Imunohistokemijsko prikazivanje molekula signalnog puta GABA-E izvan središnjeg živčanog sustava

master's thesis

• Title (english): Immunohistochemical visualization of GABA signaling pathway moleculs in non-neural tissues
• Author: Ana Bucat
• Mentor: Marija Ćurlin (mentor)
• Committee member: Srećko Gajović (predsjednik povjerenstva)
• Committee member: Dinko Mitrečić (član povjerenstva)
• Committee member: Marija Ćurlin (član povjerenstva)
• Granter: University of Zagreb; School of Medicine; (Department of Histology and Embryology); Zagreb
• Defense date and country: 2015-07-15, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Basic Medical Sciences; Cytology, Histology and Embryology

Abstract

Imunohistokemija je vrlo koristan alat u otkrivanju prisutnosti i uloge određenih bjelančevina u stanicama i tkivima. Za vjerodostojnu interpretaciju rezultata imunohistokemijske analize potrebno je postupak provesti na uzorcima koji će sigurno dati pozitivan rezultat (pozitivna kontrola) i na uzorcima koji će sigurno dati negativan rezultat (negativna kontrola). Pozitivna kontrola posebno je važna u slučaju negativnog imunohistokemijskog signala na ispitivanom tkivu. U istraživanju uloge neurotransmitera GABA (gama-aminomaslačna kiselina) u regulaciji izlučivanja sluzi u endocervikalnim sekrecijskim stanicama provedeno je imunohistokemijsko ispitivanje prisutnosti molekula signalnog puta GABA u tkivu humanog vrata maternice. Neočekivano je ova analiza dala negativan rezultat za GAD (engl. Glutamate decarboxylase) i VGAT (eng. Vesicular GABA transporter) u tkivu cerviksa. Zbog nedostatka pozitivne kontrole taj se rezultat nije mogao vjerodostojno interpretirati. ----- Cilj ovog rada bio je dobiti pozitivan signal za GAD i VGAT u tkivu za koje je poznato da se u njemu nalaze ove molekule. Za tu svrhu odabrana su tkiva mozga, pluća i testisa miša. Bilo je potrebno prilagoditi imunohistokemijski postupak za dokazivanje GAD i VGAT u parafinskim rezovima tkiva obzirom da se istraživanje provodi na parafinskim rezovima humanog tkiva cerviksa. Pozitivan rezultat imunohistokemijskog prikazivanja GAD i VGAT u tkivu mozga i testisa pokazao je da ova tkiva mogu poslužiti kao pozitivna kontrola za imunohistokemijsku analizu prisutnosti tih molekula u parafinskim rezovima tkiva. Konačno, imunohistokemijska analiza humanog tkiva cerviksa pokazala je negativan signal za GAD i VGAT u tom tkivu, dok je za GABA receptor signal bio pozitivan. Uz pozitivne kontrole uspostavljene ovim radom i odgovarajuću negativnu kontrolu za GABA receptor, rezultate tog pokusa možemo smatrati vjerodostojnima.

Abstract (english)

Immunohistochemistry is a very useful tool in the localization and functional characterization of specific proteins in cells and tissues. For an authentic interpretation of the results from the immunohistochemical analysis, it is necessary to carry out the process on the samples that would certainly give a positive result (positive control), and the samples that will certainly give a negative result (negative control). The positive control is particularly important in the case of negative immunohistochemical signal of the examined tissue. In the study of the role of neurotransmitter GABA (gamma-aminobutyric acid) in the regulation of secretion of mucus in endocervical secretory cells, an immunohistochemical examination of the presence of molecules of GABA signalling pathway in human cervical tissue was conducted. Unexpectedly, this analysis gave a negative result for GAD (Glutamate decarboxylase) and VGAT (Vesicular GABA transporter) in the cervix tissue. Due to the lack of positive controls, that result could not have been credibly interpreted. ----- The aim of this study was to get a positive signal for GAD and VGAT in the tissue where it’s known that it contains these molecules. For this purpose the selected tissues were brain, lung and testes of the mouse. It was necessary to adapt the procedure for immunohistochemical detection of GAD and VGAT in paraffin sections of tissue, since the research was carried out on paraffin sections of human tissue of the cervix. Positive immunohistochemical result showing GAD and VGAT in brain and testes tissues showed that this tissue may be used as positive controls for immunohistochemical analysis of the presence of these molecules in paraffin sections of tissue. Finally, immunohistochemical analysis of human cervical tissue showed a negative signal for GAD and VGAT in the tissue, while the GABA receptor signal was positive. With positive control established in this study and the corresponding negative control for the GABA receptor, the results of this experiment can be reliable.

• Keywords: imunohistokemijska analiza; neurostransmiter GABA (gama-aminomaslačna kiselina); imunohistokemijsko ispitivanje; GAD (Glutamate decarboxylase); VGAT (Vesicular GABA transporter)
• Keywords (english): immunohistochemical analysis; neurotransmitter GABA (gamma-aminobutyric acid); immunohistochemical examination; GAD (Glutamate decarboxylase); VGAT (Vesicular GABA transporter)
• Language: croatian
• URN:NBN: urn:nbn:hr:105:758272
• Study programme: Title: Medicine; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: doktor/doktorica medicine (doktor/doktorica medicine)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Public note: Pohraniteljica objekta dodala ključne riječi.
• Created on: 2016-08-03 11:39:22