Stem Cell Research & Therapy Lecture Notes

This set of flashcards covers key vocabulary and concepts related to stem cell research, specifically focusing on the generation and characterization of clinical-grade induced pluripotent stem cell (iPSC) lines, HLA compatibility, reprogramming methods, and quality control for clinical applications, based on a lecture about a Spanish iPSC haplobank.

Human-Induced Pluripotent Stem Cells (hiPSCs)

Cells generated from somatic cells that offer a virtually unlimited supply of specialized cells for transplantation and treating age-related diseases.

Allogeneic hiPSC collection

A collection of hiPSC lines from healthy donors that can be expanded and differentiated to treat different patients, reducing immune rejection risk if diverse and compatible homozygous HLA haplotypes are present.

Compatible HLA haplotype

Refers to the similarity or match between the Human Leukocyte Antigen (HLA) gene regions of two individuals, crucial for organ transplantation and immune compatibility.

HLA System

A group of genes on chromosome 6 encoding proteins critical for the immune system's recognition of self vs. non-self.

Haplotype

A group of alleles (gene variants) that tend to be inherited together.

HLA Typing

A process done via DNA sequencing to identify the exact HLA alleles present.

HLA Matching

The number of matches across key HLA loci (e.g., 6/6 or 10/10) that determines compatibility between individuals.

Perfect 10/10 HLA Match

A match where both individuals share both HLA haplotypes (one from each parent), leading to very similar immune systems and low risk of transplant rejection.

8/10 or 9/10 HLA Match

One or two mismatches in HLA alleles, often acceptable in less critical transplants or older patients, sometimes still referred to as 'compatible'.

Highly Immunogenic Tissues

Tissues like hematopoietic stem cells/bone marrow where HLA matching is essential; mismatching can cause graft-vs-host disease (GVHD) or graft failure.

Less Critical Transplants

Organs like the liver, heart, lung, and cornea where HLA matching is less important due to immune privilege or urgency, and immunosuppression is the main strategy.

Off-the-shelf cell therapy products

Cell derivatives from HLA-matched hiPSC banks that are easily accessible for critical acute, subacute, or new emergent diseases, reducing cost and patient immune suppression.

Cord Blood Units (CB units)

A cell type of choice for generating homozygous HLA haplotype hiPSC collections due to no collection risk, pre-existing HLA typing, fewer genetic/epigenetic risks, and established hiPSC generation methodology.

GMP conditions (Good Manufacturing Practice)

Strict quality assurance standards under which cell-based products for human therapy must be established in facilities with relevant product manufacturing license, ensuring ethical and legal compliance.

Clinical-use iPSCs

iPSCs acceptable for therapeutic use, requiring specificities for donor selection, cell sourcing, reprogramming methodology, culture/expansion, testing, and banking.

HeLa cells

The first immortal human cell line, derived from Henrietta Lacks' cervical cancer tumor in 1951 without her informed consent. Essential for biomedical research.

Hayflick Limit

A built-in limit in normal human cells (about 40–60 divisions) due to telomere shortening, leading to senescence or apoptosis.

Telomeres

The ends of chromosomes that shorten with each cell division, contributing to the Hayflick limit.

Senescence

A state of permanent cell cycle arrest.

Apoptosis

Programmed cell death.

HeLa Cell Immortality (HPV infection)

High-risk HPV-18 infection in Henrietta Lacks' cells caused viral proteins E6 and E7 to inactivate tumor suppressors (p53 and Rb), removing growth stop signals.

HeLa Cell Immortality (Telomerase reactivation)

Her cancer cells reactivated telomerase, an enzyme that rebuilds telomeres, preventing shortening and allowing continuous divisions.

HeLa Cell Immortality (Chromosomal instability)

HeLa cells are aneuploid (abnormal numbers of chromosomes), contributing to their rapid growth and adaptability in culture.

Yamanaka integrating viral transfection disadvantages

Associated with tumorigenicity (c-Myc and Oct4), insertional mutagenesis, lack of copy number control, and stimulation of immune responses.

Non-integration viral transfection disadvantages

Adenoviral and Sendai viruses can activate innate and adaptive immune responses.

Self-excising vectors disadvantages

Incomplete/inefficient excision, double-strand DNA breaks (DSBs) causing genomic instability, off-target recombination by recombinase enzymes, and potential innate immune responses.

Non-integrating non-viral approaches

Methods for iPSC generation that do not involve integrating genetic material into the host genome, such as episomal plasmids, synthetic mRNA, and synthetic protein.

Episomal Plasmids

Circular, non-integrating DNA vectors that express reprogramming genes via transfection; pros: no integration, simple; cons: moderate efficiency, multiple transfections, plasmid dilution.

Transfection

The process of introducing nucleic acids (DNA or RNA) into eukaryotic cells using non-viral methods, like chemical (lipofection) or physical (electroporation, microinjection) means.

Transduction

The process of introducing nucleic acids (DNA or RNA) into eukaryotic cells using viral infection methods.

Synthetic mRNA

Modified mRNA transcripts encoding reprogramming factors delivered daily via transfection; pros: high efficiency, no genomic DNA manipulation, transient expression; cons: labor-intensive, cytotoxic transfection, susceptible to degradation.

Synthetic Protein

Purified transcription factor proteins fused to cell-penetrating peptides; pros: extremely safe (no genetic material), no integration risk; cons: very low efficiency, difficult to produce/purify, requires frequent delivery.

Clinically Compliant iPSCs

iPSCs that require the use of xeno-free and clinical-grade reagents as well as integration-free methods of reprogramming, beyond full traceability of manufacturing.

Clinical-grade, iPSC haplobank

A bank of iPSC lines from selected donors with specific HLA haplotypes, generated and expanded under GMP-compliant methods for clinical therapeutic use, as presented in the Spanish study.

Sendai Virus Vector

A non-pathogenic, single-stranded RNA virus with high transfection efficiency that has no DNA intermediate, eliminating the risk of unwanted DNA integration for reprogramming.

Transcription Factors (SOX2, OCT4, KLF4, c-MYC)

Proteins that regulate gene expression, essential for inducing and maintaining pluripotency in stem cells.

iPSC Morphology

Characteristics of iPSC colonies including cobblestone appearance, high nucleus-to-cytoplasm ratio, and large nucleoli.

PCR analysis (Polymerase Chain Reaction)

A molecular biology technique used to amplify specific DNA sequences, in this context, to confirm the absence of Sendai virus genome and transgenes in iPSC clones.

SeV genome and transgenes

Genetic information (including reprogramming factors) introduced by Sendai virus during iPSC generation, which ideally should be absent from nascent iPSCs for clinical use.

G-Banding Karyotypes

A cytogenetic technique used to visualize chromosome banding patterns, allowing for the detection of chromosomal abnormalities (e.g., aneuploidy) in iPSC lines.

Immunocytochemistry (ICC)

A technique using antibodies to detect specific proteins or antigens within individual cells (cultured cells), used to study cell identity and protein localization.

Embryoid Body (EB) Differentiation

An in vitro technique where pluripotent stem cells form 3D aggregates (EBs) in suspension, mimicking early embryo development and inducing spontaneous differentiation into all three germ layers (ectoderm, mesoderm, endoderm).

Teratoma Assay

An in vivo functional test of pluripotency where PSCs are injected into immunodeficient mice, forming a benign tumor-like mass (teratoma) that is histologically analyzed to confirm differentiation into all three germ layers.

Chimera Assay

The most stringent in vivo functional test of pluripotency, where labeled PSCs are injected into a developing embryo to assess their ability to contribute to all tissues and potentially the germline of a resulting mosaic organism.

GMP-MCB (Master Cell Bank)

A fully characterized and tested cell bank from which working cell banks are derived.

GMP-WCB (Working Cell Bank)

A cell bank derived from the Master Cell Bank, used for routine production or clinical applications, subject to stringent quality control acceptance criteria.