Unveiling the mechanisms of breast cancer pathogenesis by proteomics methods

Patogenezu složene i heterogene bolesti kao što je to rak dojke nije moguće u potpunosti razjasniti klasičnim metodološkim pristupima koji su se koristi li posljednjih desetljeća u molekularnoj medicini. Osobito je važno otkriti nove biljege koji bi pomogli u ranoj detekciji bolesti te služili za razvoj ciljane terapije. Dosadašnji rezultati iz područja istraživanja raka dojke u kombinaciji s globalnim metodama istraživanja statusa ekspresije gena i proteina pokazali su kako je u patogenezu raka dojke uključen ljudski hormon rasta (hGH) te mehanizmi autokrine regulacije preko proteina Pax-5. Također je identi fi ciran čitav niz novih potencijalnih biomarkera kao što su to HOXA1, CHOP SH3GLB1, kazein kinaze, p53, aneksin XI, CDC25C, eIF- 4E i MAP kinaze 7, 14-3-3e, galekti n-1, aneksin-5, aneksin-1, LDH-B, GST-pi, akti n, vimenti n, HSP70, CK18, moezin, SH3GLB1, SUB1, SND1 i TRIM28, osteoponti n i osteonekti nBreast cancer is a complex and heterogenic disease. Classical molecular medical approaches cannot be successfully used to completely understand its pathogenesis. In additi on, fi nding new biological markers that would help in early detecti on and creati on of guided and specifi c therapy is crucial for the future breast cancer management. The results of breast cancer research in combinati on with those obtained by large scale methods that examine the expression status of genes and proteins, have pointed to an important molecule, namely the human growth hormone (hGH), as a major player in the pathogenesis of breast cancer. It acts through the autocrine regulati on system where the acti vati on of Pax-5 gene has been recently confi rmed as well. A large number of new biomarkers, like HOXA1, CHOP SH3GLB1, casein kinase, p53, annexin XI, CDC25C, eIF-4E, MAP kinase 7, 14-3-3e, galecti n-1, annexin-5, annexin-1, LDH-B, GST-pi, acti n, vimenti n, HSP70, CK18, moesin, SH3GLB1, SUB1, SND1 i TRIM28, osteoponti n and osteonecti n, have been recently identi fi ed as well

Evaluation of the Antioxidant Capacity, Antimicrobial and Antiproliferative Potential of Fir (Abies alba Mill.) Honeydew Honey Collected from Gorski kotar (Croatia)

U radu su ispitani antiproliferativni, antimikrobni i antioksidacijski učinci medljikovaca jele (Abies alba Mill.) prikupljenih s planinskog područja Hrvatske (Gorski kotar) a u svrhu potencijalne primjene umjesto standardnih antibiotika i kemoterapeutika. Preživljavanje stanica, annexin V test i protočna citometrijska analiza poslužili su za procjenu antiproliferativnog učinka na indukciju apoptoze i smrti humanih tumorskih staničnih linija: HeLa, MCF-7, SW620, CFPAC-1, MIA PaCa-2 i normalnih diploidnih humanih fibroblasta (BJ). Antimikrobna je aktivnost testirana na različitim sojevima bakterija Staphylococcus i Acinetobacter uporabom difuzijske i mikrodilucijske metode. DPPH˙ testom određena je sposobnost uklanjanja radikala, dok je matematičkim modelima proučavana kinetika inhibicije DPPH˙. Uočen je antiproliferativni učinak na sve ispitivane stanične linije kao i na normalne diploidne fibroblaste (BJ), kolorektalni adenokarcinom (SW620, metastatski) i adenokarcinom epitela dojke (MCF-7, metastatski). Mehanizmi antiproliferativnog učinka uključuju nakupljanje stanica u sub-G1 fazi u svim ispitivanim stanicama te indukciju apoptoze osobito u SW620 i MCF-7 stanicama. Antibakterijski testovi pokazuju da su rezistentni sojevi obaju bakterija, uključujući i multirezistentni soj A. baumannii ATCC® BAA-1605™, bili osjetljivi na sve ispitane uzorke meda. Proučavanjem kinetike uklanjanja radikala dokazano je da antioksidansi prisutni u medu posjeduju različite sposobnosti hvatanja radikala, da reagiraju s njima različitim brzinama i time uzrokuju dva stupnja reakcije. Rezultati istraživanja pokazuju da hrvatski medljikovac ima terapeutski potencijal zbog snažne biološke aktivnosti te može služiti u zaštiti ljudskog zdravlja.The paper examines the antiproliferative, antimicrobial and antioxidative effects of fir (Abies alba Mill.) honeydew honey from mountain region of Croatia (Gorski kotar) as a potential replacement for standard antibiotics and chemotherapeutic agents. Cell viability, annexin V assay and flow cytometry analysis served to analyse the antiproliferative effect on, apoptosis induction in and cell death of cancer cell lines: HeLa, MCF-7, SW620, CFPAC-1, MIA PaCa-2 and normal diploid human fibroblasts (BJ). Antimicrobial activity was tested against Staphylococcus and Acinetobacter strains by agar well diffusion and microdilution assays. The DPPH˙ assay determined the radical scavenging activity, while mathematical models helped to evaluate the kinetic data of DPPH˙ inhibition. Antiproliferative effect on all tested cell lines and the prominent effect on normal diploid human fibroblasts (BJ), colorectal adenocarcinoma (SW620, metastatic) and breast epithelial adenocarcinoma (MCF-7, metastatic) was observable. The mechanisms of antiproliferative effect included accumulation of cells in the sub-G1 phase in all tested cells and induction of apoptosis in SW620 and MCF-7 cells predominantly. The antibacterial assays showed that antibiotic resistant strains of both bacteria, including multi-resistant strain A. baumannii ATCC® BAA-1605™, were sensitive to all tested honey samples. Radical scavenging assay suggests that antioxidants present in the honey possess different radical suppressing abilities and that they react at different rates with radicals, thereby causing two steps of reaction. The results of the study indicate that Croatian fir honeydew honey has a therapeutic potential due to the strong biological activity and can serve to protect human health

Independent expression of GASP phenomenon on the mutT gene in enterobacteria Escherichia coli

Cilj istraživanja u ovom radu bio je uočiti pojave međudjelovanja enterobakterija. Proučavane su dvije vrste iz ove obitelji, Escherichia coli i Salmonella enterica serotip Typhimurium. Prednost rasta u stacionarnoj fazi (eng. ˝Growth Advantage in Stationary Phase˝ – GASP) je mehanizam koji bakterijama omogućuje da prežive stres tijekom dugotrajnog izgladnjivanja u svom okolišu. Praćena je dinamika rasta mutanta bakterije E. coli mutT TetR, u zasebnoj kulturi kao i rast u miješavinama. Miješavine su se sastojale od 10 dana starih stanica kulture bakterije E. coli mutT TetR, i mlade (24 sata) bakterijske kulture S. enterica StrR ili E. coli StrR. Dobiveni rezultati pokazuju da mutant bakterije E. coli mutT TetR u kompletnoj hranjivoj podlozi pokazuje slab fenotip GASP, no to nije slučaj za minimalnu podlogu. U minimalnoj podlozi mutant mutT E. coli pokazuje jak fenotip GASP u mješavinama s bakterijskom kulturom S. enterica ili E. coli.The aim of this study was to evaluate the interaction between Enterobacteriaceac. Two species of this bacterial family was studied: Escherichia coli and Salmonella enterica serovar Typhimurium. Growth advantage in stationary phase – GASP is a mechanism, which allows bacteria to survive during periods of stress. We were observing the dynamics of growth of bacteria E. coli mutT TetR, in the separate cultures as well as in the mixed ones. Mixed cultures consist of 10 days old bacteria E. coli mutT TetR, and young, 24 hours old, wild type of bacteria S. enterica StrR or E. coli StrR. Results indicate that bacteria E. coli mutT TetR exhibits weak GASP phenotype in the rich medium, but that is not the case for the minimal medium. Bacteria E. coli mutT exhibits strong GASP phenotype in the minimal medium in the mixed cultures with bacteria S. enterica StrR and bacteria E. coli StrR

21 days

Rad pod nazivom „21 dan“ bavi se predočavanjem života sa opsesivno - kompulzivnim poremećajem koji , uz kompulzije, uzrokuje izrazito nelagodne, zastrašujuće misli, konstantnu sumnju, osjećaj krivnje, te znatno otežava život. Na početku rada daje se uvid u poremećaj putem stručne literature. Slijede ju vlastiti primjeri i iskustvo, te praćenje poremećaja kroz dvije faze, svake u trajanju od 21 dana ( sa razmakom od još 21 dana između njih). Prva faza se temelji na izvanjskom prikazu zabilježenom autoportretima, a druga na unutarnjem, nastalom bilježenjem opsesivnih misli i poremećaju popratnih stanja depresije i anksioznosti. Naposljetku se nalazi tehnička razrada djela. Rad posjeduje i terapeutski učinak, te je priprema za buduće prošireno bavljenje društveno angažiranom umjetnosti.The work under the name „21 days“ is about representing life with obsessive – compulsive disorder which, along with compulsions, causes extremely uncomfortable, frightening thoughts, constant doubt, feelings of guilt and makes life much more difficult. At the beginning of my work, an insight is given into this disorder through professional literature on the subject. The subject is followed by personal examples and experiences along with observing the disorder through two phases, each lasting 21 days (with interval of 21 days between each phase). The first phase is based on the external representation reflected by the self-portraits, while the second phase is based on the internal representation reflected in obsessive thoughts and accompanying feelings of depression and anxiety. Finally, there is a technical elaboration of my work. It also has a therapeutic effect and is a preparation for expanded future engagement with socially engaged art

Independent expression of GASP phenomenon on the mutT gene in enterobacteria Escherichia coli

Cilj istraživanja u ovom radu bio je uočiti pojave međudjelovanja enterobakterija. Proučavane su dvije vrste iz ove obitelji, Escherichia coli i Salmonella enterica serotip Typhimurium. Prednost rasta u stacionarnoj fazi (eng. ˝Growth Advantage in Stationary Phase˝ – GASP) je mehanizam koji bakterijama omogućuje da prežive stres tijekom dugotrajnog izgladnjivanja u svom okolišu. Praćena je dinamika rasta mutanta bakterije E. coli mutT TetR, u zasebnoj kulturi kao i rast u miješavinama. Miješavine su se sastojale od 10 dana starih stanica kulture bakterije E. coli mutT TetR, i mlade (24 sata) bakterijske kulture S. enterica StrR ili E. coli StrR. Dobiveni rezultati pokazuju da mutant bakterije E. coli mutT TetR u kompletnoj hranjivoj podlozi pokazuje slab fenotip GASP, no to nije slučaj za minimalnu podlogu. U minimalnoj podlozi mutant mutT E. coli pokazuje jak fenotip GASP u mješavinama s bakterijskom kulturom S. enterica ili E. coli.The aim of this study was to evaluate the interaction between Enterobacteriaceac. Two species of this bacterial family was studied: Escherichia coli and Salmonella enterica serovar Typhimurium. Growth advantage in stationary phase – GASP is a mechanism, which allows bacteria to survive during periods of stress. We were observing the dynamics of growth of bacteria E. coli mutT TetR, in the separate cultures as well as in the mixed ones. Mixed cultures consist of 10 days old bacteria E. coli mutT TetR, and young, 24 hours old, wild type of bacteria S. enterica StrR or E. coli StrR. Results indicate that bacteria E. coli mutT TetR exhibits weak GASP phenotype in the rich medium, but that is not the case for the minimal medium. Bacteria E. coli mutT exhibits strong GASP phenotype in the minimal medium in the mixed cultures with bacteria S. enterica StrR and bacteria E. coli StrR

21 days

Rad pod nazivom „21 dan“ bavi se predočavanjem života sa opsesivno - kompulzivnim poremećajem koji , uz kompulzije, uzrokuje izrazito nelagodne, zastrašujuće misli, konstantnu sumnju, osjećaj krivnje, te znatno otežava život. Na početku rada daje se uvid u poremećaj putem stručne literature. Slijede ju vlastiti primjeri i iskustvo, te praćenje poremećaja kroz dvije faze, svake u trajanju od 21 dana ( sa razmakom od još 21 dana između njih). Prva faza se temelji na izvanjskom prikazu zabilježenom autoportretima, a druga na unutarnjem, nastalom bilježenjem opsesivnih misli i poremećaju popratnih stanja depresije i anksioznosti. Naposljetku se nalazi tehnička razrada djela. Rad posjeduje i terapeutski učinak, te je priprema za buduće prošireno bavljenje društveno angažiranom umjetnosti.The work under the name „21 days“ is about representing life with obsessive – compulsive disorder which, along with compulsions, causes extremely uncomfortable, frightening thoughts, constant doubt, feelings of guilt and makes life much more difficult. At the beginning of my work, an insight is given into this disorder through professional literature on the subject. The subject is followed by personal examples and experiences along with observing the disorder through two phases, each lasting 21 days (with interval of 21 days between each phase). The first phase is based on the external representation reflected by the self-portraits, while the second phase is based on the internal representation reflected in obsessive thoughts and accompanying feelings of depression and anxiety. Finally, there is a technical elaboration of my work. It also has a therapeutic effect and is a preparation for expanded future engagement with socially engaged art

21 days

Rad pod nazivom „21 dan“ bavi se predočavanjem života sa opsesivno - kompulzivnim poremećajem koji , uz kompulzije, uzrokuje izrazito nelagodne, zastrašujuće misli, konstantnu sumnju, osjećaj krivnje, te znatno otežava život. Na početku rada daje se uvid u poremećaj putem stručne literature. Slijede ju vlastiti primjeri i iskustvo, te praćenje poremećaja kroz dvije faze, svake u trajanju od 21 dana ( sa razmakom od još 21 dana između njih). Prva faza se temelji na izvanjskom prikazu zabilježenom autoportretima, a druga na unutarnjem, nastalom bilježenjem opsesivnih misli i poremećaju popratnih stanja depresije i anksioznosti. Naposljetku se nalazi tehnička razrada djela. Rad posjeduje i terapeutski učinak, te je priprema za buduće prošireno bavljenje društveno angažiranom umjetnosti.The work under the name „21 days“ is about representing life with obsessive – compulsive disorder which, along with compulsions, causes extremely uncomfortable, frightening thoughts, constant doubt, feelings of guilt and makes life much more difficult. At the beginning of my work, an insight is given into this disorder through professional literature on the subject. The subject is followed by personal examples and experiences along with observing the disorder through two phases, each lasting 21 days (with interval of 21 days between each phase). The first phase is based on the external representation reflected by the self-portraits, while the second phase is based on the internal representation reflected in obsessive thoughts and accompanying feelings of depression and anxiety. Finally, there is a technical elaboration of my work. It also has a therapeutic effect and is a preparation for expanded future engagement with socially engaged art

In vitro Antiproliferative and Antimicrobial Activity of the Essential Oil from the Flowers and Leaves of Helichrysum italicum (Roth) G. Don Growing in Central Dalmatia (Croatia)

In the present paper, chemical composition, in vitro antimicrobial and antiproliferative activity of the essential oil from the flowers and leaves of Helichrysum italicum (Roth) G. Don (Central Dalmatia, Croatia) as a potential replacement for standard antibiotics and chemotherapeutic agents was analyzed. Essential oil was isolated by steam distillation and analyzed by GC/MS. Antimicrobial activity was carried out by agar-well diffusion and microdilution assays with Gram (+), Gram (-) bacteria and one yeast. Antiproliferative effect, apoptosis induction and cell death on cancer cell lines: HeLa, MCF-7, SW620, CFPAC-1 and MIA PaCa-2 were analyzed by cell viability and Annexin-V assay as well as by flow cytometric analysis. In essential oil, -pinene, -curcumene and neryl acetate was found as major compounds. The antimicrobial assays, showed that essential oil had weak to moderate antimicrobial potential with S. aureus and S. epidermidis as the most sensitive bacterial strains. Essential oil treatment possessed moderate antiproliferative impact on MCF-7 and HeLa cell lines, while analyzing cell cycle treatment had no significant effect on tested cells except on MIA PaCa-2 with the highest cells increase in sub G1 phase cell cycle. However, treatment caused significantly induction of apoptosis in MCF-7 and HeLa cells, but not in MIA PaCa-2 cells. In this cell line, multiple cell death mechanism existed with involving apoptosis, senescence or necrosis. Results of the study provide a promising basis for the evaluation of the potential use of essential oil from H. italicum (Roth) G. Don as a source of alternative nature antibiotics or anticancer agents for the prevention or treatment of different disease