Determination of VEGFR-2 (KDR) 604A>G Polymorphism in Pancreatic Disorders.

Pancreatic disorders have a high prevalence worldwide. Despite the fact that screening methods became more effective and the knowledge we have nowadays about pancreatic diseases has enhanced, their incidence remains high. Our purpose was to determine whether single nucleotide polymorphism (SNP) of VEGFR-2/KDR (vascular endothelial growth factor receptor 2/kinase insert domain receptor) influences susceptibility to develop pancreatic pathology. Genomic DNA was extracted from blood samples collected from patients diagnosed with acute pancreatitis (n = 110), chronic pancreatitis (n = 25), pancreatic cancer (n = 82) and healthy controls (n = 232). VEGFR-2 (KDR) 604A>G (rs2071559) polymorphism frequency was determined with TaqMan allelic discrimination assays. Statistical assessment was performed by associating genetic polymorphism with clinical and pathological data. In both pancreatic disorders and healthy control groups the polymorphism we studied was in Hardy-Weinberg equilibrium. Association between increased risk for pancreatic disorders and studied polymorphism was statistically significant. KDR 604AG and AG + GG genotypes were more prevalent in acute pancreatitis and pancreatic cancer patients than in controls. These genotypes influence disease development in a low rate. No association was found between chronic pancreatitis and KDR 604AG and AG + GG genotypes. In Romanian cohort, we found an association between the KDR 604A→G polymorphism and acute pancreatitis and pancreatic cancer. Carriers of the -604G variant allele were more frequent among acute pancreatitis and pancreatic cancer than among controls, suggesting that KDR 604G allele may confer an increased risk for these diseases. In the future, more extensive studies on larger groups are necessary, in order to clarify the role of VEGFR2 polymorphisms in pancreatic pathology.

Hepatic pathological features in naïve patients with chronic hepatitis C who have developed thyroid disorder.

Knowing the hepatic pathological features encountered in patients with chronic hepatitis C (CHC) and the fact that extrahepatic manifestations occur only in people with certain characteristics of the immune system, we tried to evaluate, qualitatively and semi-quantitatively, the liver pathological aspects encountered in 96 patients with CHC, previously untreated with Interferon (naïve), who showed or did not show signs of thyroid disorder (TD), hospitalized in the 2nd Medical Clinic of the Emergency County Hospital, Craiova, Romania, within a period of five years (2007-2012). Following hormonal, immunological, and thyroid ultrasound investigations, 14 (14.58%) of the 96 patients showed signs of TD. The main clinical forms of TD in the studied patients with CHC were autoimmune thyroiditis and subclinical hypothyroidism. In the patients with CHC with TD, we found mild chronic hepatitis in 14.28% of cases, the appearance of moderate chronic hepatitis was found in 71.42% patients, and the appearance of severe chronic hepatitis was found in 14.28% patients, while in the patients with CHC without TD we found chronic mild hepatitis in 62.19% of cases, the appearance of moderate chronic hepatitis was met in 32.92% patients, and the appearance of severe chronic hepatitis was found in 4.87% of patients. Mild and moderate fibrosis were found only in CHC patients without TD in a percentage of 25.6% and 65.85%, respectively, while severe fibrosis was found at 12.19% among CHC patients without TD and 92.85% among CHC patients with TD. The pathological aspect of liver cirrhosis was found only in those with TD (7.14%). In conclusion, the pathological features which define the liver necroinflammatory process, as encountered at the pathological examination in CHC patients with TD are the same as in any active chronic hepatitis, the differences being represented by the higher percentage of the periportal and the preseptal necrosis (piecemeal necrosis), as well as by the higher score of portal inflammation. In addition, the severe hepatic fibrosis and the histopathological appearance of the liver cirrhosis have only defined the cases of CHC with TD.

Multidisciplinary approach to patients with manifestations and pulmonary complications of cystic fibrosis.

Cystic fibrosis (CF) is a genetic disease, with autosomal recessive transmission, multisystemic, characterized by a remarkable clinical polymorphism and significant lethal prospective. Respiratory manifestations dominate the clinical picture, being present in all patients. The aim of the paper was to analyze the incidence of clinical manifestations, especially respiratory ones, as well as the contribution of interdisciplinary consultations to the positive diagnosis of CF, in a group of 16 patients who were hospitalized and treated in the IInd Pediatric Clinic and IInd Medical Clinic of the Emergency County Hospital, Craiova, Romania, in a period of 20 years. The 16 patients diagnosed with and treated of CF had all shown increased values of sweat chloride concentration of over 60 mmol∕L. The main symptoms and clinical signs encountered in these patients were cough (75%), sputum (62.5%), dyspnea (50%), wheezing (50%), stature hypotrophy (100%), pallor (37.5%), cyanosis (25%). All 16 patients had an acute exacerbation of chronic pulmonary disease. Of the total hospitalizations, the death was recorded only in the case of one female patient. The association of some clinical aspects specific with a positive result of the sweat test or the presence of the two pathological alleles made room for determining a positive diagnosis. The multisystemic nature of this disease requires a multidisciplinary approach to these patients. Histopathologically, there was a correspondence between lung morphological lesions and the results of imaging investigations.

The association between chronic pancreatitis and the iNOS-2087A>G polymorphism.

INTRODUCTION: Chronic pancreatitis is morphologically characterized by ductal dysplasia, breeding grounds for the proliferation of the ductal cells, the degenerative changes in pancreatic acinar cells and fibrosis, and it is defined on the basis of the clinical, morphological and functional criteria. AIM: The aim of our study is to examine the existence of a possible correlation between the iNOS-2087A>G polymorphism and chronic pancreatitis by means of the genetic analysis. MATERIAL AND METHOD: We have conducted the study at the Gastroenterology Clinic and the Research Center of Gastroenterology and Hepatology of the University of Medicine and Pharmacy, Craiova, between March 2015 - September 2016. The study had a prospective character. Both for the 58 patients diagnosed with chronic pancreatitis and for the 132 patients in the witness group, the biological material was represented by blood, (around 2.5 - 5 milliliters of venous blood) let on EDTA and kept at 4°C up to the separation of the DNA molecule. All the patients were genotyped for the iNOS - 2087A>G polymorphism, by means of the Real Time PCR technique with TaqMan probes. RESULTS: Analysing the prevalence of the iNOS genotypes within the study group and witness group, we have noticed that, statistically speaking, there are no significant differences between the two groups. CONCLUSION: As a conclusion, in the study lot we can sustain that the risk of developing chronic pancreatitis is not increased by the presence of the iNOS-2087A>G polymorphism.

The Relationship of Cytokines IL-13 and IL-17 with Autoantibodies Profile in Early Rheumatoid Arthritis.

Aims. In the present study, we aimed to assess the concentrations of IL-13 and IL-17 in serum of patients with early rheumatoid arthritis (eRA), the investigation of correlation between the concentrations of these cytokines and disease activity score, and the concentration of some autoantibodies and the evaluation of the utility of IL-13 and -17 concentration measurements as markers of disease activity. Materials and Methods. Serum samples were collected from 30 patients and from 28 controls and analysed parameters. Results. The serum concentrations of IL-13, IL-17, anti-CCP, and IgM-RF were statistically significantly higher in patients with eRA, compared to the controls. IL-13 concentrations in the severe and moderate groups with eRA were statistically higher than in the mild and control groups. Also, in the case of IL-17, serum concentrations increased proportionally with the disease activity of eRA. We observe that concentrations of IL-13 and -17 did not correlate with autoantibodies. IL-17 concentration significantly positively correlated with CRP, while IL-13 concentration significantly negatively correlated with CRP. Disease activity score, DAS28, was strongly positively correlated with levels of ESR and weakly positively correlated with concentrations of anti-RA33 autoantibodies. IL-13 has a higher diagnostic utility than IL-17, CRP, ESR, IgM-RF, and anti-CCP as markers of disease activity. Conclusions. The presence of higher IL-13 and IL-17 serum levels in patients, compared with those of controls, confirms that these markers, found with high specificity, might be involved in the pathogenesis of eRA. IL-13 and IL-17 might be of better usefulness in the prediction of eRA activity status than IgM-RF and anti-CCP.

Histological factors that predict the liver fibrosis in patients with chronic hepatitis C.

In chronic hepatitis, pathologies reveal a prominent inflammatory infiltrate portal consisting mostly of lymphocytes and plasma cells invading the portal spaces, although one can also identify macrophages, neutrophils or eosinophils. In all the forms of chronic hepatitis, fibrosis starts in the portal area, namely periportally, subsequently extends towards the lobules to the central veins, causing septa, followed by fibrosis. We studied 52 patients with chronic hepatitis C, who underwent a hematological, biochemical, virological and histopathological investigation. We found that the severity degree of the portal inflammation was in direct relation to the hepatitis activity index (HAI) and to the degree of fibrosis. The portal inflammation is dependent to the degree of fibrosis. The degree of inflammation significantly changes the distribution of cases with different degrees of fibrosis (chi-square p=0.00011 <0.001). Periportal inflammation, periportal necrosis and focal necrosis are the morphological aspects of the necroinflammatory process best correlated to the occurrence and development of fibrosis.

Significance of circulating and crevicular matrix metalloproteinase-9 in rheumatoid arthritis-chronic periodontitis association.

In the recent years, statistically significant associations between rheumatoid arthritis (RA) and periodontal disease have been identified. Emerging as a chronic inflammatory joint disease, RA displays various features and pathogenetic events similar to chronic periodontitis (CP). The purpose of this study was to evaluate the utility of determining systemic and crevicular levels of metalloproteinase-9 (MMP-9) as potential biomarkers for association between RA and CP. A total of fifty-six patients were included in the study. The subjects were categorized into four groups as follows: healthy-control (n = 21), active RA (n = 16), CP (n = 14), and RA-CP association (n = 12). Assessment of serum and crevicular concentrations of total MMP-9 (active and pro-MMP-9) was based on ELISA technique. The results of this study showed statistically significant differences of serum MMP-9 between patients groups and control. Serum levels of MMP-9 were similar in RA and RA-CP associated patients. Gingival crevicular fluid (GCF) recorded increased MMP-9 levels in RA-CP association subjects as compared to CP. Considering that RA-CP association is characterized by a disregulation of the inflammatory response, MMP-9 may play a role in the pathogenesis of RA-CP association. MMP-9 is therefore a sensitive tool in the diagnosis and management of patients affected by this binomial association.

Histopathological aspects described in patients with chronic hepatitis C.

Chronic hepatitis C affects an estimated 170 million people worldwide and causes approximately 350 000 deaths each year. The current antiviral therapy allows the virus eradication or the permanent inhibition of the virus replication (sustained virological response, SVR), the reduction of the inflammation, and the prevention or the reduction of liver fibrogenesis (histological response). We studied the histopathological aspects found during percutaneous liver biopsy in patients with chronic hepatitis C viral infection who were treated and monitored over a period of two years. The assessment of the histological activity index through Ishak score determined the presence of: mild chronic hepatitis in 12 (23.1%) patients, moderate chronic hepatitis in 21 (40.4%) patients, and severe chronic hepatitis in 19 (36.5%) patients. The percutaneous liver biopsy performed on the patients with chronic viral hepatitis C showed a series of histological alterations, the most frequent being: portal inflammation, periportal necrosis, lobular inflammation, focal necrosis, and hepatic fibrosis (scarring). The severity degree of this histopathological aspect was correlated with the hepatitis activity index. The association of piecemeal with bridging necrosis is the deadline at which the antiviral treatment can still be effective. Evidence of early fibrosis represent the important moment for the antiviral treatment start. The specific histopathological aspects, but not pathognomonic, of chronic hepatitis C (hepatic steatosis, portal lymphoid infiltrates and bile duct damage) had a reduced incidence, occurring in only half (hepatic steatosis), a quarter (portal lymphoid infiltrates) and a fifth (destruction of biliary ducts) of all the patients with chronic viral hepatitis C, and these patterns was correlated with advanced degree of necroinflammatory process of the liver, particularly in the portal tracts.

Comparative study of clinical-morphological profiles of different types of gastric carcinoma.

AIM: Gastric carcinoma shows considerable variation in the histological pattern and degree of differentiation. The aim of the study was to assess especially the morphological differences between gastric carcinomas revealing one morphological feature and those including two morphological features. MATERIALS AND METHODS: Two groups of patients were selected: Group 1, including 43 patients with tumors revealing only one architectural pattern, and Group 2, including 16 patients with two architectural patterns within the tumor. In addition to gender and age, the main morphological parameters were: lesion location and macroscopic appearance on the surgical excision sample and microscopic appearance of the surgical excision sample, assessed for architectural pattern, secretory properties and prognosis based on histological features, degree of tumor extension and the degree of tumor aggressiveness, using a wide range of histological and immunohistochemical stainings. All data were compared between the two groups using statistical tests. RESULTS AND DISCUSSION: Significant differences were observed between the values and distributions of morphological parameters in the two groups and were discussed comparatively. CONCLUSIONS: Tumors with two dominant histological aspects present simultaneously are a reality that cannot be argued but whose morphological and biological profile needs to be completed and validated.

Morphological aspects in tuberculosis of oral cavity - our experience and a review of the literature attempt.

AIM: The authors continue a started series of articles about extrapulmonary tuberculosis (EPTB) with the assessment of the mycobacterial lesions discovered on tissue samples of the oral cavity structures in the Department of Pathology of the Emergency County Hospital of Craiova, Romania, and the review of the cases reported in the literature available, between 1990 and 2013. MATERIALS AND METHODS: The studied material consisted, for our series, of samples obtained by biopsy or surgical excision, including the salivary glands and excluding the lymph nodes from 17 patients histopathologically diagnosed with tuberculosis and, for review series, 190 papers selected from PubMed database. RESULTS: The number of cases reported increased throughout the studied period. Most cases came from departments connected with oral pathology but also from various medical and surgical departments. In general, patients were adults with a mean age of around 40 years, with twice as many men than women, without no information or no clinical suspicion of tuberculosis (TB) at the admission. When reported, the provisional diagnostic was oriented most often towards neoplastic proliferation. There was no information about human immunodeficiency virus (HIV) testing in more than half of the case reports but when existed the result was two-fold more frequently negative than positive. TB lesions of the oral cavity were more often primary infections than secondary. From morphological point of view, the granuloma cellular population included both epithelioid and Langhans cells in most of the cases, the necrosis, present in most of the cases, displayed the whole range of morphological features, but mainly the acidophilic, microgranular one and the perilesional fibrosis was absent in almost all of the cases. As a whole, well-differentiated granulomas were the most frequent, usually of grade II - reactive type ("homeostatic") but with a significant contingent of grade I - hyperplastic ("protective") granulomas. Local extension was usually not present and, when present, regional lymph nodes were mainly involved. Coexistence of TB lesions with a neoplastic proliferation was very rare and when present it was mainly located in the parotid gland. Apart from this general profile, particular, individual profiles were observed for each of the oral cavity segments. CONCLUSIONS: TB lesions in the oral cavity are indeed a rare event but no swelling or ulcer in the oral cavity should be disregarded by the medical practitioners because it could be tuberculosis.

E-cadherin and α-SMA expression in the epithelial-mesenchymal transition of salivary glands pleomorphic adenomas.

The pleomorphic adenoma, known also as mixed tumor of epithelioma with reshuffling stroma, is the most frequent salivary gland tumor, representing 45-75% of the total salivary gland tumors. In our study, we have investigated the immunohistochemical expression of E-cadherin and alpha-smooth muscle actin (α-SMA) in 15 cases of pleomorphic adenoma of salivary glands. We noticed the constant presence of E-cadherin reactivity at the level of luminal cells that cover the proliferative structures of ductal-cystic type, which gradually disappears to the periphery. At the level of epithelial proliferative solid areas, the reactivity for E-cadherin was inconstant and heterogeneous, while α-SMA expression increased. This aspect indicated the involving of epithelial-mesenchymal transition process in the evolution of pleomorphic adenoma.

Disseminated tuberculosis presenting as febrile seizures with fatal evolution in an infant.

Disseminated tuberculosis with the involvement of brain, liver and gut is a rare disease in immunocompetent infant. Early diagnostic and instauration of anti-tuberculosis therapy is capital because the outcome is poor. Here, we report the case of an 11-month-old boy with disseminated tuberculosis of brain, liver abdominal lymph nodes, small bowel and lung, which presented with fever, generalized tonic-clonic seizure, hemodynamic instability and a history of recurrent respiratory tract infections. His father was diagnosed with active pulmonary tuberculosis six month ago and family members completed an anti-tuberculosis chemoprophylaxis regimen.

Corpora amylacea in the brain form highly branched three-dimensional lattices.

Corpora amylacea (CA) are glycoprotein-based depositions that accumulate in the normal aging brain or consecutively to different neuro-degenerative diseases. Although controversies still exists in what it regards their origins and functions, their morphology is described simply as round basophilic entities based on bi-dimensional observations. The aim of the present study was to evaluate the three-dimensional morphology of these bodies in the brain in normal aging. We utilized here brain tissue from six aged patients, and performed an in-depth stereological analysis of CAs based on series of thin serial sections, and 50 µm-thick sections. The thin seriate sections have been counter-stained with Hematoxylin and Eosin, while the thick sections have been immunostained in fluorescence for ubiquitin and GFAP/collagen IV or aquaporin 4. Three-dimensional renderings have been obtained after aligning the serial sections, while high-resolution z-stacks have resulted after deconvolution on the thick sections. More than 70% of all the identified CAs proved to be in fact parts of larger aggregates, where the flattened individual spheroids branched and communicated with other bodies in a complex pattern, and budding of small CAs from larger CAs could be observed. There was a direct correlation between the diameter of the vessels and the number of associated CAs. Astrocyte GFAP and aquaporin 4 signals surrounded CAs, but without any colocalization with the ubiquitin areas, while perivascular CAs were sometimes enclosed in pockets of the basement membranes. In conclusion, as far as we know, this is the first study to describe the three-dimensional branching complexity of corpora amylacea in the brain.

The role of immunohistochemistry in diagnosing a synchronous colon tumor.

Simultaneous presence of an epithelial and lymphoid tumor of the digestive tract is quite rarely met in literature. In this paper, we describe a case which presented such an association. Diagnosis was established by histological study, followed by immunohistochemistry. It is a synchronous colon tumor, associating a non-Hodgkin's lymphoma to a colon adenocarcinoma. The 57-year-old male patient has been clinically diagnosed with a tumor of the left abdominal quadrant and paraclinically (imaging and endoscopic) with colon neoplasm. Exploratory laparotomy revealed two tumors: one tumor of five centimeters in the sigmoid, with firm consistency, mobile on lower plans and the second tumor in the ceco-ascending colon, measuring about 7 cm, irregular, with firm consistency, mobile on lower plans, with lymph nodes extending to retroperitoneal space. The urinary bladder, kidneys, liver and stomach were of normal aspect. Subtotal colectomy was performed with latero-lateral ileo-sigmoid anastomosis. Microscopic examination revealed sigmoid tumor as G1 adenocarcinoma and cecal tumor as B-cell type lymphoma. Immunohistochemistry established the final diagnosis of cecal localization being a diffuse immunoblastic large B-cell non-Hodgkin's malignant lymphoma. The final diagnosis of this patient was actually a synchronous manifestation of a colon adenocarcinoma and non-Hodgkin's lymphoma. This association puts into question synchronous tumors etiopathogeny matter.

Determination of autophagy gene ATG16L1 polymorphism in human colorectal cancer.

Autophagy has emerged not only as an essential repair mechanism to degrade damaged organelles and proteins but also as a major player in protection of tumor cells from multiple stresses. It was shown that autophagy gene polymorphisms are correlated with development of chronic inflammatory lesions, which represent a risk factor for colorectal tumors. In this study, we aimed to determine if ATG16L1 +898A>G (Thr300Ala) polymorphism is associated with an increased risk of developing colorectal cancer (CRC) and to establish correlations between ATG16L1 genotypes and the major clinical and morphological parameters. We observed that subjects carrying GG genotype were at a higher risk for CRC (OR 1.99, 95% CI: 1.02-3.91, p=0.039) when compared with the more frequent AA genotype, furthermore this was even more consistent in male subjects (OR 2.72, 95% CI: 1.11-6.63, p=0.019) but not in female subjects (OR 1.29, 95% CI: 0.43-3.86, p=0.652). In addition, we noticed a correlation between ATG16L1 GG genotype and tumor stage in moderately and poorly differentiated CRC cases. GG genotype carrying patients were at a higher risk for CRC (OR 5.19, 95% CI: 1.50-17.87, p=0.002) when compared with the more frequent AA genotype. Such correlation suggests a possible role of autophagy gene polymorphisms in the development of human colorectal cancer.

Tumor necrosis factor-alpha serum levels in healthy smokers and nonsmokers.

BACKGROUND: Tobacco smoking is the most important risk factor for chronic obstructive pulmonary disease (COPD) development. Inhaled cigarette smoke can induce tumor necrosis factor-alpha (TNF-alpha) production by alveolar macrophages, which in turn may enhance the production of metalloproteinases (MMPs). MMPs have been involved in mediating airway inflammation and lung destruction. OBJECTIVES: We aimed to measure the TNF-alpha serum levels in healthy heavy smokers and healthy nonsmokers to determine the dose-response relationship based on the cigarette smoke exposure. SUBJECTS AND METHODS: We included in our study 43 healthy heavy smokers and 19 healthy nonsmokers (the control group). The smokers group was classified as less than one pack, one pack, and more than one pack per day. A clinical and paraclinical evaluation was performed in both groups, without any evidence of infection or COPD. The serum levels of TNF-alpha were assessed by ELISA. RESULTS: The TNF-alpha serum levels were significantly higher for the group of smokers compared to the group of nonsmokers (P < 0.004). We also noticed an increased TNF-alpha concentration in the serum of smokers with more than one pack per day compared with those with less than one pack per day (P < 0.03). There was a positive correlation between the serum level of TNF-alpha and tobacco smoke exposure. CONCLUSIONS: The high levels of TNF-alpha in the serum of smokers suggest an imbalance between the proinflammatory and anti-inflammatory factors as a result of tobacco smoke exposure. The concentration of TNF-alpha is elevated in the serum of healthy heavy smokers in a cigarette dose-dependent manner. We speculate that the serum level of TNF-alpha might be a useful biomarker for the selection of heavy smokers with a high risk of developing smoke induced pulmonary diseases.