Minireview: Functions of the cumulus oophorus during oocyte maturation, ovulation, and fertilization.

Mucification of the cumulus layer around the oocyte is an obligatory process for female fertility. Tumor necrosis factor-induced protein-6 (TNFIP6 or TSG6) has been shown to be specifically expressed during this process. We have generated TNFIP6-deficient mice and tested the ability of their cumulus cells to undergo mucification. Cumulus cell-oocyte complexes fail to expand in TNFIP6-deficient female mice because of the inability of the cumulus cells to assemble their hyaluronan-rich extracellular matrix. The impaired cumulus matrix formation is due to the lack of covalent complexes between hyaluronan and the heavy chains of the inter-alpha-trypsin inhibitor family. As a consequence, TNFIP6-deficient females are sterile. Cultured TNFIP6-deficient cumulus cell-oocyte complexes also fail to expand when stimulated with dibutyryl cyclic AMP or epidermal growth factor. Recombinant TNFIP6 is able to catalyze the covalent transfer of heavy chains to hyaluronan in a cell-free system, restore the expansion of Tnfip6-null cumulus cell-oocyte complexes in vitro, and rescue the fertility in Tnfip6-null females. These results provide clear evidence that TNFIP6 is a key catalyst in the formation of the cumulus extracellular matrix and indispensable for female fertility.

/pdf/impaired-cumulus-mucification-and-female-sterility-in-tumor-5833q1e174.pdf

Impaired cumulus mucification and female sterility in tumor necrosis factor-induced protein-6 deficient mice

Successful ovulation requires that developmentally competent oocytes are released with appropriate timing from the ovarian follicle. Somatic cells of the follicle sense the ovulatory stimulus and guide resumption of meiosis and release of the oocyte, as well as structural remodelling and luteinization of the follicle. Complex intercellular communication co-ordinates critical stages of oocyte maturation and links this process with release from the follicle. To achieve these outcomes, ovulation is controlled through multiple inputs, including endocrine hormones, immune and metabolic signals, as well as intrafollicular paracrine factors from the theca, mural and cumulus granulosa cells and the oocyte itself. This review focuses on the recent advances in understanding of molecular mechanisms that commence after the gonadotrophin surge and culminate with release of the oocyte. These mechanisms include intracellular signalling, gene regulation and remodelling of tissue structure in each of the distinct ovarian compartments. Most critical ovulatory mediators exert effects through the cumulus cell complex that surrounds and connects with the oocyte. The convergence of ovulatory signals through the cumulus complex co-ordinates the key mechanistic processes that mediate and control oocyte maturation and ovulation.

Molecular mechanisms of ovulation: co-ordination through the cumulus complex

Aluminium adjuvants—in retrospect and prospect

Fertilization in mammals requires the successful completion of many steps, starting with the transport of gametes in the reproductive tract and ending with sperm-egg membrane fusion. In this minireview, we focus on three adhesion steps in this multistep process. The first is oocyte "pick-up," in which the degree of adhesion between the extracellular matrix of the cumulus cells and oviductal epithelial cells controls the successful pick-up of the oocyte-cumulus complex and its subsequent transfer into the oviduct. The second part of this review is concerned with the interaction between the sperm and the zona pellucida of the egg. Evidence is discussed that a plasma membrane form of galactosyltransferase on the surface of mouse sperm binds to ZP3 in the zona pellucida and initiates an acrosome reaction. Additional evidence raises the possibility that initial sperm binding to the zona pellucida is independent of ZP3. Last, we address the relationship between sperm adhesion to the egg plasma membrane and membrane fusion, especially the role of ADAM family proteins on the sperm surface and egg integrins.

Cell adhesion and fertilization: steps in oocyte transport, sperm-zona pellucida interactions, and sperm-egg fusion.

To determine the changes in patterns of 17 beta-estradiol and progesterone levels underlying abnormal cycles in bitches immunized with solubilized crude porcine zonae pellucidae (cPZP), to attempt to circumvent these problems by immunizing with a purified zona fraction (pPZP), and to test the effectiveness of different adjuvants, bitches were immunized with cPZP or pPZP 2-6 times with no adjuvant, Freund's adjuvant, alum adjuvant, or the adjuvant CP-20,961. The bitch immunized without adjuvant had a low titer with a normal cycle and fertility. Immunization with cPZP and adjuvant produced moderate to high titers of antizona antibodies and infertility. Bitches with high titers experienced abnormal estrous cycles. Estradiol rose during proestrus, but instead of falling sharply in early estrus as in controls, it remained elevated. Progesterone did not rise. The moderate-titered bitches had normal cycles and steroid patterns. Bitches immunized with pPZP had moderate titers. Cycles were normal after 3 injections, but after 6 injections one bitch had an abnormal cycle. One pPZP-immunized bitch remained fertile but the others were infertile. Alum was the mildest adjuvant, causing no injection site lesions, but the highest titers occurred with Freund's and CP-20,961 adjuvants. All three adjuvants induced titers sufficient to inhibit fertility. Infertility in bitches immunized with PZP may be due to prevention of zona penetration, because their antisera inhibited zona penetration of oocytes by spermatozoa in vitro. However, alterations in ovarian function preventing ovulation and luteinization could be involved in high-titered bitches.

/pdf/fertility-control-in-the-bitch-by-active-immunization-with-4zk6x8xyft.pdf

Fertility control in the bitch by active immunization with porcine zonae pellucidae: use of different adjuvants and patterns of estradiol and progesterone levels in estrous cycles.

The ovarian histopathology of bitches immunized with crude (cPZP) or partially purified (pPZP) porcine zona pellucida proteins was examined in order to determine the cause of abnormal estrous cycles. The majority of immunized bitches had ovarian cytes. Those immunized with cPZP had follicular cysts lined with a thin layer of granulosa cells, while in those immunized with pPZP, the cysts were lined by a basement membrane with a clump of luteinized cells. In two bitches immunized with cPZP, oocytes were present only in primordial follicles. Similar abnormalities were not found in a bitch immunized with human serum albumin or in 12 untreated bitches. Oocytes flushed from the oviducts of mated, immunized bitches were degenerating, which may have been a primary cause of infertility in such bitches. Ovaries studied 2–6 weeks after immunization showed no loss of gap junctional communication between oocytes and granulosa cells, nor was any inflammatory reaction seen. IgG was bound to the zona as revealed by fluoresceinated protein A staining of frozen sections of those ovaries. Abnormal estrous cycles in PZP-immunized bitches appear to result from follicular dysgenesis or cyst formation, but the etiology of these conditions is unresolved.

Ovarian histopathology of bitches immunized with porcine zonae pellucidae

The process whereby hamster oviductal fimbria transport ova to the ampulla was investigated in vitro and in vivo with the intent of determining whether cumulus-free ova were transported and resolving some of the parameters involved in cumulus-isolated from ovaries could not be picked up until 16–17 h before it would be ovulated. This was apparently related to the time at which the cumulus matrix begins to expand. The fimbria were not species-specific, as they readily picked up rat, mouse, and rabbit ovulated cumul, but they were partially tissue-specific as they would only pick up tissues such as the vitreous humor and loose connective tissue, which contain considerable extracellular glycosaminoglycan. Interaction of cumulus and fimbria could be prevented b treating either structure with polycatinic macromolecules such as poly-l-lysine, cationic ferritin, and protamine. Treatment of fimbria with proteolytic enzymes or hyaluronidase did not prevent pickup, but neuraminidase did. The only “artificial cumulus” the fimbria would pick up was chicken egg-white. Cumulus-free ova were not picked up in vitro, but part of them placed into the ovarian bursa were transported to the ampulla in vivo. Coating the ova with chicken egg-white allowed their pickup in vitro, but had little effect on the success of transport of ova transplanted into the ovarian bursa in vivo. The transport of ova from the ovarian bursa to the oviductal ampulla is apparently a rather complex process involving interaction of some component of the cilia surfaces with the cumulus as well as other factors not necessarily related to this interaction.

Parameters influencing ovum pickup by oviductal fimbria in the golden hamster

In a study designed to evaluate the contraceptive potential of anti-egg zona pellucida immunization, bitches were injected with isolated and solubilized zonae pellucidae of either the pig or the dog in saline and Freund's adjuvant or with saline and adjuvant alone (controls). They were boosted monthly, and serum samples were collected before the first injection and 10 days after each injection. The titers of anti-zona pellucida antibodies in each serum sample were measured by treating fresh canine oocytes with the serum, then evaluating antibody binding as indicated by indirect immunofluorescence, precipitation of the zona surface, and penetrability of the zonae by spermatozoa in vitro. The bitches were bred when they came into estrus. All three bitches immunized with porcine zonae developed high titers (1:10,000 or more by indirect immunofluorescence) of antibodies that cross-reacted with canine zonae to cause precipitation of the zona surface both in vivo and in vitro and that completely inhibited penetration of the zonae by spermatozoa in vitro. The two bitches immunized with canine zonae developed only low titers, and their sera had little or no effect on treated zonae. The two control bitches did not develop anti-zona antibody. None of the bitches immunized against porcine zonae became pregnant when bred, but one bitch immunized against canine zonae and one control did become pregnant. The bitches immunized with porcine zonae had somewhat abnormal cycles for unknown reasons. Thus, we could not establish with certainty whether the infertility resulted from specific interference with fertilization, as in vitro, or from alterations in ovarian function, or both.

Infertility in bitches induced by active immunization with porcine zonae pellucidae

Overpopulation of domestic dogs is a serious problem both in the United States and in many other countries. In the City and County of Honolulu alone, 21,614 dogs and cats were euthanized by the Humane Society from July 1983 through June 1984 simply because they were unwanted. That constituted 87.5% of the animals they took in (figures from Hawaiian Humane Society). Obviously, birth control technology is not the entire solution to this problem, because the best technology will not work unless owners of dogs can be motivated to use it. However, if humane organizations will do their job of educating people to control their pets’ fertility, it will be our job to provide a variety of contraceptive strategies to meet the variety of anticipated needs.

Cherrie A. Mahi-Brown

Papers

Fertility control in the bitch by active immunization with porcine zonae pellucidae: use of different adjuvants and patterns of estradiol and progesterone levels in estrous cycles.

Ovarian histopathology of bitches immunized with porcine zonae pellucidae

Parameters influencing ovum pickup by oviductal fimbria in the golden hamster

Infertility in bitches induced by active immunization with porcine zonae pellucidae

Prospects for Control of Fertility in the Female Dog by Active Immunization with Porcine Zona Pellucida Proteins