Ayhan Ozcan

Systemic Markers of Lipid Peroxidation and Antioxidants in Patients with Nonalcoholic Fatty Liver Disease

OBJECTIVES:The aim of the present study was to examine the systemic parameters of oxidative stress and antioxidants in patients with nonalcoholic fatty liver disease and investigate the relationship between these parameters and clinical and biochemical outcomes.METHODS:Fifty-one male patients with nonalcoholic fatty liver disease (group I), 30 age-matched and body mass index (BMI)-matched healthy male subjects, and 30 age-matched male patients with chronic viral hepatitis (group II) were enrolled in the study.RESULTS:Increased systemic levels of malondialdehyde and depletion of antioxidants such as coenzyme Q10, CuZn-superoxide dismutase, and catalase activity were observed in group I. Coenzyme Q10 and CuZn-superoxide dismutase correlated negatively with increasing necroinflammatory activity and fibrosis. Body fat was negatively associated with plasma coenzyme Q10 levels, while an inverse association was found between plasma catalase levels and TG. However, LDL was positively associated with plasma malondialdehyde levels. CuZn-superoxide dismutase levels were negatively associated with glucose, insulin, and HOMA-IR. In addition, the levels of CuZn-superoxide dismutase correlated significantly in a negative manner with BMI.CONCLUSIONS:Our results concerning correlations suggest that disturbances in BMI, body fat, and lipid metabolism may contribute to altered oxidative status in NAFLD, and insulin resistance may be related to decreased antioxidants in NAFLD as well as products of lipid peroxidation. However, although our results suggest interesting correlations, this different mostly “weak” relationships must be taken with caution.

PAX 8 expression in non-neoplastic tissues, primary tumors, and metastatic tumors: a comprehensive immunohistochemical study

PAX 8 is a transcription factor that is essential for embryonic development of the kidney, Müllerian organs, and thyroid. It may also have a role in tumor development in these organs. The diagnostic utility of PAX 8 has not been comprehensively studied. Formalin-fixed, paraffin-embedded tissue samples for non-neoplastic tissues (n=1601), primary neoplasms (n=933), and metastatic neoplasms (n=496) were subjected to PAX 8 immunostain. In non-neoplastic tissues, PAX 8 was consistently noted in glomerular parietal epithelial cells, renal collecting ductal cells, atrophic renal tubular epithelial cells regardless of nephronic segments, and epithelial cells of the endocervix, endometrium, fallopian tube, seminal vesicle, epidydimis, thyroid, pancreatic islet cells, and lymphoid cells. PAX 8 was not seen in the rest of the tissue samples. In primary neoplasms, PAX 8 was expressed by 194 of 240 (89%) renal cell neoplasms, by 238 of 267 (89%) Müllerian-type neoplasms, by 65 of 65 (100%) thyroid follicular cell neoplasms, by 8 of 8 (100%) nephrogenic adenomas, and by 17 of 17 (100%) lymphomas. Weak focal staining was noted in 5 of 12 (42%) cases of parathyroid hyperplasia/adenoma and in 6 of 17 (35%) well-differentiated neuroendocrine tumors of the pancreas. PAX 8 was not seen in other neoplasms. In metastatic neoplasms, PAX 8 was expressed by 90 of 102 (88%) metastatic renal cell carcinomas, by 57 of 63 metastatic Müllerian tumors (90%), and by 6 of 6 metastatic papillary thyroid carcinomas (100%). There was also weak focal staining for 1 of 15 metastatic small cell carcinomas and for 1 of 9 metastatic well-differentiated neuroendocrine carcinomas. PAX 8 was not seen in other metastatic neoplasms. It can be successfully identified in routinely processed tissue samples, and its expression is mostly nuclear. PAX 8 expression in non-neoplastic mature tissues is limited to the organs, the embryonic development of which depends on this transcription factor. This tissue/cell-specific expression is maintained during both neoplastic transformation and metastasis. PAX 8 is a sensitive and specific marker for tumors of renal, Müllerian, or thyroid origin in both primary and metastatic sites.

Elevated plasma homocysteine concentrations as a predictor of steatohepatitis in patients with non‐alcoholic fatty liver disease

Background:  Although steatosis is common in patients with severe hyperhomocysteinemia due to deficiency of cystathionine β‐synthase, there are no satisfactory data on homocysteine concentrations in patients with non‐alcoholic fatty liver disease. The main aim of the present study was to evaluate the clinical significance of plasma homocysteine concentrations in patients with non‐alcoholic fatty liver disease.

PAX2 and PAX8 expression in primary and metastatic müllerian epithelial tumors: A comprehensive comparison

PAX2 and PAX8 are transcription factors that are essential in embryonic development of müllerian organs. They may also play a role in tumor development in these organs. The diagnostic utility of PAX2 and PAX8 relative to one another has not been comprehensively studied. Archival tissue samples for normal or non-neoplastic tissue (251), primary epithelial neoplasms (316 for PAX2 and 357 for PAX8), and metastatic epithelial neoplasms (16), all of müllerian origin, were subjected to PAX2 and PAX8 immunostaining. The staining frequency, extent, and intensity for these markers were compared. Virtually identical PAX2 and PAX8 expressions were noted in non-neoplastic tissue. They were constantly seen in most epithelial cells (but not in stromal cells) of the endocervix, endometrium, fallopian tube, paratubal cyst, endosalpingiosis, endometriosis, and endometrial polyp. Within the primary epithelial neoplasms, PAX2 and PAX8 expression was noted in 55% and 98% of serous tumors, 25% and 94% of endometrioid tumors, 19% and 100% of clear cell tumors, 11% and 67% of transitional/undifferentiated tumors, and 10% and 22% of mucinous tumors, respectively. Regardless of histologic subtypes, PAX2 staining was noted in fewer cells and with less staining intensity compared with PAX8. No tumor showed only PAX2 staining. Within the metastatic carcinomas, PAX2 and PAX8 expression was noted in 38% and 98% of cases, respectively, with a diffuse and strong staining for PAX8, contrasting with a patchy and weak PAX2 expression. PAX2 and PAX8 are constantly expressed in normal or non-neoplastic tissue of müllerian origin. For primary and metastatic müllerian epithelial tumors, PAX8 shows strong and diffuse staining in most cases of all histologic subtypes, except in mucinous tumors. In contrast, PAX2 expression is always less than PAX8, and exclusive staining for PAX2 is not seen. PAX8 supersedes PAX2 as probably the best epithelial marker hitherto for primary or metastatic müllerian epithelial tumors.

PAX-2 expression in non-neoplastic, primary neoplastic, and metastatic neoplastic tissue: A comprehensive immunohistochemical study.

PAX-2 is a transcription factor that controls the development of the kidney, organs deriving from the mesonephric (Wolffian) duct, and those related to the Müllerian duct. Although PAX-2 is shown to be a sensitive marker for tumors derived from these organs, but whether it is specific, that is, whether other tumor types also express PAX-2, has not been systematically evaluated in either primary or metastatic tumors. Tissue sections from 937 normal or reactive tissue samples, 759 primary neoplasms, and 332 metastatic neoplasms were submitted to PAX-2 immunostain. Among the non-neoplastic tissue, PAX-2 was expressed in glomerular parietal epithelial cells, renal collecting duct cells, atrophic renal tubular cells, epithelial cells of ovarian surface, fallopian tube, endocervix, endometrium, seminal vesicle, and lymphocytes. Among the primary neoplasms, PAX-2 was noted in 104/122 (85%) of renal cell carcinoma, 31/95 carcinomas of Müllerian origin, 17/17 (100%) lymphomas, 4/4 (100%) nephrogenic adenomas, and 1/16 (6%) benign parathyroid tumors, but was negative in 477 other tumors. Among the metastatic tumors, PAX-2 was noted in 70/95 (74%) metastatic renal cell carcinomas, 14/20 (70%) metastatic tumors of Müllerian origin, 1/20 (5%) metastatic colon carcinoma of lymph nodes, 1/62 (2%) metastatic breast carcinoma of lymph nodes, but was not seen in the remaining 247 metastatic tumors. PAX-2 expression in non-neoplastic mature tissue is limited to the organs whose embryonic development depends on this transcription factor. PAX-2 is a sensitive and specific marker for tumors of renal or Müllerian origin in both primary and metastatic contexts.

Increased Acylation Stimulating Protein Concentrations in Nonalcoholic Fatty Liver Disease are Associated with Insulin Resistance

OBJECTIVES:As acylation stimulating protein (ASP) acts on adipocytes mainly as a paracrine factor to increase triglyceride synthesis and storage; hypothetically, it may play a similar role in the pathogenesis of nonalcoholic fatty liver disease (NAFLD).METHODS:Forty-six male patients with NAFLD (group A), age-matched 30 male patients with chronic viral hepatitis (group B) and 30 age-matched and body mass index (BMI)-matched healthy male subjects were enrolled in the study.RESULTS:Among the NAFLD patients, 10 patients (24.4%) had simple steatosis and 36 patients (69.6%) had nonalcoholic steatohepatitis (NASH). The mean levels of ASP, complement 3, insulin, C-peptide, HOMA-IR, triglyceride, and very low-density lipoprotein (VLDL) were significantly higher in group A patients than both controls and group B. ASP levels correlated significantly in a positive manner with BMI, insulin, and HOMA-IR.CONCLUSIONS:Dysregulation of the ASP pathway may have important metabolic consequences in NASH and is associated with insulin resistance.

Novel monoclonal antibodies detect Smad-interacting protein 1 (SIP1) in the cytoplasm of human cells from multiple tumor tissue arrays.

Smad-interacting protein 1 (SIP1, also known as ZEB2) represses the transcription of E-cadherin and mediates epithelial-mesenchymal transition in development and tumor metastasis. Due to the lack of human SIP1-specific antibodies, its expression in human tumor tissues has not been studied in detail by immunohistochemistry. Hence, we generated two anti-SIP1 monoclonal antibodies, clones 1C6 and 6E5, with IgG1 and IgG2a isotypes, respectively. The specificity of these antibodies was shown by Western blotting studies using siRNA mediated downregulation of SIP1 and ZEB1 in a human osteosarcoma cell line. In the same context, we also compared them with 5 commercially available SIP1 antibodies. Antibody specificity was further verified in an inducible cell line system by immunofluorescence. By using both antibodies, we evaluated the tissue expression of SIP1 in paraffin-embedded tissue microarrays consisting of 22 normal and 101 tumoral tissues of kidney, colon, stomach, lung, esophagus, uterus, rectum, breast and liver. Interestingly, SIP1 predominantly displayed a cytoplasmic expression, while the nuclear localization of SIP1 was observed in only 6 cases. Strong expression of SIP1 was found in distal tubules of kidney, glandular epithelial cells of stomach and hepatocytes, implicating a co-expression of SIP1 and E-cadherin. Squamous epithelium of the esophagus and surface epithelium of colon and rectum were stained with moderate to weak intensity. Normal uterus, breast and lung tissues remained completely negative. By comparison with their normal tissues, we observed SIP1 overexpression in cancers of the kidney, breast, lung and uterus. However, SIP1 expression was found to be downregulated in tumors from colon, rectum, esophagus, liver and stomach tissues. Finally we did nuclear/cytoplasmic fractionation in 3 carcinoma cell lines and detected SIP1 in both fractions, nucleus being the dominant one. To our best knowledge, this is the first comprehensive immunohistochemical study of the expression of SIP1 in a series of human cancers. Our finding that SIP1 is not exclusively localized to nucleus suggests that the subcellular localization of SIP1 is regulated in normal and tumor tissues. These novel monoclonal antibodies may help elucidate the role of SIP1 in tumor development.

The bisphosphonate zoledronic acid inhibits the development of plasmacytoma induced in BALB/c mice by intraperitoneal injection of pristane

Abstract:  Objectives: Bisphosphonates (BPs) are mostly used in the palliative care of myeloma‐associated osteolytic lesions. Recent studies have suggested that BPs may also exert direct antitumor effects on myeloma cells. We have investigated the effect of the potent bisphosphonate, zoledronic acid (ZOL), on the development of pristane (2,6,10,14‐tetramethylpentadecane)‐induced plasmacytoma (PCT) in six‐week‐old BALB/c mice. Methods: Different groups of pristane‐treated mice also received ZOL (100 μg/kg) commencing after the development of PCT or ZOL (20 μg/kg) from the first day. Control groups received pristane alone, ZOL alone (20 μg/kg), or phosphate‐buffered saline. The study was terminated on day 300, and the remaining mice were autopsied and abdominal tissues were examined histologically for PCT. Results and conclusions: Statistical analysis revealed a significant delay in PCT development in the group receiving pristane plus ZOL (20 μg/kg) from the first day compared to the groups receiving pristane alone and pristane combined with ZOL (100 μg/kg) after the appearance of PCT (Log‐rank, P = 0.0001 and 0.0001; respectively). Kaplan–Meier analysis revealed a significant difference in survival between the group treated with pristane alone and the groups receiving pristane plus ZOL (20 μg/kg) from the first day or ZOL (100 μg/kg) after the appearance of PCT (Log‐rank, P = 0.016 and 0.023; respectively). These results indicate a direct anti‐tumor effect of ZOL in pristane‐induced PCT development BALB/c mice, which may contribute to their significantly increased survival. This hypothesis should now be further investigated in clinical trials.

Beneficial Effects of N-Acetylcysteine and Ebselen on Renal Ischemia/Reperfusion Injury

Abstract Introduction: It has been demonstrated that peroxynitrite accompanies acute renal ischemia and contributes to the pathophysiology of renal damage. Therefore, we aimed to investigate the roles of N-acetylcysteine (NAC), a well-known powerful antioxidant, and ebselen (E), a scavenger of peroxynitrite, on renal injury induced by renal ischemia/reperfusion injury (IRI) of rat kidney. Materials and methods: Forty male Sprague–Dawley rats were divided into five groups: sham, renal IRI, renal IRI+NAC, renal IRI+E, and renal IRI+NAC+E. IR injury was induced by 60 min of bilateral renal ischemia followed by 6 h of reperfusion. After reperfusion, kidneys and blood samples were obtained for histopathological and biochemical evaluations. Results: Renal IR resulted in increased malondialdehyde and nitrite/nitrate levels suggesting increased lipid peroxidation and peroxynitrite production and decreased superoxide dismutase and glutathione peroxidase activities. Both NAC and E alone significantly decreased malondialdehyde and nitrite/nitrate levels and increased superoxide dismutase and glutathione peroxidase activities. Additionally in the renal IRI+NAC+E group, all biochemical results were quite close to those of sham group. Histopathologically, the kidney injury in rats treated with combination of NAC and E was found significantly less than the other groups. Conclusions: Both NAC and E are able to ameliorate IRI of the kidney by decreasing oxidative and nitrosative stresses and increasing free radical scavenger properties. Additionally, combination of NAC and E prevents kidney damage more than when each drug is used alone, suggesting that scavenging peroxynitrite nearby antioxidant activity is important in preventing renal IRI.

Protective Effects of Valproic Acid Against Hypoxic-Ischemic Brain Injury in Neonatal Rats

Although controversial, protective and therapeutic effects of valproic acid in various types of cellular injury suggest a potential role for this agent in hypoxic-ischemic brain injury. We therefore investigated the effects of valproic acid in an experimental model of neonatal hypoxic-ischemic brain injury. To examine the effect of valproic acid in this condition, hypoxic-ischemic brain injury was induced in 7-day-old rat pups by ligation of the right common carotid and then the pups were exposed to 1 hour of hypoxia in 8% oxygen. Low (200 mg/kg/day) and high (400 mg/kg/day) doses of valproic acid were administered in a 5-day regimen. Neuropathologic evaluation was performed using the hippocampus, cerebral cortex, and basal ganglia in the coronal plane. The 5-day regimen of valproic acid administration resulted in some protective and therapeutic effects on the brain damage and neuronal apoptosis in both hemispheres in a dose-dependent manner. Administration of valproic acid also decreased the percentage of apoptotic neurons in the contralateral hemisphere (P < .05). These results suggest that valproic acid can have therapeutic and protective effects in hypoxic-ischemic brain injury. (J Child Neurol 2005;20:582—587).