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Aberrant methylation of the adenomatous polyposis coli (APC) gene promoter occurs in about 40% of breast tumours and has been correlated with reduced APC protein levels. To what extent epigenetic alterations of the APC gene may differ according to specific breast cancer phenotypes, remains to be elucidated. Our aim was to explore the role of APC methylation in the inflammatory breast cancer (IBC) phenotype. The status of APC gene promoter hypermethylation was investigated in DNA from normal breast tissues, IBC and non-IBC by both conventional and real-time quantitative methylation-specific PCR (MSP). APC methylation levels were compared with APC mRNA and protein levels. Hypermethylation of the APC gene promoter was present in 71% of IBC samples (n=21) and 43% of non-IBC samples (n=30) by conventional MSP (P=0.047). The APC gene also showed an increased frequency of high methylation levels in IBC (in 74% of cases, n=19) vs non-IBC (in 46% of cases, n=35) using a qMSP assay (P=0.048). We observed no significant association between APC methylation levels by qMSP and APC mRNA or protein expression levels. In conclusion, for the first time, we report the association of aberrant methylation of the APC gene promoter with the IBC phenotype, which might be of biological and clinical importance.

Purpose: A fibrotic focus, the scar-like area found in the center of an invasive breast tumor, is a prognostic parameter associated with an expansive growth pattern, hypoxia, and (lymph)angiogenesis. Little is known about the molecular pathways involved., Experimental Design: Sixty-five patients were selected of whom microarray data of the tumor and H&E slides for histologic analysis were available. The growth pattern and the presence and size of a fibrotic focus were assessed. Differences in biological pathways were identified with global testing. The correlations of growth pattern and fibrotic focus with common breast cancer signatures and with clinicopathologic variables and survival were investigated., Results: Tumors with a large fibrotic focus showed activation of Ras signaling and of the hypoxia-inducible factor-1alpha pathway. Furthermore, unsupervised hierarchical cluster analysis with hypoxia- and (lymph)angiogenesis-related genes showed that hypoxia-inducible factor-1alpha, vascular endothelial growth factor A, and carbonic anhydrase 9 were overexpressed. The presence of a fibrotic focus, especially a large fibrotic focus, was associated with the basal-like subtype (P = 0.009), an activated wound-healing signature (P = 0.06), and a poor-prognosis 76-gene signature (P = 0.004). The presence of a fibrotic focus (P = 0.02) and especially of a large fibrotic focus (P = 0.004) was also associated with early development of distant metastasis., Conclusions: Our results sustain the hypothesis that hypoxia-driven angiogenesis is essential in the biology of a fibrotic focus. Ras and Akt might play a role as downstream modulators. Our data furthermore suggest that vascular endothelial growth factor A does not only drive angiogenesis but also lymphangiogenesis in tumors with a fibrotic focus. Our data also show an association between the presence of a fibrotic focus and infaust molecular signatures.

A fibrotic focus is a scar-like area in the centre of a carcinoma and can be regarded as a focus of exaggerated reactive tumour stroma formation. Although modern surgical pathology uses different histopathological and molecular markers to assess the aggressiveness and predict the behaviour of malignant tumours, markers reflecting stromal cell behaviour and interactions between epithelial cells and stromal cells are scarce. In this review we summarize all studies investigating the value of a fibrotic focus as a prognostic factor and as a surrogate marker for hypoxia and (lymph)angiogenesis in patients with breast cancer. These data show that a fibrotic focus can be used as a practical, easily assessable and reproducible integrative histological prognostic parameter in breast cancer. We propose a consensus methodology to assess the fibrotic focus in breast cancer.

Purpose: Lymph node (LN) lymphangiogenesis has recently been shown to be important in the premetastatic niche of sentinel LNs. To study its role in the further metastatic spread of human breast cancer, we investigated the association of angiogenesis and lymphangiogenesis in sentinel LN metastases with the presence of nonsentinel LN metastases in breast cancer patients with a positive sentinel LN., Experimental Design: Angiogenesis and lymphangiogenesis--quantified as endothelial cell proliferation fraction (ECP%) and lymphatic ECP fraction (LECP%)--were assessed in sentinel LN metastases of 65 T(1)/T(2) patients with breast cancer using CD34/Ki67 and D2-40/Ki67 immunohistochemical double stains. Correlations were analyzed between nonsentinel LN status, LECP%, and other clinicopathologic variables (number of involved sentinel LNs, size of the primary tumor and LN metastasis, presence of lymphovascular invasion in the primary tumor, and of extracapsular growth in the sentinel LN metastasis)., Results: Thirty seven out of 65 patients (56.9%) had at least one involved nonsentinel LN. Size of the sentinel LN metastasis (P = 0.001), lymphovascular invasion (P = 0.02), extracapsular growth (P = 0.02), and LECP% (P = 0.01) were correlated with a positive nonsentinel LN status. The multivariate logistic regression model retained high LECP% (odds ratios = 4.2, P = 0.01) and the presence of extracapsular growth (odds ratios = 3.38, P = 0.04) as independently associated with the presence of nonsentinel LN metastases., Conclusions: Increased sentinel LN metastasis lymphangiogenesis is associated with metastatic involvement of nonsentinel axillary LNs. These are the first data sustaining the hypothesis that sentinel LN lymphangiogenesis is involved in further metastatic spread of human breast cancer.

We studied the presence of lymphangiogenesis in lymph node (LN) metastases of breast cancer. Lymph vessels were present in 52 of 61 (85.2%) metastatically involved LNs vs 26 of 104 (25.0%) uninvolved LNs (P<0.001). Furthermore, median intra- and perinodal lymphatic endothelial cell proliferation fractions were higher in metastatically involved LNs (P<0.001). This is the first report demonstrating lymphangiogenesis in LN metastases of cancer in general and breast cancer in particular.

Schistosomiasis is characterised by periovular granuloma formation within the portal tract and presinusoidal venules. As inflammation wanes, continued attempts to wall off and repair hepatic injury, lead to the development of extensive fibrosis. The codependence of chronic inflammation and angiogenesis is a well-known phenomenon. Neovascularisation is a complex process of endothelial cell proliferation and remodelling of the extracellular matrix. Previous studies demonstrated the ability of schistosome soluble egg antigens (SEAs) to stimulate endothelial cell activation in vitro. In the present study, we investigated the angiogenic potential of SEA in Swiss and BALb/c mice, after infection with Schistosoma mansoni or S. haematobium and by implanting SEA-coated beads into the murine liver. Anti-CD34 and anti-Ki-67 immunohistochemical stainings demonstrated newly formed blood vessels within and at the periphery of the granulomas. However, in one third of the granulomas the pre-existing portal stroma was not destroyed by the granulomatous inflammation, angiogenesis was minimal or absent and further growth of the granuloma was prevented. In C57BL/6J and C3H/HeN inbred mice, this polarisation was even more pronounced. In 91% of the granulomas in C57BL6/J mice the portal stroma was preserved. These mice had significantly smaller granulomas, less fibrosis and less mortality as compared to the high pathology C3H/HeN mice, where 87% of the granulomas were of the angiogenic type with destruction of the pre-existing stroma, leading to more severe chronic pathology. Thus, host's genetic mechanisms regulating the degree of angiogenesis and fibrosis, determine the severity of schistosome-induced pathology.

Recently, peritumoural (lympho)vascular invasion, assessed on haematoxylin-eosin (HE)-stained slides, was added to the St Gallen criteria for adjuvant treatment of patients with operable breast cancer (BC). New lymphatic endothelium-specific markers, such as D2-40, make it possible to distinguish between blood (BVI) and lymph vessel invasion (LVI). The aim of this prospective study was to quantify and compare BVI and LVI in a consecutive series of patients with BC. Three consecutive sections of all formalin-fixed paraffin-embedded tissue blocks of 95 BC resection specimens were (immuno)histochemically stained in a fixed order: HE, anti-CD34 (pan-endothelium) and anti-D2-40 (lymphatic endothelium) antibodies. All vessels with vascular invasion were marked and relocated on the corresponding slides. Vascular invasion was assigned LVI (CD34 [plus sign in circle] or [minus sign in circle]/D2-40 [plus sign in circle]) or BVI (CD34 [plus sign in circle]/D2-40 [minus sign in circle]) and intra- (contact with tumour cells or desmoplastic stroma) or peritumoural. The number of vessels with LVI and BVI as well as the number of tumour cells per embolus were counted. Results were correlated with clinico-pathological variables. Sixty-six (69.5%) and 36 (37.9%) patients had, respectively, LVI and BVI. The presence of 'vascular' invasion was missed on HE in 20% (peritumourally) and 65% (intratumourally) of cases. Although LVI and BVI were associated intratumourally (P=0.02), only peritumoural LVI, and not BVI, was associated with the presence of lymph node (LN) metastases (p(peri)=0.002). In multivariate analysis, peritumoural LVI was the only independent determinant of LN metastases. Furthermore, the number of vessels with LVI was larger than the number of vessels with BVI (P=0.001) and lymphatic emboli were larger than blood vessel emboli (P=0.004). We demonstrate that it is possible to distinguish between BVI and LVI in BC specimens using specific lymphatic endothelium markers. This is important to study the contribution of both processes to BC metastasis. Furthermore, immunohistochemical detection of lymphovascular invasion might be of value in clinical practice.

Enteric or intestinal-type neoplasms of the vagina are extremely rare. The present report describes a 55-year-old woman who presented with a 6-week history of postmenopausal bleeding. On clinical examination, a lesion on the posterior vaginal wall was noticed. Rectovaginal examination suggested a tumor in the rectovaginal septum. Biopsy revealed an adenocarcinoma of the intestinal type, with a small remnant of a villous adenoma. The histologic interpretation pointed in the direction of a metastatic gastrointestinal tumor; yet, clinical examination, rectoscopy, colonoscopy, magnetic resonance imaging (MRI) of the abdomen, and positron emission tomography (PET) scanning excluded this possibility. This led to the conclusion that the lesion was a primary intestinal-type adenocarcinoma of the vagina that had arisen from a vaginal villous adenoma. It is important to be aware of this tumor type and to distinguish them from metastatic colorectal adenocarcinoma in order to plan appropriate treatment.

We present the case of a 2-year-old girl who was referred to the ENT Department of the Antwerp University with a bony obstruction of the left side of the nose. The lesion originated from the ethmoid roof and the middle turbinate and extended into the nasopharynx. A partial functional resection was performed using an endoscopic approach. A combination of clinical, radiographic and histological information permitted a diagnosis of a benign fibro-osseous lesion: sinonasal hamartoma. The benign nature of this tumor justified a conservative follow-up.

Hypoxia and angiogenesis are important factors in breast cancer progression. Little is known of hypoxia and angiogenesis in lymph node metastases of breast cancer. The aim of this study was to quantify hypoxia, by hypoxia-induced marker expression levels, and angiogenesis, by endothelial cell proliferation, comparing primary breast tumours and axillary lymph node metastases. Tissue sections of the primary tumour and a lymph node metastasis of 60 patients with breast cancer were immunohistochemically stained for the hypoxia-markers carbonic anhydrase 9 (CA9), hypoxia-inducible factor-1alpha (Hif-1alpha) and DEC-1 and for CD34/Ki-67. Endothelial cell proliferation fraction (ECP%) and tumour cell proliferation fraction (TCP%) were assessed. On haematoxylin-eosin stain, the growth pattern and the presence of a fibrotic focus were assessed. Hypoxia-marker expression, ECP% and TCP% in primary tumours and in lymph node metastases were correlated to each other and to clinico-pathological variables. Median ECP% and TCP% in primary tumours and lymph node metastases were comparable (primary tumours: ECP%=4.02, TCP%=19.54; lymph node metastases: ECP%=5.47, TCP%=21.26). ECP% correlated with TCP% (primary tumours: r=0.63, P<0.001; lymph node metastases: r=0.76, P<0.001). CA9 and Hif-1alpha expression were correlated (primary tumours P=0.005; lymph node metastases P<0.001). In primary tumours, CA9 and Hif-1alpha expression were correlated with DEC-1 expression (P=0.05), presence of a fibrotic focus (P<0.007) and mixed/expansive growth pattern (P<0.001). Primary tumours and lymph node metastases with CA9 or Hif-1alpha expression had a higher ECP% and TCP% (P<0.003); in primary tumours, mixed/expansive growth pattern and fibrotic focus were characterised by higher ECP% (P=0.03). Furthermore, between primary tumours and lymph node metastases a correlation was found for ECP%, TCP%, CA9 and Hif-1alpha expression (ECP% r=0.51, P<0.001; TCP r=0.77, P<0.001; CA9 and Hif-1alpha P<0.001). Our data demonstrate that the growth of breast cancer lymph node metastases is angiogenesis dependent and that angiogenesis and hypoxia in the primary tumour predict angiogenesis and hypoxia in the lymph node metastases. Together with previous findings in breast cancer liver metastases, which grow in 96% of cases angiogenesis independently, these data suggest that both the intrinsic growth characteristics and angiogenic potential of breast cancer cells and the site-specific tumour microenvironment determine angiogenesis and hypoxia in breast cancer.

Purpose: At the time of diagnosis, metastatic dissemination of tumor cells via the lymphatic system has occurred in nearly all patients with inflammatory breast cancer (IBC). The objective of this study was twofold: (a) to determine which is the most suitable marker of lymph vessels in primary breast tumors and (b) to compare histomorphometric lymph vessel variables in IBC and non-IBC., Experimental Design: Serial sections of 10 IBCs and 10 non-IBCs were immunostained for D2-40, LYVE-1, podoplanin, and PROX-1. Relative lymph vessel area, lymph vessel perimeters, and counts and lymphatic endothelial cell proliferation (LECP) were then measured in D2-40/Ki-67 double-immunostained sections of 10 normal breast tissues, 29 IBCs, and 56 non-IBCs., Results: D2-40 was the most suitable antibody for staining peritumoral and intratumoral lymph vessels. D2-40-stained intratumoral lymph vessels were present in 80% of non-IBCs and 82.8% of IBCs (P = 0.76). In non-IBC, lymph vessels located in the tumor parenchyma were smaller and less numerous than those at the tumor periphery (P < 0.0001) whereas in IBC, intratumoral and peritumoral variables were not significantly different. The mean relative tumor area occupied by lymph vessels was larger in IBC than in non-IBC (P = 0.01). LECP at the tumor periphery was higher in IBC than in non-IBC: median LECP was 5.74% in IBC versus 1.83% in non-IBC (P = 0.005)., Conclusions: The high LECP in IBC suggests that lymphangiogenesis contributes to the extensive lymphatic spread of IBC.

We validated with univariate and multivariate (Cox) analysis, the prognostic value of the mitotic activity index (MAI), the fibrotic focus (FF) and other prognosticators in 448 patients with lymph node-negative (LN-) invasive breast cancer <55 years without adjuvant systemic treatment (72.5 months median follow-up, range 4-119). Of these patients, 24.8% developed distant and 1.6% loco-regional recurrence. FF showed excellent inter-observer reproducibility (kappa = 0.93). Strong prognosticators were MAI, grade, nuclear atypia, FF and the St. Gallen criterion (SG). The subgroup with excellent survival selected by SG was only 16% of all patients, implying over-treatment of more than 70% of all LN- patients when using SG as adjuvant therapy selection criterion. If MAI <10, 13% showed distant metastases, contrasting with 41% if MAI > or = 10. FF was prognostic in the ductal and mixed ductal cancers, but not in the lobular and other subtype cancers. Patients with invasive (mixed) ductal cancers with FF absent, FF < 1/3 or FF > 1/3 of the tumour area, had distant metastasis rates of 17%, 35% and 48%; in MAI < 10 and FF absent, FF < 1/3 or FF > 1/3, metastasis rates were 11%, 13% and 42% and if MAI > or = 10, metastasis rates were 31%, 48% and 50%, respectively. In the 12 patients with MAI < 10 and a large FF > 1/3, event-free survival was similar to patients with MAI > or = 10. With multiple regression MAI < 10 versus > or = 10 is the strongest prognosticator (also stronger than the SG). The FF may be important as it has additional prognostic value to the MAI in the small subgroup of invasive ductal or mixed-ductal breast cancer patients with combined MAI < 10 and an FF > 1/3 of the tumour area.

Background: Myxoid leiomyosarcoma (MLMS) of the vulva is a mesenchymal tumor with only five reported cases in the literature., Case: We report an 85-year-old woman with a unilateral nonulcerating, painless vulvar mass. According to the patient, the mass slowly enlarged. Initial biopsies were benign. However, clinically, this lesion was suspicious for a soft tissue tumor. Therefore, the mass was removed by a wide local excision. Definitive histology revealed a myxoid leiomyosarcoma of the vulva. At present, 25 months after the operation, the patient is well with no sign of recurrence., Conclusions: Vulvar myxoid leiomyosarcomas are rare and can be confused with other benign or malignant tumors. It is important to be aware of this rare tumor variant, in order to plan appropriate treatment.

Purpose: Inflammatory breast cancer is a distinct and aggressive form of locally advanced breast cancer with unique clinical and pathological features. Recently, histologic evidence of intense angiogenesis was found in inflammatory breast cancer specimens. The aim of this study was to confirm the angiogenic phenotype of inflammatory breast cancer and to investigate its potential to induce lymphangiogenesis., Experimental Design: Real-time quantitative reverse transcriptase-PCR was used to measure levels of mRNA of tumor angiogenesis and lymphangiogenesis-related factors [vascular endothelial growth factor (VEGF)-A, VEGF-C, VEGF-D, Flt-1, KDR, Flt-4, Ang-1, Ang-2, Tie-1, Tie-2, cyclooxygenase-2, fibroblast growth factor-2 (FGF-2), Egr-1, Prox-1, and LYVE-1] in tumor specimens of 16 inflammatory breast cancer and 20 noninflammatory breast cancer patients. Tissue microarray technology and immunohistochemistry were used to study differential protein expression of some of the angiogenic factors in inflammatory breast cancer and noninflammatory breast cancer. Active lymphangiogenesis was further assessed by measuring lymphatic endothelial cell proliferation., Results: Inflammatory breast cancer specimens had significantly higher mRNA expression levels than noninflammatory breast cancer specimens of the following genes: KDR (P = 0.033), Ang-1, (P = 0.0001), Tie-1 (P = 0.001), Tie-2 (P = 0.001), FGF-2 (P = 0.002), VEGF-C (P = 0.001), VEGF-D (P = 0.012), Flt-4 (P = 0.001), Prox-1 (P = 0.005), and LYVE-1 (P = 0.013). High mRNA levels of FGF-2 and cyclooxygenase-2 corresponded to increased protein expression by immunohistochemistry. Inflammatory breast cancer specimens contained significantly higher fractions of proliferating lymphatic endothelial cells than noninflammatory breast cancer specimens (P = 0.033)., Conclusions: Using real-time quantitative reverse transcriptase-PCR and immunohistochemistry, we confirmed the intense angiogenic activity in inflammatory breast cancer and demonstrated the presence of active lymphangiogenesis in inflammatory breast cancer. This may help explain the high metastatic potential of inflammatory breast cancer by lymphatic and hematogenous route. Both pathways are potential targets for the treatment of inflammatory breast cancer.

Background: Cyclooxygenase (COX)-2 expression and tumour-induced angiogenesis appear to be increased in squamous cell carcinoma (SCC) of the skin. In other cancers, COX-2 is a pro-angiogenic factor. The association between angiogenesis and COX-2 has not been studied in skin cancer., Objectives: To assess the onset of increased COX-2 expression and angiogenesis in the multistage carcinogenesis of SCC as well as the correlation between those two parameters., Patients/methods: We performed a retrospective paired immunohistochemical analysis of normal skin, actinic keratosis (AK), Bowen's disease (BD) and SCC among 35 individuals. Specimens were considered COX-2 immunopositive when 5% or more of the tumour cells showed clear evidence of immunostaining. To quantify active angiogenesis, we used a Ki-67-CD34 double-labelling immunohistochemical stain and calculated the fraction of proliferating endothelial cells. The Chalkley method was used to determine the microvessel density. To detect hypoxia, a carboanhydrase IX immunostain was used., Results: Compared with normal epidermis (0%), AK (31%), BD (22%) and SCC (40%) were significantly more likely to be COX-2 immunopositive (P < 0.01). The fraction of proliferating endothelial cells and the Chalkley scores paralleled multistage carcinogenesis (P < 0.05 between different stages). COX-2 immunopositivity was fairly correlated with hypoxia and higher proliferating endothelial cell fractions but not with Chalkley counts., Conclusions: Induction of COX-2 expression and angiogenesis are both early events in the development of SCC. In addition to ultraviolet light, hypoxia and COX-2 may be involved in skin tumour angiogenesis.

Aims: Inflammatory breast cancer (IBC) is an aggressive subtype of breast cancer with poor prognosis. The mechanisms responsible for the aggressive clinical evolution are incompletely understood. We constructed a tissue microarray (TMA) and validated its use in translational IBC research. Differential expression of proteins that might play a role in causing the IBC phenotype was studied., Methods and Results: A TMA containing 34 IBC and 41 non-stage matched non-IBC tumours was constructed. Five core biopsies were taken for each IBC and three cores for each non-IBC tumour. The TMA was validated using three approaches: (1) the excellent concordance between immunohistochemical results of the initial pathological examination and the results obtained with the TMA for ER, PR and HER2/neu (kappa > 0.74); (2) the known differential expression between IBC and non-IBC for four bio-markers in IBC (ER, PR, p53 and HER2/neu) was confirmed ( p < 0.01); (3) the HER2/neu status using three different antibodies (CB11, TAB250 and HercepTest) was highly concordant (kappa > 0.75). Furthermore, the overexpression of E-Cadherin and RhoC GTPase in IBC ( p < 0.05) was confirmed. We did not find a differential expression pattern for carbonic anhydrase IX (CA IX) and EGFR., Conclusions: Using different approaches, we have validated the use of our TMA for studying differential protein expression in IBC and non-IBC. We confirm the overexpression of E-Cadherin and RhoC GTPase in IBC. The lack of differential expression for CA IX and EGFR might suggest the pathways are equally utilised in both types of breast cancer.

Although angiogenesis is a prerequisite for the growth of most human solid tumours, alternative mechanisms of vascularisation can be adopted. We have previously described a non-angiogenic growth pattern in liver metastases of colorectal adenocarcinomas (CRC) in which tumour cells replace hepatocytes at the tumour-liver interface, preserving the liver architecture and co-opting the sinusoidal blood vessels. The aim of this study was to determine whether this replacement pattern occurs during liver metastasis of breast adenocarcinomas (BC) and whether the lack of an angiogenic switch in such metastases is due to the absence of hypoxia and subsequent vascular fibrinogen leakage. The growth pattern of 45 BC liver metastases and 28 CRC liver metastases (73 consecutive patients) was assessed on haematoxylin- and eosin-stained tissue sections. The majority of the BC liver metastases had a replacement growth pattern (96%), in contrast to only 32% of the CRC metastases (P<0.0001). The median carbonic anhydrase 9 (CA9) expression (M75 antibody), as a marker of hypoxia, (intensity x % of stained tumour cells) was 0 in the BC metastases and 53 in the CRC metastases (P<0.0001). There was CA9 expression at the tumour-liver interface in only 16% of the BC liver metastases vs 54% of the CRC metastases (P=0.002). There was fibrin (T2G1 antibody) at the tumour-liver interface in only 21% of the BC metastases vs 56% of the CRC metastases (P=0.04). The median macrophage count (Chalkley morphometry; KP-1 anti-CD68 antibody) at the interface was 4.3 and 7.5, respectively (P<0.0001). Carbonic anhydrase 9 score and macrophage count were positively correlated (r=0.42; P=0.002) in all metastases. Glandular differentiation was less in the BC liver metastases: 80% had less than 10% gland formation vs only 7% of the CRC metastases (P<0.0001). The liver is a densely vascularised organ and can host metastases that exploit this environment by replacing the hepatocytes and co-opting the vasculature. Our findings confirm that a non-angiogenic pattern of liver metastasis indeed occurs in BC, that this pattern of replacement growth is even more prevalent than in CRC, and that the process induces neither hypoxia nor vascular leakage.

The value of the fibrotic focus (FF) as a marker of intra-tumoral hypoxia in invasive breast carcinoma was assessed by studying its relationship with the expression of the hypoxia-induced carbonic anhydrase IX (CA IX), angiogenesis indices and prognosis. CA IX expression was immunohistochemically detected in 2 independent study groups, totaling 184 patients, and correlated with tumor characteristics, angiogenesis related parameters and patient outcome by univariate analysis. CA IX immunostaining scores in carcinoma cells and in tumoral fibroblasts were significantly higher in expansively growing tumors (p = 0.0001 and p < 10(-4), respectively), containing an FF (p = 0.0004 and p < 10(-4)) and showing high histological grade (p = 0.016 and p = 0.0006). Microvessel density, quantified by Chalkley counting, was correlated with CA IX expression both in the carcinoma cells and in the fibroblasts (p = 0.0076 and p = 0.0025) and with the presence and relative size of an FF (p = 0.006). The fraction of proliferating endothelial cells was positively correlated with CA IX scores in the fibroblasts (r = 0.4, p = 0.02) and with the presence of an FF (p = 0.02). CA IX scores in the fibroblasts--and to a lesser extent in the carcinoma cells--were associated with a higher relapse rate (p = 0.006) and a worse overall survival (p = 0.003). The highest CA IX immunostaining scores were found in the fibroblasts of large FF occupying more than one-third of the tumor. A large FF was associated with worse overall survival in a consecutive patient group (p = 0.01) and with shorter disease-free (p = 0.02) and overall survival (p = 0.0005) in T1-2N0 breast cancer patients. The strong association of CA IX expression with the presence of an FF shows that the latter is a marker of intra-tumoral hypoxia. FF is useful as a surrogate marker of hypoxia-driven ongoing angiogenesis and is associated with a higher relapse rate and a worse overall survival.

Aims: We postulated that skin metastases and cutaneous local recurrences from breast adenocarcinoma show different growth patterns with distinct angiogenic profiles., Methods and Results: Fifty-one surgically resected dermal breast cancer deposits were evaluated for growth pattern, E-cadherin expression, presence of necrosis and a fibrotic focus, fibrin deposition, carbonic anhydrase IX expression (CA IX), microvessel density, endothelial cell proliferation and blood vessel immaturity. Growth patterns were infiltrative, with carcinoma cells infiltrating the dermis without significant disturbance of the pre-existing architecture, expansive, meaning that a nodule of carcinoma cells and desmoplastic tissue pushed aside the pre-existing dermal structures, or mixed. All lobular carcinomas showed an infiltrative growth and lacked membranous E-cadherin expression. Different growth patterns in the ductal carcinomas were not correlated with differences in E-cadherin expression. The presence of necrosis and/or a fibrotic focus and the expression of the hypoxia marker CA IX were significantly associated with an expansive growth. Fibrin was present in all expansive deposits and less frequently in the other growth patterns. There was a positive association between fibrin deposition, CA IX expression and microvessel density. The latter was significantly higher in the expansive and mixed growth patterns than in the infiltrative pattern. Endothelial cell proliferation was highest in the expansive growth pattern and was positively correlated with the presence of a fibrotic focus and with fibrin deposition. The maximum percentage of immature blood vessels was higher in the expansive and mixed growth patterns than in the infiltrative one., Conclusion: The recognition of different subgroups of cutaneous breast cancer deposits with different degrees of hypoxia-driven angiogenesis may have important implications for the usefulness of anti-angiogenic therapy.

Inflammatory breast cancer (IBC) is the most aggressive form of breast cancer. Improved understanding of the mechanisms responsible for the differences between IBC and non-IBC might provide novel therapeutic targets. We studied 35 consecutive patients with IBC, biopsied prior to the initiation of chemotherapy. Angiogenesis was evaluated by Chalkley counting and by assessment of endothelial cell proliferation (ECP) and vessel maturity. The presence of fibrin, expression of the hypoxia marker carbonic anhydrase IX (CA IX) and epithelialcadherin (E-cadherin) expression were immunohistochemically detected. The same parameters were obtained in a group of 104 non-IBC patients. Vascular density, assessed by Chalkley counting (P<0.0001), and ECP (P=0.01) were significantly higher in IBC than in non-IBC. Abundant stromal fibrin deposition was observed in 26% of IBC and in only 8% of non-IBC (P=0.02). Expression of CA IX was significantly less frequent in IBC than in non-IBC with early metastasis (P=0.047). There was a significant positive correlation between the expression of CA IX and ECP in IBC (r=0.4, P=0.03), implying that the angiogenesis is partly hypoxia driven. However, the higher ECP in IBC and the less frequent expression of CA IX in IBC vs non-IBC points at a role for other factors than hypoxia in stimulating angiogenesis. Strong, homogeneous E-cadherin expression was found at cell-cell contacts in all but two IBC cases, both in lymphovascular tumour emboli and in infiltrating tumour cells, challenging our current understanding of the metastatic process. Both the intense angiogenesis and the strong E-cadherin expression may contribute to the highly metastatic phenotype of IBC.

Familial amyloidotic polyneuropathy is a systemic amyloidosis with a dismal prognosis for which a surgical treatment exists. Awareness of the clinical characteristics of the disease is critical for early genetic diagnosis and timely referral for liver transplantation. In this report we describe the history of a 49-year-old man in whom non-AA amyloidotic infiltration of the heart and the intestinal tract was diagnosed. Initially, inappropriate identification of the etiology of the disease led to maltreatment.

Restrictive dermopathy (RD) is a rare, fatal, autosomal recessive genodermatosis in which tautness of a translucent thin skin is the major clinical observation. This causes an intrauterine fetal akinesia deformation sequence (FADS) resulting in polyhydramnios, reduced fetal movements at around 31 weeks gestation, dysmorphic facies, arthrogryposis, and early neonatal death because of respiratory insufficiency. The characteristic histologic abnormalities of the skin are located in a thin dermis, consisting of compactly arranged collagen fibers, scant elastic fibers, and poorly developed skin appendages. The epidermal rete ridges are flattened and the dermal-hypodermal border is remarkably straight. The etiology of these changes remains unclear. We tested several existing hypotheses and could not confirm them. These included fibroblast dysfunction, abnormal keratin composition, desmosomal changes, and increased proinflammatory cytokines [tumor necrosis factor (TNF)-alpha and interleukin (IL)-6]. We conclude that RD is a relatively easy clinical and pathologic diagnosis, but that the pathogenesis of the disease is not clarified.

Tumor samples obtained from 106 primary breast cancer patients were examined biochemically (DCCA) and immunohistochemically (IHC) for estrogen (ER) and progesterone receptors (PR) to assess a quantitative relationship between both assays and to study the influence of the tumor-stroma ratio on this quantitative relationship. We used a model of logit transformation of IHC values (% of positive cells) and logarithmic transformation of DCCA values (fmol receptor/mg cytosolic protein). Tumors were subdivided into three categories according to the tumor-stroma ratio (more (t > s), equal amounts (t = s) or less (t < s) tumor than stroma), and the influence of the tumor-stroma ratio was studied using multiple regression analysis. We report a mathematical relationship between the results of the biochemical and immunohistochemical assays for the determination of ER status and PR status in primary breast cancer patients (ER: log DCCA(fmol/mg) = 0.369 logit (IHC(%pos cells)) + 2.328 (r = 0.573; p < 0.0001); PR: log DCCA (fmol/mg) = 0.474 logit (IHC(%pos cells)) + 0, 00 (r = 0.634; p < 0.0001)). In tumors overexpressing ER immunohistochemically (>10% nuclear positivity), median ER-DCCA is significantly higher if the tumor-stroma ratio is greater than 1. As these patients respond to hormonal treatment, depending on the degree of expression of both receptors, this study suggests that the biochemical assay be avoided because this technique is hampered by false-negative or falsely low results due to the loss of morphological information on the tumor-stroma ratio.

Idiopathic granulomatous mastitis is a very rare benign breast disease, which mimics breast cancer both clinically and mammographically. Most cases have an unknown aetiology, however, we found an alpha-1-antitrypsin deficiency. A literature review is presented and the controversies in diagnosis and management are discussed.

Thyroid acropachy is a rare manifestation of autoimmune thyroid disease, in the form of soft tissue swelling of the hands and feet with insidious onset, associated with clubbing and characteristic periosteal reactions. It is usually part of a syndrome consisting of a typical triad of thyroid acropachy, exophthalmos, and pretibial myxedema. The purpose of this case report is to demonstrate the imaging features of this typical triad in a 65-year-old-woman. This case is the first in which the MRI features of thyroid dermopathy are documented.

This case report describes the devastating consequences of spontaneous coronary dissection in a 36-year-old female patient. Surgical revascularization was attempted, but diffuse myocardial infarction developed. The patient was bridged to heart transplantation but died secondary to multiple organ failure. To our knowledge, this is the only reported case of spontaneous dissection of the three main coronary arteries due to severe cystic medial necrosis.

Subcutaneous granuloma annulare (SGA) is little known to radiologists. Better knowledge of this lesion may prompt accurate diagnosis. A typical case is presented with plain radiography, ultrasound and MR imaging, and is confirmed by histology. When an otherwise healthy child presents with a rapidly growing, solitary, nontender, subcutaneous soft tissue mass, located on the scalp or extensor aspect of the limbs, that radiologically presents as an indistinct radiodense and hypoechoic mass, isointense to muscle on T1- and slightly hypointense to fat on T2-weighted MR images, without calcifications, bone involvement or extracompartmental invasion, SGA should be suspected.

Severe heart failure in acute rheumatic myocarditis is rare. It may be rapidly reversible with treatment, so maximal medical treatment and, if necessary, mechanical support should be given before heart transplantation is considered.

A 44-year-old woman complaining of thoracic back pain was found to have a large mediastinal leiomyosarcoma. This was completely excised. No adjuvant treatment was given. Follow-up after 24 months showed no evidence of disease. For these rare tumours surgery offers the best treatment, although local recurrence and metastases do occur.

In isolated feline right ventricular papillary muscles, with intact or damaged (exposure to dry air for 30 s) endocardium, the effects of cholic acid were studied. At 35 degrees C and a [Ca2+] of 1.25 mM, isometric twitches and the maximal unloaded velocity of shortening (Vmax) were registered. Endocardial endothelium (EE) and myocardial ultrastructure were evaluated on scanning and transmission electron microscopy. In muscles with intact EE, low concentrations of cholic acid (3 x 10(-9) to 3 x 10(-7) M) or short incubation (30 min) with 3 x 10(-8) M induced a mild positive inotropic response, manifested as an increased peak isometric twitch tension (TT) and shortened twitch duration, without changes in Vmax. This inotropic response was abolished by damaging the EE. It was also absent after the addition of propranolol to muscles with intact EE. Higher concentrations (3 x 10(-6) to 3 x 10(-4) M) or prolonged incubation at 3 x 10(-8) M caused extensive morphologic damage of the EE, without detectable impairment of subjacent myocardium. This was accompanied by a progressive decrease in the TT and further twitch abbreviation, without changes in Vmax, resembling twitches obtained from muscles with experimentally damaged EE. These observations suggest that cholic acid induces an EE-dependent and beta-receptor-mediated positive inotropic effect, while simultaneously causing damage to EE cells.