Biomedical subjects

B C Su

Publications and source records attributed to B C Su.

14 recordsLinked to original sources

PubMed1995

Sperm as a noninvasive gene delivery system for preimplantation embryos.

OBJECTIVE: To determine if sperm could be manipulated to be a noninvasive transport carrier for the delivery of gene fragments to the blastocyst. DESIGN: Sperm cells carrying foreign DNA fragments from human papillomavirus (HPV) types 16, 18, 31, and 33 were allowed to migrate from one end of an artificial reproductive tube and to come in contact with hatching mouse blastocysts at the other end of the tube. The blastocysts were then washed and analyzed for the presence of the foreign DNA fragments. SETTING: Clinical and academic research environment. MAIN OUTCOME MEASURE: Detection of amplified products from transferred foreign DNA using the polymerase chain reaction and primers targeted at the E6-E7 region for different HPV types. RESULTS: Polymerase chain reaction analyses showed transference of DNA HPV type 18 to the blastocysts. Not all types of DNA fragments were transferred equally. CONCLUSION: The results suggested the possibility of using sperm as a noninvasive gene delivery system for passing on gene fragments to preimplantation embryos. It was demonstrated that certain DNA fragments were easier to deliver than others, indicating the necessity for exploring all the factors involved in the mechanism of the transference process. The study also serves to highlight the possibility of unintentional transmission of viral or bacterial DNA to the developing embryo via the sperm.

Animals↗

PubMed1994

Human papillomavirus gene sequences in washed human sperm deoxyribonucleic acid.

The present study demonstrated the presence of HPV gene sequences in Percoll-washed sperm cells using polymerase chain reaction primers targeting smaller gene regions. Up to 64% of the sperm specimens were shown to contain gene sequences indicative of the presence of HPV. Human papillomavirus type 16 was detected about twice as often as HPV type 18. The results suggest the possible role of sperm as a vector for HPV.

DNA, Viral↗

PubMed1994

White blood cells in semen affect hyperactivation but not sperm membrane integrity in the head and tail regions.

The presence of high numbers of peroxidase-positive PML in ejaculated semen significantly reduced sperm HA, an important step leading to sperm capacitation. Sperm membranes at both the head and tail regions, as assessed by the hypo-osmotic viability parameter and the hypo-osmotic sperm swelling test, respectively, were not affected by peroxidase-containing leukocytes. Sperm motility was not affected, but sperm curvilinear and straight line velocity parameters were reduced in the presence of high concentrations of leukocytes in the ejaculate. The results suggested that the effect of leukocytes on sperm was through a reduction in sperm hyperactive motility but not through alterations in the sperm head and tail membranes.

Cell Membrane↗

PubMed1994

The relationship between human sperm fertilizing capacity and histocompatibility linked antigen (HLA) alleles gene sequences.

Heterodimer membrane glycoproteins expressed from hypervariable genes located in the histocompatibility linked antigen (HLA) class II genes on chromosome 6 have been shown to induce activation of lymphocytes and are involved in human sperm binding processes. The objective was to identify an association between HLA-DQA1, -DRB1 or -DPB1 genes and sperm kinematic parameters and sperm penetration of oocytes. The results showed reduced sperm hyperactivation and decreased sperm penetration of zona-free oocytes when the HLA-DRB1 allele was present. The reduced hyperactive motility was not attributed to alterations in sperm kinematic parameters. In contrast, the HLA-DPB1 gene only affected sperm count, linearity of movement and sperm head dimensions. HLA-DQA1 had no effect on the sperm parameters. The data suggest a link between HLA-DRB1 and HLA-DPB1 genes and sperm concentration, sperm movement characteristics and fertilizing capacity.

Alleles↗

PubMed1992

Uptake of exogenous human papilloma virus L1 DNA by oocytes and detection by the polymerase chain reaction.

OBJECTIVE: The purpose of this study was to determine if oocytes were capable of taking up exogenous DNA such as human papillomaviral (HPV) DNA and evaluate the zona pellucida as a barrier to the entry of foreign DNA into the oocyte. METHODS: The experiment consisted of four groups of hamster oocytes exposed to HPV DNA fragments: Group A, zona-free oocytes (n = 5); Group B, oocytes with an intact zona pellucida (n = 5); Group C, oocytes fixed in 4% buffered formalin solution for 20 min (n = 5); and Group D, zona-free oocytes (n = 4). Group C oocytes served as an internal control to ensure adequate washing of the oocytes after incubation. RESULTS: The zona pellucida was not a barrier to foreign DNA molecules. The PCR did not detect L1-HPV and beta-globin gene sequences in the untreated hamster oocyte. Uptake of the smaller DNA fragments such as that amplified from the beta-globin region was independent of active oocyte cell processes. CONCLUSION: Oocytes cultured in vitro can passively take up exogenous DNA fragments. The results suggest a possible role of oocytes as vectors for foreign DNA.

Animals↗

PubMed1992

Rapid identification of deoxyribonucleic acid sequence differences in cytochrome P-450 21-hydroxylase (CYP21) genes with denaturing gradient gel blots.

Denaturing gradient gel electrophoresis was used to identify single-base differences in the cytochrome P-450 21-hydroxylase (CYP21) genes of 132 unrelated control individuals and family members of three unrelated patients with 21-hydroxylase deficiency. The salt-wasting variety was caused by gene deletion and gene conversion/deletion mutations in affected members of two families studied. The simple virilizing form, present in the third family, was caused by an apparent point mutation not detectable by routine Southern blots. We have detected many restriction fragment melting polymorphisms in the CYP21 genes of the members of both salt-wasting families and normal individuals with denaturing gradient gel electrophoresis. We also identified a restriction fragment melting polymorphism specific for the simple virilizing patient in the third family. The data demonstrate that the CYP21 genes are highly polymorphic and that denaturing gradient gel electrophoresis is useful for genomic deoxyribonucleic acid analysis of patients with 21-hydroxylase deficiency.

Adrenal Hyperplasia, Congenital↗

PubMed1992

Sperm hyperactivation as quality control for sperm penetration assay.

The sperm penetration assay (SPA) is subject to considerable variation, and controls are needed to verify the accuracy of the results. It is proposed that sperm hyperactivation (HA) can serve as a quality control check for the SPA. The objective was to determine if there was an association between the SPA outcome and sperm HA measured at various times during the SPA procedure. The data showed a significant correlation between percent sperm HA and percent zona-free oocyte penetration by sperm preincubated for three hours prior to sperm-oocyte interaction (short preincubation). Some sperm hyperactivity was observed in liquefied raw semen samples, but this was insignificantly related to SPA results. Low correlation was observed between SPA results and sperm HA determined immediately after centrifuge washing of sperm. The results suggest that it is possible to utilize sperm HA measured immediately after the sperm-oocyte interaction period as a quality control check of SPA results.

Female↗

PubMed1992

Assessment of sperm for cryopreservation using the hypoosmotic viability test.

In summary, the hypoosmotic viability parameter was significantly correlated with the outcome of the thawed sperm motility. The prefreeze supravital staining for sperm viability and the hypoosmotic sperm swelling test were not predictive of the thawed sperm total motility. The hypoosmotic viability parameter was not correlated to the postwarmed sperm motility after refrigeration. The results indicated that the integrity of the sperm membranes at the head were more important than the tail membrane.

Cell Survival↗

PubMed1991

Identification of male-factor semen samples prior to insemination and in vitro fertilization.

Semen analyses carried out as part of the clinical in vitro fertilization or intrauterine insemination protocols provide important information that determine the type of clinical treatment of the male partner and the sperm processing method. It is postulated that the sperm of male-factor patients cannot survive hypoosmotic stress conditions because of defective sperm membrane function. To test this, 0.1 ml of semen from each of 102 patients was placed in 1.0 ml of 150 mosmol/liter eosin citrate fructose solution and incubated for 30 min at 37 degrees C. The percentage viability of the sperm cells was then determined. The results indicated that patients with two or more abnormal semen parameters had a significantly lower percentage viability while in the hypoosmotic solution (40.6 +/- 4.7%), in contrast to non-male-factor patients (69.0 +/- 1.6%). Donor sperm (N = 32) serving as controls (73.3 +/- 2.1%) had a viability in hypoosmotic solution similar to that of non-male-factor patients. The data suggest that sperm of male-factor patients are less able to survive the hypoosmotic stress conditions as shown by the percentage viability in hypoosmotic solution and emphasize the importance of using less stressful sperm processing methods for in vitro fertilization or insemination in these patients.

Cell Membrane↗

PubMed1991

Combined supravital staining and hypoosmotic swelling test.

The parameter of sperm viability in hypoosmotic solution (VHOS) provides information concerning the membrane integrity of the sperm head. The objective of this study was to determine the association between the VHOS parameter and the sperm penetration assay. The VHOS parameter correctly predicted 70.0-71.4% of the failed sperm penetration assay samples in the short duration preincubation groups. A combination of both the VHOS parameter and the hypoosomotic sperm swelling (HOS) test significantly reduced the number of false negative results. In general, a sperm sample with an abnormal VHOS result and an abnormal HOS test result would be associated with a negative sperm penetration assay. Washing by centrifugation appeared to weaken the sperm head membranes while the swim-up method selected for sperm with strong head and tail membranes. After the various processing methods unique changes in the integrity of the sperm head and tail membranes for each sperm sample may help to identify the optimal method of preparation for individual patients undergoing the newer assisted reproductive technologies such as sperm microinjection.

Cell Membrane↗

PubMed1988

Temporal and constitutive expression of homeobox-2 gene (Hu-2), human heat shock gene (hsp-70), and oncogenes C-sis and N-myc in early human trophoblast.

Recent studies of the genetic basis of animal development indicate that homeobox genes, protooncogenes, and some heat shock genes may play a role in early embryogenesis. To investigate the possibility that these genes function in early human embryonic development, we monitored the expression of a human homeobox gene (Hu-2), two human protooncogenes (C-sis and N-myc), and a human heat shock gene (hsp-70) in human trophoblasts at 7 to 13 weeks gestational age. All these genes were found to be expressed in the tissues analyzed. The hsp-70 gene was expressed at nearly constant levels throughout the development period surveyed, whereas N-myc, C-sis, and Hu-2 showed a coordinated pattern of regulated expression. These results are consistent with a functional role of these genes in the early course of human development.

Blotting, Northern↗

PubMedDate not supplied

Diadenosine tetraphosphate (Ap4A) and triphosphate (Ap3A) signaling of human sperm motility.

The ubiquitous dinucleotide polyphosphate, diadenosine tetraphosphate (Ap4A), has been shown to be a signal molecule for DNA replication in mammalian cells. In this study, Ap4A and a related compound, diadenosine triphosphate (Ap3A), were tested for possible signaling functions in human spermatozoa. A computerized automated semen analyzer was used to detect changes in spermatozoa motility parameters. Cryopreserved-thawed donor spermatozoa were washed and incubated in 0.1 mM Ap4A, 0.1 mM Ap3A, or control medium. The data indicated that both Ap4A and Ap3A decreased the percentage of motile spermatozoa after 4 or more hours of incubation in vitro. The two dinucleotide polyphosphates caused an increase in the amplitude of lateral spermatozoa head displacement parameter only at the start of incubation. The other spermatozoa kinematic parameters were unaffected. No opposing ying-yang dual actions of Ap4A to Ap3A were seen. From the results, Ap4A and Ap3A were observed to be potential inhibitory signals of spermatozoa motility after prolonged exposure.

Dinucleoside Phosphates↗

PubMedDate not supplied

A double method sperm wash for artificial insemination.

Recently, there have been concerns regarding the presence of reactive oxygen species (ROS) produced during sperm processing for insemination. However, the sperm wash methods that yielded low ROS levels also had low sperm recovery after processing. The objective of this study was to compare sperm recovery after swim-up from pellet, overlay, and 2-layer Percoll wash methods with the recovery after the double method wash. The latter method consisted of a combination of 2 sperm wash methods, namely, the overlay and the Percoll method. Motile sperm were first collected through the overlay method. The leftover semen was then processed through the 2-layer Percoll method to scavenge motile sperm and the resultant pellet combined with the pellet from the overlay method. In this manner, the level of ROS was kept to a minimal, sperm recovery was improved, and a mixture of sperm with different surface properties was produced as a result of using different processing methods. The results indicated an improvement in sperm recovery and in total sperm motility in noncryopreserved sperm after using the double method wash when compared with the other wash methods. The study suggests that the double method wash is a feasible method for processing sperm for insemination.

Cell Separation↗

PubMedDate not supplied

Clinical pregnancy rate after the double method wash and intrauterine insemination.

The protocol for intrauterine insemination (IUI) involves sperm processing using different methods that have produced varying results. These sperm wash methods do not take into consideration the problems of the exact timing of ovulation and the requirements of sperm cells at different stages of capacitation. The objectives of this study were 1) to use the double method wash previously reported to produce a mixture of different populations of sperm cells and determine the pregnancy outcome after IUI and 2) to compare the sperm kinematic parameters after the double method wash with those after the centrifuge (or whole-population) wash method. Patients were divided into either the double method group (n = 119) or the centrifuge method group (n = 76). The Hamilton-Thorn HTM-C automated sperm motility analyzer (Hamilton-Thorn Research, Danvers, MA) was used to analyze sperm motility parameters. Pregnancy outcomes were evaluated after controlled ovarian stimulation and IUI. An almost 2-fold increase was seen in the pregnancy rate with the double method wash compared with the centrifuge method wash. Sperm motility and velocity were also enhanced in the double method wash groups. The results support the usefulness of the double method wash for the preparation of sperm for IUI.

Female↗