Jacob (Yaqub) Hanna - يعقوب حنا
@jacob-hanna.bsky.social
Palestinian Stem Cell Scientist. Palestinian and LQBTQ+ rights advocate. #Arab&BlackLivesMatter #EndOccupation #FreePalestine #PeaceNOW #CeasefireNOW #BringThemHomeNow! Personal account.
EEE meeting is BACK! Early Embryogenesis & Epigenetics conference in Berlin 02/2026.
Checkout great program and over 12 slots for (not so) short talks for submitted abstracts!. Early registration now open -
w.molgen.mpg.de/embryo2026
Checkout great program and over 12 slots for (not so) short talks for submitted abstracts!. Early registration now open -
w.molgen.mpg.de/embryo2026
September 28, 2025 at 12:00 AM
EEE meeting is BACK! Early Embryogenesis & Epigenetics conference in Berlin 02/2026.
Checkout great program and over 12 slots for (not so) short talks for submitted abstracts!. Early registration now open -
w.molgen.mpg.de/embryo2026
Checkout great program and over 12 slots for (not so) short talks for submitted abstracts!. Early registration now open -
w.molgen.mpg.de/embryo2026
12/n last one (plenty more but u should get the point by now) - iEFCs=60h in Preprint are shown to have a mix of PrE/Epi/TE , while in Paper they are homogeneously labeled in Magenta?
August 12, 2025 at 6:45 AM
12/n last one (plenty more but u should get the point by now) - iEFCs=60h in Preprint are shown to have a mix of PrE/Epi/TE , while in Paper they are homogeneously labeled in Magenta?
11/n in relation to the above concern, another panel of Paper in Fig.3F 32C ICM are marked (left) and according to this analysis on dataset used they also highly overlap with 16-Cell labeled in green? yet again why drown in scRNA-seq when immunostainings r so easy to solve this ?
August 12, 2025 at 6:44 AM
11/n in relation to the above concern, another panel of Paper in Fig.3F 32C ICM are marked (left) and according to this analysis on dataset used they also highly overlap with 16-Cell labeled in green? yet again why drown in scRNA-seq when immunostainings r so easy to solve this ?
10/n some meta-analysis was done to claim iEFC fall on 8 cell. Upon close inspection - 8C and 32ICM r coloured similarly! and there is NO arrow pointing to 32ICM on the UMAP. it seems 32ICM cells are nicely overlapping with iEFCs? where is 64cell ICM? But again, simple scRNA-seq above says it all!
August 12, 2025 at 6:44 AM
10/n some meta-analysis was done to claim iEFC fall on 8 cell. Upon close inspection - 8C and 32ICM r coloured similarly! and there is NO arrow pointing to 32ICM on the UMAP. it seems 32ICM cells are nicely overlapping with iEFCs? where is 64cell ICM? But again, simple scRNA-seq above says it all!
9/n Straightforward scRNA-seq analysis of 60h=iEFC in Preprint also showed cells are not going backwards in development, but forward. Gata6+ cells express Sox17 and Pdgfra which are not 8-16cell markers but of good old E4.5 PrE cells.
August 12, 2025 at 6:43 AM
9/n Straightforward scRNA-seq analysis of 60h=iEFC in Preprint also showed cells are not going backwards in development, but forward. Gata6+ cells express Sox17 and Pdgfra which are not 8-16cell markers but of good old E4.5 PrE cells.
8/n Strangely straightforward scRNA-seq analysis of 60h=iEFC in Preprint showing rare CDX2/GATA6 coexpression & and that iEFC is basically a mix of Epi/PrE/TE, was NOT included in the published Paper although same dataset was used in both. & Nanog is reduced in TE-&PrE-like cells.
August 12, 2025 at 6:43 AM
8/n Strangely straightforward scRNA-seq analysis of 60h=iEFC in Preprint showing rare CDX2/GATA6 coexpression & and that iEFC is basically a mix of Epi/PrE/TE, was NOT included in the published Paper although same dataset was used in both. & Nanog is reduced in TE-&PrE-like cells.
7/n Preprint: 3/3 iEFC stainings show no/rare CDX2+/GATA6+ (include the same 2 above) and one in Preprint Fig1C, which was replaced in Paper Fig 1E with one that shows fuzzy coexpression. In sum 3 out of 4 immunostainings shown, rare CDX2/GATA6 co-expression!
August 12, 2025 at 6:42 AM
7/n Preprint: 3/3 iEFC stainings show no/rare CDX2+/GATA6+ (include the same 2 above) and one in Preprint Fig1C, which was replaced in Paper Fig 1E with one that shows fuzzy coexpression. In sum 3 out of 4 immunostainings shown, rare CDX2/GATA6 co-expression!
6/n why drown in intracellular stainings? Immunostaining for these markers r very easy to do in EFC=60h (names of same between Paper & Preprint tinyurl.com/tu6cv3cd). No quantitative & wideview stainings r shown. 2/3 iEFC stainings in Paper show no/rare CDX2+/GATA6+.
August 12, 2025 at 6:42 AM
6/n why drown in intracellular stainings? Immunostaining for these markers r very easy to do in EFC=60h (names of same between Paper & Preprint tinyurl.com/tu6cv3cd). No quantitative & wideview stainings r shown. 2/3 iEFC stainings in Paper show no/rare CDX2+/GATA6+.
3/n Intracellular FACS staining for TFs is a "dirty assay", each sample must have its OWN unstained and ISOTYPE MATCHED CONTROL to ensure true gating 4 positive staining. Wasn't done! & Preprint had GATING showing low%, was CHANGED in the Paper resulting in boost up to >85%!
August 12, 2025 at 5:52 AM
3/n Intracellular FACS staining for TFs is a "dirty assay", each sample must have its OWN unstained and ISOTYPE MATCHED CONTROL to ensure true gating 4 positive staining. Wasn't done! & Preprint had GATING showing low%, was CHANGED in the Paper resulting in boost up to >85%!
2/n 2/n Li et al. purported a concept of transiently capturing EFCs based on coexpression of OCT4/CDX2/GATA6 in 8-16cell embryos, after 60h of chemical treatment. Main proof shown is based on triple+ FACS intracellular staining for these genes (& NOT using triple reporter ESC)
August 12, 2025 at 5:51 AM
2/n 2/n Li et al. purported a concept of transiently capturing EFCs based on coexpression of OCT4/CDX2/GATA6 in 8-16cell embryos, after 60h of chemical treatment. Main proof shown is based on triple+ FACS intracellular staining for these genes (& NOT using triple reporter ESC)
end / Thanks to all Hanna lab members, great collaborators, funders and our lead Heroes from Turkey,
Gulben and Alperen (BSKY less) for the hard work despite of all the noise around.
Gulben and Alperen (BSKY less) for the hard work despite of all the noise around.
August 7, 2025 at 6:23 PM
end / Thanks to all Hanna lab members, great collaborators, funders and our lead Heroes from Turkey,
Gulben and Alperen (BSKY less) for the hard work despite of all the noise around.
Gulben and Alperen (BSKY less) for the hard work despite of all the noise around.
15/n But rather, naive PSCs have minimal plasticity in 2i/LIF shown by Brickman tinyurl.com/yhzrducz & our conditions boost amplify this propensity by using alternative signaling conditions. Naive PSCs are de facto totipotent and have "multiple personalities!" . "The Matrix" continues to inspire :)
August 7, 2025 at 6:17 PM
15/n But rather, naive PSCs have minimal plasticity in 2i/LIF shown by Brickman tinyurl.com/yhzrducz & our conditions boost amplify this propensity by using alternative signaling conditions. Naive PSCs are de facto totipotent and have "multiple personalities!" . "The Matrix" continues to inspire :)
14/n In summary, embryo founder-like cells r neither induced nor involved in TF-SEM formation. there is no evidence that mouse nPSCs need to first go to an earlier "embryo founder cells" & only then give rise to TE/PrE/Epi.
August 7, 2025 at 6:14 PM
14/n In summary, embryo founder-like cells r neither induced nor involved in TF-SEM formation. there is no evidence that mouse nPSCs need to first go to an earlier "embryo founder cells" & only then give rise to TE/PrE/Epi.
13/n AC-MEF cells were blastoid competent! and TF-SEM competent. They also lacked any 2C-like or 8-16 cell-morula-like embryo founder cells during expansion!
August 7, 2025 at 6:14 PM
13/n AC-MEF cells were blastoid competent! and TF-SEM competent. They also lacked any 2C-like or 8-16 cell-morula-like embryo founder cells during expansion!
12/n Naive ESCs in AC-MEF conditions generated advanced E8.5 TF-SEMs. So we developed a rapid induction regimen , and also along-term induction and maintenance based route to make mouse TF-SEMs.
August 7, 2025 at 6:13 PM
12/n Naive ESCs in AC-MEF conditions generated advanced E8.5 TF-SEMs. So we developed a rapid induction regimen , and also along-term induction and maintenance based route to make mouse TF-SEMs.
11/n My favorite result, is that we could MAINTAIN in this naive alternative condition (AC) media, OCT4/NANOG+ naive PSC colonies that contained OCT4+/Cdx2+ TE primed or OCT4/GATA6 PrE primed double positive cells that self renew for >30 passages. Triple+ cells were very rare!
August 7, 2025 at 6:13 PM
11/n My favorite result, is that we could MAINTAIN in this naive alternative condition (AC) media, OCT4/NANOG+ naive PSC colonies that contained OCT4+/Cdx2+ TE primed or OCT4/GATA6 PrE primed double positive cells that self renew for >30 passages. Triple+ cells were very rare!
10/n i never get bored from watching them change day by day
August 7, 2025 at 5:47 PM
10/n i never get bored from watching them change day by day
9/n We then used electronically controlled ex utero culture device & EUCM conditions we previously developed in Aguilera-Castrejon 2021 tinyurl.com/2nuddrnw & generated E8.5-E8.75 mouse TF-SEMs at enhanced efficiency, from more lines, from ESCs & iPSCs & with better quality & number of somites...
August 7, 2025 at 5:46 PM
9/n We then used electronically controlled ex utero culture device & EUCM conditions we previously developed in Aguilera-Castrejon 2021 tinyurl.com/2nuddrnw & generated E8.5-E8.75 mouse TF-SEMs at enhanced efficiency, from more lines, from ESCs & iPSCs & with better quality & number of somites...
8/n consistently given no emergence of new cell populations besides PrE and TE in the induction phase, defining 16cell-morula-like embryo founder master markers genes (tinyurl.com/4n23x9h9) were not induced .
August 7, 2025 at 5:41 PM
8/n consistently given no emergence of new cell populations besides PrE and TE in the induction phase, defining 16cell-morula-like embryo founder master markers genes (tinyurl.com/4n23x9h9) were not induced .
7/n Naive pluripotent cells just partially drift towards PrE and TE as can be easily seen below.
August 7, 2025 at 5:40 PM
7/n Naive pluripotent cells just partially drift towards PrE and TE as can be easily seen below.
6/n 4-day rapid induction & maturation regimen from naive mouse ESCs showed TF induction of TE & PrE that correspond to E3.5-E4.5. Pls note PrE & TE are the only newly induced populations over 4 day time-course!, & there are NO induced 8-16-morula-like embryo founder cells.
August 7, 2025 at 5:39 PM
6/n 4-day rapid induction & maturation regimen from naive mouse ESCs showed TF induction of TE & PrE that correspond to E3.5-E4.5. Pls note PrE & TE are the only newly induced populations over 4 day time-course!, & there are NO induced 8-16-morula-like embryo founder cells.
5/n we aimed for "confusing mixed signaling" that partially promote mouse TE & PrE, but retain some PSCs. We tested molecules reported to promote TE or PrE but also boosted iPSCs (alongside HDACi). Briefly, Deng chemical iPSC molecules were top hits (see Fig1 for screen details) tinyurl.com/3bsatkat
August 7, 2025 at 5:22 PM
5/n we aimed for "confusing mixed signaling" that partially promote mouse TE & PrE, but retain some PSCs. We tested molecules reported to promote TE or PrE but also boosted iPSCs (alongside HDACi). Briefly, Deng chemical iPSC molecules were top hits (see Fig1 for screen details) tinyurl.com/3bsatkat
4/n i)In Oldak et al. 2023 we found that enhancers of extra-embryonic master regulators were largely open in human nPSCs, but not mouse nPSCs, possibly explaining why human SEMs could be made W/O Tg. ii)Consistently, Chambers & co reported HDACi enables TF mouse TSCs tinyurl.com/27z9tuz4.
August 7, 2025 at 5:20 PM
4/n i)In Oldak et al. 2023 we found that enhancers of extra-embryonic master regulators were largely open in human nPSCs, but not mouse nPSCs, possibly explaining why human SEMs could be made W/O Tg. ii)Consistently, Chambers & co reported HDACi enables TF mouse TSCs tinyurl.com/27z9tuz4.
3/n c)Human blastoids were (Wu, Rivron..) made from naive PSCs by simultaneous induction of PrE & TE by CONFUSING the aggregate with a mix of cytokines/small molecules that promote PrE/TE yet maintain some nPSCs. Thus, we wanted to make mouse transgene free TF-SEMs & induce PrE/TE in the same mix?
August 7, 2025 at 5:00 PM
3/n c)Human blastoids were (Wu, Rivron..) made from naive PSCs by simultaneous induction of PrE & TE by CONFUSING the aggregate with a mix of cytokines/small molecules that promote PrE/TE yet maintain some nPSCs. Thus, we wanted to make mouse transgene free TF-SEMs & induce PrE/TE in the same mix?
2/n a)We previously made mouse SEMs solely from naive PSCs (nPSCs), by use of transgenes to separately induce TE & PrE (Tarazi et al. 2022, tinyurl.com/yc7d35tb) b)However, we could make human SEMs from naive PSCs (nPSCs) without transgenes (Oldak et al. 2023 tinyurl.com/yc623v48)?
August 7, 2025 at 4:58 PM
2/n a)We previously made mouse SEMs solely from naive PSCs (nPSCs), by use of transgenes to separately induce TE & PrE (Tarazi et al. 2022, tinyurl.com/yc7d35tb) b)However, we could make human SEMs from naive PSCs (nPSCs) without transgenes (Oldak et al. 2023 tinyurl.com/yc623v48)?