Pyocyanin (Synonyms: Sanasin,Sanazin,Pyocyanine)

Cell lines

Human embryonic lung epithelial cell line, L-132

Preparation Method

Cell cultures in 96 well plates were developed by adding 0.2 ml cell suspension in growth medium containing approximately 5 × 105 cells ml−1 and incubating for 12 h at appropriate temperature.

Reaction Conditions

Pyocyanin was prepared with different concentrations in growth medium. Then added to the wells to attain final strength ranging from 6.25 to 200 µg ml−1 for XTT (mitochondrial activity), neutral red up take (plasma membrane damage) and SRB (protein synthesis), 0–200 µg ml−1 for LDH and H2O2, and 25–200 µg ml−1 for glucose consumption in triplicate for each concentration. Incubate for 24 h.

Applications

The cytotoxicity of pyocyanin could be assessed by this experiment, facilitating pyocyanin’s safe usage toxicity. Higher concentrations of pyocyanin (175 and 200 mg l−1) only caused significant morphological changes such as clumping, and necrosis as visualized microscopically in L-132 cell line. Human cell membrane was found to be susceptible to oxidative damage induced by pyocyanin.

Animal models

Male C57BL/6J mice (8-10 weeks, 20-30 g)

Preparation Method

Mice were housed under controlled laboratory conditions, maintained on a 12 h day and night cycle. Then mice were acclimatized for 5 days to the lab conditions and handling prior to the actual beginning of the experiments. Mice were administered 0.9% saline as control, and pyocyanin by intranasal route. Another group was treated with LPS (P.aeruginosa) by intraperitoneal route to mimic the effects of ongoing infection on tissue permeability, and pyocyanin was instilled by intranasal route 3h after LPS injection.

Dosage form

Pyocyanin (50 µg/50 µL in 0.9% saline); 0.9% saline (50 µL); LPS (3 mg/kg)

Applications

Pyocyanin could be diffused into systemic circulation, which was not influenced by the pre-exposure to pseudomonal infestation. This experiment detected the plasma concentration of intranasally administered Pyocyanin. Furthermore, localized administration of Pyocyanin was able to elicit changes to behavior and a systemic pro-inflammatory and pro-oxidant effect.

References:

[1]. Priyaja P, et al. Pyocyanin induced in vitro oxidative damage and its toxicity level in human, fish and insect cell lines for its selective biological applications. Cytotechnology. 2016 Jan;68(1):143-155.

[2]. Arora D, et al. Pyocyanin induces systemic oxidative stress, inflammation and behavioral changes in vivo. Toxicol Mech Methods. 2018 Jul;28(6):410-414.