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Both adipose-derived stem cells (ADSCs) and fat grafting promote burn wound healing, but whether adipogen-derived cells using various inducers such as 3-isobutyl-1-methylxanthine (IBMX) and insulin affect wound healing is unknown. Herein, ADSC-differentiated adipogenic lineages were used in rat burn wounds to evaluate wound healing potential. ADSCs were cultivated using six different adipogenic differentiation conditions (IBMX ± insulin, IBMX for 5 days, high and low Dulbecco's modified Eagle's medium) and in vitro morphological changes and cell proliferations during adipogenic differentiation were recorded. Intermediate burn wounds were inflicted in 15 Wistar male rats. Afterwards, the rats were divided into five groups for subcutaneous injections under the wounds: control; ADSCs; differentiated adipocytes (-IBMX+INSULIN and +IBMX[D1-5]+INSULIN) and fat prepared by Coleman technique. Macroscopic changes and histology were documented for 3 weeks. Repeated measures analysis of variance was performed to analyze cell growth and wound healing with a statistical level set of P < .05. Induction cocktails significantly reduced proliferation and induced lipid droplet accumulation. Conditioning without insulin induced the least lipid accumulation, while discontinuing IBMX generated larger adipocytes (P < .001). Adipogenic differentiated ADSCs had similar wound healing abilities with ADSC and fat injections, but differentiated adipocytes (+IBMX[D1-5]+INSULIN) and fat grafting accelerated the early healing process relative to ADSC (P < .001). Reduced fibrosis and mild inflammatory infiltration limited to superficial dermis were observed in +IBMX(D1-5)+INSULIN and fat injection groups, while those reactions were mild to moderate in ADSC group. Differentiated adipocytes achieve similar wound healing results compared with ADSC and fat injections, but differentiated adipocytes (+IBMX[D1-5]+INSULIN) and fat grafting accelerate early healing relative to ADSC.

Magnetic hyperthermia is a highly motivating subject for the medical community focused on cancer therapy methods.

Obiectiv: Limfocitele T se constituie în factori celulari importanţi ai răspunsului imun. Leucemia limfocitară cronică B se caracterizează prin defecte ale sistemului imun. Studiul nostru îşi propune caracterizarea diferitelor subseturi maturative şi funcţionale limfoide T şi NK la pacienţi aflaţi în stadii distincte de boală. Material şi metoda: au fost utilizate celule mononucleate separate în gradient de densitate de la 43 de pacienţi cu LLC-B (16 femei, 27 bărbaţi, vârsta 68±10 ani, 8 în Rai0, 22 în Rai1/2 şi 13 în Rai3/4) în vederea analizei prin citometrie in flux multiparametrică. Au fost identificate şi comparate între diferite stadii Rai, subseturile limfoide T CD4+ (naive, memorie centrală, memorie periferică/efector, reglatorii, foliculare-TFH, CXCR3 şi/sau CCR4+), CD8+ (naive+memorie, efectorii, senescente) şi NK (CD57 şi/sau CD94+). Rezultate: Limfocitele T totale circulante se găsesc în numere crescute la pacienţi în stadii avansate ale bolii. Majoritatea limfocitelor T CD4+ sunt celule ce au întâlnit antigenul, cu subseturi crescute de tip reglator, folicular şi CXCR3+ (asociat profilului Th1). Expansiunea limfocitelor citotoxice CD8+ este corelată cu un subset diferenţiat terminal, senescent, CD57+. În ceea ce priveşte subseturile limfocitare NK, nu s-au identificat diferente semnificative între diferite stadii ale bolii. Concluzii: Comportamentul leucemiei limfocitare cronice B pare a fi asociat cu o creştere în valori absolute a limfocitelor T CD4+ reglatorii şi foliculare circulante, dar şi cu expansiunea limfocitelor T citotoxice senescente. Modularea răspunsului T la pacienţii cu LLC-B joacă un rol important în evoluţia bolii şi se constituie în eveniment cheie al compensarii imunodeficientei caracteristice stadiilor avansate ale bolii.

The renin-angiotensin-aldosterone system (RAAS) is a hormonal system which contributes to the regulation of both arterial pressure and extra cellular fluids volume. The increase of RAAS, especially at angiotensin II (Ang II) level, affects the target organs and increases the risk of cardio-vascular issues, by increasing arterial pressure and through the direct effect of Ang II upon the vascular endothelium and the renal and cardiac tissue. Ang II reduces the renal capacity of sodium excretion and initiates a set of events which increase arterial pressure. Increase of arterial pressure is necessary for re-establishing sodium excretion, being realized by the pressure-natriuresis relationship. Arterial hypertension affects the target organs (heart, kidneys) and leads to a vicious circle which contributes to maintaining a high arterial pressure. Materials and Method: Male Wistar rats subjected on a normal diet, received either a sham operation (n=9) or continuous angiotensin II (Ang II) infusion (300ng/kgc/ min) subcutaneously, via mini pumps. Water ingestion and systolic blood pressure were measured for 14 days, after which the animals were sacrificed under anesthesia with ketamin, and the xylasin body weight, water ingestion, heart mass, right and left ventricular mass, right and left kidney mass were measured. Results: After 14 days of Ang II infusion, bodily weight decreased, systolic blood pressure increased, heart and left ventricular mass indexed to body weight were significantly enhanced compared with the sham group, and kidneys mass indexed to body weight was similar in the two groups.

We have tried to establish a possible connection between breast cancer and a series of individual factors: age of the menopause, woman's age at the first delivery, number of pregnancies and the number of born children, the age when the first symptom appeared (mammary tumor), environment, hereditary and collateral antecedents. The data have been analysed with the help of statistics methods and electronic computer programmes used to solve most complex problems. The conclusions regarding some risk factors, dependent on the statistic behaviour of the elements analysed, indicate a maximum frequency of the neoplasm incidence immediately after menopause, an increased risk for the aged primiparas and the protecting role of an early pregnancy, as well as a bimodal aspect of the frequency of this disease depending on the age at which the first symptom appears and the age of the menopause, with two maximums, the principal one placed within the first ten years after the setting in of menopause, and the second one occuring at about twenty years after menopause.

Purpose To assess the value of aggressive loco-regional surgery in desperate situations with locally advanced breast cancer. Methods In this study there were considered 31 patients with locally advanced breast cancer who underwent surgery in a 5-year period. 10 of them received 3 cycles of chemotherapy and radiotherapy before surgical intervention; the rest of the 21 patients had systemic or local contra-indications for neo-adjuvant therapy. We describe clinical aspects and technical difficulties. Surgical intervention focused on tumour removal and lymph node dissection. Skin defect was covered with flaps according to the Mortimer-Show technique. Postoperatively, the outcome was influenced in a favourable way by the use of Detralex, a micronized flavonoid; all but 2 patients received chemotherapy and locoregional radiotherapy. Results 25 patients survived free of disease; from 6 patients who suffered recurrence, 2 are still living and 4 have died. Conclusions In some forms of locally advanced breast cancer, aggressive surgery offers improvement in the quality of life and increases survival.

În acest studiu am analizat modalitatea optimă de poziţionare a cut off-ului în cadrul analizei cantitative a infiltratului inflamator în cancerul mamar cu ajutorul sistemului HistoQuest. Am utilizat specimene de ţesut mamar tumoral, care au fost marcate imunohistochimic cu markerii CD68 (pentru macrofage) şi CD8 (pentru limfocitul T citotoxic) şi ulterior analizate automat pe regiunile IT (intratumoral), PT (peritumoral) şi FI (front de invazie) cu ajutorul sistemului soft HistoQuest. Pentru evidenţierea populaţiei de celule pozitive pe histograme şi scatergrame, este necesară poziţionarea cut off-ului (discriminator). Pentru optimizarea tehnicii, am comparat 5 modalităţi de poziţionare a cut off-ului. Rezultatele obţinute statistic pentru markerul CD8 pentru toate cele 5 tipuri de cutoff aplicate pe regiunile IT, PT şi FI nu au înregistrat diferenţe semnificative statistic (p < 0,05). Pentru markerul CD68 s-au înregistrat diferenţe semnificative statistic (p

Introduction. Metabolic syndrome (MS) is a cluster of distinct metabolic alterations with an increased cardiovascular risk. Peroxisome Proliferator-Activated Receptor - Alpha (PPARα), member of the nuclear receptor superfamily of transcription factors, is critically involved in the management of lipid metabolism during homeostasis or inflammatory stresses in various cell types and represents one of the therapeutic targets in MS. We analysed the PPARα expression in leukocytes of pacients with MS, in order to address PPARα involvement in these group of diseases. Material and method. Our study included 57 adult patients recruited under informed voluntary consent, investigated in order to establish whether they present MS, according to International Diabetes Federation (IDF) European guidelines and grouped in 2 lots: the MS Lot (26 patients) and control group, non-MS Lot (31 subjects). Common clinical and laboratory parameters targeted in MS evaluation were determined for all the studied cases. The expression levels of 2 molecules, PPARα and CD36 were evaluated in various circulating leukocyte populations of these patients by an optimized flow cytometry method. Statistic analysis clarifying the significance of value differences for various parameters measured was performed under SPSS and simple statistical tests (Pearson, t-Student, Chi -test). Results and discussion. The fluorescence staining for PPARα were significantly dimmer when comparing the cellular expression in eosinophils (p

Traffic of tumorand normal cells through the peripheral blood (PB) of patients with B-cell chronic lymphocytic leukemia (B-CLL) to the lymph nodes (LN) or spleen/ liver sites is governed by specific changes in surface and intracellular molecule expression. The study aims to investigate the potential association between different lymphocyte subsets, chemokine receptors or genetic alterations and specific clinical signs in a group of B-CLL patients. Forty-three patients were included in the study. The expression of CCR7, CXCR5, CXCR3, CCR4, CD3, CD4, CD8, CD27, CD28, CD45RA, CD25, CD127, CD38 was tested by multiparameter flow cytometry. Genetic alterations were determined by MLPA. We found increased frequency of CD38+ B-CLL cells directly correlated with the presence of LN>5cm. CXCR5 and CCR7 are homogenously expressed by monoclonal B-CLL cells. CCR4+ B-CLL cell frequency is found to be lower in the PB of patients presenting particular LN involvement. Heterogeneous and complex genetic alterations were found and only the presence of trisomy 12 associated with less frequent axillary LN involvement. We also report a significant increase in the frequency of total T cells and T cell subsets (effectorand central memory CD4+ T cells, regulatory T cells, follicular T helper cells, distinct functional CD8+ T cells) with the occurrence of specific clinical manifestations. Chemokine receptor expression on circulating CD4+ T cell subsets was augmented in connection to some specific LN locations. Consequently, clinical manifestations in B-CLL are linked to both, factors intrinsic to the monoclonal B cells, and external influences coming from the microenvironment.

Background and Aim: Breast cancer shows poor prognosis when tied with chronic inflammation mediated by tumor associated macrophages (TAM). However, TAM’s multiple role, localization and prognosis are still debatable. Our study aims to quantify macrophage area and compare defined tumor morphological compartments: tumor associated stroma (TAS), tumor parenchyma and invasive front. Material and Methods: Automated digital macrophage area quantification was standardized using a tissue cytometry system (TissueFAXS) on slides originated from 50 patients with invasive breast carcinoma. Macrophages were stained using immunohistochemistry with anti-CD68 marker. The three tumor compartments were manually delineated on the digital slides. Results: Macrophage area was 3 times higher in lymph node-negative metastasis patients (pN0 group) when compared to lymph node-positive group in tumor and invasive front compartments. Highly significant strong correlations were found between the three compartments only in pN0 group (i.e. R >0.90, P

Recent studies are focusing on complementary prognostic and predictive markers that could complete the predictive TNM staging and one of the most promising directions is the study of tumor immune infiltrates.Our 2-year retrospective study includes resection specimens from the primary tumors of 23 patients presenting to our clinic for a local or a distant relapse after colon or rectal cancer. From every primary tumor specimen we obtained immunohistochemically stained slides in order to assess cd3, cd4, cd8, cd45ro and cd68 infiltrates. Digital analysis assessed the density and percentage of positively stained cells in the normal peritumoral tissue, invasive margin and center of the tumor.A small density of cd8 positive cells in the peritumoral region was strongly correlated with a longer disease-free interval (p=0.009) and the Kaplan-Meier survival analysis showed that the percentage of cd8+ T cells could be used to stratify patients in terms of relapse risk (p=0.006). We found no correlation between invasion front infiltrates and intra tumoral infiltrates and the disease-free interval.Our study concludes that cytotoxic T-cell infiltrates in the normal peritumoral tissue could be used to predict a more aggressive tumor in terms of the relapse risk.

Background/Aim. Tumour angiogenesis defined by microvessel density (MVD) is generally accepted as a prognostic factor in breast cancer. However, due to variability of measurement systems and cutoffs, it is questionable to date whether it contributes to predictive outline. Our study aims to grade vascular heterogeneity by comparing clear-cut compartments: tumour associated stroma (TAS), tumour parenchyma, and tumour invasive front.Material and Methods. Computerized vessel area measurement was performed using a tissue cytometry system (TissueFAXS) on slides originated from 50 patients with breast cancer. Vessels were marked using immunohistochemistry with CD34. Regions of interest were manually defined for each tumour compartment.Results. Tumour invasive front vascular endothelia area was 2.15 times higher than that in tumour parenchyma and 4.61 times higher than that in TAS (P<0.002). Worth to mention that the lymph node negative subgroup of patients show a slight but constant increase of vessel index in all examined compartments of breast tumour.Conclusion. Whole slide digital examination and region of interest (ROI) analysis are a valuable tool in scoring angiogenesis markers and disclosing their prognostic capacity. Our study reveals compartments’ variability of vessel density inside the tumour and highlights the propensity of invasive front to associate an active process of angiogenesis with potential implications in adjuvant therapy.

Introducere. Pacienţii cu leucemie acuta limfoblastică (LAL) neonatală prezintă în mod frecvent, o tran- slocaţie particulară, t(4;ll)(q21;q23), care duce la fuziunea genei MLL (mixed lineage leukemia) de pe cromo- zomul 11 cu gena AF4 de pe cromozomul 4. Deşi în literatura de specialitate au fost descrise anterior cazuri de LAL neonatală caracterizate prin prezenţa unor produşi de fuziune MLL-AF4 cu variate puncte de ruptura, cazul prezentat în acest material este inedit, atât prin complexitatea fuzatului MLL-AF4 identificat, cât şi prin particu- larităţile sale clinico-biologice. Materiale şi metode. Intr-o probă de sânge periferic provenită de la un nou-năs- cut în vârstă de 21 de zile a fost testată, prin biologie moleculară, prezenţa genelor de fuziune: TEL-AML1, BCR-ABL(plQO). E2A-PBX1 si MLL-AF4. Fragmentul MLL-AF4 identificat, având o lungime neobişnuită, a fost purificat şi secvenţiat. Evaluarea imunofenotipică a blaştilor leucemici circulanţi s-a efectuat prin citometrie în flux, multiparametrică, utilizându-se un citometru FACSCanto-II (Becton-Dickinson). Rezultate. S-a identificat un transcript de fuziune MLL-AF4 cu o secvenţă particulară (fuziunea exonului 12 de pe gena MLL cu exonul 4 de pe gena AF4). Evaluarea imunofenotipică a evidenţiat, de asemenea, o evoluţie particulară: la diagnostic s-a des- cris o clonă malignă cu fenotip de precursor limfoid B, în timp ce la recădere s-a evidenţiat expansiunea predominantă a unei clone monocitoide. Prezenţa aceluiaşi transcript MLL-AF4(el2-e4) a fost evidenţiată şi la recădere, în absenţa oricăror altor fuzaţi caracteristici pentru lineajul mieloid. Concluzie. Considerăm că particularităţile punctelor de ruptură care au dus la generarea rearanjamentului MLL-AF4 în acest caz sunt inedite, nemaifiind menţionate anterior în literatura de specialitate. Detecţia sa timpurie are potenţial predictiv pentru switch-ul imunofenotipic şi implicaţii în optimizarea strategiilor terapeutice.

It has been known for quite a lot of time that tumours are cellular aggregates of different cells, mainly malignant cells but also immune cells--of which the most well-known are the tumour infiltrating lymphocytes (TIL) and the tumour associated macrophages (TAM). By observing hematoxylin--eosin stained or immunohistochemical stained slides belonging to different areas of the tumour it is clear that there are clusters of malignant cells within the tumour itself that seem to behave differently from the rest of the tumour. Another fact is that different areas of the tumour contain different inflammatory cells which may promote carcinogenesis or may help to confine it. Whereas different immune cells can be recognised by using immunohistochemistry, a satisfactory characterization of the molecular characteristics of the malignant clusters of the tumour cannot be made without further use of different molecular techniques such as different PCR techniques or microarray methods. Laser microdissection thus comes as a valuable aid in choosing exactly which cells will be analyzed further on.Laser microdissection is based on using the energy of a focused laser beam to cut through the thickness of the tissue that is placed on a microscope slide in order to obtain cell samples previously selected by the pathologist through special software.To be able to cut the cells that you want and analyze them further without having them contaminated with other cells means that you can get more insight into the progression of the mutations that occur in these malignant cells, mutations that cause them to become more aggressive or multidrug-resistant. This could in time lead to the discovery of new molecular targets for cancer therapy.

Bone marrow mesenchymal stem cells are important for both research and clinical purpose. A number of culture methods for these cells are available on the market, many of them consisting of specialized growing media in combination with growth factors. Our goal was to optimize a less expensive culture method for bone marrow mesenchymal cells.Eight samples of bone marrow aspirates from patients were used. Out these 8 samples 2 were from healthy people, 3 from chronic granulocytic leukemia patients, 2 from multiple myeloma patients and 2 from patients with myelodysplastic syndrome. Bone aspirates from healthy people were used to optimize the culture method and the rest were used for testing the optimized method. Two methods were tried: 1. Cell culture starting from whole bone marrow, 2) cell culture after bone marrow separation in density gradient with Histopaque.Cell culture starting from whole bone marrow gives better yields for mesenchymal stem cells than methods which include gradient density separation of mononuclear cells with Ficoll-Histopaque.We have optimised a less expensive cell culture method for bone marrow mesenchymal cells.

There are almost no data concerning the involvement of endoplasmic reticulum stress (Ca2+ fluxes) in the apoptosis of the pro-B cell type Ba/F3. Thus, we aimed the characterization of thapsigargin-induced effects on Ba/F3 cells in vitro.For some experiments Ba/F3 cells were treated with 1 microM thapsigargin for 24 hours. To compare, we used as positive control the effects of valinomycin 10 microM. The negative control Ba/F3 cells received no treatment for 24 hours. After that, all batches of Ba/F3 cells were incubated in the presence of 1 mimcroM JC-1 (Sigma-Aldrich) at 37 degrees C for 30 minutes.From the normal Ba/F3 JC-1-control cells (20.000 events gated by flow cytometry) 77.61 +/- 2.90% are associating high and 22.39 = 2.90% lower mitochondrial membrane potential. In the case of thapsigargin, 75.49 +/- 1.78% of the Ba/F3 cells are associating high and 24.51 +/- 1.78% lower mitochondrial membrane potential.While mitochondrial permeability transition pore (MPTP) opening necessarily correlates with a loss of mitochondrial membrane potential (Psi(mt)), a decrease in Psi(mt) does not necessarily indicate MPTP opening. Also, endoplasmic reticulum stress regulation of high cytosolic calcium levels by thapsigargin may prompt the opening of the MPTP without necessarily triggering irreversible pore activation or subsequent apoptogenic factors in Ba/F3 cells.

Myelodysplastic syndromes (MDSs) are clonal disorders of hematopoietic stem cells (HSCs) characterized by ineffective hematopoiesis. MDSs are responsible for 1 or several peripheral cytopenias. The evidence accumulated in recent years demonstrates that in addition to HSC defects, a particular role is also played by stromal microenvironment dysfunctions, which mediate the direct contact with hematopoietic precursor cells (HPCs). These interactions help regulate different adhesion-related processes, such as progenitor cell proliferation, apoptosis, clonogenic growth, and maintenance in in vitro cultures. As previously reported, these interactions are responsible for altering the microenvironment in MDS. Herein, we present a novel selection protocol for obtaining a standards-compliant mesenchymal stromal cell (MSC) preparation. This method allowed us to comparatively analyze 2 subpopulations of bone marrow MSCs (BM-MSCs) in terms of their adhesion profiles and growth abilities: BM-MSCs selected from MDS settings and their normal counterparts. Functional assays revealed that the MSCs from MDS are intrinsically pathological, thus showing a continuous decline of proliferation and a reduced clonogenic capacity during 14 days of culture and in the absence of signals from hematopoietic cells. The MSC growth defects were significantly correlated with decreases in CD44 adhesion molecules and CD49e (α5-integrin).

Despite recent advances in understanding the immune mechanisms of cervical cancer (CC), relapse remains still an actual issue and recognition of new predictive biomarkers is essential.The purpose of this retrospective study was to investigate neo-angiogenesis in CC and its possible utility as prognostic biomarker.Paraffin-embedded tissue samples from 61 consecutive women with CC were immunostained for CD34 and E-cadherin. Statistical analysis was performed in SPSS-12 software, p0.05.Statistically significant differences between CD34 distribution among three interest tumor regions: micro-vessels density increase from central to peripheral area (chi(2), p0.05); statistically significant correlation between CD34 expression, particularly in stromal and peripheral sites, E-cadherin (Spearman r1=-0.321) and lymphatic invasion (Spearman r2=0.455) (p0.05) were reported. Overall five-year survival is clearly dependent on level and distribution of tumor angiogenesis among defined area of interest as suggested by Kaplan-Meier analysis.Angiogenesis is essential for guiding CC evolution and prognosis, particularly in squamous invasive types.

The last two decades brings many data about white adipose tissue capacity to secrete hormones, named adipokines, which could mediate the relationship between obesity and lung diseases. In this paper we presented some data about adipokines involvement on pulmonary function, with special emphasis on leptin, adiponectin, tumor necrosis factor alpha, vascular endothelial growth factor, resistin, hepatocyte growth factor, interleukin-6, angiotensinogen and apelin.

Genital cancers represent an important issue regarding women health, as they produce a large number of cases, invalidities and deaths.The evaluation of individual risks for a group of women with genital neoplasia.We conducted a pilot case-control study to validate the questionnaire for the evaluation of risk factors in genital cancers, in a number of 40 female subjects (20 cases and 20 controls). We have realized the data processing using the SPSS 16 and EpiInfo 3.5.1. soft wares.For questionnaire validation we have evaluated the reproducibility, the validity, the acceptability and the practicability of the questionnaire. In order to establish the test reproducibility, we have calculated k factors for inter-investigator (k = 0.34) and intra-investigator (k = 0.72) variation. To evaluate the questionnaire validity we calculated the sensitivity (cases = 94%; controls = 97%), specificity (cases = 67%, controls = 75%), positive predictive value (cases = 94%; controls = 94%), negative predictive value (cases = 67%; controls = 86%). The risk factors most frequently identified were genital infectious in clinical records (75% comparing to 52.5% - p = 0.036), and also the presence of other pathological records in genital area (65% comparing to 22.5% - p = 0.0001).For an accurate identification of individual risk at women with genital cancers subsequently studies on greater number of subjects is necessary.

Breast and ovarian cancer are common pathologies in women, with increasing incidences worldwide. In hereditary breast and ovarian cancer (HBOC) families, a large percentage of cases are attributable to hereditary factors compatibles with dominant autosomal transmission of a major tumour suppressor gene with incomplete penetrance. Screening for BRCA1 mutations is now standard practice for HBOC cases in western world, and permits medical follow-up and genetic counselling. Over 300 BRCA1 germinal mutations are stored in the Breast Cancer Information Core (BIC) mutation database.Estimates in different countries range from 5 to 15% the BRCA1 related cases of hereditary breast cancer due to copy number changes of one or more exons of this gene. Exon deletions and amplifications will usually not be detected by sequence analysis of the complete BRCA1 gene, therefore MLPA screening is needed.Here we describe Multiplex Ligation-dependent Probe Amplification technique (MLPA) implementation for BRCA1 large genomic rearrangements.We did not detect any BRCA1 mutation by analysis of 15 HBOC recruited patients.

Leukemic cells have unique aberrant phenotypes, which permit identification of this cells at diagnose and in evolution of the disease. Signaling molecules with other cells and bone marrow stroma are part of the leukemic cells phenotype. Genetic and molecular abnormalities have the main prognostic significance and confer the leukemic cell status. The main aim of the current study is to identify correlation between recognized prognostic factors in acute myeloid leukemia (AML) patients and other phenotypic markers.Imunophenotypic analysis (BDFACS CantoII, FACSDiva Software) was performed on peripheral blood/bone marrow aspirate samples of 56 patients diagnosed with AML (9 M0, 3 M1, 10 M2, 4 M3, 28 M4/M5, 1 M6, 1 M7) between 2007-2009 in Hematology Department of "Sf. Spiridon" Hospital Iaşi. We used an extended panel of monoclonal antibodies and we determined the level of expression of cytokines receptors (IL3Ra, IL7R) and chemokines (CXCR4, CKR5).In our study, IL7R expression on AML blasts was significant correlate with low WBC count at diagnosis (p = 0.04) and with multilinear displasia (p = 0.01), high CXCR4 expression was correlate (p = 0.05) with lack of response at first induction therapy and CD123 (IL3Ra) expression was correlate with M4 FAB phenotype. Survival was negative influenced by presence of IL3R on AML blasts, but flt3 mutations, CXCR4, IL7R expression on leukemic cells, other phenotypic aberrancies did not influenced treatment response and survival in our patients population.Complete investigation of leukemic cells phenotype extended with cytokines/chemokines receptors at diagnostic is useful for correct characterization of the disease, for discover new prognostic categories and for better identification of minimal residual disease.

In this paper we want to study the changes in the expression of cell proliferation factor Ki67 in the respiratory epithelium of patients diagnosed with bronchial asthma.Twenty one patients with bronchial asthma having different degrees of severity (according to GINA 2002) have been included in our study. Fragments of bronchial mucosa were obtained through fiberbronchoscopy, prepared for histologic examination in view of immune marking with anti-Ki67 antibodies.Microscopic examination revealed a progressive increase in the number of Ki-67 cells in the respiratory epithelium corresponding to the severity of asthma (following FEV1 parameters).The increase in the cell proliferation in the respiratory epithelium represents one of the mechanisms that can help in rebuilding the epithelial structure destroyed by the chronic irritation.

Despite recent advances in the immune mechanisms of cervical cancer (CC), the relapse still remains an actual issue and recognition of new predictive biomarkers is essential.The purpose of this retrospective study was to investigate possible differences in the primary, in situ, cellular immune response between cervical carcinoma with and without relapse.Paraffin-embedded tissue samples from 61 consecutive women with CC (34 with and 27 without relapse) were immunostained for CD3, CD20 and CD45 cells. Immune cell profile densities were further assessed, assigning scores between 0 and 3: "0" meaning the absence of inflammatory infiltrate, "1+" low, "2+" intense and "3+" intense infiltrate with lymphoid follicles. Statistical analysis was performed in SPSS-13 software, p0.05.Statistically significant intra- and peri-tumoral low numbers of several immune cell subtypes are strongly associated with relapse of disease within three and five years in patients with CC (p0.05); moreover, statistical significant correlations between immune cells and both free survival (CD3: r=0.382; CD20: r=0.404; CD45: r=0.376) and relapse (CD3: r=-0.408; CD20: r=-0.355; CD45: r=-0.354) have been demonstrated. Only CD3 was reported as predictive biomarker of relapse in CC (ANOVA, t-Student, p0.05).Major differences in the cellular immune response among patients with cervical cancer with and without relapse within three and five years have been demonstrated. CD3 may be used as potential prognostic biomarkers, whereas the results are promising for adjuvant immunotherapy.

Despite recent advances in the immune mechanisms of cervical cancer (CC) and complex management opportunities, relapse remains still an actual issue. While predictive factors are required, current research is directed towards proliferation and tumor aggressiveness biomarkers as potential negative factors in CC. The main objectives were to assess tumor proliferation and invasiveness biomarkers (Ki-67, E-cadherin) and to identify potential correlation between biomarkers and classic prognostic factors in CC. Radical hysterectomy specimens from 61 consecutive CC were immunohistochemically investigated for Ki-67 and E-cadherin. Nuclear immunostaining for Ki-67 proliferation index was assigned scores 1 to 3, "+" meaning low (10-30%), "++" moderate (30-50%), "+++" high-proliferation rate (50%); cell membrane E-cadherin staining was either negative or positive. Statistical analysis was performed in SPSS-13 software, p0.05.no significant correlation between Ki-67 and classical prognostic factors (p0.05) was reported; however, in relapsed CC, Ki-67 correlates with tumor grading (r=0.386, p0.05). Significant correlation between E-cadherin and tumor size (r=-0.280, p=0.029), relapse (r=-0.386, p=0.002) and disease free survival (r=0.374, p=0.003) were demonstrated. Indirect statistically significant moderate correlation between Ki-67 and E-cadherin (r=-0.461, p0.00001) was shown, mainly in invasive squamous CC (r=-0.549, p=0.0001), stage IB (r=-0.578, p=0.009), IIB (r=-0.585, p=0.003), relapsed CC (r=-0.525, p0.01), HPV-infection (r=-0.504, p=0.033).CC aggressiveness, particularly in invasive squamous carcinoma, either 16 or 18 HPV-positive cases, FIGO stage IB and IIB, and cases with relapse, depends on two pivotal factors, tumor proliferation rate (Ki-67) and tumor invasiveness (E-cadherin).

Recurrent abortions, defined as more than three consecutive pregnancy losses, are associated with genetic, anatomic and endocrine causes. Whenever investigations fail to detect a cause, immunological dysfunctions are incriminated. The paper shortly reviews the main causes, investigation protocols and immune therapeutic attempts that have been made. Active immunization with allogenic leukocytes or trophoblastic membranes, passive immunization with intravenous immunoglobulins and immunomodulation with glucocorticoids in unexplained recurrent abortions generated contradictory results and was banned by reproductive immunology scientists as empirical, risky for the mother and harmful for the fetus. Confronted with desperate couples, clinicians use the immunotherapy with paternal lymphocytes or intravenous immunoglobulins in healthy women with unexplained recurrent abortions or in cases where accepted therapies failed.

The immunohistochemistry represents a very effective tool for describing the biological phenomena that characterize the primary or secondary neoplastic development. By using a complex immuno-morphologic approach, our study aims to develop a characterization of the metastatic liver microenvironment, based on the following features: similarities and differences in the proliferation activity, particular aspects of vascularity and immune response, the cellular dialogue generated at the level of the parenchyma - neoplastic tissue interface, the structure of the associated stroma.Ten cases of liver metastases have been studied, including five treated exclusively by surgery (group 1) and five with recurrent metastases occurred after a previous treatment by thermonecrosis (group 2). The cases were investigated by routine histopathological exam and by immunohistochemistry. The choice of the antibodies was motivated by the follow-up of the parameter classes presented below: (i) proliferation/apoptosis; (ii) structure; (iii) angiogenesis and lymphangiogenesis; (iv) immunological reactivity; (v) cytokines.The elements defining the histopathologic and immunohistochemical patterns allowed a comparative evaluation of the metastasis models considered for the two groups of studied cases. The analysis of proliferation / apoptosis parameters proved a high aggressiveness in tumor proliferation for both types of metastases. Recurrent metastases showed an increased angiogenesis and a moderate lymphangiogenesis, as well as a massive matrix reshaping. The evaluation of the inflammatory infiltrate suggested that the development of the liver metastases is not accompanied by significant immunologic rejection, but it rather induces the tissue remodeling at the invasion border level. The recurrence corresponds to a typical interval of 6 months or longer, and is associated with intense fibrogenesis, angiogenesis, and inconsistent inflammatory infiltrate.Therefore, the efficiency of the thermonecrosis procedure with water vapors seems to drastically depend on the complete necrosis of the target tumor, plus the adjacent cells of the parenchyma in a neighborhood of some millimeters. Moreover, the therapy must include anti-angiogenic resources as well as inflammation modulators, since the vascular supply and the leukocytic activity support the metastasis development.

Aiming to detect reliable markers indicating protection from or susceptibility to tuberculosis infection, we investigated both Th1/Th2 cytokines and total IgE plasma levels in health care workers occupationally exposed to M. tuberculosis, in patients with pulmonary tuberculosis and in healthy persons.The study groups have included 15 health care workers in close contact with TB patients, patients with active pulmonary tuberculosis at diagnosis and after treatment (12 advanced and 10 moderate TB, of which 6 had also pleurisy) and 20 healthy volunteers. Peripheral blood mononuclear cells (PBMC) were stimulated with PPD for 7 days and the release of six cytokines (IL-2, IFN-gamma, TNFalpha, IL-4, IL-5, IL-10) was simultaneously quantified by cytometric bead array (CBA) in culture supernatants. The same method was used to determine the cytokine level in plasma and pleural effusions from TB patients. Six neoplastic pleurisies were included in this investigation as a control group. Total plasma IgE level was measured by chemiluminescence technique.Plasma and pleural fluid cytokine analysis at the outset of tuberculosis disease reflect the same Th1 response dominated by IFN-gamma. In opposition, very low IFN-gamma levels were recorded in neoplastic pleural fluids. Both types of cytokines (Th1 and Th2) were secreted in response to in vitro PPD stimulation of PBMCs and had different evolution in moderate and advanced TB. Thus, IFN-gamma, TNFalpha, IL-4, and IL5 production after 6 months-treatment decreased in moderate TB and increased in severe disease (p0.05). Moreover, total IgE plasma levels were higher than the normal value (87 IU/ml) in health care workers and significant amounts were recorded in patients, especially in advanced TB after 6 months of treatment (p = 0.00).Our results confirm that the quantification of IFNă could be a good marker for the diagnosis of TB pleural effusions but raised the question whether plasma IgE levels might be a reliable marker indicating the transition to disease. Further studies are needed to understand the complex interaction between pro- and anti-inflammatory cytokines that might play an Key words: TUBERCULOSIS, CYTOKINES

Developing countries like Romania have a high incidence of tuberculosis. Literature data suggest that people in these countries have an important Th2-type immune background which prevents a protective Th1 response of the host against Mycobacterium tuberculosis. Our study is the first attempt in Romania to identify cytokine patterns in active tuberculosis. The study included 15 patients with active pulmonary tuberculosis and 11 healthy volunteers. Peripheral blood mononuclear cells (PBMC), stained with carboxyfluoresceine-diacetate-succinimidylester (CFSE), were incubated for 7 days with purified protein derivate (PPD) or with medium alone. Intracellular synthesis of IFNgamma and IL-4 in proliferated (CFSE(low)) T cells was detected by flowcytometry. The results showed that both Th1 (IFNgamma) and Th2 (IL-4) cytokines are produced in response to in vitro PPD-stimulation of PBMC from pulmonary tuberculosis patients and healthy controls. Moreover, the proportion between IFNgamma and IL-4 is tilted in favour of IL-4 in PPD-activated (CD3+ CFSE(low)) cells from healthy persons, who did not develop active tuberculosis during the two-year study time interval. This predominance of Th2 effectors points to the need to further investigate the role of IL-4 in the M. tuberculosis infection.

This study was designed to detect the presence of the p27(Kip1) protein expression in breast carcinomas and corresponding axillary lymph node metastases, as well as any potential correlation of p27(Kip1) with factors of tumor progression and prognosis (estrogen receptors-ER, progesterone receptors-PR, c-erbB-2, p53).44 primary breast cancers and positive axillary lymph nodes from 24 cases were examined by immunohistochemistry.19 out of 44 (43.18%) tumor specimens expressed the p27 protein, whereas 25 (56.82%) specimens expressed it only at low levels or were negative. Absence of p27 correlated significantly (p0.05) with a negative status for ER and PR receptors. The presence of p27 protein in primary tumors was always associated with positive expression in the corresponding nodal metastases.Our results indicate a correlation between lack or low levels of p27 protein expression and the absence of hormone receptors. The p27 phenotype is preserved from the primary breast tumors to the corresponding axillary lymph node metastases.

Malignant transformation is the result of genetic events, translated into sequential acquisitions of multiple abnormalities in the control of cellular growth and cell cycle regulation. We determined the expression of the estrogen receptor (ER), progesterone receptor (PR), c-erbB-2 and p53 gene products in a patient with mixed (ductal and lobular) invasive breast carcinoma bearing different coexisting lesions. The purpose of the study was to establish a possible correlation between the expression pattern for these molecules and the histological appearance of the breast tumor. Our results showed no positivity for ER. PR expression was restricted to normal epithelium, simple hyperplasia and in situ carcinoma. c-erbB-2 was detected in all lesions with the exception of normal epithelium and immunostaining for p53 was found positive only in in situ and invasive carcinoma. These findings support the hypothesis of tumorigenesis as a multistep process and as a sum of changes, each representing an advantageous acquisition for the malignant cells' behavior. The loss of hormone receptors' expression occurred as an early event in this case, while the p53 mutations were found only in more advanced neoplastic lesions.

The aim of this study was to assess breast cancer incidence, diagnostic possibilities in clinical treatable stages and the prognosis in a series of breast cancer patients repeatedly treated in the same unit, by the same team. Diagnostic difficulties in the early stage of the contralateral cancer constitute a proven reality even in our cases. The detection of the second metachronous breast tumor was done and evaluated mostly in an more advanced stage than the initial tumours. This might be explained by the delayed visit to the doctor. Most BBC were lobular carcinoma. Adjuvant therapy--chemotherapy, antiestrogens, ovariectomy--do not prevent the appearance of the second tumoral site.

The heterogeneity of the evolution of breast cancer complicates patient management. The use of vascular markers as prognostic factors is a new and promising tool in medical oncology. Research data of the current decade demonstrate that angiogenesis plays substantial role in growth and spread of malignant tumor. At present, immunohistochemical determination of intratumoral microvascular density represents one of the more promising new prognostic indicators in breast cancer that needs to be further investigated to identify and standardize the method of choice to be tested in prospective clinical studies. Consequently markers of angiogenic activity have receiving increasing attention. By analyzing the evolution of 209 cases of breast cancer enrolled in a prospective study reaching the 5th year of follow up, we provide herein data supporting that Factor VIII and CD34 could be reliable markers for prognosis of DFS. Tumors with lower expression of Factor VIII and CD34 have a better prognostic and lower potential metastatic. The Factor VIII comparative with CD34 represents a more faithful prognostic marker. Angiogenesis markers have also become a putative therapeutic target.

HLA antigens are involved in inducing either susceptibility or resistance to different diseases. Many studies reported various associations between HLA antigens and tuberculosis, depending on race, ethnic group and geographic area.Our purpose was to identify HLA class I antigens inducing susceptibility to tuberculosis in population from North-Eastern Romania.The study group consisted of 50 tuberculosis patients and the control group included 90 healthy people. HLA-A and HLA-B antigens were determined using the CDC-NIH (complement-dependent-cytotoxicity-National Institute of Health) assay. A comparison was made between the frequency of HLA antigens expression in the two studied groups.HLA-B18 and HLA-A29(19) were expressed more frequently in tuberculosis patients. The difference was statistically significant only for HLA-B18 antigen. HLA-B7 and -B61(40) antigens were expressed with statistically significant higher frequency in controls compared to tuberculosis patients. The frequency of other HLA-A and HLA-B antigens was either comparable in the two groups or without statistical significance. CONCLUSIONS We found a positive association between HLA-B18 antigen and tuberculosis, while HLA-B7 and HLA-B61(40) antigens seem to protect against the disease.

We have investigated the cellular and serum CK18 in 26 non-treated primary ductal invasive breast carcinomas. The soluble CK18 (TPS) was detected by chemiluminescent assay, and the cellular CK18 and PCNA expression by immunocytochemistry. Flow-cytometry was used to estimate the amount of DNA in malignant cells. There was a significant correlation between soluble CK18 and the pre-menopausal status (p0.05), characterized in our group by a PCNA estimated low proliferation index. We have also found a significant correlation between soluble CK18 and the DNA index (p0.01). The intracellular CK18 has correlated with the PCNA expression (p0.05), while no correlation could be found between cellular and serum CK18. The values of soluble CK18 may offer information about the treatment-induced cell death, if monitored, while isolated measurements should be interpreted cautiously. Elevated levels of serum CK18 in non-treated carcinomas may rather reflect a high tumor turn-over or perhaps a more intensive tumor cell killing.

Angiogenesis represents an essential event required by tumors to support their growth. The aim of our study was to investigate the presence of neovascularization in 44 primary breast carcinomas and in 24 axillary lymph nodes metastases and to establish a possible correlation between the presence of tumor angiogenesis, some clinical and pathological features of the cases and the expression of p53, an important cell cycle regulator. To identify the new blood vessels, we used immunohistochemistry for von Willebrand factor, a marker of the endothelial cells. The results showed that 77.27% of the primary breast carcinomas and 75% of the lymph nodes metastases are positive for von Willebrand factor and this positivity is significantly correlated (p0.05) with the expression of p53, supporting the idea that angiogenesis is a marker for tumor aggressiveness and p53 could be involved in this process.

Fas (CD95/APO-1) and its natural ligand, FasL, are molecules expressed on cellular membranes, being involved in the induction of programmed cells death or apoptosis. Recently, it has been shown that malignant cells originating from solid tumors tend to inhibit the expression of Fas, as an escape mechanism from the immune cells' attack and to express FasL, as a counterstrike mechanism against the immune effector cells. The purpose of this study was to investigate, by immunohistochemistry, the presence of Fas and FasL in 15 breast carcinomas and to establish possible associations between the expression of these molecules and the histological type and grading of the tumors. Our results showed that 7 breast tumors have lost the expression of Fas and 11 tumors were positive for the Fas-ligand expression, important arguments for the mechanisms of immune escape and tolerance induction. Furthermore, 7 of the 11 FasL+ tumors were poorly differentiated invasive ductal carcinomas, suggesting a possible association between FasL expression and tumor aggressivity.

The etiology and pathogenesis of acne vulgaris are not yet completely understood. Therefore we have investigated 5 patients with different clinical forms of disease, including the rare form of acne fulminans. Taking into consideration the four factors that are currently incriminated in the development of acne, sebaceous hypersecretion, hyperkeratosis of the pilosebaceous infundibulum, bacterial colonisation and perifollicular inflammation, we have focused our study on a set of cells involved in the chronic inflammatory process. We have evidenced by immunohistochemistry methods, using appropriate monoclonal antibodies, the presence of T lymphocytes and macrophages, while the B cells could be evidenced only in the severe forms. We were also interested to investigate the occurrence of new capillary formation, as an accompanying phenomenon of the inflammatory process. The presence and histological distribution of these cells highly supports the hypothesis that the mechanisms underlying the development of acne vulgaris belong to the Delayed Type Hypersensitivity.

Both innate and adaptive immunity against Listeria monocytogenes are essential in circumventing or fighting the disease, although there is no clear delineation of the precise role of either in listeriosis. A consistent amount of in vivo experiments have generated a complex picture over the impact of listerial infection on the hosts, and they are shortly reviewed in this paper.

The aim of the study was to identify HLA class I types associated with susceptibility to psoriasis among population of Northeast region of Romania.27 psoriatic patients and 89 controls were typed serologically for HLA-A and HLA-B lymphocyte expression using CDC-NIH (complement dependent-cytotoxicity--National Institute of Health) method. The Terasaki plates used for HLA class I typing were prepared in our laboratory using antisera of known specificities.psoriatic patients in the group of study frequently express HLA-B57 phenotype. The relative risk induced by this phenotype was highly statistically significant (p = 0.0016) while HLA-B13 was not associated with a significant risk for developing psoriasis in patients under study compared with controls (p = 0.48). Also, HLA-B27 (p = 0.96) and HLA-B44 (p = 0.99), reported by others to be associated with late psoriasis, do not meet (a particular) disease susceptibility between psoriatic patients under investigation.

The lymph nodes status is one of the most important prognostic factors in breast cancer. The routine hematoxilin-eosin staining is efficient for the metastases detection only when there is a large number of tumor cells, while a small number of metastatic cells can easily remain undetectable. For those situations, the immunohistochemistry for cytokeratins, markers of the epithelial cells, is a very sensitive method. We have investigated the cytokeratin 8 expression in 10 primary breast carcinomas and in the corresponding axillary lymph nodes, comparing with hematoxilin-eosin. The routine examination has detected axillary lymph nodes metastases in six cases, confirmed by the immunohistochemistry for cytokeratin 8. Four cases were diagnosed as negative for the axillary lymph nodes metastases by the hematoxilin-eosin staining. In all those four cases, immunohistochemistry for cytokeratin 8 has detected a small number of tumor cells, either spread in the lymph nodes tissues, either confluent as small islets.

Breast tumors growth is regulated by female sex steroid hormones. The level of the estrogen and progesterone receptors (ER and PR) expression by the malignant cells is important for the evaluation of the tumor prognostic and the benefit of a hormonal therapy. The aim of our study was to identify the expression of estrogen and progesterone receptors in primary breast tumors and in the corresponding axillary lymph nodes metastases, in 24 cases. The results showed that more than 30% of poorly differentiated breast carcinomas lost their expression of hormone receptors from the primary tumors to axillary metastases, an event which can be associated with an aggressive tumoral behaviour and resistance to hormonal therapy.

Oncogenes, the abnormal forms of proto-oncogenes, were shown to be involved in malignant transformation and in tumor progression. c-erbB2/HER2/neu is member of EGFR family and encodes the p185 protein, which functions as a tyrosine-kinase. Gene amplification and/or p185 overexpression were reported to be associated with poor prognostic in cancer. Our purpose was to investigate p185 immunohistochemical expression in breast carcinomas and in the corresponding axillary lymph nodes metastases and to identify possible correlation between p185 and other factors of poor prognostic, such as loss of hormonal receptors expression. In our study, 40.91% of cases were erbB-2 positive, p185 expression being maintained from the primary tumors to axillary metastases and associated with positive nodal status and with the absence of hormonal receptors expression (p0.05). These findings support the hypothesis the c-erbB2 is an advantageous acquisition for the aggressive behavior of the tumor cell and for its ability to invade and metastasize.

The aim of this study is to justify an individual therapeutical attitude in breast cancer, related to diversity of breast tumors, aggressiveness grade and metastatic potential. Between January 2000--December 2001, 150 patients were admitted with breast cancer (stage II and III) and underwent surgery in our department. We selected 75 cases in our study. In 51 (68%) cases the first therapeutical method was surgery, in 15 (20%) cases surgery was performed after chemotherapy, in 2 (2.66%) cases after radiotherapy and after chemotherapy and radiotherapy in 7 (9.33%) cases. We evaluated several classical factors and new immunohistochemical markers with an important value for diagnosis, prognosis and therapy: oestrogen and progesterone receptors, c-erb B2, pS2 and p53 proteins, von Willebrand factor. Several factors had a predictive role regarding the response to chemotherapy. These predictive factors will improve the histopathological diagnosis. The oncoproteins and hormonal receptors also will evaluate with more accuracy the metastatic risk and will assure a better therapy decision.

During normal pregnancy, the maternal immune response achieves a split tolerance state towards the fetal allograft, expressed by immunophenotypic profile. This may be impaired in the preterm birth. Our study aimed to analyze the immunophenotypic profiles of the peripheral and retroplacental blood in women with normal pregnancy and preterm birth. Blood leukocyte populations were assessed by flowcytometry in nine normal at term pregnancies, five preterm births and eight nonpregnant controls. Our results showed in normal pregnant women as compared to controls, a significant decreased number of lymphocytes, monocytes and B cells, while granulocytes, T-lymphocytes and the T CD8+ subsets were elevated. The CD4/CD8 ratio was significantly decreased, especially in the retroplacental blood. The preterm birth group demonstrated a significantly elevated number of lymphocytes and B cells, lower T CD8+ cells and an increased CD4/CD8 ratio, in comparison with normal pregnancies group.Normal pregnancy determines a suppressive immunophenotypic profile that is impaired in the preterm birth suggesting its immunological involvement.

The fetus was compared to an allograft because paternal antigens are also expressed. Even the analogy is not a real one, the main steps of the rejection phenomenon are relevant for the understanding of the complex materno-fetal interactions as the large granular lymphocytes are the first immune cells which invade the graft. Due to the important role of NK cells in the cell mediated immune response, the purpose of this paper was to assess the NK cell dependent cytotoxicity during normal pregnancy. The study was performed on a test group of 52 normal gravidas, in the Ist, IInd and IIIrd trimester and a control group of 20 healthy nonpregnant women with similar age and parity averages. The cytotoxic activity of the NK cells was measured in a 51Cr release assay against K 562 cell line. The results were expressed as the cytotoxicity index (%) and showed a statistically significant decrease for all gravidas compared to controls, mostly for the IInd trimester. The trend of the NK cytotoxic activity during normal pregnancy suggests the deep involvement of the NK cells in the mechanisms acting at the fetomaternal interface.

The authors studies 280 rectal cancers treated in the first surgical clinic of Iaşi in the period 1981-1994. Although surgery is the main treatment of these tumors, it cannot be and it must be not used alone. Surgery must be associated with radio- and chemotherapy, because is the only way possible to increase the number of resections for rectal tumors and the survival rate. For the 263 tumors operated the resectability was 73, 5%: Milles's operation 101 (36, 7%), Dixon's operation 81 (30, 79%), other operation 91 (33, 11%). We consider that Dixon's operation is the best choice for the tumors in the stages A and B of Dukes classifications. Preoperative radiotherapy can influence the tumor volume (downstaging) and decrease local recurrences to about 10-15%. Surgery alone must be practiced only in emergencies. Postoperative chemotherapy is mandatory and it reduces local recurrences and delays metastasis evolution.