Effects of streptozotocin- and alloxan-induced diabetes mellitus on mouse follicular and early embryo development

Mice were made diabetic by intraperitoneal injection of streptozotocin or alloxan. Germinal vesicle breakdown in the ovarian follicles at 8 h after hCG in control animals (57%) was significantly greater than in streptozotocin-(24%) and alloxan-(42%) diabetic animals (P less than 0.001). This delay in oocyte maturation was reversible by in-vivo insulin administration to diabetic mice. A developmental delay was also found for embryos recovered from diabetic mice. This developmental delay extended into the 72 h in-vitro cultures. Compared to control embryos, those from alloxan- and streptozotocin-treated mice demonstrated marked impairment in development as assessed by (1) distribution of developmental cell stages at each observation period and (2) rates of development which increasingly diverged at each observation period. In diabetic mice treated with insulin in vivo, the percentage of 2-cell embryos recovered increased. Furthermore, in streptozotocin- and alloxan-animals treated with insulin, the rate of in-vitro development of embryos, as well as their developmental stage distribution improved. We therefore suggest that uncontrolled diabetes mellitus, as well as contributing to the development of congenital malformations, may deleteriously affect reproductive performance both before fertilization and at the very earliest gestational stages.     

Analysis of post-implantation mouse embryos after maternal heat stress during meiotic maturation

The effects of heat stress during oocyte maturation were studied in post-implantation mouse embryos. Virgin ICR mice were exposed to 35 +/- 1 degree C and 65 +/- 3% RH for 12.5 h beginning immediately after synchronization of ovulation with PMSG and hCG. Embryos of heat-stressed dams were developmentally heterogeneous and showed significant delays in development with as much as 48 h delayed development. Nearly 6% of these embryos were triploid, and another 2% were hyper-diploid. Development of triploid embryos was delayed more than 24 h. Nine embryos with severe developmental delay had heterogeneous chromosome constitutions. Embryo mortality before and after implantation was higher in heat-stressed dams than in controls.     

Maternal diabetes and oocyte quality

Maternal diabetes has been demonstrated to adversely affect preimplantation embryo development and pregnancy outcomes. Emerging evidence has implicated that these effects are associated with compromised oocyte competence. Several developmental defects during oocyte maturation in diabetic mice have been reported over past decades. Most recently, we further identified the structural, spatial and metabolic dysfunction of mitochondria in oocytes from diabetic mice, suggesting the impaired oocyte quality. These defects in the oocyte may be maternally transmitted to the embryo and then manifested later as developmental abnormalities in preimplantation embryo, congenital malformations, and even metabolic disease in the offspring. In this paper, we briefly review the effects of maternal diabetes on oocyte quality, with a particular emphasis on the mitochondrial dysfunction. The possible connection between dysfunctional oocyte mitochondria and reproductive failure of diabetic females, and the mechanism(s) by which maternal diabetes exerts its effects on the oocyte are also discussed.     

The endocannabinoid system modulates the ovarian physiology and its activation can improve in vitro oocyte maturation

The cannabinoid (CB) system has been involved in many aspects of reproduction and it is known that the systemic chronic use of exogenous CBs are deleterious to reproductive processes. Even so, it is not known what happens in relation to the physiology of the ovary when CB receptors are absent. The present study investigated the effect of the lack of CB1 and CB2 receptors in mice ovarian morphology, folliculogenesis, oocyte retrieval, and oocyte maturation and evaluated the use of Δ9-tetrahydrocannabinol (THC) on oocyte in vitro maturation (IVM) by comparing classical IVM and two-step IVM by analyzing the meiotic competence of the oocytes and their evolution toward embryos. Thus, when CB1 and CB2 receptors were missed, the ovary area and volume was significantly less and the action of the equine chorionic gonadotropin (eCG) hormone was diminished. In addition, the mutant genotypes had fewer ovarian follicles and they were less competent after eCG administration compared with wild-type mice, and this lack of CB receptors showed a mismatch of oocyte maturation. However, the in vitro use of THC showed improvements in oocytes IVM after a Pre-IVM step for 48 hr, as those oocytes reached a significantly higher polar body rate, a larger diameter and the best result on blastocysts rate was achieved when THC was used during the IVM step.     

Altered meiotic regulation in oocytes from diabetic mice

In the present study, we have utilized a streptozotocin-induced diabetic mouse model to examine how the diabetic condition and different glucose concentrations affect several parameters of reproductive physiology. We report that oocyte maturation is altered under all experimental conditions examined. In cumulus cell-enclosed oocytes (CEO) from diabetic mice, spontaneous maturation was accelerated but the FSH-mediated delay of spontaneous maturation was suppressed. Higher glucose levels in the culture medium suppressed spontaneous maturation but did not influence the transient arrest mediated by FSH. Meiotic arrest in CEO by hypoxanthine and dibutyryl cAMP (dbcAMP) was less effective at higher glucose concentrations. In addition, both FSH-induced maturation in vitro and hCG-induced maturation in vivo were reduced by the diabetic condition. The ovulation rate was lowered by about 50% in diabetic mice and fewer ovulated ova had reached metaphase II. Despite the decreased number of ova at metaphase II, in vitro cultures showed the oocytes were capable of completing meiotic maturation at control levels. Insulin treatment reversed the detrimental effects of diabetes on meiotic induction, ovulation, and completion of meiotic maturation. Cultures of pronuclear-staged embryos confirmed a negative effect of diabetes and hyperglycemia on development to the blastocyst stage. These data suggest that defects in meiotic regulation brought about by the diabetic condition are due to decreased communication between the somatic and germ cell compartments, and it is concluded that such conditions may contribute to postfertilization developmental abnormalities.     

Effect of diabetes mellitus on mouse pre-implantation embryo development.

Fifteen spontaneously diabetic, non-obese mice (NOD strain), 17 non-diabetic NOD mice (in which diabetes had not yet developed) and 9 diabetic NOD mice were treated with insulin. All animals were superovulated with 5 iu of pregnant mares' serum gonadotrophin followed 48 h later by 5 iu human chorionic gonadotrophin (hCG) and mated overnight with NOD males of proven fertility. To assess in-vitro and early in-vivo development, 23 NOD mice were killed 72 h after hCG treatment. Embryos were recovered from oviduct flushings and cultured in Ham's F-10 medium with 0.1% bovine serum albumin at 37 degrees C in an atmosphere of 5% O2, 5% CO2, and 90% N2. Development was assessed at intervals of 24 h for 72 h. Compared with embryos from non-diabetic NOD mice (n = 81), embryos from diabetic NOD mice (n = 68) demonstrated marked impairment in growth assessed by distribution of developmental stages at each observation period (24, 48, 72 h, all P less than 0.001) and by overall rates of progression of developmental stages (P less than 0.01). In diabetic NOD mice treated with insulin, embryo development (n = 7) was not significantly different from that of embryos from non-diabetic NOD mice (n = 81), but was significantly faster than in embryos from diabetic NOD mice not treated with insulin (n = 68) (P less than 0.001, for all periods, overall rate P less than 0.01). To assess late in-vivo growth, 18 NOD mice were killed 120 h after hCG.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pre- and post-implantation losses of embryos in aging female IVCS mice

Mated female mice of the IVCS strain, aged 90 (control group), 180, 210, 240, 270, 300, 330, 360 and 420 days old, were studied for pre- and post-implantation loss of embryos. The percentage of pre-implantation loss in mice aged 90 to 210 days was 1.7/11.8 (14.4%) to 2.7/11.7 (23.1%). In mice aged 240 to 300 days it increased significantly as compared to the controls (46.5-90.2% versus 14.4%). It reached 100% in 300-days-old mice. The post-implantation loss of embryos and/or fetuses in mice aged 90 to 240 days was 1.0/10.2 (9.8%) to 2.5/9.0 (27.8%). In mice aged 270 to 300 days it increased significantly compared to the controls (100% versus 9.8%). The decrease in reproductive activity appeared first in a decrease in litter size, followed by a decrease in the number of blastocyst and implantation sites, and finally by anovulation during the process of aging in IVCS mice.     

ADAMTS proteases in fertility

The reproductive organs are unique among adult organs in that they must undergo continual tissue remodelling as a key aspect of their normal function. The processes for persistent maturation and release of new gametes, as well as fertilisation, implantation, placentation, gestation and parturition involve cyclic development and regression of tissues that must continually regenerate to support fertility. The ADAMTS family of proteases has been shown to contribute to many aspects of the tissue morphogenesis required for development and function of each of the reproductive organs. Dysregulation or functional changes in ADAMTS family proteases have been associated with reproductive disorders such as polycystic ovarian syndrome (PCOS) and premature ovarian failure (POF). Likewise, proteolytic substrates of ADAMTS enzymes have also been linked to reproductive function. New insight into the roles of ADAMTS proteases has yielded a deeper understanding of the molecular mechanisms behind fertility with clinical potential to generate therapeutic targets to resolve infertility, develop biomarkers that predict dysfunction of the reproductive organs and potentially offer targets for development of non-hormonal male and female contraceptives.     

Nutritional effects on oocyte and embryo development in mammals: implications for reproductive efficiency and environmental sustainability

The environment in which a breeding female lives prior to conception and during the early stages of her pregnancy has striking effects on oocytes developing in the ovarian follicle and on early embryos in the reproductive tract. Of the various environmental factors known to affect oocyte and embryo development, altered nutrition during this critical period has been particularly well studied. Alterations in the quantity of food consumed or the composition of the diet imposed solely during the pre-mating period affect oocyte maturity, blastocyst yield, prenatal survival and the number of offspring born alive. Importantly, nutrition at this time also affects the quality of embryos and resultant offspring, with increasing evidence from a variety of species showing that peri-conception nutrition can alter behaviour, cardiovascular function and reproductive function throughout post-natal life. In livestock species, it is important to devise nutritional strategies that improve reproductive efficiency and the quality of offspring but that do not add to the environmental footprint of the production system and which recognize likely changes in feedstuff availability arising from predicted changes in climate.     

Roles of insulin-like growth factor Ⅱ in regulating female reproductive physiology

The number of growth factors involved in female fertility has been extensively studied, but reluctance to add essential growth factors in culture media has limited progress in optimizing embryonic growth and implantation outcomes, a situation that has ultimately led to reduced pregnancy outcomes. Insulin-like growth factor Ⅱ(IGF-Ⅱ) is the most intricately regulated of all known reproduction-related growth factors characterized to date, and is perhaps the predominant growth factor in human ovarian follicles. This review aims to concisely summarize what is known about the role of IGF-Ⅱ in follicular development, oocyte maturation, embryonic development, implantation success, placentation, fetal growth, and in reducing placental cell apoptosis, as well as present strategies that use growth factors in culture systems to improve the developmental potential of oocytes and embryos in different species. Synthesizing the present knowledge about the physiological roles of IGF-Ⅱ in follicular development, oocyte maturation, and early embryonic development should, on the one hand, deepen our overall understanding of the potential beneficial effects of growth factors in female reproduction and on the other hand support development(optimization) of improved outcomes for assisted reproductive technologies.     

Pancreas-infiltrating Th1 cells and diabetes develop in IL-12-deficient nonobese diabetic mice.

IL-12 and IL-12 antagonist administration to nonobese diabetic (NOD) mice accelerates and prevents insulin-dependent diabetes mellitus (IDDM), respectively. To further define the role of endogenous IL-12 in the development of diabetogenic Th1 cells, IL-12-deficient NOD mice were generated and analyzed. Th1 responses to exogenous Ags were reduced by approximately 80% in draining lymph nodes of these mice, and addition of IL-12, but not IL-18, restored Th1 development in vitro, indicating a nonredundant role of IL-12. Moreover, spontaneous Th1 responses to a self Ag, the tyrosine phosphatase-like IA-2, were undetectable in lymphoid organs from IL-12-deficient, in contrast to wild-type, NOD mice. Nevertheless, wild-type and IL-12-deficient NOD mice developed similar insulitis and IDDM. Both in wild-type and IL-12-deficient NOD mice, approximately 20% of pancreas-infiltrating CD4+ T cells produced IFN-gamma, whereas very few produced IL-10 or IL-4, indicating that IDDM was associated with a type 1 T cell infiltrate in the target organ. T cell recruitment in the pancreas seemed favored in IL-12-deficient NOD mice, as revealed by increased P-selectin ligand expression on pancreas-infiltrating T cells, and this could, at least in part, compensate for the defective Th1 cell pool recruitable from peripheral lymphoid organs. Residual Th1 cells could also accumulate in the pancreas of IL-12-deficient NOD mice because Th2 cells were not induced, in contrast to wild-type NOD mice treated with an IL-12 antagonist. Thus, a regulatory pathway seems necessary to counteract the pathogenic Th1 cells that develop in the absence of IL-12 in a spontaneous chronic progressive autoimmune disease under polygenic control, such as IDDM.     

Long-Lasting Expression of HO-1 Delays Progression of Type I Diabetes in NOD Mice

Heme oxygenase-1 (HO-1) is crucial in regulating oxidative injury. The present study was designed to assess whether HO-1 upregulation by cobalt protoporphyrin IX (CoPP) moderates or prevents the diabetic state in non-obese diabetic (NOD) mice, an animal model for Type 1 diabetes (T1D). HO-1 expression and HO activity were upregulated in the pancreas by the intermittent administration of CoPP. This was associated with decreases in blood glucose and pancreatic O2-, but increased pAKT and BcL-XL and cell survival. A considerable number of beta cells were preserved in the islets of CoPP-treated NOD mice, while none were found in untreated diabetic mice. The number of CD11c+ dendritic cells was decreased in the pancreas of CoPP-treated NOD mice (p     

Temporal dynamics of oocyte development, plasma sex steroids and somatic energy reserves during seasonal ovarian maturation in captive Murray cod Maccullochella peelii peelii.

The temporal dynamics of oocyte growth, plasma sex steroids and somatic energy stores were examined during a 12 month ovarian maturation cycle in captive Murray cod Maccullochella peelii peelii under simulated natural photothermal conditions. Ovarian function was found to be relatively uninhibited in captivity, with the exception that post-vitellogenic follicles failed to undergo final maturation, resulting in widespread pre-ovulatory atresia. Seasonal patterns of oocyte growth were characterised by cortical alveoli accumulation in March, deposition of lipids in April, and vitellogenesis between May and September. Two distinct batches of vitellogenic oocytes were found in Murray cod ovaries, indicating a capacity for multiple spawns. Plasma profiles of 17beta-oestradiol and testosterone were both highly variable during the maturation period suggesting that multiple roles exist for these steroids during different stages of oocyte growth. Condition factor, liver size and visceral fat stores were all found to increase prior to, or during the peak phase of vitellogenic growth. Murray cod appear to strategically utilise episodes of high feeding activity to accrue energy reserves early in the reproductive cycle prior to its deployment during periods of rapid ovarian growth.     

The quest for the sea urchin egg receptor for sperm

The incretin glucagon-like peptide-1 (GLP-1) and other GLP-1 receptor agonists have been shown to cause both antiapoptotic as well as regenerative effects on beta-cells in different animal models for diabetes. Our aim of this study was to test the hypothesis that spontaneously diabetic non obese diabetic (NOD) mice show an altered expression of GLP-1 compared to normoglycemic age-matched controls as a consequence of a diabetic state. To do this we used an ELISA prototype for mouse GLP-1 to measure plasma total GLP-1 from recently diabetic NOD mice as well as from age-matched normoglycemic NOD mice (controls). We also stained sections of pancreatic glands for GLP-1 from diabetic NOD mice and controls. We found increased levels of plasma total GLP-1 in diabetic NOD mice, when compared to control mice, both from non-fasted mice and from mice fasted for 2h. Furthermore, diabetic NOD mice displayed a higher GLP-1 response to an oral glucose tolerance test, compared to control mice. We also found that sections of pancreatic glands from diabetic NOD mice had an increased GLP-1 positive islet area in regard to relative islet area (i.e. total islet area / total pancreas area of the sections) compared to control mice. To our knowledge, this study is the first to show increased levels of GLP-1 in plasma in spontaneously diabetic NOD mice. We suggest that these results might represent a compensatory mechanism of the diabetic NOD mice to counteract beta-cell loss and hyperglycemia.     

Generation of viable embryos and embryonic stem cell-like cells from cultured primary follicles in mice

Primary follicles retrieved from B6CBAF1 prepubertal mice were cultured in a stepwise manner in an alpha-minimum essential medium-based medium to generate viable embryos and embryonic stem cell (ESC)-like cells. A significant increase in follicle growth and oocyte maturation accompanied by increased secretion of 17beta-estradiol and progesterone was achieved by exposing primary follicles to 100 or 200 mIU of follicle-stimulating hormone (FSH) during culture. More oocytes developed into blastocysts following in vitro fertilization (IVF) or parthenogenetic activation after culture with 200 mIU of FSH during the entire culture period than with 100 mIU. Eleven ESC-like cell lines, consisting of four heterozygotic and seven homozygotic phenotypes, were established from 25 trials of primary follicle culture combined with IVF or parthenogenetic activation. In conclusion, primary follicles can potentially yield developmentally competent oocytes, which produce viable embryos and ESC-like cell lines following in vitro manipulation. We suggest a method to utilize immature follicles, which are most abundant in ovaries, to improve reproductive efficiency and for use in regenerative medicine.     

Cryopreservation of oocyte and ovarian tissue

Cryopreservation of reproductive cells (i.e., oocytes, spermatozoa) and tissues (i.e., ovarian and testicular tissue) is a developing technology that has tremendous implications for rapid advancement of biomedical research in general. Since the early 1980s, advances have been made in establishing optimal conditions for in vitro oocyte maturation, fertilization, and culture of resulting embryos. These in vitro systems have contributed significantly to the utilization of these cells and tissues after thawing and have made it possible to evaluate protocols designed to cryopreserve such biomaterials more effectively. Although cryopreservation of preimplantation embryos from various species including mouse, human, and farm animals has been successful, cryopreservation of oocytes from most mammalian species has been more challenging due to their extreme sensitivity to suboptimal conditions during the cryopreservation process. Cryopreservation on mouse oocytes have been well documented and have resulted in greater success than studies with other mammalian species. Ovarian tissue cryopreservation and transplantation techniques have recently received much scientific and public attention due to their great potential use in human infertility treatment, in safeguarding the reproductive potential of the endangered species, and in genome banking of genetically important lab animal strains. A review of past and current research in the field of oocyte and ovarian tissue cryopreservation and transplantation and discussion of possible strategies for oocyte and ovarian tissue banking are provided.     

Chromosomal Aberrations in In-Vitro Matured Oocytes Influence Implantation and Ongoing Pregnancy Rates in a Mouse Model Undergoing Intracytoplasmic Sperm Injection

Implantation failure and early pregnancy loss have been reported to be closely related to the quality of mammalian oocytes; however, the pregnant outcome of embryos from in-vitro matured (IVM) oocytes remains unknown. In this study we examined spindle assembly and chromosome segregation during differentiation, and the duration of IVM of mouse oocytes. The resulting implantation and pregnancy outcomes were analyzed to clarify the relationship between the spindle and chromosomes of IVM oocytes and implantation and early pregnancy. Cumulus-enclosed germinal vesicle oocytes were collected and randomly cultured in IVM medium with different IVM durations. One part of IVM oocytes were analyzed the spindle and chromosome morphology by immunofluorescence method, and the other part of them were fertilized by intracytoplasmic sperm injection. The resulting embryos were transferred into pseudo-pregnant female mice, and the post-implantation and full term development was observed. The chromosome aberrations and incorrect spindle assembly seems not affect the early development and blastocyst cell number derived from IVM oocytes, however the development potential of the resulting embryos after implantation were significant decreased with the ratio increasing of chromosome aberrations and incorrect spindle assembly. Accordingly, the full-term development was also decreased. In conclusion, the present study showed the spindle assembly of in vitro-matured oocytes was one of the most important factors that affected the implantation and ongoing pregnancy rates of IVM oocytes, and the improvement by an appropriate duration of maturation in vitro will enhance the post-implantation development potential of the resulting embryos, and decrease implantation failure and early pregnancy loss.     

Endometrial receptivity defects and impaired implantation in diabetic NOD mice

Implantation failure is a major hurdle to a successful pregnancy. The high rate of postimplantation fetal loss in nonobese diabetic (NOD) mice is believed to be related to an abnormal decidual production of interferon (IFN)gamma. To address whether diabetes alters the natural events associated with successful implantation, certain morphological and molecular features of uterine receptivity in diabetic NOD (dNOD) mice were examined in normally mated pregnancy and in concanavalin A (ConA)-induced pseudopregnancy. As opposed to normoglycemic NOD (cNOD) mice, dNOD mice expressed retarded maturation of their uterine pinopodes and overexpressed MUC1 mucin at implantation sites (P < 0.001). Uterine production of leukemia inhibitory factor (LIF) and phosphorylation of uterine NFkappaBp65 and STAT3-Ty705 were found to be low (P < 0.01) during Day 4.5 postcoitum, whereas IFNgamma was aberrantly overexpressed. Loss of temporal regulation of progesterone receptor A (PR A) and PR B, together with aberrantly increased expression of the protein inhibitor of activated STAT-y (PIASy) (P < 0.01) and reduced recruitment (P < 0.01) of the latter to nuclear progesterone receptor sites were prominent features of decidualization failure occurring at peri-implantation in dNOD mice. In conclusion, the aberrant expression of endometrial IFNgamma in dNOD mice is associated with a nonreceptive endometrial milieu contributing to peri-implantation embryo loss in type 1 diabetes.