Lecture 15- Clocks and Immunity I

Question 1

Innate immunity

Answer 1

First line of defense; non-specific and rapid, same response on each encounter with antigen
E.g. external barriers, receptors that recognise a broad spectrum of pathogens

Question 2

Adaptive immunity

Answer 2

Acquired immunity; creates immunological memory; pathogen specific; i.e lifelong immunity after measles infection (basis of vaccinations)

Question 3

Cells involved in innate immunity

Answer 3

Macrophages, mast cells

dendritic cell, granulocytes, natural killer cells

Question 4

Cells involved in adaptive immunity

Answer 4

B cell, T cell

Question 5

Cells involved in both kinds of immune response

Answer 5

γδ T cell, natural killer T cell

Question 6

Macrophages (overview)

Answer 6

Phagocytic cells;
Detect, engulf and destroy pathogens; Antigen presenting cells; Initiate inflammation via release of cytokines (e.g. tumour necrosis factor alpha, TNF⍺) that activate other cells

Question 7

LPS into peritoneal cavity of rat, collecting peritoneal macrophages (2hrs) and performing RNA analysis shows…

Answer 7

… time of day affects macrophage responses to LPS (IL6 + IL2 higher at CT12 than CT0)

Question 8

LPS into peritoneal cavity of rat, collecting blood sample (4hrs) and performing ELISA…

Answer 8

…quantifies cytokine response in serum –> time of day affects systemic responses to LPS (again IL6 + IL2 higher at CT12)

Question 9

Using Cre-lox system to specifically delete clocks in macrophages (LysM-Bmal1-/-)…

Answer 9

…renders mice macrophages arhythmic while mice remain behaviourally rhythmic

Question 10

Deletion of the macrophage clock…

Answer 10

…abolishes time of day response

Question 11

Endotoxin dosing

Answer 11

The same dose of endotoxin survived by most animals in the middle of subjective night is highly lethal if given 8-12 hours earlier or later

Question 12

Rev-erb alpha -/-

Answer 12

No rhythm in IL6 response –> mediates circadian regulation of macrophage responses

Question 13

Mast cell overview

Answer 13

Type of granulocyte, full of granules which contain inflammatory mediators, such as histamine; Histamine is a vasoactive amine, increasing blood vessel diameter; Important in defense against parasitic infection and allergic reactions

Question 14

Mast cell activates…

Answer 14

Histamine, cytokines (IL4, IL13, TNF alpha) and lipid mediators (leukotrienes, prostaglandins)

Question 15

Histamine (blood vessels and smooth muscle contraction)

Answer 15

Increases vascular permeability; Smooth muscle contraction

Question 16

IL4 + IL13

Answer 16

Promotes Th2 differentiation; Promotes IgE production

Question 17

Leukotrienes and prostaglandins

Answer 17

Increases vascular permeability; Causes smooth muscle contraction; Stimulates mucus secretion; Chemoattractant for T cells, eosinophils, mast cells and basophils

Question 18

Histamine release in Clock delta 19 mice vs WT

Answer 18

Is generally lower and lacks the rhythm that remains consistent +/- serum shock in WT (though obvs raised + shock) –> histamine release from mast cells in circadian

Question 19

How can the clock be disrupted?

Answer 19

Environmental: lighting, feeding, jet lag, shift work

Genetic (via targeting): deletion of core clock genes (Bmal1 deletion in macrophages, Clock mutation in mast cells)

Question 20

Jet-lag protocol in mice

Answer 20

6 hour phase advance by shortening dark period once/week;

Show increased pro-inflammatory response to LPS challenge

Question 21

Shiftwork and disease risk

Answer 21

Circadian misalignment;
Increased risk of cancer, metabolic disease and obesity, chronic inflammatory disease, cardio vascular disease, gastro-intestinalproblems

Question 22

IL6 in circadian misalignment

Answer 22

Increased circulating IL6 in human volunteers

Question 23

Immune homeostasis

Answer 23

Natural turnover of immune cells and searching for pathogens

Question 24

Lymphocyte trafficking (immune homeostasis)

Answer 24

Lymphocytes (white blood cells) T cells and B cells; Survey antigen by circulating through blood
and lymph;
In mouse, lymphocyte numbers peak ZT13 (start of active phase) (previously thought to be continuous release)

Question 25

S1pr1

Answer 25

Induces lymphocyte egress, expression higher during the day

Question 26

CCR7

Answer 26

Induces lymphocyte homing, expression higher during the night

Question 27

EAE model

Answer 27

experimental autoimmune encephalitis, animal model of brain inflammation; Mice immunized with myelin oligodendrocyte glycoprotein (MOG); Results in brain and spinal cord demyelination and eventually
paralysis; Depends on T cells responding to MOG within the lymph nodes, mounting inflammatory response

Question 28

EAE circadian response

Answer 28

At ZT8, LN T cell number highest and thus increased disease severity weeks later (lowest at ZT20)