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Connexin expression and gap junctional coupling in human cumulus cells: contribution to embryo quality.

Wang HX, Tong D, El-Gehani F, Tekpetey FR, Kidder GM - J. Cell. Mol. Med. (2008)

Bottom Line: Multiple connexins (Cx), the subunits of gap junction channels, have been found within ovarian follicles in several species but little is known about the connexins in human follicles.The strength of gap junctional conductance varied between patients and was significantly and positively correlated with Cx43 level, but neither was correlated with patient age.Interestingly, Cx43 level and intercellular conductance were positively correlated with embryo quality as judged by cleavage rate and morphology, and were significantly higher in patients who became pregnant than in those who did not.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, ON, Canada.

ABSTRACT
Gap junctional coupling among cumulus cells is important for oogenesis since its deficiency in mice leads to impaired folliculogenesis. Multiple connexins (Cx), the subunits of gap junction channels, have been found within ovarian follicles in several species but little is known about the connexins in human follicles. The aim of this study was to determine which connexins contribute to gap junctions in human cumulus cells and to explore the possible relationship between connexin expression and pregnancy outcome from in vitro fertilization (IVF). Cumulus cells were obtained from IVF patients undergoing intra-cytoplasmic sperm injection (ICSI). Connexin expression was examined by RT-PCR and confocal microscopy. Cx43 was quantified by immunoblotting and gap junctional coupling was measured by patch-clamp electrophysiology. All but 5 of 20 connexin mRNAs were detected. Of the connexin proteins detected, Cx43 forms numerous gap junction-like plaques but Cx26, Cx30, Cx30.3, Cx32 and Cx40 appeared to be restricted to the cytoplasm. The strength of gap junctional conductance varied between patients and was significantly and positively correlated with Cx43 level, but neither was correlated with patient age. Interestingly, Cx43 level and intercellular conductance were positively correlated with embryo quality as judged by cleavage rate and morphology, and were significantly higher in patients who became pregnant than in those who did not. Thus, despite the presence of multiple connexins, Cx43 is a major contributor to gap junctions in human cumulus cells and its expression level may influence pregnancy outcome after ICSI.

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Example of an RT-PCR survey of human cumulus cells for connexin mRNAs. Primers and amplification conditions were optimized for each of 20 connexin sequences. All PCR products were run on a 1.2% agarose gel. All but Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47 mRNAs were detected.
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fig01: Example of an RT-PCR survey of human cumulus cells for connexin mRNAs. Primers and amplification conditions were optimized for each of 20 connexin sequences. All PCR products were run on a 1.2% agarose gel. All but Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47 mRNAs were detected.

Mentions: RT-PCR was used to survey cumulus cells for the presence of mRNAs encoding 20 connexins (Cx25, Cx26, Cx30, Cx30.2, Cx30.3, Cx31, Cx31.1, Cx31.9, Cx32, Cx36, Cx37, Cx40, Cx40.1, Cx43, Cx45, Cx46, Cx47, Cx50, Cx59 and Cx62). A representative gel illustrating the PCR products is shown in (Fig. 1). All but five connexin mRNAs (Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47) were detected, and this result was consistent for all 11 samples surveyed by RT-PCR. No specific bands were found in negative control samples, and all PCR products were confirmed by sequencing (data not shown).


Connexin expression and gap junctional coupling in human cumulus cells: contribution to embryo quality.

Wang HX, Tong D, El-Gehani F, Tekpetey FR, Kidder GM - J. Cell. Mol. Med. (2008)

Example of an RT-PCR survey of human cumulus cells for connexin mRNAs. Primers and amplification conditions were optimized for each of 20 connexin sequences. All PCR products were run on a 1.2% agarose gel. All but Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47 mRNAs were detected.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3823412&req=5

fig01: Example of an RT-PCR survey of human cumulus cells for connexin mRNAs. Primers and amplification conditions were optimized for each of 20 connexin sequences. All PCR products were run on a 1.2% agarose gel. All but Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47 mRNAs were detected.
Mentions: RT-PCR was used to survey cumulus cells for the presence of mRNAs encoding 20 connexins (Cx25, Cx26, Cx30, Cx30.2, Cx30.3, Cx31, Cx31.1, Cx31.9, Cx32, Cx36, Cx37, Cx40, Cx40.1, Cx43, Cx45, Cx46, Cx47, Cx50, Cx59 and Cx62). A representative gel illustrating the PCR products is shown in (Fig. 1). All but five connexin mRNAs (Cx30.2, Cx31.9, Cx40.1, Cx46 and Cx47) were detected, and this result was consistent for all 11 samples surveyed by RT-PCR. No specific bands were found in negative control samples, and all PCR products were confirmed by sequencing (data not shown).

Bottom Line: Multiple connexins (Cx), the subunits of gap junction channels, have been found within ovarian follicles in several species but little is known about the connexins in human follicles.The strength of gap junctional conductance varied between patients and was significantly and positively correlated with Cx43 level, but neither was correlated with patient age.Interestingly, Cx43 level and intercellular conductance were positively correlated with embryo quality as judged by cleavage rate and morphology, and were significantly higher in patients who became pregnant than in those who did not.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, ON, Canada.

ABSTRACT
Gap junctional coupling among cumulus cells is important for oogenesis since its deficiency in mice leads to impaired folliculogenesis. Multiple connexins (Cx), the subunits of gap junction channels, have been found within ovarian follicles in several species but little is known about the connexins in human follicles. The aim of this study was to determine which connexins contribute to gap junctions in human cumulus cells and to explore the possible relationship between connexin expression and pregnancy outcome from in vitro fertilization (IVF). Cumulus cells were obtained from IVF patients undergoing intra-cytoplasmic sperm injection (ICSI). Connexin expression was examined by RT-PCR and confocal microscopy. Cx43 was quantified by immunoblotting and gap junctional coupling was measured by patch-clamp electrophysiology. All but 5 of 20 connexin mRNAs were detected. Of the connexin proteins detected, Cx43 forms numerous gap junction-like plaques but Cx26, Cx30, Cx30.3, Cx32 and Cx40 appeared to be restricted to the cytoplasm. The strength of gap junctional conductance varied between patients and was significantly and positively correlated with Cx43 level, but neither was correlated with patient age. Interestingly, Cx43 level and intercellular conductance were positively correlated with embryo quality as judged by cleavage rate and morphology, and were significantly higher in patients who became pregnant than in those who did not. Thus, despite the presence of multiple connexins, Cx43 is a major contributor to gap junctions in human cumulus cells and its expression level may influence pregnancy outcome after ICSI.

Show MeSH
Related in: MedlinePlus