Stem Cells & Cell Differentiation

Vow Company

start-up company using stem cells to create "clean meat"

Stem Cell

A cell that can renew(divide) or differentiate

Stem cell niche

what triggers the differentiation of stem cells into dif cells, AKA stem cell microenvironment, critical to controlling cell division vs. microenvironment

# of doublings of stem cells

100-200 doublings possible, more than regular somatic cell

adipose (fat) derived stem cells (adMSCs)

most popular type of adult stem cells, are in 700+ stem cell therapy trials rn

Fetal stem cells

amniotic, placental; umbilical cord are the most commonly used ones

embryonic stem cells

hESCs and hPSCs, hESCs have been in US clinical trials as of 2010

Differentiation

cell becomes more specialized, can be partial or full

"progenitor" cells

have "restricted lineage," limited to differentiating into only 1 or 2 types of cells

transdifferentiation (direct reprogramming)

converting a differentiated cell into another type of differentiated cell without going through an embryonic step

RNA-seq

one metric to compare iPSC generated hepatocytes to another one

Dedifferentiation & redifferentiation

ability of a cell to become more embryo-like and differentiate into another cell type, occurs in tandem!

reversine

chemical that can induce dedifferentiation

Eastern Red Spotted Newt

organism where dedifferentiation and redifferentiation has been observed to grow back limbs and lens of eyes

When was transdifferentiation first done experimentally?

1987, but several cells have been generated since

What does the stem cell niche include?

neighboring cells, ECM, local growth factors, physical environment (pH, oxygen tension, pressure)

Totipotent

Can differentiate into all cell types, typically only a fertilized egg counts as this

pluripotent

can differentiate into many cell types, some restricted stemness

Multipotent

can differentiate into several cell types, stemness even more restricted

Unipotent

can differentiate into only 1 cell type, would be a progenitor cell

Blastocyst

late pre-implantation stage embryo

Where do hESCs originate from?

inner cell mass of blastocysts

Purpose of chimera test

Designed to prove/disprove totipotency of cells, only way to do so

Chimera test experiment

Label test stem cell with GFP, implant it into the blastocyst, then put it in a surrogate mother, then track which tissues and organs have GFP labeled cells

Stimulus Triggered Acquisition of Pluripotency (STAP) cells

claimed cells treated with some sort of acid could turn somatic cells back into embryonic cells, which could then redifferentiate; turned out to be fraudulent

Biodistribution & Homing

ability of stem cells to find "home" (AKA the target tissue)

how does biodistribution/homing work

damaged/compromised tissue release factors that cause endogeneous MSCs to home to damaged site

Importance of biodistribution/homing

important to see if stem cells in therapies are actually going where they're going, has been found to occur in vivo (patient cardiomyctes found in transplanted hearts)

Fusogenic

ability of stem cells to spontaneously fuse with each other and form a tetraploid cell (which could generate cancer stem cells)

when could fusion of stem cells occur?

when they're injected into patient, more injections = increase mechanical stress, which causes fusion

SCID (Sever combined immuno deficiency) mice

have a compromized immune system (no B or T cells)

use of SCID mice

to determine if injected candidate stem cell can differentiate into multiple types of tissues/cells in vivo & if candidate human cancer cell can generate tumors in vivo

Somatic Cell Nuclear Transfer (SCNT)

enucleation of a nucleus from an egg and injecting a somatic nucleus into it

Importance of SCNT?

showed that cytoplasmic factors in the egg can reprogram a somatic cell nucleus and make it totipotent

Sir Ian Wilmut

cloned the sheep Dolly in 1996 using SCNT

Rhesus Monkeys

have been successfully cloned with SCNT, useful for testing drugs

potential application of SCNTs

therapeutic cloning to use hESCs as an autograft

Challenge of SCNT

thousands SCNTs required for one implantable embryo

Parthenogenesis

birth of a fully functional organism without using sperm

John Gurdon

First to clone frogs using SCNT in 1960

hPSCs

human parthenogenetic stem cells

benefit of parthenogenesis

only 200-300 eggs required to generate hPSCs that could match anyone in the world

Limitation of parthenogenesis

all alleles will be homozygous bc of no sperm, increases chance of phenotypic expression of a mutation

discovery of parthenogenesis?

by Loeb in 1913 using unferitilized sea urchin and unfertilized starfish eggs, changed osmolarity of surrounding medium and used dilute acid respectively

induced pluripotent stem cells (iPSCs)

adult somatic cells that were induced to become pluripotent stemm cells using 4 reprogramming factors

Shinya Yamanaka & James Thomson

Both discovered that somatic human cells could be converted to true stem cells with only 4 additional genes

How can you see if there was true differentiation in culture?

using RT-PCR

SCID mouse test for iPSCs

saw that they form teratomas in mice

Use of iPSCs

basic research on differentiation, drug discovery (make patient specific cells of individuals carrying genetic defects), future source of cells for stem cell therapy

iPSCs vs. adMSCs (from SCNT)

iPSCs more pluripotent

safety issues of stem cells

tumorigenicity, immunogenicity, inappropriate differentiation

Tumorigenicity

propensity to form tumors such as teratocarcinomas, applies to stem cells bc they can divide many more times than normal cells bc of long telomeres

immunogenicity

propensity to trigger an immune response, risk of triggering one increases with more frequent stem cell injections

inappropriate differentiation

risk of stem cells differentiating into cells that weren't intended, and not native to the target organ

Cord blood use

could be used for blood replacement and stem cell therapy

C. elegans

model organism to use to study stem cells, non-parthenogenic roundworm

Robert Horvitz

first discovered apoptotic genes in C. elegans

Victor Ambros and Gary Ruvkun

discovered first microRNA in C. elegans using heterochronic mutant

Benefit of using C. elegans

translucent, simple organism, limited number of cells, cells have been coded, can predict the differentiation patterns