Red, somatic cells. Scale bar: 10 μm. ( F ) Tracing the oocyte movement in the 4D tracing images (from the time point 1 to point 2 of Fig. 2E). Showing connected oocytes moved in different directions, and most of the cyst-like structures were actually assembled by separated single oocytes in the ovaries after c-17.5 dpc. Cyan arrows: oocyte movement direction. Scale bar: 10 μm. ( G ) Quantifying the ratio of separated oocytes in total oocytes from c-17.5 dpc to c-PD1. By continuously tracing the oocyte movement in the 4D time-lapse imaging, the ratio of oocytes with separation events in total oocytes was counted every 24 h. Data were presented as the mean ± SD. n = 6 ovaries ( n > 40 oocytes per ovary). c-18.5 dpc vs. c-17.5 dpc: p value = 0.213; c-19.5 dpc vs. c-17.5 dpc: p value = 0.0003; c-PD1 vs. c-17.5 dpc: p value = 5.20E-06; c-19.5 dpc vs. c-18.5 dpc: p value = 0.0033; c-PD1 vs. c-18.5 dpc: p value = 7.60E-05; c-PD1 vs. c-19.5 dpc: p value = 0.011. ( H ) Identifying the movement of labeled oocytes in the Oct4-CreER T2 ;mTmG ovaries after low dosage of tamoxifen treatment. Only a small portion of cysts were labeled in the ovaries, with most of them breaking down into single oocytes at c-17.5 dpc. Green, oocytes; Red, somatic cells. Scale bar: 20 μm. ( I ) The ratio of single oocytes in total labeled oocytes from c-17.5 dpc to c-PD1 in ovaries with low labeling density of oocytes. Data were presented as the mean ± SD. n ≥ 6 ovaries ( n > 15 oocytes per ovary). c-18.5 dpc vs. c-17.5 dpc: p value = 3.01E-05; c-19.5 dpc vs. c-17.5 dpc: p value = 0.0001; c-PD1 vs. c-17.5 dpc: p value = 7.64E-08; c-19.5 dpc vs. c-18.5 dpc: p value = 0.0208; c-PD1 vs. c-18.5 dpc: p value = 0.0007; c-PD1 vs. c-19.5 dpc: p value = 0.608. Statistical significance was determined by unpaired one-way ANOVA tests. P (a, b) < 0.05, P (a, c) < 0.05, P (b, c) < 0.05. .

Article Snippet: To obtain the subcellular structures on oocytes in the imaging, the Z-stack images of time-lapse imaging were projected for about 30 μm by Imaris or ImageJ software ( http://rsbweb.nih.gov/ij ) to reconstitute 3D oocyte live images.

Techniques: In Vitro, Imaging, Labeling

Journal: EMBO Reports

Article Title: Cyst-independent oocyte phagocytosis builds the female reproductive reserve in mice

doi: 10.1038/s44319-025-00663-7

Figure Lengend Snippet: ( A ) A representative image showing the developing cellular characteristics of surviving oocytes. Highlighting the formation of filopodia-like structures (FLs, arrows) and surrounded by various oocyte debris (OD, arrowheads). Scale bar: 10 μm. ( B ) Tracing the derivation of ODs. The sacrificed oocyte was colored red to highlight the progress of OD formation. Scale bar: 10 μm. ( C ) The frequency of surviving oocytes forming FLs (green line) and sacrificed oocytes forming ODs (red line), showing a consistent peaking period of two events (brown region) from c-19.5 dpc to c-PD2. Data were presented as the mean ± SEM. n = 4 ovaries (more than 40 oocytes in each time point per ovary). ( D ) The FLs from surviving oocytes extended towards the ODs to assist them in moving towards the surviving oocyte and then attached ODs. Scale bar: 10 μm. ( E ) Representative time-lapse images showing the detailed progress of how surviving oocytes absorbed the ODs. Scale bar: 10 μm. ( F ) FLs on the oocytes and ODs were observed in fresh transparent ovaries at PD1. Scale bar: 10 μm. ( G ) The graphic model illustrates the cyst-independent oocyte phagocytosis in mouse ovaries. The sacrificed oocyte sacrifices itself to form various ODs containing organelles, enriched cytoplasm. The ODs are caught by the surviving oocyte formed FLs to move forward to the surviving oocyte. Then the surviving oocyte absorbs the ODs to enrich the cytoplasm and organelles for survival. Green, surviving oocyte; Red, sacrificed oocyte; Dark red, ODs. .

Article Snippet: To obtain the subcellular structures on oocytes in the imaging, the Z-stack images of time-lapse imaging were projected for about 30 μm by Imaris or ImageJ software ( http://rsbweb.nih.gov/ij ) to reconstitute 3D oocyte live images.

Techniques:

Journal: EMBO Reports

Article Title: Cyst-independent oocyte phagocytosis builds the female reproductive reserve in mice

doi: 10.1038/s44319-025-00663-7

Figure Lengend Snippet: ( A ) 3D reconstructed images of Oct4-CreER T2 ;Rainbow ovaries are shown, displaying random expressions of RFP (red) or CFP (blue) in oocyte cytoplasm. Cytoplasm exchange leads to a mixture of fluorescence in the oocyte (purple, arrow). Scale bar: 10 μm. ( B ) Analysis of oocyte fluorescence reveals an increased number of oocytes with mixed fluorescent cytoplasm (purple, arrows) from c-17.5 dpc to c-PD3. Scale bar: 10 μm. ( C ) The quantification of the ratio of purple oocytes in total oocytes from c-17.5 dpc to c-PD4, showing that the peak of cytoplasm exchanges occurs between c-PD1 to c-PD3. Data were presented as the mean ± SD. n = 4 ovaries. More than 200 oocytes were measured at each time point. ( D ) Tracing the attachment between oocytes with different fluorescence in the time-lapse imaging. No cytoplasm exchange occurs after attachments (arrowheads) between oocytes. Scale bar: 5 μm. ( E ) The reconstructed 3D images demonstrate the existence of ODs (arrowheads) surrounding the oocyte (arrow) in the Oct4-CreER T2 ;Rainbow ovaries. ( F ) The time-lapse images trace the progress of a blue surviving oocyte (arrow) absorbing red ODs (arrowheads), leading to a gradual change of cytoplasm color from blue to purple. The bottom line highlights the color change of the surviving oocyte cytoplasm. Scale bar: 10 μm. ( G ) Analyzing the absolute area of mitochondria in oocytes (red: mitochondria, green: oocytes). Showing a significantly increased average mitochondria area in oocytes at c-PD2 compared to that in oocytes at c-19.5 dpc. Scale bar: 10 μm. More than 50 oocytes were measured at each time point. Data were presented as the mean ± SD. p value = 3.21E-11. *** P < 0.001, by two-tailed unpaired Student’s t -test. ( H ) Comparing the relative density of mitochondria in oocytes and ODs (red: mitochondria, green: oocytes or ODs) at c-PD1 (left). A great variation of mitochondria density was observed in ODs but not in oocytes (right). Scale bar: 10 μm. Oocytes: n = 23; ODs: n = 107. Data were presented as the mean ± SD. .

Article Snippet: To obtain the subcellular structures on oocytes in the imaging, the Z-stack images of time-lapse imaging were projected for about 30 μm by Imaris or ImageJ software ( http://rsbweb.nih.gov/ij ) to reconstitute 3D oocyte live images.

Techniques: Fluorescence, Imaging, Two Tailed Test