Recreating Anatomy for Regenerative Medicine – Stem Cells

Recreating Anatomy for Regenerative Medicine – Stem Cells

Module Title: 4BBA1010 Fundamentals of Anatomy and Developmental Biology

  • Prof. Francesca M Spagnoli

  • Date: 10th February 2025

  • Faculty: Life Sciences & Medicine

  • Location: Centre for Gene Therapy & Regenerative Medicine, Guy's Campus

Overview of Key Topics

  • Key properties of Stem Cells
       - Self-renewal
       - Differentiation
       - Role in organ contribution and maintenance of tissue
       

  • Cell-cell communication: Niche & Microenvironment

  • Applications of Stem Cells in Regenerative Medicine

  • Recreating Anatomy in Regenerative Medicine
       - Self-organization vs guided engineering approaches
       - Example 1: Intestinal organoids and engineering the niche
       - Example 2: Bioengineering pancreatic cell niche(s)

Overview on Stem Cells

Definition of Stem Cells

  • Stem Cell: An undifferentiated cell with the ability to self-renew and differentiate into specialized cell types.
       - Self-renewal: Process through which stem cells replicate while maintaining their undifferentiated state, maintaining the stem cell pool
       - Differentiation: Process through which stem cells specialize into differentiated cells contributing to organ formation and maintenance.
          - Specialized cells replace dead or damaged cells enable tissue to respond to physiological demands.

Types of Stem Cells

1. Embryonic Stem Cells (ESCs)

  • Originates from the inner cell mass of the blastocyst.

  • Characteristics:
       - Self-renewal: Ability to replicate indefinitely.
       - Pluripotency: Can differentiate into over 250 cell types. gives rise to tissue stem cells for the 3 germ layers so can form any cell in the body

  • Applications involve generating mouse chimeras and various tissue types.

  • generation of 254 cell types originating adult tissue

2. Adult Stem Cells (ASCs)

  • Derived during ontogeny and persist in niches within most adult tissues/organs.

  • undifferentiated cell found in some adult organs

  • self-renew and differentiate to become most or all the specialized cell types within the specific organ

  • no ethical issues, restricted plasticity, limited quantities, hard to identify

  • Characteristics:
       - Self-renewal: Capability of prolonged replication.
       - Multipotency: Ability to differentiate into multiple tissue types. cells have the ability to develop into specific types of cells

  • typically present in organs that undergo constant renewal

  • Key for maintaining tissue homeostasis under both physiological and pathological conditions.

3. Induced Pluripotent Stem Cells (iPSCs)

  • Created from somatic differentiated cells after transduction by transcriptional growth factors like Oct4, c-Myc, Sox2, and Klf4.

  • FGF preserves them in a pluripotent state, once removed they can differentiate into any cell type

  • Mature differentiated cells can be reprogrammed to become pluripotent

  • once established can self-renew and differentiate into any cell type

  • Characteristics:
       - Self-renewal: Similar to ESCs.
       - Pluripotency: Ability to generate various cell types.

  • Applications include patient-specific stem cells for research and therapy.

  • generation of mouse chimeras

  • important for study of development, pathogenic study, stem cell-based regeneration

Establishment of Cell Identity

  • extrinsic signals (growth factors, e.g. LIF and FGF)

  • morphogenesis and cell interactions

  • Key transcription factors involved include:
       - Oct4, Nanog, LIF

  • requires both intrinsic and extrinsic signals to cause differentiation into specific cell type

  • unraveling control mechanisms of cell identity has direct clinical and translational relevance

Stem Cell Niche and Microenvironment

The Stem Cell Niche Concept

  • A Cellular Niche:
       - Provides a supportive environment for controlled self-renewal and differentiation through cell-cell interactions.

  • adult and pluripotent cells reside in a microenvironment which preserves the cells in an uncommitted state and helps them to support their self-renewal

  • where adult cells interact with other cell (bone marrow, intestine)

  • e.g. reside in hair follicles within basal layer of epidermis

  • e.g. intestinal SCs found at base of crypts
     

  • A Non-cellular Niche:
       - Extracellular matrix (ECM) plays a crucial role in providing structural support and signaling for stem cells.

    • reside in baso-lamina (ECM)

    • e.g. muscle (satellite cells)

  • cellular niche:

    • composed of other cell types (eg. bone marrow or intestine)

Intestinal Stem Cell Niche

  • Anatomy:
       - Villus and Crypt Structure:
          - Cell shedding leads to differentiation and migration of cells.
       - Short turnover time of small intestinal epithelium: ~5 days.
       - Proliferative progenitors include goblet, entero-endocrine, absorptive cells, and Paneth cells.
          - Total mitotic renewal takes 24-36 hours from crypts to villus apex.

  • Lgr5 + SCs → give rise to transit amplifying cells → differentiate into: goblet cells, enteroendocrine cells, absorptive epithelial cells, paneth cells

Applications of Stem Cells in Regenerative Medicine

General Applications of Pluripotent Stem Cells

  • Cell Therapy:
       - Types of Cells Derived:
          - Hepatocytes, pancreatic β-cells, lung and airway epithelial cells, cholangiocytes.
       - Limiting factors: Cell signaling pathways (Activin, BMP4, FGF2, HGF, etc.).
       - Diseases targeted include liver disease, diabetes, and cystic fibrosis.

  • Disease Modeling:
       - iPSC-derived models used to understand genetic disorders, infections, and gastrointestinal disorders.

  • Drug Screening:
       - Used for safety and efficacy testing in hepatotoxicity and other diseases.

    • need to be able to directly differentiate pluripotent cells to find cell of interest - directed differentiation

Progress in iPSC-based Therapies

Focus Areas:

  • allow us to model disease, define cell therapies, drug screening, etc.

  • New drug development and target validation via patient-specific stem cells.

  • Organogenesis:
       - Organoids are created for patient-specific drug responses, leading to personalized medicine approaches.

Recreating Anatomy in Regenerative Medicine

Key Components to Consider

  • cell types are the building blocks of our organs stuck together by the ECM which is fundamental for mechanical reason and signals important for survival, proliferation and differentiation of the cell

reading:

  1. Components of tissue architecture

  2. Ratios of different cell types

  3. Spatial distribution and overall tissue architecture

writing:

  1. Assembling components into a functional unit

  2. Maturation into mature tissues

  3. Preservation of functionality

Objective

  • The ultimate goal is to recapitulate the native organ formation to address tissue loss or dysfunction.

Organoid Technology

Definition

  • Organoids are three-dimensional (3D) culture systems derived from stem cells that reflect simplified versions of organs and their complex structures.

Self-organization Process

Self-organization: A process by which local interactions between cells that are initially disordered lead to the emergence of patterns and functions
   

  • Lgr5+ organoid technology

    • biopsies are taken from patent, which contain intestinal crypts (containing LGR5+ SCs)

    • intestinal crypts are isolated and added to suitable culture medium → mimics intestinal SC niche to allow them to grow

    • LGR5+ will proliferate and self-organise to form organoids → organoids can be expanded to provide renewable source of SC

    • SCs can be transplanted back into patient to repair damaged intestinal tissue

Guided engineering

  • an approach that involves using biomaterials and growth factors to engineer tissue or organs

    • involved seeding SCs onto scaffold made of biocompatible material which mimics the natural environment of the tissue or organ

    • SCs grow in patterned tubular matrix organize into crypt-like structures and then are guided to differentiate into the desired cell type using growth factors or signaling molecules

    • e.g. bioengineering pancreatic cells for the treatment of diabetes, the pancreas lacks adult SC therefore islet transplantation is used to treat diabetes

Engineering Pancreatic Niche(s)

Developmental Stages

  • Analysis of embryo stages E12.5 to E14.5 focusing on mesenchymal cells, vascular networks, and integration of these structures into a functional pancreas.

Tissue Architecture

  • The exocrine pancreas comprises ~90% of the overall pancreatic mass while endocrine tissues comprise ~1-2%.

Pancreatic organ development

  • pancreatic lineage fate decision:

    • requires TF for guide differentiation

    • FGF10 drives primary transition

    • promotes proliferation and differentiation of pancreatic progenitor cells into tip and trunk progenitor populations

    • endocrine/exocrine/ductal lineages

  • notch and retinoic acid signaling pathway

    • drives secondary transition

    • tip progenitors → acinar exocrine cells

    • trunk progenitors → give rise to endocrine and ductile lineages

    • determines fate of progenitor cells to ensure proper differentiation, regulating balance between progenitor cell maintenance and differentiation

Challenges in Pancreatic Regeneration

Statistics on Diabetes and Pancreatic Cancer

  • Diabetes:
       - Affects approximately 422 million adults globally.
       - Leads to approximately 3.7 million deaths annually due to complications.
       - Connection to obesity, family history, and lifestyle factors.

  • Pancreatic Cancer:
       - Mortality rates are increasing, with very low survival rates post-diagnosis (~4.6 months).
       - Symptoms include pain, nausea, jaundice, and weight loss.

Therapeutic Options

  • Islet Transplantation: A potential solution to restore insulin production in diabetes patients.

  • Regenerative cell therapies for diabetes:

    • cells can be attacked after transplantation by immune system

    • pluripotent cells generated from embryos or patient somatic cells

    • Differentiate into: Pancreatic progenitors, B-like cells, mature B-cells

    • To prevent immune rejection, cells are enxcapsulated

      • macro-encapsulation: many cells in a single device

      • micro-encapsulation: each cell/group encapsulated

    • transplanted into the patient → aiming to restore insulin production and reverse hyperglycemia

Advances in Engineering Functional Pancreatic Tissue

Techniques

  • Includes three-dimensional bioprinting and encapsulation methods for delivering pancreatic progenitors into a functional structure.

Conclusion

  • Fundamental properties of stem cells are pivotal for applications in regenerative medicine.

  • The comparison between self-organization and guided engineering approaches is crucial for effective applications in organoid and pancreatic engineering.