Browsing by Author "Serpa Carangui, Erika Margoth"

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Eficacia de tres protocolos de criopreservación de espermatozoides equinos con diferentes velocidades de enfriamiento sobre la calidad espermática postdescongelación
(Universidad de Cuenca, 2022-09-29) Serpa Carangui, Erika Margoth; De la Cruz Tandazo, Francisco Javier; Soria Parra, Manuel Elías
The cooling rate determines intracellular and extracellular ice formation and, consequently, sperm cryosurvival. This work evaluated the efficacy of a cryopreservation protocol based on two freezing ramps with accelerating cooling rates compared with two commercial freezing protocols: one conventional with decelerating cooling rates, and another with programmed cooling rates by the biological freezer. For this, 24 ejaculates from 4 sexually adult Arabian stallions were collected with an artificial vagina, diluted with Botusemen Gold® + 5% dimethylformamide, and frozen by exposing the straws (0.5 mL) to liquid nitrogen (LN2) in three different protocols: [1] PCC-1: Conventional (Minitube®) with 1 freezing ramp at 5 cm from the LN2 level for 10 min; [2] PC2R-2: two ramps at 17 cm (for 4 min) and 2 cm (for 2 min) from the level of LN2 content inside a cryobox of 31 x 31 x 30 cm (length, width, and height, respectively); and [3] PCBP-3: programmable biological freezer TK-4000 (Tecnologia®). Ice nucleation and initial cooling rate occurred at -13.8ºC to 63.5ºC/min, at -8.5ºC to 19ºC/min, and at -5.9ºC to -11.4ºC/min for PCC-1, PC2R-2, and PCBP-3, respectively. The cryopreservation process reduced (P<0.01) the kinematic parameters and integrity of sperm membranes, regardless of the protocol used. There were no significant differences (P>0.05) neither kinematics nor plasma membrane integrity between freezing protocols. However, the PC2R-2 and PCBP-3 protocols produced higher post-thaw acrosomal integrity than the PCC-1 protocol. In conclusion, the PC2R-2 protocol produced accelerating cooling rates and cryosurvival of equine spermatozoa similar to commercial protocols of conventional freezing or programmed cooling rates, even improving acrosomal integrity. The use of the two-ramp freezing protocol is recommended as a successful alternative to cryopreserve stallion sperm due to its easy application, efficacy, and less consumption of LN2.
Optimization of cryopreservation of arabian stallion sperm using dimethylformamide, glycerol, and different freezing protocols
(Wiley-Blackwell, 2022) Serpa Carangui, Erika Margoth; Samaniego Campoverde, Jorge Xavier; Galarza Lucero, Diego Andrés; Soria Parra, Manuel Elías; Ochoa Guillén, Juan Diego; Álvarez Palacios, David Alejandro; De la Cruz Tandazo, Francisco Javier; Galarza Lucero, Diego Andrés
This study evaluated the effect of penetrating cryoprotectant agents (CPA) and the cryosurvival of three freezing protocols on the kinematics and integrity of membranes of frozen-thawed stallion sperm. Twenty-four ejaculates of four adult Arabian horses were collected in six weekly sessions (six ejaculates/horse). Each ejaculate was divided into two aliquots. With the first aliquot, three CPA treatments were conformed: 5% glycerol (GLY), 5% dimethylformamide (DMF), and 3%–3% DMF–GLY combination, and the sperm samples were frozen exposing them to liquid nitrogen (LN2) vapors. The second aliquot was diluted with freezing medium plus 5% DMF and the sperm samples were frozen in three freezing protocols: (P1) Styrofoam cryo-box (30 × 29 × 31 cm of length, width, and height, respectively) with two ramps (at 17 and 7 cm above LN2); (P2) freezing unit® (Minitüb, Germany); and (P3) programmable TK 4000-freezer® (Compacta, Brazil). The DMF-GLY combination and DMF yielded higher (p<.05) post-thaw values than the GLY regarding the motility (SM: 54.2±2.25 and 50.2±1.80 vs. 41.4±2.35%, respectively), curvilinear velocity (VCL: 58.0±1.71 and 54.0±1.58 vs. 42.3±1.60 µm/s), and the proportion of sperm with intact plasma and intact acrosome (IPIA: 58.0±1.11 and 52.6±0.99 vs. 42.5±1.07%). Furthermore, the P1 protocol produced a similar (p>.05) post-thaw SM, VCL, and IPIA than the other protocols. Indeed, the P1 and P3 protocols yielded lower proportion (p<.05) of sperm with damaged plasma and damaged acrosome than the P2 protocol after thawing (3.7±0.18 and 3.1±0.18 vs. 6.1±0.44%, respectively). In conclusion, the addition of DMF or combined with GLY to freezing medium, and the freezing with Styrofoam cryo-box with two ramps increase the cryosurvival of Arabian stallion spermatozoa.