ROMANIAN JOURNAL of MORPHOLOGY and EMBRYOLOGY

Vol. 51 No. 2, 2010

1. Matrix metalloproteinases involvement in pathologic conditions

Cornelia Amalinei, Irina-Draga Caruntu, Simona Eliza Giusca, Raluca Anca Balan

Matrix metalloproteinases (MMPs) have a great variability that provides a complex intervention in pathophysiological conditions. MMPs roles in pathology may be grouped into the following main types: (1) tissue destruction, as in cancer invasion and metastasis, rheumatoid arthritis, osteoarthritis, different types of ulcers, periodontal disease, brain injury and neuroinflammatory diseases; (2) fibrosis, as in liver cirrhosis, fibrotic lung disease, otosclerosis, atherosclerosis, and multiple sclerosis; (3) weakening of matrix, as in dilated cardiomyopathy, epidermolysis bullosa, aortic aneurysm and restenotic lesions. Recent data also adds new MMPs functions in angiogenesis and apoptosis. Interesting opposite intervention in escaping mechanisms vs. antitumor defensive mechanisms had been also reported. As MMP-7 is expressed by tumor cells of epithelial and mesenchymal origin, it may be used as a biological marker of an aggressive phenotype and as a target of therapeutic intervention. MMPs play a pivotal role in the pathogenesis of arthritis, atherosclerosis, pulmonary emphysema, and endometriosis. Although MMP involvement in pathology is more than simple excessive matrix degradation, or an imbalance between them and their specific tissular inhibitors (TIMPs), MMP inhibition may be of therapeutic benefit, so synthetic MMPs inhibitors had been developed and are currently under clinical testing.

Corresponding author: Cornelia Amalinei, Associate Professor, MD, PhD, e-mail: cornelia_amalinei@yahoo.com

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2. Diagnostic and clinical significance of D2-40 expression in the normal human thymus and thymoma

M. Raica, A. Kondylis, L. Mogoanta, Svetlana Encica, Anca Maria Cimpean

Human thymus development and thymoma behavior remain elusive, in spite of many acquisitions in the field in last decades. In the present paper, we analyzed the immunohistochemical expression of D2-40 in the normal human thymus and thymoma. In both fetal and postnatal normal thymus, we found a strong expression of D2-40 in the subcapsular and cortico-medullary epithelial cells, and lack of expression in the thymus of involution. These findings support a role for podoplanin in the proliferation of some subtypes of epithelial cells of the normal thymus stroma. In thymoma, the expression of D2-40 was detected in neoplastic cells in 18 from 26 cases (69.23%). No correlation was found between D2-40 expression and histological types of thymoma, but strong correlation was noticed with tumor stage. Based on these results, it is suggested that D2-40 expression is a good predictor of invasion and can be considered as a potential target for therapy in selected cases.

Corresponding author: Marius Raica, Professor, MD, PhD, e-mail: raica@umft.ro

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3. Hepatocyte steatosis in patients infected with genotype 1 hepatitis C virus

Carmen Fierbinteanu-Braticevici, Maria Mohora, Laura Tribus, Ana Petrisor, Sanda M. Cretoiu, D. Cretoiu, R. Usvat, L. Ionita

Background: Recent findings suggest a higher prevalence of hepatic steatosis in patients with chronic hepatitis C, estimated at 50%. Both host and viral factors contribute to the development of steatosis in chronic hepatitis C. Steatosis is an initial stage, which promotes hepatic fibrosis through oxidative stress. Aim: To assess the pathogenic mechanism of genotype 1 hepatitis C virus in steatosis and to evaluate the correlation between the degree of steatosis and the level of oxidative stress. Patients and Methods: The study was carried out on 50 patients (29 males, 21 females) with genotype 1 HCV and liver biopsy proven chronic hepatitis C. Patients with other etiology of chronic liver disease were excluded. We statistically correlated the degree of steatosis with clinical (age, sex, waist circumferences) and biological parameters (alaninaminotransferase, gammaglutamyltranspeptidase - GGT, insulin, ferritin, serum viral load, oxidative stress). Insulin resistance (IR) was determined by the homeostasis model assessment (HOMA) method. The oxidative stress was estimated by serum malondialdehyde (MDA) and glutathione (GSH). Results: 27 patients presented steatosis (57%): 14 out of 29 men (48%) and 14 out of 21 women (66%); in two thirds of them, steatosis was moderate. Univariate analysis identified five parameters that significantly influenced steatosis: age >45 years, sex - female, IR (HOMA>2.5), BMI, central adiposity (as reflected by waist circumferences and high GGT-values). Multivariate analysis identified four significant parameters: sex - female, insulin resistance (HOMA>2.5), BMI>30 kg/m2 and GGT>2N. No relationship was found between steatosis and viral replication. The study demonstrated a significant correlation between steatosis and IR on the one hand and between steatosis and liver fibrosis on the other hand (p<0.05). Liver fibrosis was significant correlated with the increase levels of free radicals (MDA>250 nmol/dL). Conclusions: The pathogenic mechanism of genotype 1 HCV in steatosis is independent from viral replication and it may be linked to virus induced metabolic abnormalities such as IR. More women (66%) than men (48%) developed steatosis. Increased levels of free radicals, correlated with moderate and severe steatosis suggest the intervention of oxidative stress in determining the hepatic lesions associated with steatosis.

Corresponding author: Carmen Fierbinteanu-Braticevici, MD, PhD, e-mail: cfierbinteanu@yahoo.com

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4. Intratemporal course of the facial nerve: morphological, topographic and morphometric features

Nicoleta Maru, A. C. Cheita, Carmen Aurelia Mogoanta, B. Prejoianu

The purpose of this study is to present some morphological and morphometric aspects of the facial nerve and especially of the tympanic and mastoid segments of this nerve. The authors follow up a mesoscopic study concerning the tract (length, angulation, width) of these segments and the anatomic relations with the important structures of the middle ear. At the same time, some anatomical variations which involve the canal of the facial nerve (dehiscences, tract deviation or other anatomical deviations) are presented. To evaluate the risk of the facial nerve injury during operations for chronic otitis media with or without cholesteatoma, stapedectomy in otosclerosis, exploratory tympanotomy, tympanoplasty, canaloplasty, osteomas surgery or other otologic surgery that involve facial nerve area. The intricate course of the facial nerve through the temporal bone is of vital concern to all otologic surgeons, since it often traverses the surgical field. Therefore, authors will review the course of the facial canal through the petrosal portion of the temporal bone from the internal auditory meatus to the stylomastoid foramen, paying particular attention to its relations to adjacent structures.

Corresponding author: Nicoleta Maru, Senior Lecturer, MD, PhD, e-mail: nicoletamaru@yahoo.com

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5. The immunohistochemical expression of the p53-protein in gastric carcinomas. Correlation with clinicopathological factors and survival of patients

Daniela Lazar, Sorina Taban, I. Sporea, Alis Dema, Marioara Cornianu, Elena Lazar, A. Goldis, Iulia Ratiu, C. Vernic

Background: P53-tumor suppressor gene has an essential role in controlling cell cycle and initiating carcinogenesis. In the case of gastric cancer, the role of p53-protein accumulation as prognostic factor is controversy. Various results are due to the different methods of study regarding patients' selection, immunohistochemical techniques used and the quantifying systems for immunoreactions. Aim: Assessment of p53-immunohistochemical expression in 61 patients with gastric carcinomas and the correlation with clinicopathological factors (gender, age, location, macroscopic, and histological type, degree of tumor differentiation and TNM-stage) and patients' survival. Material and Methods: From the total number of 265 patients (186 males and 79 females) diagnosed with gastric cancer in the period 1998-2002, 61 operated patients were selected. On this group, we performed a prospective study regarding the evolution and aggressiveness of gastric cancer, on a duration of five years. Survival time was calculated from the month of the surgical intervention until the month of death or confirmation of survival, and survival rate was represented by the percentage of survivals at the end of the observed interval (in years and months). We used the monoclonal antibody DO7 that detects the wild and mutant form of p53-protein, by EnVision technique and DAB-visualization. We considered positive reaction only in the presence of brown staining of the nuclei. Results: P53-immunoreactions were positive in 25 gastric cancers (41%). We obtained positive stainings in 41.9% cases in men and 38.9% women. We found positive p53-immunoreactions in all the carcinomas developed in the upper third of the stomach (100%), in 53.3% of the corporeal tumors, 50% of the pangastric tumors; according to Lauren's classification, we noticed a significantly increased immunoreaction of p53 in the intestinal-type carcinomas. Among histological types, papillary, mucinous, anaplastic and tubular adenocarcinomas presented a relatively increased percentage of p53-positive immunoreactions. P53-positive stainings are more frequently encountered in moderate/poor differentiated carcinomas and those associated with lymphovascular invasion; according to pT- and pN-stage, we remarked a significantly increase of the number of p53-positive cases (p=0.02291 and p=0.038264). Five-year survival rate for patients with p53-positive carcinomas was significantly lower in comparison to the patients p53-negative (8% vs. 22.2%, p=0.0326). Conclusions: Immunohistochemical evaluation of p53-protein represents in our study an important prognostic factor, allowing the selection of a group of patients with an aggressive therapeutic indication, such as extensive lymphadenectomy and adjuvant chemotherapy.

Corresponding author: Daniela Lazar, Assistant, MD, PhD, e-mail: lazar_daniela@yahoo.com

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6. Enhanced chemoresistance and tumor sphere formation as a laboratory model for peritoneal micrometastasis in epithelial ovarian cancer

Olga Soritau, C. I. Tomuleasa, Emoke Pall, Piroska Virag, Eva Fischer-Fodor, V. Foris, Otilia Barbos, Corina Tatomir, G. Kacso, A. Irimie

Background and Purpose: Ovarian cancers are composed of heterogeneous cell populations, including highly proliferative immature precursors and differentiated cells that may belong to different lineages. The main reason why epithelial ovarian cancer is difficult to treat is the unusual mechanism of dissemination that involves local invasion of pelvic and abdominal organs. But, unlike many other carcinomas, initial dissemination rarely requires blood or lymph vessels. Because it has been proven that aggregates of malignant cells within the ascites of patients diagnosed with ovarian cancer represent an impediment to cure such cancers, in the present study we adopted suspension culture combined with anti-cancer regimens as a laboratory strategy for research of the initial process of peritoneal micrometastasis. Experimental Design: MLS human ovarian cancer cells were cultured in serum-free medium. Cells of passage eight were treated in combination with the anticancer agent doxorubicin at different peak plasma concentrations for 24 hours, and then maintained under suspension culture. The acquired increased aggressiveness properties was confirmed by multidrug resistance assays and by their ability to grow in an anchorage-independent manner in vitro as tumor spheroids. Results: Cells selected after chemotherapy had a increased proliferative potential, eliminated Rhodamine 123 in culture and also formed spheroids in suspension. Conclusions: Here we present direct evidence that the metastasis of human ovarian cancer may be a result of transformation and dysfunction of immature precursor cells in the ovary. Also, spheroid formation may represent a key component of chemotherapy recurrence and a better understanding of these 3D structures can contribute to the development of new treatments for metastatic carcinoma.

Corresponding author: Ciprian Tomuleasa, PhD, e-mail: ciprian.tomuleasa@gmail.com

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7. Immunohistochemical comparative study of fibrosis and biliary ductular reaction in alcoholic and viral chronic hepatitis

I. Egyed-Zsigmond, I. Jung, Ileana Egyed-Zsigmond, Gabriella Marton, Simona Gurzu, T. Mezei

Our study includes 102 cases of liver biopsy previously diagnosed with chronic alcoholic hepatitis and also B and C viral hepatitis. In these cases, we analyzed the extension of fibrosis with two different methods. First, we evaluated fibrosis with the subjective Knodell score; secondly, we used digital image analysis to achieve this. We also used immunohistochemical methods to mark those cells positive at Smooth Muscle Actin (SMA) and Glial Fibrillary Acidic Protein (GFAP). We have observed that the extension of fibrosis was most predominant in cases with B viral chronic hepatitis, while the number of cells responsible of fibrosis (stellate cells, myofibroblasts) was highest in C viral chronic hepatitis. These differences help clinician to divide patients into those who may be treated with interferon and those treatable with antiviral therapy. We observed ductular reaction (as shown by cytokeratin 7 immunostaining) within the lobular structure more frequently in alcohol related chronic hepatitis, whilst in C viral chronic hepatitis this reaction was more readily seen in portal spaces. We have concluded that patients with C viral hepatitis can benefit most from a correctly indicated hepatic biopsy since in these cases the lesions might be observed in an early and potentially curable phase.

Corresponding author: Emeric Egyed-Zsigmond, MD, PhD, e-mail: iegyed@clicknet.ro

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8. The gastric mucosa in portal hypertension: structural and ultrastructural observations

Alina Draghia, Fl. Draghia, L. Coman

The gastric mucosal lesions represent a frequent cause of hemorrhage in the portal hypertension (PHG) and in the hepatic cirrhosis. This study was undertaken to assess the structural and ultra structural modifications in the intimal lamina of the stomach in this pathology. The cells of mucosa show graded alterative transformations. In the gastric mucosa, some of the chief (enzymatic) cells present a quasi-normal histological organization; others increased alterations such as irregular and heterochromatic nuclei, fewer cytoplasm organelles, numerous clear vesicles and heterogeneous lysosomes. The parietal (oxyntic) cells show in their apical cytoplasm wide dilatations of the intracellular canalicles, vesicles, a reduced number of organelles and irregular nuclei. The enteroendocrine cells (APUD) present an increased number of granules and organelles in the cytoplasm. The connective inter-glandular tissue contains active fibroblasts, micro inflammatory zones, blood vessels with hypertrophied endothelium, irregular and dilated lumen. Subsequent to these alterations in the structures of the gastric mucosa in portal hypertension, the function of synthesis/excretion of the pepsinogen, lipase, hydrochloric acid, intrinsic factor, serotonin, etc. are affected.

Corresponding author: Alina Draghia, Assistant Professor, MD, PhD, e-mail: dfraghia@yahoo.com, dfraghia@gmail.com

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9. Clinical significance of Her2/neu overexpression in urothelial carcinomas

Aurora Alexa, Flavia Baderca, Delia Elena Zahoi, Rodica Lighezan, D. Izvernariu, M. Raica

HER2/neu is a defective transmembrane tyrosine kinase receptor, homologue to the epidermal growth factor receptor, showing overexpression in a large variety of tumor cells. There are no studies published so far regarding HER2/neu overexpression and sensitivity of the urothelial tumors of the urinary bladder to anti-HER2/neu therapy. There are a relatively high number of articles in the literature referring to HER2/neu expression in urothelial tumors of the urinary bladder, but only two of them had investigated HER2/neu expression in patients with urothelial tumors of the upper urinary tract. We have studied HER2/neu overexpression in 59 patients with urothelial carcinomas of the urinary tract by immunohistochemistry. Normal urothelium and the elements of the neighboring renal parenchyma were negative. Out of the 59 cases of urothelial carcinomas, 38 were negative (0 and +1) and 21 were positive: eight were moderately and 13 were intensely positive (+2 and +3). The percentage of positive cases was 35.59%. The negative cases were mostly well-differentiated, G1 tumors, no matter the T-tumor stage. Most of the cases were diagnosed as papillary or, rarely, infiltrative. There is no correlation between HER2/neu overexpression and the tumor stage. The same was true for the lymph node status. The expression intensity, however, was significantly correlated with the differentiation grade. Overexpression was most likely present in tumors with high differentiation grade (p<0.05).

Corresponding author: Dragos Izvernariu, Assistant Professor, MD, PhD, e-mail: dragos_izve@yahoo.com

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10. The correlation of genetic markers with anatomoclinical and histopathological forms in Hirschsprung's disease

V. Broasca, Camelia Ciobotaru, Iuliana Dimofte, Mariana Aschie, A. Pruna, Beatrice Severin

Hirschsprung's disease is a birth defect that affects about one out of 5000 newborns. It is one of the most common causes of intestinal obstruction at the babies. The objectives of this study are to evaluate the characteristics of Hirschsprung's disease in Dobrogea area, test of genetic markers in families and single cases, estimate the value of the test in the diagnosis and for evolution. We made a case-control study for the period 1995-2006 and analyzed 21 cases of Hirschsprung's disease, which were treated in the Emergency County Hospital, Constanta. The diagnostic methods comprised clinical and paraclinical examination. The chromosomal markers used in the study are represented by four categories of chromosome abnormalities: Trisomy 21, Del 10q, Del 13q, Del 17q. The molecular markers investigated by us are represented by: RET, EDNRB and EDN3. We made the correlation of genetic markers with the anatomopathological and histopathological forms, by measuring the level of association, expressed by the calculated relative risk (OR) and using the correlation index Fi. Based on data obtained from the group investigated, we found that the indices of association and correlation are consistently higher compared to DNA-markers with chromosomal markers, both for anatomopathological forms as well as histopathological. We noticed that no chromosomes markers were recorded with indices of correlation with negative values, which means that these chromosomal abnormalities are involved with a particular quota to the release of disease.

Corresponding author: Valentin Broasca, MD, e-mail: broascav@yahoo.com

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11. Neoangiogenesis in cervical cancer: focus on CD34 assessment

Codrina Ancuta, E. Ancuta, Fl. Zugun-Eloae, E. Carasevici

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. Aim: The purpose of this retrospective study was to investigate neo-angiogenesis in CC and its possible utility as prognostic biomarker. Material and Methods: 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, p<0.05. Results: Statistically significant differences between CD34 distribution among three interest tumor regions: micro-vessels density increase from central to peripheral area (chi-square, p<0.05); 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) (p<0.05) 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. Conclusions: Angiogenesis is essential for guiding CC evolution and prognosis, particularly in squamous invasive types.

Corresponding author: Codrina Ancuta, Lecturer, MD, PhD, e-mail: alexia02ro@yahoo.com

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12. Cutaneous leiomyomas and leiomyosarcomas: an immunohistochemical study with p53

A. Fernandez-Flores

Background: Cutaneous smooth muscle tumors are rare and sometimes the differential diagnosis between leiomyoma and leiomyo-sarcoma is difficult and based in very subtle criteria. We therefore tried to investigate the use of p53 in such a conundrum. This marker has rarely been reported in cutaneous leiomyomas and even in more rare occasions, in cutaneous leiomyomas. Material and Methods: We studied 30 benign cutaneous smooth muscle tumors, including angioleiomyomas, common leiomyomas and a symplastic leiomyoma, as well as four leiomyosarcomas and one cutaneous metastasis of leiomyosarcoma. All cases were reviewed in order to confirm the diagnosis, before the cases were included in the study. In all cases, we performed an immunohistochemical study in all cases with p53 and the percentage of positive cells was estimate counting a total of 1000 cells per case. Results: Six cases from the 31 (19.35%) benign cutaneous smooth muscle tumors showed some expression of p53. The expression of it varied from only occasional cells to 1% of the cells. On the contrary, all leiomyosarcomas investigated showed expression of p53, and in three of the four cases (75%), the marker was expressed by at least 80% of the tumoral cells. Only in one leiomyosarcoma, the marker was expressed by a low percentage (0.5%) of cells. No expression of p53 was found in the only case of symplastic leiomyoma, which was investigated. The case of a cutaneous metastasis of leiomyosarcoma showed expression of p53 by 20% of cells. Conclusions: We conclude that expression of p53 by a high percentage of cells in a cutaneous smooth muscle cell tumor should be considered as highly suspicious for malignancy.

Corresponding author: Angel Fernandez-Flores, MD, PhD, e-mail: gpyauflowerlion@terra.es

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13. Atomic force microscopy study regarding the influence of etching on affected and sclerotic dentine

A. Georgescu, Gianina Iovan, Simona Stoleriu, Cl. Topoliceanu, S. Andrian

Aim of study is to compare the effect of etching with ortho-phosphoric acid on sound dentine, affected dentine and sclerotic dentine through AFM analysis. Material and Methods. The group study included 30 extracted third molars, 20 with acute and chronic carious lesions and 10 intact teeth. Teeth were sectioned in long axe to prepare sections with carious lesions surrounded by sound dentine. The sound teeth were sectioned to a depth of dentine comparable with carious lesions depth. The sections were etched with 37% ortho-phosphoric acid. The surface roughness was determined initially and after etching using AFM analysis. The results were expressed as relative variation of squared roughness. Results. The values of relative roughness indices were between 2.78 and 3 for sclerotic dentine, 3.18 and 3.26 for sound dentine, 3.32 and 3.38 for affected dentine. The highest values of roughness index were recorded for the affected dentine samples. Significant statistically values were recorded when comparing relative roughness indices for sclerotic dentine with relative roughness indices for affected dentine and sound dentine. Conclusions. Sclerotic dentine has significant higher resistance to the action of ortho-phosphoric acid than affected dentine and sound dentine. The lowest resistance to the action of etching agent was recorded for the affected dentine.

Corresponding author: Andrei Georgescu, Junior Assistant, DMD, e-mail: andgeorgescu@yahoo.com

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14. Immunohistochemical expression of growth factors in the exocrine pancreas of patients with chronic liver diseases

Garofita Mateescu, Maria Comanescu, Rodica Mehedinti, Zizi Niculescu, Adriana Bold, Laurentia Panduru, Daniela Cernea

Acute viral hepatitis has been reported to cause acute pancreatitis. It was also reported that exocrine pancreatic function is damaged in chronic liver disease (CLD). Growth factors stored in the extracellular matrix and released in the course of pancreatic degradation are major mediators of inductive processes. The immunostaining technique was used to evidence the changes of the expression of the growth factors in different pancreatic cells. VEGF and FGF-beta are involved in the angiogenesis processes and in the evolution of the pancreatic interstitial tissue in case of chronic pancreatitis. Theses markers can also be used for the diagnosis of pancreatitis, but their value is variable. They stimulate the pancreatic star cells, the myofibroblasts and play an important role in the genesis of the extracellular matrix and in the repairing of the tissue after the aggression. TGF beta is important for its role in cellular differentiation and growth and in the development of the fibrosis in liver and other organs. The present paper studies the immunohistochemical expression of these growth factors in pancreatic cells.

Corresponding author: Garofita Mateescu, Assistant Professor, MD, PhD, e-mail: garo2963@yahoo.com

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15. Aspects of oral morphology as decision factors in mini-implant supported overdenture

Elena Preoteasa, Marina Melescanu-Imre, Cristina Teodora Preoteasa, Mihaela Marin, Henriette Lerner

Aim: Evaluation of some morphological oral aspects perceived as decision factors in complete edentulism treatment by mini-implants overdenture. Patients, Material and Methods: An observational study was conducted on a sample of 24 patients (average age of 61 years), through clinical and imagistic methods. The variables taken into consideration were: age, gender, alveolar mucosa status, bone offer, mini-implants characteristics, insertion torque and loading type. Results: 117 mini-implants were applied, 59% to the lower jaw and 41% to the upper jaw. On average, there were inserted 5.33 implants to maxilla and 4.6 to mandible. Fifty-seven percent of implants were applied in the interforaminal area. The implants used had 10 mm (28.2%) and 13 mm (71.8%). In only 41% patients, the ridge width was higher than 5 mm (value necessary for conventional implant application). Diameters of implant used were: 1.8 mm (2.56%), 2.1 mm (25.64%) and 2.4 mm (71.8%). Smaller diameters were used in mandible. Only in 16.67% of patients was possible the transmucosal insertion. Bone density was D2 and D3 for men, and D3 and D4 for women. Immediate loading was achieved in 20.5% patients. Conclusions: Mini-implant supported overdenture can be an alternative to conventional denture and conventional implant overdenture. Its advantages derives from implants' characteristics (smaller diameter, variable length, O-ring retention system), which adapts better to the particular edentulous conditions. Insertion technique implies less surgical trauma. Choosing implants' size, number, topography, and the loading method have a great variety, depending on anatomical feature (bone offer, mucosa and relationship with the nearby anatomical structures), functional features and patients' wishes.

Corresponding author: Elena Preoteasa, Professor, DMD, PhD, e-mail: dr_elena_preoteasa@yahoo.com

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16. In vitro experiment of the modular orthopedic plate based on Nitinol, used for human radius bone fractures

Daniela Tarnita, D. N. Tarnita, L. Hacman, C. Copilus, C. Berceanu

Shape memory alloys (SMAs) and in particular Ni-Ti alloys are commonly used in bioengineering applications as they join important qualities as resistance to corrosion, biocompatibility, fatigue resistance, MR compatibility, kink resistance with two unique thermo-mechanical behaviors: the shape memory effect and the pseudoelastic effect. They allow Ni-Ti devices to undergo large mechanically induced deformations and then to recover the original shape by thermal loading or simply by mechanical unloading. Diaphyseal fractures of the radius and ulna present specific problems not encountered in the treatment of fractures of the shafts of other long bones. The adaptive modular implants based on smart materials represent a superior solution in the osteosynthesis of the fractured bones over the conventional implants known so far. To realize the model of the implant module we used SolidWorks software. The small sizes of the modules enable the surgeon to make small incisions, using surgical techniques minimally invasive, having the following advantages: reduction of soft tissues destruction; eliminating intra-operator infections; reduction of blood losses; the reduction of infection risk; the reduction of the healing time. Numerical simulations of the virtual modular implant are realized using Visual Nastran software. The stress diagrams, the displacements diagram and the strain diagram are obtained. An in vitro experiment is made, simulating the osteosynthesis of a transverse diaphyseal fracture of human radius bone. The kinematical parameters diagrams of the staple are obtained, using SIMI Motion video capture system. The experimental diagram force-displacement is obtained.

Corresponding author: Daniela Tarnita, Professor, Eng, PhD, e-mail: dtarnita@yahoo.com

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17. Amplification of HER-2 gene in breast cancer: immunohistochemical and FISH assessment

Alina Belengeanu, Anca Muresan, Dorina Stoicanescu, Elena Lazar

The invasive mammary cancer is the most frequent malignant tumor of women. Different inherited or acquired molecular genetic alterations have been identified in human breast cancers. A fraction of these cancers, as part of their development, undergoes gene amplification. Among the potential prognostic factors are included the biomarkers which measure or are associated with biological processes involved in tumor progression. Evaluation of HER-2 status is important in the management of patients with breast carcinoma, especially for the identification of those who are eligible for immunotherapy. The aim of our study was to evaluate HER-2 amplification status of human breast cancers by FISH and immunohistochemistry. From the total of 50 tumors included in the study, 17 (34%) presented different degrees of positivity; 33 (66%) did not express the oncoprotein HER-2. HER-2 gene and chromosome 17 status were tested in HER-2 2+ cases using FISH technique. FISH analysis may be useful to better evaluate HER-2 status in breast cancer in uncertain cases, where the immunohistochemistry score is 2+. HER-2 testing results have an important role in the clinical management of breast cancer patients. The identification of HER-2 positive tumors is certainly crucial in order to identify patient candidates for anti-HER-2 therapies.

Corresponding author: Alina Belengeanu, Assistant Professor, MD, PhD candidate, e-mail: alinabele@yahoo.com

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18. VEGF-induced corneal neovascularisation in a rabbit experimental model

L. Coman, Oana Andreia Coman, H. Paunescu, Fl. Draghia, I. Fulga

Introduction and Purpose: Various conditions may cause vascularization of the normally avascular cornea. The aim of the present study was to create a reproducible experimental model that could enable the investigation of the phenomena leading to corneal vascularization. This involved creating a software to record the experimental data, enabling a subsequent digital analysis based on the growth models. The VEGF-induced pattern of neovascularization was also investigated. Material and Methods: Twenty-seven rabbits divided in groups were used for the purposes of the present study. Some of them underwent intracorneal implants with or without vascular endothelial growth factor (VEGF) pellets, using an original microsurgical technique. Central and peripheral corneal burns were induced to other groups of animals in order to mimic the neovascularization process induced by inflammation. Finally, Dexamethasone (Maxidex) was given intraocularly, on days 1 and 3 after the onset of neovascularisation, in rabbit groups with both corneal burns and VEGF-implants. Video recording and data analysis of the corneal vascularization were made with an advanced biomicroscope, a computerized imaging system and a special software. A histochemical study of the animals' eyes was also carried out. Results and Discussion: The recorded data showed the simplicity and reproducibility of the present experimental model. The results showed the importance of VEGF as an initiator and promoter of corneal vascularization through a non-inflammatory mechanism, quite different from the inflammation illustrated by the corneal burn. At the same time, Dexamethasone therapy proved its effectiveness in corneal angiogenesis induced by thermal burn, but not by VEGF-implant.

Corresponding author: Laurentiu Coman, Lecturer, MD, PhD, e-mail: laurentiucmn@yahoo.com

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19. Microanatomical and immunohistochemical study of the human anterior branch of the medial antebrachial cutaneous nerve of forearm at the antecubital fossa and its clinical implications

S. Chakravarthy Marx, P. Kumar, S. Dhalapathy, Anitha C. Marx

Purpose: Poor prognosis of nerve repair in patients may be due to changes in intraneural anatomy with age. Also, chances of Complex Regional Pain Syndrome-Type I (CRPS-I) secondary to peripheral nerve injury are comparatively high. The present study is to find the fascicular pattern of the anterior branch of the medial antebrachial cutaneous nerve of forearm (MACN) (at antecubital fossa), microanatomic morphometric characteristics of its connective tissue components (adipose tissue) and changes with age and study of intraneural sympathetic fiber content. Material and Methods: Sixty six human (37-88-year-old) cadaveric anterior branch of MACN have been collected from antecubital fossa and the study has been performed at magnifications (5x, 10x, 20x, and 40x objective) after routine histological (Hematoxylin & Eosin stain, Masson's trichrome stain) processing was done for morphometric analysis (total cross-sectional, fascicular and non-fascicular area) and immunohistochemical (tyrosine hydroxylase) processing for sympathetic fibers. Results and Conclusions: The anterior branch of the MACN's average total cross section area was 1.150 mm2 on right side and 1.156 mm2 on left side. There was significant increase in non-fascicular connective tissue area. In non-fascicular area, there was very less amount of adipose tissue in 86.37% of cases and more adipose tissue in 13.63% (elderly) cases. The average sympathetic fiber area is 0.0109 mm2 without definite relationship with age. Our study makes an attempt to build a normal data base for MACN which might be helpful during the application of diagnostic and surgical nerve graft procedures.

Corresponding author: S. Chakravarthy Marx, PhD, e-mail: marxmanipal@gmail.com

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20. Morphology of the medial meniscus of the knee in human fetuses

B. V. Murlimanju, Narga Nair, Mangala M. Pai, A. Krishnamurthy, X. Chandra Philip

Objectives: The objectives were to study the morphology of the medial menisci in human fetal knee joints and to verify the developmental etiology of the discoid meniscus. Material and Methods: The study was carried out on 106 fetal knee joints and the morphological variants of the shapes of the medial menisci were macroscopically noted and classified. Results: From our observations, 46.2% of the medial menisci were crescentic shaped, 23.6% were sided V-shaped, 13.2% had sided U-shape, 9.4% of them were sickle shaped, and 7.5% were C-shaped. In 54.71% of the cases, the medial meniscus shape was different on either side knees of the fetuses. No discoid medial menisci were seen. Conclusions: The majority of the knees were having crescentic or semi lunar shaped medial menisci and the shape of the medial meniscus was different on right and left side knee joint of the fetuses in more than fifty percent of the cases. The observations favor Kaplan's theory, as there were no discoid medial menisci observed from the fetuses of various gestational ages. Even the youngest fetus (CRL 88 mm, 14 weeks of gestation) exhibited the medial tibial plateau incompletely covered by the menisci, which did not exhibit the discoid shape. The discoid meniscus must be an abnormal finding and is due to abnormal morphogenesis.

Corresponding author: B. V. Murlimanju, Assistant Professor, MD, e-mail: flutesnowmm@gmail.com

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21. The composite lymphoma: chronic lymphocytic leukemia - classic Hodgkin's lymphoma

M. Badea, Camelia Dobrea, Daniela Badea, Amelia Genunche-Dumitrescu, P. Mitrut, Doriana Duta

The composite lymphoma (CL) is defined by the presence in the same tissue or organ of two distinct histological aspects of non-Hodgkin's lymphoma (NHL), or NHL and Hodgkin's lymphoma (HL). The definition of the CL has evolved, requesting the identification of the immunophenotypic pattern and clonal distinct aspects for the two-lymphoproliferative lesions. We present a case of a 73-year-old farmer who presented with B-symptoms and multiple adenomegaly. The biopsy of a left cervical lymph node reveal a CL: a histological and immunophenotypic aspect of HL-mixed cellularity (CD15+, CD30+, CD20-) and a diffuse small cell infiltrate which meet the criteria for B-CLL (CD20+, CD23+, and CD5+). The lymphocytes in peripheral blood over 15000/mm3 and marrow infiltrate with small lymphocytes also sustain the B-CLL diagnosis. The relationship between the two lymphoproliferations is discussed reported to the case above, but also considering the literature data. In most of the cases the two proliferative processes are clonal related which means they have a commune lymphoid progenitor, pre-GC or early-GC with individual detachment and transit through GC (also, the afferent related processes). It is also possible that the two proliferations, which form the composite lesion to have different cellular origins, possibility sustained by the analysis of the IgH rearrangements and of the somatic mutations identified in the two clones. The EBV-role in HL-pathogeny is related to the way of salvage or/and initiation of a clonal process in a GC-cell which has major deletions in the variable part of IgH.

Corresponding author: Mihail Badea, Professor, MD, PhD, e-mail: mihbadea@yahoo.com

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22. Thyroid regional metastasis from a giant cell malignant fibrous histiocytoma of the larynx in a patient with history of trichinellosis and tuberculosis

Corina Lichiardopol, I. Osman, S. D. Enache, Camelia Foarfa, Violeta Comanescu, Mirela Ghilusi, M. Popescu

Sarcomas represent less than 1% of malignant laryngeal tumors and giant cell malignant fibrous histiocytoma is exceptionally rare. Diagnosis is histologically based and immunohistochemistry allows differentiation from other fibro-histiocytic neoplasms. We present the case of a 53-year-old male patient with positive medical history for trichinellosis and tuberculosis, and a laryngeal tumor invading the thyroid and causing respiratory distress by airway obstruction. Total laryngectomy and thyroidectomy were performed followed by thyroxine replacement therapy and radiotherapy. Histologically, the tumor consisted of spindle shaped cells with prominent mitoses, and abundant, osteoclast-like, multinucleated giant cells. Similar lesions were identified in the thyroid, adipose and muscular tissues. Parasitic elements were present in muscles. Tumoral cells showed positive immunostaining for Ki67 (40-50%) and vimentin and negative for AE1/AE3, CD31, S100 and myoglobin; the giant multinucleated cells were CD68-positive. Chronic infection might have had a pathogenic significance.

Corresponding author: Corina Lichiardopol, MD, PhD, e-mail: corinalich@gmail.com

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23. Pancreatic metaplasia of gastric mucosa associated with gastroduodenal ulcer

L. Mogoanta, C. T. Streba, D. Pirici, Rodica Dirnu, B. Oprea

Metaplasia represents the process of transforming a well-differentiated adult tissue into another type of adult tissue. Pancreatic metaplasia of the gastric mucosa represents the process in which the normal mucosa of the stomach is replaced with pancreatic formations, which mimic the structure of pancreatic acini. We describe the case of a male patient aged 39 who was admitted for abdominal pain, vomiting, hematemesis, melena, pale teguments, intense perspiration and nausea. The patient underwent surgery for suturing a perforated duodenal ulcer five years prior to this episode (2002). A gastric ulcer complicated with superior digestive bleeding and a chronic duodenal ulcer complicated with partial stenosis and perivisceritis were found during surgery. Gastric wall fragments were harvested and underwent usual histological techniques and immunohistochemistry. We found an ulceration from the gastric mucosa to the submucosa, covered by fibrino-leukocytic detritus. In the mucosal chorion we found numerous round or oval shaped nested formations which occupied the lower two thirds of the chorion, to the muscularis mucosae. Some metaplasic acini contain cells variable in shape, color and immunophenotype. Surrounding the nested acini we found tubular formations, formed of cubic shaped cells, representing excretory canals which were continued by gastric glands or opened directly in the crypts of the gastric epithelium.

Corresponding author: Laurentiu Mogoanta, Professor, MD, PhD, e-mail: laurentiu_mogoanta@yahoo.com

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24. A simple prosthetic restorative solution of a single peg-shaped upper central primary incisor in a case of ectodermal dysplasia

Anca Maria Raducanu, Mihaela Pauna, I. V. Feraru

Introduction: This clinical report describes the oral rehabilitation of a 7-year-old male diagnosed with hypohidrotic ectodermal dysplasia and anodontia. Background: Ectodermal dysplasia (ED) comprises a large, heterogeneous group of inherited disorders, which are defined by primary defects in the development of two or more tissues derived from the embryonic ectoderm (skin, hair follicles, nails, teeth, sebaceous and sweat glands). Our patient exhibited the typical general and cranio-facial features of ED as well as emotional and behavioral problems. The patient presents only one conical primary central incisor. Clinical management: The treatment was focused to improve his esthetic needs and oral functions and included the fabrication of removable prostheses and of an acrylic crown on the single tooth existing in his mouth, manufactured by an original simple method. Six months follow-up points out that restorative treatment provided psychosocial and functional comfort to the young boy. Conclusions: This clinical report demonstrates the importance of the prosthodontic treatment for oral rehabilitation in children with ED. The partial and complete removable dentures associated with the acrylic crown can be a reversible and inexpensive method of treatment for ED patients.

Corresponding author: Anca Maria Raducanu, Associate Professor, MD, PhD, e-mail: raducanu2000@yahoo.com

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25. Capsular origin of the long head of the biceps brachii tendon: a clinical case

J. M. Egea, Consolacion Melguizo, J. Prados, Antonia Aranega

The biceps brachii tendon arises directly from the superior glenoid labrum with the remainder usually attached to the supraglenoid tubercle. Although some cases of anomalous origin of this tendon have been described, these anomalies are rarely encountered in daily practice. We report a patient with a capsular origin of the LHBT as a congenital anomaly and present the clinical, magnetic resonance and arthroscopic findings. Recognition of this anatomic variation may be important to explain the patient's clinical data and to aid both diagnosis and surgery.

Corresponding author: Jose Prados, MD, e-mail: jcprados@ugr.es

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26. Axillary and perianal leiomyosarcoma: report of two cases

C. Mesina, I. Vasile, I. D. Vilcea, C. C. Vere, Claudia Valentina Georgescu, Mirela Ghilusi, M. Pasalega, H. Parvanescu, F. Calota, S. S. Mogoanta

Soft tissue leiomyosarcoma is a relatively rare malignant tumor. It may be difficult to be distinguished from gastrointestinal stromal tumors and Schwann cell neoplasms. To make a correct identification of soft tissue leiomyosarcoma, immunostaining with several smooth muscle differentiation markers (actin, calponin and desmin), and negative staining results with S100 (to rule out Schwann cell neoplasm), c-kit and CD34 (to rule out gastrointestinal stromal tumors) is needed. Prompt diagnosis and referral are desirable, since the size of the tumor at presentation is a continuous variable for the risk of local recurrence and metastatic disease. Chemosensitivity varies according to the tumor subtype, and the tumor grade, the patient's age, performance status, and the timing of metastatic disease further influence the likelihood of a response and survival. Chemotherapy is palliative for most patients with unresectable or metastatic disease. Ifosfamide and doxorubicin are routinely used in this setting; doxorubicin as a single agent is considered the drug of choice.

Corresponding author: Cristian Mesina, Assistant Professor, MD, PhD, e-mail: mesina.cristian@doctor.com

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27. Isolate vertigo crisis revealing an endolymphatic sac tumor

C. Codreanu, P. Tran Ba Huy

Endolymphatic sac tumors are rare entities that have a destructive potential on the temporal bone. They are aggressive tumors presenting as low-grade papillary adenocarcinoma, but there are no reports of metastasis in the literature. The Von Hippel-Lindau disease is a hereditary condition caused by germinal mutations of the tumor-suppressor VHL-gene. We present the case of an endolymphatic sac tumor associated with the Von Hippel-Lindau disease at a 46-year-old patient revealed by an isolate vertigo crisis, discussing the management of the tumor and the clinical, imaging, genetic and histopathologic features of it. Conclusions: Endolymphatic sac tumors have recently been described as part of the Von Hippel-Lindau disease, a genetic disorder involving the development of hypervascular tumors. The treatment depends on the size of the tumor, however surgical approach is the most successful choice and can be associated or not with radiotherapy.

Corresponding author: Corneliu Codreanu, MD, e-mail: tatoi53@yahoo.com

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28. A case of visceral leishmaniasis in Oltenia region (Romania)

Amelia Gaman, Camelia Dobrea, G. Gaman

Visceral leishmaniasis is produced by a protozoan parasite that belongs to the genus Leishmania. Transmission is made through sting, the vector being represented by a species of the genus Phlebotomus. The first case of visceral leishmaniasis in Romania was reported by Manicatide (1912). In 1934, it was described a focus of visceral leishmaniasis in Oltenia region (24 cases).The symptoms of disease are unspecific: fatigue, feverishness, cephalalgia, anorexia, nausea, obnubilation status. The fever is irregular, with high oscillations. Clinical, a sallow pallor of the skin, enlarge lymph nodes, hepatomegaly, splenomegaly, weight loss have been observed. Laboratory exams showed frequently severe anemic syndromes or other cytopenias, erythrocytes sedimentation rate was increased, hypergammaglobulinemia with monoclonal peak has been found. Immunolectrophoresis showed hyper-IgG and hyper-IgM. Bone marrow biopsy showed lympho-plasmocyte infiltration, histiocytes, Leishman-Donovan bodies intracellular or extracellular. The prognosis of the disease is unfavorable in the absence of specific treatment with antimony. In case of resistance, it is used immunotherapy, amphotericin or miltefosine.

Corresponding author: Amelia Gaman, Associate Professor, MD, PhD, e-mail: gamanamelia@yahoo.com

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29. Chondrohumeralis and axillary arch of Langer: a rare combination of variant muscles with unique insertion

Polly Lama, B. K. Potu, K. M. R. Bhat

During routine dissection, we found a rare case of an aberrant muscular slip originating from the pectoralis major called as the "chondroepitrochlearis / thoracoepicondylaris / costohumeralis" and an anomalous slip from the latissimus dorsi called as "axillary arch of Langer / musculus dorsoepitrochlearis" in the same axilla. Interestingly, these two slips found to have a common insertion after arching superficial to the axillary neurovascular bundle, into the fascia covering the biceps brachi and to the lateral lip of the intertubercular sulcus of the humerus. Presence of combination of these two rare variant muscles and their unique insertion pattern has not been reported earlier. The knowledge of these muscle variations is important for the clinicians and physiotherapist for better diagnosis and treatment. Here in this report, in addition to the case presentation, we also discuss their clinical significance and the review of literature in detail.

Corresponding author: Kumar M. R. Bhat, Associate Professor, MSc, PhD, e-mail: kummigames@yahoo.com

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30. A rare origin of abductor pollicis longus

Rajalakshmi Rai, Anu V. Ranade, Mamatha T, Jiji PJ, Sujatha D'Costa, Maheshwari C

Abductor pollicis longus is known for its variations in the form of multiple tendons at its insertion. The present case reports about a variation in the origin of abductor pollicis longus (APL) which has not been cited in the medical literature until date. It has been shown that the intersection area where the APL and extensor pollicis brevis cross over the tendons of the extensor carpi radialis (ECR) tendons in the forearm can contribute to Intersection syndrome. This can be applied to the present variation where the two bellies of APL entrap the ECR tendons. An awareness of this unusual finding may be clinically relevant for surgical approaches in the forearm region.

Corresponding author: Rajalakshmi Rai, Senior Lecturer, e-mail: rajalakshmi.rai@manipal.edu

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31. Hepato-spleno-mesenteric trunk: a case report

K. Sridhar Varma, Narendra Pamidi, Venkata Ramana Vollala, Sreenivas R. Bolla

Hepatic, splenic and left gastric arteries are the "classical branches" of celiac trunk. The authors report a rare variation, hepato-spleno-mesenteric trunk with two classical branches of celiac trunk and superior mesenteric artery having common origin from the abdominal aorta. The third classical branch of the celiac trunk (left gastric artery) was directly arising from the abdominal aorta. Knowledge of variations concerning the celiac trunk and superior mesenteric arteries are of great important for both surgical approaches and angiographic examinations.

Corresponding author: Narendra Pamidi, Senior Grade Lecturer, e-mail: pommidi_narendra@yahoo.co.in

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