Proceeding of Veterinary and Animal Science Days 2017, 6th- 8th June, Milan, Italy 

HAF © 2013 

Vol. IV, No. 1s   ISSN: 2283-3927 

   

 

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Keywords 

Tumor microenvironment, 

Mouse models, Histology, 

Immunohistochemistry, Digital 

image analysis 

 

CORRESPONDING AUTHOR 

Lucia Minoli  

lucia.minoli@unimi.it 

 

JOURNAL HOME PAGE 

riviste.unimi.it/index.php/haf 

Tumor microenvironment in experimental 

models of human cancer: morphological 

investigational approaches 

Lucia Minoli1,2* , Eugenio Scanziani1,2 

 

 

 

 

1University of Milan, Department of Veterinary Medicine, Italy  

2Fondazione Filarete, Mouse and Animal Pathology Laboratory (MAPLab), Milan, Itlay 

 

Abstract 

Tumor microenvironment (TME) is defined as the non-tumoral part of tumors. It is composed of 

different cell populations and structures (such as tumor-associated vasculature, immune-

inflammatory cells, fibroblasts). TME could either promote or antagonize tumor growth and has a 

great potential as target for novel therapeutic strategies (Hanahan and Coussens, 2012). Along with 

several methods (i.e. molecular assays), morphological techniques allow to evaluate the components 

of TME in the setting of their action. The aim of this work was to define valuable morphological 

approaches useful to investigate the TME. 

Histological and immunohistochemical techniques, along with digital image analysis, were tested on 

experimental mouse models (both xenograft and genetically engineered mice) of four human tumors 

(ovarian cancer, pancreatic ductal adenocarcinoma (PDAC), colon adenocarcinoma, thyroid 

carcinoma). 

Concerning the vascular compartment, CD31 immunostaining and double-immunofluorescence with 

CD31 and α-SMA (pericytes marker) allowed to respectively quantify vessels and evaluate their 

maturation degree. Immunohistochemical detection of previously administrated Pimonidazole, 

revealed variable extended areas of hypoxia in a consistent pattern between frozen and formalin-

fixed paraffin-embedded samples. 

Concerning the stromal component, anti-human MHC I and species-specific markers for Vimentin 

demonstrated the host-derivation of stroma in xenotumors, while Sirius Red histochemical staining 

allowed the quantification of desmoplasia in models of PDAC. 

Concerning immune-inflammatory cells, an immunohistochemical panel with CD3 (T-lymphocytes), 

B220 (B-lymphocytes), MPO (neutrophils) and Iba-1 (macrophages), showed high reliability in 

characterizing the tumoral infiltrate. 

 Moreover, the application of markers specific for different macrophage subsets confirmed the higher 

prevalence of M2 (Arginase I positive) on M1 (iNOS positive) macrophages. YM1 demonstrated low 

performance in detecting the M2 population (Fig. 1). 

http://creativecommons.org/licenses/by-nc-nd/3.0/deed.en


Proceeding of Veterinary and Animal Science Days 2017, 6th- 8th June, Milan, Italy 

HAF © 2013 

Vol. IV, No. 1s   ISSN: 2283-3927 

 

 

 

 

 

 

 

 

 

 

 

Due to the microenvironmental heterogeneity which influence tumor development and behavior, a 

sole quantification is unreliable for characterizing the TME. Considering that, morphological 

techniques proved to be a valuable approach, allowing the evaluation of spatial distribution and 

mutual interaction between the different elements. 

 

Acknowledgments 

The authors wish to thanks Dr. Giavazzi (IRCCS-Istituto di Ricerche Farmacologiche Mario Negri, Milan, 

Italy), Dr. Alessandra Capuano (Centro di Riferimento Oncologico, Aviano (PN), Italy), Dr. Greco 

(IRCCS-Istituto Nazionale dei Tumori, Milan, Italy) and their teams. 

 

 

 

Fig.1: Mouse, subcutaneous xenograft of thyroid carcinoma, same area, 200x. Hematoxylin&Eosin (A). 
Immunohistochemistry for Iba-1 (B) revealed a macrophage infiltrate, which turn out to be also Arginase I positive (C), 
demonstrating the prevalent M2 polarization of tumor-associated macrophages. Conversely, immunohistochemistry for 
iNOS (D) show the absence of M1-polarized macrophages. 

 

References 

Hanahan, D. and Coussens, L. M., 2012. Accessories to the crime: functions of cells recruited to the tumor 
microenvironment. Cancer Cell. 21, 309-322 

http://creativecommons.org/licenses/by-nc-nd/3.0/deed.en