Jeng-Dong Hsu

Inhibitory effect of Hibiscus protocatechuic acid on tumor promotion in mouse skin

Hibiscus protocatechuic acid (PCA), a phenolic acid isolated from Hibiscus sabdariffa L., was evaluated for its ability to inhibit the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced promotion in skin tumors of female CD-1 mice. Topical application of PCA (5, 10 or 20 micromol) 5 min prior to TPA (15 nmol) treatment twice weekly for 20 weeks to mice which were initiated with benzo[a]pyrene (B[a]P) inhibited the incidence of tumors in mice to 81.3, 62.5 and 56.3%, respectively, while all mice in the TPA-treated group developed tumors. The average number of tumors in mice pretreated with PCA was 2-4 and that of mice treated only with TPA was 6.6. The protection effects of PCA were also presented by its significant suppression on the TPA-induced hyperplasia in the skin and edema of mouse ears by 65 and 73% at doses of 10 and 20 micromol, respectively. When it was applied to the dorsal surface of CD-1 mice before TPA application, PCA (5, 10 or 20 micromol) inhibited the induction of epidermal ornithine decarboxylase (ODC) activity by 5 nmol TPA and myeloperoxidase (MPO) activity by 6.5 nmol TPA. The same doses of PCA also reduced the formation of hydrogen peroxide in the mouse skin to an inhibition of 61, 84 and 89%, respectively, when compared with that of the TPA-treated group. These results indicate that PCA possesses potential as a cancer chemopreventive agent against tumor promotion.

Scoring mechanisms of p16INK4a immunohistochemistry based on either independent nucleic stain or mixed cytoplasmic with nucleic expression can significantly signal to distinguish between endocervical and endometrial adenocarcinomas in a tissue microarray study

BackgroundEndocervical adenocarcinomas (ECAs) and endometrial adenocarcinomas (EMAs) are malignancies that affect uterus; however, their biological behaviors are quite different. This distinction has clinical significance, because the appropriate therapy may depend on the site of tumor origin. The purpose of this study is to evaluate 3 different scoring mechanisms of p16INK4a immunohistochemical (IHC) staining in distinguishing between primary ECAs and EMAs.MethodsA tissue microarray (TMA) was constructed using formalin-fixed, paraffin-embedded tissue from hysterectomy specimens, including 14 ECAs and 24 EMAs. Tissue array sections were immunostained with a commercially available antibody of p16INK4a. Avidin-biotin complex (ABC) method was used for antigens visualization. The staining intensity and area extent of the IHC reactions was evaluated using the semi-quantitative scoring system. The 3 scoring methods were defined on the bases of the following: (1) independent cytoplasmic staining alone (Method C), (2) independent nucleic staining alone (Method N), and (3) mean of the sum of cytoplasmic score plus nucleic score (Method Mean of C plus N).ResultsOf the 3 scoring mechanisms for p16INK4a expression, Method N and Method Mean of C plus N showed significant (p-values < 0.05), but Method C showed non-significant (p = 0.245) frequency differences between ECAs and EMAs. In addition, Method Mean of C plus N had the highest overall accuracy rate (81.6%) for diagnostic distinction among these 3 scoring methods.ConclusionAccording to the data characteristics and test effectiveness in this study, Method N and Method Mean of C plus N can significantly signal to distinguish between ECAs and EMAs; while Method C cannot do. Method Mean of C plus N is the most promising and favorable means among the three scoring mechanisms.

Specific induction of glutathione S-transferase GSTM2 subunit expression by epigallocatechin gallate in rat liver.

The antitumor effect of green tea polyphenols has been well characterized in numerous papers. However, the mechanism of their action is still poorly defined. In this study, epigallocatechin gallate (EGCG), the main ingredient of green tea extract, was studied for its effect on the expression of glutathione S-transferases (GSTs) in rat liver to examine the mechanism of action. Liver samples were collected from Sprague-Dawley rats treated with EGCG in H(2)O by portal vein perfusion and examined for total GST activity and GST expression. The results showed that the induction of GST activity by EGCG was dose- and time-dependent. GST activity was increased about 28-fold at 12 hr after treatment. Three GST subunits (GSTA1/2, GSTM1, and GSTM2) were examined by Western blot for changes in protein level affected by EGCG (1 mg/kg weight). Only GSTM2 revealed a significant time-dependent increase, with a maximal induction of approximately 2.0-fold. The differential effect of EGCG on GST subunit expression was also verified by immunocytochemical examination and showed strong induction of the GSTM2 (but not the GSTA1/2 and GSTM1) level in liver section. This induction occurred as early as 3 hr after treatment and extended gradually outward from the hepatic veins as treatment time increased. The change in the GSTM2 protein level was accompanied by a corresponding alteration in mRNA quantity ( approximately 2.0-fold of control). Our report is the first to demonstrate a specific induction of the GSTM2 subunit by a chemopreventor and suggests a primary influence of EGCG on GSTM2 gene expression.

Gallic Acid Induces G2/M Phase Arrest of Breast Cancer Cell MCF-7 through Stabilization of p27Kip1 Attributed to Disruption of p27Kip1/Skp2 Complex

Gallic acid (GA), 3,4,5-trihydroxybenzoic acid, is a natural polyphenolic acid and widely found in gallnuts, tea leaves and various fruits. Previous studies have shown that GA possesses anti-inflammatory, antiallergic and anticarcinogenic activity. In the present study, we aim to investigate the antitumor effects of GA on breast cancer cell. Our results revealed that GA treatment significantly reduced the cell growth of human breast cancer cell MCF-7 in a dose-dependent manner. Further flow cytometric analysis showed that GA induced significant G2/M phase arrest but slightly affected the population of sub-G1MCF-7 cells. Therefore, levels of cyclins, cyclin-dependent kinases (CDKs), and their regulatory proteins involved in S-G2/M transition were investigated. Our findings revealed that levels of cyclin A, CDK2, cyclin B1 and cdc2/CDK1 were diminished; in contrast, levels of the negative regulators p27(Kip1) and p21(Cip1) were increased by GA treatment. Additionally, Skp2, a specific ubiquitin E3 ligase for polyubiquitination of p27(Kip1) was reduced by GA treatment. Further investigation showed that GA attenuated Skp2-p27(Kip1) association and diminished polyubiquitination of p27(Kip1) in MCF-7 cells. Moreover, knockdown of p27(Kip1) but not p21(Cip1) significantly alleviated GA-induced accumulation of G2/M phase. These findings indicate that GA may upregulate p27(Kip1) level via disruption of p27(Kip1)/Skp2 association and the consequent degradation of p27(Kip1) by proteosome, leading to G2/M phase arrest of MCF-7 cell. It is suggested that GA should be beneficial to treatment of breast cancer and p27(Kip1)-deficient carcinomas.

Polyphenols extracted from Hibiscus sabdariffa L. inhibited lipopolysaccharide-induced inflammation by improving antioxidative conditions and regulating cyclooxygenase-2 expression.

Oxidative stress and inflammation are related to several chronic diseases including cancer and atherosclerosis. Hibiscus sabdariffa Linnaeus has been found to possess antioxidant effects. In this study, polyphenols extracted from Hibiscus sabdariffa L. (HPE) were used to detect anti-inflammatory effects on nitrite and prostaglandin E2 (PGE2) in lipopolysaccharide (LPS) treated RAW264.7 cells. Sequentially, an animal model examination was performed to confirm the effects of HPE on LPS-induced hepatic inflammation. The results showed that HPE reduced 94.6% of xanthine oxidase activity in vitro, and decreased nitrite and PGE2 secretions in LPS-induced cells. In LPS-treated rats, HPE significantly decreased the serum levels of alanine and aspartate aminotransferase. In the liver, lipid peroxidation and liver lesions decreased, and catalase activity and glutathione increased. The study also revealed that down-regulation of cyclooxygenase-2 (COX-2), p-c-Jun N-terminal kinase (p-JNK) and p-P38 might have been involved. In sum, this study found an anti-inflammatory potency of HPE both in vitro and in vivo.

Increased expression of iNOS and c-fos via regulation of protein tyrosine phosphorylation and MEK1/ERK2 proteins in terminal bronchiole lesions in the lungs of rats exposed to cigarette smoke

Abstract. Epidemiological evidence suggests that smoking is a major cause of human lung cancer. However, the mechanism by which cigarette smoke induces the cancer remains unestablished. To evaluate the effects of cigarette smoke on the expression of inducible nitric oxide synthase (iNOS), nuclear protooncogenes and related mitogen-activated protein kinases (MAPKs) in rat lung tissue, a histopathological study of the effects of gas-phase cigarette smoke on rat lung tissue were carried out. The terminal bronchioles were found to be infiltrated predominantly by lymphocytes in the peribronchiolar region and a mild to moderate degree of emphysema was noted in the alveolar spaces. The terminal bronchioles also showed marked lipid peroxidation, dilatation, and peribronchiolar fibrosis. Immunohistochemical evaluation showed that the expression of iNOS, NF-κB, MAPKs (MEK1, ERK2), phosphotyrosine protein and c-fos was increased in the terminal bronchioles but protein kinase C (PKC), MEKK-1, c-jun, p38 and c-myc showed no change. These results provide evidence to suggest that exposure to cigarette smoke results in oxidant stress which leads to the stimulation of iNOS and c-fos together with the induction of protein tyrosine phosphorylation and MEK1/ERK2 which in turn may promote lung pathogenesis.

Gaseous Nitrogen Oxide Promotes Human Lung Cancer Cell Line A549 Migration, Invasion, and Metastasis via iNOS-Mediated MMP-2 Production

Gaseous nitrogen oxide (gNO) is an important indoor and outdoor air pollutant. Many studies have indicated gNO causes lung tissue damage by its oxidation properties and free radicals. However, there are considerably few data on the association between lung cancer and gNO exposure. The purpose of this study was to examine whether gNO could contribute to the process of malignant progression of lung cancer. The results of wound-healing assay and in vitro transwell assay revealed that gNO-induced dose and time dependently the migration and invasion of A549 cells, a human lung cancer cell line, under noncytotoxic concentrations. gNO was able to induce release of NO from A549 cells, an effect that was mediated via the activation of inducible nitric oxide synthases (iNOS), but not constitutive isoforms, during the same treatment period. An increased expression of matrix metalloproteinase (MMP) and a coincided reduction in repress tissue inhibitors of metalloprotease-2 were observed upon the treatment of gNO. The gNO-mediated MMP-2 induction appeared to be a consequence of nuclear factor kappa B and activation protein-1 activation, because that their DNA binding activity was enhanced by gNO. All these influences of gNO were efficiently repressed by the pretreatment of a NOS inhibitor (N(G)-nitro-L-arginine methyl ester). Using a mouse model, we showed that gNO promoted A549 metastasis to the lung through a mechanism involving the iNOS-dependent MMP-2 activity. Our data imply that gNO exposure, which in turn led to iNOS activation and the enhancement of MMP-mediated cellular events, was related to lung cancer development.

Paeonia lactiflora Pall inhibits bladder cancer growth involving phosphorylation of Chk2 in vitro and in vivo

ETHNOPHARMACOLOGICAL RELEVANCE Extracts of Paeonia lactiflora Pall (RPA), a traditional Chinese medicines has been shown to treat cancers. AIM OF THE STUDY The purpose of this study is to evaluate the anticancer effect of RPA in urinary bladder carcinoma in vitro and in vivo. MATERIALS AND METHODS The cell viability was analyzed with DAPI. Flow cytometry and Western blot were used to study the apoptosis and cell cycle related mechanism. A rat model of bladder cancer was induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (OH-BBN). Tumors were analyzed with immunohistochemical analysis. RESULTS Our data suggested that RPA inhibits growth of bladder cancer via induction of apoptosis and cell cycle arrest. Treatment of TSGH-8301 cells with RPA resulted in G2-M phase arrest that was associated with a marked decline in protein levels of cdc2, cyclin B1, cell division cycle 25B (Cdc25B) and Cdc25C. We also reported that RPA-mediated growth inhibition of TSGH-8301 cells was correlated with activation of checkpoint kinase 2 (Chk2). Herein, we further evaluated urinary bladder cancer using a model of bladder cancer induced by OH-BBN. Analysis of tumors from RPA-treated rats showed significant decrease in the expression of Bcl2, cyclin D1, and PCNA, and increase in the expression of p-Chk2 (Thr-68), Bax, and Cip1/p21. CONCLUSION Our data provide the experimental evidence that RPA could modulate apoptosis in models of bladder cancer.

Primary tuberculous infection of breast: experiences of surgical resection for aged patients and review of literature

Primary mammary tuberculosis is a rare entity that usually occurs in female of reproductive age. Herein three such patients including two males with ages over 80 years, who underwent surgical resection, are reported. Fine needle biopsy failed to achieve specific diagnosis before surgical operation. All of their conditions got satisfactory improvement and anti-tuberculosis chemotherapy was administered postoperatively. Previous literature related to the epidemiology, diagnosis and treatment for mammary tuberculosis will be also reviewed. Mammary tuber culosis is usually related to breast feeding women and is extremely rare in aged man. The possible mechanisms resulting in this disease in our three patients, including direct extension, reactivation, or transmitted by staffs or peers of the nursing home, would also be discussed.

Promotion of colon carcinogenesis through increasing lipid peroxidation induced in rats by a high cholesterol diet

To examine the influence of hypercholesteremia on 1,2-dimethylhydrazine (DMH)-induced rat colon cancer, Sprague-Dawley rats received dietary cholesterol (CH, 0-2%) and cholic acid (CA, 0.25%) with or without DMH (20 mg/kg, s.c. injection) for 18 weeks. The rats receiving dietary cholesterol and cholic acid all significantly increased total serum cholesterol and lipids but only a high cholesterol diet (2% CH plus 0.25% CA) decreased the activity of glutathione peroxidase (GSH-Px) and increased the formation of peroxides in the colon (P < 0.01). The rats that received the combination of DMH and high cholesterol diet enhanced these effects. At the end of the experiment, the diet group administered DMH and high cholesterol (2% CH plus 0.25% CA) developed colon adenoma at 50% of incidence in pathological examination, but no colon adenoma formed in the rats treated with high cholesterol alone. It is supposed that a non-carcinogenic agent like cholesterol may potentiate the carcinogenicity of DMH in rats via an increase of lipid peroxidation and decrease in the activity of peroxidase in the target organ.