The wide use of cell technologies in clinical practice requires a large amount of cell material, which has led to improvement in culture conditions, making it possible to obtain more cell material in a shorter period of time. Thus, the purpose of our paper was to study the effects of different concentrations of an insulin-like growth factor (IGF-1), a fibroblast growth factor (FGF-2),| a growth hormone (rhGH), and Biolaminin 521 LN (LN 521) on the proliferative activity and genetic stability of stem cell cultures derived from the cat bone marrow, adipose tissue, and myocardium. Cell cultures for the experiment were obtained from the adipose tissue, bone marrow, and myocardium of a cat. Differences were found in the effects of the various growth promoters on the proliferative activity of cells in the culture. The IGF-1 demonstrated a positive effect on the proliferative activity of all cultures. The addition of the rhGH to the bone marrow-derived cell culture increased the size of the cells and decreased the proliferation index relative to the control group. The addition of the growth factors to the culture medium did not significantly increase the number of cells with altered karyotype in any of the cultures relative to the control group.
According to the International Embryo Technology Society, the number of bovine embryos produced by in vitro fertilization technology is increasing every year. However, despite the large volumes of their production, the effectiveness of this method is still low and needs to be improved. Therefore, the purpose of this study was to compare the effectiveness of two commercial media – Origio Sequential Series (Origio, Denmark) and a panel of products manufactured by Minitube (Germany) in terms of oocyte maturation and development of bovine embryos in vitro. At the first stage of the study, a comparative evaluation of oocyte maturation media was performed: based on TCM 199 (Minitube) and Universal (Origio) culture media. At the second stage, the protocols for culturing bovine embryos were compared: Minitube and the two-stage Origio culture protocol with changing media. Therewith, it was found that the use of TCM 199 medium for oocyte maturation is more effective compared to Universal. Thus, at 48 hours of cultivation (the initial stage of embryo development), 64.3 ± 1.0 and 60.3 ± 1.4% of 2-8 cell embryos were obtained, and on Day 8 – 25.3 ± 1.0 and 20.0 ± 0.6% of blastocysts, respectively. The results of a comparison of bovine embryo culture protocols showed that when using both Minitube and Origio media, the percentage of division and the percentage of resulting embryos corresponded to their known values. It was found that the Minitube cultivation protocol is more effective than Origio. At 48 hours, the number of embryos obtained using the Minitube culture protocol was 1.3% higher compared to Origio, on Day 6 – by 7.8%, and on Day 8 – by 3.8%. The results obtained are a necessary component of the development of successful processes to produce bovine embryos in vitro with further implementation in the ruminant reproduction biotechnology
Producing embryos in vitro is an important technology used to improve the genetic potential of cattle and perfect the programs of their breeding. Regardless of the way they are produced, all embryos that had not been used for transplantation to recipients must be conserved. Because of significantly increased interest in the problem of cryoconservation of embryos, both coming from scientists and businesses, there are emerging new commercial environments that allow the facilitation of cryoconservation and the increase in the embryo survival. Oocyte-cumulus complexes obtained from the ovaries of slaughtered clinically healthy cows matured in 22–24 h in in vitro conditions. The oocytes were co-cultured with spermatozoids in Fertilization medium, and the obtained zygotes were cultured in Culture medium with Sodium-Pyruvate for 4 or 7 days up to the stage of morula or blastocyste, respectively. For the vitrification of cow embryos, we used a commercial kit for the vitrification of human embryos, having compared the duration of equilibration. According to the results of the studies, we observed high efficiency of cryoconservation of cow embryos using the commercial kit for vitrification of human embryos. The results revealed the significant effect of equilibration on survival and further development of embryos. In addition, we described the dependence of development stage of cattle embryo on the duration of the contact of embryo with equilibration solution. Therefore, optimal time of contact of cattle embryos at the morula stage with equilibration solution was 12 minutes. On the 24th h after thawing, 46.7 ± 3.3% of the embryos were observed to undergo blastulation, and on 48th h, this parameter increased to 96.7 ± 3.3%, which corresponded to the parameters in the group of embryos that had not been subjected to cryoconservation. In the conditions of further cultivation, the percentage of blastocystes that hatched in the experimental group was no different from that of the control. At the same time, the highest efficiency of vitrification of blastocystes of cows was seen after the contact with the equilibration solution for 15 min, since the percentage of hatched blastocystes was the same as in the control group. Therefore, using the commercial kit for vitrification of human embryos is beneficial, for it promotes the parameters of cow embryos after vitrification/thawing that are similar to such of intact embryos (without freezing). The data we analyzed and presented in the paper could help to increase the efficiency of cryoconservation of cattle embryos for both scientific and commercial purposes.
VETERINARYАнотація. У тканинах дорослого організму, окрім спеціалізованих клітин, містяться незрілі, недиференційовані і низькодиференційовані клітин, так звані стовбурові. Не виключенням є кістковий мозок, жирова тканина та підшлу-нкова залоза. Саме ці клітини здатні до адгезії та проліферації в умовах in vitro утворюючи клітинні культури. У статті описано вплив культур клітин отриманих з різних тканин (підшлункова залоза, кістковий мозок та жирова тканина) на перебіг експериментального цукрового діабету у щурів. Досліджено, що оптимальним методом введення клітинного матеріалу, є трансплантація його під капсулу підшлункової залози. Встановлено, що після введення куль-тури клітин отриманих із перерахованих вище тканин, у тварин-реципієнтів із експериментальним цукровим діабе-том наступає позитивний терапевтичний ефект у вигляді збільшення загального об'єму острівкової тканини (у порі-внянні з контрольною групою), зниження рівня глюкози у сироватці крові. Ключові слова: культура клітин, цукровий діабет, кістковий мозок, жирова тканина, підшлункова залоза, острі-вці Лангерганса.
Досліджено вплив фактору росту фібробластів (FGF-2) та інсуліноподібного фактору росту (IGF-1) у різних концентраціях на проліферативну активність та генетичну стабільність стовбурових клітин отриманих з кісткового мозку, жирової тканини та міокарду кота. Встановлено, що інсуліноподібний фактор росту та фактор росту фібробластів позитивно впливає на проліферативну активність всіх досліджуваних культур. За даними цитогенетичного аналізу встановлено, що додавання факторів росту у культуральне середовище не призводить до достовірного збільшення кількості генетичних помилок (у порівнянні з контролем) у всіх досліджуваних культурах. Ключові слова: фактор росту фібробластів (FGF-2), інсуліноподібний фактор росту (IGF-1), стовбурові клітини, культура клітин кісткового мозку, культура клітин жирової тканини, культура клітин міокарда, коти, цитогенетичний аналіз.
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