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miR-451 deficiency is associated with altered endometrial fibrinogen alpha chain expression and reduced endometriotic implant establishment in an experimental mouse model.

Nothnick WB, Graham A, Holbert J, Weiss MJ - PLoS ONE (2014)

Bottom Line: MicroRNAs, small non-coding RNAs that post-transcriptionally regulate gene expression, are mis-expressed in endometriosis but a functional role in the disease pathogenesis remains uncertain.After induction of the disease, we evaluated the impact of this deficiency on implant development and survival.Loss of miR-451 expression resulted in a lower number of ectopic lesions established in vivo.

View Article: PubMed Central - PubMed

Affiliation: University of Kansas Medical Center, Department of Molecular & Integrative Physiology, Kansas City, Kansas, United States of America.

ABSTRACT
Endometriosis is defined as the growth of endometrial glandular and stromal components in ectopic locations and affects as many as 10% of all women of reproductive age. Despite its high prevalence, the pathogenesis of endometriosis remains poorly understood. MicroRNAs, small non-coding RNAs that post-transcriptionally regulate gene expression, are mis-expressed in endometriosis but a functional role in the disease pathogenesis remains uncertain. To examine the role of microRNA-451 (miR-451) in the initial development of endometriosis, we utilized a novel mouse model in which eutopic endometrial fragments used to induce endometriosis were deficient for miR-451. After induction of the disease, we evaluated the impact of this deficiency on implant development and survival. Loss of miR-451 expression resulted in a lower number of ectopic lesions established in vivo. Analysis of differential protein profiles between miR-451 deficient and wild-type endometrial fragments revealed that fibrinogen alpha polypeptide isoform 2 precursor was approximately 2-fold higher in the miR-451 donor endometrial tissue and this elevated expression of the protein was associated with altered expression of the parent fibrinogen alpha chain mRNA and protein. As this polypeptide contains RGD amino acid "cell adhesion" motifs which could impact early establishment of lesion development, we examined and confirmed using a cyclic RGD peptide antagonist, that endometrial cell adhesion and endometriosis establishment could be respectively inhibited both in vitro and in vivo. Collectively, these results suggest that the reduced miR-451 eutopic endometrial expression does not enhance initial establishment of these fragments when displaced into the peritoneal cavity, that loss of eutopic endometrial miR-451 expression is associated with altered expression of fibrinogen alpha chain mRNA and protein, and that RGD cyclic peptide antagonists inhibit establishment of endometriosis development in an experimental mouse model suggesting that this approach may prove useful in the prevention of endometriosis establishment and survival.

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Development of experimental endometriosis is associated with the level of implant miR-451 expression.Experimental endometriosis was induced and development assessed as described under “Materials and Methods”. A. The number of implants of each genotype that developed in wild-type host mice (N = 8/genotype). Data are presented as the mean ± SEM and were analyzed by one-way ANOVA followed by post-hoc analysis. Different letters indicate statistical significance (P<0.05). B. Elevated miR-451 expression is associated with wild-type “implant” tissue that develops ectopically. Uterine fragments were obtained from wild-type mice at the time (0 h) of PMSG administration or 44 h later (the time of endometrial fragment harvest). miR-451 and miR-144 expression was determined by qRT-PCR. Different letters indicate statistical significance (P<0.05) between groups (by unpaired t-test). ND indicates that miR-144 levels were not detectable by qRT-PCR.
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pone-0100336-g002: Development of experimental endometriosis is associated with the level of implant miR-451 expression.Experimental endometriosis was induced and development assessed as described under “Materials and Methods”. A. The number of implants of each genotype that developed in wild-type host mice (N = 8/genotype). Data are presented as the mean ± SEM and were analyzed by one-way ANOVA followed by post-hoc analysis. Different letters indicate statistical significance (P<0.05). B. Elevated miR-451 expression is associated with wild-type “implant” tissue that develops ectopically. Uterine fragments were obtained from wild-type mice at the time (0 h) of PMSG administration or 44 h later (the time of endometrial fragment harvest). miR-451 and miR-144 expression was determined by qRT-PCR. Different letters indicate statistical significance (P<0.05) between groups (by unpaired t-test). ND indicates that miR-144 levels were not detectable by qRT-PCR.

Mentions: Based upon the initial demonstration that miR-451 expression was reduced in eutopic endometrium from women with endometriosis, coupled with the postulated factors which miR-451 may regulate and their role in endometriosis establishment, we hypothesized that the reduced miR-451 expression by eutopic endometrium may enhance the ability of this displaced endometrial tissue to establish ectopically and develop into endometriosis. To test this hypothesis, we transferred endometrial tissue fragments from miR-451+/+, miR-451+/− and miR-451−/− donor mice into 2 to 4 month-old wild-type (miR-451+/+) females and assessed the number of implants which developed 4 weeks after transfer. Surprisingly, transfer of tissue fragments from miR-451−/− mice resulted in significantly less implants developing in wild-type recipients compared to the number of developed implants derived from miR-451+/− and miR-451+/+ donors (Fig. 2A). qRT-PCR was performed on endometrial fragment tissue from wild-type and miR-451−/− mice to confirm induction of miR-451 in wild-type mice as well as assess miR-144 expression. As depicted in Figure 2B, PMSG treatment induced a significant increase in miR-451 expression in the wild-type mice. miR-451 expression was not detected in the mice as expected (data not shown). miR-144 was not detected in endometrial fragments from wild-type mice regardless of treatment (Fig. 2B), while miR-144 was not detected in the miR-451/144−/− mice as expected (data not shown).


miR-451 deficiency is associated with altered endometrial fibrinogen alpha chain expression and reduced endometriotic implant establishment in an experimental mouse model.

Nothnick WB, Graham A, Holbert J, Weiss MJ - PLoS ONE (2014)

Development of experimental endometriosis is associated with the level of implant miR-451 expression.Experimental endometriosis was induced and development assessed as described under “Materials and Methods”. A. The number of implants of each genotype that developed in wild-type host mice (N = 8/genotype). Data are presented as the mean ± SEM and were analyzed by one-way ANOVA followed by post-hoc analysis. Different letters indicate statistical significance (P<0.05). B. Elevated miR-451 expression is associated with wild-type “implant” tissue that develops ectopically. Uterine fragments were obtained from wild-type mice at the time (0 h) of PMSG administration or 44 h later (the time of endometrial fragment harvest). miR-451 and miR-144 expression was determined by qRT-PCR. Different letters indicate statistical significance (P<0.05) between groups (by unpaired t-test). ND indicates that miR-144 levels were not detectable by qRT-PCR.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4061076&req=5

pone-0100336-g002: Development of experimental endometriosis is associated with the level of implant miR-451 expression.Experimental endometriosis was induced and development assessed as described under “Materials and Methods”. A. The number of implants of each genotype that developed in wild-type host mice (N = 8/genotype). Data are presented as the mean ± SEM and were analyzed by one-way ANOVA followed by post-hoc analysis. Different letters indicate statistical significance (P<0.05). B. Elevated miR-451 expression is associated with wild-type “implant” tissue that develops ectopically. Uterine fragments were obtained from wild-type mice at the time (0 h) of PMSG administration or 44 h later (the time of endometrial fragment harvest). miR-451 and miR-144 expression was determined by qRT-PCR. Different letters indicate statistical significance (P<0.05) between groups (by unpaired t-test). ND indicates that miR-144 levels were not detectable by qRT-PCR.
Mentions: Based upon the initial demonstration that miR-451 expression was reduced in eutopic endometrium from women with endometriosis, coupled with the postulated factors which miR-451 may regulate and their role in endometriosis establishment, we hypothesized that the reduced miR-451 expression by eutopic endometrium may enhance the ability of this displaced endometrial tissue to establish ectopically and develop into endometriosis. To test this hypothesis, we transferred endometrial tissue fragments from miR-451+/+, miR-451+/− and miR-451−/− donor mice into 2 to 4 month-old wild-type (miR-451+/+) females and assessed the number of implants which developed 4 weeks after transfer. Surprisingly, transfer of tissue fragments from miR-451−/− mice resulted in significantly less implants developing in wild-type recipients compared to the number of developed implants derived from miR-451+/− and miR-451+/+ donors (Fig. 2A). qRT-PCR was performed on endometrial fragment tissue from wild-type and miR-451−/− mice to confirm induction of miR-451 in wild-type mice as well as assess miR-144 expression. As depicted in Figure 2B, PMSG treatment induced a significant increase in miR-451 expression in the wild-type mice. miR-451 expression was not detected in the mice as expected (data not shown). miR-144 was not detected in endometrial fragments from wild-type mice regardless of treatment (Fig. 2B), while miR-144 was not detected in the miR-451/144−/− mice as expected (data not shown).

Bottom Line: MicroRNAs, small non-coding RNAs that post-transcriptionally regulate gene expression, are mis-expressed in endometriosis but a functional role in the disease pathogenesis remains uncertain.After induction of the disease, we evaluated the impact of this deficiency on implant development and survival.Loss of miR-451 expression resulted in a lower number of ectopic lesions established in vivo.

View Article: PubMed Central - PubMed

Affiliation: University of Kansas Medical Center, Department of Molecular & Integrative Physiology, Kansas City, Kansas, United States of America.

ABSTRACT
Endometriosis is defined as the growth of endometrial glandular and stromal components in ectopic locations and affects as many as 10% of all women of reproductive age. Despite its high prevalence, the pathogenesis of endometriosis remains poorly understood. MicroRNAs, small non-coding RNAs that post-transcriptionally regulate gene expression, are mis-expressed in endometriosis but a functional role in the disease pathogenesis remains uncertain. To examine the role of microRNA-451 (miR-451) in the initial development of endometriosis, we utilized a novel mouse model in which eutopic endometrial fragments used to induce endometriosis were deficient for miR-451. After induction of the disease, we evaluated the impact of this deficiency on implant development and survival. Loss of miR-451 expression resulted in a lower number of ectopic lesions established in vivo. Analysis of differential protein profiles between miR-451 deficient and wild-type endometrial fragments revealed that fibrinogen alpha polypeptide isoform 2 precursor was approximately 2-fold higher in the miR-451 donor endometrial tissue and this elevated expression of the protein was associated with altered expression of the parent fibrinogen alpha chain mRNA and protein. As this polypeptide contains RGD amino acid "cell adhesion" motifs which could impact early establishment of lesion development, we examined and confirmed using a cyclic RGD peptide antagonist, that endometrial cell adhesion and endometriosis establishment could be respectively inhibited both in vitro and in vivo. Collectively, these results suggest that the reduced miR-451 eutopic endometrial expression does not enhance initial establishment of these fragments when displaced into the peritoneal cavity, that loss of eutopic endometrial miR-451 expression is associated with altered expression of fibrinogen alpha chain mRNA and protein, and that RGD cyclic peptide antagonists inhibit establishment of endometriosis development in an experimental mouse model suggesting that this approach may prove useful in the prevention of endometriosis establishment and survival.

Show MeSH
Related in: MedlinePlus