Wednesday, May 1, 2013
Correction to q22 Stem Cells Final Review Qs
Thanks Sydney for pointing out that Dr Brown did include mention of evidence of cancer stem cells in breast cancer -> on one of his slides, there is described an experiment of transplanting cells taken from existing breast cancer into a mammary pad and observing that it grows into another tumor.
Sunday, April 28, 2013
Stem Cells Final Review Q
First off, I've started putting down the lectures that you'd look at to figure out the answers. As I progress on my own last minute studying, I'll try and add bulletpoints of key things you probably could mention in your answers. Also if you have suggestions I'm happy to put them up too (comment below!)
https://docs.google.com/document/d/1IyCV-7EdsXEt_QiSA2cCjiJC_ckiD8yRuJ8W1t3KR38/edit?usp=sharing
Good luck!
https://docs.google.com/document/d/1IyCV-7EdsXEt_QiSA2cCjiJC_ckiD8yRuJ8W1t3KR38/edit?usp=sharing
Good luck!
Saturday, April 27, 2013
More Research Paper Sumups!
I will be focusing on the study guide from here on out. If I am done that early and have extra time, I will continue the research paper sum-ups. Until then, good luck with studying!
There's evidence that GDNF is important for regulating population size of spermatogonial stem cells.
What is the evidence? For a long term culture, you need GDNF to maintain and expand stem spermatogonia (how do you know they are stem spermatogonia? by putting them in a testis w/o germ cells and restoring spermatogenesis).
But wait, is GDNF just a survival factor or does it actually promote the self-renewing kind of replication for stem cells? Let's see. Mouse stem spermatogonia were deprived of GDNF for 3 or 6 days, but the population still grew as if it had GDNF. One would infer from this that GDNF does not promote self-renewal over differentiation. So that's the in vitro study.
What about in vivo? We are kinda limited to knocking out or messing with GDNF expression from time of birth or earlier, but not in adult tissue, nor can we try to restore it after knocking it out to see what happens. Overexpression of GDNF -> lots of undifferentiated spermatogonia that just died by apoptosis and led to infertile adult. Knockout of GDNF -> lost all germ cells in a week and the few left did not replicate.
Lots of questions, but how to answer them? We made a new system where we could inhibit GDNF and also reverse the inhibition. Let's look at the signaling pathway. GDNF binds Gfr-alpha1 which activates Ret and phosphorylates Ret (also part of the same receptor). Our transgenic mice had 1 mutation in Ret but it doesn't affect Ret activity. BUT it increases affinity for a competitive inhibitor NA-PP1. So by treating mice with NA-PP1, it competes with ATP and prevents the binding of GDNF from phosphorylating RET. When you take away NA-PP1, things go back to normal.
hypothesis: GDNF is essential for maintaining spermatogonial stem cell pool and also stimulates self-renewing replication of the stem cells. prediction if hypothesis is true: stem spermatogonia will be lost if GDNF signaling is inhibited for duration of 1 cell cycle (2 days).
Results:
the mutation in Ret works: baby mice with underdeveloped kidneys die with the mutation (proves mutation works), but it doesn't affect adults (proves it's usable for studying adult tissue)
ok, so treatment with NA-PP1 for 5, 11, 20 days and looking at Gfr-alpha1 positive cells and Zbtb16+ cells (these are regular spermatogonia, not stem spermatogonia). the Gfra1+ cells are less and less by 5 and 11 days and basically gone by 20. The Zbtb16+ aren't changed at 5 but are reduced by 11 and gone by 20 as well. This doesn't happen in mice w/o the Ret mutation. similar results seen with mRNA levels of Ret, Gfra1, and Zbtb16.
Still, after 20 days, those transcripts are still detectable (just reduced). Did we lose functional stem cells or did we just reduce them to numbers so low they're hardly detectable? Let's do 30 days treatment and collect the tissue either then or 35 days later. Histology sections show Ret-mutated mice lost ALL their spermatogonial stem cells from 30 days of treatment with NA-PP1. Further testing showed this happened over a course of several days (not all at once)
After the 11 day treatment though, some still remain! 35 days after the 11 day NA-PP1 trtmt, testes were back to normal. What? We saw 3% of tubules active in spermatogenesis and there were dense clusters of Gfra1+ cells. We also saw that even with a short 2 day NA-PP1 trtmt, some stem spermatogonia are gone.
Discussion:
This is the first use of a chemical-genetic approach to study adult stem cell regulation via growth factor. This method has 3 advantages:
1) you start with totally normal pool of stem cells (existing in normal organism, not in weird imitation culture, etc)
2) you get normal in vivo response to whatever you are doing
3) you can reverse what you just did and go back to normal conditions
We showed GDNF is required for maintaining stem cells in adult testis. Also we saw stem cell loss with absence of GDNF, which is opposite to the stem cell growth mentioned earlier in that in vitro study. But some survive even past 11 days, so there are definitely other factors involved.
This may be due to a) varying normal concentrations of GDNF for various stages of the tubules, b) the cells normally do not replicate at the same time all together, c) it has been hypothesized that there are quiescent ones also who enter the cell cycle only when tissue damage has occurred.
When NA-PP1 treatment ceases, cells migrate and refill empty areas, and new refilled niches will start producing spermatogonia again with time.
A previous study isolated cells from new baby rat testes that could divide w/o differentiating or divide and produce testosterone (normal Leydig cell function). And when they were put into normal testis, the cells also could differentiate -> evidence that these baby rat cells were Leydig stem cells.
This study looks for stem Leydig cells in the adult testis. Since it is possible to separate seminiferous tubules and interstitial compartment of a testis, it made it feasible to go looking for these stem cells and their physical niche.
Results:
Testicular cells were isolated from Leydig-depleted testis and grown in culture. Without LH (luteinizing hormone), cells kept growing and growing for about 1.5 yr (325 population doublings). They didn't produce testosterone. When cultured with LH, they produced testosterone.
Seminiferous tubules and interstitial compartment were separated from Leydig-depleted testis and cultured separately. Cells appeared on surface of cultured tubules but not cultured interstitial compartment. These cells stained for 3B-HSD (enzyme in pathway of making testosterone) and the tubule culture, when treated with LH, started producing testosterone (neither tubule culture w/o LH nor interstitial culture w/ or w/o LH made testosterone). Therefore, the Leydig precursor cells came from the tubules.
Are they stem cells? These cells were selectively labeled with EdU, and then given EDS, which stripped the 3B-HSD plus label cells. Some time later, they came back. If you measured testosterone, its presence and absence correlated with the presence/absence of the 3B-HSD cells.
Discussion:
Had been known previously that Leydig cells came back after depletion and it was the point of this study to see if there were stem cells ( as opposed to quiescent progenitor cells ) and where they are and how they are regulated.
If they were stem cells, we should have been able to isolate them and have them grow for a long time without trying to repress any Leydig cell markers and be able to differentiate into T-producing cells. We accomplished this.
There have not been any Leydig stem cell markers identified, so we had to go a different way about locating them. Our results with the separation of tissue in culture experiment demonstrated they were the cells growing on the surface of the tubules.
If they were stem cells, they should be able to come back again if they suffered another depletion by EDS. The EDS in the tissue culture experiment showed that. Also we had an unpublished experiment where we treated the tubules with collagenase and dispase to cut anything attached to surface of tubules off, and any stem-activity was lost (couldn't make cells that produce T).
In terms of regulation, more research is needed. But considering that they could grow on tubule without the interstitial compartment present suggests the blood vessels associated with interstitial compartment are definitely not important, but they might still have a function in vivo.
Also the tubule culture method is a great way to keep answering questions about niche components in regulating adult stem cells.
Savitt J, Singh D, Zhang C, Chen L, Folmer J, Shokat K, and W Wright. "The In Vivo Response of Stem and Other Undifferentiated Spermatogonia to the Reversible Inhibition of GDNF Signaling in the Adult." Stem Cells 4, 2012: 732-740.
Intro:There's evidence that GDNF is important for regulating population size of spermatogonial stem cells.
What is the evidence? For a long term culture, you need GDNF to maintain and expand stem spermatogonia (how do you know they are stem spermatogonia? by putting them in a testis w/o germ cells and restoring spermatogenesis).
But wait, is GDNF just a survival factor or does it actually promote the self-renewing kind of replication for stem cells? Let's see. Mouse stem spermatogonia were deprived of GDNF for 3 or 6 days, but the population still grew as if it had GDNF. One would infer from this that GDNF does not promote self-renewal over differentiation. So that's the in vitro study.
What about in vivo? We are kinda limited to knocking out or messing with GDNF expression from time of birth or earlier, but not in adult tissue, nor can we try to restore it after knocking it out to see what happens. Overexpression of GDNF -> lots of undifferentiated spermatogonia that just died by apoptosis and led to infertile adult. Knockout of GDNF -> lost all germ cells in a week and the few left did not replicate.
Lots of questions, but how to answer them? We made a new system where we could inhibit GDNF and also reverse the inhibition. Let's look at the signaling pathway. GDNF binds Gfr-alpha1 which activates Ret and phosphorylates Ret (also part of the same receptor). Our transgenic mice had 1 mutation in Ret but it doesn't affect Ret activity. BUT it increases affinity for a competitive inhibitor NA-PP1. So by treating mice with NA-PP1, it competes with ATP and prevents the binding of GDNF from phosphorylating RET. When you take away NA-PP1, things go back to normal.
hypothesis: GDNF is essential for maintaining spermatogonial stem cell pool and also stimulates self-renewing replication of the stem cells. prediction if hypothesis is true: stem spermatogonia will be lost if GDNF signaling is inhibited for duration of 1 cell cycle (2 days).
Results:
the mutation in Ret works: baby mice with underdeveloped kidneys die with the mutation (proves mutation works), but it doesn't affect adults (proves it's usable for studying adult tissue)
ok, so treatment with NA-PP1 for 5, 11, 20 days and looking at Gfr-alpha1 positive cells and Zbtb16+ cells (these are regular spermatogonia, not stem spermatogonia). the Gfra1+ cells are less and less by 5 and 11 days and basically gone by 20. The Zbtb16+ aren't changed at 5 but are reduced by 11 and gone by 20 as well. This doesn't happen in mice w/o the Ret mutation. similar results seen with mRNA levels of Ret, Gfra1, and Zbtb16.
Still, after 20 days, those transcripts are still detectable (just reduced). Did we lose functional stem cells or did we just reduce them to numbers so low they're hardly detectable? Let's do 30 days treatment and collect the tissue either then or 35 days later. Histology sections show Ret-mutated mice lost ALL their spermatogonial stem cells from 30 days of treatment with NA-PP1. Further testing showed this happened over a course of several days (not all at once)
After the 11 day treatment though, some still remain! 35 days after the 11 day NA-PP1 trtmt, testes were back to normal. What? We saw 3% of tubules active in spermatogenesis and there were dense clusters of Gfra1+ cells. We also saw that even with a short 2 day NA-PP1 trtmt, some stem spermatogonia are gone.
Discussion:
This is the first use of a chemical-genetic approach to study adult stem cell regulation via growth factor. This method has 3 advantages:
1) you start with totally normal pool of stem cells (existing in normal organism, not in weird imitation culture, etc)
2) you get normal in vivo response to whatever you are doing
3) you can reverse what you just did and go back to normal conditions
We showed GDNF is required for maintaining stem cells in adult testis. Also we saw stem cell loss with absence of GDNF, which is opposite to the stem cell growth mentioned earlier in that in vitro study. But some survive even past 11 days, so there are definitely other factors involved.
This may be due to a) varying normal concentrations of GDNF for various stages of the tubules, b) the cells normally do not replicate at the same time all together, c) it has been hypothesized that there are quiescent ones also who enter the cell cycle only when tissue damage has occurred.
When NA-PP1 treatment ceases, cells migrate and refill empty areas, and new refilled niches will start producing spermatogonia again with time.
Stanley E, Lin C, Jin S, Liu J, Sottas C M, Ge R, Zirkin B, and H Chen. "Identification, Proliferation, and Differentiation of Adult Leydig Stem Cells." Endocrinology 153, 2012: 5002-5010
When testes get depleted of their Leydig cells (testes interstitial compartment) with treatment of EDS, they come right back! There must be cells that make Leydig cells then, but very little is known about them.A previous study isolated cells from new baby rat testes that could divide w/o differentiating or divide and produce testosterone (normal Leydig cell function). And when they were put into normal testis, the cells also could differentiate -> evidence that these baby rat cells were Leydig stem cells.
This study looks for stem Leydig cells in the adult testis. Since it is possible to separate seminiferous tubules and interstitial compartment of a testis, it made it feasible to go looking for these stem cells and their physical niche.
Results:
Testicular cells were isolated from Leydig-depleted testis and grown in culture. Without LH (luteinizing hormone), cells kept growing and growing for about 1.5 yr (325 population doublings). They didn't produce testosterone. When cultured with LH, they produced testosterone.
Seminiferous tubules and interstitial compartment were separated from Leydig-depleted testis and cultured separately. Cells appeared on surface of cultured tubules but not cultured interstitial compartment. These cells stained for 3B-HSD (enzyme in pathway of making testosterone) and the tubule culture, when treated with LH, started producing testosterone (neither tubule culture w/o LH nor interstitial culture w/ or w/o LH made testosterone). Therefore, the Leydig precursor cells came from the tubules.
Are they stem cells? These cells were selectively labeled with EdU, and then given EDS, which stripped the 3B-HSD plus label cells. Some time later, they came back. If you measured testosterone, its presence and absence correlated with the presence/absence of the 3B-HSD cells.
Discussion:
Had been known previously that Leydig cells came back after depletion and it was the point of this study to see if there were stem cells ( as opposed to quiescent progenitor cells ) and where they are and how they are regulated.
If they were stem cells, we should have been able to isolate them and have them grow for a long time without trying to repress any Leydig cell markers and be able to differentiate into T-producing cells. We accomplished this.
There have not been any Leydig stem cell markers identified, so we had to go a different way about locating them. Our results with the separation of tissue in culture experiment demonstrated they were the cells growing on the surface of the tubules.
If they were stem cells, they should be able to come back again if they suffered another depletion by EDS. The EDS in the tissue culture experiment showed that. Also we had an unpublished experiment where we treated the tubules with collagenase and dispase to cut anything attached to surface of tubules off, and any stem-activity was lost (couldn't make cells that produce T).
In terms of regulation, more research is needed. But considering that they could grow on tubule without the interstitial compartment present suggests the blood vessels associated with interstitial compartment are definitely not important, but they might still have a function in vivo.
Also the tubule culture method is a great way to keep answering questions about niche components in regulating adult stem cells.
Two more paper sumups: Germline Stem Cells
Eun S H, Gan Q, and X Chen. "Epigenetic Regulation of Germ Cell Differentiation." COCeBi 22, 2010: 1-7.
Epigenetics modify chromatin w/o affecting DNA sequence. There are proteins who "write" on the modifications, "read" and act on the modifications, or "erase" the modifications.
Germ cells are "immortal" b/c they produce the next generation. Epigenetics in germ cells might be involved in
1) meiosis and terminal differentiation
2) keeping the genome information good
3) erasing bad information
GSCs (germline stem cells) initiate gametogenesis. Their chromatin structure maintains self-renewing and blocks differentiation. Evidence: ISWI "Imitation Switch" is a chromatin remodeling factor. it is essential for preventing differentiation (in mutants for iswi, the bag gene is expressed and induces differentiation). One example is the NURF complex that contains the ISWI factor in male drosophila.
histone modifications also functions in maintaining stem-cellness. Evidence: scny gene removes ubiquitin from histone H2B. scny mutant leads to super acetylated H3 and super ubiquitinated U2B => open or loose chromatin -> active transcription of differentiation genes.
also, this chromatin landscape isn't permanently blocked from differentiation but described as "poised" to start differentiation upon stimuli. in mammalian: have both activating H3K4methyl3 and repressive H3K27methyl3 going on and a paused RNA Pol II. it prevents DNA methylation, which is a more stable block and would be difficult to remove for differentiation to happen at a snap. in drosophila: not bivalent (i.e. not an active and a repressive histone modification at the same time) and not associated with paused RNA Pol II, but probably b/c drosophila doesn't have methylase anyway.
only germ cells do meiosis. with meiosis, chromatin regulators and opposite-fxn histone modifying enzymes are downregulated. at same time, celltype specific factors are unregulated (i.e. the tTAFs testis-specific TBP-associated facotrs in drosophila spermatocytes)
same/similar-fxn histone modifying enzymes work together though, maybe separated by time (i.e. in drosophila ovary, one isoform is in early germ cells and GSCs while the similar isoform is needed in later stage germ cells). but in mice, meiosis was found to have both functional enzymes that are opposite fxn.
also during mammalian gametogenesis is when imprinting (DNA methylation) happens. This might happen via a sort of weird transcription process that goes over modified genes in order to "read" and match it to the new DNA strands. This happens while oocytes are arrested in meiosis but we're not sure about spermatocytes. More research needs to be done.
in spermiogenesis in drosophila and mammals, histones are kicked out and replace with Tnps and Prms to super condense the DNA in super nuclei. This process is also regulated by epigenetics. Some examples include:
1) H4 S1A substitution/mutation messes up phosphorylation of H4S1 and results in failed sporulation in yeast
2) demethylation of H3K9me2/1 is necessary to increase expression of the Tnps and Prms in mice.
3) (see more in the paragraph)
in human sperm, about 4% of the genome still has regular histone nucleosomes, no Tnps or Prms or anything. Specially, the Hox genes and imprinted genes that are necessary for early embryonic development. but things like Nanog that are necessary for ESC pluripotency are off.
after fertilization, Prms get kicked out and given maternal histones. some chromatin regulating proteins are involved in helping out. the cells that will be the germ cells for this zygote's babies are the PGCs (primordial germ cells). They get a genome-wide erasure of DNA methylation so as to get rid of any epimutations that happened between their ancestor's PGCs and this fertilization.
Proper histone modification control takes care of differentiation genes and fertility, but they also have been observed to ensure proper lifespan.
During the journey of germ cells, they need to maintain a correct epigenome, which is ensured by dynamic control, erasure, and re-establishment of the epigenome. More needs to be done in the field, except it is hard to get a good amount of these GCs and PGCs (ethics!) so if we can has new technology that don't need as many cells, we will be good to go. Also this will be useful for diseases due to germ cell differentiation fail (infertility, germ cell tumors, etc) but also regenerative medicine.
Tran V, Lim C, Xie J, and X Chen. "Asymmetric Division of Drosophila Male Germline Stem Cell Shows Asymmetric Histone Distribution." Science 338: 2012: 679-682.
question: stem cell activity is regulated by epigenetic, but do stem cells retain their epigenetic info?
male drosophila GSCs have been well characterizes and are easily identified. they divide asymmetrically and can be examined at the single cell level.
histones are major carriers of the epigenome. we created this switchable dual-color method to label and identify what is a new histone versus and old histone. can be controlled spatially or temporally (by heat shock!). labeled histones can get switched from green to red.
how? heat shock causes the recombinant gene to irreversibly shut down GFP-labeled old histone expression and initiate expression of new mKO-labeled histones. if old histones are distributed btw daughter cells equally, over time any green color will be replaced by red color. if old histones are kept in the GSC and new histones are kept in the daughter fated cell, then green will be specifically retained.
before heat shock: testes with transgenes showed green nucleus but almost no red. after heat shock, red started appearing. of all GSCs, about 70% of them will be in G2 phase, 21% in s phase, and <2% in mitosis. entire cycle length is 16 hrs. plus the two daughters will be connected after incomplete cytokinesis and are called "spectrosomes." examine testes 16-20 hrs after heat shock, specifically the spectrosomes.
results: green preferentially held in daughter GSC, while red was preferentially delivered to fated daughter cell. in contrast, for symmetrically dividing GSCs, both daughters had equal red and green. also in contrast, this was observed with canonical H3 transgene, but not with variant H3.3 transgene. also looking at during mitosis, anaphase pulled green chromatids toward future daughter GSC while the spindle pulled red chromatids towards future fated daughter cell.
change it up: ectopic activation of JAK-STAT signaling pathway. previously shown that this causes overpopulation of GSCs. they did it and found that the asymmetric green-red distribution didn't happen. so equal distribution of old and new histones is also associated with this forced symmetrical divisions.
proposed mechanism: old and new histones are incorporated into separate chromatids during S, and are preferentially segregated at the spindle.
this may go towards understanding how epigenetic is maintained in SCs and reset in sibling cells that will differentiate.
Prepping for Final Exam of Stem Cells Biology
Ok, so the doc has been updated with ALL the lectures of this semester now, hopefully typo free. Again, comment or email me if you have questions or something looks funny.
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
Sumups of the other research papers (16 more to do, homg) and a study guide for the exam coming either later today or tomorrow.
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
Sumups of the other research papers (16 more to do, homg) and a study guide for the exam coming either later today or tomorrow.
Thursday, April 4, 2013
Audio of Stem Cell Lecture 7, Lecture Transcript updates
Here's the audio for lecture 7 with Dean Gregory Ball. I will be discontinuing the audio recordings b/c I simply don't have the equipment for good recordings (I did Ball b/c he has such a booming voice anyway).
https://dl.dropbox.com/u/22080433/balllecture.mp3
But, also I have updated the lecture transcript with Lecture 7 and 8, Dean Ball and Dr. Alan Meeker.
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
https://dl.dropbox.com/u/22080433/balllecture.mp3
But, also I have updated the lecture transcript with Lecture 7 and 8, Dean Ball and Dr. Alan Meeker.
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
Sunday, March 17, 2013
Audio of Stem Cell Lecture 6 and Reminder Link of LectureTranscripts
Well, here's audio lecture of Number 6, Dr. Daniella Drummond-Barbosa (JHSPH) on Stem Cells in Drosophila Ovary
https://dl.dropbox.com/u/22080433/march13lecture.mp3
and the link again for the lecture transcripts:
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
https://dl.dropbox.com/u/22080433/march13lecture.mp3
and the link again for the lecture transcripts:
https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
Wednesday, February 27, 2013
Audio Recording of Stem Cell Lecture (attempt 2)
So here I have attempted to simplify by just taking an audio recording of the lecture by Dr. Barry Zirkin (this time on Leydig stem cells). I have to forewarn you that my computer's Audacity program also succinctly captured the sound of my typing (as I transcribe the lecture as well). Apologies if that annoys you :(. Still, it captures the lecture all the same.
https://dl.dropbox.com/u/22080433/StemCell2013Lecture5.mp3
Also I still update the transcribed lectures on the google doc (see link some posts earlier) and in two weeks, we have our first exam, so in one week, look out for some study guides! :D
https://dl.dropbox.com/u/22080433/StemCell2013Lecture5.mp3
Also I still update the transcribed lectures on the google doc (see link some posts earlier) and in two weeks, we have our first exam, so in one week, look out for some study guides! :D
Lecture 2 Paper Sumup: induced Pluripotent Stem Cells
Takahashi K and Yamanaka S. "Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors." Cell 126, 2006: 663-676.
Introduction:
ES cells are pluripotent: they come from mammalian embryo and can make cells from all 3 germ layers.
- super useful for treating diseases
- but ethically controversial (can we use human embryos for this purpose?)
- and physically difficult if implanted tissue is rejected by host
possible solution? get pluripotent cells from host themselves!
- take normal body cell (somatic cell) DNA and stick it in an oocyte or fuse the cell with an ES cell
- the other cell contents of ES cells/oocyte have been shown to contain some factors that make somatic cells pluripotent!
- we know some factors that maintain pluripotency, but maybe these factors also INDUCE pluripotency
what do we know about factors that maintain pluripotency:
- Oct3/4, Nanog handle maintenance
- Stat3, E-Ras, c-myc, Klf4, Beta-catenin are highly expressed in tumors--they also help maintain long term ES cell phenotype and proliferation
Results:
We tried out 24 genes that could have been the factors that induce pluripotency.
- experiment: if gene X induces pluripotency, cell will be resistant to G418 (a molecule that inhibits protein synthesis).
- ergo, if we see cell resistant to G418, then the gene that we unregulated in the cell is a factor that induces pluripotency.
step 1: in mouse, knock out the gene Fbx15. This gene is super important for maintaining pluripotency in mouse development.
- ES cell with knocked out Fbx15 resist G418
- somatic cells with knocked out Fbx15 cannot resist G418
step 2: one by one, insert each of the 24 candidate genes into these knockout embryonic mice cells
- no resistance observed
- ergo, these genes cannot induce pluripotency by themselves
step 3: upregulate all 24 genes in these knockout cells
- get lots of resistant colonies!
- some of these look very similar to ES cells (morphology, proliferation traits, gene expression markers, etc)
step 4: upregulate all but 1 gene in these knockout cells
- found 10 genes that, once you did NOT upregulate them in the cells, you did not get resistant colonies
- ergo, these are super important factors that you can't NOT put in in order to induce pluripotency
step 5: upregulate all 10 special genes in knockout cells
- get lots of resistant colonies!
- many of these look similar to ES cells
step 6: upregulate all except 1 of these 10 special genes in knockout cells
- not including either Oct3/4 or Klf4 resulted in no resistant colonies
- not including Sox2 resulted in very very few resistant colonies
- not including c-myc resulted in weird looking resistant colonies
- not including any of the others produced all resistant colonies, so the others were not as important as the above 4.
step 7: upregulate only those 4 super special genes in knockout cells
- get same result as step 5
- culture and confirm that these are iPSC (induced pluripotent stem cells)
step 8: upregulate pairs and triplets of these 4 super special genes in knockout cells
- no two of them could form any resistant colonies
- 2 triplets produced a few colonies but they did not survive further culturing
- 2 other triplets produced more colonies but looked weird (different from ES or previously determined iPSC)
step 9: do gene expression analysis of iPSCs induced with the various combos
- the 4 combo and the 10 combo cells, both are similar to ES cell expression profiles but not exactly the same
- the 3 combo cells were very very different
step 10: try to make teratomas with these various combo-formed iPSCs
- there was inconsistent data: some of the 10 combos and the 4 combos made teratomas with cell types of all 3 layers, but some of the same combos could only form 2 or 1 of the germ layers
- so conclude the majority of the 10-combo and 4-combo cells are pluripotent, but not all
- tumors from 3-combo cells did not differentiate = not pluripotent!
- similar results when trying to form embryoid bodies in culture as opposed to forming teratomas in vivo.
step 11: introduce the 4 combo gene into mouse tail fibroblasts (somatic cells) and then inject these cells into blastocyst
- were able to observe that these injected cells helped form some of the germ layers and baby mice were actually born from these blastocysts that received injections!
step 12: compare gene expression levels of these 4 factors with protein expression levels btw iPSC and ES
- saw that while some of these genes were higher or lower in iPSC cells than in normal ES cells, the protein levels (Western blot!) were about the same!
step 13: try to grow iPSC without them differentiating in culture
- they always differentiated unless they were provided "feeder cells" in the same culture
Discussion:
- Oct3/4, Sox2, Nanog are essential for maintaining pluripotency
- Oct 3/4 and Sox 2 are essential for MAKING iPSCs
- Nanog is not important for that
- c-Myc Klf4 are also essential
- c-Myc upregulates genes for proliferation and transformation
- it affects some histone modifying enzymes (histone acetyltransferase, for example)
- there are a LOT (upt to 25000) of sites for c-Myc binding in mammal genome
- this is way more than what we'd guess for Oct3/4 or Sox2 binding sites
- it could be that c-Myc causes global histone acetylation, causing a lot of the genome to open up, so that Oct3/4 and Sox2 can find all their target binding sites
- what about Klf4? represses p53
- okay, what does p53 do? It suppresses Nanog during differentiation
- so if you repress p53, you enable Nanog, which should normally NOT be active for differentiation.
- this might contribute to making the iPSC or at least ES-like cell phenotype
- Klf4 activates p21 which suppresses proliferation, and c-Myc suppresses p21. This opposites relationship of c-Myc and Klf4 might be important (in other words, we're just guessing)
- one important question: which cells of the tissue given these four factors are becoming iPSCs?
- only a small portion of cells treated with the 4 factors become iPSCs
- maybe it's the progenitor/stem cells that already exist in tissue that are kinda multipotent but not pluripotent that transform into pluripotent cells
- the frequency doesn't change when we try this out with bone marrow, which should have a high percentage of progenitor/stem cells to begin with to change
- so it can't be those cells..
- maybe getting the right expression level of each factor in the cells is important
- experimental evidence: just a 50% increase or decrease in Oct3/4 proteins in an ES cell causes it to differentiate and lose pluripotency
- we know our iPSC clones overexpress RNA levels but their protein levels of the 4 factors are just right
- but these cells must be able to regulate that, b/c high high levels are necessary to become ES-cells but in order to stay ES-like, too much of the 4 factors is badddd
- they might need some chromosomal alterations too to stay ES-like
- this may be spontaneous or induced by some of the 4 factors
- where the retrovirus brings in the transgenes to overexpress the 4 factors also matters: could have impacted the expression of any native genes depending on how the transgene got shoved into the genome
- another question: are these 4 factors also important when we're trying to reprogram somatic cells by fusion with ES cells or plucking out nuclei and putting them in oocytes?
- the precise roles of Klf4 and c-Myc are also confusing and vague. they aren't essential for mouse development before the egg implants. c-Myc isn't detectable in oocytes at all. Hmm??
- well, related proteins, L-myc and Klf17 and Klf7 do exist. maybe Klf4 and C-myc's real properties are being supplanted by these relatives in wildtype development
- some other questions that this paper brings up...
- still unsure if these 4 factors can make pluripotent cells out of human somatic cells.
- testing/experimental process is going to require super specific culture environments
- but this is all really cool in the search for the tools to control pluripotency, and one day we might be able to make pluripotent cells from a patient's somatic cells.
Introduction:
ES cells are pluripotent: they come from mammalian embryo and can make cells from all 3 germ layers.
- super useful for treating diseases
- but ethically controversial (can we use human embryos for this purpose?)
- and physically difficult if implanted tissue is rejected by host
possible solution? get pluripotent cells from host themselves!
- take normal body cell (somatic cell) DNA and stick it in an oocyte or fuse the cell with an ES cell
- the other cell contents of ES cells/oocyte have been shown to contain some factors that make somatic cells pluripotent!
- we know some factors that maintain pluripotency, but maybe these factors also INDUCE pluripotency
what do we know about factors that maintain pluripotency:
- Oct3/4, Nanog handle maintenance
- Stat3, E-Ras, c-myc, Klf4, Beta-catenin are highly expressed in tumors--they also help maintain long term ES cell phenotype and proliferation
Results:
We tried out 24 genes that could have been the factors that induce pluripotency.
- experiment: if gene X induces pluripotency, cell will be resistant to G418 (a molecule that inhibits protein synthesis).
- ergo, if we see cell resistant to G418, then the gene that we unregulated in the cell is a factor that induces pluripotency.
step 1: in mouse, knock out the gene Fbx15. This gene is super important for maintaining pluripotency in mouse development.
- ES cell with knocked out Fbx15 resist G418
- somatic cells with knocked out Fbx15 cannot resist G418
step 2: one by one, insert each of the 24 candidate genes into these knockout embryonic mice cells
- no resistance observed
- ergo, these genes cannot induce pluripotency by themselves
step 3: upregulate all 24 genes in these knockout cells
- get lots of resistant colonies!
- some of these look very similar to ES cells (morphology, proliferation traits, gene expression markers, etc)
step 4: upregulate all but 1 gene in these knockout cells
- found 10 genes that, once you did NOT upregulate them in the cells, you did not get resistant colonies
- ergo, these are super important factors that you can't NOT put in in order to induce pluripotency
step 5: upregulate all 10 special genes in knockout cells
- get lots of resistant colonies!
- many of these look similar to ES cells
step 6: upregulate all except 1 of these 10 special genes in knockout cells
- not including either Oct3/4 or Klf4 resulted in no resistant colonies
- not including Sox2 resulted in very very few resistant colonies
- not including c-myc resulted in weird looking resistant colonies
- not including any of the others produced all resistant colonies, so the others were not as important as the above 4.
step 7: upregulate only those 4 super special genes in knockout cells
- get same result as step 5
- culture and confirm that these are iPSC (induced pluripotent stem cells)
step 8: upregulate pairs and triplets of these 4 super special genes in knockout cells
- no two of them could form any resistant colonies
- 2 triplets produced a few colonies but they did not survive further culturing
- 2 other triplets produced more colonies but looked weird (different from ES or previously determined iPSC)
step 9: do gene expression analysis of iPSCs induced with the various combos
- the 4 combo and the 10 combo cells, both are similar to ES cell expression profiles but not exactly the same
- the 3 combo cells were very very different
step 10: try to make teratomas with these various combo-formed iPSCs
- there was inconsistent data: some of the 10 combos and the 4 combos made teratomas with cell types of all 3 layers, but some of the same combos could only form 2 or 1 of the germ layers
- so conclude the majority of the 10-combo and 4-combo cells are pluripotent, but not all
- tumors from 3-combo cells did not differentiate = not pluripotent!
- similar results when trying to form embryoid bodies in culture as opposed to forming teratomas in vivo.
step 11: introduce the 4 combo gene into mouse tail fibroblasts (somatic cells) and then inject these cells into blastocyst
- were able to observe that these injected cells helped form some of the germ layers and baby mice were actually born from these blastocysts that received injections!
step 12: compare gene expression levels of these 4 factors with protein expression levels btw iPSC and ES
- saw that while some of these genes were higher or lower in iPSC cells than in normal ES cells, the protein levels (Western blot!) were about the same!
step 13: try to grow iPSC without them differentiating in culture
- they always differentiated unless they were provided "feeder cells" in the same culture
Discussion:
- Oct3/4, Sox2, Nanog are essential for maintaining pluripotency
- Oct 3/4 and Sox 2 are essential for MAKING iPSCs
- Nanog is not important for that
- c-Myc Klf4 are also essential
- c-Myc upregulates genes for proliferation and transformation
- it affects some histone modifying enzymes (histone acetyltransferase, for example)
- there are a LOT (upt to 25000) of sites for c-Myc binding in mammal genome
- this is way more than what we'd guess for Oct3/4 or Sox2 binding sites
- it could be that c-Myc causes global histone acetylation, causing a lot of the genome to open up, so that Oct3/4 and Sox2 can find all their target binding sites
- what about Klf4? represses p53
- okay, what does p53 do? It suppresses Nanog during differentiation
- so if you repress p53, you enable Nanog, which should normally NOT be active for differentiation.
- this might contribute to making the iPSC or at least ES-like cell phenotype
- Klf4 activates p21 which suppresses proliferation, and c-Myc suppresses p21. This opposites relationship of c-Myc and Klf4 might be important (in other words, we're just guessing)
- one important question: which cells of the tissue given these four factors are becoming iPSCs?
- only a small portion of cells treated with the 4 factors become iPSCs
- maybe it's the progenitor/stem cells that already exist in tissue that are kinda multipotent but not pluripotent that transform into pluripotent cells
- the frequency doesn't change when we try this out with bone marrow, which should have a high percentage of progenitor/stem cells to begin with to change
- so it can't be those cells..
- maybe getting the right expression level of each factor in the cells is important
- experimental evidence: just a 50% increase or decrease in Oct3/4 proteins in an ES cell causes it to differentiate and lose pluripotency
- we know our iPSC clones overexpress RNA levels but their protein levels of the 4 factors are just right
- but these cells must be able to regulate that, b/c high high levels are necessary to become ES-cells but in order to stay ES-like, too much of the 4 factors is badddd
- they might need some chromosomal alterations too to stay ES-like
- this may be spontaneous or induced by some of the 4 factors
- where the retrovirus brings in the transgenes to overexpress the 4 factors also matters: could have impacted the expression of any native genes depending on how the transgene got shoved into the genome
- another question: are these 4 factors also important when we're trying to reprogram somatic cells by fusion with ES cells or plucking out nuclei and putting them in oocytes?
- the precise roles of Klf4 and c-Myc are also confusing and vague. they aren't essential for mouse development before the egg implants. c-Myc isn't detectable in oocytes at all. Hmm??
- well, related proteins, L-myc and Klf17 and Klf7 do exist. maybe Klf4 and C-myc's real properties are being supplanted by these relatives in wildtype development
- some other questions that this paper brings up...
- still unsure if these 4 factors can make pluripotent cells out of human somatic cells.
- testing/experimental process is going to require super specific culture environments
- but this is all really cool in the search for the tools to control pluripotency, and one day we might be able to make pluripotent cells from a patient's somatic cells.
Sunday, February 24, 2013
Stem Cells: Lecture 4 Video
Here is my first attempt at recording. I did it less for the purpose of showing slides (because we get that provided at our school) and more for the purpose of audio learners, who don't do well just reading the transcripts of the lectures. I might just try an audio recording instead of video and audio next time.
This is the lecture by Dr. William Wright on the subject of stem spermatogonia.
https://www.youtube.com/watch?v=br7STDDbx24
This is the lecture by Dr. William Wright on the subject of stem spermatogonia.
https://www.youtube.com/watch?v=br7STDDbx24
Thursday, February 21, 2013
Papers, videos, and transcripts
Hello my Stem Cell Biology people!
First of all, remember to keep checking the google doc of the lecture transcripts. I always update it ASAP and I've been trying to correct my spelling mistakes and add in a few more explanatory words in case I'm not sure the lecturer was clear on something.
Also I'm experimenting with recording the lectures, b/c for some, they are better audio learners. Bear with me as I struggle with technology.
Finally, we have one or two papers assigned for every lecture, so I'll post a sum-up of the papers. I still recommend you read the papers and then read the sum-up, because 1) you need practice if you ever want to be able to digest the condensed jargon of science writing and 2) the more you read the material over and over again the more it sticks in your head.
Here it is from Lecture 1 (Please excuse my casual language. It is my opinion that to make science available to the general, you must speak "layman"...seriously though. I am not trying to condescend to anyone, I just want to make sure people can understand, because that's the foundation to building up knowledge later on.)
Also remember that every paper wants to tell you something, but it is always up to you if you choose to follow or take it for its word. Always ask questions (is this right? did they do this correctly? did they consider this? etc etc) Whatever I write below is not an expression of something I believe in, but just a paraphrase of what I interpreted the paper was trying to say. I'm always open to discussion! (hence, comment box)
Spradling, Allan C. "The living-tissue microscope: the importance of studying stem cells in their natural, undisturbed microenvironment." J Pathol 2011; 225: 161-162.
We need tools to look at cells in their daily lives without disturbing them.
Why? Because we would find out all about normal cell lives, diseased cell lives, new cell to cell and cell to gene and gene to gene pathways that we can't see from frozen or dead cells.
We have the tools to do this:
Small multicell structures with labels can be seen live, for a long period of time (1-3 hr) and at the level of a single cell.
approach referenced from Gaisa et al (see citation 4 in this paper) where they used mitochondrial DNA mutations to trace where each cell came from (lineage analysis)
This is super important for stem cell research.
Why? they are rare and hard to find and hard to identify and basically impossible to label with gene expression tag
But you can do it with lineage analysis!
How did they do it back in the day without being able to creep on cells in vivo?
well, they took tissue from active stem cell tissues (like bone marrow or testis), and put them in a host with little to no stem cell activity, and then voila, cells were being made there.
researchers figured out that there needed to be an area "stem cell niche" to for stem cells to be enriched and function
BUT they couldn't tell if all the cells they transplanted were actually stem cells or just some of them were…
some later studies in flies and mice showed that even daughter cells from stem cells that had begun differentiating (progenitor cells we call them now) could reverse and go back to being stem cell if they found a niche-->hmm, it was now supposed that tissues often contain lots of potential stem cells as well as stem cells.
But we still can't pick out and identify specific actual stem cells until we can look at them in vivo without disturbance. We still don't know where they are in most mammalian tissues.
But we can fix that! We need everyone to do lineage tracing in all of the organs!
it used to be that people searched for stem cells on guesses and unfounded assumptions:
myth: most stem cells are quiet and reproduce only sporadically.
assumption: it must be cells that are still labeled with BrdU (thymidine analog, gets into DNA) after a long time--because they are quiet--must be stem cells.
truth**: at least six types of stem cells have been shown to divide continuously
myth: stem cells spawn huge numbers of daughter cells and these may even be extremely diverse cells (different cell types) once you stick it into tissue culture
assumption: if we stick cells into tissue and they don't suddenly reproduce a crap ton of cells, they must not be stem cells…
truth: most stem cells are maintained as stem cells only by their niche. if they leave or are pushed out, they stop being stem cells and differentiate.
example: mouse intestinal stem cells need specific cytokines and actual niche cells and other stuff in order to successfully propagate.
(**Keep in mind that I only say "truth" to mean that there is evidence to the contrary, not that this is the as-laid-down-by-the-laws-of-nature-factual-truth. Remember always to have a healthy dose of skepticism in science.)
we only figured all that out after the cells were identified by lineage analysis and we could characterize their niche and signals in the niche
unless we have an exact replica of the niche, stem cell in tissue culture will look nothing like stem cells in vivo.
This is applicable to fields outside stem cell research.
embryo cells have constant signaling and interaction with their environment during development
cells in culture also have change in genes/epigenetics, even if their overall karyotype is still the same.
this is all likely due to the fact that they are stressed in an environment not like what they should be in (different signals, treatments, etc)
this is why tissue culture instead of just cell culture is better for multicellular biology
lessons learned from stem cell research:
if we still want to use cell culture but we want to look at things that don't happen just by themselves, we need new tools
we have to look at the events in vivo or in tissue culture
when we figure stuff out about the cells of interest and their environment, then we can create a replica cell culture
then we gotta double-check our model to make sure cells in vitro are behaving just as we had already observed in vivo
you have to follow these steps if you want accurate replication of the crazy complicated system of biology that every cell activity depends on.
good thing new developments in live imaging and lineage analysis are going to make this easier
just remember that you don't want to destroy the very biological events/systems that you wanted to study in the first place.
First of all, remember to keep checking the google doc of the lecture transcripts. I always update it ASAP and I've been trying to correct my spelling mistakes and add in a few more explanatory words in case I'm not sure the lecturer was clear on something.
Also I'm experimenting with recording the lectures, b/c for some, they are better audio learners. Bear with me as I struggle with technology.
Finally, we have one or two papers assigned for every lecture, so I'll post a sum-up of the papers. I still recommend you read the papers and then read the sum-up, because 1) you need practice if you ever want to be able to digest the condensed jargon of science writing and 2) the more you read the material over and over again the more it sticks in your head.
Here it is from Lecture 1 (Please excuse my casual language. It is my opinion that to make science available to the general, you must speak "layman"...seriously though. I am not trying to condescend to anyone, I just want to make sure people can understand, because that's the foundation to building up knowledge later on.)
Also remember that every paper wants to tell you something, but it is always up to you if you choose to follow or take it for its word. Always ask questions (is this right? did they do this correctly? did they consider this? etc etc) Whatever I write below is not an expression of something I believe in, but just a paraphrase of what I interpreted the paper was trying to say. I'm always open to discussion! (hence, comment box)
Spradling, Allan C. "The living-tissue microscope: the importance of studying stem cells in their natural, undisturbed microenvironment." J Pathol 2011; 225: 161-162.
We need tools to look at cells in their daily lives without disturbing them.
Why? Because we would find out all about normal cell lives, diseased cell lives, new cell to cell and cell to gene and gene to gene pathways that we can't see from frozen or dead cells.
We have the tools to do this:
Small multicell structures with labels can be seen live, for a long period of time (1-3 hr) and at the level of a single cell.
approach referenced from Gaisa et al (see citation 4 in this paper) where they used mitochondrial DNA mutations to trace where each cell came from (lineage analysis)
This is super important for stem cell research.
Why? they are rare and hard to find and hard to identify and basically impossible to label with gene expression tag
But you can do it with lineage analysis!
How did they do it back in the day without being able to creep on cells in vivo?
well, they took tissue from active stem cell tissues (like bone marrow or testis), and put them in a host with little to no stem cell activity, and then voila, cells were being made there.
researchers figured out that there needed to be an area "stem cell niche" to for stem cells to be enriched and function
BUT they couldn't tell if all the cells they transplanted were actually stem cells or just some of them were…
some later studies in flies and mice showed that even daughter cells from stem cells that had begun differentiating (progenitor cells we call them now) could reverse and go back to being stem cell if they found a niche-->hmm, it was now supposed that tissues often contain lots of potential stem cells as well as stem cells.
But we still can't pick out and identify specific actual stem cells until we can look at them in vivo without disturbance. We still don't know where they are in most mammalian tissues.
But we can fix that! We need everyone to do lineage tracing in all of the organs!
it used to be that people searched for stem cells on guesses and unfounded assumptions:
myth: most stem cells are quiet and reproduce only sporadically.
assumption: it must be cells that are still labeled with BrdU (thymidine analog, gets into DNA) after a long time--because they are quiet--must be stem cells.
truth**: at least six types of stem cells have been shown to divide continuously
myth: stem cells spawn huge numbers of daughter cells and these may even be extremely diverse cells (different cell types) once you stick it into tissue culture
assumption: if we stick cells into tissue and they don't suddenly reproduce a crap ton of cells, they must not be stem cells…
truth: most stem cells are maintained as stem cells only by their niche. if they leave or are pushed out, they stop being stem cells and differentiate.
example: mouse intestinal stem cells need specific cytokines and actual niche cells and other stuff in order to successfully propagate.
(**Keep in mind that I only say "truth" to mean that there is evidence to the contrary, not that this is the as-laid-down-by-the-laws-of-nature-factual-truth. Remember always to have a healthy dose of skepticism in science.)
we only figured all that out after the cells were identified by lineage analysis and we could characterize their niche and signals in the niche
unless we have an exact replica of the niche, stem cell in tissue culture will look nothing like stem cells in vivo.
This is applicable to fields outside stem cell research.
embryo cells have constant signaling and interaction with their environment during development
cells in culture also have change in genes/epigenetics, even if their overall karyotype is still the same.
this is all likely due to the fact that they are stressed in an environment not like what they should be in (different signals, treatments, etc)
this is why tissue culture instead of just cell culture is better for multicellular biology
lessons learned from stem cell research:
if we still want to use cell culture but we want to look at things that don't happen just by themselves, we need new tools
we have to look at the events in vivo or in tissue culture
when we figure stuff out about the cells of interest and their environment, then we can create a replica cell culture
then we gotta double-check our model to make sure cells in vitro are behaving just as we had already observed in vivo
you have to follow these steps if you want accurate replication of the crazy complicated system of biology that every cell activity depends on.
good thing new developments in live imaging and lineage analysis are going to make this easier
just remember that you don't want to destroy the very biological events/systems that you wanted to study in the first place.
Sunday, February 17, 2013
Cell Biology Study Guides
So I realized I must have shared my Cell Bio exam guides elsewhere or in person, b/c it's not on this blog!
So here they are! Cell Bio exam 1 guide and exam 2 guide. There's already a post for exam 3 somewhere on this blog.
exam 1: https://docs.google.com/file/d/0B99-sSwVe231OWM2YjA3OGItOTU3Ni00MGFiLWEzYzEtMzViNTczMTljYmY5/edit?usp=sharing&authkey=CLKk8NQB
exam 2: https://docs.google.com/file/d/0B99-sSwVe231NTI4Mzk1MWMtMGQwYi00M2JlLWFkM2YtMGRkYTM4NDNhYTFl/edit?usp=sharing&authkey=COCampkH
Again, let me know if the links don't work! (via comment or something)
So here they are! Cell Bio exam 1 guide and exam 2 guide. There's already a post for exam 3 somewhere on this blog.
exam 1: https://docs.google.com/file/d/0B99-sSwVe231OWM2YjA3OGItOTU3Ni00MGFiLWEzYzEtMzViNTczMTljYmY5/edit?usp=sharing&authkey=CLKk8NQB
exam 2: https://docs.google.com/file/d/0B99-sSwVe231NTI4Mzk1MWMtMGQwYi00M2JlLWFkM2YtMGRkYTM4NDNhYTFl/edit?usp=sharing&authkey=COCampkH
Again, let me know if the links don't work! (via comment or something)
Sunday, February 10, 2013
Stem Cells
For the purpose of the class I'm taking, I'll be making three kinds of posts:
1) script -> the class at our university does not have podcasting, so I will be transcribing a misspelling-prone version of near-verbatim of the lectures. Please find it here: https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
I have little time to spellcheck or grammar check, so please comment if you find something confusing that I can clarify!
2) study guide -> this will be prepared prior to every exam. hopefully it helps!
3) list of papers -> a significant portion of this class involves reading primary documentation, so I will be listing citations of the papers we read (and that will contain info that I will probably include in my study guides).
Ready for stem cells!
1) script -> the class at our university does not have podcasting, so I will be transcribing a misspelling-prone version of near-verbatim of the lectures. Please find it here: https://docs.google.com/document/d/1iFMdbzr9tr0M8PuvGmHvYGXVrPMIAV78OSxW2cGsQxM/edit?usp=sharing
I have little time to spellcheck or grammar check, so please comment if you find something confusing that I can clarify!
2) study guide -> this will be prepared prior to every exam. hopefully it helps!
3) list of papers -> a significant portion of this class involves reading primary documentation, so I will be listing citations of the papers we read (and that will contain info that I will probably include in my study guides).
Ready for stem cells!
Thank you!
Hello! I've been missing for a while (read as "not taking science classes"). This semester I will be back with Stem Cell Biology.
I've just discovered that this blog has rocketed in pageviews over the course of the past several months. I'm not sure what happened, but at least half of my total pageviews have come from Belgium. Not sure who's reading up on science, but I certainly hope it helps! It's very encouraging to know people are looking at what I post, since the purpose of this is to try and share science in a more understandable and explained way than the condensed jargon-dense speech of textbooks.
Cheers!
I've just discovered that this blog has rocketed in pageviews over the course of the past several months. I'm not sure what happened, but at least half of my total pageviews have come from Belgium. Not sure who's reading up on science, but I certainly hope it helps! It's very encouraging to know people are looking at what I post, since the purpose of this is to try and share science in a more understandable and explained way than the condensed jargon-dense speech of textbooks.
Cheers!
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