title: Inoculated mammary carcinoma-associated fibroblasts: contribution to hormone independent tumor growth.
abstract: BACKGROUND
  Increasing evidence has underscored the role of carcinoma associated fibroblasts (CAF) in tumor growth. However, there are controversial data regarding the persistence of inoculated CAF within the tumors. We have developed a model in which murine metastatic ductal mammary carcinomas expressing estrogen and progesterone receptors transit through different stages of hormone dependency. Hormone dependent (HD) tumors grow only in the presence of progestins, whereas hormone independent (HI) variants grow without hormone supply. We demonstrated previously that CAF from HI tumors (CAF-HI) express high levels of FGF-2 and that FGF-2 induced HD tumor growth in vivo. Our main goal was to investigate whether inoculated CAF-HI combined with purified epithelial (EPI) HD cells can induce HD tumor growth.
  
  
  METHODS
  Purified EPI cells of HD and HI tumors were inoculated alone, or together with CAF-HI, into female BALB/c mice and tumor growth was evaluated. In another set of experiments, purified EPI-HI alone or combined with CAF-HI or CAF-HI-GFP were inoculated into BALB/c or BALB/c-GFP mice. We assessed whether inoculated CAF-HI persisted within the tumors by analyzing inoculated or host CAF in frozen sections of tumors growing in BALB/c or BALB/c-GFP mice. The same model was used to evaluate early stages of tumor development and animals were euthanized at 2, 7, 12 and 17 days after EPI-HI or EPI-HI+CAF-HI inoculation. In angiogenesis studies, tumor vessels were quantified 5 days after intradermal inoculation.
  
  
  RESULTS
  We found that admixed CAF-HI failed to induce epithelial HD tumor growth, but instead, enhanced HI tumor growth (p < 0.001). Moreover, inoculated CAF-HI did not persist within the tumors. Immunofluorescence studies showed that inoculated CAF-HI disappeared after 13 days. We studied the mechanisms by which CAF-HI increased HI tumor growth, and found a significant increase in angiogenesis (p < 0.05) in the co-injected mice at early time points.
  
  
  CONCLUSIONS
  Inoculated CAF-HI do not persist within the tumor mass although they play a role during the first stages of tumor formation promoting angiogenesis. This angiogenic environment is unable to replace the hormone requirement of HD tumors that still need the hormone to recruit the stroma from the host.
authors.affiliation: Laboratory of Hormonal Carcinogenesis, Instituto de Biología y Medicina Experimental (Consejo Nacional de Investigaciones Científicas y Técnicas - CONICET), Vuelta de Obligado 2490, Buenos Aires, Argentina.
authors.forename: Victoria T
authors.initials: VT
authors.lastname: Fabris
authors.forename: Ana
authors.initials: A
authors.lastname: Sahores
authors.forename: Silvia I
authors.initials: SI
authors.lastname: Vanzulli
authors.forename: Lucas
authors.initials: L
authors.lastname: Colombo
authors.forename: Alfredo A
authors.initials: AA
authors.lastname: Molinolo
authors.forename: Claudia
authors.initials: C
authors.lastname: Lanari
authors.forename: Caroline A
authors.initials: CA
authors.lastname: Lamb
mesh_terms: D000818:Animals; D001943:Breast Neoplasms / Q000098:blood supply / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D018270:Carcinoma, Ductal, Breast / Q000098:blood supply / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D002450:Cell Communication; D049109:Cell Proliferation; D002470:Cell Survival; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology*; D012739:Gonadal Steroid Hormones / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D008822:Mice, Transgenic; D009368:Neoplasm Transplantation; D009389:Neovascularization, Pathologic / Q000378:metabolism / Q000473:pathology; D013997:Time Factors; D047368:Tumor Burden; D014407:Tumor Cells, Cultured
chemical_list: D012739:Gonadal Steroid Hormones; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins