Utjecaj agonista TLR receptora na vijabilnost THP-1 humanih monocitnih stanica

master's thesis

• Title (english): Effect of TLR receptor agonists on THP-1 cell viability
• Author: Tina Smoljanović
• Mentor: Lada Rumora (mentor)
• Committee member: Lada Rumora (predsjednik povjerenstva)
• Committee member: Anita Somborac Bačura (član povjerenstva)
• Committee member: Petra Turčić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of medical biochemistry and haematology); Zagreb
• Defense date and country: 2017-07-07, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Medical Biochemistry

Abstract

Kronična opstrukcijska plućna bolest karakterizirana je pojačanom kroničnom upalom dišnih putova i pluća izazvanom stranim česticama i plinovima koja dovodi do stvaranja ispljuvka i razaranja plućnog parenhima. TLR receptori uključeni su u regulaciju urođenog i stečenog imuniteta te imaju ključnu ulogu u održavanju homeostaze pluća. Važni su u obrani od patogena, ali ne dozvoljavaju razvijanje pretjeranog upalnog odgovora koji bi mogao dovesti do oštećenja plućnog tkiva. Smatra se da cigaretni dim i drugi štetni čimbenici, u kombinaciji s genskom predispozicijom, djeluju na TLR receptore, dovode do njihove aktivacije, nakupljanja stanica imunosnog sustava u većem broju u dišnim putovima, otpuštanja upalnih citokina te promjena u ekspresiji gena za TLR receptore što pridonosi razvoju kronične upale.
U ovom istraživanju ispitivala se toksičnost različitih koncentracija agonista TLR receptora LPS-a (agonist TLR4) i LTA (agonist TLR2) na THP-1 humanim monocitnim stanicama. Na taj se način pokušao oponašati upalni odgovor u bolesnika s KOPB-om. Vijabilnost stanica određivala se tripanskim plavilom, određivanjem katalitičke aktivnosti LDH i MTS testom.
Dobiveni rezultati su pokazali da 0,1 μg/ml, 0,5 μg/ml, 1 μg/ml i 5 μg/ml LPS i LTA uzrokuju porast aktivnosti LDH-a u mediju iznad stanica, odnosno dovode do gubitka integriteta stanične membrane. Prema MTS testu, taj gubitak integriteta stanične membrane ne utječe značajno na metaboličku aktivnost stanica. Stoga zaključujemo da LPS i LTA ipak ne djeluju citotoksično na THP-1 stanice u zadanim koncentracijama.

Abstract (english)

Chronic obstructive pulmonary disease is characterized by enhanced chronic airway and lung inflammation caused by inhalation of particles and gases which leads to sputum production and destruction of lung parenchyma. TLR receptors are involved in regulation of innate and acquired immunity and have a key role in maintenance of lung homeostasis. They play important part in defending against infections, but prevent excessive inflammatory response which could lead to lung destruction. It is believed that cigarette smoke and other risk factors, combined with genetics, affect TLR receptors, activate them and lead to enhanced recruitment of immune cells in the airway, pro-inflammatory cytokine production and changes in TLR receptor gene expression which contributes to development of chronic inflammation.
In this research we investigated toxicity of different concentrations of TLR receptor agonists LPS (TLR4 agonist) and LTA (TLR2 agonist) on THP-1 human monocytic cells by mimicking inflammatory responses in COPD patients. Cell viability was determined with Trypan blue, LDH and MTT assays.
According to the results obtained, 0.1 μg/ml, 0.5 μg/ml, 1 μg/ml i 5 μg/ml LPS and LTA cause increase of LDH activity in cell medium and lead to a significant loss of cell membrane integrity. On the other hand, MTS assay showed that the metabolic activity of cells was not significantly altered despite of disturbed cell membrane integrity. Therefore we can conclude that LPS and LTA do not provoke a significant cytotoxic effect on THP-1 cells.

• Keywords: lipopolisaharid; lipoteikoična kiselina; TLR receptori; kronična opstrukcijska plućna bolest; THP-1 stanice; vijabilnost
• Keywords (english): lipopolysaccharide; lipotheicoic acid; TLR receptors; chronic obstructive pulmonary disease; THP-1 cells; viability
• Language: croatian
• URN:NBN: urn:nbn:hr:163:111975
• Study programme: Title: Pharmacy; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra farmacije (magistar/magistra farmacije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-11-16 12:27:48