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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with <t>ImageLab</t> software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="250" height="auto" />
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Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with ImageLab software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see <xref ref-type= Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm. " width="100%" height="100%">

Journal: Frontiers in Endocrinology

Article Title: Tailor-made 3D in vitro maturation of early antral follicles uncovers cumulus-cell transcriptomic driver signature to predict oocyte competence

doi: 10.3389/fendo.2025.1629815

Figure Lengend Snippet: Spatial distribution and expression of cumulus cell genes linked to oocyte maturity. Validation of drivers through immunoblotting and immunofluorescence (A, B) analyses. HAS2 was shown for Network 1(MII Endpoint - GV Startpoint ); CASP3 for Network 2(GV Endpoint -GV Startpoint ); SEMA3A for Pairwise 3 (MII Endpoint - GV Endpoint . (A) Immunoblotting of the selected drivers. Blot quantification was made with ImageLab software by Bio-Rad. Data (mean ± SD) represent 3 independent sets of experiments (n = at least 3 biological replicates in each group per set; each biological replicate assayed in at least 3 technical replicates). * and *** Statistically significant values between the different studied groups (p<0.05 and p<0.001respectively). (B) Representative confocal images of co-immunofluorescence staining of nuclei (Hoechst), Alexa-Fluor 488 ( SEMA3A or CASP3 ) or Alexa-Fluor 568 ( HAS2 ) in CCs enclosing oocytes in the different nuclear stage. HAS2 and SEMA3A reactions for the MII Endpoint group were carried out on serial sections immediately adjacent to the equatorial plane of the same EAf. The equatorial section itself was used to document the oocyte nuclear stage (see Supplementary Datasheet 10 ). Likewise, CASP3 and SEMA3A reactions for the GV Endpoint group were performed on sections contiguous to the equatorial plane of the same EAf. Scale bar: 25 µm.

Article Snippet: Densitometric analysis for protein quantification was conducted using ImageLab software analyzer (ImageLab v. 6.1.0 Bio-Rad Laboratories, Milan, Italy).

Techniques: Expressing, Biomarker Discovery, Western Blot, Immunofluorescence, Software, Staining