Phylogeny of phylum Porifera

U znanosti postoji težnja za rješavanjem evolucijske povijesti taksonomskih skupina živih (i poznatih izumrlih) organizama. Ovim se kompleksnim zadatkom bavi filogenija. Koljeno Porifera - spužve - pokazalo se kao iznimno zanimljiva skupina čija filogenija nije potpuno ustanovljena niti nakon više od stotinu godina istraživanja i unatoč razvitku novih metoda istraživanja. Povijesno gledajući, morfologija spikula te cjelokupnog skeleta imala je važnu ulogu u postavljanju prvotne sistematike spužvi, a njen se značaj ne smije zanemariti ni danas. Uspostavljanje filogenije isključivo pomoću morfologije nije bilo moguće. Nešto kasnije, citologija je donijela novitete u pristupu i jasne odgovore kod razlučivanja pojedinih porodica. Također je vrlo važna metoda kod istraživanja spužvi bez spikula. U pojedinim slučajevima, citologija se pokazala efikasnijom od molekularne filogenetike. Istraživanja sekundarnih metabolita pokazala su se problematičnima zbog brojnosti endosimbionata u spužvi, iako potencijalno nude koristan set informacija za filogeniju. Razvitak metoda molekularne filogenije 1990-ih rezultirao je opsežnom primjenom na spužvama, međutim postoji još niz neriješenih pitanja osobito unutar skupine Demospongiae. Ipak, postoje poteškoće u primjeni tih novih metoda kad je potrebno odrediti filogeniju vrsta čija se specijacija odvijala u evolucijski kratkom periodu ili kad je proces odvajanja evolucijski „star“. Problem najčešće proizlazi iz naše nemogućnosti da sa sigurnošću detektiramo homoplaziju (tj. ortologe) i sekundaran gubitak karakteristika. Dio poteškoća bit će moguće riješiti kroz pažljiv odabir većeg broja gena s filogenetski važnom informacijom u ulazni set podataka za formiranje rodoslovnog stabla. Probleme filogenije spužvi stoga nije moguće riješiti jedinstvenom metodom niti izoliranim setom informacija. Njima treba pristupiti multidisciplinarno uz racionalno korištenje što većeg broja dostupnih metoda/informacija.There is a strong tendency in science towards solving the evolutionary history of taxonomic ranks of all living (and known fossil) organisms. That complex task is assigned to phylogeny. The phylum Porifera - sponges - has turned out to be a very interesting group whose phylogeny has not yet been resolved despite more than hundred years of research and emergence of new scientific methods. From a historical point of view, morphology of spicules and overall skeleton shape as a whole had an important role in defining the early systematics of Porifera and it is not to be ignored even at this point in time. On the other hand, using morphology as the only key to assemble phylogeny was not sufficient. Appearing a bit later, cytology has brought novelty in approach and clear answers regarding some Poriferan genera. Also, it plays a very important role in studying sponges without spicules. In some cases cytology has proved to be more effective in yielding information than molecular phylogeny. Including secondary metabolites in analyses, although potentially useful, has shown to be rather difficult due to a great number of endosymbionts living in sponges. The development of molecular phylogeny methods in 1990's resulted with their wide use in research of Porifera. Still, many questions remain unanswered, especially regarding the class Demospongiae. These new methods have been less effective in search of connections among species that evolved rather quickly in an evolutionary sense of time and/or have separated early in the evolution. The problem most commonly arises from our inability to detect homoplasy (or orthologues) and a secondary character loss with certainty. A part of the solution is expected to be reached through inclusion of more carefully selected genes that carry useful phylogenetic information which would be included in the formation of a phylogenetic tree. In the end, problems of modern phylogeny cannot be solved with any unique method or single set of information. They have to be approached from a multidisciplinary point of view with rational use of as many methods/information as possible

Phylogeny of phylum Porifera

U znanosti postoji težnja za rješavanjem evolucijske povijesti taksonomskih skupina živih (i poznatih izumrlih) organizama. Ovim se kompleksnim zadatkom bavi filogenija. Koljeno Porifera - spužve - pokazalo se kao iznimno zanimljiva skupina čija filogenija nije potpuno ustanovljena niti nakon više od stotinu godina istraživanja i unatoč razvitku novih metoda istraživanja. Povijesno gledajući, morfologija spikula te cjelokupnog skeleta imala je važnu ulogu u postavljanju prvotne sistematike spužvi, a njen se značaj ne smije zanemariti ni danas. Uspostavljanje filogenije isključivo pomoću morfologije nije bilo moguće. Nešto kasnije, citologija je donijela novitete u pristupu i jasne odgovore kod razlučivanja pojedinih porodica. Također je vrlo važna metoda kod istraživanja spužvi bez spikula. U pojedinim slučajevima, citologija se pokazala efikasnijom od molekularne filogenetike. Istraživanja sekundarnih metabolita pokazala su se problematičnima zbog brojnosti endosimbionata u spužvi, iako potencijalno nude koristan set informacija za filogeniju. Razvitak metoda molekularne filogenije 1990-ih rezultirao je opsežnom primjenom na spužvama, međutim postoji još niz neriješenih pitanja osobito unutar skupine Demospongiae. Ipak, postoje poteškoće u primjeni tih novih metoda kad je potrebno odrediti filogeniju vrsta čija se specijacija odvijala u evolucijski kratkom periodu ili kad je proces odvajanja evolucijski „star“. Problem najčešće proizlazi iz naše nemogućnosti da sa sigurnošću detektiramo homoplaziju (tj. ortologe) i sekundaran gubitak karakteristika. Dio poteškoća bit će moguće riješiti kroz pažljiv odabir većeg broja gena s filogenetski važnom informacijom u ulazni set podataka za formiranje rodoslovnog stabla. Probleme filogenije spužvi stoga nije moguće riješiti jedinstvenom metodom niti izoliranim setom informacija. Njima treba pristupiti multidisciplinarno uz racionalno korištenje što većeg broja dostupnih metoda/informacija.There is a strong tendency in science towards solving the evolutionary history of taxonomic ranks of all living (and known fossil) organisms. That complex task is assigned to phylogeny. The phylum Porifera - sponges - has turned out to be a very interesting group whose phylogeny has not yet been resolved despite more than hundred years of research and emergence of new scientific methods. From a historical point of view, morphology of spicules and overall skeleton shape as a whole had an important role in defining the early systematics of Porifera and it is not to be ignored even at this point in time. On the other hand, using morphology as the only key to assemble phylogeny was not sufficient. Appearing a bit later, cytology has brought novelty in approach and clear answers regarding some Poriferan genera. Also, it plays a very important role in studying sponges without spicules. In some cases cytology has proved to be more effective in yielding information than molecular phylogeny. Including secondary metabolites in analyses, although potentially useful, has shown to be rather difficult due to a great number of endosymbionts living in sponges. The development of molecular phylogeny methods in 1990's resulted with their wide use in research of Porifera. Still, many questions remain unanswered, especially regarding the class Demospongiae. These new methods have been less effective in search of connections among species that evolved rather quickly in an evolutionary sense of time and/or have separated early in the evolution. The problem most commonly arises from our inability to detect homoplasy (or orthologues) and a secondary character loss with certainty. A part of the solution is expected to be reached through inclusion of more carefully selected genes that carry useful phylogenetic information which would be included in the formation of a phylogenetic tree. In the end, problems of modern phylogeny cannot be solved with any unique method or single set of information. They have to be approached from a multidisciplinary point of view with rational use of as many methods/information as possible

Phylogeny of phylum Porifera

U znanosti postoji težnja za rješavanjem evolucijske povijesti taksonomskih skupina živih (i poznatih izumrlih) organizama. Ovim se kompleksnim zadatkom bavi filogenija. Koljeno Porifera - spužve - pokazalo se kao iznimno zanimljiva skupina čija filogenija nije potpuno ustanovljena niti nakon više od stotinu godina istraživanja i unatoč razvitku novih metoda istraživanja. Povijesno gledajući, morfologija spikula te cjelokupnog skeleta imala je važnu ulogu u postavljanju prvotne sistematike spužvi, a njen se značaj ne smije zanemariti ni danas. Uspostavljanje filogenije isključivo pomoću morfologije nije bilo moguće. Nešto kasnije, citologija je donijela novitete u pristupu i jasne odgovore kod razlučivanja pojedinih porodica. Također je vrlo važna metoda kod istraživanja spužvi bez spikula. U pojedinim slučajevima, citologija se pokazala efikasnijom od molekularne filogenetike. Istraživanja sekundarnih metabolita pokazala su se problematičnima zbog brojnosti endosimbionata u spužvi, iako potencijalno nude koristan set informacija za filogeniju. Razvitak metoda molekularne filogenije 1990-ih rezultirao je opsežnom primjenom na spužvama, međutim postoji još niz neriješenih pitanja osobito unutar skupine Demospongiae. Ipak, postoje poteškoće u primjeni tih novih metoda kad je potrebno odrediti filogeniju vrsta čija se specijacija odvijala u evolucijski kratkom periodu ili kad je proces odvajanja evolucijski „star“. Problem najčešće proizlazi iz naše nemogućnosti da sa sigurnošću detektiramo homoplaziju (tj. ortologe) i sekundaran gubitak karakteristika. Dio poteškoća bit će moguće riješiti kroz pažljiv odabir većeg broja gena s filogenetski važnom informacijom u ulazni set podataka za formiranje rodoslovnog stabla. Probleme filogenije spužvi stoga nije moguće riješiti jedinstvenom metodom niti izoliranim setom informacija. Njima treba pristupiti multidisciplinarno uz racionalno korištenje što većeg broja dostupnih metoda/informacija.There is a strong tendency in science towards solving the evolutionary history of taxonomic ranks of all living (and known fossil) organisms. That complex task is assigned to phylogeny. The phylum Porifera - sponges - has turned out to be a very interesting group whose phylogeny has not yet been resolved despite more than hundred years of research and emergence of new scientific methods. From a historical point of view, morphology of spicules and overall skeleton shape as a whole had an important role in defining the early systematics of Porifera and it is not to be ignored even at this point in time. On the other hand, using morphology as the only key to assemble phylogeny was not sufficient. Appearing a bit later, cytology has brought novelty in approach and clear answers regarding some Poriferan genera. Also, it plays a very important role in studying sponges without spicules. In some cases cytology has proved to be more effective in yielding information than molecular phylogeny. Including secondary metabolites in analyses, although potentially useful, has shown to be rather difficult due to a great number of endosymbionts living in sponges. The development of molecular phylogeny methods in 1990's resulted with their wide use in research of Porifera. Still, many questions remain unanswered, especially regarding the class Demospongiae. These new methods have been less effective in search of connections among species that evolved rather quickly in an evolutionary sense of time and/or have separated early in the evolution. The problem most commonly arises from our inability to detect homoplasy (or orthologues) and a secondary character loss with certainty. A part of the solution is expected to be reached through inclusion of more carefully selected genes that carry useful phylogenetic information which would be included in the formation of a phylogenetic tree. In the end, problems of modern phylogeny cannot be solved with any unique method or single set of information. They have to be approached from a multidisciplinary point of view with rational use of as many methods/information as possible

Expression of proteins sFRP3 and DVL3, key molecules of Wnt signaling pathway in glioblastoma

Oboljeli od glioblastoma uz trenutno dostupne terapije imaju medijan preživljenja kraći od godinu dana. Cilj diplomskog rada je identificirati status ključnih proteina signalnog puta Wnt (sFRP3 I DVL3) u stanicama ljudskog glioblastoma, provjeriti postoji li poveznica statusa s kliničkim karakteristikama bolesnika i ukazati na eventualne nove mete za razvoj ciljnih terapija. Na preparatima ljudskog glioblastoma proteini su detektirani metodom imunohistokemije. Tkiva su analizirana pod svjetlosnim mikroskopom (Olympus BH-2). Razina ekspresije proteina određena je semikvantitativnom metodom u 3 stupnja jačine signala uz oznaku tkivne i subcelularne lokalizacije. Oba proteina su eksprimirana u stanicama glioblastoma. Nema korelacije između njihove ekspresije, kao ni korelacije jačine signala s kliničkim parametrima. Dominacija jakog signala u stanicama dovedena je u vezu sa simultanom lokalizacijom proteina DVL3 u citoplazmi i nukleusu, te proteina sFRP3 simultano u citoplazmi i ekstracelularno uz izostanak signala u nukleusu. Prognoze oboljelih od glioblastoma su loše čak i uz kombiniranje dostupnih terapija, jer je glioblastom jedan od agresivnijih i malignijih tipova tumora, često neoperabilan zbog lokalizacije. Glioblastom je vrlo heterogenog porijekla i histopatološke slike, specifičan za oboljelog, bitno je razviti metode kojima će se detektirati individualna slika stanja. Tek će detaljnije poznavanje mreže signalnih puteva moći ukazati na optimalne pristupe terapijskim rješenjima.Glioblastoma patients have a median survival of less than a year. Thesis aim is to identify key proteins' (sFRP3 and DVL3) status in Wnt signaling pathway in human glioblastoma samples, search for their connection with clinical data and point out possible new target therapy objects. Proteins are detected by immunohistochemistry. Tissues are analyzed under a light microscope (Olympus BH-2). The expression levels are determined by the semiquantitative method in the 3-stage signal strength with tissue and subcellular localization labels. Both proteins are expressed in glioblastoma. There is no correlation between their expression, as well as signal strength correlation with clinical parameters. Strong signal dominance was brought into connection with simultaneous localization of DVL3 in the cytoplasm and nucleus, and sFRP3 simultaneously in the cytoplasm and extracellularly along with the absence of signal in the nucleus. Prognosis for glioblastoma patients are bad even with the combination of all available therapies. It is one of the most aggressive and malignant tumor types, often inoperable due to localization. Glioblastoma is highly heterogeneous in origin and histopathological images so developing methods for detecting individual condition traits are essential. Only a more detailed knowledge of signaling pathways network could give the optimal therapeutic solutions

Expression Levels and Localizations of DVL3 and sFRP3 in Glioblastoma

The expression patterns of critical molecular components of Wnt signaling, sFRP3 and DVL3, were investigated in glioblastoma, the most aggressive form of primary brain tumors, with the aim to offer potential biomarkers. The protein expression levels and localizations in tumor tissue were revealed by immunohistochemistry and evaluated by the semiquantitative method and immunoreactivity score. Majority of glioblastomas had moderate expression levels for both DVL3 (52.4%) and sFRP3 (52.3%). Strong expression levels were observed in 23.1% and 36.0% of samples, respectively. DVL3 was localized in cytoplasm in 97% of glioblastomas, of which 44% coexpressed the protein in the nucleus. sFRP3 subcellular distribution showed that it was localized in the cytoplasm in 94% of cases. Colocalization in the cytoplasm and nucleus was observed in 50% of samples. Wilcox test indicated that the domination of the strong signal is in connection with simultaneous localization of DVL3 protein in the cytoplasm and the nucleus. Patients with strong expression of DVL3 will significantly more often have the protein in the nucleus (P=6.33×10−5). No significant correlation between the two proteins was established, nor were their signal strengths correlated with epidemiological parameters. Our study contributes to better understanding of glioblastoma molecular profile