hIPSCs Flashcards
(4 cards)
iPSC vs ESC
source esc- embryo
high rate of proliferation
high availability
spontaneous differentiation- yes
capacity to produce diverse cell types- high
exact same for ipscs except source- somatic cell
multipotent stem cells source- adult, perinatal and fetal tissues
rate- medium to low
availability- low
spontaneous- no
capacity to produce diverse cell types- low
cellular reprogramming
in esc, chromatin is globally decondensed (open). open chromatin is transcriptionally active. upon differentiation, chromatin becomes more condensed and trasncriptionally inactive.
reprogramming re opens chromatin, making it more active
chromatin re-opening during reprogramming may not always be complete and thus leaves an epigenetic memory of the original cell type.
ipsc: yamanka factors= oct2/3, sox2, klf4, myc, nanog, lin28
fibroblasts, peripheral blood mononuclear cells, retinal tubular epithelial cells and hair keratinocytes can be reprogrammed.
methods of reprogramming can be
integrating methods- retrovirus and lentivirus
or non-integrating methods- lentrivirus with loxp alleles, adenovirus, sendai virus, episomal plasmids, microrna, mrna, proteins
new mrna and microrna reprogramming kits can reprogram the cells in 10 days
applications of ipsc
disease modelling, drug screening and discovery, cardiac, neural, liver toxicity tests, can create stem cells from a patient
ipscs in modeling cardiac disease: skin i=fibroblasts—> retroviruses reprogramming method—> ipsc—-> forms embryoid bodies. differentiation into cardiomyocytes
can model arrthymia
neural disease modeling: same process. forms neurons. further differentiation into glutamatergic neurons (with synpases). reduced glutamergic synpase number and cell soma size. treat with IGF1 and forms cell with increased glutaminergic synpase number and cell soma size
can do autologous transplantation however, it is impractical. takes a long time to establish ipsc lines, validate and differentiate into target tissue. the patient’s genetic background may harbor other deleterious mutations, prohibitvely expensive
solution: allogenic transplantation from human leukocyte antigen (HLA)- matching donors
Title: Clinically Relevant HLA Loci for Transplantation
The loci A, B, and DRB1 are the most important HLA genes to match for transplantation. These are the ones your immune system uses to recognize self vs. non-self.
Case 1: Heterozygous Stem Cell Donor
Patient is HLA-heterozygous, meaning they have two different versions of each HLA gene (e.g. A1/A2, B7/B8, DRB103/04).
Stem cell is also heterozygous, but not a perfect match.
➤ Immune rejected? Yes.
➤ Why? The patient’s immune system can recognize non-self HLA from the donor and attack it.
Case 2: Homozygous Stem Cell Donor
The iPSC-derived stem cell line is HLA-homozygous, meaning it only expresses one version of each HLA gene (e.g. A1/A1, B7/B7, DRB103/03).
If the patient shares at least one HLA haplotype, their immune system is less likely to see it as foreign.
➤ Immune rejected? Much reduced.
➤ Why? Because the donor cells lack the extra set of HLA variants that would trigger rejection.
➤ Patient may not recognise foreign antigen: since the HLA molecules present antigens to immune cells, the reduced diversity in HLA may limit immune recognition of non-self.
Big Picture (What the Slide Is Trying to Say):
Using HLA-homozygous iPSCs can reduce immune rejection, especially if matched to the patient’s HLA type.
That’s why creating iPSC banks from HLA-homozygous donors is being explored — you can match more patients with fewer immune issues.
stringent criteria for donors: homozygocity of atleast 3 hla loci: A,B,DR. blood type O. clean medical and family history.
HLA type known—> blood cord banking
IPSC quality control
exome sequencing.
🔬 What is Genome Editing in iPSCs?
Genome editing (like using CRISPR/Cas9) refers to modifying the DNA of iPSCs to:
Correct genetic mutations (e.g. in patient-derived iPSCs for gene therapy).
Delete or alter specific HLA genes to reduce immune rejection.
Improve safety, e.g., by removing tumorigenic mutations or adding suicide genes for safety switches.
🧬 What are Universal Donor iPSCs?
These are iPSC lines that have been genetically modified to be compatible with many or all patients, regardless of their HLA type.
How is this done?
Delete/knock out HLA class I and II genes to avoid T cell recognition.
Insert immune-inhibitory molecules like HLA-E or CD47 to prevent attack by NK cells (which would normally kill cells that lack HLA).
The result: a “stealth” iPSC that can be used in many patients without rejection.
🧠 Think of it like this:
Instead of matching every patient’s HLA, you create an immune-invisible iPSC that no immune system can easily reject.
🧪 What does Exome Sequencing / iPSC Quality Control Have to Do With This?
Exome sequencing = sequencing all the coding regions of the genome.
It’s used for quality control of iPSC lines to:
Detect any mutations (e.g., cancer-related).
Ensure genomic integrity after reprogramming or genome editing.
Avoid transplanting cells with harmful mutations.
So:
Exome sequencing = QC tool.
Genome editing = engineering tool.
Universal donor iPSCs = engineered iPSCs that escape immune rejection.
✅ Summary You Can Use on a Slide:
Genome Editing & Universal Donor iPSCs
Genome editing tools (e.g., CRISPR/Cas9) are used to:
Correct disease-causing mutations.
Knock out HLA genes to reduce immunogenicity.
Universal donor iPSCs are engineered to avoid rejection in all patients by:
Deleting HLA genes.
Adding immunomodulatory genes (e.g., CD47, HLA-E).
Exome sequencing is used for quality control to ensure genetic safety of edited iPSCs.
