Cancer associated fibroblasts express pro-inflammatory factors in human breast and ovarian tumors.

Inflammation has been established in recent years as a hallmark of cancer. Cancer Associated Fibroblasts (CAFs) support tumorigenesis by stimulating angiogenesis, cancer cell proliferation and invasion. We previously demonstrated that CAFs also mediate tumor-enhancing inflammation in a mouse model of skin carcinoma. Breast and ovarian carcinomas are amongst the leading causes of cancer-related mortality in women and cancer-related inflammation is linked with both these tumor types. However, the role of CAFs in mediating inflammation in these malignancies remains obscure. Here we show that CAFs in human breast and ovarian tumors express high levels of the pro-inflammatory factors IL-6, COX-2 and CXCL1, previously identified to be part of a CAF pro-inflammatory gene signature. Moreover, we show that both pro-inflammatory signaling by CAFs and leukocyte infiltration of tumors are enhanced in invasive ductal carcinoma as compared with ductal carcinoma in situ. The pro-inflammatory genes expressed by CAFs are known NF-κB targets and we show that NF-κB is up-regulated in breast and ovarian CAFs. Our data imply that CAFs mediate tumor-promoting inflammation in human breast and ovarian tumors and thus may be an attractive target for stromal-directed therapeutics.

Differences Between Subclinical Ruminators and Reflectors in Narrating Autobiographical Memories: Innovative Moments and Autobiographical Reasoning.

Reasoning may help solving problems and understanding personal experiences. Ruminative reasoning, however, is inconclusive, repetitive, and usually regards negative thoughts. We asked how reasoning as manifested in oral autobiographical narratives might differ when it is ruminative versus when it is adaptive by comparing two constructs from the fields of psychotherapy research and narrative research that are potentially beneficial: innovative moments (IMs) and autobiographical reasoning (AR). IMs captures statements in that elaborate on changes regarding an earlier personal previous problem of the narrator, and AR capture the connecting of past events with other parts of the narrator's life or enduring aspects of the narrator. A total of N = 94 university students had been selected from 492 students to differ maximally on trait rumination and trait adaptive reflection, and were grouped as ruminators (N = 38), reflectors (N = 37), and a group with little ruminative and reflective tendencies ("unconcerned," N = 19). Participants narrated three negative personal experiences (disappointing oneself, harming someone, and being rejected) and two self-related experiences of more mixed valence (turning point and lesson learnt). Reflectors used more IMs and more negative than positive autobiographical arguments (AAs), but not more overall AAs than ruminators. Group differences were not moderated by the valence of memories, and groups did not differ in the positive effect of narrating on mood. Trait depression/anxiety was predicted negatively by IMs and positively by AAs. Thus, IMs are typical for reflectors but not ruminators, whereas the construct of AR appears to capture reasoning processes irrespective of their ruminative versus adaptive uses.

Isolation of normal and cancer-associated fibroblasts from fresh tissues by Fluorescence Activated Cell Sorting (FACS).

Cancer-associated fibroblasts (CAFs) are the most prominent cell type within the tumor stroma of many cancers, in particular breast carcinoma, and their prominent presence is often associated with poor prognosis. CAFs are an activated subpopulation of stromal fibroblasts, many of which express the myofibroblast marker α-SMA. CAFs originate from local tissue fibroblasts as well as from bone marrow-derived cells recruited into the developing tumor and adopt a CAF phenotype under the influence of the tumor microenvironment. CAFs were shown to facilitate tumor initiation, growth and progression through signaling that promotes tumor cell proliferation, angiogenesis, and invasion. We demonstrated that CAFs enhance tumor growth by mediating tumor-promoting inflammation, starting at the earliest pre-neoplastic stages. Despite increasing evidence of the key role CAFs play in facilitating tumor growth, studying CAFs has been an on-going challenge due to the lack of CAF-specific markers and the vast heterogeneity of these cells, with many subtypes co-existing in the tumor microenvironment. Moreover, studying fibroblasts in vitro is hindered by the fact that their gene expression profile is often altered in tissue culture. To address this problem and to allow unbiased gene expression profiling of fibroblasts from fresh mouse and human tissues, we developed a method based on previous protocols for Fluorescence-Activated Cell Sorting (FACS). Our approach relies on utilizing PDGFRα as a surface marker to isolate fibroblasts from fresh mouse and human tissue. PDGFRα is abundantly expressed by both normal fibroblasts and CAFs. This method allows isolation of pure populations of normal fibroblasts and CAFs, including, but not restricted to α-SMA+ activated myofibroblasts. Isolated fibroblasts can then be used for characterization and comparison of the evolution of gene expression that occurs in CAFs during tumorigenesis. Indeed, we and others reported expression profiling of fibroblasts isolated by cell sorting. This protocol was successfully performed to isolate and profile highly enriched populations of fibroblasts from skin, mammary, pancreas and lung tissues. Moreover, our method also allows culturing of sorted cells, in order to perform functional experiments and to avoid contamination by tumor cells, which is often a big obstacle when trying to culture CAFs.