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Biomedical subjects

J R Correa

Publications and source records attributed to J R Correa.

At least 19 recordsLinked to original sources
PubMed2005

Adiabatic formation of quasibound states of antihydrogen.

The classical trajectory of an initially unbound positron within the electric field of an antiproton and a uniform magnetic field is simulated in three dimensions. Several simulations are run incorporating experimental parameters used for antihydrogen production, which has been achieved by two different groups [M. Amoretti, Nature (London) 419, 456 (2002); G. Gabrielse, Phys. Rev. Lett. 89, 213401 (2002)]. The simulations indicate that temporary bound states of antihydrogen can form at positive energies, where the energy of the system is defined to be zero when the positron and antiproton are at rest with infinite separation. Such quasibound states, which form only when the magnetic field is present, are typically smaller than in a dimension perpendicular to the magnetic field. An analytical model is developed for a formation cross section, and it is found that quasibound states may form more frequently than stable Rydberg states.

Journal Article↗
PubMed1998

Effects of seminal plasma from cigarette smokers on sperm viability and longevity.

OBJECTIVE: To evaluate the effects of cigarette smoking on the ability of seminal plasma (SP) to maintain sperm viability. DESIGN: Clinical randomized study. Spermatozoa from cigarette smoking or nonsmoking subjects were reconstituted in SP from smokers and nonsmokers and in modified Ham's F-10 medium, followed by sperm quality assessment during a 48-hour incubation period. SETTING: Andrology Institute of Lexington, Lexington, Kentucky. PATIENT(S): Twenty men who had been smoking cigarettes for longer than 3 years (30 cigarettes per day or more) and 20 nonsmokers participated in this study. MAIN OUTCOME MEASURE(S): Improvement in sperm viability by removal of SP--and associated detrimental factors present in the SP--from smoker subjects. RESULT(S): The results obtained indicate that the quality of spermatozoa obtained from nonsmokers was superior to that of smokers. The SP from the two patient groups had a definite effect on their respective sperm quality, i.e., beneficial effects for the nonsmokers, detrimental effects for the smokers. Exposure of spermatozoa from the nonsmokers to SP from the smokers resulted in a significant reduction in sperm viability. However, exposure of spermatozoa from the smokers to SP from the nonsmokers or to Ham's F-10 medium yielded significant improvements in sperm viability. CONCLUSION(S): The detrimental effects of smokers' SP on nonsmokers' spermatozoa was prominent and a rather unique phenomenon. The results generated in this study could be of clinical significance since removal of smokers' SP and subsequent reconstitution and incubation in physiological media seems to enhance the viability, longevity, and possibly the fertilizing ability of these spermatozoa for use in various assisted reproductive technologies.

Adult↗
PubMed1998

Antisperm antibody treatment mode: levels of antisperm antibodies after incubation with TEST-yolk buffer and filtration using the SpermPrep II method.

OBJECTIVE: To assess whether incubation in TEST-yolk buffer (TYB) or human tubal fluid (HTF) could alter the sperm membrane characteristics and its relationship to antisperm antibodies (ASA) and/or antigen detachment from the sperm membrane and to evaluate the filtration of those specimens and possible recovery of ASA-free spermatozoa. DESIGN: A prospective clinical study. SETTING: Andrology Institute of Lexington, Lexington, Kentucky. PATIENT(S): Twenty patients undergoing infertility treatment. MAIN OUTCOME MEASURE(S): Recovery of spermatozoa with reduced levels or antisperm antibody-free sperm after treatment with TYB or HTF, followed by filtration using the SpermPrepII method (Sephadex based). RESULT(S): Assessment of ASA using the direct immunobead test showed no significant differences between specimens incubated for 2 hours in seminal plasma (fresh) or HTF with regard to levels of IgA and IgG. The percentage binding of anti-IgA and anti-IgG immunobeads was significantly reduced in specimens incubated for 2 hours in TYB compared with specimens incubated in seminal plasma or HTF. Furthermore, selection of spermatozoa using the SpermPrepII filtration method significantly reduced the percentage binding of anti-IgA and anti-IgG immunobeads compared with specimens incubated in HTF. CONCLUSION(S): The results suggest that TYB either altered the sperm membrane properties so that there was a decreased affinity at the antibody and/or antigen sites or that the egg yolk proteins were absorbing the antibodies and/or antigens complexes from the sperm membrane surface. Incubation of spermatozoa in TYB followed by filtration with the SpermPrepII method improved the recovery of ASA-free spermatozoa by selectively entrapping spermatozoa with ASA bound to its surface.

Antibodies, Anti-Idiotypic↗
PubMed1998

An electron microscope study of the axonemal ultrastructure in human spermatozoa from male smokers and nonsmokers.

OBJECTIVE: To investigate possible abnormalities or deterioration of the sperm axonemal ultrastructure in men who have smoked a large quantity of cigarettes (> 20 per day) for a prolonged period. DESIGN: Semen specimens were collected by patients via masturbation; qualitative characteristics of the sperm were assessed and ultrastructural analysis of the sperm axoneme was performed using standard operating procedures for electron transmission microscopy. SETTING: The Andrology Institute of Lexington, Lexington, Kentucky, and the Department of Histology and Embryology, University of Salonika, Greece (collaborative effort). PATIENT(S): Twenty-nine men (mean age +/- SD, 30.7 +/- 2.1 years) who smoked a mean (+/- SD) of 30.7 +/- 2.1 cigarettes per day for 10.7 +/- 0.7 years and 15 men who never smoked (mean age +/- SD, 30.4 +/- 2.2 years) participated in this study. MAIN OUTCOME MEASURE(S): Ultrastructural organization of the sperm axoneme in male smokers and nonsmokers. RESULT(S): Changes in the number and the arrangement of axonemal microtubules were noted in the smoker group when compared to the nonsmoker group. The incidence of axonemal abnormalities was higher in spermatozoa from smokers compared with that in spermatozoa from nonsmokers. CONCLUSION(S): Smoking a large quantity of cigarettes per day, under the conditions of the current study, severely affected the ultrastructure of the flagellum and, more specifically, it affected the axoneme of the human spermatozoon.

Humans↗
PubMed1998

Assessment of a tablet drug delivery system incorporating nonoxynol-9 coprecipitated with polyvinylpyrrolidone in preventing the onset of pregnancy in rabbits.

OBJECTIVE: To assess the in vivo efficacy of the tablet drug delivery system containing nonoxynol-9 coprecipitated with polyvinylpyrrolidone by delivering the spermicidal agents vaginally and evaluating their ability to prevent the onset of pregnancy in rabbits. DESIGN: Controlled clinical study. SETTING: Division of Laboratory and Animal Resources, College of Pharmacy, University of Kentucky. ANIMAL(S): Forty-two New Zealand White female rabbits. INTERVENTION(S): The rabbits were artificially inseminated at various intervals after vaginal insertion of the tablet drug delivery system containing either polyvinylpyrrolidone only (0 minutes) or nonoxynol-9 coprecipitated with polyvinylpyrrolidone (polyvinylpyrrolidone/nonoxynol-9; 0, 3, 30, 180, and 360 minutes). The rabbits were induced to ovulate 6 hours before insemination by i.m. injection of hCG (200 IU). MAIN OUTCOME MEASURE(S): The onset of pregnancy in the rabbits was evaluated after insertion of the tablet drug delivery system containing polyvinylpyrrolidone only or polyvinylpyrrolidone/nonoxynol-9 at various intervals, followed by artificial insemination. RESULT(S): The onset of pregnancy was not reduced significantly when the tablet drug delivery system containing polyvinylpyrrolidone or polyvinylpyrrolidone/nonoxynol-9 was used and insemination was performed immediately after tablet insertion (time 0). However, pregnancy rates (PRs) were reduced significantly in the rabbits that received the tablet drug delivery system containing polyvinylpyrrolidone/nonoxynol-9 and were inseminated at 3, 30, 180, and 360 minutes after tablet insertion. The highest PR reduction occurred between 30 and 180 minutes after insertion of the tablet drug delivery system containing polyvinylpyrrolidone/nonoxynol-9. CONCLUSION(S): The tablet drug delivery system is an efficient method of delivering the tested spermicidal agents vaginally. The design and dosage used in preparing the tablet drug delivery system provide short- and long-term release of the spermicidal agents, which results in almost immediate and extended enhancement of their contraceptive properties.

Administration, Intravaginal↗
PubMed1998

Fertilization potential and qualitative characteristics of human spermatozoa after short-term cryostorage at 5 degrees C in two different TEST-yolk buffer preparations.

Use of the media TEST-yolk buffer (TYB) in semenology today enables the short-term incubation and cryostorage of spermatozoa and its subsequent use in the various assisted reproductive technologies (ART). Preparation of TYB media involves the addition of egg yolk (20% v/v) to a physiological solution of the zwitterion buffers TES and Tris. The TYB is usually thermoprecipitated to remove the majority of the egg yolk globules and other macromolecules from the medium. However, removal of these egg yolk constituents could possibly eliminate or reduce essential factors that could enhance the sperm viability and fertilization potential after short-term dilution and storage. Improvements in the quality of the TYB could add greater benefits to those techniques employed in the various forms of ART. The objectives of the investigation were 1) to study the sperm qualitative characteristics following short-term cryostorage at 5 degrees C in either thermoprecipitated (T-TYB) or non-thermoprecipitated (NT-TYB), and 2) to compare the fertilizing potential of spermatozoa stored for 24 hours at 5 degrees C in the two TYB preparations. In Experiment 1, semen specimens from 15 patients were collected, assessed and split into two aliquots. Sperm specimens were processed by diluting 1:1 (v/v) with the T-TYB or NT-TYB, followed by centrifugation and reconstitution of the specimen to its initial volume with the corresponding TYB medium. Sperm specimens were cryostored for 1, 2, 24, 48 and 72 hours. Samples were taken at each interval and placed in a 37 degrees C water bath and allowed to warm for 15 minutes after each cryostorage interval. Semen specimens were assessed for percentage and grade of motility. The results of this study indicated that, although the NT-TYB yielded better results than the T-TYB, overall those differences were not statistically significant. In Experiment 2, the fertilization potential of spermatozoa recovered after 24 hours of cryostorage in the two TYB preparations and further prepared via filtration, was assessed by the sperm penetration assay (SPA) using zona-free hamster oocytes. The average penetration rate (PR) and penetration index (PI) were significantly better for the NT-TYB than for the T-TYB. The PR was 54% vs. 25%, and the PI 0.78 and 0.27 for spermatozoa incubated in the NT-TYB vs. T-TYB. The range of penetration was also much lower for the T-TYB (6 to 100%) preparation when compared to the NT-TYB (22 to 100%). The highest penetrator showed 100% for both preparations. However, the lowest penetrator showed 6% for the T-TYB and 22% for the NT-TYB. The data obtained in this study suggest that both TYB preparations can be employed in short-term cryostorage (5 degrees C) of human spermatozoa and can adequately maintain the qualitative characteristics of those spermatozoa. The data also showed that the NT-TYB preparation yielded sperm samples of higher fertilization potential, thus possibly establishing the superior usefulness of the NT-TYB in an ART program.

Animals↗
PubMed1998

Evaluation of techniques for the cryopreservation of washed spermatozoa: comparisons between Ham's F-10 and TEST-yolk media.

The objective of this study was to develop new techniques for the cryopreservation of washed spermatozoa. Two media (Ham's F-10 and nonthermoprecipitated TEST-yolk buffer [NT-TYB]) containing 7% (v/v) glycerol were compared to semen cryopreservation by adding glycerol directly to the semen. Twenty four men collected a semen specimen each after 4 days of sexual abstinence via the use of a semen collection device at intercourse. Specimens were assessed for volume (ml), count (x 10(6)), percentage and grade of motility, morphology (% normal) and acrosomal status (% intact acrosomes). Each ejaculate was split into 3 aliquots (Aliquots 1 to 3) and processed for freezing. Aliquot 1 was prepared for cryopreservation by adding glycerol (7% [v/v] final concentration) directly via a dropwise mode. Aliquot 2 and 3 were diluted 1:1 (v/v) with Ham's F-10 and NT-TYB, respectively. Aliquots 2 and 3 were then centrifuged (400 x g for 10 minutes) and resuspended into the corresponding media containing 7% (v/v) glycerol to complete the sperm wash procedure. All aliquots were frozen in 0.5 ml french straws. Sperm specimens were frozen in liquid nitrogen (LN2) vapor from +23 degrees C to -68 degrees C at a slow rate (2.3 degrees C/minute), after which the specimens were plunged directly into LN2 and stored for 30 days. The quality of the spermatozoa were monitored throughout each step of the overall procedure by measuring the motility characteristics of the spermatozoa. Straws corresponding to each aliquot were thawed in a water bath at 37 degrees C for 2 minutes, followed by assessment of sperm motility and acrosomal status. The percentage of motility after thawing was 31.6 +/- 5.6%, 32.8 +/- 1.8% and 37.3 +/- 1.9% in Aliquots 1 to 3, respectively. Similarly, the grade of motility was 2.4 +/- 0.2, 2.6 +/- 0.1 and 3.0 +/- 0.1 in Aliquots 1 to 3, respectively. The acrosomal status (% intact acrosomes) in Aliquots 1 to 3 was 41.2 +/- 2.6, 43.1 +/- 3.6 and 51.6 +/- 4.5, respectively. The results suggest that the characteristics of spermatozoa washed and frozen in NT-TYB (Aliquot 3) were improved over those spermatozoa prepared via direct addition of glycerol to the semen (Aliquot 1) or by using Ham's F-10 (Aliquot 2). The most significant reduction noted during freezing was in the loss of acrosomal integrity. The results obtained in this study point out that washed spermatozoa can be cryopreserved with some success and that the recovered spermatozoa could be used for intrauterine insemination in an artificial insemination program using husband's or donor sperm, or for the various assisted reproductive technology procedures. It is the opinion of the authors that the information generated in this study is of importance for those scientists and clinicians involved in the handling and manipulation of cryopreserved spermatozoa.

Buffers↗
PubMed1997

Relationships among frozen-thawed sperm characteristics assessed via the routine semen analysis, sperm functional tests and fertility of bulls in an artificial insemination program.

Frozen semen specimens from 22 Holstein bulls representing a wide range of field fertility levels or nonreturn rates (NRR) were used in this study. Semen specimens were thawed at 37 degrees C for a minimum of 30 sec, followed by assessment via a routine semen analysis (RSA) and other sperm functional tests. The RSA was performed by assessing sperm count, motility and morphological characteristics. Other sperm functional tests were performed by assessing the acrosomal membrane integrity, sperm penetration into the cervical mucus and the sperm membrane functional integrity. Following assessment of sperm characteristics, the fertility data of the various bulls were compared to the RSA and the functional tests results. Bulls of high and low fertility were similar in terms of sperm count and progressive motility (P > 0.05). Other characteristics measured by the RSA and functional tests were significantly higher in high fertility bulls (P < 0.05). Correlation coefficients among the various sperm characteristics and fertility of bulls were highly significant (P < 0.01). The highest correlation coefficients between sperm characteristics and fertility were obtained for motility (r = 0.53; P < 0.01), normal morphology (r = 0.59; P < 0.01) and swollen spermatozoa (r = 0.57; P < 0.01). Analysis of specific sperm swelling patterns showed that those patterns considered to reflect maximal sperm swelling were indicative of high fertility.

Journal Article↗
PubMed1997

Vaginal delivery of new formulations of nonoxynol-9 coprecipitated with polyvinylpyrrolidone in rabbits. Comparisons between two formulation delivery systems.

The objective of this study was to assess the efficacy of two formulation delivery systems (FDS) in preventing the onset of pregnancy in rabbits. Nonoxynol-9 (N-9) was coprecipitated with polyvinylpyrrolidone (PVP), which yields PVP/ N-9, and prepared as capsules or tablets. Semen specimens were collected (from eight male rabbits), pooled, and used for in vitro spermicidal assessment or artificial insemination (AI). In vitro spermicidal assessment was performed by introducing and mixing the FDS containing PVP or PVP/N-9 with 1.0 mL of semen, followed by incubation at 37 degrees C for 6 h. Semen samples were taken at various time intervals to determine killing of spermatozoa and dissolution of the FDS. The efficacy of the FDS in preventing the onset of pregnancy was assessed by inserting the FDS vaginally. The does were artificially inseminated at 0, 0.5, and 6 h after insertion of the FDS vaginally. The number of pregnant does and newborn rabbits was recorded. In the in vitro spermicidal trial, semen specimens exposed to both FDS containing PVP/N-9 were killed within 10 to 15 min of incubation. Tablets containing PVP only or PVP/N-9 dissolved completely after 3 h of incubation. However, capsules did not dissolve completely by 6 h of incubation. The results obtained in the in vivo trial showed that both FDS exhibited some variations in preventing the onset of pregnancy over the various time intervals following the insertion of tablets or capsules and AI. The tablet seemed to be a more efficient delivery system than the capsule, yielding significantly lower pregnancy rates at all three time intervals assessed. The tablet FDS, as applied in this study, was found to be the most efficient mode of delivery of the tested spermicidal formulations.

Administration, Intravaginal↗
PubMed1997

Sperm membrane functional integrity and response of frozen-thawed bovine spermatozoa during the hypoosmotic swelling test incubation at varying temperatures.

The objective of this study was to assess the sperm membrane integrity and permeability of frozen-thawed bovine spermatozoa, processed at varying temperatures during and after thawing, by exposing the spermatozoa to standardized hypoosmotic conditions. The hypoosmotic swelling (HOS) test was employed to measure changes in sperm membrane functional status and permeability. Frozen specimens (from 5 bulls) were thawed at 37h degrees C for 10 sec and transferred to a water bath at 37 (Aliquot 1), 21 (Aliquot 2) or 5 degrees C (Aliquot 3) to complete thawing (1 to 2 min). The specimens were maintained and processed at these temperatures for additional 5 to 10 min. Specimens were slowly diluted 1:1 (v/v) and washed with Ham's F-10 media containing 3% (w/v) BSA. The HOS test was performed by adding 0.1 ml of the sperm specimen to 1.0 ml of a 100 mOsm/L HOS diluent. The following treatments were performed: 1) Aliquot 1 (control), specimens were incubated in HOS solutions at 37 degrees C for 5 min; 2) Aliquot 2, specimens were incubated in HOS solutions at 21 or 37 degrees C for 5 min; and 3) Aliquot 3, specimens were incubated in HOS solutions at 5 or 37 degrees C for 5 min. Samples were obtained from the sperm specimen-HOS diluent mixtures at 1 min intervals (during the 5 min incubation period), fixed and assessed for sperm swelling patterns. The sperm response to the HOS test for specimens processed at temperatures below 37 degrees C was higher when samples were incubated in HOS diluents at 37 degrees C. This finding indicates that the potential for sperm swelling (measurement of sperm membrane functional status) can be maintained when spermatozoa are processed at temperatures below 37 degrees C. The highest response to the HOS test was observed in spermatozoa processed at 21 degrees C and incubated in a HOS solution at 37 degrees C. The response to the HOS test was superior to the one observed in specimens maintained and processed at 37 degrees C throughout. Thawing of spermatozoa at 37 degrees C, followed by processing at 21 degrees C seems to reduce the negative effects associated with osmotic shock and results in the preservation of the sperm membrane functional status during the in vitro handling of frozen-thawed bovine spermatozoa.

Journal Article↗
PubMed1997

Quantitative and qualitative characteristics of frozen-thawed bovine spermatozoa recovered via a conventional and a standardized swim-up technique.

The objective of this study was to use the bovine as a model to evaluate the recovery of frozen-thawed spermatozoa via a conventional and a standardized swim-up technique. Frozen-thawed semen specimens (3 bulls) were washed and reconstituted with 2.9% (w/v) sodium citrate extender containing 20% (v/v) chicken egg yolk (SC-EY). Reconstituted sperm specimens were used for selection via conventional swim-up and the standardized ZSC method. The swim-up method consisted of overlayering the sperm specimen with 0.7 to 1.0 ml of isolation media (Ham's F-10), followed by 1 hr of incubation. The ZSC consisted of a conical cavity on the bottom of a glass column. The sperm specimen was placed into the conical cavity until the surface of the specimen was at the same level as the upper boundaries of the conical cavity. The surrounding periconical and epiconical areas were filled with 0.7 to 1.0 ml of isolation media followed by 1 hr of incubation. The isolation media was removed (harvesting) from swim-up (80% volume) and ZSC specimens (100% volume) at the end of incubation. Recovered specimens were assessed for volume (ml), sperm concentration (x10(6) spermatozoa/ml), the percentage and grade of motility (0 to 4), the occurrence of osmotic shock and the percentage of spermatozoa reactive to the hypoosmotic swelling (HOS) test. Swim-up and ZSC selected specimens were qualitatively similar to each other. However, higher numbers of spermatozoa were recovered when sperm specimens were processed via the ZSC method (1.6 fold increase) than with the conventional swim-up technique. Because the ZSC method enabled the recovery of up to 100% of the overlayered media, it also enabled the recovery of most of the spermatozoa that migrated from the sperm specimen into the isolation media with no possibility of mixing the two, which was the case with the swim-up method, and which could also contaminate the recovered specimen with dead and immotile spermatozoa. Thus, the ZSC technique enabled the harvesting of the medium closest to the underlayered sperm specimen, which contributed to maximize the number of sperm recovered. When all assessed parameters were noted and all clinical improvements and efficiency of the method were compared to the swim-up technique, the sperm manipulation procedure of choice was clearly the ZSC method.

Animals↗
PubMed1996

Preparation and recovery of frozen-thawed bovine spermatozoa via various sperm selection techniques employed in assisted reproductive technologies.

A number of semen manipulative techniques are currently available to remove the undesirable spermatozoa, debris and other factors and to increase sperm quality. The use of motility stimulants such as caffeine or others could optimize the recovery and quality of frozen-thawed spermatozoa processed by a variety of sperm selection techniques. Frozen-thawed specimens from 5 bulls were slowly diluted and washed with Ham's F-10 medium containing 3% BSA (w/v) and 0 or 2 mM caffeine. Aliquots containing approximately 50 x 10(6) total sperm cells were used for conventional sperm wash, swim-up, Percoll density gradient centrifugation (80, 70, 55 and 40% Percoll gradients) and Sephadex (SpermPrep I) filtration. Quantitative and qualitative characteristics of selected spermatozoa included: total sperm (x 10(6)), percentage and grade (0 to 4) of motility, percentage of spermatozoa with coiled tails and response to the hypoosmotic swelling (HOS) test (percentage of swollen spermatozoa). When compared to washed specimens, fewer spermatozoa were recovered via the swim-up, Percoll and SpermPrep I filtration methods. Quantitative and qualitative characteristics of these spermatozoa improved further after processing with Ham's F-10 containing 2 mM caffeine, followed by selection via the various techniques. Enhancement of sperm motility, in conjunction with the most appropriate sperm selection technique, represents an efficient method for the recovery of spermatozoa with improved qualitative characteristics.

Journal Article↗
PubMed1996

Comparative spermicidal performance of iodinated and non-iodinated contraceptive formulations of nonoxynol-9 co-precipitated with polyvinylpyrrolidone.

The objective of this study was to evaluate the spermicidal qualities of various combinations of nonoxynol-9 (N-9; whole molecule = oligomers 1-18) and its isolated fractions (oligomers 8-10, 4-6 and 1-3), co-precipitated with non-iodinated and/or iodinated (Io) polyvinylpyrrolidone (PVP) as possible vaginal contraceptives. Spermicidal qualities of known equimolar concentrations of various combinations of PVP/N-9 and PVP-Io/N-9 were tested via a modified Sander-Cramer test (SCT) using human spermatozoa. Spermicidal agents and semen samples were mixed 1:1 (v/v) and evaluated for sperm viability. Spermicidal activity was reported as the minimal concentration (microgram/mL) of spermicide capable of killing all spermatozoa within 20 sec after exposure to the spermicide. The spermicidal activity of PVP/N-9 and PVP-Io/N-9 preparations containing N-9 oligomers 1-18 and/or 8-10 was similar, and these preparations were more efficient in killing the spermatozoa than the ones containing N-9 oligomers 4-6 and 1-3. Polyvinyl-pyrrolidone proved to be an effective vehicle for PVP/N-9 and PVP-Io/N-9 preparations, especially those containing N-9 oligomers 4-6 and 1-3. Incorporation of Io into the spermicidal preparations brought about additional efficacy. The current findings could be of clinical significance in future studies when preparing and delivering those selected co-precipitates vaginally.

Cell Survival↗
PubMed1996

Thawing and processing of cryopreserved bovine spermatozoa at various temperatures and their effects on sperm viability, osmotic shock and sperm membrane functional integrity.

The objective of this study was to evaluate the effects of thawing and processing temperatures on post-thaw sperm viability, occurrence of osmotic shock and sperm membrane functional status. The occurrence of osmotic shock, characterized by increased spermatozoa with coiled tails, eventually results in reduced sperm viability and sperm membrane integrity. The effects of different thawing temperatures were assessed by thawing frozen specimens at 37, 21 or 5 degrees C for 1 to 2-min, followed by processing at these temperatures. A subset of frozen specimens were thawed at 37 degrees C for 10 to 15-sec and transferred to a water bath at 21 or 5 degrees C for 1 to 2-min to complete thawing, followed by processing at these temperatures. Sperm processing (washing) consisted of dilution, centrifugation and resuspension to remove glycerol from the medium and to gradually return the spermatozoa to isotonic conditions. Post-thawed specimens (0.5 mL) were slowly diluted 1:1 (v/v) at a rate of 0.1 mL/min, centrifuged, and resuspended to 0.5 mL (37 degrees C). Diluted specimens were equilibrated for 1 to 2-min after dilution and for 5-min after resuspension. The specimens were then incubated for 2-h (37 degrees C) and assessed at 60-min intervals for the percentage of motility, for progressive motility (Grades 0 to 4), for the percentage of spermatozoa with coiled tails, and for the percentage of swollen spermatozoa. The percentage of swollen spermatozoa (measurement of sperm membrane integrity) was assessed by exposing spermatozoa to a modified hypoosmotic swelling (HOS) test. The results obtained seem to indicate that physiological thawing and processing temperatures (37 degrees C) are required to maintain sperm motility. However, thawing and processing at lower temperatures (< 37 degrees C) seems to prevent the occurrence of osmotic shock and to maintain sperm membrane functional integrity. In this study, thawing at 37 degrees C (10 to 15-sec) and transfer to a water bath at 21 degrees C (1-min) to complete thawing, followed by processing at 21 degrees C, yielded better results in terms of increased sperm viability, reduced occurrence of osmotic shock and higher reactivity to the HOS test.

Journal Article↗
PubMed1996

Measurement of the sperm motility index via the sperm quality analyzer and its relationship to other qualitative sperm parameters.

Sperm parameters such as the concentration and percentage of motile spermatozoa are commonly used to assess semen quality. The sperm quality analyzer (SQA) is a device that detects variations in the optical density of motile spermatozoa, providing a sperm motility index (SMI) that is based on various sperm parameters including the concentration, morphology and acrosomal status of motile spermatozoa. The relationship between SMI values of frozen-thawed bovine spermatozoa undergoing swelling in a hypoosmotic medium (100 mOsm/L) and other sperm parameters were evaluated. Frozen semen specimens from 3 bulls were thawed and washed with Ham's F-10 supplemented with 3% BSA and split into 3 (0.2 mL) aliquots. The aliquots were diluted with 1.0 mL of Ham's F-10 (Aliquot 1), isotonic sodium citrate (Aliquot 2), and hypotonic sodium citrate (Aliquot 3). The osmotic pressure of the media used for dilution of Aliquots 1 and 2 was 300 mOsm/L, while that for Aliquot 3 was 100 mOsm/L. Following dilution, the aliquots were incubated for 30 min and manually assessed at 5-min intervals for the percentage and grade of motility (Grades 0 to 4) as well as for the percentage of swollen spermatozoa. Sperm samples were simultaneously evaluated by SQA to obtain the SMI values at the same 5-min intervals during the 30-min incubation. Significant correlations were observed between SMI values and other sperm parameters in Aliquot 3 (P < 0.05). The results indicated that the SMI values obtained from frozen-thawed bovine spermatozoa exposed to a 100 mOsm/L diluent, which causes optimal swelling of spermatozoa, are highly correlated to other sperm parameters. The SQA unit, as applied in this study, can be used for rapid and reliable screening of sperm samples.

Journal Article↗