Usporedba učinka normoksije i hipoksije na astrocite miša

master's thesis

• Title (english): Comparison of the effect of normoxia and hypoxia on mouse astrocytes
• Author: Paula Stančin
• Mentor: Dinko Mitrečić (mentor)
• Mentor: Dubravka Hranilović (sumentor)
• Committee member: Dubravka Hranilović (predsjednik povjerenstva)
• Committee member: Sandra Hudina (član povjerenstva)
• Committee member: Mirta Tkalec (član povjerenstva)
• Committee member: Sofia Ana Blažević (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2020-07-20, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Hipoksija se nalazi u patofiziološkoj pozadini nekih od najčešćih bolest čovjeka, kao što su srčani i moždani udar. Budući da osim živčanih stanica, neurona, u normalnom funkcioniranju živčanog sustava brojne uloge imaju i astrociti, u ovom istraživanju istražen je izražaj dva intermedijarna filamentna citoskeletna proteina astrocita: nestina, koji se smatra markerom živčanih matičnih stanica i glijalnog fibrilarnog kiselog proteina (GFAP) koji je marker zrelih astrocita. Izražaj je istražen u uvjetima normoksije (21% kisika) i hipoksije (1% kisika). Također je cilj bio usporediti primjenu dva programa za poluatomatsku analizu slika, Cell profiler i Fiji u obradi jačine imunohistokemijskog signala. Nakon 8 dana diferencijacije od živčanih matičnih stanica su dobiveni zreli astrociti. Dio stanica je premješten u inkubator s 1 % kisika na 24 sata, dok je drugi dio stanica ostao u inkubatoru na ambijentalnoj koncentraciji kisika kao kontrola. Imunhistokemijska metoda je pokazala kako je nestin najviše izražen na početku diferencijacije, a dalje postupno opada. Hipoksija je statistički značajno smanjila izražaj nestina. Istovremeno, izražaj GFAP-a raste sa stupnjem diferencijacije astrocita. Hipoksija nije djelovala na izražaj GFAP. Usporedba rezultata mjerenja jačina imunohistokemijskog signala dobivenih računalnim programima Cell profiler i Fiji je pokazala kako oba programa daju gotovo identične razine statističkih razlika između uspoređivanih skupina.

Abstract (english)

Hypoxia stands in the pathophysiological background of some of the most common human diseases, such as heart attack and stroke. Since in addition to neurons, astrocytes also play a number of roles in the normal functioning of the nervous system, in this paper is investigated the expression of two intermediate filamentous cytoskeletal proteins of astrocytes: nestin, which is considered a marker of neural stem cells and glial fibrillar acidic protein (GFAP) which is a marker of mature astrocytes. Expression was investigated under conditions of normoxia (21% oxygen) and hypoxia (1% oxygen). In addition, the aim of this study was to investigate the application of two programs for semi-automatic signal analysis, Cell profiler and Fiji and to compare the results of quantification of the signal obtained by immunohistochemistry. After 8 days of differentiation from nerve stem cells, mature astrocytes were obtained. Half of these cells were transferred to an incubator with 1% oxygen for 24 hours while another part of the cells remained in the incubator at ambient oxygen concentration as a control. The immunohistochemical method revealed that the expression of nestin peaked at the beginning of differentiation and then gradually decreased. Hypoxia influenced expression of nestin and it was reduced. At the same time, the expression of GFAP increased in parallel to astrocyte differentiation. Hypoxia did not affect GFAP expression. Comparison of the results of immunohistochemical signal measurements obtained by the computer programs Cell profiler and Fiji showed that both programs give almost identical levels of statistical differences between the compared groups.

• Keywords: GFAP; nestin; astrociti; imunohistokemija; ishemija
• Keywords (english): GFAP; nestin; astrocytes; immunohistochemistry; ischemia
• Language: croatian
• URN:NBN: urn:nbn:hr:217:108582
• Study programme: Title: Experimental Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra eksperimentalne biologije (magistar/magistra eksperimentalne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2020-08-07 09:04:50