Aging Increases Susceptibility to Ovarian Cancer Metastasis in Murine Allograft Models and Alters Immune Composition of Peritoneal Adipose Tissue.

Ovarian cancer, the most deadly gynecological malignancy in U.S. women, metastasizes uniquely, spreading through the peritoneal cavity and often generating widespread metastatic sites before diagnosis. The vast majority of ovarian cancer cases occur in women over 40 and the median age at diagnosis is 63. Additionally, elderly women receive poorer prognoses when diagnosed with ovarian cancer. Despite age being a significant risk factor for the development of this cancer, there are little published data which address the impact of aging on ovarian cancer metastasis. Here we report that the aged host is more susceptible to metastatic success using two murine syngeneic allograft models of ovarian cancer metastasis. This age-related increase in metastatic tumor burden corresponds with an increase in tumor infiltrating lymphocytes (TILs) in tumor-bearing mice and alteration of B cell-related pathways in gonadal adipose tissue. Based on this work, further studies elucidating the status of B cell TILs in mouse models of metastasis and human tumors in the context of aging are warranted.

Methods for the visualization and analysis of extracellular matrix protein structure and degradation.

This chapter highlights methods for visualization and analysis of extracellular matrix (ECM) proteins, with particular emphasis on collagen type I, the most abundant protein in mammals. Protocols described range from advanced imaging of complex in vivo matrices to simple biochemical analysis of individual ECM proteins. The first section of this chapter describes common methods to image ECM components and includes protocols for second harmonic generation, scanning electron microscopy, and several histological methods of ECM localization and degradation analysis, including immunohistochemistry, Trichrome staining, and in situ zymography. The second section of this chapter details both a common transwell invasion assay and a novel live imaging method to investigate cellular behavior with respect to collagen and other ECM proteins of interest. The final section consists of common electrophoresis-based biochemical methods that are used in analysis of ECM proteins. Use of the methods described herein will enable researchers to gain a greater understanding of the role of ECM structure and degradation in development and matrix-related diseases such as cancer and connective tissue disorders.

Quantitative proteomic analysis of murine white adipose tissue for peritoneal cancer metastasis.

Cancer metastasis risk increases in older individuals, but the mechanisms for this risk increase are unclear. Many peritoneal cancers, including ovarian cancer, preferentially metastasize to peritoneal fat depots. However, there is a dearth of studies exploring aged peritoneal adipose tissue in the context of cancer. Because adipose tissue produces signals which influence several diseases including cancer, proteomics of adipose tissue in aged and young mice may provide insight into metastatic mechanisms. We analyzed mesenteric, omental, and uterine adipose tissue groups from the peritoneal cavities of young and aged C57BL/6J mouse cohorts with a low-fraction SDS-PAGE gelLC-MS/MS method. We identified 2308 protein groups and quantified 2167 groups, among which several protein groups showed twofold or greater abundance differences between the aged and young cohorts. Cancer-related gene products previously identified as significant in another age-related study were found altered in this study. Several gene products known to suppress proliferation and cellular invasion were found downregulated in the aged cohort, including R-Ras, Arid1a, and heat shock protein ß1. In addition, multiple protein groups were identified within single cohorts, including the proteins Cd11a, Stat3, and Ptk2b. These data suggest that adipose tissue is a strong candidate for analysis to identify possible contributors to cancer metastasis in older subjects. The results of this study, the first of its kind using uterine adipose tissue, contribute to the understanding of the role of adipose tissue in age-related alteration of oncogenic pathways, which may help elucidate the mechanisms of increased metastatic tumor burden in the aged. Graphical abstract We analyzed mesenteric, omental, and uterine adipose tissue groups from the peritoneal cavities of young and aged C57BL/6J mouse cohorts with a low-fraction SDS-PAGE gelLC-MS/MS method. These fat depots are preferential sites for many peritoneal cancers. The results of this study, the first of its kind using uterine adipose tissue, contribute to the understanding of the role of adipose tissue in age-related alteration of oncogenic pathways, which may help elucidate the mechanisms of increased metastatic tumor burden in the aged.

Multiparity activates interferon pathways in peritoneal adipose tissue and decreases susceptibility to ovarian cancer metastasis in a murine allograft model.

Ovarian cancer is the fifth leading cause of cancer deaths in U.S. women and the deadliest gynecologic malignancy. This lethality is largely due to the fact that most cases are diagnosed at metastatic stages of the disease when the prognosis is poor. Epidemiologic studies consistently demonstrate that parous women have a reduced risk of developing ovarian cancer, with a greater number of births affording greater protection; however little is known about the impact of parity on ovarian cancer metastasis. Here we report that multiparous mice are less susceptible to ovarian cancer metastasis in an age-matched syngeneic murine allograft model. Interferon pathways were found to be upregulated in healthy adipose tissue of multiparous mice, suggesting a possible mechanism for the multiparous-related protective effect against metastasis. This protective effect was found to be lost with age. Based on this work, future studies exploring therapeutic strategies which harness the multiparity-associated protective effect demonstrated here are warranted.

Obesity Contributes to Ovarian Cancer Metastatic Success through Increased Lipogenesis, Enhanced Vascularity, and Decreased Infiltration of M1 Macrophages.

Epithelial ovarian cancer (EOC) is the leading cause of death from gynecologic malignancy, with high mortality attributable to widespread intraperitoneal metastases. Recent meta-analyses report an association between obesity, ovarian cancer incidence, and ovarian cancer survival, but the effect of obesity on metastasis has not been evaluated. The objective of this study was to use an integrative approach combining in vitro, ex vivo, and in vivo studies to test the hypothesis that obesity contributes to ovarian cancer metastatic success. Initial in vitro studies using three-dimensional mesomimetic cultures showed enhanced cell-cell adhesion to the lipid-loaded mesothelium. Furthermore, in an ex vivo colonization assay, ovarian cancer cells exhibited increased adhesion to mesothelial explants excised from mice modeling diet-induced obesity (DIO), in which they were fed a "Western" diet. Examination of mesothelial ultrastructure revealed a substantial increase in the density of microvilli in DIO mice. Moreover, enhanced intraperitoneal tumor burden was observed in overweight or obese animals in three distinct in vivo models. Further histologic analyses suggested that alterations in lipid regulatory factors, enhanced vascularity, and decreased M1/M2 macrophage ratios may account for the enhanced tumorigenicity. Together, these findings show that obesity potently affects ovarian cancer metastatic success, which likely contributes to the negative correlation between obesity and ovarian cancer survival.