9. motako subtilisina/kexina proproteina konbertasaren (PCSK9) inhibizioaren eragina koloneko minbizi-zelula ametan (MZA)

Abstract

[EUS] Minbizi-zelula amak (MZA) minbizi askoren erresistentziaren eta berragerpenen atzean daudela uste da, eta beraz, horiek aztertzea ezinbestekoa da etorkizunean tratamendu eraginkorrak sortzeko. Munduan minbiziak eragiten dituen heriotzetatik, kolon-ondestekoa bigarrena da maiztasunari dagokionez. Halaber, minbizi horren zelula parentaletan PCSK9k babes funtzioa duela frogatu da. Horrez gain, koloneko MZAetan PCSK9aren gainadierazpena ikusi da. Ikerketa honetan, MZAen hazkuntzan PCSK9ak zer eragina duen aztertu nahi izan da. Horretarako CRISPR/Cas9 bidez PCSK9a erauzitako SW620 zelulak aztertu dira eta diluzio klonala egin ondorengo kolonia puruetatik abiatuz, zelulen edizioa PCR eta elektroforesiz bermatu da. Azkenik, PCSK9 gabeko SW620 zeluletatik MZAk sortu dira kolonosferak sortuz eta ondoren, horien proliferazioa aztertu da XTT saio kolorimetrikoa erabiliz. Tamalez, ikerketa taldearen kontroletik haratago egon diren arazo logistikoek, ikerketa geldiarazi dute. Hala ere, egindako ikerketa bibliografikotik, hepatokartzinomaren PCSK9aren adierazpena aldakorra izan daitekeela ikusi da. Dirudienez, minbiziaren fase goiztiarretan PCSK9aren adierazpena gutxituta dago eta fase aurreratuetan ordea, handituta. Bestalde, talde honen aurreko ikerketan PCSK9rik gabeko kolon-ondesteko SW620 zelula parentaletan hazkuntza moteltzen dela ikusi bada ere, LDL aberatseko medioan edo PCSK9 aberatseko medioan hazteak izan ditzaken ondorioak aztertzearen beharra aldarrikatu da. Gainera, SW620 zelulen jatorri metastasikoak lortutako emaitzetan eragina izan dezakeela ere arrazoitu da. Azkenik, literatura zientifikoa eta ikerketa taldearen aurreko ikerketetatik abiatuz, MZAetan PCSK9 ezak hazkuntza moteldu beharko zuela teorizatu da. Hala ere, ikerketa gehiagoren beharra dago (in vitro zein in vivo) konbertasak minbizian dituen funtzio zehatzak aztertzeko, bai PCSK9ren portaera eta adierazpenean, bai zelula parental zein MZAetan.

[EN] Cancer stem cells (CSCs) are an important reason for the resistance to treatment and recurrence of many cancers. As such, studying CSCs is essential to create effective future treatments. Colorectal cancer (CRC) causes many deaths, ranking second when taking stock of worldwide cancer deaths. In CRC, PCSK9 has been linked to increased survival. Moreover, an overexpression of PCSK9 has been observed in CSCs. This study set out to analyse the impact of PCSK9 on the growth of the CSCs. To do this, SW620 cells were used and PCSK9 was deleted using CRISPR/Cas9. After obtaining pure colonies through clonal dilution, we verified that PCSK9 had been deleted using PCR and electrophoresis. Finally, after we developed colonospheres to create CSCs from the edited PCSK9 SW620 cells, growth was analysed using the XTT colorimetric assay. Unfortunately, for logistical reasons beyond our control, we had to halt the investigation. Nevertheless, we theorise, based on the bibliographic study we carried out, that PCSK9 expression in hepatocarcinoma may vary from early stages to advanced stages of cancer. In early stages, PCSK9 expression seems to be decreased; whereas in advanced stages, PCSK9 seems to be overexpressed. In addition, we claim that the effects of PCSK9-deleted colorectal parental cells should be monitored in an LDL-enriched medium or in a PCSK9-enriched one to find out whether growth halts, as previously reported. We also find out that the metastatic origin of the SW620 parental cells could have influenced those results. Finally, based on the existing literature and data from previous studies, we theorise that non-PCSK9 CSCs’ growth should have been impeded if the investigation would have been concluded. Nevertheless, the exact function of the convertase in cancer needs further study (in vitro and in vivo) to account for PCSK9’s varying behaviour and expression in both parental cells and CSCs.