Image analysis of immune cell patterns in the human mammary gland during the menstrual cycle refines lymphocytic lobulitis



To improve microscopic evaluation of immune cells relevant in breast cancer oncoimmunology, we aim at distinguishing normal infiltration patterns from lymphocytic lobulitis by advanced image analysis. We consider potential immune cell variations due to the menstrual cycle and oral contraceptives in non-neoplastic mammary gland tissue.


Lymphocyte and macrophage distributions were analyzed in the anatomical context of the resting mammary gland in immunohistochemically stained digital whole slide images obtained from 53 reduction mammoplasty specimens. Our image analysis workflow included automated regions of interest detection, immune cell recognition, and co-registration of regions of interest.


In normal lobular epithelium, seven CD8\(^{+}\) lymphocytes per 100 epithelial cells were present on average and about 70% of this T-lymphocyte population was lined up along the basal cell layer in close proximity to the epithelium. The density of CD8\(^{+}\) T-cell was 1.6 fold higher in the luteal than in the follicular phase in spontaneous menstrual cycles and 1.4 fold increased under the influence of oral contraceptives, and not co-localized with epithelial proliferation. CD4\(^{+}\) T-cells were infrequent. Abundant CD163\(^{+}\) macrophages were widely spread, including the interstitial compartment, with minor variation during the menstrual cycle.


Spatial patterns of different immune cell subtypes determine the range of normal, as opposed to inflammatory conditions of the breast tissue microenvironment. Advanced image analysis enables quantification of hormonal effects, refines lymphocytic lobulitis, and shows potential for comprehensive biopsy evaluation in oncoimmunology.

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The authors thank Prof. Dr. Dr. U. Welsch, Department for Cell Biology, Biomedical Center Munich, for his kind permission to use materials from previous studies that were collected during his term as Head of the Department for Microscopic Anatomy and Cytology (Chair II), Institute for Anatomy, Ludwig Maximilians University (LMU) Munich, Germany. The authors thank Pia Unterberger for organizing and storing archival material of the reduction mammoplasty series. The authors thank Gregory Apou for collaboration and scientific discussions on image analysis. The authors thank Priv.-Doz. Dr. M. Christgen, Institute of Pathology, Hannover Medical School, for breast pathology expertise. The authors thank the team of the Immunohistochemistry Laboratory, Institute for Pathology, Hannover Medical School for technical assistance, and the team of microDimensions, Munich, for help with 3D histology reconstruction.


This study was funded by the German Federal Ministry of Education and Research (BMBF) for the collaborative SYSIMIT project, grant number 01ZX1308A–D.

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Author contribution

NSS and FF designed the study, analyzed the results, and prepared the manuscript. NSS, RS, AG, GF, and CW contributed to the concept and implementation of the image analysis workflow. NSS performed the statistical analysis. JCLA, HHK, and HH contributed to study design and edited the manuscript. NK established the staining and image acquisition workflow. MS and FF designed the collection protocol for the reduction mammoplasty material and obtained patient histories and informed patient consent. All authors approved the final manuscript.

Corresponding author

Correspondence to Friedrich Feuerhake.

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Conflict of interest

Dr. Ralf Schönmeyer is a full-time employee of Definiens AG, Munich. All other authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving use of surplus tissue samples and clinical information on human participants were in accordance with the ethical standards of the institutional research committee (Reference number of the Ethical Committee/institutional review board #2063–2013) and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study. This article does not contain any studies with animals performed by any of the authors.

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Schaadt, N.S., Alfonso, J.C.L., Schönmeyer, R. et al. Image analysis of immune cell patterns in the human mammary gland during the menstrual cycle refines lymphocytic lobulitis. Breast Cancer Res Treat 164, 305–315 (2017).

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  • Lymphocytic lobulitis
  • Digital pathology
  • Oncoimmunology
  • Object-based image analysis
  • Menstrual cycle
  • Hormonal fluctuations