- Email: [email protected]
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RESULTS: We utilized Function Express to compare expression between diabetic and control CEOs. A total of 14 genes were significantly altered in the diabetic CEOs compared to control including 11 that were downregulated with a false discovery rate of 10%: SPARC related modular calcium binding 2, UDP glucose dehydrogenase, Cathespin Z, male sterility domain containing 2, 18 day embryo whole body gene, Kruppel-like factor 3, interleukin 1 receptor type 1, eukaryotic translation factor 5, leucine rich repeat containing 8 family, lectin, galactose binding gene soluble 1, and A kinase(PRKA) anchor protein 2. Three genes were downregulated with a false discovery rate of 0% including lysosomal membrane glycoprotein 2, integrin beta 1, and adult male stomach gene. CONCLUSION: Interestingly, one gene that is directly involved with glucose metabolism, UDP glucose dehydrogenase, was down regulated. Glycogen levels in diabetic oocytes have been found to be elevated compared to control (unpublished data). If UDP glucose dehydrogenase activity is slowed, UDP glucose substrate levels rise, and thus more glycogen may be produced via glycogen synthase. UDP glucose activity will be measured in diabetic and non-diabetic oocytes for confirmation. Other genes that were affected were involved with varied cellular processes including cytokine release, trafficking, and cytoskeletal structure. These broad changes may display the varied effects of maternal hyperglycemia on the developing, preovulatory oocyte. These findings represent changes in gene expression at the 6 hour timepoint following hCG administration. The microarray analysis will also be repeated at the 18 hour timepoint, after completion of metaphase I. These findings will be confirmed by realtime PCR and changes in protein expression will also be examined using immunofluorescent microscopy on individual CEOs and Western immunoblot analysis on pooled CEOs. Supported by: This work was supported in part by the Berlex Foundation Award in Basic Science.
differed in OMD in all groups. In group A, the diabetic mice had significantly fewer oocytes with minimal OMD and significantly more oocytes with moderately increased OMD (P ⫽ 0.001). Whereas in group B, diabetic mice had significantly fewer oocytes with minimal and moderate OMD but significantly more oocytes with markedly increased OMD (P ⬍ 0.0001). Diabetic mice also displayed significantly fewer oocytes with intact CG (P ⫽0.028) and significantly more oocytes with minimal and marked CG loss respectively (P ⬍ 0.0001).
Wednesday, October 25, 2006 3:45 pm O-201 OOCYTE AGING IN DIABETES MELLITUS. P. T. Goud, A. P. Goud, M. P. Diamond, B. Gonik, H. M. Abu-Soud. Wayne State Univ, Detroit, MI. OBJECTIVE: The impact of diabetes on reproduction is profound, as seen by diminution in fertility and increase in reproductive losses. Elevated glucose levels before conception and during pregnancy are associated with spontaneous miscarriages, congenital malformations and abnormalities in fetal growth and development. However, the mechanisms that contribute to these outcomes are poorly understood. Diabetes influences the process of oocyte maturation and preimplantation embryo development. Developmental abnormalities in DM therefore, could be related to oocyte dysfunction and aging that may occur more often in DM. Current study investigates the oocyte aging phenomena in diabetic and non-diabetic mice. DESIGN: Oocyte aging phenomena were studied using confocal microscopy and compared between oocytes from diabetic and non-diabetic oocytes retrieved at different post-ovulatory ages. MATERIALS AND METHODS: Young and relatively old oocytes were retrieved from superovulated ALS/LtJ diabetic mice and age/weight matched B6D2F1 non-diabetic control mice at 13.5, 16 and 18 h post-hCG (groups A, B and C, n⫽83). Oocyte aging phenomena of zona pellucida dissolution time (ZPDT), ooplasmic microtubule dynamics (OMD) after taxol stimulation and premature cortical granule loss (CGL) were studied with confocal microscopy and other techniques described before by an observer blinded to the treatment groups (1). The data on ZPDT were analyzed using the Student’s unpaired t test, two-way ANOVA and the Student-Neuman-Keuls post-hoc test. Frequency data within each test and control groups were analyzed using the Chi square test. Frequencies of OMD and CGL were compared with respective controls using the Fisher’s exact test (SPSS version 11.0, SPSS Inc.®, Chicago, IL). RESULTS: The ZP dissolution time in oocytes from ALS/LtJ diabetic mice was similar to those from the non-diabetic B6D2F1 mice in group A. However, the ZPDT increased significantly with postovulatory aging in oocytes from both, diabetic and non-diabetic mice. Thus, oocytes from diabetic mice exhibited significantly increased ZPDT in groups B and C (P ⬍ 0.0001). Moreover, increase in ZPDT with postovulatory age was significantly higher in diabetic versus non-diabetic mice (F ⫽ 23.7, df ⫽ 2, 77, P ⬍ 0.0001). The oocytes from diabetic and non-diabetic mice however,
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CONCLUSION: Oocytes from DM mice exhibit enhanced post-ovulatory aging, indicating a possible narrowing of the temporal window for optimal fertilization in uncontrolled DM [(1) Goud AP et al., Biochemistry 2005: 44: 11361-8]. Supported by: NIH RO1 HL066367 (to H. M. A-S) and the Dept OB/Gyn, Wayne State University, Detroit, MI, USA.
Wednesday, October 25, 2006 4:00 pm O-202 AN IN VITRO MODEL FOR PCOS RELATED INSULIN RESISTANCE: THE EFFECTS OF TESTOSTERONE ON PHOSPHORYLATION OF INTRACELLULAR INSULIN SIGNALING PROTEINS IN RAT SKELETAL MUSCLE PRIMARY CULTURE. M. C. Allemand, Y. Asmann, K. Klaus, L. Tatpati, C. C. Coddington, K. S. Nair. Mayo Clinic Rochester, Rochester, MN. OBJECTIVE: Polycystic ovarian syndrome (PCOS) is a heterogeneous disorder in which hyperandrogenism and insulin resistance are prominent features. The mechanism of how hyperandrogenism results in insulin resistance (IR) has not been elucidated. Our aim was to study the mechanism of IR in hyperandrogenism, and to test whether testosterone exposure results in aberrant phosphorylation of insulin signaling pathway proteins. DESIGN: Exposure to insulin in rat primary skeletal muscle culture after overnight incubation in testosterone in comparison with no testosterone incubation.
Vol. 86, Suppl 2, September 2006
differed in OMD in all groups. In group A, the diabetic mice had significantly fewer oocytes with minimal OMD and significantly more oocytes with moderately increased OMD (P ⫽ 0.001). Whereas in group B, diabetic mice had significantly fewer oocytes with minimal and moderate OMD but significantly more oocytes with markedly increased OMD (P ⬍ 0.0001). Diabetic mice also displayed significantly fewer oocytes with intact CG (P ⫽0.028) and significantly more oocytes with minimal and marked CG loss respectively (P ⬍ 0.0001).
Wednesday, October 25, 2006 3:45 pm O-201 OOCYTE AGING IN DIABETES MELLITUS. P. T. Goud, A. P. Goud, M. P. Diamond, B. Gonik, H. M. Abu-Soud. Wayne State Univ, Detroit, MI. OBJECTIVE: The impact of diabetes on reproduction is profound, as seen by diminution in fertility and increase in reproductive losses. Elevated glucose levels before conception and during pregnancy are associated with spontaneous miscarriages, congenital malformations and abnormalities in fetal growth and development. However, the mechanisms that contribute to these outcomes are poorly understood. Diabetes influences the process of oocyte maturation and preimplantation embryo development. Developmental abnormalities in DM therefore, could be related to oocyte dysfunction and aging that may occur more often in DM. Current study investigates the oocyte aging phenomena in diabetic and non-diabetic mice. DESIGN: Oocyte aging phenomena were studied using confocal microscopy and compared between oocytes from diabetic and non-diabetic oocytes retrieved at different post-ovulatory ages. MATERIALS AND METHODS: Young and relatively old oocytes were retrieved from superovulated ALS/LtJ diabetic mice and age/weight matched B6D2F1 non-diabetic control mice at 13.5, 16 and 18 h post-hCG (groups A, B and C, n⫽83). Oocyte aging phenomena of zona pellucida dissolution time (ZPDT), ooplasmic microtubule dynamics (OMD) after taxol stimulation and premature cortical granule loss (CGL) were studied with confocal microscopy and other techniques described before by an observer blinded to the treatment groups (1). The data on ZPDT were analyzed using the Student’s unpaired t test, two-way ANOVA and the Student-Neuman-Keuls post-hoc test. Frequency data within each test and control groups were analyzed using the Chi square test. Frequencies of OMD and CGL were compared with respective controls using the Fisher’s exact test (SPSS version 11.0, SPSS Inc.®, Chicago, IL). RESULTS: The ZP dissolution time in oocytes from ALS/LtJ diabetic mice was similar to those from the non-diabetic B6D2F1 mice in group A. However, the ZPDT increased significantly with postovulatory aging in oocytes from both, diabetic and non-diabetic mice. Thus, oocytes from diabetic mice exhibited significantly increased ZPDT in groups B and C (P ⬍ 0.0001). Moreover, increase in ZPDT with postovulatory age was significantly higher in diabetic versus non-diabetic mice (F ⫽ 23.7, df ⫽ 2, 77, P ⬍ 0.0001). The oocytes from diabetic and non-diabetic mice however,
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Abstracts
CONCLUSION: Oocytes from DM mice exhibit enhanced post-ovulatory aging, indicating a possible narrowing of the temporal window for optimal fertilization in uncontrolled DM [(1) Goud AP et al., Biochemistry 2005: 44: 11361-8]. Supported by: NIH RO1 HL066367 (to H. M. A-S) and the Dept OB/Gyn, Wayne State University, Detroit, MI, USA.
Wednesday, October 25, 2006 4:00 pm O-202 AN IN VITRO MODEL FOR PCOS RELATED INSULIN RESISTANCE: THE EFFECTS OF TESTOSTERONE ON PHOSPHORYLATION OF INTRACELLULAR INSULIN SIGNALING PROTEINS IN RAT SKELETAL MUSCLE PRIMARY CULTURE. M. C. Allemand, Y. Asmann, K. Klaus, L. Tatpati, C. C. Coddington, K. S. Nair. Mayo Clinic Rochester, Rochester, MN. OBJECTIVE: Polycystic ovarian syndrome (PCOS) is a heterogeneous disorder in which hyperandrogenism and insulin resistance are prominent features. The mechanism of how hyperandrogenism results in insulin resistance (IR) has not been elucidated. Our aim was to study the mechanism of IR in hyperandrogenism, and to test whether testosterone exposure results in aberrant phosphorylation of insulin signaling pathway proteins. DESIGN: Exposure to insulin in rat primary skeletal muscle culture after overnight incubation in testosterone in comparison with no testosterone incubation.
Vol. 86, Suppl 2, September 2006