Background: Tertiary lymphoid structures (TLSs) mediate local antitumor immunity, and interest in them significantly increased since cancer immunotherapy was implemented. We examined TLS− tumor stromal blood vessel interplay for each breast cancer (BC) molecular subtype related to recurrence, lymphovascular invasion (LVI), and perineural invasion (PnI). Methods: TLSs were quantified on hematoxylin and eosin stain specimens followed by CD34/smooth muscle actin (SMA) double immunostaining for stromal blood vessel maturation assessment. Statistical analysis linked microscopy to recurrence, LVI, and PnI. Results: TLS negative (TLS−) subgroups in each BC molecular subtype (except to Luminal A) have higher LVI, PnI, and recurrence. A significant rise in LVI and PnI were observed for the HER2+/TLS− subgroup (p < 0.001). The triple negative breast cancer (TNBC)/TLS− subgroup had the highest recurrence and invasion risk which was also significantly related to tumor grade. PnI but not LVI significantly influenced recurrence in the TNBC/TLS+ subgroup (p < 0.001). TLS−stromal blood vessel interrelation was different amongst BC molecular subtypes. Conclusion: BC invasion and recurrence are strongly influenced by TLS presence and stromal blood vessels, especially for HER2 and TNBC BC molecular subtypes.

Background: Breast cancer (BC) stroma has CD34- and αSMA-positive cancer-associated fibroblasts (CAFs) differently distributed. During malignant transformation, CD34-positive fibroblasts decrease while αSMA-positive CAFs increase. The prevalence of αSMA-positive CAFs in BC stroma makes microscopic examination difficult without digital image analysis processing (DIA). DIA was used to compare CD34- and αSMA-positive CAFs among breast cancer molecular subgroups. DIA-derived data were linked to age, survival, tumor stroma vessels, tertiary lymphoid structures (TLS), invasion, and recurrence. Methods: Double immunostaining for CD34 and αSMA showed different CAF distribution patterns in normal and BC tissues. Single CD34 immunohistochemistry on supplemental slides quantified tumor stroma CD34_CAFs. Digital image analysis (DIA) data on CAF density, intensity, stromal score, and H-score were correlated with clinico-pathologic factors. Results: CD34/αSMA CAF proportion was significantly related to age in Luminal A (LA), Luminal B (LB), and HER2 subtypes. CD34_CAF influence on survival, invasion, and recurrence of LA, LB-HER2, and TNBC subtypes was found to be significant. The CD34/αSMA-expressing CAFs exhibited a heterogeneous impact on stromal vasculature and TLS. Conclusion: BC stromal CD34_CAFs/αSMA_CAFs have an impact on survival, invasion, and recurrence differently between BC molecular subtypes. The tumor stroma DIA assessment may have predictive potential to prognosis and long-term follow-up of patients with breast cancer.

One controversial issue in pituitary pathology is the simultaneous proliferation of PitNETs and endothelial cells. No previous studies have compared the MIB1 Labeling Index (MIB1 LI) of PitNETs and stromal endothelial compartments and its connection with VEGF protein and gene expression. Simultaneous PitNETs proliferation index assessment in tumor and endothelial cells is related to VEGF protein and gene expression, and by using the automated QuPath platform for digital image analysis (DIA), it can be determined whether this dual proliferation specifically characterizes certain PitNETs subtypes. A total of 109 PitNETs were immunostained for endothelial cells (CD34) and proliferation (MIB1). VEGF was assessed by using IHC and RNA scopes. QuPath_DIA measured hormone-dependent MIB1 nuclear expression in tumor and stromal endothelial cells. MIB1 LI correlated with VEGF_mRNA and protein expression. PRL-secreting and non-functioning PitNETs had a high MIB1 LI in stromal endothelial cells. MIB1-positive tumor cell (%MIB1 LI.T) and endothelial cell (%MIB1 LI.E) percentages were substantially correlated (p = 0.01). The profiles of VEGF and hormones significantly and heterogeneously impact the MIB1-LI of tumor and endothelial cells. Tumor–endothelial cell proliferative interaction is specific to PRL-secreting and non-functioning PitNETs. These findings suggest that digital analysis of MIB1 and VEGF expression may serve as a valuable tool for risk stratification in PitNETs.

The expression and function of podoplanin (PDPN) in the normal human placenta has been debated in placental evaluation. This study emphasizes the importance of a multimodal approach of PDPN expression in normal human placentas. A complete examination is performed using immunohistochemistry, RNAscope and automated Digital Image examination (DIA) interpretation. QuPath DIA-based analysis automatically generated the stromal and histological scores of PDPN expression for immunohistochemistry and RNAscope stains. The umbilical cord’s isolated fibroblasts and luminal structures expressed PDPN protein and PDPN_mRNA. RNAscope detected PDPN_mRNA upregulation in syncytial placental knots trophoblastic cells, but immunohistochemistry did not certify this at the protein level. The study found a significant correlation between the IHC and RNAscope H-Score (p = 0.033) and Allred Score (p = 0.05). A successful multimodal strategy for PDPN assessment in human placentas confirmed PDPN expression heterogeneity in the full-term human normal placenta and umbilical cord at the protein and mRNA level. In placental syncytial knots trophoblastic cells, PDPN showed mRNA overexpression, suggesting a potential role in placenta maturation.

Mediastinal hemangiomas, particularly those of thymic origin, are rare phenomena. Due to its rarity, this pathologic condition is not characterized as related to the angiogenic profile of hemangioma endothelial cells. The diagnosis is challenging clinically and radiologically, and biopsies may not yield a definitive answer. Surgical resection offers the material for histologic diagnosis, relieves symptoms, and has a favorable long-term prognosis for such benign tumors. Sometimes, such benign tumors may have aggressive behavior and repeated recurrences but the causes responsible for this unpredictable evolution are not actually known. A case of intrathymic hemangioma diagnosed in a 16-year-old girl is presented here. We focused equally on a multidisciplinary approach to this challenging diagnosis but also on the characterization of the hemangioma endothelial cells profile not previously performed for such type of vascular anomalies. To define an antibodies panel for the evaluation of intrathymic hemangiomas may help in the full characterization of this rare vascular lesion, and subsequently focus on the new therapeutic targets which may be applied for cases with aggressive behavior.