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Background: Overexpression of chloride intracellular channel protein 1 (CLIC1) in tumor cells has been confirmed, but it has received less attention in the tumor blood vessel endothelium. Aim: The assessment of CLIC1 expression in ccRCC tumor blood vessels and its relationship with TNM parameters and tumor cell CLIC1 expression. Methods: CLIC1 immunostaining in ccRCC was evaluated in 50 cases in both malignant cells and tumor blood vessels (CLIC1 microvessel density-CLIC1-MVD) and was correlated with TNM staging parameters. Results: CLIC1-MVD was observed in approximately 65% of cases, and CLIC1 co-localization in both tumor and endothelial cells was observed in 59% of cases. ccRCC was classified into four groups (Classes 0–3) based on the percentage of positive tumor cells, with each group including sub-groups defined by CLIC1 expression in the endothelium. Class 3 (60–100% positive tumor cells) had the highest CLIC1-MVD, with an impact on T and M parameters (p value = 0.007 for T, and p value = 0.006 for M). For cases with CLIC1 intracellular translocation, there was a strong correlation between CLIC1-MVD and M (p value < 0.001). Conclusions: Co-expression of ccRCC tumor and endothelial cells promotes tumor progression and metastasis and should be investigated further as a potential therapeutic target for ccRCC and other human malignancies.

Background/Aim: Renal cell carcinoma is strongly vascularized, and formation of new blood vessels is a complex and multi-step process. In this study, we analysed the subtypes of intermediate blood vessels, as shown by double immunohistochemistry. Materials and Methods: Tumour-associated blood vessels were identified by double immunostaining based on CD34 and smooth muscle cell actin. Blood vessels were classified both quantitatively and qualitatively based on the expression of the aforementioned two markers. The main criteria to sub-classify intermediate blood vessels was the presence, distribution, and arrangement of perivascular cells. Results: We described three subtypes of intermediate blood vessels found particularly in the tumour area: Subtype 1 lacked perivascular cells, subtype 2 showed scattered pericytes attached to the vascular wall, and subtype 3 showed a continuous layer of perivascular cells on one side. Conclusion: We describe for the first time three subtypes of renal cell carcinoma-associated intermediate blood vessels, which could be important in prognosis and as potential targets for anti-vascular therapy.

Background/aim The histopathological variability of each type of pituitary adenoma (PA) that causes growth hormone (GH) excess influences the phenotype, radiological characteristics and therapy response of acromegaly patients. We correlated the immunohistochemical (IHC) features of GH-secreting PAs with their clinical, laboratory and imaging data. Patients and methods We included 32 patients with documented acromegaly; tumour specimens were histologically and IHC examined: anterior pituitary hormones, pituitary-specific transcription factor-1 (PIT-1), Ki-67 labelling index were evaluated. Results Macroadenomas represented 93.75%. Post-surgery disease control negatively correlated with the maximum initial tumour diameter (p=0.04). Ki-67 did not predict remission. No correlation was found between GH serum levels and IHC expression (p=0.45). PIT-1 was positive in all specimens, two had a weak expression. Four were considered PIT-1 positive plurihormonal adenomas and several had unusual IHC combinations. Conclusion PIT-1 accurately classifies GH-secreting PAs. The IHC classification as well as radiological dimensions and extent influence disease control, probably being the best prognosis factors.

Background/Aim: Chloride intracellular channel protein 1 (CLIC1) activates inflammasomes in rheumatoid (RA) and psoriatic (PsA) arthritis. We studied CLIC1 expression in RA and PsA patients’ skin with vasculitis and its variability depending on the therapy used. Materials and Methods: CLIC1 immunoexpression was evaluated in the vascular (CLIC1-V) and stromal (CLIC1-S) compartments of the RA and PsA skin biopsies of patients treated with methotrexate (MTX), leflunomid (LFN), corticotherapy (CT), or biological therapies. Results: MTX significantly reduced CLIC1-S expression (p=0.016), whereas LFN decreased CLIC1-V (p

Background/aim Angiogenic growth factors expression is not known in the normal cornea. The aim was to study corneal gene expression profile of VEGF and PDGF pathways influencing the avascular state of cornea. Materials and methods cDNA synthesis was performed from mRNA extracted from five fresh pig corneas followed by cDNA synthesis and analysis of VEGF and PDGF pathways by TaqMan Array gene expression profile. Results Normal pig cornea lacks VEGFR2 and VEGFR3 gene expression. MK2 and AKT1 genes were significantly overexpressed (p=0.000684, p=0.050995, respectively). Six PDGF pathway genes were overexpressed: TIAM1 (p=0.047), PIK3CA (p=0.00005), IKBKG (p=0.000006), PAK4 (p=0.034), RAC1 (p=0.000006 and PTGS2, p=0.00375). PDGF A was up-regulated, but not with a statistical significance (p=0.79911), while PDGFRα was down-regulated and PDGFRβ was not expressed. Conclusion Normal cornea avascularity is given by growth factor receptors down-regulation. Rapid corneal neovascularisation is induced by activation of the main angiogenic growth factors that induce angiogenic cascade and vessel recruitment.

Background/aim Invasive bladder cancer mortality remains high despite progresses made in early diagnosis and surgical procedures. Thus, there is a need to define new markers for bladder cancer. CLIC1 has not been previously studied in bladder cancer and thus, we aimed to assess its immunohistochemical expression in relation to different stages of bladder cancer development. Materials and methods Immunohistochemistry for CLIC1 was applied in 50 cases of muscle invasive bladder cancer. Results CLIC1 was not expressed in the normal urothelium, but a strong reaction was observed in dysplastic urothelium, carcinoma in situ and in 94% of the cases with invasive urothelial carcinoma; however, it was not expressed in squamous cell carcinoma cases. No correlation was found between the immunohistochemical expression of CLIC1 and the stage and grade of the tumour. Conclusion CLIC1 was overexpressed in urinary bladder dysplastic epithelium, carcinoma in situ and invasive carcinoma. CLIC1 constitutes a new potential marker of invasive bladder cancer.

Chronic stress is regarded as a significant factor in the etiology of the many diseases. Numerous methods have been developed through which the effect of chronic stress is examined. The aim of this study is to demonstrate the new experimental model for analysis of immuno-suppression induced by chronic restraint stress, through challenge with conditionally tumorigenic cell line BHK-21/C13. 20 male NMRI mice were randomly divided into 2 groups-control and experimental. Each mouse was subcutaneously inoculated with BHK-21/C13 cells. Stress in the experimental group was induced for 20 days. After the experiment, tumor masses were removed, and analyzed using histology and immunohistochemistry techniques. We found a statistically significant difference (p = 0.034) in tumor expression and tumor volumes (p = 0.0061) between groups, as well as in immunopositivity on Ki67, cytochrome C and matrix metalloproteinase 9. Absence of immune infiltrate was noticed in experimental, and the presence of inflammatory infiltrate at tumor invasion front in control group.

Background – digitalization in eastern European medical education is a stringent need of teaching and assessment processes, which must be updated to challenges continuously launched by medical students. In this context, our university implemented an e-learning and e-assessment system in histology, a preclinical education subject. Aim – to evaluate the impact of replacing a classic learning and assessment methods with a completely digitized one. Methodology – the new system impact on students’ skills and teacher’s performance, was evaluated by using an opinion pool questionnaire including as assessment criteria system compliance, student-teacher interaction, online access improvement and new facilities regarding e-assessment and time management was performed. Results – “computer gaming” was delightful for students. Management of information’s given to students was improved by a higher standardization and this, helped them to pass evaluations easier. Better stress management of examination induced the increase of attendance to first presentation. Graduation percentage and students’ confidence in the accuracy of the exam results were improved. Time for evaluation was significantly reduced giving to teachers the opportunity for a better management of their time dedicated to research or academic development. Teaching and exam module software was periodically updated, being adapted to our curriculum. Conclusions – e-learning and e-assessment had a quick positive impact on students and teacher’s despite of controversial issues which commonly come with any change

Background/Aim: We previously described four different vascular patterns (reticular, diffuse, fasciculate, and trabecular) in renal cell carcinoma (RCC) suggesting an early and heterogeneous acquisition of perivascular cells most probably due to a particular PDGF pathway gene expression profile. The aim of the study was to study PDGF pathway gene expression profiles, separately for each vascular pattern. Materials and Methods: TaqMan assay for the PDGF pathway was performed on twelve cases of ccRCC previously evaluated by histopathology, immunohistochemistry, and RNAscope. Gene expression profile was correlated with grade, invasion, vascular patterns, and VEGF. Results: PIK3C3 and SLC9A3 genes were overexpressed in all vascular patterns, but they were significantly correlated with high VEGF mRNA in the reticular and diffuse pattern. STAT1, JAK2, SHC2, SRF and CHUK (IKK) were exclusively overexpressed in cases with diffuse vascular pattern. SLC9A3, CHUK and STAT3 were overexpressed in G2 tumors. Conclusion: Three ccRCC subgroups were defined: 1) PIK3C3 (VSP34)/SLC9A3 which may be proper for anti PIK3C3 inhibitors; 2) VEGF(high) subgroup where association of anti VEGF may be a benefit and 3) JAK2/STAT1 subgroup, potentially being eligible for anti JAK/STAT therapy associated with IKK inhibitors.

Chloride intracellular channel protein 1 (CLIC1) is known as a promoter of cancer progression, metastasis, and angiogenesis. Thus, CLIC1 could be a future therapeutic target. This study aimed to evaluate the effect of anti-CLIC1 antibodies on tumour cells and vessels of human renal cell carcinoma (RCC) in rabbit cornea and chick embryo chorioallantoic membrane (CAM) models.Human cc-RCC xenografts on rabbit cornea and CAM surface were performed. Anti-CLIC1 antibodies were applied for 5 consecutive days on both tumor models. We comparatively evaluated treated and untreated tumors by combining ultrasonography with microscopic techniques.RCC implants rapidly recruited blood vessels and had an exponential growth rate on both tumor models. Anti-CLIC1 antibodies suppressed tumor growth by inducing tumor cell necrosis. Tumor vessels regressed rapidly but not completely during anti-CLIC1 antibodies based therapy.Anti-CLIC1 antibodies induced tumor necrosis and tumor vasculature regression in human cc-RCC xenografts in both in vivo experimental models.

Cushing's disease represents a rare endocrine disorder caused by hypercortisolism due to an ACTH-secreting pituitary tumor. Pituitary adenomas (PAs) are mostly benign tumors but may have features of invasiveness and aggressiveness. They are clinically classified by their hormone-secreting characteristics. Expression of pituitary hormones in adenohypophyseal cells is controlled by several transcription factors (PIT1 for GH, PRL and TSH lineage, SF1 for gonadotroph lineage and TPIT for ACTH lineage). Eight cases of Cushing's disease (CD) were included in the current study. The diagnostic was sustained by clinical features, hormonal assessment, and radiological findings (pituitary CT or MRI). All patients underwent surgical removal of pituitary tumor using transsphenoidal resection. All tissue specimens were evaluated using histological routine staining (hematoxylin and eosin) and immunohistochemical staining for vimentin, pituitary hormones (GH, PRL, TSH, FSH, LH, ACTH), and transcription factors (PIT1, TPIT, SF1). The intensity IHC scores ranged from 0 to 3+ and a staining higher than 30% (2+) was considered positive for interpreting the results. Based on the histopathological and immunohistochemical data correlated with the evolution of the disease (remission versus persistent disease), five molecular subgroups were identified. Subgroups 1 to 3 had a positive expression of ACTH and TPIT, with variations regarding SF1 expression: there was no expression in subgroup 1, nuclear positivity in 2, and cytoplasmic reaction in 3. Subgroups 4 and 5 consisted of plurihormonal adenomas with unusual immunohistochemical combination. They had IHC positive score for ACTH, GH, TPIT and were differentiated by the expression of SF1: lack of expression in subgroup 4 and intense cytoplasmic reaction in 5. The expression of SF1 was linked to the evolution of the disease, patients from subgroups which did not have SF1 cytoplasmic reaction (1, 2, 4) had remission of CD, while patients in subgroups 3 and 5 had persistent disease (including one case who evolved to exitus due to cardiovascular complications of the severe hypercortisolism). Transcription factors evaluation is important for classifying pituitary adenomas and finding clinicopathological correlations with the aim of providing a better therapeutical approach. In the current study, although SF1 is a marker of gonadotroph lineage (1), its aberrant cytoplasmic expression in CD may contribute to an aggressive evolution and to other cell lineage differentiation (PIT1, TPIT). Neou M et al, Pangenomic Classification of Pituitary Neuroendocrine Tumors. Cancer Cell. 2020 Jan 13;37(1): 123-134. e5. doi: 10.1016/j. ccell.2019.11. 002. Epub 2019 Dec 26. PMID: 31883967. Presentation: No date and time listed

ObjectiveControversial, heterogeneous, and inconsistent responses to beta-blockers have been reported in some cases of infantile proliferative hemangiomas. On the basis of these clinical observations, we aimed to examine the β1 adrenergic receptor (β1-AR) protein expression distribution among different types of pediatric vascular anomalies.MethodsImmunohistochemistry (IHC) was performed for β1-AR on 43 surgical specimens.ResultsWe found positive β1-AR IHC staining in all intramuscular hemangiomas, capillary–lymphatic, lymphatic, venous, and combined malformations, and Masson’s tumor cases, as well as in 7 of 10 cases of proliferative infantile hemangiomas.ConclusionsOur research demonstrates, for the first time, the degree of heterogeneous expression of β1-AR among pediatric vascular malformations. Our results support the need for β1-AR assessment in pediatric vascular anomalies to select cases with a robust response to β1-selective blockers. β1-AR assessment may have a strong impact on therapeutic refinement for pediatric vascular anomalies by selecting cases with a stronger response to beta-blockers.

Purpose. Clinically apparent anastomotic leakage (AL) after low anterior rectal resection (LAR

Background/aim Colon cancer liver metastases with desmoplastic growth pattern (dGP) have a highly heterogeneous therapy response. The aim of the study was to evaluate the dGP liver metastasis molecular profile from a chemo-naive patient by mimicking metastatic process on an experimental chick embryo chorioallantoic membrane (CAM) model. Materials and methods Three successive CAM passages of dGP human colorectal liver metastases were immunophenotyped for keratin (K) 8, and 20, CLIC1, VEGF, EGFR, CD34, podoplanin, Ki67, E-cadherin and vimentin. Results Metastatic cells gradually lost K20 while K8, E-cadherin and vimentin heterogeneously increased during passages. VEGF, CLIC 1, EGFR expression increased in metastatic cells especially at the tumor graft periphery. Scattered proliferating and non-proliferating podoplanin-positive tumor cells, lymphatic and blood vessels were heterogeneously detected in tumor xenografts depending on passage stage. Conclusion By mimicking repetitive metastatic processes we proved that metastatic cells change their phenotype. This may explain why not all metastases have a similar response to therapy.

Keeping students motivated in conditions of social distancing is a real challenge for teachers. The use of alternative methods that are not perceived as a conventional way of learning increases student’s curiosity. The game is a method of relaxation at any age, nowadays often combined with modern technology. Gamification is an alternative learning method increasingly applied in higher medical education for developing medical students’ practical skills. The online distance learning has challenged us to apply gamification in histology. We launched a competition for 2nd year medical students having as first award Romanian edition of the worldwide known histology book. In May 2020, in the middle of COVID 19 pandemic, we asked students to review our Case-Center histological slides from their own perspective, to select those tissues which resemble to anything from real life and to do art using modern technology. A Facebook page was created for posting students works. Students created high quality works using tissue pieces, their unlimited imagination and modern technology. We received 181 pictures, one animation and 3 videos with a brief message attached. A stunning compilation of Art in Histology competition works created a book entitled "181 HISTOries from 2nd year medical students during COVID19 pandemic."

We compared two E-assessment platforms and we evaluated the impact of online versus on-site examination by analysing students’ outcomes and classifying them into performance groups. Two groups of 480 (G1/2019) and 527 (G2/2020) 2nd year medical students were included. G1 was assessed by using E-School Exam Module on-site, while G2 used FlexiQuiz assessment platform online. G2 received 3 partial examinations during the semester while G1 was evaluated by o single final exam at the end of the semester. Students were classified in four performance categories as high (8-10, HPG), medium (6-7.99, MPG), low (5-5.99, LPG) and fail (less than 4, FPG) based on final grade. The analysis was applied for both theoretical and practical exams. A significant increase of HPG (79% vs 4% for theory and 42% vs 16% for practice) was noted for G2 compared with G1. MPG remained relatively the same between groups while LPG decreased for G2 compared with G1 (22% vs 1% for theory, 25% vs 4% for practice). The G2-FPG significantly decreased but remained higher for practical exam. HGP, LPG, FPG were strongly influenced by applying different assessment methods while MPG remained similar for G1 and G2. Factors influencing these variations will be discussed here.

Background/aim Chloride intracellular channel protein (CLIC1), E- and P-cadherin (Ecad, Pcad) are certified factors of aggressivity, but they have not been studied in breast cancer to date. The aim was to study CLIC1, Ecad and Pcad impact on breast cancer in terms of defining new high-risk subgroups. Materials and methods Ninety-seven breast cancer biopsies were immunohistochemically evaluated for CLIC1, Ecad and Pcad expression related to molecular subtypes. CLIC1 expression was assessed in both tumor cells (CLIC1T) and blood vessels (CLIC1V). Results For 23% of Luminal A cases, both cadherins and CLIC1V were positive. Luminal B/HER2 subtype, had two specific phenotypes: Ecad-/Pcad-/CLIC1T-/CLIC1V+ and Ecad+/Pcad-/CLIC1T-/CLIC1V+. All TNBC cases were clustered into two subgroups: 60% were Ecad+/Pcad+/CLIC1T+/CLIC1V+) while 40% were Ecad+/Pcad+/CLIC1T+/CLIC1V-). Conclusion CLIC1, Ecad and Pcad association stratifies molecular types of breast cancer in subgroups that may explain different response to therapy and different aggressiveness previously observed by other authors within the same molecular subtype.

To teach histology with the computer instead of the microscope seems to be like “a fight against the wave” especially for those countries with a “frozen in time” learning methods. Such changes seem to have an unexpected impact on teachers, students and society being paved with challenges and pitfalls solvable by own “step by step” experience overlapped on young generation actual needs. Present work started in 2017 at Department of Histology, by acquisition of E-School System. Conventional microscopes were replaced by computers and glass slides were scanned. We evaluated the e-Learning and eAssessment impact on student’s skills and performance and on teachers and tutors work. “Computer gaming” with histology was delightful for students. By contrast, teachers were afraid initially of losing jobs. Standardized information’s given to students helped them to pass evaluations easier. Exams stress decreased while attendance increased. Students’ confidence in the accuracy of the exam was improved. Time for evaluation was significantly reduced giving to teachers more time for research. Teaching and exam module software was periodically updated, being adapted to our curriculum. Conclusions. Implemented for the first time in preclinical medical education in Romania, eLearning and e-Assessment had a quick positive impact on students and teachers.

The frequency of the squamous cell carcinoma of the head and neck is constantly increasing, with over 550.000 new cases registered globally each year. The conventional histopathological diagnosis most commonly indicates the squamous cell carcinoma as tumor type and G2 as differentiation grade. Despite of this relative morphological uniformity, there is a great heterogeneity in the molecular profile, the therapeutic response and prognosis. Most probably, this entity includes many diseases, similar in basic morphologic features, but different in the biological behavior. Trying to answer this question and to show discrepancies when they exist, we have evaluated in this book chapter, our own results and data from the literature in terms of molecular profile at the protein level, including the spectrum of proliferation markers, growth factors and their receptors, stromal proliferation, angiogenesis and lymphangiogenesis. These data will allow to identify some major criteria for a better stratification of cases, selected for gene analysis and personalized therapy as a future perspective and direction.

Most of angiogenesis assays were designed and developed during Folkman's era. But the growth of new blood vessels in several pathologic conditions as tumor development or inflammation were observed long time ago.The development of new blood vessels was early observed by ancient Egyptians who tried to destroy them by applying empirical methods. From the first observations regarding angiogenesis to a personalized therapy targeting newly formed blood vessels a lot of experimental in vitro and in vivo angiogenesis assays have been developed. The present work will overview the oldest and less known part of angiogenesis assays development, and in addition, it will present the newest data in the experimental field of angiogenesis which is rapidly improved by the needs of new antiangiogenic and antivascular therapy development.

Most of angiogenesis assays were designed and developed during Folkman's era. But the growth of new blood vessels in several pathologic conditions as tumor development or inflammation were observed long time ago.The development of new blood vessels was early observed by ancient Egyptians who tried to destroy them by applying empirical methods. From the first observations regarding angiogenesis to a personalized therapy targeting newly formed blood vessels a lot of experimental in vitro and in vivo angiogenesis assays have been developed. The present work will overview the oldest and less known part of angiogenesis assays development, and in addition, it will present the newest data in the experimental field of angiogenesis which is rapidly improved by the needs of new antiangiogenic and antivascular therapy development.

Background/aim E- and P-cadherin (E-cadh, P-cadh) control tumor cell invasion, metastatic or stemness potential and chemotherapy resistance. The study aimed to assess E- and P-cadherin expression in breast cancer molecular subtypes. Materials and methods Immunohistochemistry for E-cadh and P-cadh was performed for 97 breast cancer cases. Membrane (M), cytoplasmic (C) or mixed (MC) patterns of E-cadh and P-cadh were considered in our evaluation. Results E-cadh and P-cadh C pattern was significantly correlated in the HER2 subtype (p=0.031). P-cadh M pattern was highly specific for the HER2 subtype (p=0.002). Only P-cadh C characterized the triple negative breast cancer subtype (p=0.015). For Luminal B/HER2 cases, P-cadh M pattern was strongly coexpressed with the E-cadh MC pattern (p=0.012). Progesterone receptor (PR) expression influenced E-cadh M pattern in the Luminal B/HER2 subtype (p=0.042). Conclusion E- and P-cadherins define distinct subgroups within breast cancer molecular subtypes. Our findings support the inclusion of E- and P-cadherin into breast cancer molecular classification.

Background/aim Human mesenchymal stem cells (hMSC) represent a versatile cell population, able to modulate the tumor microenvironment. Our aim was to recreate an open scene for the in vivo interaction between hMSC and the MCF-7 breast cancer cells (MCF-7), in order to enlighten the intimate involvement of hMSC in tumor vasculogenesis and angiogenesis. Materials and methods hMSC and MCF-7 were seeded onto the chick embryo chorioallantoic membrane (CAM) and incubated for 7 days. Consecutively, the morphology and the immunohistochemical profile of CAM were assessed. Results Following this complex interaction, MCF-7 acquired a more aggressive phenotype, hMSC switched to a vascular precursor phenotype, while CAM underwent a major reset to an earlier stage, with hotspots of angiogenesis, vasculogenesis and hematopoiesis. Conclusion The hallmark of this study was the establishment of a veritable in vivo experimental model of MSC involvement in tumor vasculogenesis and angiogenesis, allowing further analysis in the field.

PDGFs/PDGFRs axis is documented as an important tumor-promoting agent and potential therapeutic target for several human carcinomas, including breast cancer. However, little is known about the role played by the PDGF family members in the normal development of the breast tissue, breast carcinogenesis and tumor-microenvironment dynamics Despite its potent pro-lymphangiogenic effects, PDGF-B/PDGFR-beta axis remains controversial and incompletely elucidated in the field of breast cancer, with emphasis to its differential implications in breast cancer molecular subtypes. Although some data are available concerning this aspect, little or no information is found regarding the role of the PDGF-B/PDGFR-beta axis in rare and aggressive types of breast cancers, such as triple negative breast cancers (TNBCs) and its associated subtypes This review attempted to gather as many data as possible concerning PDGFs family members in the normal breast tissue and in breast carcinogenesis with special focus on their role in diagnosis and therapeutic approach.

Aggressive tumor cells can adopt an endothelial cell-like phenotype and contribute to the formation of a tumor vasculature, independent of tumor angiogenesis. This adoptive mechanism is referred to as vascular mimicry and it is associated with poor survival in cancer patients. To what extent tumor cells capable of vascular mimicry phenocopy the angiogenic cascade is still poorly explored. Here, we identify pericytes as important players in vascular mimicry. We found that pericytes are recruited by vascular mimicry-positive tumor cells in order to facilitate sprouting and to provide structural support of the vascular-like networks. The pericyte recruitment is mediated through platelet-derived growth factor (PDGF)-B. Consequently, preventing PDGF-B signaling by blocking the PDGF receptors with either the small tyrosine kinase inhibitor imatinib or blocking antibodies inhibits vascular mimicry and tumor growth. Collectively, the current study identifies an important role for pericytes in the formation of vascular-like structures by tumor cells. Moreover, the mechanism that controls the pericyte recruitment provides therapeutic opportunities for patients with aggressive vascular mimicry-positive cancer types. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference. Fil: Nowak Sliwinska, Patrycja. Université of Lausanne; Suiza. Univeristé of Geneve; Suiza Fil: Alitalo, Kari. Katholikie Universiteit Leuven; Bélgica Fil: Allen, Elizabeth. Katholikie Universiteit Leuven; Bélgica Fil: Anisimov, Andrey. Katholikie Universiteit Leuven; Bélgica Fil: Aplin, Alfred C.. University of Washington; Estados Unidos Fil: Auerbach, Robert. University of Wisconsin; Estados Unidos Fil: Augustin, Hellmut G.. Heidelberg University; Alemania. German Cancer Consortium; Alemania Fil: Bates, David O.. University of Nottingham; Reino Unido Fil: Beijnum, Judy R. van. Cancer Center Amsterdam; Países Bajos Fil: Bender, R. Hugh F.. University of California; Estados Unidos Fil: Bergers, Gabriele. Katholikie Universiteit Leuven; Bélgica Fil: Bikfalvi, Andreas. Universite de Bordeaux; Francia Fil: Bischoff, Joyce. Harvard Medical School; Estados Unidos Fil: Böck, Barbara C.. Heidelberg University; Alemania. German Cancer Consortium; Alemania Fil: Brooks, Peter C.. Maine Medical Center Research Institute; Estados Unidos Fil: Bussolino, Federico. Università di Torino; Italia. Candiolo Cancer Institute; Italia Fil: Cakir, Bertan. 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University of Amsterdam; Países Bajos Fil: Kleinmann, Hynda K.. The George Washington University; Estados Unidos Fil: Koolwijk, Pieter. Fondation Asile des Aveugles; Suiza. Universida de Lausanne; Suiza Fil: Kuczynski, Elisabeth. University of Toronto; Canadá Fil: Kwak, Brenda R.. University of Geneva; Suiza Fil: Koen, Marien. HistoGeneX; Bélgica Fil: Melero Martin, Juan M.. University of Liège; Bélgica Fil: Munn, Lance L.. Harvard Medical School; Estados Unidos Fil: Nicosia, Roberto F.. VA Puget Sound Health Care System; Estados Unidos Fil: Noel, Agnes. University of Liège; Bélgica Fil: Nurro, Jussi. University of Eastern Finland; Finlandia Fil: Olsson, Anna-Karin. Uppsala University; Suiza Fil: Petrova, Tatiana V.. Ludwig Institute for Cancer Research Lausanne; Suiza Fil: Pietras, Kristian. Division of Translational Cancer Research; Suecia Fil: Pili, Roberto. Indiana University Simon Cancer Center; Estados Unidos Fil: Pollard, Jeffrey W.. University of Edinburgh; Reino Unido Fil: Post, Mark J.. Maastricht University; Países Bajos Fil: Quax, Paul H. A.. Einthoven Laboratory for Experimental Vascular Medicine; Países Bajos Fil: Rabinovich, Gabriel Adrián. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; Argentina Fil: Raica, Marius. Victor Babes University of Medicine and Pharmacy; Rumania Fil: Randi, Anna M.. Imperial College London; Reino Unido Fil: Ribatti, Domenico. Università degli Studi di Bari; Italia Fil: Ruegg, Curzio. University of Fribourg; Suiza Fil: Schlingemann, Reinier O.. University of Amsterdam; Países Bajos Fil: Schulte Merker, Stefan. Institute of Cardiovascular Organogenesis and Regeneration; Alemania Fil: Smith, Lois E. H.. Harvard Medical School; Estados Unidos Fil: Song, Jonathan W.. Ohio State University; Estados Unidos Fil: Stacker, Steven A.. University of Melbourne; Australia Fil: Stalin, Jimmy. Institute of Cardiovascular Organogenesis and Regeneration; Alemania Fil: Stratman, Amber N.. National Institutes of Health; Estados Unidos Fil: Van de Velde, Maureen. University of Liège; Bélgica Fil: van Hinsbergh, Victor W. M.. Universida de Lausanne; Suiza Fil: Vermeulen, Peter B.. HistoGeneX; Bélgica. University of Antwerp; Bélgica Fil: Waltenberger, Johannes. University of Münster; Alemania Fil: Weinstein, Brant M.. National Institutes of Health; Estados Unidos Fil: Xin, Hong. University of California at San Diego; Estados Unidos Fil: Yetkin Arik, Bahar. University of Amsterdam; Países Bajos Fil: Yla Herttuala, Seppo. University of Eastern Finland; Finlandia Fil: Yoder, Mervin C.. Indiana University; Estados Unidos Fil: Griffioen, Arjan W.. VU University Medical Center; Países Bajos

Aim To characterize baby hamster kidney fibroblast (BHK 21/C13) cells and test the effects of antibodies against podoplanin and disodium cromolyn on BHK 21/C13 cell line-derived tumors grown on chick embryo chorioallantoic membrane (CAM). Material and methods BHK 21/C13 cell-derived fibrosarcomas developed in hamsters were implanted on CAM and treated with anti-podoplanin antibodies and disodium cromolyn. BHK 21/C13 cell immunophenotype was assessed. Results Fibrosarcoma cells were positive for vimentin, CD117, smooth muscle actin, vascular endothelial growth factor epidermal growth factor receptor, homebox prospero gene 1 and negative for platelet-derived growth factor B, neuron-specific enolase, S100, CD34, Ewing sarcoma and podoplanin. CAM-grown fibrosarcomas were highly sensitive to disodium cromolyn and anti-podoplanin antibodies. Conclusion Immunophenotyping BHK 21/C13 cells and their response to drugs represent the first step in revealing cell line utility and a reliable tool for experimental cancer research.