Assessment of cyclooxygenase inhibitors using in vitro assay systems

Various inhibitors of cyclooxygenase are known to mediate cancer chemopreventive effects. We currently describe two in vitro assay systems for measuring cyclooxygenase activity. These assays can be used in combination and thereby provide a rapid, reliable, and economical approach that is applicable for large-scale evaluation of test samples. This approach employs peroxidase co-substrate oxidation and oxygen consumption assays. The former system, adapted to a 96-well plate format, detects inhibitors that function as a radical scavengers or interact with the enzyme directly. The latter system specifically monitors cyclooxygenase inhibitors that interact with the enzyme itself. Thus, the peroxidase co-substrate oxidation assay serves as a pre-screening method, whereas the oxygen consumption assay is used subsequently to investigate the mode of action mediated by samples which test positive.     

A model of corrective gene transfer in X-linked ichthyosis

Single gene recessive genetic skin disorders offer attractive prototypes for the development of therapeutic cutaneous gene delivery. We have utilized X-linked ichthyosis (XLI), characterized by loss of function of the steroid sulfatase arylsulfatase C (STS), to develop a model of corrective gene delivery to human skin in vivo. A new retroviral expression vector was produced and utilized to effect STS gene transfer to primary keratinocytes from XLI patients. Transduction was associated with restoration of full-length STS protein expression as well as steroid sulfatase enzymatic activity in proportion to the number of proviral integrations in XLI cells. Transduced and uncorrected XLI keratinocytes, along with normal controls, were then grafted onto immunodeficient mice to regenerate full thickness human epidermis. Unmodified XLI keratinocytes regenerated a hyperkeratotic epidermis lacking STS expression with defective skin barrier function, effectively recapitulating the human disease in vivo. Transduced XLI keratinocytes from the same patients, however, regenerated epidermis histologically indistinguishable from that formed by keratinocytes from patients with normal skin. Transduced XLI epidermis demonstrated STS expression in vivo by immunostaining as well as a normalization of histologic appearance at 5 weeks post-grafting. In addition, transduced XLI epidermis demonstrated a return of barrier function parameters to normal. These findings demonstrate corrective gene delivery in human XLI patient skin tissue at both molecular and functional levels and provide a model of human cutaneous gene therapy.      

Induction of B cell apoptosis by co-cross-linking CD23 and sIg involves aberrant regulation of c-myc and is inhibited by bcl-2

A novel system to study the effects of co-cross-linking CD23/FceRII and sIg on murine B lymphocytes utilizes a highly multivalent form of anti- Ig prepared by covalently linking anti-Ig antibodies to a DNP-dextran backbone. CD23-sIg co-cross-linking is accomplished by the addition of DNP-specific monoclonal IgE. Previous studies demonstrated that co- cross-linking CD23 and sIg significantly inhibited mouse B cell proliferation, especially at high doses of the multivalent anti-Ig. Interestingly, examination of early activation signals reveals no difference in B cells subjected to co-cross-linking conditions as compared to B cells activated with anti-Ig alone. Total cellular protein tyrosine phosphorylation levels are unchanged by co-cross- linking. Analysis of B cell mRNA reveals that co-cross-linking the receptors does not alter the expression levels of ornithine decarboxylase 8 h after stimulation as compared to the controls. In contrast, levels of the proto-oncogene c-myc were significantly elevated 1 h after inducing B cell activation under co-cross-linking conditions. However, it remains unclear whether this aberrant c-myc regulation plays any role in inducing apoptosis. In addition, on day 3 after stimulation, the co-cross-linking of CD23 and sIg resulted in the formation of apoptotic B cells, determined by both photomicroscopy of the B cell cultures and FACS analysis of B cell nuclei. B cells obtained from bcl-2 transgenic mice proliferated as well as controls, and failed to undergo apoptosis when CD23 and sIg were co-cross-linked on their surface. These studies indicate that co-cross-linking of CD23 with B cell sIg inhibits B cell proliferation by a mechanism that is distinct from that seen by co-cross-linking of the Fc gamma RII and sIg. In addition, these results suggest a means by which antigen- specific IgE can down-regulate additional B cell activation and IgE synthesis.      

Malgun (clear) cell change in Helicobacter pylori gastritis reflects epithelial genomic damage and repair

Cancers may develop in the background of genomic instability with accumulated mutations. Helicobacter pylori gastritis is characterized by acute foveolitis of the proliferative zone, which is found in any stage of the gastritis as long as the infection persists. Because acute foveolitis targets specifically the proliferative zone of pits, the proliferating epithelial cells are under severe and persistent mutagenic pressure. In H. pylori gastritis, a characteristic morphological change of epithelial cells, the malgun (clear) cell change is frequently present in association with acute foveolitis. Malgun cells have enlarged euchromatic nuclei and abundant cytoplasm. The expression of proliferating cell nuclear antigen and cytokeratin 8 are typically up-regulated in them indicating that they are mitotically and metabolically active. Here, we report evidence for DNA damage and repair in malgun cells. Significant double-strand DNA breaks were shown by the consistent terminal dUTP nick-end labeling in the nuclei of malgun cells. Proteins related to DNA damage and repair, such as Ku, poly(ADP-ribosyl) polymerase, OGG1, and MSH2 were selectively up-regulated in malgun cells. Inducible nitric oxide synthase was also up-regulated. There were occasional bcl2- and p53-expressing cells suggesting that further steps of carcinogenesis took place at the single cell level. Our results suggest that the malgun cell change represents a characteristic morphological sign of cellular genomic damage and repair, and may be implicated in an early stage of carcinogenesis. It is suggested that acute foveolitis of the proliferative zone is a major pathogenetic step of gastric carcinogenesis in H. pylori gastritis.     

Alterations in extracellular matrix components in transplant glomerulopathy

The distribution pattern of extracellular matrix (ECM) components in transplant glomerulopathy was studied in relation to light microscopic features, actin expression of mesangial cells, and intraglomerular inflammatory cells. Nine cases of mild (group I) and nine cases of severe (group II) transplant glomerulopathy were stained with antisera against fibronectin (FN), tenascin (TN), collagen types III and IV, smooth muscle actin, CD45RO, CD68, and Ki-67 antigen. The composition of ECM was similar in the two groups. The expanded mesangium was diffusely stained by type-IV collagen, FN and TN, and focally and weakly stained by type-III collagen and smooth muscle actin. Type-IV collagen was linearly stained along the capillary walls, imparting a double-contour feature, whereas FN and TN showed granular staining along the capillary walls. CD68 positive cells were increased in severe transplant glomerulopathy, but this increase was not related to ECM deposition. These findings suggest that increased glomerular deposition of normal and abnormal ECM components participate in the evolution of transplant glomerulopathy. Received: 5 October 1999 / Accepted: 17 January 2000     

First serologic evidence of human spotted fever group rickettsiosis in Korea

To investigate the prevalence of spotted fever group rickettsioses in Korea, a serosurvey of Japanese spotted fever rickettsiosis in patients with acute febrile illness was conducted with an indirect immunofluorescence assay. Overall, 19.88% of the patients were found to have polyvalent antibody against Rickettsia japonica. This study is the first documentation of spotted fever group rickettsiosis in Korea.     

医药类专业本科物理课程的设置及目标与定位探讨

目的：探讨物理课程在高等医药类专业教育中的作用与地位。方法：对全国部分城市不同类别医院从业医务工作者进行调查，综合国际医学教育标准和我国现实医学教育以及医疗服务行业状况进行分析。结果：探讨了我国高等医药类专业教育中物理课程的设置及目标与定位问题，给出了明确的课程定位与设置标准。结论：现代医学高等教育不应忽视或淡化物理课程在医学人才培养中素质教育和专业水平提高的积极作用与基础地位，在有限的学时空间内合理安排基础性内容和与专业素质培养相关的应用内容。     

Different modifying responses of capsaicin in a wide-spectrum initiation model of F344 rat

The modifying potential of capsaicin (CAP) on lesion development was examined in a rat multiorgan carcinogenesis model. Groups 1 and 2 were treated sequentially with diethylnitrosamine (DEN) (100 mg/kg, ip, single dose at commencement), N-methylnitrosourea (MNU) (20 mg/kg, ip, 4 doses at days 2, 5, 8, and 11), and N,N-dibutylnitrosamine (DBN) (0.05% in drinking water during weeks 3 and 4). Group 3 received vehicles without carcinogens during the initiation period. Group 4 served as the untreated control. After this initiating procedure, Groups 2 and 3 were administered a diet containing 0.01% CAP. All surviving animals were killed 20 weeks after the beginning of the experiment and the target organs examined histopathologically. The induction of GST-P+ hepatic foci in rats treated with carcinogens was significantly inhibited by treatment with CAP. CAP treatment significantly decreased the incidence of adenoma of the lung but increased the incidence of papillary or nodular (PN) hyperplasia of the urinary bladder. The tumor incidence of other organs, such as the kidney and thyroid, was not significantly different from the corresponding controls. These results demonstrated that concurrent treatment with CAP not only can inhibit carcinogenesis but can also enhance it depending on the organ. Thus, this wide-spectrum initiation model could be used to confirm organ-specific modification potential and, in addition, demonstrate different modifying effects of CAP on liver, lung, and bladder carcinogenesis.     

Clinical Comparison of Distal Pancreatectomy with or without Splenectomy

The spleen may be preserved during distal pancreatectomy (DP) for benign disease. The aim of this study was to compare the perioperative and postoperative courses of patients with conventional DP and spleen-preserving distal pancreatectomy (SPDP) for benign lesions or tumors with low-grade malignant potential occurred at the body or tail of the pancreas. A retrospective analysis was performed for the hospital records of all the patients undergoing DP and SPDP between January 1995 and April 2006. One-hundred forty-three patients underwent DP and 37 patients underwent SPDP. There were no significant differences in age, sex, indications of operation, estimated blood loss, operative time, and postoperative hospital stay between the two groups. Pancreatic fistula occurred in 21 (13.3%) patients following DP and in 3 (8.1%) following SPDP without a significant difference (p=0.081). Portal vein thrombosis occurred in 4 patients after DP. Splenic infarction occurred in one patient after SPDP. Overwhelming postosplenectomy infection was observed in one patient after DP. SPDP can be achieved with no increase in complication rate, operative time, or length of postoperative hospitalization as compared to conventional DP. Additionally, it has the advantage of reducing the risk of overwhelming postsplenectomy infection and postoperative venous thrombosis.     

Clinical prognostic values of vascular endothelial growth factor, microvessel density,and p53 expression in esophageal carcinomas

Vascular endothelial growth factor (VEGF) is known to play a key role in tumor angiogenesis. The tumor-suppressor gene p53 has been thought to regulate VEGF. We investigated the effect of VEGF on esophageal carcinoma and the correlation between VEGF and p53. Tissue samples were taken from 81 patients with esophageal carcinoma after surgery. VEGF and p53 expressions were examined by immunohistochemical staining. Microvessels in the tumor stained for CD34 antigen were also counted. VEGF and p53 expressions were observed in 51.3% (41/80) and 51.9% (41/79), respectively. The microvessel density was 70.9+/-6.7 (mean+/-SE) in VEGF-positive group and 68.7+/-5.1 in VEGF-negative group. However, no correlation was noted between VEGF and p53 expression. Whereas the tumor size, nodal status, depth of invasions, and tumor stage were associated with poor overall survival, VEGF expression or p53 expression was not. These results indicate that VEGF and p53 are highly expressed in esophageal carcinomas. Since the VEGF expression is not correlated with the p53 expression, microvessel density or clinicopathological findings, further studies with other angiogenic molecules are needed to determine the role in esophageal carcinomas.