Apoptosis and FACS Analysis Quiz

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Questions and Answers

Which process is indicated by the presence of FITC labeled Annexin V binding to phosphatidylserine (PS)?

Mitosis

Autophagy

Necrosis

Apoptosis (correct)

In the FACS plot, which quadrant represents live cells that are negative for both Annexin V and PI?

Left upper quadrant

Left lower quadrant (correct)

Right upper quadrant

Right lower quadrant

What does the presence of a double membrane structure indicate in the context of cellular processes?

Autophagy (correct)

Mitosis activity

Cellular necrosis

Apoptotic bodies

Which statement correctly describes how the interpretation of a FACS plot changes when treating cells with chemotherapy?

Reduced population of green positive cells may indicate apoptotic response Signup and view all the answers

What is the primary effect of the inability to cleave pro-caspase 9 in the studied cell line?

The cells cannot activate caspase 3, blocking intrinsic apoptosis. Signup and view all the answers

Which mechanism best explains the action of the new drug developed for aggressive breast cancer?

MAP-kinase phosphorylates and activates Hid, leading to IAP protein activation. Signup and view all the answers

Which statement accurately describes how excessive apoptosis can lead to disease?

It plays a major role in autoimmune diseases by targeting healthy cells. Signup and view all the answers

How does dysregulation of apoptosis often manifest in cancer?

Through the overexpression of anti-apoptotic factors. Signup and view all the answers

What role does cytochrome c play in the intrinsic pathway of apoptosis?

It promotes the formation of the apoptosome. Signup and view all the answers

What is the primary function of phagolysosomes in the context of cellular processes?

To degrade old cellular components Signup and view all the answers

What signifies that a cell is undergoing death based on the authors' explanation?

The recognition of specific external signals Signup and view all the answers

Which statement best describes the role of the signals mentioned by the authors?

They trigger intracellular pathways for cell death. Signup and view all the answers

What is meant by the term 'coryolysis' as mentioned in the content?

The melting of chromatin Signup and view all the answers

In the context of cell death, what is the significance of the receptors mentioned?

They recognize specific external signals necessary for cell death. Signup and view all the answers

Which process occurs as a result of the activation of specific intracellular pathways?

Cellular apoptosis is initiated. Signup and view all the answers

What purpose do the authors aim to achieve with their illustration regarding cell death?

To demonstrate the need for specific signals for cell death. Signup and view all the answers

How do the authors depict the concept of recycling in cellular processes?

By showing how old components are used to create new materials. Signup and view all the answers

What is the primary function of apoptosis?

To remove damaged or unnecessary cells Signup and view all the answers

Which of the following correctly contrasts apoptosis and necrosis?

Necrosis is often uncontrolled, whereas apoptosis is regulated Signup and view all the answers

How do pro-apoptotic proteins function?

By initiating the apoptosis cascade Signup and view all the answers

Which best describes autophagy in relation to apoptosis?

It is a process of recycling cellular components Signup and view all the answers

What differentiates primary literature from secondary literature?

Primary literature contains original findings, while secondary summarizes past research Signup and view all the answers

What can result from dysregulation of apoptosis?

Development of various diseases Signup and view all the answers

For which purpose would primary literature be most suitable?

To read about original experiments and findings Signup and view all the answers

Which statement is true regarding apoptosis during development?

It is essential for normal developmental processes Signup and view all the answers

What role does Annexin V-FITC play in the identification of apoptotic cells?

It binds to phosphatidylserine exposed on the outer membrane. Signup and view all the answers

In flow cytometry, what does the left lower quadrant represent?

Cells that are alive and not undergoing apoptosis. Signup and view all the answers

What does the term 'apoptosis' refer to?

Programmed cell death that involves a series of biochemical events. Signup and view all the answers

What role does Bcl-2 play in the apoptosis process?

It inhibits cytochrome c release, preventing apoptosis. Signup and view all the answers

Why does only dead cells take up PI in flow cytometry?

PI cannot penetrate live cell membranes. Signup and view all the answers

What happens as a result of mutations in Bax?

It can impair the process of apoptosis. Signup and view all the answers

Which protein is primarily responsible for activating caspase 9?

Cytochrome c Signup and view all the answers

What occurs to phosphatidylserine (PS) during apoptosis?

PS flips from the inner to the outer membrane. Signup and view all the answers

How is fluorescent intensity measured in flow cytometry?

By detecting the light emitted by dyes bound to DNA. Signup and view all the answers

What is the main function of executioner caspase 3?

To cleave cellular proteins and initiate apoptosis. Signup and view all the answers

What does the right upper quadrant represent in flow cytometry results?

Apoptotic cells that are positive for both Annexin V and PI. Signup and view all the answers

Which statement about p53 is true?

It stimulates apoptosis if DNA damage cannot be repaired. Signup and view all the answers

What role does the FAS ligand play in apoptosis?

It facilitates the binding to death receptors, activating apoptosis. Signup and view all the answers

Which of the following statements correctly describes the cleavage involved in apoptosis?

It leads to the formation of apoptotic bodies. Signup and view all the answers

How do IAPs affect the apoptosis process?

They block the action of pro-apoptotic signals. Signup and view all the answers

What is the consequence of mutations in phosphatidylserine?

Random destruction or survival of cells. Signup and view all the answers

Study Notes

Apoptosis and Cell Death Mechanisms

Apoptosis is programmed cell death, crucial for development and maintaining cellular health.
Apoptosis is an energy-dependent process characterized by specific morphological changes.
Apoptosis pathways include intrinsic (internal cellular stress) and extrinsic (external signals).
Apoptosis doesn't induce inflammation.
Cells shrink, condense, and form apoptotic bodies, which are then engulfed.

Learning Objectives

Students should compare and contrast apoptosis, necrosis, and autophagy by understanding cellular changes.
Students should explain why apoptosis is a normal process and provide examples.
Students should learn the intrinsic and extrinsic apoptosis pathways.
Students should explain how pro- and anti-apoptotic proteins function.
Students should describe how necrosis and apoptosis dysregulation lead to diseases and give examples.

Disclaimer

The lecture format will be different, prioritizing interactive activities.
Lecture slides are provided but will be explained minimally.
Students must use supplemental articles to answer activity questions.
Class notes must be submitted for homework in a specific format.

Activity 1 – Articles Review

Articles aim to discuss types of apoptosis, benefits, and how different pathways function.
Different articles target different audiences (general vs. science-focused).
One article may be easier to understand based on simpler terminology, aiding in clarity for a wider range of learners.
Primary literature presents original findings, while secondary summarizes existing research.

Activity 2 – Apoptosis, Necrosis, and Autophagy

Apoptosis: Programmed cell death, no inflammation, occurs when cell function is complete or the cell is misbehaving. Examples include embryonic development and cancer.
Necrosis: Uncontrolled and passive cell death from injury (lack of oxygen, physical trauma), resulting in inflammation and releasing cell components. An example is a burn victim.
Autophagy: Cellular self-digestion, maintaining cell health, occurring when there is starvation - cell breakdown and recycling of components for new cell function/repair.

Activity 2 - Illustration and Signaling

The illustration metaphorically compares the cell's processes to daily life objects, like a cellular signal.
The author aims to describe cell death pathways' complexity and interplay of external and internal signals in cell processes.
Authors use analogies, such as cell communication or the signal transmission, to convey the pathways and steps involved.

Activity 2 - Proteases & Cell Death in 1996

ICE-like proteases are now more commonly called caspases.
Caspase-1 is part of this proteolytic family.

Apoptosis, Necrosis, and Autophagy (Continued)

Morphological features (cell size, cytoplasmic density, organelle distribution): used to distinguish between apoptosis, necrosis, and autophagy.
Morphological features for each - cell death process
- Apoptosis: smaller, dense cytoplasm, condensed organelles
- Necrosis: large, swollen cells
- Autophagy: unchanged cytoskeleton, vacuoles
Process importance: Understanding differences is vital for applying appropriate interventions or treatment.

Additional Activity 2 Information

This section includes descriptions of each process, how to distinguish them, and why these processes matter.
Key morphological changes in each process to aid in identification
Apoptosis: cell shrinkage, chromatin condensation
Necrosis: cell swelling, loss of membrane integrity
Autophagy: compartmentalization, organelles surrounded by membranes.

Activity 3 – Cell Death/Flow Cytometry

PS(phosphatidylserine) flips to the outer membrane, making the cell green in labeling; PI or 7AAD uptake by dead cells gives pink color
This helps distinguish dead from living cells and early and late apoptotic/necrotic stages through cellular staining.
Quantify apoptotic and necrotic cells using FACs (fluorescence activated cell sorter) data/plots

Activity 3 – Cell Death/Flow Cytometry (Continued)

Flow cytometry uses fluorescent dyes (annexin V, PI, 7AAD) to categorize cells based on their apoptosis status.
Analyzing flow cytometry graphs helps clinicians and researchers understand cell death populations and stages.

Activity 3- Organelle Identification

Microscopic (EM/Electron Microscopy) images depict organelles within a cell, aiding cellular processes that involve the degradation and recycling of organelles.
Autophagy shows organelles within double membranes, used for cell digestion and component recycling/regeneration.

Activity 4 – Dysregulated Apoptosis

Excessive apoptosis can lead to diseases like AIDS, type 1 diabetes, Alzheimer's, Parkinson's.
Excessive apoptosis is seen when cells "die too quickly" thus affecting function.
Insufficient apoptosis leads to uncontrolled cell growth and cancer (overexpression of Bcl2).

Muddiest Point

The student found the intrinsic pathway description to be challenging.

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Description

Test your knowledge on apoptosis processes and FACS analysis with this quiz. Explore key concepts like Annexin V binding, cell viability interpretation, and the impact of chemotherapy on cell fate. Ideal for students studying cell biology and related fields.