stemm cells
Stem Cells: Fundamentals and Types
Definition of Stem Cells
A stem cell is a unique type of cell characterized by its ability to self-renew and differentiate into various specialized cell types.
Distinction from Terminally Differentiated Cells:
Terminally differentiated cells have reached their final state of maturity and function.
Key Characteristics of Stem Cells
Not Terminally Differentiated:
They have not yet reached their final mature state.
Unlimited Division:
Stem cells can divide essentially without limit for the lifetime of an organism, demonstrating a form of cellular immortality.
Self-Renewal:
Upon division, stem cells produce two daughter cells; crucially, one daughter cell preserves the stem cell-like characteristics while the other typically differentiates.
Maintenance of Undifferentiated State:
They retain the ability to remain unspecialized until prompted by signals to differentiate.
Self-Renewal Mechanism
The self-renewal process ensures that the stem cell pool is maintained.
A division produces two daughter cells, with only one maintaining stem cell characteristics.
Comparison with Differentiation:
If both daughter cells committed to differentiation, it would reduce the available stem cell number.
Adult Stem Cells
Definition:
Stem cells present in adult organisms, essential for tissue repair and maintenance throughout life.
Presence and Abundance
Widespread Presence:
Adult stem cells are found in a greater number of tissues than previously believed.
Small Numbers:
They exist in very small quantities within these tissues, such as:
Example: Hematopoietic Stem Cells in the bone marrow.
Hematopoietic Stem Cells (HSCs)
They are responsible for generating all cells of the immune system and red blood cells in the bloodstream.
Prevalence:
Approximately 1 in 10,000 to 1 in 15,000 cells in the bone marrow is a bona fide hematopoietic stem cell.
This small number is sufficient to renew all necessary blood cells continuously.
Locations in the Body
Adult stem cells can be found in various tissues such as:
Bone Marrow
Skin Epithelium (for skin cell replacement)
Digestive System
Blood Vessels
Dental Pulp
Brain (especially in the hippocampus, responsible for learning and memory, replenishing neurons and glial cells).
Division Rate and Control
Characteristics:
Adult stem cells typically divide slowly and in a highly controlled manner.
Slow Rate:
They do not exhibit rapid division.
Controlled Division:
Their division is meticulously regulated, avoiding uncontrolled proliferation.
Purpose:
Stem cells replace differentiated cells that are no longer capable of division, ensuring safety by reducing the risk of retaining older cells that may harbor mutations.
Potency: Embryonic vs. Adult Stem Cells
Potency Definition:
Refers to the ability of a stem cell to differentiate into different cell types.
Embryonic Stem Cells (ESCs):
Categorized as Pluripotent:
Pluripotent cells possess the potential to become all different tissues in the body.
Adult Stem Cells (ASCs):
Generally classified as Multipotent:
They possess a limited capacity for differentiation compared to ESCs since they have already started differentiating.
Example:
A hematopoietic stem cell can generate all blood cells but cannot produce cells found in the stomach.
In Vitro vs. In Vivo:
Scientists may coax adult stem cells to show more plasticity in laboratory conditions (in vitro), yet their inherent potential (in vivo) remains restricted.
Mechanisms of Cell Fate Determination
Fundamental to stem cell biology is how a single stem cell can generate daughter cells with differing fates.
Divisional Asymmetry:
Refers to the unequal distribution of cytoplasmic contents during cell division resulting in different fates for daughter cells.
Unequal Cytoplasmic Factors:
Although both daughter cells share identical genetic content (genome), they receive different subsets of cytoplasmic factors (including proteins and gene regulatory factors).
Differential Gene Regulation:
Various factors lead to different gene expressions in each daughter cell, culminating in different protein production and distinct cell identities from the first division.
Environmental Asymmetry:
Occurs when daughter cells experience different microenvironments impacting their developmental paths.
Differential Signaling:
One daughter cell may remain attached to its original microenvironment, receiving signals necessary to preserve stem cell identity, whereas the other may detach and lose critical signaling inputs, propelling it towards differentiation.
Signal Transduction:
The attachment to the extracellular matrix (ECM) transmits signals influencing cellular behavior, while detachment alters the environmental context, leading to different outcomes in development.
Generating Large Numbers of Differentiated Cells
Despite the slow division of stem cells, they can generate vast numbers of differentiated cells required by the body.
Mechanism:
The original stem cell does not rapidly divide; instead, the daughter cell, known as a transit amplifying cell, can undergo multiple rapid divisions.
Transit Amplifying Cells:
The initial stem cell divides infrequently, while transit amplifying cells may divide rapidly (e.g., up to 30 times) before halting to establish a large population of terminally differentiated cells.
Hematopoietic Stem Cells (HSCs): A Key Example
Definition:
HSCs are multipotent adult stem cells found in the bone marrow capable of giving rise to all blood cell types.
Lineage Specification from HSCs
HSCs differentiate into two primary progenitor cell types:
Common Lymphoid Progenitor (CLP):
Gives rise to lymphatic cells:
T cells: Develop in the bone marrow but mature primarily in the thymus.
B cells: Develop chiefly in the bone marrow.
Dendritic cells: Can arise from CLP lineage.
Natural Killer (NK) cells: Also part of this lineage.
Common Myeloid Progenitor (CMP):
Gives rise to myeloid cells:
Granulocytes: Neutrophils, Eosinophils, Basophils.
Other myeloid cells, including macrophages, red blood cells, and platelets.
Multipotency of Hematopoietic Stem Cells
HSCs can generate all blood cell types, depending on received specific signals.
However, their multipotency remains restricted within the blood lineage and does not typically extend to unrelated cell types (e.g., neurons, epithelial cells).
Committed Progenitor Cells: HSCs lead to progenitor cells specifically designed to produce a limited number of blood cell types, narrowing differentiation potential.
Experimental Evidence: Mouse Studies
Scientific experiments in mice illustrated the regenerative capability of HSCs:
Irradiation Experiment:
Mice were subjected to irradiation to eliminate circulating blood cells, risking death through immune system collapse.
Bone Marrow Transplantation:
Bone marrow cells were extracted from a donor mouse and transplanted into the irradiated mouse, successfully repopulating its blood system.
Compatibility Requirement:
Donor and recipient mice must be MHC-compatible (analogous to HLA in humans) to prevent immune system rejection.
Remarkable Efficiency:
As few as 5 stem cells successfully reinstated an entire hematopoietic system, showcasing that even rare dividing stem cells can generate large differentiated populations.
Clinical Applications: Human Stem Cell Transplantation
Allogeneic Stem Cell Transplantation:
Pioneered by Dr. Donnall Thomas who won a Nobel Prize for the key findings.
1977 Study:
Involved 100 patients with blood cancers (e.g., leukemia) using HLA-matched sibling donors.
Initial marrow collection was painful as it involved accessing the interior of bones.
Patient Preparation:
Patients underwent total body irradiation and chemotherapy to ablate the existing blood system before transplantation.
Outcome:
Despite challenges, 13 out of the 100 patients survived, marking a successful initial application of stem cell transplantation.
Immune Attack:
Even with HLA matching, potential for Graft-versus-Host Disease remains.
Modern Advances:
Improved techniques now offer less toxic methods for ablating the patient’s blood system, enhancing safety and efficacy.
Autologous Stem Cell Transplantation
Process:
Patient’s own stem cells are collected and preserved.
After undergoing chemotherapy and radiation, patients receive their previously collected stem cells back.
Advantages:
Significantly safer as there are no rejection issues since cells are genetically identical to the patient.
Suitability:
Effective if the patient's cancer mutation is not present in their stem cells.
Stem Cell Transplantation: Autologous vs. Allogeneic Considerations
Congenital Mutations:
An autologous transplant would be ineffective if a mutation is present in all cells from birth.
Acquired Mutations:
Autologous transplantation is possible if the mutation occurs later in life and is non-congenital, allowing for the use of healthy stem cells.
Goal:
Use potent hematopoietic stem cells to repopulate the bloodstream with healthy components.
Applications of Stem Cells: The "Skin Gun" for Burn Victims
Issue with Severe Burns:
Address the urgent demand for extensive skin replacement in burn cases to prevent infections, often the leading cause of death in victims.
"Holy Grail" of Burn Surgery:
The goal is to generate sufficient healthy skin within one week.
Solution:
Dr. George Garlock developed a Skin Cell Gun that uses stem cells to expedite healing:
Isolate healthy skin cells (stem cells) from a small biopsy.
Suspend in a water solution.
Utilize a computer-controlled device to spray onto the burned area.
Advantages of the Skin Cell Gun:
Speed: Complete preparation in 1.5 hours.
Healing: Patients demonstrate rapid recovery time with severe burns healing within days.
Case Study:
Matt Euro, treated with the skin cell gun, achieved complete healing of severe burns in just days.
Cloning: Reprogramming Differentiated Cells and Dolly the Sheep
Cellular Plasticity Debate:
Exploration of whether fully differentiated cells could be reprogrammed to develop a new organism inspired cloning research.
The Dolly the Sheep Experiment
Conducted in 1996 by Scientists Ian Wilmut and Keith Campbell in Scotland, demonstrating that a terminally differentiated cell could give rise to a new organism.
Process of Somatic Cell Nuclear Transfer (SCNT) for Dolly:
Donor Cell Preparation:
White-Faced Ewe: A terminally differentiated mammary cell was harvested.
Cells were semi-starved to remodel their chromatin and arrest in a non-dividing state.
Enucleated Egg Cell Preparation:
An egg was harvested from another ewe; its nucleus was removed.
The remaining cytoplasm contained essential components for early development.
Cell Fusion:
The nucleus from the mammary cell was fused with the enucleated egg cell.
Embryo Development and Implantation:
The fused cell was cultured into early embryos and then implanted into surrogate mothers.
Birth of Dolly:
Of thirteen implanted embryos, only one resulted in a lamb, Dolly, who was genetically identical to the donor mammary cell.
Significance of Dolly
Proof of Nuclear Totipotency:
Demonstrated that differentiated somatic cell nuclei can direct the development of an entire organism.
Reversibility of Differentiation:
Showed that changes in gene expression and chromatin structure during differentiation are potentially reversible.
Monumental Achievement:
Despite low efficiency, it was a major milestone in biological science.
Controversy and Limitations of Dolly
Ethical Concerns:
Dolly's creation ignited debates over cloning ethics.
Low Efficiency:
The method's inefficiency posed technical challenges.
Health Issues:
Dolly was euthanized at age 6 due to lung disease, indicative of potential accelerated aging.
Reproductive Capacity:
Despite health issues, Dolly gave birth to four lambs, showcasing her ability to reproduce.
Other Cloning Developments Post-Dolly
Several other species have been cloned including cattle, cats, dogs, deer, mules, etc.
Somatic Cell Nuclear Transfer (SCNT) Process Summary (Example: Pig)
Egg preparation by enucleation.
Donor cell preparation from a differentiated somatic cell.
Fusion and activation of the reconstructed egg.
Cell division leading to pre-implantation embryos.
Implantation in surrogate mother leading to the birth of a genetically identical animal.
Introduction to Embryonic Stem Cells (ESCs)
Definition:
ESCs are pluripotent cells able to differentiate into any cell type from the three germ layers (ectoderm, mesoderm, endoderm).
Culturing:
Can be maintained indefinitely in controlled lab environments.
Source:
Derived from the Inner Cell Mass (ICM) of a blastocyst, typically 5-7 days old.
Culturing and Differentiation of ESCs
Milestones:
Mouse ESCs cultured in 1981; human ESCs in 1998, emphasizing species-specific conditions for ESC maintenance.
Directed Differentiation:
Scientists can direct differentiation through signaling molecules.
Example 1: Retinoic acid for neuron differentiation.
Example 2: Cytokines for macrophages.
Controversy and Ethical Considerations of Human ESC Research
Embryo Destruction:
Ethical concerns regarding the destruction of blastocysts during ESC extraction.
NIH Funding Guidelines:
Research requiring federal funding must use ESC lines from government-approved registries, currently around 500 lines available.
Therapeutic Promise of ESCs
Potential:
Inner cell mass cells can differentiate into any tissue type in the body.
In Vitro Differentiation:
Can be manipulated to form various cell types for therapeutic applications.
Chimeric Mice Demonstration:
Injecting ESCs into different blastocysts proves their capacity to form all approximately 220 cell types, contributing to all germ layers.
Controversy Ongoing:
Despite immense potential, ESC use is contentious due to embryo destruction.
Dr. Shinya Yamanaka and Induced Pluripotent Stem Cells (iPSCs)
Ethical Dilemma Resolution:
Yamanaka aimed to create pluripotent cells without embryo destruction.
Approach:
Reprogramming somatic cells back to an embryonic stem-like state rather than through nuclear transfer.
Key Characteristics of iPSCs:
Indefinite proliferation, capacity for multiple lineage differentiation, contribution to all three germ layers in live animals.
Timeline of Discovery
2006: Successful mouse reprogramming.
2007: Successful human skin cell reprogramming.
2012: Nobel Prize awarded for breakthroughs.
Mechanism of iPSC Reprogramming: The Yamanaka Factors
Reprogramming Factors Discovery:
Identification of key genes that induce pluripotency by acting as transcription factors.
The four Yamanaka factors: Oct4, Sox2, c-Myc, KLF4.
Reprogramming Process:
Delivery of these factors to somatic cells leads to chromatin remodeling and reactivation of embryonic stem cell characteristics, resulting in iPSCs.
Delivery Method:
Initially via a retrovirus to insert reprogramming genes.
Characteristics and Challenges of iPSCs
Similarities to ESCs:
Self-renewable, pluripotent, capable of in vivo differentiation.
Differences from ESCs:
Minor variations in gene expression patterns compared to ESCs.
Major Challenge—Safety Concerns:
Retroviral integration can lead to uncontrolled gene expression and oncogenic potential, raising cancer risks.
Challenges with Early iPSC Reprogramming Methods
Oncogenic Potential and Tumor Formation:
Random integration can disrupt important genes, increasing tumor formation risk.
Teratoma Formation:
Tumors may contain immature tissues, signaling incomplete differentiation of injected iPSCs.
Advancements in iPSC Delivery Methods
Scientists developed new methods to deliver reprogramming factors to minimize genomic integration and tumor risks.
Non-Integrating Viral Vectors:
Temporary delivery without permanent genomic changes.
RNA Delivery:
RNA delivery avoids genomic integration but allows transient action for nucleic reprogramming.
Episomal Vectors:
Plasmid-based delivery avoids modification to host DNA while remaining replicative outside chromosomes.
Clinical Potential and Applications of iPSCs
Patient-Specific Disease Models:
iPSCs from a patient matched genetically, aiding in drug screening and personalized medicine.
Avoiding Immune Rejection:
Autologous transplantation using patient-derived iPSCs eliminates the immune response risk.
Ongoing Research:
Direct reprogramming methods are explored to convert one cell type into another directly.
Applications and Challenges of iPSCs
Research Applications:
Drug Screening and Disease Modeling:
Evaluation of potential drugs using patient-specific iPSC derivatives.
Therapeutic Applications:
Replacing damaged or destroyed cells, though considerable risks exist including DNA integration issues, cell immaturity, and unforeseen behaviors upon transplantation.
Feasibility and Future Outlook
Cost Reduction:
With rapid advancements, costs for patient-specific iPSC lines are decreasing significantly.
Ongoing Trials:
Numerous studies are underway evaluating iPSCs; early trials have shown promise with patient-specific therapies.
Potential for Future Therapies:
Improving methods holds promise for deeper insights and innovations in treating human diseases.