Aspirin and Plants

Question 1

Willow (uses)

Answer 1

• Salix species
• indegenous Americans use for colds, influenza, indigestion
• Sumerian clay tablets descrive willow bark for use in rheumatism
• ancient egyptians use lead and bark as painkiller
• chinese doctors use by 500BC
• Hippocrates uses for women in labor for aches and fever

Question 2

Reverend Edward Stone (1763)

Answer 2

• has epiphany while relaxing under a willow tree
• bitter taste of bark reminds him of quinine used for malaria
• performs experiments on willow bark to see if it can be used to also treat Malaria

Question 3

Chemical Compounds found in Willow

Answer 3

• astringent quality due to a glycoside called salicin
• salicin breaks down into salicylic acid and sugar in the intestine and liver

Question 4

Felix Hoffman (Bayer, 1897)

Answer 4

• german chemist
• finds that the addition of an acetyl group to salicylic acid reduces its irritant properties in the gut

Question 5

Why is acetylsalicylic acid named Aspirin?

Answer 5

• “A” : represents the added acetyl group
• “spir” : is derived from Spiraea ulmaria (meadowsweet), which yields salicin
• “in” : common suffic used for drugs at the time

Question 6

Aspirin use in Modern Medicine

Answer 6

• 1998: used to treat hypertension with the greatest benefit in preventing heart attacks
• 2005: primary prevention tool to reduce risk of stroke increasing risk of heart attack or death from cardiovascular causes
• 2013/2014: reduces risk of colorectal, stomach and oesophageal cancers

Question 7

Where does Salicylic acid accumulate?

Answer 7

• in both local and systemic tissues (not the systemic signal)
• has a number of analogs/conjugates

Question 8

Salicylic Acid signalling

Answer 8

• triggers a change in the cells redox potential
• this causes the translocation of non-expresser of PR1 (NPR gene) to the nucleus to activate SA-regulated genes

Question 9

SA ‘time-of-day’ expression

Answer 9

• peaks in dark
• consistent with expression of genes affecting SA levels which also show circadian oscillations
• occurs even when there is no pathogen attack
• under pathogen attack clock can be regulated by NPR1

Question 10

Hypersensitive Response (HR)

Answer 10

1. burst of oxygen reactive species around infection site
2. synthesis of antimicrobial phytoalexins
3. accumulation of SA
4. directly kill and damage pathogens
5. strengthen cell walls, and trigger apoptosis
6. restrict pathogen from spreading
7. rapid and local

Question 11

Systemic Acquire Resistance (SAR)

Answer 11

1. secondary response
2. systemic
3. broad-range resistance
4. leads to pathogenesis related (PR) gene expression
5. signals: SA to JA, ethylene

Question 12

What are potential approaches to figure out what enzymes are responsible for producing SA?

Answer 12

• radiolabelling studies
• genetic studies
• correlative (genomic) studies
• expression of genes/enzymes in a heterologous system
• gene silencing (VIGs) or genome editing (Crispr- CAS9)
• putting in new pathways which increase flux or siphon off compound into another pathway

Question 13

Pathways of SA biosynthesis in plants?

Answer 13

1. isotope feeding experiments suggest that plants synthesize SA from cinnamate produced by PAL
2. genetic studies have indicated that the bulk of SA is produced from isochorismate
3. genes PBS3 and EPS1 are identified as important for pathogen-induced SA production

Question 14

Mutants affecting SA synthesis in elevated SA accumulation

Answer 14

mpk4 (MAP protein kinase 4): constitutive SA accumulation

Question 15

Mutants affecting SA synthesis in reduced SA accumulation

Answer 15

• eds1 (enhanced disease susceptibility 1): lipase homolog - immune receptor
• pad4 (phytoalexin deficient 4): another lipase homolog
• sid1 and sid2 (salicylic acid induction-deficient): defects in chorismate pathway

Question 16

Usage and Value of ASA

Answer 16

• ASA valuable in the discovery of the mechanism of COX 1 and 2 and the newer Cox inhibitors which can be specific to one variant or the other
• COX enzymes produce prostaglandins
• popularity of ASA declined in the 1960s as other drugs (ibuprofen and acetiminophen) came on line
• ASA is a NSAID

Question 17

What are Eicosanoids?

Answer 17

signalling molecules created from the breakdown of arachidonic acid

Question 18

Properties of COX-1 enzymes

Answer 18

• located in the kidney, stomach and platelets
• expression: constitutive (activated by physiologic stimuli)
  role: “housekeeping” and maintenance

Question 19

Properties of COX-2 enzymes

Answer 19

• located in macrophages, leukocytes and fibroblasts
• expression: inducible by pro-inflammatory stimuli
• role: pro-inflammatory and mitogenic functions

Question 20

COX-1 interacting with ASA

Answer 20

• aspirin is unique as an inhibitor as it irreversibly inactivates COX1 by acetylating the serine of the enzyme
• low doses inhibit COX-1 for the lifetime of the platelet
• higher doses inhibits both isoforms of COX

Question 21

Issues and Potential side effects with COX1 interacting with ASA

Answer 21

• rapidly metabolized and rendered inert by the body
• efficacy reduced by other drugs
• can trigger asthma
• GI upset
• large doses - salicylism : vertigo, vomiting and hearing loss
• extreme doses: CNS failure, coma, death

Question 22

What is the common precursor for the five principal prostaglandins

Answer 22

PGH2.

Question 23

Biological roles of prostaglandins

Answer 23

• aggregation of platelets
• induction of labor
• regulate cell growth
• regulate fever

Question 24

Discovery of Prostaglandins

Answer 24

• 1930 Kurzrok & Lieb
• 1936 von Euler: Thought that the acidic lipid was produced in prostate, Called them: PROSTAGLANDINS
• 1960-1962 Vane, Bergstorm and Samuelsson: Find the effect is from several closely related compounds and Elucidated structures of PGE1 & PG F2α from sheep
• 1964 Bergstorm : Biosynthesis of PGE2
  from Arachidonic acid

Question 25

Medicinal 5’-Lipoxygenase Inhibitors

Answer 25

1. baicalein: root - scutellaria laterioflora
2. caffeic acid: bark (Eucalyptus globulus), freshwater fern (Salvinia molesta) , mushroom (Phellinus linteus)
3. Curcumin : Curcuma longa (tumeric)
4. Hyperforin: Hypericum perforatum (St John’s wort)

Question 26

Prostaglandin Receptors

Answer 26

• ligand binds = exchange of GTP for GDP
• activated G-protein forms separate alpha and beta-gamma complexes
• both active forms move along PM to activate/inhibit down-stream targets
• eventually, the alpha subunit hydrolyzes GTP to GDP and resets the process

Question 27

Effect of Painkillers on COX enzymes (NSAIDs vs. Acetaminophen)

Answer 27

• NSAIDS targeted inhibition of COX1/2 synthesis of prostaglandins (anti-inflammatory capabilities
• acetaminophen targeted inhibition of COX 2/3 synthesis of prostaglandins (no anti-inflammatory effects)

Question 28

Acetaminophen Mechanism of Action

Answer 28

1. acetaminophen enters system
2. penetrates bbb
3. blocks cycloxygenase (COX3) in brain
4. blocks the formation and release of PGE in the CNS
5. inhibit the action of endogenous pyrogens on the heat-regulating centers in the brain
6. antipyretic effects