Journal of Steroid Biochemistry and Molecular Biology, 1997
Estrogen is implicated in sexual differentiation of the avian gonad. Expression of the estrogen r... more Estrogen is implicated in sexual differentiation of the avian gonad. Expression of the estrogen receptor and aromatase genes was therefore examined at the time of gonadal sex differentiation in chicken embryos, using reverse transcription and the polymerase chain reaction (RT-PCR). Estrogen receptor (cER) transcripts were detected in the gonads of both presumptive sexes at embryonic days 4.5, 5.5 and 6.5, and in female but not male urogenital tissues at day 3.5. Aromatase (cAROM) transcripts were detected in female but not male gonads from day 6.5 of embryogenesis, and in adult gonads of both sexes. Both female and male embryos thus express cER mRNA before morphological differentiation of the gonads, which begins on day 5, whereas cAROM expression begins at or shortly after the onset of differentiation and is female-specific. Examination of other tissues showed that, in 5.5-day-old embryos, cER expression was limited to the gonads; no transcripts were detected in the mesonephric kidney, liver, brain, hindlimb or heart of either sex. In 9.5-day-old female embryos, cER and cAROM transcripts were present in both the left (ovarian) and the right (regressing) gonads. Altogether, these observations imply that the gonads of both sexes develop the capacity to respond to estrogens early in embryogenesis, before morphological differentiation, whereas the capacity to synthesize estrogens is female-specific and occurs later, at the time of differentiation. These observations are consistent with estrogens having a key role in ovarian development.
Journal of Assisted Reproduction and Genetics, 1988
A rapid bioassay procedure is described for quality-control testing water and apparatus used in t... more A rapid bioassay procedure is described for quality-control testing water and apparatus used in the preparation of media for gamete and embryo culture. This bioassay is based on the sensitivity of hamster epididymal spermatozoa to contaminants present in water and/or in the culture apparatus. The bioassay is usually performed using a modified Tyrode's solution as the sperm culture medium, although complex media can be used. The sensitivity of this test is greatly enhanced by the absence of protein in the medium. The bioassay has been used to detect impurities in water prepared by a standard cartridge filtration system and to verify that reverse-osmosis pretreatment of water could eliminate the problem. It has also detected toxic contaminants leached from syringe filters during medium sterilization. The bioassay is simple to perform and can be completed in 1 working day. It may be a useful alternative to the conventional mouse embryo tests that are in widespread use in human in vitro fertilization (IVF) laboratories.
By using a chemically defined (protein-free) culture medium that supports sperm viability but not... more By using a chemically defined (protein-free) culture medium that supports sperm viability but not capacitation or the acrosome reaction, we have determined that hamster spermatozoa can be chemically capacitated in vitro by the divalent cation chelators D-penicilla-mine, L-histidine, and L-cysteine in the absence of bovine serum albumin (BSA). Washed cauda epididymal spermatozoa were preincubated (1–2 × 106 sperm/ml) for 3, 4, or 6 hr at 37°C in 5% CO2 in air. The basic culture medium used for sperm preincubation and for sperm:egg coincubation was a modified Tyrode's solution (protein-free) containing 10 mM sodium lactate, 100 μM sodium pyruvate, and 1.0 mg/ml polyvinylalcohol (TLP-PVA). Sperm viability was maintained in all preincubation and coincubation media with PHE (20 μM D-penicillamine, 100 μM hypotaurine, and 1.0 μM epinephrine). The low control sperm preincubation medium consisted of TLP-PVA. In some cases the high control preincubation medium also contained 3 mg/ml BSA (TALP-PVA). The experimental preincubation medium was TLP-PVA with additional D-penicillamine (125 or 500 μM), or L-histidine (10, 100, or 1,000 μM) or L-cysteine (25, 75, or 125 μM). After preincubation, sperm were coincubated (2 × 104 sperm/ml) with cumulus-free hamster eggs in TALP-PVA ± additional D-penicillamine, L-histidine, or L-cysteine for 1.5 hr, fixed, and evaluated for percent egg penetration as an index of sperm capacitation. The results demonstrate that hamster spermatozoa can be chemically capacitated in vitro with D-penicillamine (500 μM: range of mean penetration values, 53.6%–84.3%), L-histidine (100 μM: range of mean values, 24.8%–56.3%) or L-cysteine (75 μM: 51.3%) in the absence of exogenous protein.
The endpoint for sperm capacitation occurs when spermatozoa become able to penetrate intact zonae... more The endpoint for sperm capacitation occurs when spermatozoa become able to penetrate intact zonae pellucidae of unfertilized homologous eggs. Activation of eggs stimulated by sperm fusion or by gamete aging initiates changes in zonae pellucidae that bar further sperm entry. This zona block mechanism reduces the usefulness of eggs as indicators for sperm capacitation. Egg storage in concentrated salt solutions destroys the zona block mechanism while retaining the biological sperm receptor/activator functions of the zona pellucida [Yanagimachi et al., Fertil Steril 31:562–574, 1979]. We have developed techniques for the quantitative assay of capacitation using multiple sperm penetration into zonae and have evaluated the sperm-response characteristics of hamster eggs stored in a modified salt solution (STOR). Sperm, preincubated in capacitating or noncapacitating treatments (CAP or NOCAP, respectively), were coincubated with unfertilized and fertilized STOR-treated zonae for 4 hr. Then zonae were stripped of externally bound sperm and the sperm heads in the perivitelline space (PVS) were fluorescently labeled with Hoechst dye 33342. Entry of CAP sperm into the PVS of STOR-treated unfertilized eggs was highly correlated with sperm concentration (Rw equals; .859) and was log linear from 1 × 103–2 × 105 CAP sperm/ml. At 2 × 105 CAP sperm/ml, the mean number of PVS sperm was 63.5 and the maximum observed was 158. NOCAP sperm very rarely penetrated unfertilized zonae (2 sperm into 75 eggs). Few CAP sperm entered the PVS of STOR-treated fertilized eggs (two sperm into 54 eggs at 2 × 105 sperm/ml) unless the sperm concentration was raised to high levels. Differences between replicates were due to male but not female (ie, egg) differences. We conclude that (1) STOR-treated unfertilized zonae can be used to accurately quantitate differences between sperm capacitation and/or fertility states and (2) the availability of large numbers of homogeneous “shelf-stable” zonae will make it feasible to perform hamster sperm capacitation bioassays on a large scale.
Psychological disorders are highly prevalent in patients with inflammatory bowel disease (IBD). A... more Psychological disorders are highly prevalent in patients with inflammatory bowel disease (IBD). Anxiety and depression are known to independently affect quality of life and may additionally impair quality of life in IBD over and above the IBD itself. Some researchers have further proposed that anxiety and depression may influence the clinical course of IBD. However, despite the potential for anxiety and depression to play an important role in the clinical picture of IBD, there is little prospective well-controlled research in this area. Probably because of this lack of clear data, researchers dispute the actual role of these psychological disorders in IBD, with a number of conflicting opinions expressed. This article reports on a review of the literature in this field. Herein we discuss the five main areas of controversy regarding IBD and the specific psychological comorbidities of depression and anxiety: 1) the relative rate of cooccurrence of these psychological disorders with IBD; 2) the cooccurrence of these psychological disorders with particular phase of IBD; 3) the cooccurrence of these psychological disorders with the specific type of IBD; 4) the rate of these psychological comorbidities compared both to healthy subjects and to other disease states; and 5) the timing of onset of psychological comorbidity with respect to onset of IBD. Methodological weaknesses of the reviewed studies make it impossible to resolve these controversies. However, the results clearly show that anxiety/depression and IBD frequently interact. Given the long-term illness burden patients with IBD face, further prospective, appropriately controlled studies are needed to adequately answer the question of the precise interplay between anxiety/depression and IBD.(Inflamm Bowel Dis 2007)
Journal of Steroid Biochemistry and Molecular Biology, 1997
Estrogen is implicated in sexual differentiation of the avian gonad. Expression of the estrogen r... more Estrogen is implicated in sexual differentiation of the avian gonad. Expression of the estrogen receptor and aromatase genes was therefore examined at the time of gonadal sex differentiation in chicken embryos, using reverse transcription and the polymerase chain reaction (RT-PCR). Estrogen receptor (cER) transcripts were detected in the gonads of both presumptive sexes at embryonic days 4.5, 5.5 and 6.5, and in female but not male urogenital tissues at day 3.5. Aromatase (cAROM) transcripts were detected in female but not male gonads from day 6.5 of embryogenesis, and in adult gonads of both sexes. Both female and male embryos thus express cER mRNA before morphological differentiation of the gonads, which begins on day 5, whereas cAROM expression begins at or shortly after the onset of differentiation and is female-specific. Examination of other tissues showed that, in 5.5-day-old embryos, cER expression was limited to the gonads; no transcripts were detected in the mesonephric kidney, liver, brain, hindlimb or heart of either sex. In 9.5-day-old female embryos, cER and cAROM transcripts were present in both the left (ovarian) and the right (regressing) gonads. Altogether, these observations imply that the gonads of both sexes develop the capacity to respond to estrogens early in embryogenesis, before morphological differentiation, whereas the capacity to synthesize estrogens is female-specific and occurs later, at the time of differentiation. These observations are consistent with estrogens having a key role in ovarian development.
Journal of Assisted Reproduction and Genetics, 1988
A rapid bioassay procedure is described for quality-control testing water and apparatus used in t... more A rapid bioassay procedure is described for quality-control testing water and apparatus used in the preparation of media for gamete and embryo culture. This bioassay is based on the sensitivity of hamster epididymal spermatozoa to contaminants present in water and/or in the culture apparatus. The bioassay is usually performed using a modified Tyrode's solution as the sperm culture medium, although complex media can be used. The sensitivity of this test is greatly enhanced by the absence of protein in the medium. The bioassay has been used to detect impurities in water prepared by a standard cartridge filtration system and to verify that reverse-osmosis pretreatment of water could eliminate the problem. It has also detected toxic contaminants leached from syringe filters during medium sterilization. The bioassay is simple to perform and can be completed in 1 working day. It may be a useful alternative to the conventional mouse embryo tests that are in widespread use in human in vitro fertilization (IVF) laboratories.
By using a chemically defined (protein-free) culture medium that supports sperm viability but not... more By using a chemically defined (protein-free) culture medium that supports sperm viability but not capacitation or the acrosome reaction, we have determined that hamster spermatozoa can be chemically capacitated in vitro by the divalent cation chelators D-penicilla-mine, L-histidine, and L-cysteine in the absence of bovine serum albumin (BSA). Washed cauda epididymal spermatozoa were preincubated (1–2 × 106 sperm/ml) for 3, 4, or 6 hr at 37°C in 5% CO2 in air. The basic culture medium used for sperm preincubation and for sperm:egg coincubation was a modified Tyrode's solution (protein-free) containing 10 mM sodium lactate, 100 μM sodium pyruvate, and 1.0 mg/ml polyvinylalcohol (TLP-PVA). Sperm viability was maintained in all preincubation and coincubation media with PHE (20 μM D-penicillamine, 100 μM hypotaurine, and 1.0 μM epinephrine). The low control sperm preincubation medium consisted of TLP-PVA. In some cases the high control preincubation medium also contained 3 mg/ml BSA (TALP-PVA). The experimental preincubation medium was TLP-PVA with additional D-penicillamine (125 or 500 μM), or L-histidine (10, 100, or 1,000 μM) or L-cysteine (25, 75, or 125 μM). After preincubation, sperm were coincubated (2 × 104 sperm/ml) with cumulus-free hamster eggs in TALP-PVA ± additional D-penicillamine, L-histidine, or L-cysteine for 1.5 hr, fixed, and evaluated for percent egg penetration as an index of sperm capacitation. The results demonstrate that hamster spermatozoa can be chemically capacitated in vitro with D-penicillamine (500 μM: range of mean penetration values, 53.6%–84.3%), L-histidine (100 μM: range of mean values, 24.8%–56.3%) or L-cysteine (75 μM: 51.3%) in the absence of exogenous protein.
The endpoint for sperm capacitation occurs when spermatozoa become able to penetrate intact zonae... more The endpoint for sperm capacitation occurs when spermatozoa become able to penetrate intact zonae pellucidae of unfertilized homologous eggs. Activation of eggs stimulated by sperm fusion or by gamete aging initiates changes in zonae pellucidae that bar further sperm entry. This zona block mechanism reduces the usefulness of eggs as indicators for sperm capacitation. Egg storage in concentrated salt solutions destroys the zona block mechanism while retaining the biological sperm receptor/activator functions of the zona pellucida [Yanagimachi et al., Fertil Steril 31:562–574, 1979]. We have developed techniques for the quantitative assay of capacitation using multiple sperm penetration into zonae and have evaluated the sperm-response characteristics of hamster eggs stored in a modified salt solution (STOR). Sperm, preincubated in capacitating or noncapacitating treatments (CAP or NOCAP, respectively), were coincubated with unfertilized and fertilized STOR-treated zonae for 4 hr. Then zonae were stripped of externally bound sperm and the sperm heads in the perivitelline space (PVS) were fluorescently labeled with Hoechst dye 33342. Entry of CAP sperm into the PVS of STOR-treated unfertilized eggs was highly correlated with sperm concentration (Rw equals; .859) and was log linear from 1 × 103–2 × 105 CAP sperm/ml. At 2 × 105 CAP sperm/ml, the mean number of PVS sperm was 63.5 and the maximum observed was 158. NOCAP sperm very rarely penetrated unfertilized zonae (2 sperm into 75 eggs). Few CAP sperm entered the PVS of STOR-treated fertilized eggs (two sperm into 54 eggs at 2 × 105 sperm/ml) unless the sperm concentration was raised to high levels. Differences between replicates were due to male but not female (ie, egg) differences. We conclude that (1) STOR-treated unfertilized zonae can be used to accurately quantitate differences between sperm capacitation and/or fertility states and (2) the availability of large numbers of homogeneous “shelf-stable” zonae will make it feasible to perform hamster sperm capacitation bioassays on a large scale.
Psychological disorders are highly prevalent in patients with inflammatory bowel disease (IBD). A... more Psychological disorders are highly prevalent in patients with inflammatory bowel disease (IBD). Anxiety and depression are known to independently affect quality of life and may additionally impair quality of life in IBD over and above the IBD itself. Some researchers have further proposed that anxiety and depression may influence the clinical course of IBD. However, despite the potential for anxiety and depression to play an important role in the clinical picture of IBD, there is little prospective well-controlled research in this area. Probably because of this lack of clear data, researchers dispute the actual role of these psychological disorders in IBD, with a number of conflicting opinions expressed. This article reports on a review of the literature in this field. Herein we discuss the five main areas of controversy regarding IBD and the specific psychological comorbidities of depression and anxiety: 1) the relative rate of cooccurrence of these psychological disorders with IBD; 2) the cooccurrence of these psychological disorders with particular phase of IBD; 3) the cooccurrence of these psychological disorders with the specific type of IBD; 4) the rate of these psychological comorbidities compared both to healthy subjects and to other disease states; and 5) the timing of onset of psychological comorbidity with respect to onset of IBD. Methodological weaknesses of the reviewed studies make it impossible to resolve these controversies. However, the results clearly show that anxiety/depression and IBD frequently interact. Given the long-term illness burden patients with IBD face, further prospective, appropriately controlled studies are needed to adequately answer the question of the precise interplay between anxiety/depression and IBD.(Inflamm Bowel Dis 2007)
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