In Vitro and in Vivo Antitumor Effects of Plant-Derived Miliusanes and Their Induction of Cellular Senescence.

Our earliest phytochemical separation of Miliusa sinensis aided us in the isolation of a class of unique miliusanes, which were demonstrated as anticancer lead molecules. In the present study, we isolated 19 miliusanes (1-19), including 11 novel ones (5 and 10-19) from another Miliusa plant ( M. balansae), and synthesized additional derivatives to elucidate the structure-activity relationship of miliusanes. When extrapolated to various carcinoma xenograft mouse models, miliusol (1) and its derivatives 20, 26, and 27 (7.5-40 mg/kg) were demonstrated with tumor inhibitory efficacy comparable or even superior to the mainstay chemotherapeutics paclitaxel or fluorouracil. To gain a molecular insight into their anticancer mechanism, 1-3 (GI50 0.03-4.79) were administered to a wide spectrum of human cancer cell lines, including those with specific drug resistance. We further revealed that the antiproliferative properties of miliusanes in carcinoma cells were highly associated with the p21-dependent induction of cellular senescence.

Potent Inhibitor of Drug-Resistant HIV-1 Strains Identified from the Medicinal Plant Justicia gendarussa.

Justicia gendarussa, a medicinal plant collected in Vietnam, was identified as a potent anti-HIV-1 active lead from the evaluation of over 4500 plant extracts. Bioassay-guided separation of the extracts of the stems and roots of this plant led to the isolation of an anti-HIV arylnaphthalene lignan (ANL) glycoside, patentiflorin A (1). Evaluation of the compound against both the M- and T-tropic HIV-1 isolates showed it to possess a significantly higher inhibition effect than the clinically used anti-HIV drug AZT. Patentiflorin A and two congeners were synthesized, de novo, as an efficient strategy for resupply as well as for further structural modification of the anti-HIV ANL glycosides in the search for drug leads. Subsequently, it was determined that the presence of a quinovopyranosyloxy group in the structure is likely essential to retain the high degree of anti-HIV activity of this type of compounds. Patentiflorin A was further investigated against the HIV-1 gene expression of the R/U5 and U5/gag transcripts, and the data showed that the compound acts as a potential inhibitor of HIV-1 reverse transcription. Importantly, the compound displayed potent inhibitory activity against drug-resistant HIV-1 isolates of both the nucleotide analogue (AZT) and non-nucleotide analogue (nevaripine). Thus, the ANL glycosides have the potential to be developed as novel anti-HIV drugs.

Efficient Semisynthesis of (-)-Pseudoirroratin A from (-)-Flexicaulin A and Assessment of Their Antitumor Activities.

Accumulating evidence indicates that natural ent-kaurane diterpenoids show great potential for medical treatment of different pathological conditions including cytotoxicity, antibacterial, and anti-inflammatory activity. Among a variety of diterpenoids tested, (-)-pseudoirroratin A displayed a promising antitumor property in vitro and in vivo. However, this diterpenoid could merely be isolated in a limited amount from a rare source of Isodon pseudoirrorata. To overcome such scanty source, we developed a novel, facile, and efficient semisynthetic strategy to prepare (-)-pseudoirroratin A from natural (-)-flexicaulin A, which can be expediently obtained from I. flexicaulis in a great quantity. The three-dimensional structure and the absolute configuration of our synthetic diterpenoid have been determined and confirmed with the X-ray crystallographic analysis. More importantly, we demonstrated for the first time that pseudoirroratin A exerted significant cytotoxicity against human colorectal carcinoma cells via an induction of apoptosis, as well as a remarkable suppression on tumor growth in a colon cancer xenograft mouse model.

Anti-HIV diphyllin glycosides from Justicia gendarussa.

In a search for new anti-HIV active leads from over several thousands of plant extracts, we have identified a potent plant lead. The active plant is determined as Justicia gendarussa (Acanthaceae), a medicinal plant that has been used for the treatment of injury, arthritis and rheumatism in Asia including China. Our bioassay-guided fractionation of the methanol extract of the stems and barks of the plant led to the isolation of two anti-HIV compounds, justiprocumins A and B. The compounds are identified as new arylnaphthalide lignans (ANL) glycosides. We further determined that the ANL glycosides are the chemical constituents that contribute to the anti-HIV activity of this plant. Justiprocumin B displayed potent activity against a broad spectrum of HIV strains with IC50 values in the range of 15-21 nM (AZT, IC50 77-95 nM). The compound also displayed potent inhibitory activity against the NRTI (nucleoside reverse transcriptase inhibitor)-resistant isolate (HIV-11617-1) of the analogue (AZT) as well as the NNRTI (non-nucleoside reverse transcriptase inhibitor)-resistant isolate (HIV-1N119) of the analogue (nevaripine).

Discovery of antifungal constituents from the Miao medicinal plant Isodon flavidus.

ETHNOPHARMACOLOGICAL RELEVANCE: Leigong Mountain is an area in the Southwest of China where there is a high incidence rate of athlete's foot, but the Miao people, a Chinese minority who reside in this mountainous area have suffered less from this disease due to their use of the herbal medicine Isodon flavidus (Hand.-Mazz.) H. Hara. AIM OF THE STUDY: The present study is to identify the active chemical constituents responsible for antifungal effects of the folk medicine plant. MATERIALS AND METHODS: The natural compounds were separated from the methanol extract of the twigs and leaves of I. flavidus by phytochemical study using chromatographic methods, and their chemical structures were determined by analysis of the spectroscopic data including 1D and 2D NMR spectra. The absolute configuration of fladin A (1) was further confirmed by X-ray crystallographic analysis. The compounds were evaluated for their antifungal activity against the athlete's foot fungus Trichophyton rubrum. They were further evaluated for their antimicrobial and anti-biofilm activity against the dental pathogens Streptococcus mutans, Porphyromonas gingivalis and Candida albicans. RESULTS: Phytochemical and biological studies of I. flavidus led to the discovery of two antifungal compounds, fladin A (1) and lophanic acid (2). Fladin A (1) is a novel diterpene with an unprecedented cyclic ether group formed between C-4 and C-9. Lophanic acid (2) displayed inhibition activity against the athlete's foot fungus Trichophyton rubrum with an MIC value of 7.8µg/mL, and fladin A (1) also showed inhibition activity against the fungus with a MIC value of 62.5µg/mL. CONCLUSIONS: Our identification of two antifungal compounds provided strong evidence for the Miao people to use I. flavidus as a medicinal plant for treatment of athlete's foot disease. The very different chemical structures of the active compounds from those in the market presents them as potential antifungal lead compounds for follow-up study.

Bioassay-Guided Isolation and Structural Modification of the Anti-TB Resorcinols from Ardisia gigantifolia.

Tuberculosis (TB) is a highly contagious disease mainly caused by Mycobacterium tuberculosis H37 RV . Antitubercular (anti-TB) bioassay-guided isolation of the CHCl3 extract of the leaves and stems of the medicinal plant Ardisia gigantifolia led to the isolation of two anti-TB 5-alkylresorcinols, 5-(8Z-heptadecenyl) resorcinol (1) and 5-(8Z-pentadecenyl) resorcinol (2). We further synthesized 15 derivatives based on these two natural products. These compounds (natural and synthetic) were evaluated for their anti-TB activity against Mycobacterium tuberculosis H37 RV . Resorcinols 1 and 2 exhibited anti-TB activity with MIC values at 34.4 and 79.2 µm in MABA assay, respectively, and 91.7 and 168.3 µm in LORA assay, respectively. Among these derivatives, compound 8 was found to show improved anti-TB activity than its synthetic precursor (2) with MIC values at 42.0 µm in MABA assay and 100.2 µm in LORA assay. The active compounds should be regarded as new hits for further study as a novel class of anti-TB agents. The distinct structure-activity correlations of the parent compound were elucidated based on these derivatives.

Inhibition of pancreatic oxidative damage by stilbene derivative dihydro-resveratrol: implication for treatment of acute pancreatitis.

Trans-resveratrol is a natural stilbenoid possessing multifarious pharmacological benefits; however, when orally consumed, it is rapidly metabolised by colonic microflora and converted to dihydro-resveratrol. Thus, this microbial metabolite is of great therapeutic relevance. In the present study, upon the oral administration of dihydro-resveratrol (10-50 mg/kg), the severity of acute pancreatitis in the cerulein-treated rats was significantly ameliorated as evidenced by decreased α-amylase activities in the plasma and lessened oedema formation in the pancreatic parenchyma. In addition, the generation of intracellular reactive oxidative products, including malondialdehyde and protein carbonyls, was accordingly reduced, so as the production of pro-inflammatory cytokines. While inhibiting the activities of NADPH oxidase and myeloperoxidase, the depletion of glutathione was considerably restored. Importantly, the attenuation of pancreatic oxidative damage by dihydro-resveratrol was associated with a down-regulation of the nuclear factor-kappaB and phosphatidylinositol 3'-kinase-serine/threonine kinase signalling pathways. Furthermore, we demonstrated that the solubility of dihydro-resveratrol was at least 5 times higher than trans-resveratrol whilst exhibiting a much lower cytotoxicity. Collectively, the current findings accentuate new mechanistic insight of dihydro-resveratrol in pancreatic oxidative damage, and advocate its therapeutic potential for the management of acute pancreatitis, particularly for patients unresponsive to trans-resveratrol due to the lack of proper microbial strains.

Dihydro-Resveratrol Ameliorates Lung Injury in Rats with Cerulein-Induced Acute Pancreatitis.

Acute pancreatitis is an inflammatory process originated in the pancreas; however, it often leads to systemic complications that affect distant organs. Acute respiratory distress syndrome is indeed the predominant cause of death in patients with severe acute pancreatitis. In this study, we aimed to delineate the ameliorative effect of dihydro-resveratrol, a prominent analog of trans-resveratrol, against acute pancreatitis-associated lung injury and the underlying molecular actions. Acute pancreatitis was induced in rats with repetitive injections of cerulein (50 µg/kg/h) and a shot of lipopolysaccharide (7.5 mg/kg). By means of histological examination and biochemical assays, the severity of lung injury was assessed in the aspects of tissue damages, myeloperoxidase activity, and levels of pro-inflammatory cytokines. When treated with dihydro-resveratrol, pulmonary architectural distortion, hemorrhage, interstitial edema, and alveolar thickening were significantly reduced in rats with acute pancreatitis. In addition, the production of pro-inflammatory cytokines and the activity of myeloperoxidase in pulmonary tissues were notably repressed. Importantly, nuclear factor-kappaB (NF-κB) activation was attenuated. This study is the first to report the oral administration of dihydro-resveratrol ameliorated acute pancreatitis-associated lung injury via an inhibitory modulation of pro-inflammatory response, which was associated with a suppression of the NF-κB signaling pathway.

Inhibition of glucose-transporter 1 (GLUT-1) expression reversed Warburg effect in gastric cancer cell MKN45.

Glucose transporter-1 (GLUT-1) plays critical roles in cancer development and progression. Warburg effect (aerobic glycolysis) contributes greatly to tumorigenesis and could be targeted for tumor therapy. However, published data on the relationship between GLUT-1 and Warburg effect are scarce. In this study, gastric cancer cell, MKN45, was transfected with GLUT-1 shRNA using Lipofectamine 2000. Oxygen consumption, LDH activity, lactate production and cytoplasmic pyruvate were detected after MKN45 cells with GLUT-1 knockdown. In the last, hexokinase 1 (HK1), HK2, and pyruvate kinase M2 (PKM2) expression were detected by using western blot. In this study, we showed that inhibition of GLUT-1 expression reversed Warburg effect in MKN45 cells, and induced apoptosis.

Determination of base binding strength and base stacking interaction of DNA duplex using atomic force microscope.

As one of the most crucial properties of DNA, the structural stability and the mechanical strength are attracting a great attention. Here, we take advantage of high force resolution and high special resolution of Atom Force Microscope and investigate the mechanical force of DNA duplexes. To evaluate the base pair hydrogen bond strength and base stacking force in DNA strands, we designed two modes (unzipping and stretching) for the measurement rupture forces. Employing k-means clustering algorithm, the ruptured force are clustered and the mean values are estimated. We assessed the influence of experimental parameters and performed the force evaluation for DNA duplexes of pure dG/dC and dA/dT base pairs. The base binding strength of single dG/dC and single dA/dT were estimated to be 20.0 ± 0.2 pN and 14.0 ± 0.3 pN, respectively, and the base stacking interaction was estimated to be 2.0 ± 0.1 pN. Our results provide valuable information about the quantitative evaluation of the mechanical properties of the DNA duplexes.

BRCA1 regulates insulin-like growth factor 1 receptor levels in ovarian cancer.

Breast cancer 1 (BRCA1) and insulin-like growth factor 1 receptor (IGF1R) are critical in ovarian cancer progression. However, the crosstalk between the BRCA1 and IGF1R signaling pathways in ovarian cancer remains largely unknown. The effects of BRCA1 on IGF1R were assessed in 121 serous ovarian cancer patients (BRCA1 mutation, n=30; non-BRCA1 mutation, n=32; hypermethylated BRCA1 promoter, n=28; and non-methylation, n=31). BRCA1 promoter methylation was analyzed via bisulfite sequencing using primers focused on the core promoter region. The expression levels of BRCA1 and IGF1R were assessed by immunohistochemistry and real-time polymerase chain reaction. Knockdown and overexpression of BRCA1 were achieved using a lentiviral vector in 293T and SKOV3 ovarian cancer cells, and primary non-mutated and BRCA1-mutated ovarian cancer cells. The present study demonstrated that IGF1R expression is increased in non-BRCA1-mutated ovarian cancer when compared with adjacent normal tissue. Furthermore, IGF1R levels are additionally significantly elevated in BRCA1 inactivation ovarian cancer (BRCA1 mutation or hypermethylated BRCA1 promoter). In addition, BRCA1 knockdown was found to be an effective method of activating IGF1R expression in non-BRCA1-mutated ovarian cancer cells. The observations of the current study indicate that BRCA1 may be a potential trigger that is involved in the transcriptional regulation of IGF1R in the development of ovarian cancer.

The pathobiological behaviors and prognosis associated with Japanese gastric adenocarcinomas of pure WHO histological subtypes.

Japan is a high-risk region for gastric carcinoma with a comparatively early stage and favorable prognosis. To clarify the pathobiological behaviors and prognosis of Japanese gastric adenocarcinoma, we analyzed the clinicopathological characteristics of different WHO subtypes of carcinomas. The expression of ki-67, CPP32, p53, FHIT, maspin, parafibromin, GRP78, GRP94, EMMPRIN, VEGF, P-GSK3beta-ser9, fascin, cortactin, Arp2, Arp3 MUC-2, MUC-5AC and MUC-6 was examined using immunohistochemistry and tissue microarrays. The majority of cases were well-, poorly-, or moderately-differentiated subtype, whereas the minority were papillary or signet ring cell carcinoma (SRC). Patients with poorly-differentiated or SRC carcinoma were predominantly young and female. Poorly-differentiated and mucinous carcinomas were larger, with deeper invasion, more venous or lymphatic invasion, frequent lymph node involvement and peritoneal dissemination, or higher staging. The SRC group exhibited weaker expression of ki-67, CPP32, p53, parafibromin, GRP78, GRP94, P-GSK3beta-ser9, VEGF or cortactin. The moderately-differentiated subtype exhibited lower expression of FHIT and Arp3 positivity. The poorly-differentiated group showed weaker expression of CPP32, EMMPRIN, MUC-2, MUC-5AC, and MUC-6. Survival analysis indicated that the patients with poorly-differentiated or mucinous subtypes had a lower cumulative survival rate than those with papillary, well-, moderately-differentiated, or SRC carcinomas (P<0.05). The age, invasive depth, lymphatic invasion, peritoneal dissemination, and WHO classification were independent prognostic factors for carcinoma patients (P<0.05). It was suggested that poorly-differentiated and mucinous subtypes are more aggressive and of unfavorable prognosis among Japanese gastric carcinomas. Lower levels of proliferation and apoptosis, as well as alterations in tumor suppressor genes, mucin production and ER stress protein played important roles in the pathogenesis of poorly-differentiated and SRC carcinomas.

Mapping the history and current situation of research on John Cunningham virus - a bibliometric analysis.

BACKGROUND: John Cunningham virus (JCV) constitutes a family of polyoma viruses, which plays important roles in the progressive multifocal leukoencephalopathy (PML) and tumorigenesis. However, no bibliometric investigation has been reported to guide the researchers and potential readers. METHODS: Papers were collected from database Sci-expanded and Pubmed until May 22, 2008. The highly-productive authors, institutes and countries, highly-cited authors and journals were ranked. The highly-cited articles were subjected to co-citation and chronological analysis with highly-frequent MeSH words for co-occurrence analysis. RESULTS: Until now, 1785 articles about JCV were indexed in Sci-expanded and 1506 in Pubmed. The main document type was original article. USA, Japan and Italy were the largest three producers about JCV. Temple University published 128 papers and ranked the top, followed by University of Tokyo. Khalili K and Yogo Y became the core authors due to more than 20 documents produced. Journal of Neurovirology published more than 15 papers and ranked the top. Padgett BL and Berger JR were the first two highly-cited authors. Journal of Virology and Journal of Neurovirology respectively ranked to the first two highly-cited journals. These top highly-cited articles were divided into 5 aspects: (1) The correlation between JC virus and tumors; (2) Causal correlation of JCV with PML; (3) Polyoma virus infection and its related diseases in renal-allograft recipients; (4) Detection of JCV antibody, oncogene and its encoding protein; (5) Genetics and molecular biology of JCV. The MeSH/subheadings were classified into five groups: (1) JCV and virus infectious diseases; (2) JCV pathogenicity and pathological appearance of PML; (3) JCV isolation and detection; (4) Immunology of JCV and PML; (5) JCV genetics and tumors. CONCLUSION: JCV investigation mainly focused on its isolation and detection, as well as its correlation with PML and tumors. Establishment of transgenic animal model using JCV T antigen would be a hopeful and useful project in the further study.

Arp2/3 overexpression contributed to pathogenesis, growth and invasion of gastric carcinoma.

BACKGROUND: Tumor metastasis depends on cell adhesion, motility and deformability, resulting from quantitative alterations and rearrangement of actin-related protein (Arp) 2 and 3. The aim of this study was to clarify the roles of both molecules in tumorigenesis and progression of gastric carcinomas. PATIENTS AND METHODS: Immunohistochemistry (IHC) was employed on tissue microarray containing gastric carcinomas, adjacent metaplasia and gastritis using antibodies against Arp2 and Arp3 with a comparison of their expression with clinicopathological parameters of carcinomas. Gastric carcinoma cell lines (MKN28, AGS, MKN45, KATO-III and HGC-27) were studied for Arp2 and Arp3 protein by IHC. RESULTS: Both proteins were expressed at low levels in gastritis compared with carcinomas (p < 0.05). Arp2 was more frequently expressed in intestinal metaplasia than in carcinoma and gastritis (p < 0.05). Most gastric carcinoma cell lines showed expression at different levels. Expression was positively correlated with tumor size, depth of invasion, venous invasion, Union Internationale Contre le Cancer (UICC) staging and expression of cortactin or fascin (p < 0.05), but not with age, sex, lymphatic invasion or lymph node metastasis (p > 0.05). There was stronger positivity of Arp3 in intestinal- than diffuse-type carcinomas (p < 0.05). A positive relationship between Arp2 and Arp3 proteins was noted (p < 0.05). Univariate analysis indicated that the cumulative survival rate of patients with positive Arp2 or Arp3 expression was not different from those without their expression (p > 0.05). Multivariate analysis showed that age, depth of invasion, lymphatic invasion, lymph node metastasis, UICC staging and Lauren's classification were independent prognostic factors for carcinomas (p < 0.05). CONCLUSION: Aberrant expression of Arp2 and Arp3 is possibly involved in pathogenesis, growth, invasion and progression of gastric carcinomas. Distinct Arp3 expression underlies the molecular mechanisms for the differentiation of intestinal- and diffuse-type carcinomas. They were considered as objective and effective markers to indicate the pathobiological behaviors of gastric carcinomas.

Overexpression of GRP78 and GRP94 are markers for aggressive behavior and poor prognosis in gastric carcinomas.

Glucose-related proteins (GRPs) are ubiquitously expressed in endoplasmic reticulum and able to assist in protein folding and assembly; consequently, they are considered as molecular chaperones. GRP78 and GRP94 expression was induced by glucose starvation and up-regulated in the malignancies. To clarify the roles of both molecules in tumorigenesis and progression of gastric carcinomas, immunohistochemistry was used on tissue microarray containing gastric carcinomas, adenomas, and nonneoplastic mucosa using the antibodies against GRP78 and GRP94, with a comparison of their expression with clinicopathological parameters of carcinomas. Gastric carcinoma cell lines (MKN28, AGS, MKN45, KATO-III, and HGC-27) were studied for both proteins by immunohistochemistry and Western blot. There was more expression of both proteins in gastric carcinoma and adenoma than in nonneoplastic mucosas (P < .05). All gastric carcinoma cell lines showed their expression at different levels. They were positively correlated with tumor size, depth of invasion, lymphatic and venous invasion, lymph node metastasis, and Union Internationale Contre le Cancer staging (P < .05), with positive relationship between both proteins (P < .05). Univariate analysis indicated the postsurgical cumulative survival rate of patients with positive GRP78 or GRP94 expression to be lower than that in those without GRP78 or GRP94 expression (P < .05), but the close link disappeared if stratified according to depth of invasion (P > .05). Multivariate analysis showed that age, depth of invasion, lymphatic invasion, lymph node metastasis, Union Internationale Contre le Cancer staging, and Lauren classification (P < .05), but not GRP78 and GRP94 expression, were independent prognostic factors for carcinomas (P > .05). Up-regulated expression of GRP78 and GRP94 was possibly involved in pathogenesis, growth, invasion, and metastasis of gastric carcinomas. They were considered objective and effective markers for the aggressive behavior and poor prognosis in gastric carcinomas.

The clinicopathological and prognostic significance of MUC-1 expression in Japanese gastric carcinomas: an immunohistochemical study of tissue microarrays.

BACKGROUND: MUC-1 is synthesized as a single polypeptide that then undergoes proteolytic cleavage, and is associated with the epidermal growth factor receptor tyrosine kinases. In malignancies, MUC-1 may function as an anti-adhesion molecule, but can also promote adhesion and presumably metastasis. MATERIALS AND METHODS: Expression of MUC-1, -2, -4 and -5AC was evaluated on tissue microarrays of gastric carcinomas (n = 237) and adjacent non-cancerous mucosa specimens (n = 89) by immunohistochemistry and compared with clinicopathological parameters and survival time of the patients. RESULTS: MUC-1 was found to be highly expressed in gastric carcinomas in comparison with noncancerous mucosa (p < 0.05) and positively correlated with depth of invasion, lymphatic and venous invasion, lymph node metastasis, UICC staging and MUC-4 expression (p < 0.05), but not with age, tumor size, MUC-2 or MUC-5AC expression (p > 0.05). Intestinal-type carcinomas showed more MUC-1 expression than their diffuse-type counterparts (p < 0.05). Kaplan-Meier analysis indicated that the cumulative survival rate of patients with no MUC-1 expression was significantly higher than those with weak, moderate or strong expression in gastric carcinomas (p < 0.05), but no difference was observed when tumors were stratified according to the depth of invasion (p > 0.05). Cox's analysis showed three independent prognostic factors, depth of invasion, lymphatic invasion and venous invasion, to affect the relationship between MUC-1 expression and prognosis. CONCLUSION: Up-regulation of MUC-1 expression may be involved in pathogenesis, invasion, metastasis and differentiation of gastric carcinoma. Altered expression might therefore be employed as an indicator of pathobiological behavior of gastric carcinoma. MUC-1 expression was found to be a prognostic factor for gastric carcinoma patients, albeit not independent of parameters of invasion.

Mixed-type gastric carcinomas exhibit more aggressive features and indicate the histogenesis of carcinomas.

To investigate the pathobiological behaviors of gastric mixed-type (MT) carcinomas and gastric carcinogenesis, the clinicopathological characteristics of MT carcinomas were analyzed and compared with intestinal-type (IT) and diffuse-type (DT) carcinomas. The expression of Ki-67, caspase-3, p53, fragile histine triad (FHIT), maspin, extracellular matrix metalloproteinase inducer (EMMPRIN), vascular growth factor (VEGF), MUC-2, 4, 5AC and 6, CD44, E-cadherin, beta-catenin, and phosphorylated glycogen synthase kinase 3beta-ser9 (P-GSK3beta-ser9) was examined on tissue microarrays using immunohistochemistry. It was found that MT carcinomas exhibited large size, deep invasion, frequent local invasion, and lymph node metastasis in comparison with IT and DT carcinomas (p < 0.05). All the markers except MUC-5AC showed higher expression in IT than DT carcinomas (p < 0.05). The expression of maspin, EMMPRIN, VEGF, MUC-4, and membrane E-cadherin was stronger in MT intestinal than diffuse component (p < 0.05). Immunoreactivities to Ki-67, EMMPRIN, and VEGF were weaker in IT carcinoma than in the MT intestinal portion (p < 0.05), while the opposite was true for CD44, MUC-2, and MUC-6 (p < 0.05). The MT diffuse component displayed a higher expression of FHIT, VEGF, and P-GSK3beta-ser9 than DT carcinoma (p < 0.05). The accumulative survival rate of the IT carcinoma patients was higher than the other types (p < 0.05). The invasive depth, venous invasion, lymph node, peritoneal or liver metastasis, and Lauren's classification were independent prognostic factors for gastric carcinomas (p < 0.05). These findings suggested that MT carcinomas were also indicated to be more aggressive than IT and DT carcinomas. Significant differences were observed in the proliferation, apoptosis, angiogenesis, mucin secretion, and cell adhesion between IT and DT carcinomas, whereas only a few of these characteristics showed differences between the MT intestinal and diffuse parts, thus suggesting that both the MT components might originate from the stem cells with similar genetic traits, but follow different histogenic pathways.

Downregulated parafibromin expression is a promising marker for pathogenesis, invasion, metastasis and prognosis of gastric carcinomas.

Parafibromin is a protein encoded by the hyperparathyroidism 2 oncosuppressor gene and its downregulated expression is involved in pathogenesis of parathyroid carcinomas. To clarify the roles of parafibromin expression in tumourigenesis and progression of gastric carcinomas, it was examined by immunohistochemistry (IHC) on tissue microarray containing gastric carcinomas (n = 508), adenomas (n = 45) and gastritis (n = 49) with a comparison of its expression with clinicopathological parametres of carcinomas. Gastric carcinoma cell lines (MKN28, AGS, MKN45, KATO-III and HGC-27) were studied for parafibromin expression by IHC and western blot. Parafibromin expression was localised in the nucleus of gastric epithelial cells, adenoma, carcinoma cells and cell lines. Its expression was gradually decreased from gastritis to gastric carcinoma, through gastric adenomas (p < 0.05) and inversely correlated with tumour size, depth of invasion, lymphatic invasion, lymph node metastasis and Union Internationale Contre le Cancer (UICC) staging (p < 0.05) but not with sex or venous invasion (p > 0.05). Parafibromin was strongly expressed in older carcinoma patients compared with younger ones (p < 0.05). There was stronger positivity of parafibromin in intestinal-type than diffuse-type carcinomas (p < 0.05). Univariate analysis indicated cumulative survival rate of patients with positive parafibromin expression to be higher than without its expression (p < 0.05). Multivariate analysis showed that age, tumour size, depth of invasion, lymphatic invasion, lymph node metastasis, UICC staging and Lauren's classification but not sex, venous invasion or parafibromin expression were independent prognostic factors for carcinomas(p < 0.05). Downregulated parafibromin expression possibly contributed to pathogenesis, growth, invasion and metastasis of gastric carcinomas. It was considered as a promising marker to indicate the aggressive behaviours and prognosis of gastric carcinomas.