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PubMed2006

Chance, choice and control: lay debate on prenatal social sex selection.

Assisted reproductive technologies are typically positioned as increasing the range of choices open to the healthcare consumer, thereby enhancing 'reproductive freedom'. In this paper, we question the equivalence of reproductive choice and personal freedom in ethical theory, using results from a project investigating how lay people make ethical evaluations about the new genetic and reproductive technologies. We took the topic of social sex selection by preimplantation genetic diagnosis (PGD), and used group discussions and interviews in the north-east of England to trace how lay people develop and express their ethical evaluations, and to identify the implicit or explicit normative framework that gave rise to their opinions on prenatal sex selection. There was a striking level of ambivalence towards choice in general and reproductive choice in particular. Participants offered few positive statements and numerous reasons why reproductive choice might be problematic. Our participants' argumentation shares with mainstream bioethical analysis the weighing of the possible harms of prenatal sex selection for social reasons against the harm of restricting reproductive freedom. However, unlike most secular-liberal bioethicists, many of our participants concluded that prenatal sex selection is undesirable because it is an expression of parental preference instead of a response to the future child's need. Our interpretation of their reasoning is that they work from an ideal of "good parents", one of the features of which is the relinquishing of control over their children, except to protect them from harm. This voluntary self-limitation does not indicate reduced autonomy, because parental autonomy can only operate within the limits set by this relational framework. We suggest that a model of relational autonomy captures our lay participants' framing of the problem better than a more traditional understanding of autonomy. Our study also shows that in appropriately structured discussion of bioethical issues, lay people can articulate reasons for their opinions that are grounded in sophisticated and morally relevant concepts.

Adolescent↗
PubMed2004

Production of piglets with sexed semen employing a non-surgical insemination technique.

The aim of the present study was to ascertain whether multiparous sows could successfully be inseminated with sexed semen non-surgically. Spermatozoa were stained with Hoechst 33342 and separated flowcytometrically in X- and Y-chromosome bearing sperm populations employing the Beltsville Sperm Sexing Technology (BSST). After weaning, estrus was induced in sows with PMSG and hCG. Animals were inseminated once per estrus non-surgically with a specially designed catheter into the tip of the uterine horn, employing 50x10(6) of either sexed or non-sexed spermatozoa diluted in 2 ml Androhep. Pregnant sows were allowed to go to term. Mean pregnancy rate from inseminations with unsexed spermatozoa was 54.5% whereas inseminations with sexed spermatozoa resulted in 33.3% pregnant sows. All but one piglet born after insemination with sexed semen were of the predicted sex. The sex of those piglets born after inseminations with non-sexed spermatozoa was 61.1% for male and 38.9% for female sex. It is concluded that non-surgically inseminations with flowcytometrically sexed spermatozoa can be conducted successfully.

Animals↗
PubMed2005

In vitro production of bovine embryos using sex-sorted sperm.

The objective of this study was to investigate the suitability of sex-sorted sperm for producing viable in vitro embryos for subsequent transfer into recipient cows and heifers on commercial dairy farms. From August 2002 to June 2003, ovaries were collected from 104 producer-nominated Holstein donor cows on seven Wisconsin farms via colpotomy or at slaughter. Oocytes (N=3526) were aspirated from these ovaries, fertilized 22+/-0.2h later, and cultured to the morula or blastocyst stage. The fluorescence-activated cell sorting ("Beltsville") approach was used to produce (primarily) X-bearing sperm from the ejaculates of three young Holstein sires, and 365 transferable embryos were produced. On average, 3.6+/-0.3 (means+/-S.E.M.) transferable embryos were produced per donor, including 1.4+/-0.2 (Grade 1), 1.5+/-0.2 (Grade 2), and 0.7+/-0.1 (Grade 3) embryos. Number of usable oocytes per donor (33.9+/-3.3) and percent cleavage (51.1+/-1.9) were significant predictors of the number of blastocysts that developed. Mean conception rates for the resulting in vitro embryos were 34.2+/-1.6% in yearling heifer recipients and 18.2+/-0.7% in lactating cow recipients. Additional oocytes (N=3312) from ovaries of anonymous donors (N unknown) collected at a commercial abattoir were fertilized using unsorted sperm, and the percentage of these that developed to blastocyst stage (20.1+/-2.9) was greater (P<0.05) than the corresponding percentage (12.2+/-2.3) achieved with sex-sorted sperm using oocytes (N=1577) from the same source. In summary, we inferred that in vitro embryo production may be a promising application of sex-sorted sperm in dairy cattle breeding, but that the biological causes of impaired embryo development in vitro and compromised conception rates of transferred embryos should be further investigated.

Animals↗
PubMed2005

Flow cytometric sexing of mammalian sperm.

This review reexamines parameters needed for optimization of flow cytometric sexing mammalian sperm and updates the current status of sperm sexing for various species where this technology is currently being applied. Differences in DNA content have provided both a method to differentiate between these sex-determining gametes and a method to sort them that can be used for predetermining sex in mammals. Although the DNA content of all cells for each mammalian species is highly conserved, slight but measurable DNA content differences of sperm occur within species even among cattle breeds due to different sizes of Y-chromosomes. Most mammals produce flattened, oval-headed sperm that can be oriented within a sorter using hydrodynamic forces. Multiplying the percentage the difference in DNA content of the X- or Y-chromosome bearing sperm times the area of the flat profile of the sperm head gives a simple sorting index that suggests that bull and boar sperm are well suited for separation in a flow sorter. Successful sperm sexing of various species must take into account the relative susceptibilities of gametes to the stresses that occur during sexing. Sorting conditions must be optimized for each species to achieve acceptable sperm sexing efficiency, usually at 90% accuracy. In the commercial application of sperm sexing to cattle, fertility of sex-sorted bull sperm at 2 x 10(6)/dose remains at 70-80% of unsexed sperm at normal doses of 10 to 20 x 10(6) sperm. DNA content measurements have been used to identify the sex-chromosome bearing sperm populations with good accuracy in semen from at least 23 mammalian species, and normal-appearing offspring have been produced from sexed sperm of at least seven species.

Animals↗
PubMed2005

Application of sexed semen technology to in vitro embryo production in cattle.

Use of sexed semen in conjunction with in vitro embryo production is a potentially efficient means of obtaining offspring of predetermined sex. For thousands of years, livestock owners have desired a methodology to predetermine the sex of offspring for their herds. The ability to sort individual sperm cells into viable X- and Y-chromosome-bearing fractions made producers' sex selection dreams reality in the 1990s and now semen can be sexed with greater than 90% accuracy with use of a flow cytometric cell sorter. Several concerns regarding the implementation of sexed semen technology include the apparent lower fertility of sorted sperm, the lower survival of sorted sperm after cryopreservation and the reduced number of sperm that could be separated in a specified time period. These issues are discussed in this review. There are also a number of issues that appear to influence the success rates of using sexed semen to produce bovine embryos in vitro. These issues include reductions in fertilization rates, lower cleavage rates, blastocyst rates and pregnancy rates, partial capacitation of the sperm, dilute sperm samples and sire variation. These subjects are also addressed in this paper. Finally, we will describe a recent field trial in which female Holstein embryos produced using the combined technologies of sex-selected semen and microfluidics were transferred either as single or bilateral twin embryos into beef cattle recipients, demonstrating these technologies' contributions to viable embryo production. The results indicate that large-scale transfer of in vitro produced, Holstein heifer embryos to beef recipients is a feasible production scheme.

Animals↗
PubMed2006

The influence of antioxidant, cholesterol and seminal plasma on the in vitro quality of sorted and non-sorted ram spermatozoa.

In an effort to improve the number of functional spermatozoa following sex-sorting and cryopreservation, the effects on in vitro sperm characteristics of the additives: (i) catalase (pre-sorting); (ii) cholesterol-loaded cyclodextrins (CLCs; pre-sorting); and (iii) seminal plasma (post-thawing) were investigated. For all experiments, spermatozoa (three males, n=3 ejaculates/male) were processed using a high speed flow cytometer before cryopreservation, thawing and incubation for 6h. Catalase had no effect (P>0.05) on post-thaw motility characteristics (as measured by CASA) of sex-sorted ram spermatozoa, but pre-sort addition of CLCs reduced (P<0.05) sperm quality after post-thaw incubation for 0 h (motility), 3h (motility, average path velocity, viability and acrosome integrity) and 6h (motility, average path and curvilinear velocity, straightness, linearity, viability and acrosome integrity). Seminal plasma had a differential effect (P<0.001) on sex-sorted and non-sorted spermatozoa. Post-thaw supplementation of increasing levels of seminal plasma caused all motility characteristics of sex-sorted, frozen-thawed spermatozoa to decline (P<0.05); conversely, non-sorted, frozen-thawed spermatozoa exhibited improvements (P<0.05) in motility, viability, acrosome integrity and mitochondrial respiration. In summary, incorporation of catalase, CLCs and seminal plasma into the sorting protocol failed to improve post-thaw sperm quality and, consequently efficiency of sex-sorting of ram spermatozoa. The paradoxical effect of seminal plasma supplementation on the in vitro characteristics of ram spermatozoa provides further evidence that sex-sorting by flow cytometry produces a selected population of cells with different functions compared with non-sorted spermatozoa.

Acrosome↗
PubMed2006

Birth of offspring of pre-determined sex after artificial insemination of frozen-thawed, sex-sorted and re-frozen-thawed ram spermatozoa.

The fertility of ram spermatozoa cryopreserved prior to, and following, sex-sorting by flow cytometry was assessed after insemination of mature Merino ewes at a synchronised estrus. Ewes were inseminated with spermatozoa from three rams, split into four treatment groups: 50 x 10(6) motile non-sorted, frozen-thawed (Control50), 15 x 10(6) motile non-sorted, frozen-thawed (Control15), 15 x 10(6) motile sex-sorted, frozen-thawed (SF15) or 15 x 10(6) motile frozen-thawed, sex-sorted, re-frozen-thawed (FSF15) ram spermatozoa. Separation of SF15 and FSF15 treatments into X- and Y-chromosome-bearing populations was achieved using a high-speed sperm sorter. The percentage of ewes lambing after insemination was similar for Control15 (36/74; 48.6%), SF15 (35/76; 46.1%) and FSF15 (26/72; 36.1%) groups (P>0.05). A higher percentage of ewes produced lambs in the Control50 (38/70; 54.3%) than the FSF15 group (P<0.05). Fifty-one of the 55 (92.7%) lambs derived from fresh, sex-sorted frozen-thawed spermatozoa were of the predicted sex, as were 41/43 (95.3%) lambs derived from frozen-thawed, sex-sorted, re-frozen-thawed spermatozoa. This study demonstrated for the first time in any species that frozen-thawed spermatozoa, after sex-sorting and a second cryopreservation step, are capable of producing offspring of the predicted sex following artificial insemination.

Animals↗
PubMed2006

Pregnancy percentage following deposition of sex-sorted sperm at different sites within the uterus in estrus-synchronized heifers.

Our objective was to assess the effect on heifer pregnancy rate of deposition at three sites within the uterus of frozen-thawed sex-sorted sperm at a fixed time after estrus synchronization. Estrus was synchronized in 209 heifers by administration of PGF2a 14 days apart. At 80-82 h after the second PGF2a injection, X-chromosomes bearing fractions of semen with 2.2 x 10(6) sperm in insemination dose were used for single insemination into the uterine body (UB-AI, n=91) or for intracornual deposition in the middle of the uterine horn (MH-AI, n=57) or close to the utero-tubal junction (UTJ-AI, n=61). The overall pregnancy rate was 43.1%. Pregnancy rates did not differ (P>0.05) among sites of sperm sperm deposition, between the two farms at which the heifers were kept or between the two bulls producing the semen. Within UB-AI, MH-AI and UTJ-AI treatments, pregnancy rates were 41.8%, 49.1% and 39.3%, respectively (P>0.05). Pooled across classes for deposition site, pregnancy rate was 25.1% higher (P<0.01) for heifers showing strong signs of estrus than for heifers showing weak signs of estrus (45.9 versus 20.8%, respectively). Embryonic and fetal loss from diagnosis of pregnancy to term and at calving equalled 5.6%. Of 88 calves of identified sex, 93.2% were female. In conclusion, pregnancy rates of heifers did not differ significantly following deposition of 2.2 x 10(6) sex-sorted sperm 80-82 h after the second PGF2a injection near the utero-tubal junction, in the middle of the horn or into the uterine body.

Animals↗
PubMed1997

Evaluating sex chromosome content of sorted human sperm samples with use of dual-color fluorescence in situ hybridization.

OBJECTIVE: Although most methods for selecting the sex of offspring by sorting spermatozoa are ineffective at shifting the ratio of Y- to X-containing cells, some commercial sources continue to offer such services. Our objective was to evaluate commercially "sorted" samples with use of dual-color fluorescence in situ hybridization and to identify variations in assessment by comparing motile and total sperm populations, donors, observers, and fluorescence in situ hybridization probes. STUDY DESIGN: Cryopreserved sperm from seven anonymous donors were processed as for insemination. Sperm cells from each total sample or motile subfraction were prepared for fluorescence in situ hybridization by incubation with disulfide-reducing agents to expand sperm nuclei. Two sets of X and Y chromosome-specific, fluorophore-labeled deoxyribonucleic acid probes were used. At least 400 nuclei from each preparation were classified independently by three blinded observers. Hybridization efficiency, aneuploidy, and sex chromosome content were evaluated in subsets of five unsorted, five female-oriented, and five male-oriented samples. Total and motile subfractions were compared with eight samples. Fluorescence in situ hybridization probes were compared in five paired unsorted samples. RESULTS: No differences were detected between washed samples and paired motile subfractions. No differences in hybridization and aneuploidy were detected between groups of sorted samples. The Y/X ratio was significantly different between the sorted groups. However, male-oriented samples had a lower Y/X ratio than female-oriented samples did. Observer and probe choice accounted for small but significant variations that did not alter conclusions about the X/Y ratio for sorted samples. CONCLUSION: In a series of 10 sorted samples from one commercial source, dual-color fluorescence in situ hybridization demonstrated a small but significant shift in the sex chromosome ratios among samples. However, this shift was opposite to that expected by the orientation of the sorted samples.

Analysis of Variance↗
PubMed1976

The effect of ion-exchange column chromatography on separation of X and Y chromosome-bearing human spermatozoa.

Separation of X and Y chromosome-bearing spermatozoa has been attempted using ion-exchange column chromatography, with cation- and anion-exchange resins of low, intermediate, and high ionic strength. Examination of F-bodies on the Y chromosome of treated human sperm and progeny resulting from insemination of treated rabbit spermatozoa indicates that in none of the cases investigated did the treatment cause a separation of X and Y chromosome-bearing spermatozoa. The treatment does appear to filter out dead rabbit (and bull) spermatozoa, but the possible beneficial effects of this phenomenon are as yet uninvestigated.

Animals↗
PubMed1980

Maternal serum human chorionic gonadotropin concentrations and fetal sex prediction.

The feasibility of using maternal serum human chorionic gonadotropin (hCG) concentrations for prenatal sex prediction was examined. hCG was mesured in 822 serum samples from 560 women with uncomplicated pregnancies. Significantly higher hCG concentrations were found in the serum of women bearing female fetuses than in the serum of women bearing male fetuses during the third trimester, especially during the 10th lunar month. The data were utilized to construct probability graphs for fetal sex prediction based upon a single maternal serum hCG determination during the third trimester and during the 10th lunar month. However, the utility of these graphs is limited by the small proportion of pregnant women with serum hCG concentrations that were high or low enough to allow a prediction with high probability.

Chorionic Gonadotropin↗