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PubMed2006

Denuding and centrifugation of maturing bovine oocytes alters oocyte spindle integrity and the ability of cytoplasm to support parthenogenetic and nuclear transfer embryo development.

The effects of cumulus cell removal and centrifugation of maturing bovine oocytes on nuclear maturation and subsequent embryo development after parthenogenetic activation and nuclear transfer were examined. Removal of cumulus cells at 4, 8, and 15 hr after in vitro maturation (IVM) or the centrifugation of denuded oocytes had no effect on maturation rates. Oocytes treated at 0 hr of IVM had a lower expulsion rate (50%) of the first polar body (PB1). The removal of cumulus cells and centrifugation affected the pattern of spindle microtubule distribution and division of chromosomes. There were almost no spindle microtubules allocated to PB1 and the spindles were swollen in anaphase I and telophase I oocytes. Approximately 20% of PB1 oocytes contained tripolar or multipolar spindles. After activation, oocytes denuded with or without centrifugation at 8 hr of IVM resulted in the lowest rate of development (3.0%). Denuded oocytes at 4, 15, and 24 hr of IVM with centrifugation or not resulted in similar blastocyst development rates (9.6%-13.2%). However, centrifugation of oocytes denuded at the beginning of IVM resulted in lower blastocyst development rate (8.1%, P < 0.05) than the noncentrifuged oocytes (17.3%). After nuclear transfer, the blastocyst development rates of oocytes denuded and centrifuged at 0, 4, and 8 hr of IVM were not different when compared to the same patch of noncentrifuged oocytes. However, oocytes denuded and centrifuged at 15 hr of IVM resulted in lower (P < 0.05) blastocyst development rates than the noncentrifuged oocytes. The results of this study suggest that removal of cumulus cells and centrifugation of denuded oocytes affect the spindle pattern. Embryo development of denuded and centrifuged oocytes may differ depending on the time of removal of cumulus cells.

Animals↗
PubMed1986

High-frequency twinning of Xenopus laevis embryos from eggs centrifuged before first cleavage.

In embryos of the frog Xenopus laevis, the dorsal structures normally develop from regions of the egg opposite the side of sperm entry. Gravity and centrifugal force, applied at an angle of 90 degrees to the animal-vegetal axis of the egg, can override this topographic relationship and can cause the dorsal structures to be positioned according to the force vector (S. Black and J. Gerhart, 1985, Dev. Biol. 108, 310-324). We report here that at time 0.40 (40% of the first cleavage interval), an average of 60% of eggs centrifuged at 30g for 4 min in this orientation form conjoined twins with one body axis arising from the centripetal side of the egg and one arising from the centrifugal side of the egg. This positioning is observed regardless of the orientation of the side of sperm entry in the centrifugal field. If, after the 0.40 centrifugation, the eggs are inclined with the centripetal side up, they do not make twins; instead, they make only a single axis at the centripetal side. This indicates that the second axis in twins is caused to form by postcentrifugation gravity-driven internal rearrangements of materials that were displaced by the centrifugation. Twins also form at high frequency in eggs centrifuged twice, first at an inclination of 90 degrees, and then at an inclination of 0 degrees. The second centrifugation yields secondary axes even when it is begun midway in the second cell cycle, well after the time of grey crescent formation. Double centrifugation also causes twinning ("double rescue") of uv-irradiated eggs which otherwise would not develop axial structures. This suggests that the internal displacements caused by the centrifugations can substitute for a step in the normal axis specification process that is impaired in irradiated eggs.

Ambystoma mexicanum↗
PubMed1998

Increased recovery of viable spermatozoa through oscillating centrifugation.

OBJECTIVE: To compare the number and quality of spermatozoa recovered following the standard centrifugation method with those recovered following oscillation of the relative centrifugal field during the centrifugation method. DESIGN: Prospectively controlled in vitro study. SETTING: Private andrology laboratory. PATIENT(S): Seventeen healthy sperm donors. INTERVENTION(S): Equal halves of the same semen sample were evaluated following the standard centrifugation or following oscillation of the relative centrifugal field during centrifugation. MAIN OUTCOME MEASURE(S): Number of sperm, sperm motility, progressive sperm motility at 0 hours and after 3-hour of incubation at 5 degrees C in TES and Tris (buffer) (TEST)-yolk. RESULT(S): The number of sperm, sperm motility, progressive sperm motility at 0 hours and after 3-hour of incubation at 5 degrees C in TEST-yolk were significantly higher for the sperm recovered following oscillating centrifugation compared with standard centrifugation. CONCLUSION(S): Oscillating the relative centrifugal field during centrifugation yields higher number of viable spermatozoa.

Cell Survival↗
PubMed1984

Isolation and subfractionation of human peripheral blood mononuclear cells (PBMC) by density gradient centrifugation on Percoll.

The use of Percoll for isolation and subfractionation of PBMC and T-lymphocytes by discontinuous and continuous density gradient centrifugation is described: PBMC were isolated from human peripheral blood by discontinuous density gradient centrifugation on Percoll. The use of Percoll instead of Ficoll-Isopaque has the advantage that Percoll, in contrast to Ficoll-Isopaque, does not alter the density of monocytes. Therefore, a better separation of lymphocytes and monocytes was achieved after subsequent continuous density gradient centrifugation on Percoll. E-RFC were isolated by discontinuous density gradient centrifugation after a first low speed centrifugation step banding lymphocytes and SRBC on a Percoll-Ficoll cushion, and a subsequent high speed centrifugation step separating high density rosettes and SRBC from low density non-E-RFC. The advantage of this procedure is the short time of performance and that there is no need to resuspend the lymphocyte/SRBC pellet. PBMC, nph.PBMC T-lymphocytes were further subfractionated by continuous density gradient centrifugation on Percoll. The method described here resulted in a good separation of lymphocytes and monocytes. However, to obtain lymphocyte fractions with minute numbers of contaminating monocytes, a depletion of monocytes prior to further subfractionation of the lymphocytes by continuous density gradient centrifugation is recommended. A marker analysis of T-lymphocytes subfractionated by continuous density gradient centrifugation on Percoll shows that high density T-lymphocytes are enriched in ANAE positive lymphocytes of type 1 and depleted of ANAE positive lymphocytes of type 2. Low density T-lymphocytes are enriched in ANAE type 2 cells and depleted of ANAE type 1 cells. On the other hand, no considerable differences were found when analyzing the T-cells from different fractions for differentiation antigens by means of monoclonal antibodies (anti Lyt 3, OKT4, and OKT8). The results may indicate that subfractionation of T-lymphocytes by continuous density gradient centrifugation on Percoll provided T-cells in different functional states rather than T-cells of distinct subclasses.

Antibodies, Monoclonal↗
PubMed1993

Effect of centrifugal force on growth of mouse osteoblastic MC3T3-E1 cells in vitro.

The effect of biomechanical force on growth of skeletal tissue was studied in monolayer cultures of mouse osteoblastic MC3T3-E1 cells which were centrifuged at 320 g for 15 min to 72 h in a CO2 incubator. Centrifugation of the cells for 30 min in low concentrations (0.3 or 1%) of fetal bovine serum (FBS) caused a two-fold increase of [3H]thymidine incorporation at 20 h from the start of centrifugation. However, centrifugation under 10% FBS caused no increase in [3H]thymidine incorporation into DNA. Under 0.3% FBS, [3H]thymidine incorporation increased in a manner dependent on the period of centrifugation and reached a maximum when the cells were centrifuged for 3 h. Stimulation of DNA synthesis by centrifugation was abolished in the presence of H-7, an inhibitor of protein kinase C. Moreover, conditioned medium collected from the centrifuged cultures increased [3H]thymidine incorporation by two-fold over the basal when added to a quiescent culture of MC3T3-E1 cells. These results suggest that centrifugal force stimulates growth of osteoblastic cells through autocrine secretion of some diffusible growth-promoting activity. On the other hand, centrifugation of the cells inhibited induction by FBS of alkaline phosphatase activity and calcium-uptake, two indices of the differentiated phenotype of osteoblasts.

3T3 Cells↗
PubMed1999

Influence of centrifugation and hindlimb suspension on testosterone and corticosterone excretion in rats.

BACKGROUND: Alterations in mass loading have been shown to stimulate the pituitary-gonadal and pituitary-adrenal axes resulting in changes in testosterone (T) and corticosterone (B) production which may account for the previously reported variations in muscle mass under these conditions. HYPOTHESIS: Centrifugation and hindlimb suspension will induce opposing effects on testosterone and corticosterone excretion. METHODS: Effects of mass loading (centrifugation at -2Gx) and unloading (hindlimb suspension, HLS) on excreted T and B levels, and testicular mass were examined in Sprague-Dawley rats. For 12 d, 16 (8 controls and 8 centrifuged) rats and 20 (10 unsuspended controls and 10 suspended) rats were studied during the centrifugation and suspension portions of the study, respectively. Following the completion of each study, testicular mass was measured. RESULTS: Absolute testicular mass was not different, following 12 d of centrifugation. Excreted T was elevated between days 1 and 6 of centrifugation. Excreted B levels were elevated for the first 4 d of centrifugation. Suspension reduced absolute and relative testicular mass. Excreted T levels were reduced from days 3-12 of suspension, while excreted B levels were only elevated on the first day of suspension. CONCLUSIONS: Centrifugation stimulated an increase in T excretion despite a lack of a change in absolute testicular mass. Conversely, suspension resulted in a reduction in T excretion which may have been associated with a decrease in absolute testicular mass. The centrifugation-induced increase in T excretion is consistent with previously reported increases in lean body mass following centrifugation, while the HLS-induced reduction in T excretion correlates with previously reported decreases in lean body mass following suspension.

Adrenal Glands↗
PubMedDate not supplied

Influence of centrifugation regimes on motility, yield, and cell associations of mouse spermatozoa.

Mouse sperm are exceptionally sensitive to mechanical forces associated with pipetting and mixing. This characteristic raised the question of the sensitivity of mouse sperm to centrifugation, a step necessary in the removal of cryoprotectants and a common component in the general manipulation of sperm suspensions for experimental purpose. Epididymal spermatozoa from ICR mice were isolated and manipulated to minimize pipetting and mixing damage. The centrifugal accelerations studied were 200, 400, 600, and 800 x g (measured with a stroboscope) for 5, 10, or 15 minutes of centrifugation time. The number of cells and the number of motile cells were counted. The percent motility and longevity, total yield, and motile yield were calculated. Centrifugation at 200 and 400 x g for short times (5 minutes) caused only a small loss in either immediate or 2.5-hour motility, but centrifugation at 600 and 800 x g for 15 minutes produced up to a fivefold loss. Low speed/short time centrifugation pelleted only about half of the cells; the others were lost when the supernatant was removed. The maximum number of motile sperm (motile yield) was obtained at intermediate centrifugal forces (approximately 400 x g for 10-12 minutes), and it is the total number of motile sperm (and not the percent motility) that is important in the use of cryopreserved sperm to regenerate cryopreserved mutant lines. Relative centrifugal force and centrifugation time exhibit reciprocity (e.g., 200 x g for 10 minutes produces similar results to 400 x g for 5 minutes). The spermatozoa must be centrifuged under carefully defined conditions to minimize the damage and to maximize the recovery of viable cells.

Animals↗
PubMed1999

Centrifugal pumps: description of devices and surgical techniques.

BACKGROUND: Because of simplicity of application, universal access, and low cost, centrifugal pumps are commonly used for short-term mechanical cardiac assist. Indications and techniques for application of this technology continue to evolve. METHODS: The clinical experience with 151 patients undergoing centrifugal mechanical cardiac assist at the University of Missouri-Columbia has been reviewed. We have compared commonly available centrifugal pumping systems in vitro and in vivo for characteristics that might distinguish them. RESULTS: Centrifugal pumps have been found to be well suited for use in surgery on the thoracic aorta, for extracorporeal membrane oxygenation and for postcardiotomy cardiac mechanical assist. Complications associated with centrifugal mechanical assist are predictable and common but potentially can be reduced by improved surgical techniques and anticoagulation strategies. In vitro and in vivo experimentation with available centrifugal pumps reveals nuances characteristic of each of the devices. CONCLUSIONS: All centrifugal pumps presently available are less destructive to blood cellular elements compared with roller pumps. With familiarity, all can function satisfactorily for short-term mechanical assist with no compelling evidence that favors any particular centrifugal pump system clinically available. Centrifugal pumps are ideally suited for left heart bypass during surgery on a thoracic aorta and for short-term application as may be required for postcardiotomy mechanical assist. Centrifugal pump technology should be part of the armamentarium of all cardiothoracic surgeons.

Animals↗
PubMed2000

Selective retension of active cells employing low centrifugal force at the medium change during suspension culture of Chinese hamster ovary cells producing tPA.

The effect of centrifugal force applied for cell separation at the medium change on the growth, metabolism and tissue plasminogen activator (tPA) productivity of Chinese hamster ovary (CHO) cells suspension culture was investigated. The viability of the precipitated cells increased exponentially as the centrifugal force decreased. However, the cell recovery was lower than 91% when centrifugal forces applied for 5 min was less than 67 x g. In cultures incubated for 474 h with 7 medium changes employing centrifugal forces ranging from 67 to 364 x g, a centrifugal force lower than 119 x g resulted in higher specific rates of growth, glucose consumption, and lactate and tPA production during the whole culture period. On the other hand, daily centrifugation at 67 to 537 x g without discarding the supernatant had no effect on the specific rates. The cultures inoculated with cells precipitated at a centrifugal force of 67 x g showed apparently higher specific rates of metabolism compared to those inoculated with cells in the supernatant. The cells in the supernatant and the precipitate obtained following centrifugation at 67 x g have average diameters of 15.5 and 17.4 microm, respectively. The intracellular contents of amino acids, especially nonessential amino acids, of the precipitated cells were markedly higher than those of the cells in the supernatant. These results indicate that large cells with high amino acid content and metabolic activity were selectively retained in the culture by means of centrifugation at low forces such as 67 x g. Consequently, application of a low centrifugal force is recommended for medium change in order to maintain higher specific productivity of suspended mammalian cells in perfusion culture.

Journal Article↗
PubMed1995

Conductometric study of erythrocytes during centrifugation. II. Erythrocyte deformability.

Erythrocyte deformability was studied by continuous reading of sediment conductance during centrifugation. The decrease in sediment conductivity during centrifugation reflects erythrocyte deformation in the pellet. The degree of erythrocyte deformation depends on the duration of centrifugation and the magnitude of centripetal acceleration. When constant centrifugal force is applied over an extended period of time, a gradual decrease in pellet conductivity occurs. Stepwise enhancement of centripetal acceleration during centrifugation induces a rapid increase in erythrocyte deformation. After centrifugation, the relaxation of erythrocyte deformation is observed. However, the relaxation and the recovery of cell shape are incomplete. The difference in compressibility of previously centrifuged and noncentrifuged cells demonstrates that centrifugation causes irreversible alteration in erythrocyte deformability. The results show that the time-dependent resistance of erythrocyte sediment during centrifugation may serve as a useful index for the kinetics of erythrocyte deformation.

Blood Sedimentation↗
PubMed1988

Further studies on the mechanism of centrifugal enhancement of cytomegalovirus infectivity.

The phenomenon of centrifugal enhancement of infectivity, previously shown to be an intrinsic property of murine and human cytomegalovirus (CMV) was studied further in an attempt to understand the underlying mechanism of action and to try to influence the phenomenon by means of extrinsic factors. The sand-rat herpes virus also displayed centrifugal enhancement in rat, mouse, and human cells. Further investigations were performed with murine CMV. The property of enhancement was not restricted to a specific phase of cell cycle. A variety of factors, known to influence cellular physiology, did not significantly affect the ratio of centrifugal infectivity to standard infectivity i.e., the c/s ratio. However, it was necessary to centrifuge the infected cultures within one hour of the establishment of infection, since centrifugation after this delay failed to yield the enhancement; but there was no partial effect; the cultures either showed complete centrifugal enhancement or no such effect. Thus, it appears that some herpes viruses, during their initial encounter with susceptible cells, can be forced into preferential productive infection by the process of centrifugation, but after a lapse of an hour or more, centrifugation can no longer do this and most of the infected cells fail to replicate virus. Consequently we continue to recommend low speed centrifugation (1000 x g, for 30 min) in order to enhance CMV detection and to optimize infectivity assays.

Animals↗
PubMed1982

[Controlled centrifugation by a w2dt integrator. Application to blood transfusion].

The centrifugation constitutes a fundamental stage in the blood components preparation as fresh frozen plasma and platelets rich plasma. The measure of the total effect of centrifugation (VT) is achieved with an integrator "w2dt". This parameter characterizes, in an unique way, the preparation procedure of a blood component. The value VT is determined and then sticked on the integrator, which made the rotation stop as soon as the preselected effect of centrifugation is reached. Thanks to this accessory, the authors have analysed the relative importance of the three data: acceleration, constant speed and deceleration, on the total effect of centrifugation. This analysis has induced them to a new approach of the centrifugation. For the authors, the classic notion of rotation time at constant speed, become insufficient. In blood transfusion, several applications are possible. The authors have determined the optimal conditions of platelets rich plasma (yield higher than 80% during a minimum real time) and platelets concentrate preparation with two different centrifuges, Sorvall RC3B and Jouan K 110 SX, equipped with Sorvall Integrator. The reproductibility of the experiments is satisfying when the centrifuged weight is comparatively constant. The standardization of centrifugation technics will be easier when the rotation radius will be included in the calculation of the total effect of centrifugation, thanks to integration.

Blood Platelets↗
PubMed2001

Effect of centrifugation on early embryonic development and parthenogenetic activation of bovine oocytes matured in vitro.

This study examined the fertilization, early developmental competence and capacity for parthenogenetic activation of bovine oocytes matured in vitro after centrifugation. Immature oocytes were cultured in tissue culture medium 199 supplemented with 10% fetal bovine serum and 75 mIU mL(-1) FSH + LH at 5% CO2 to facilitate maturation. After culture for 24 or 30 h, the metaphase-II stage oocytes were centrifuged at 3000, 5000, 7000 or 10000g for 5 min before in vitro fertilization or parthenogenetic activation. Frozen-thawed bull semen was used for in vitro fertilization. For parthenogenetic activation, the oocytes were exposed to 20 microM calcium ionophore A23187 for 5 min at room temperature. Fertilization rates were not different between control and treatment groups (87.7% v. 74.6%, 73.4%, 75.9% and 76.4% respectively). Also, there were no differences in early embryonic development between control and treatment groups (rates of blastocyst formation were 21.1% v 20.2%, 28.8%, 31.2% and 24.1% respectively). When the oocytes were centrifuged at various speeds alone, the activation rate of oocytes was significantly higher (P < 0.05) in the 10000g treatment group compared with control (10.8% v 0.0%). There were no differences in the activation rates of oocytes between control and treatment groups at speeds up to 7000g (70.9% v. 71.9%, 78.3% and 77.2% respectively) after centrifugation and stimulation with Ca(2+)-ionophore. However, the activation rate of oocytes was significantly higher (P < 0.05) in the 10000g treatment group compared with control (70.9% v. 83.1%). In addition, the percentage of activated oocytes with diploid formation was significantly higher in the oocytes after centrifugation at 10000g and stimulation with calcium ionophore A23187 than in the control (18.4% v 7.1%). These results indicate that centrifugation of oocytes matured in vitro has no detrimental effect on fertilization and subsequent early embryonic development. They also indicate that the oocytes might be parthenogenetically activated after centrifugation and that high-speed centrifugation may induce activation of some oocytes. The results suggest that the optimal speed for centrifugation of bovine oocytes might be < or = 7000g to enhance the visibility of nuclear elements for further micromanipulation.

Animals↗
PubMed1999

Urea and osmotic excretion in rats exposed to chronic centrifugation.

BACKGROUND: A reduction in vasopressin was attributed to the initial diuresis reported in rats exposed to chronic centrifugation. However, it was suggested that urea may also contribute an osmotic component to this observed diuresis. HYPOTHESIS: Increased urea excretion will contribute to osmotic excretion during chronic centrifugation, which may be partly responsible for the initial diuresis previously observed. METHODS: Eight Sprague-Dawley rats were centrifuged (12 d at -2Gx) and eight were used as a control group. Daily urine samples were collected and an aliquot measured for excreted solutes and aldosterone. RESULTS: Urine volume was elevated over the first 7 d of centrifugation with a peak on day 4. Urea and osmotic excretion were elevated over the first 5 d. Excreted Na+ was elevated on days 1 and 2, which coincided with an increase in excreted aldosterone over the first 3 d of centrifugation. Urea excretion accounted for up to 54% of the increase in osmotic excretion during the initial portion of centrifugation suggesting that urea was, in part, responsible for the observed increase in urine output despite a reduction in water consumption. Following the first day of centrifugation, aldosterone appears to regulate Na+ as suggested by the reduction in Na+ excretion between days 2 and 3 when aldosterone excretion was elevated. CONCLUSIONS: It would appear that centrifugation induced an acute increase in protein catabolism as indicated by the increase in urea excretion which resulted in an increase in obligatory water loss. This increased diuresis may have acute consequences on the hydration state of centrifuged rats.

Aldosterone↗
PubMed2004

[Clinical application of intro-aortic balloon pump and centrifugal pump in low cardiac output syndrome after coronary artery bypass grafting].

OBJECTIVE: To explore the clinical application of Intra-aortic balloon pump (IABP) and centrifugal pump in low cardiac output syndrome (LCOS) after coronary artery bypass grafting (CABG). METHODS: From April 2000 to January 2004, 5 patients suffered serious LCOS after CABG in our department. Because maximum vasoactive agent had no significant effect, we supported these 5 patients with IABP and centrifugal pumps. The centrifugal pumps were connected with cannulas of right superior pulmonary vein and ascending aorta. The flow rate of the centrifugal pumps were increased to 3-4 L/min gradually. The dosage of vasoactive agent and flow rate of the centrifugal pumps were decreased gradually after hemodynamics stabilized. RESULTS: All 5 patients' hemodynamics were improved significantly after the left ventricular assist by IABP and centrifugal pumps. Five patients were weaned from the centrifugal pumps after 4 to 7 days successfully. But some degree damage to blood cells and renal function were detected. Renal function of 3 patients were recovered gradually after centrifugal pumps removed and discharged successfully, One patient died of acute renal failure, 1 patient died of multiple organ failure, 5 patients need transfusion of whole blood or concentrated erythrocytes and 4 patients need transfusion of platelets. CONCLUSIONS: There were significant improvements in hemodynamics after support with IABP and centrifugal pumps in the patients who suffered serious LCOS with less effect of maximum vasoactive agent after CABG. Some complications on blood cells and renal function should be paid more attention.

Aged↗
PubMed1983

The effect of centrifugal accelerations on the polarity of statocytes and on the graviperception of cress roots.

The structural polarity of statocytes of Lepidium sativum L. is converted to a physical stratification by a root-tip-directed centrifugal acceleration. Sedimentation of amyloplasts and nucleus to the centrifugal (distal) cell pole and the lateral displacement of the distal endoplasmic reticulum (ER) complex occur after centrifugation for 20 min at an acceleration of 50 g. With higher doses (20 min, 100-2,000 g), smaller organelles become increasingly displaced. From the centrifugal to the centripetal cell pole, the following stratification is observed: 1) amyloplasts with mitochondria; 2) nucleus with mitochondria and a few dictyosomes, as well as laterally located ER; 3) dictyosomes with a few mitochondria; 4) vacuoles; and 5) lipid droplets. Within the first 7.5 min, after the roots have been returned to 1 g, the original arrangement of the amyloplasts sedimented on the underlying ER complex is reestablished in 66% of the statocytes. When roots previously centrifuged in an apical direction are exposed in a horizontal position to 1 g, the latent period of the graviresponse is increased by 7.5 min relative to the non-centrifuged controls. The kinetics of the response are identical to the controls. Roots centrifuged first in an apical direction and then for 2 h in a lateral direction (1,000 g) have statocytes with a physical stratification perpendicular to the root axis. A gravitropic curvature does not take place during the lateral centrifugation. These results support the hypothesis that the distal ER complex is necessary and sufficient for graviperception.

Brassicaceae↗
PubMed2006

Evaluation of a cushioned method for centrifugation and processing for freezing boar semen.

The purpose of this investigation was to evaluate the use of an iodixanol cushion during centrifugation on sperm recovery and yield after centrifugation (sperm recovery, sperm motility, viability, membrane lipid disorder, acrosome reaction and ROS generation); and to investigate how this procedure affects sperm function after freezing-thawing (sperm motility, membrane lipid disorder, acrosomal status and homologous in vitro penetration test). The sperm-rich fractions from fertile boars were centrifuged under two centrifugation régimes: 800xg for 10min (standard method) and 1000xg for 20min with an iodixanol (60% w/v) cushion at the bottom of the centrifuge tubes (Cushion method). The highest recovery was achieved using the cushion method (sperm loss for cushion method was 0.50%+/-0.18 versus 2.97%+/-0.43 for standard method, P<0.01) and sperm quality was not significantly affected by the centrifugation régime. The motion parameters (% progressive motility, % motility, VCL, VSL, VAP, ALH, BCF, P<0.05) of frozen-thawed samples showed higher values using the standard method. However, a higher number of viable spermatozoa with lower lipid disorders were found in spermatozoa processed with the cushion method. The in vitro penetration assay showed that the individual boar influenced the parameters studied but there were no differences between the two centrifugation régimes used. Our results support the hypothesis that the proportion of sperm loss in frozen-thawed semen was significantly influenced by the centrifugation régime. Therefore, the iodixanol cushion method is a suitable tool for cryopreservation of boar semen in order to reduce sperm loss without affecting sperm quality.

Acrosome Reaction↗
PubMed2000

Effect of lipid polarization by centrifugation at different developmental stages on post-thaw survival of bovine in vitro produced 16-cell embryos.

The developmental rate to the blastocyst stage of frozen-thawed bovine in vitro produced embryos at stages earlier than Day 6 morula is not sufficiently high for practical utilization. The present study was undertaken to determine the effect of polarization of lipid droplets in the cytoplasm of bovine in vitro produced embryos from zygotes to the 8-cell stage, by centrifugation without following micromanipulation, on the survival rate of Day 4 16-cell embryos. After centrifugation at 15,500 x g in medium containing cytochalasin D, embryos were cultured to the 16-cell stage, classified as either mostly or partially delipidated by degree of lipid droplet removal, and then frozen. Embryos centrifuged at the 2-cell stage developed to the 16-cell stage similarly to those centrifuged at the 8-cell stage. The developmental rate to blastocysts after freezing of the mostly delipidated 16-cell embryos centrifuged at the 2-cell stage was higher than that of those centrifuged at the zygote stage, those that were partially delipidated at the 2-cell stage, and those that were not centrifuged. The results demonstrate that polarization of lipid droplets at the 2-cell stage by centrifugation without micromanipulation improved the survival rate of mostly delipidated 16-cell embryos after freezing.

Animals↗