Hyaluronic Acid Binding Ability Research Articles

The alteration of PLCζ protein expression in unexplained infertile and asthenoteratozoospermic patients: A potential effect on sperm fertilization ability

Failed oocyte activation has been observed in unexplained infertile (UI) and asthenoteratozoospermic (AT) men. The deficiency of phospholipase C-zeta (PLCζ) could be a possible reason for such failures and has not been studied yet. We investigated the expression and localization of PLCζ protein in the sperms of patients with UI and AT conditions. The relationships between PLCζ-related parameters with male age, sperm characteristics, DNA integrity, and cellular maturity were assessed. Semen samples were collected from fertile (n = 40), UI (n = 40), and AT (n = 40) men. Subsequently, semen analysis, DNA fragmentation, hyaluronic acid-binding ability, and PLCζ level along with its distribution were evaluated using computer-assisted sperm analyzer, sperm chromatin structure assay (SCSA), hyaluronic acid-binding assay (HBA), western blot analysis and immunofluorescence microscopy, respectively. Unlike SCSA, the values of HBA, and PLCζ expression were significantly reduced in UI and AT patients compared to fertile men, whereas no significant differences were observed among the experimental groups in terms of PLCζ localization patterns. The regression analysis also showed that HBA is the only variable associated with PLCζ levels. Furthermore, the correlation of male age with PLCζ localization in postacrosomal, equatorial, and acrosomal+postacrosomal+equatorial (A+PA+E) patterns, as well as the relation of normal morphology, with the (A+PA+E) pattern, remained in the regression model. Our findings indicated that reduced PLCζ level along with the increased DNA fragmentation and impaired maturation may be possible etiologies of decreased fertilization in the studied subjects.

Evaluation of normal morphology, DNA fragmentation, and hyaluronic acid binding ability of human spermatozoa after using four different commercial media for density gradient centrifugation.

ObjectiveDensity gradient centrifugation (DGC) is frequently used to isolate high-motility fractions of spermatozoa. We compared the efficacy of four DGC media in terms of the percentage of morphologically normal spermatozoa, DNA fragmentation level, and hyaluronic acid (HA) binding ability.MethodsThirty men with a total motile spermatozoa count >80 million participated. Semen samples were divided into four aliquots, which were processed using PureSperm, PureCeption, Sidney, and SpermGrad media, respectively. The DNA fragmentation level was measured using the Halosperm assay kit and HA binding ability was measured using the HBA assay kit.ResultsThe mean percentage of morphologically normal spermatozoa was significantly enhanced after DGC using all four media (10.3%, 9.9%, 9.8%, and 10.7%, respectively; p<0.05 for each when compared with 6.9% in raw semen). The DNA fragmentation level was significantly reduced after DGC using PureSperm, PureCeption, and SpermGrad media (6.0%, 6.5%, and 4.9%, respectively; p<0.05 for each when compared with 11.2% in raw semen), but not after DGC using Sidney media (8.5%, p>0.05). HA binding ability did not change after DGC using any of the four media.ConclusionThe four media were equally effective for obtaining a sperm fraction with highly motile, morphologically normal sperm. PureSperm, PureCeption, and SpermGrad media were equally effective for acquiring a sperm fraction with less DNA fragmentation.

Efficiency of Hyaluronic Acid Binding Ability to Improve Sperm Selection in Intracytoplasmic Sperm Injection (ICSI)

Background: intra Cytoplasmic Sperm Injection (ICSI) is a reliable method for single sperm selection and injection to a large number of infertile patients. Human oocytes are naturally surrounded by cumulus cells embedded in the intracellular matrix made primarily of hyaluronic acid (HA), which plays a role as physiological selector for Intra-Cytoplasmic Sperm Injection (PICSI). Mature spermatozoa are able to bind to and digest HA for the best chance of reaching the oocyte and to maintain fertilization. The intact human spermatozoa are bound to immobilize HA surrounding oocyte in vitro and this leads to reduced risk of chromosomal imbalance or chromatin diseases. Selection of spermatozoa by HA before ICSI may help to optimize the outcomes of the treatment; thus, our study aimed to compare ICSI results, based on the hyaluronic acid or traditional method for sperm selection. Setting: this study conducted at International Islamic Center for Population Studies and Research, Assisted Reproduction Unit, Al-Azhar University from January to October 2015. Patients and Methods:120 couples with male factor infertility and normal wives conducted at the Assisted Reproduction Unit in the International Islamic Center for Population Studies and Research (IICPSR), Al-Azhar University, Egypt, during the period from January to October 2015. All cases were clinically evaluated and eligible for analysis by using inclusion and exclusion criteria. All studied men subjected to ejaculated sperm prepared by traditional method and another prepared by PICSI method. ICSI performed simultaneously for different oocytes from the same woman using an ejaculated sperm by two methods from the same husband. ICSI outcomes compared to the two sequential attempts performed, respectively. Number and quality of oocytes, fertilization rates, rate of embryo cleavage and pregnancy outcomes were recorded. The collected data were tabulated and statistically analyzed. Results: the present study showed a significant increase in the incidence of fertilization rate, the percentage of embryos with top grade in teratozoospermia and thawed semen groups with PICSI technique compared to the traditional method of ICSI groups. Finally, the percentage of pregnancy rate an increase in all groups manipulated with PICSI technique and this increase was statistically significant. Conclusion: this study shed more light on the physiological sperm selection method (PICSI) which may improve fertilization and pregnancy rates compared to the traditional selected sperm in ICSI. It is concluded that PICSI technique in Assisted Reproductive Technology (ART) is one of the very important technique that improve fertilization in case of low fertilization rate after ICSI.

Is sperm hyaluronic acid binding ability predictive for clinical success of intracytoplasmic sperm injection: PICSI vs. ICSI?

Although intracytoplasmic sperm injection (ICSI) is now a widely-used technique, it is still of interest to improve our knowledge as to which is the best spermatozoon to be selected for ICSI. Infertile men have increased risks of producing aneuploid spermatozoa. Using hyaluronic acid (HA)-binding sperm selection may reduce the genetic risks such as chromosomal aberrations of offspring. In the present study we examined the clinical success of ICSI with HA-selected sperm (‘physiologic’ ICSI, PICSI) compared to conventional ICSI, as well as the necessity to differentiate patients according to the initial HA-binding assay result (HBA score) and whether the sperm concentration or HBA score can provide additional information. We observed a significantly higher fertilization rate (FR) of the PICSI group with >60% HBA, implantation rate (IR) of the PICSI group with ≤60% HBA, and clinical pregnanacy rate (CPR) in every PICSI group compared to the ICSI groups (p < 0.01). We also observed a significantly higher life birth rate (LBR) in the PICSI group with ≤60% HBA compared to ICSI patients with ≤60% HBA (p < 0.001). The pregnancy loss rate (PLR) was significanly lower in PICSI patients compared to the ICSI group (p < 0.0001). The FR, IR, CPR, and LBR of the PICSI group with <50% HBA were significantly higher and the PLR was lower than in the ICSI group with <50% HBA (p < 0.01). A statistically significant correlation was found between the sperm concentration and the HA-binding capacity (r = 0.62, p < 0.001). We found a closer relationship between HBA score and FR (r = 0.53, NS) than between sperm concentration and FR (r = 0.14, NS). HBA could be considered for sperm selection prior to ICSI because of its success and apparant ability to reduce genetic complications. However, this must be extended to a larger study.

Evaluation of sperm selection procedure based on hyaluronic acid binding ability on ICSI outcome

To compare the efficiency of routine sperm selection method with HA-selection procedure for fertilization rate, embryo development, implantation and pregnancy rates as well as evaluating the relationship between HA-binding ability with sperm protamine deficiency and DNA fragmentation. Semen samples were obtained from the 50 couples undergoing ICSI. The percentage of fertilization rate, cleavage and quality of embryos compared between two procedures (routine sperm selection and HA-binding selection). The semen samples were assessed for DNA fragmentation and protamine deficiency by sperm chromatin dispersion (SCD) test and Chromomycin A3 (CMA3) staining, respectively. A significant inverse correlation was observed between percentage of HA binding with protamine deficiency, DNA fragmentation and abnormal sperm morphology (P < 0.05). Furthermore, in current study, oocytes inseminated by HA sperm selection procedure had significantly higher fertilization rate (P < 0.05). While the pregnancy and implantation rates were insignificantly increased. The results suggest that normal sperm have higher chance to bind HA and therefore, HA sperm selection procedure may select sperm with normal protamine content and low DNA fragmentation, but to confirm the effect of HA sperm selection on the ICSI outcome requires further studies.

Murine SPAM1 is secreted by the estrous uterus and oviduct in a form that can bind to sperm during capacitation: acquisition enhances hyaluronic acid-binding ability and cumulus dispersal efficiency

Sperm uptake of epididymal sperm adhesion molecule 1 (SPAM1) in vitro has recently been shown to be a marker of sperm maturation, since acquisition of this surface hyaluronidase increases cumulus dispersal efficiency. Here, we demonstrate that this glycosyl phosphatidylinositol-linked sperm antigen, previously shown to be expressed during estrous in the female reproductive tract, is secreted in the uterine and oviductal fluids (ULF and OF respectively) in a 67 kDa form, which can bind to sperm. We show that it can be acquired by caudal sperm from Spam1 null, Spam1-deficient mutant, and wild-type (WT) mice in vitro during incubation in ULF or OF at 37 degrees C, as detected by immunocytochemistry and flow cytometry. SPAM1 binding after ULF incubation was localized predominantly to the acrosome and the mid-piece of the flagella of Spam1 null sperm in a pattern identical to that of WT sperm. After ULF incubation, WT sperm demonstrated a significantly (P<0.001) enhanced hyaluronic acid-binding ability, and the involvement of SPAM1 in this activity was shown by a significant (P<0.001) decrease in binding when sperm were exposed to SPAM1 antiserum-inhibited ULF. Importantly, when Spam1 null sperm were exposed to ULF with SPAM1 accessible (in the presence of pre-immune serum) or inaccessible (in the presence of SPAM1 antiserum) for uptake, there was a significant difference in cumulus dispersal efficiency. Taken together, these results suggest that in the sperm surface remodeling that occurs prior to and during capacitation, the fertilizing competence of sperm is increased via acquisition of SPAM1, and likely other hyaluronidases, from the female tract.

Hyaluronic acid binding ability of human sperm reflects cellular maturity and fertilizing potential: selection of sperm for intracytoplasmic sperm injection

The current concepts of sperm biochemical markers and the central role of the HspA2 chaperone protein, a measure of sperm cellular maturity and fertilizing potential, are reviewed. Because HspA2 is a component of the synaptonemal complex, low HspA2 levels and increased frequency of chromosomal aneuploidies are related in diminished maturity sperm. We also suggest a relationship between HspA2 expression in elongating spermatids and events of late spermiogenesis, such as cytoplasmic extrusion and plasma membrane remodeling that aid the formation of the zona pellucida binding and hyaluronic acid binding sites. The presence of hyaluronic acid receptor on the plasma membrane of mature sperm, coupled with hyaluronic acid coated glass or plastic surfaces, facilitates testing of sperm function and selection of single mature sperm for intracytoplasmic sperm injection. The frequencies of sperm with chromosomal disomy are reduced approximately fourfold to fivefold in hyaluronic acid selected sperm compared with semen sperm, comparable to the increase in such abnormalities in intracytoplasmic sperm injection offspring. Hyaluronic acid binding also excludes immature sperm with cytoplasmic extrusion, persistent histones, and DNA chain breaks. Hyaluronic acid mediated sperm selection is a novel technique that is comparable to sperm zona pellucida binding. Hyaluronic acid selected sperm will also alleviate the risks related to intracytoplasmic sperm injection fertilization with sperm of diminished maturity that currently cause worldwide concern.

Cutting Edge: An Inducible Sialidase Regulates the Hyaluronic Acid Binding Ability of CD44-Bearing Human Monocytes

Previous studies established that variable degrees and types of glycosylation can account for differences in the ability of CD44 to function as a receptor for hyaluronic acid. We have now used neuraminidase treatment to conclude that sialylation negatively regulates CD44 on the human monocytic cell line THP-1 and peripheral blood monocytes. Both of these cell types displayed increased receptor activity after overnight culture with LPS. Of particular interest, the sialidase inhibitor 2-deoxy-2, 3-dehydro-N-acetylneuraminic acid completely blocked the LPS induced recognition of hyaluronic acid by THP-1 cells. Furthermore, acquisition of this characteristic paralleled induction of one type of sialidase activity. Monocytes may be capable of enzymaticly remodeling cell surface CD44, altering their ability to interact with the extracellular matrix.

Evaluation of hyaluronic acid binding ability of hepatic sinusoidal endothelial cells in rats with liver cirrhosis

BACKGROUND & AIMS: In liver cirrhosis, the binding and degradation of hyaluronic acid in the hepatic sinusoidal endothelial cells are considered to be reduced by development of hepatic sinusoidal capillarization, resulting in high serum hyaluronic acid concentration. The aim of this study is to clarify the cause of high blood hyaluronic acid concentration in liver cirrhosis. METHODS: Liver cirrhosis was induced in rats by thioacetamide administration. In vivo observation of sinusoidal capillarization, in vitro immunolocalization of factor VIII- related antigen and CD44, and [14C]hyaluronic acid binding in cultured sinusoidal endothelial cells were determined. RESULTS: Basement membranes were observed on the basal side of sinusoidal endothelial cells. The fenestrae and fluorescent intensity of anti-CD44 bound to the cells decreased with progression of hepatic fibrosis. Immunofluorescent reactive products of factor VIII-related antigen were more abundant in the cirrhotic rats compared with the controls. Amount of [14C]hyaluronic acid binding was significantly decreased in the cirrhotic group compared with the controls. CONCLUSIONS: One reason that the blood hyaluronic acid concentration increases markedly in liver cirrhosis is considered to be the reduction in hyaluronic acid receptors of hepatic sinusoidal endothelial cells and in the amount of hyaluronic acid binding to the cells. (Gastroenterology 1996 Oct;111(4):1049-57)

Involvement of CD44 variant isoforms in hyaluronate adhesion by human activated T cells

The standard, 85-95-kDa form of the hyaluronic acid (HA) receptor CD44 and a number of CD44 mRNA splice variants play important roles in immune responses and tumor metastasis. Variants carrying exon 6 (v6), or 9 (v9) products are transiently expressed on activated human T cells. Here, modulation experiments with specific monoclonal antibodies (mAb) indicate that v6 and v9 are expressed independently on distinct sets of CD44 molecules, and that their combined expression is necessary for HA adhesion. Moreover, the finding that mAb-mediated cross-linking of v6 and v9 promoted cytosolic free Ca2+ mobilization and co-stimulated CD3-triggered T cell proliferation indicates that v6 and v9 possess signaling and effector function activation ability. Finally, HA-mediated signaling appears to be required for variant-dependent adhesion to HA. The observation that soluble HA promoted cytosolic free Ca2+ mobilization indicates that HA-induced Ca2+ mobilization can occur during T cell-HA interaction. Since Ca2+ mobilization was inhibited by pretreatment of cells with an anti-CD44 mAb directed against the HA-binding domain of CD44, CD44 receptors appear to be involved in HA-mediated signal transduction. The requirement of cytosolic free Ca2+ for adhesion is shown by the fact that ionomycin (a Ca2+ ionophore) stimulated, and EGTA (a Ca2+ chelator), inhibited HA adhesion. In addition, cytoskeletal functional activation is required for cell adhesion to HA, since drugs that block actin polymerization, such as cytochalasin B, or actomyosin contraction, such as the calmodulin antagonist W-7, inhibited cell adhesion to HA. As this adhesion is also ADP ribosylation-sensitive, it may involve a GTP-dependent function of CD44v, i.e. ankyrin binding. Our data indicate that there is a functional hierarchy among the CD44 molecules expressed on human peripheral blood T cells and that the splice variants, as compared to the standard form, exhibit a greater HA binding ability which involves CD44-mediated signaling and effector function activation.

Hyaluronic acid binding ability of hepatic sinusoidal endothelial cells in liver cirrhosis: [formula omitted]. Second Department of Medicine, Kurame University School of Medicine, Kurume, Japan

Hyaluronic acid binding ability of hepatic sinusoidal endothelial cells in liver cirrhosis: [formula omitted]. Second Department of Medicine, Kurame University School of Medicine, Kurume, Japan