Effectiveness of immunization with multi-component bacterial immunomodulator in foals at 35th day of life.

The aim of the study was to investigate the mechanisms leading to immunization through the use of a multicomponent bacterial immunomodulator and to find out the relationship between the TLR 4 receptor with selected parameters of innate immunity and to acquire immunity. The study was conducted on 18 Polish Pony Horses foals divided into two study groups: control (n = 9) and experimental (n = 9). Foals from the experimental group received intramuscular duplicate injection of 5 ml of multi-component bacterial immunomodular at 35 and 40 days of age. RNA isolated from venous blood was used to evaluate the expression of TLR4 genes using RT-PCR. Concentration of Il-6, IL-10, IgM and IgG2 was determined by the ELISA method in blood plasma. Immunostimulation had a significant impact on the level of genes expression for TLR4 expression and IL-6 concentration. No effect of stimulation on IgM and IgG2 concentrations was found. The expression of TLR4 genes as well as the levels of interleukins could be modulated by stimulation with a pharmacological agent multi-component bacterial immunomodulator. The experiment demonstrated a strong positive correlation between TLR4 gene expression and IL-6 concentration and between TLR4 gene expression and IgM concentration.

In vitro maturation of equine oocytes followed by two vitrification protocols and subjected to either intracytoplasmic sperm injection (ICSI) or parthenogenic activation.

The aim of this study was determine the viability and developmental competence of equine oocytes after IVM and vitrification using the Rapid-I method, as part of an effort to develop an effective equine oocyte vitrification protocol. Equine oocytes were collected by scraping ovarian follicles of slaughtered mares. A total of 1052 ovaries were used in this study, from which 3135 oocytes were obtained. Of the 2853 oocytes retrieved, 2557 underwent in vitro maturation for approximately 36 h. After in vitro culture, 1202 oocytes (47%) had a first polar body. To evaluate the toxicity of the solutions (Experiment I), oocytes were exposed to vitrification media without cryopreservation. Of all the experimental groups evaluated, the best results were obtained for IVM oocytes exposed to EquiproVitKit media (IVM + TOX EquiVitKit), with a viability rate of 69.5%. In the Experiment II, oocytes, either freshly collected from the ovary or after in vitro maturation (IVM), were vitrified using either the EquiPro VitKit or an in-house medium containing 18% Ficoll, 40% ethylene glycol and 0.3 M sucrose. Oocytes were stained with fluorescein diacetate and ethidium bromide to evaluate viability. In vitro matured oocytes vitrified using EquiproVitKit media (IVM + VIT EquiVitKit) had a cryosurvival rate of 63%. In the last part of the study (Experiment III), vitrified IVM oocytes were activated by 7.5 µM ionomycin in TCM-199 for 5 min TCM 199 (5 min) combined with 2 mM 6-DMAP in TCM-99 with 10% FBS (4.5 h) or in vitro fertilized using ICSI. Development of potential embryos after activation in TCM-199 medium, showed a cleavage rate was 10.2%, compared to 22.5% of oocytes cultured in G1/G2 medium. ICSI of vitrified IVM oocytes resulted in 20% embryo development to the 16-cell stage, compared to 33.3% in the control. The vitrification of oocytes after IVM by Rapid-I method is a good way to preserve genetic material in horses.