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In a unique serum- and protein-free chemically defined in vitro culture model of postimplantation mammalian development the epidermis differentiates regularly, although the differentiation of other tissues is impaired due to the lack of the serum. The present study in that model was done to estimate more carefully the degree of epidermal differentiation in defined media supplemented with some growth- or differentiation-stimulating substances. The main objective was to discover by grafting in vivo to the richer environment whether simple protein-free culture conditions restrict an inherent embryonic potential for differentiation of skin appendages. Embryonic parts of E9.5 gastrulating Fischer rat embryos were cultivated for 2 weeks in the protein-free Eagle’s minimum essential medium supplemented with holotransferrin, apotransferrin, insulin and/or Na2SeO3 and in controls cultivated in protein-free medium or in serum-supplemented medium. In all experiments there was a high incidence of differentiation of the epidermis. A high level of epidermal differentiation was confirmed for the first time at the ultrastructural level. A well-differentiated cornified layer and cells connected with desmosomes containing keratohyaline masses and cytokeratin filaments were found. A strong immunohistochemical signal for the proliferating cell nuclear antigen was always detected in the basal layer of the epidermis showing that those cells were still able to proliferate. Finally, embryos precultivated for 1 or 2 weeks in the protein-free medium and media supplemented with apotransferrin or serum were grafted under the kidney capsule for an additional 2 weeks. It was discovered that even after spending 2 weeks in the simple protein-free medium in vitro, embryos retained their developmental potential for differentiation of skin appendages (hair and sebaceous glands).

We have been using an original serum-free chemically defined model system for investigation of developmental processes in gastrulating rat embryo. By this method 9, 5 days old rat embryos without extraembryonic membranes are cultivated on the air-liquid boundary for two weeks in serum-free medium without protein supplements. In such system parameters as survival, growth and differentiation of embryos can be followed by simple methods. 5-azacytidine (5azaC), demethylating agent and teratogen, was applied to the serum- and protein-free culture medium as well as to serum-free medium supplemented with human transferrin. Only one dosage of 5azaC impaired survival, growth and differentiation of embryos in serum-free medium, while in our recent as well as in previous experiments in serum-supplemented medium only repeated application of 5-azacytidine caused changes of development. We might conclude that in simple chemically defined medium it was easier to discover activity of 5-azacytidine than in medium with serum . So, this serum-free in vitro method can probably be used further on to screen for teratogenic or embryotoxic substances at the stage of gastrulation which is the most critical stage of mammalian development.

Prikaz knjige fra Ante Grbeša, Vrijeme kad se svjetiljka puni uljem Teovizija, Zagreb 2003.

Kod nekoliko kalusnih linija bundeve određena je aktivnost perok- sidaze i dobivena elektroforetska slika njenih izoenzima. Najviša aktivnost enzima utvrđena je kod kalusnnih linija DE i NA3, koje rastu uz dodatak regulatora rastenja i pokazuju visoku sposobnost zametanja embrioida. Kod prilagođenog tkiva (linija Ž5b/T) znatno je niža aktivnost peroksidaze, međutim pojavi embrioida prethodi porast aktivnosti enzima. Kalusne linije razlikuju se međusobno u elektroforetskoj slici izoenzima. Na tu sliku utječu regulatori rastenja, međutim ona je također odraz određenog razvojnog stanja u kulturi.

The summer aspect of seaweed communities of the island of Lokrum and the opposite Dubrovnik coastline was studied by the technique of free diving. Algal specimens were collected in August—September 1960 and 1961, May 1962, and in July 1974 and 1979, along many vertical and horizontal profiles within the upper, middle, and lower infralittoral zones. Among the specimens collected 234 taxa of Thallophyta and 1 species of Angiospermae were determined, belonging to the following phylla: Rhodophyta 157 taxa, Phaeophyta 38 taxa, and Chlorophyta 39 taxa. On the ground of its geographic location, its vicinity to the coast, and the strong influence of the open sea, Lokrum seems to display transitional characteristics between the southern coastal facies and the facies of the open sea., Ljetni aspekt zajednica bentoskih algi otoka Lokruma i nasuprotnog dubrovačkog primorja ispitivan je tehnikom slobodnog ronjenja. Alge su skupljane od kolovoza do rujna 1960. i 1961. godine, u svibnju 1962. godine i u srpnju 1974. i 1979. godine, uz veći broj vertikalnih i horizontalnih profila unutar gornjeg, srednjeg i donjeg infralitorala. Među skupljenim uzorcima određena je 1 vrsta angiosperma i 234 vrsta talofita, i to Rhodophyta 157 vrsta, Phaeophyta 38 vrsta i Chlorophyta 39 vrsta. Svojim geografskim položajem, blizinom kopna, a unatoč tome pod jakim utjecajem otvorena mora, Lokrum svojom bentoskom florom pokazuje prijelazni oblik ozmeđu južnog priobalnog facijesa i facijesa otvorenog mora.

Peroxidase isozymes and their total activity have ben studied in pumpkin embryogenic callus lines DE and NA3 in order to state how peroxidases are influenced by different kinds of exogenous growth regulators. Peroxidase bands Rm 0.46 and 0.54—0.56 were present in both lines regardless of the growth regulator. NAA and adenine sulphate induced the enzyme activity at the position Rm 0.35 which disappeared if the tissue was growing on 2,4-D. The changes in isoperoxidases pattern preceded any morphological change in the culture. The total peroxidase activity increased with time in line NAs, however, it remained low in line DE. Protein content changed inversely with peroxidase activity., Embriogene kalusne linije bundeve (Cucurbita pepo L.) koje rastu uz dodatak 2,4-D (DE) odnosno NAA i adenin sulfata (NA3) uspoređene su s obzirom na elektroforetsku sliku peroksidaza i njihovu ukupnu aktivnost. Peroksidazne pruge Rm 0,46 i 0,54—0,56 utvrđene su kod obje linije bez obzira na vrstu regulatora rastenja u hranidbenoj podlozi. NAA i adenin sulfat inducirali su enzimsku aktivnost u položaju Rm 0,35 koja je registrirana prije vanjskih morfoloških promjena u kulturi (embrioidi u kasnim razvojnim stadijima i biljčice). Peroksidazna aktivnost u položaju Rm 0,68 bila je naročito izražena uz dodatak NAA i adenin sulfata, a izostala je u mlađim kulturama linije DE na podlozi s 2,4-D. Aktivnost peroksidaza naglo je rasla tijekom starenja kultura linije NAa, dok je kod linije DE ostala niska. Sadržaj topivih proteina mijenjao se obrnuto proporcionalno peroksidaznoj aktivnosti.

The crown gall tumor test system on potato tuber discs was used for the study of antiumor activity of mistletoe protein extracts. By estimating the number and weight of tumors we found that crude mistletoe protein extract inhibits tumor induction and growth. The refined protein fraction was even more efficient. The mistletoe proteins also showed stimulative effects on rhizogenesis., Crown-gall tumori na tkivu gomolja krumpira korišteni su kao test-sistem za ispitivanje antitumorske aktivnosti proteina u ekstraktu imele. Određivanje broja i mase tumora pokazalo je, sa statističkom sigurnošću, da već sirovi ekstrakt inhibira tumorsku indukciju i rast. Inhibitomo djelovanje pročišćenih proteinskih frakcija još je izrazitije. Proteinski ekstrakt stimulirao je rizogenezu u tumorskom tkivu.