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Loss of Cdh1 and Trp53 in the uterus induces chronic inflammation with modification of tumor microenvironment.

Stodden GR, Lindberg ME, King ML, Paquet M, MacLean JA, Mann JL, DeMayo FJ, Lydon JP, Hayashi K - Oncogene (2014)

Bottom Line: Mice with conditional ablation of Cdh1 and Trp53 (Cdh1(d/d)Trp53(d/d)) clearly demonstrate architectural features characteristic of type II ECs, including focal areas of papillary differentiation, protruding cytoplasm into the lumen (hobnailing) and severe nuclear atypia at 6 months of age.Our microarray analysis found that most of the genes differentially regulated in the uteri of Cdh1(d/d)Trp53(d/d) mice were involved in inflammatory responses.CD163 and Arg1, markers for tumor-associated macrophages, were also detected and increased in the uteri of Cdh1(d/d)Trp53(d/d) mice, suggesting that an inflammatory tumor microenvironment with immune cell recruitment is augmenting tumor development in Cdh1(d/d)Trp53(d/d) uteri.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Southern Illinois University School of Medicine, Carbondale, IL, USA.

ABSTRACT
Type II endometrial carcinomas (ECs) are estrogen independent, poorly differentiated tumors that behave in an aggressive manner. As TP53 mutation and CDH1 inactivation occur in 80% of human endometrial type II carcinomas, we hypothesized that mouse uteri lacking both Trp53 and Cdh1 would exhibit a phenotype indicative of neoplastic transformation. Mice with conditional ablation of Cdh1 and Trp53 (Cdh1(d/d)Trp53(d/d)) clearly demonstrate architectural features characteristic of type II ECs, including focal areas of papillary differentiation, protruding cytoplasm into the lumen (hobnailing) and severe nuclear atypia at 6 months of age. Further, Cdh1(d/d)Trp53(d/d) tumors in 12-month-old mice were highly aggressive, and metastasized to nearby and distant organs within the peritoneal cavity, such as abdominal lymph nodes, mesentery and peri-intestinal adipose tissues, demonstrating that tumorigenesis in this model proceeds through the universally recognized morphological intermediates associated with type II endometrial neoplasia. We also observed abundant cell proliferation and complex angiogenesis in the uteri of Cdh1(d/d)Trp53(d/d) mice. Our microarray analysis found that most of the genes differentially regulated in the uteri of Cdh1(d/d)Trp53(d/d) mice were involved in inflammatory responses. CD163 and Arg1, markers for tumor-associated macrophages, were also detected and increased in the uteri of Cdh1(d/d)Trp53(d/d) mice, suggesting that an inflammatory tumor microenvironment with immune cell recruitment is augmenting tumor development in Cdh1(d/d)Trp53(d/d) uteri. Further, inflammatory mediators secreted from CDH1-negative, TP53 mutant endometrial cancer cells induced normal macrophages to express inflammatory-related genes through activation of nuclear factor-κB signaling. These results indicate that absence of CDH1 and TP53 in endometrial cells initiates chronic inflammation, promotes tumor microenvironment development following the recruitment of macrophages and promotes aggressive ECs.

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The effect of Cdh1 and Trp53 ablation on proliferation, angiogenesis and steroid hormone receptors. (a) Immunoreactive Ki67 (MKI67) as a marker of cell proliferation, CD31 (PECAM1) as an endothelial cell marker, and steroid hormone receptors (ESR1 and PGR) were detected in the uteri of control and Cdh1d/dTrp53d/d mice at 6 and 12 months of age. (b) Semiquantitatively scored Ki67 by ImmunoRatio, and CD31 by positively stained cell numbers. Cell specific ESR1 and PGR positive cells were counted and the percentage (positive cells/total cells) is shown. ImmunoRatio calculates the percentage of positively stained nuclear cells/total cells.
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Figure 3: The effect of Cdh1 and Trp53 ablation on proliferation, angiogenesis and steroid hormone receptors. (a) Immunoreactive Ki67 (MKI67) as a marker of cell proliferation, CD31 (PECAM1) as an endothelial cell marker, and steroid hormone receptors (ESR1 and PGR) were detected in the uteri of control and Cdh1d/dTrp53d/d mice at 6 and 12 months of age. (b) Semiquantitatively scored Ki67 by ImmunoRatio, and CD31 by positively stained cell numbers. Cell specific ESR1 and PGR positive cells were counted and the percentage (positive cells/total cells) is shown. ImmunoRatio calculates the percentage of positively stained nuclear cells/total cells.

Mentions: Next, to determine whether an alteration in cell proliferation is occurring in Cdh1d/dTrp53d/d uteri, we performed immunohistochemical staining for Ki67. Nuclear staining of Ki67 in 12-mo uterine sections was analyzed by ImmunoRatio.38 In agreement with the lack of changes in uterine gross morphology and histology, only a few cells were Ki67 positive in the uteri from control, Cdh1d/d, Trp53d/d, Cdh1d/d Trp53d/+, Cdh1d/+Trp53d/d mice, and a significant number of Ki67 positive cells were observed in only the Cdh1d/dTrp53d/d uteri (Supplementary Figure 2d). Therefore, we focused further analyses to control and Cdh1d/dTrp53d/d mice (Figure 3). The uteri of Cdh1d/dTrp53d/d mice showed significantly increased Ki67 positive cells compared to control uteri, starting at 2-mo (data not shown). Immunostaining of CD31, a marker of endothelial cells, indicated that angiogenesis is more active in Cdh1d/dTrp53d/d uteri than control. Nuclear staining of ESR1 and PGR in different cell types was quantified, and the percentage of positive cells from total cells was shown. Epithelial ESR1 and PGR were decreased following ablation of both Trp53 and Cdh1, although stromal PGR in Cdh1d/dTrp53d/d uteri were increased, suggesting that loss of Trp53 and Cdh1 leads to abnormal proliferation, angiogenesis and loss of or abnormal steroid hormone activity.


Loss of Cdh1 and Trp53 in the uterus induces chronic inflammation with modification of tumor microenvironment.

Stodden GR, Lindberg ME, King ML, Paquet M, MacLean JA, Mann JL, DeMayo FJ, Lydon JP, Hayashi K - Oncogene (2014)

The effect of Cdh1 and Trp53 ablation on proliferation, angiogenesis and steroid hormone receptors. (a) Immunoreactive Ki67 (MKI67) as a marker of cell proliferation, CD31 (PECAM1) as an endothelial cell marker, and steroid hormone receptors (ESR1 and PGR) were detected in the uteri of control and Cdh1d/dTrp53d/d mice at 6 and 12 months of age. (b) Semiquantitatively scored Ki67 by ImmunoRatio, and CD31 by positively stained cell numbers. Cell specific ESR1 and PGR positive cells were counted and the percentage (positive cells/total cells) is shown. ImmunoRatio calculates the percentage of positively stained nuclear cells/total cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4551401&req=5

Figure 3: The effect of Cdh1 and Trp53 ablation on proliferation, angiogenesis and steroid hormone receptors. (a) Immunoreactive Ki67 (MKI67) as a marker of cell proliferation, CD31 (PECAM1) as an endothelial cell marker, and steroid hormone receptors (ESR1 and PGR) were detected in the uteri of control and Cdh1d/dTrp53d/d mice at 6 and 12 months of age. (b) Semiquantitatively scored Ki67 by ImmunoRatio, and CD31 by positively stained cell numbers. Cell specific ESR1 and PGR positive cells were counted and the percentage (positive cells/total cells) is shown. ImmunoRatio calculates the percentage of positively stained nuclear cells/total cells.
Mentions: Next, to determine whether an alteration in cell proliferation is occurring in Cdh1d/dTrp53d/d uteri, we performed immunohistochemical staining for Ki67. Nuclear staining of Ki67 in 12-mo uterine sections was analyzed by ImmunoRatio.38 In agreement with the lack of changes in uterine gross morphology and histology, only a few cells were Ki67 positive in the uteri from control, Cdh1d/d, Trp53d/d, Cdh1d/d Trp53d/+, Cdh1d/+Trp53d/d mice, and a significant number of Ki67 positive cells were observed in only the Cdh1d/dTrp53d/d uteri (Supplementary Figure 2d). Therefore, we focused further analyses to control and Cdh1d/dTrp53d/d mice (Figure 3). The uteri of Cdh1d/dTrp53d/d mice showed significantly increased Ki67 positive cells compared to control uteri, starting at 2-mo (data not shown). Immunostaining of CD31, a marker of endothelial cells, indicated that angiogenesis is more active in Cdh1d/dTrp53d/d uteri than control. Nuclear staining of ESR1 and PGR in different cell types was quantified, and the percentage of positive cells from total cells was shown. Epithelial ESR1 and PGR were decreased following ablation of both Trp53 and Cdh1, although stromal PGR in Cdh1d/dTrp53d/d uteri were increased, suggesting that loss of Trp53 and Cdh1 leads to abnormal proliferation, angiogenesis and loss of or abnormal steroid hormone activity.

Bottom Line: Mice with conditional ablation of Cdh1 and Trp53 (Cdh1(d/d)Trp53(d/d)) clearly demonstrate architectural features characteristic of type II ECs, including focal areas of papillary differentiation, protruding cytoplasm into the lumen (hobnailing) and severe nuclear atypia at 6 months of age.Our microarray analysis found that most of the genes differentially regulated in the uteri of Cdh1(d/d)Trp53(d/d) mice were involved in inflammatory responses.CD163 and Arg1, markers for tumor-associated macrophages, were also detected and increased in the uteri of Cdh1(d/d)Trp53(d/d) mice, suggesting that an inflammatory tumor microenvironment with immune cell recruitment is augmenting tumor development in Cdh1(d/d)Trp53(d/d) uteri.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Southern Illinois University School of Medicine, Carbondale, IL, USA.

ABSTRACT
Type II endometrial carcinomas (ECs) are estrogen independent, poorly differentiated tumors that behave in an aggressive manner. As TP53 mutation and CDH1 inactivation occur in 80% of human endometrial type II carcinomas, we hypothesized that mouse uteri lacking both Trp53 and Cdh1 would exhibit a phenotype indicative of neoplastic transformation. Mice with conditional ablation of Cdh1 and Trp53 (Cdh1(d/d)Trp53(d/d)) clearly demonstrate architectural features characteristic of type II ECs, including focal areas of papillary differentiation, protruding cytoplasm into the lumen (hobnailing) and severe nuclear atypia at 6 months of age. Further, Cdh1(d/d)Trp53(d/d) tumors in 12-month-old mice were highly aggressive, and metastasized to nearby and distant organs within the peritoneal cavity, such as abdominal lymph nodes, mesentery and peri-intestinal adipose tissues, demonstrating that tumorigenesis in this model proceeds through the universally recognized morphological intermediates associated with type II endometrial neoplasia. We also observed abundant cell proliferation and complex angiogenesis in the uteri of Cdh1(d/d)Trp53(d/d) mice. Our microarray analysis found that most of the genes differentially regulated in the uteri of Cdh1(d/d)Trp53(d/d) mice were involved in inflammatory responses. CD163 and Arg1, markers for tumor-associated macrophages, were also detected and increased in the uteri of Cdh1(d/d)Trp53(d/d) mice, suggesting that an inflammatory tumor microenvironment with immune cell recruitment is augmenting tumor development in Cdh1(d/d)Trp53(d/d) uteri. Further, inflammatory mediators secreted from CDH1-negative, TP53 mutant endometrial cancer cells induced normal macrophages to express inflammatory-related genes through activation of nuclear factor-κB signaling. These results indicate that absence of CDH1 and TP53 in endometrial cells initiates chronic inflammation, promotes tumor microenvironment development following the recruitment of macrophages and promotes aggressive ECs.

Show MeSH
Related in: MedlinePlus