SIO2004 Animal Cell Culture Lecture 5

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Questions and Answers

What happens to the average telomere length in humans from birth to old age?

Declines from approximately 11 kilobases to less than 4 kilobases (correct)

Increases slightly and then decreases significantly

Remains constant at around 11 kilobases

Increases to more than 20 kilobases

Which of the following is NOT a function of telomerase?

Preventing DNA replication errors (correct)

Lengthening telomere repeats

Allowing cells to exceed the Hayflick limit

Preventing degradation of chromosomal ends

In which types of cells is telomerase most commonly expressed?

All cells in the human body

Somatic cells

Mature somatic cells

Fetal tissues, adult germ cells, and stem cells (correct)

What occurs when a cell reaches a critical telomere length?

The cell experiences growth arrest and may become senescent or undergo apoptosis Signup and view all the answers

What is the Hayflick limit in cellular biology?

The maximum number of times a normal somatic cell can replicate before senescence Signup and view all the answers

What typically happens to most normal cells when cultured over time?

They have a finite lifespan Signup and view all the answers

Which phase of the cell cycle is often associated with the restriction point (R point)?

G1 phase Signup and view all the answers

What role does telomerase play in cell division?

It extends telomeres which allows cells to divide indefinitely Signup and view all the answers

What outcome usually occurs if DNA repair mechanisms fail during the cell cycle?

The cell enters the quiescent phase (G0) Signup and view all the answers

Which of the following proteins is primarily known as the master regulator of the cell cycle?

p53 Signup and view all the answers

What commonly contributes to declining cell division rates in normal cells?

Environmental factors like temperature and pH changes Signup and view all the answers

What is typically a sign of cell senescence in cultured cells?

Loss of the ability to divide Signup and view all the answers

How do telomeres affect the replicative lifespan of a cell?

Telomeres shorten with each division, limiting the lifespan Signup and view all the answers

What defines the number of times a cell line has been sub-cultured?

Passage number Signup and view all the answers

What is the average number of divisions after which human cells typically enter senescence?

52 divisions Signup and view all the answers

What protects the ends of chromosomes from deterioration during cell division?

Telomeres Signup and view all the answers

Which of the following statements about telomeres is incorrect?

They increase the lifespan of a cell. Signup and view all the answers

How many population doublings do human fibroblasts typically cease dividing?

25-40 doublings Signup and view all the answers

What role does telomerase play in relation to telomeres?

It elongates telomeres and helps maintain cell division. Signup and view all the answers

Which factor contributes primarily to the replicative lifespan of a primary cell in culture?

Telomere length Signup and view all the answers

What is the primary characteristic of replicative cell senescence?

Cells enter a non-dividing state. Signup and view all the answers

Study Notes

SIO2004 Animal Cell and Tissue Culture, Lecture 5

Course: Biotechnology Program
University: Universiti Malaya
Instructor: Dr. Nuradilla Mohamad Fauzi

Maintaining Cell Cultures

Maintaining cell cultures involves keeping cultured cells alive.
Media changes are crucial for replenishing nutrients and removing harmful metabolic byproducts and dead cells.
Adherent cultures have media removed and replaced through aspiration.
Suspension cultures require separation of cells from the media via centrifugation, then resuspension in fresh media.

Checking Cell Morphology

Regularly examine cell morphology (shape and appearance) under a microscope to assess successful cell culture experiments.
Healthy cells exhibit no granularity around the nucleus, remain attached to the substrate, and have no cytoplasmic vacuolation.
Signs of deterioration (granularity, detachment, vacuolation) should signal potential issues such as contamination, senescence, toxins in the medium, or the need for a medium change.

Cell Types

Primary culture: Freshly isolated from a tissue source.
Cell lines:
- Finite cell lines have a limited lifespan and die after several subcultures.
- Continuous cell lines are "immortal" and cancer-derived or transformed, able to be passaged and maintained.
- General rule: Cells that can divide can be passaged and maintained (not all).

Contact Inhibition and Confluence

Contact inhibition: Cell types avoiding each other, slowing proliferation.
Confluence: Cell layer covers the dish surface; cells stop dividing.
Confluence is used to estimate the number of adherent cells.

Subculturing / Passaging

At high confluence, cells become crowded, nutrients become limited, and cells may undergo differentiation or death. Passaging is needed.
Methods for detaching adherent cells include:
- Enzymatic methods (Trypsin, Trypsin-EDTA, Dispase)
- Mechanical methods (scraping, cell scrapers)
Suspension cells are subcultured by taking a small sample and diluting to fresh medium.

Cryopreservation

Freeze cells to maintain them when not needed.
Cryoprotectants like DMSO prevent ice crystal formation during the process.
Freeze at 1°C per minute.
Store in liquid nitrogen.

Cell Cycle

Cells regenerate from existing cells.
Eukaryotic cell cycle is an orderly sequence of events where cells duplicate contents then divide.
The cell cycle control system (regulatory proteins) governs progression and responds to internal and external signals.

Cell Cycle Phases

Key phases (from figures): G1, S, G2, M.
G0 phase: Cells that aren't dividing.
Senescence: A non-dividing state entered by cells.

Telomeres

Telomeres are repetitive sequences at chromosome ends.
Protect chromosomal ends from degradation or fusion.
Telomeres shorten with each cell division.
Telomeres reach a critical length when cells stop dividing.
Some cells activate telomerase enzyme to extend telomeres, therefore allowing continued cell division.

Telomerase

Telomerase is a reverse transcriptase enzyme adding to telomeres.
Prevents degradation of chromosomal ends.
Its expression regulates development.
Post-natal somatic cells often lack telomerase activity.

Cells that Express Telomerase

Fetal tissues, adult germ cells, stem cells, cancer, transformed cells, express telomerase to extend telomere lifespans.

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Description

Explore the essential techniques in maintaining animal cell cultures in this lecture from the Biotechnology Program at Universiti Malaya. Understand the processes of media changes and the importance of monitoring cell morphology for successful cultures. The quiz will cover adherent and suspension cultures and their specific requirements.