BME 4710 Lecture 2 SLIDES

Cells in Tissue Engineering

Cell Types Derived from Tissue

  • Over 200 different cell types exist.

  • Organized into various tissue types:

    • Epithelia

    • Connective tissue

    • Nervous tissue

    • Musculoskeletal

    • Blood

    • Sensory

    • Stem cells

Cell Type Based on Culture

Primary Culture

  • Cells are isolated from tissue and plated to confluence.

  • Considered "primary" if used after first plating.

  • Features:

    • Heterogeneous cell population: Culture often contains multiple cell types.

    • Heterogeneity degree varies based on:

      • Type/location of tissue.

      • Technique/expertise of procedure.

      • Culture conditions (cell attachment, media additives, type).

Cell Sources

  • Sources include:

    • Epidermis

    • Dermis

    • Subcutis

Cell Isolation Techniques

  • Biopsy: Involves collecting tissue samples.

  • Liposuction: Technique used for fat removal.

  • Use of Collagenase I (0.5% and 0.075% concentrations) for cell separation from tissues.

  • Centrifuge steps used (1500 rpm for 5 mins, 600 rpm for 10 mins).

Cell Type Based on Culture (Continued)

Cell Lines

  • Defined as cells subcultured from initial culture.

  • Procedures:

    • Lifted from culture plastic upon confluence.

    • Rendered into single-cell suspension for re-plating.

    • Ability to expand number from one flask to multiple.

Finite vs Continuous Cell Lines

  • Finite cell lines: Will senesce (lose proliferative ability) due to shortened telomeres.

  • Continuous (immortal) cell lines: Can proliferate indefinitely, potentially induced virally or occurring spontaneously, such as in tumor cells.

Examples of Cell Lines

  • BALB/c: Derived from albino mice.

  • MC3T3: Osteoblast precursor from mouse calvaria; multiple subclones with specific traits.

  • NIH3T3: Fibroblast from mouse; favored for gene transfection.

  • Saos-2: Osteosarcoma cell; well-documented, predictable osteoblast-like behavior.

  • HeLa: Cervical cancer cells from Henrietta Lacks (1951); durable and maintainable through numerous passages; controversial usage.

Utility of Primary Cells in Tissue Engineering

Benefits

  • Accurate assessment of cell behavior similar to in vivo performance.

  • Fewer mutations and more “life-like” results.

  • Greater acceptance in scientific community due to authenticity.

Drawbacks

  • High cost involved in animal sourcing and research.

  • Time-consuming isolation procedures.

  • Issues with heterogeneity can complicate results interpretation.

  • Limitations on cell quantity and experimental size.

Utility of Cell Lines in Tissue Engineering

Benefits

  • Cost-effective and quick to obtain.

  • Versatile and robust, allowing multiple experimental rounds with the same population.

  • Generally homogenous in cell type.

Drawbacks

  • Less indicative of native cell behavior due to potential mutations.

  • Results face increased scrutiny and may mutate over time.

Primary Cells vs Cell Lines

  • Scenarios for using cell lines vs. primary isolation should be carefully considered based on research goals and requirements.

Stem Cells

Definition

  • A stem cell possesses the ability to self-replicate and differentiate into various cell types of the body.

Stem Cell Behavior

  • The fate of daughter cells is influenced by various unknown factors.

  • Requires specific culture conditions to maintain proliferation and prevent differentiation.

Stem Cells to Form Tissues

Embryonic Stem Cells

  • Isolated from the inner mass of the blastocyst and expanded in culture to maintain properties.

Culturing Methods

  • Plates are treated with feeder layers providing necessary components for stem cell survival.

Types of Stem Cells

  • Totipotent: Can develop into any tissue type.

  • Pluripotent: Can become any cell type from three germ layers (excludes extra-embryonic tissues).

  • Multipotent: Can differentiate into a limited range of cell types (e.g., mesenchymal stem cells).

  • Induced Pluripotent Stem Cells (iPS cells): Reprogrammed somatic cells into a pluripotent state.

The Three Germ Layers

  • Endoderm: Leads to internal organs (i.e., lungs, thyroid).

  • Mesoderm: Forms structures like bones and muscles.

  • Ectoderm: Develops into the skin and nervous system.

Utility of Stem Cells in Tissue Engineering (Mesenchymal)

Benefits

  • Provide a good reflection of the in vivo environment and assist in the development of inductive materials.

Drawbacks

  • More effort required for isolation and maintenance than cell lines.

Utility of Stem Cells in Tissue Engineering (iPS)

Drawbacks

  • Risk of tumorigenesis leading to uncontrolled growth rates.

  • Issues with complete differentiation and low efficiency.

Cell Culture Processes

Cell Acquisition/Isolation and Maintenance

  • Focus on methods to maintain homogenous cell populations using cell banks and filtration techniques.

Differentiation of Cells

Induction of Differentiation

  • Can be triggered via growth factors, co-culturing, physical manipulation of environment.

  • Differentiation often marked by a decrease in proliferation.

Cell Signaling in Tissue Engineering

Mechanism

  • Cellular communication initiated by binding of ligands which start signaling cascades.

Integrin Signaling

  • Integrins mediate interactions between cells and the extracellular matrix (ECM).

  • This signaling is crucial for cell behavior and tissue responses.

Impact of Molecular Knowledge on Tissue Engineering

  • Understanding of molecular responses aids material design and stimulation approaches for improved cell interactions.