Controlled Release Matrix Formulation of Paliperidone in Concurrence with Regulatory Requirements

Paliperidone is the 9-hydroxy metabolite (9-hydroxy) of risperidone and is a psychotropic drug of the atypical antipsychotic family. Paliperidone has the racemates (+)- and (-)-paliperidone. It is a dopamine D2 antagonist with serotonergic 5-HT2A antagonistic action that acts centrally. ALZA OROS® osmotic medication release technology is used to create Invega ER tablets. It is a tri-layer longitudinally compressed tablet based on a sophisticated osmotic delivery method that is meant to administer the paliperidone in a defined way over 24 hours. This research aims to create a generic controlled-release single-layer matrix tablet of paliperidone. Different combinations of Polyox and hypromellose in the core were used, followed by coating, to assist/build a stable and strong formulation. All strengths have similar in-vitro dissolution profiles. Freeze formulation was assessed for nitrosamine risk assessment as well as challenge for alcohol dose dumping study. Paliperidone is a basic compound with a pKa1 of 8.2 (piperidine moiety) and a pKa2 of 2.6 (pyrimidine moiety). As a result, a substantial portion of the molecule is ionized at physiological pH. It is relatively insoluble in water (0.003 g/100 mL water at pH 7.4). The solubility decreases at higher pH (0.001 g/100 mL at pH 12.9) and significantly increases at lower pH (3 g/100 mL at pH 5.3). The partition coefficient octanol/water (log P) is 2.39. Hence, discriminating media was identified as pH 2.75 buffer. The Higuchi model was used for expressing the in-vitro release profile through matrix composition. Formulation withstands 0–40% alcoholic conditions under in-vitro release tests. It is easy to formulate, stable and cost-effective. The manufacturing process involves dry blending followed by compression and coating so there will be the least chemical interaction of an active substance with other excipients. Hence, there is a negligible possibility to generate nitrosamine impurity in the formulation. The formulation is classified as rugged against dose dumping.

Optimization of pretreatment methods for N-nitrosamine compounds in ready-to-eat aquatic products / 预防医学

Objective@#To optimize the pretreatment method of N-nitrosamine compounds in ready-to-eat aquatic products. @*Methods@#Market-sold ready-to-eat aquatic products were collected, homogenized and distilled by steam. The samples were extracted for 10 minutes using dispersive liquid-liquid microextraction (DLLME) with ethanol, trichloromethane and sodium chloride (3.0 g). After centrifugation, the organic phase in the lower layer was collected and subjected to gas chromatography-tandem mass spectrometry (GC-MS/MS). The six common N-nitrosamine compounds were determined in ready-to-eat aquatic products using multiple reaction monitoring mode (MRM) and quantified by the internal standard method. @*Results@#The optimized method exhibited a good linear relationship at concentrations of 10.0 to 500 μg/L for determination of 6 N-nitrosamine compounds (correlation coefficient of greater than 0.999), with 0.05 to 0.60 μg/kg limit of detection, 0.15 to 1.60 μg/kg limit of quantitation, mean spiked recovery rates of 71.8% to 108.9%, and relative standard deviations of 1.4% to 8.6%. N-Nitrosodimethylamine showed the highest detection rate in 20 market-sold ready-to-eat aquatic products (90%), and the detection rates of N-Nitrosopyrrolidine, N-Nitrosodiethylamine and N-dibutylnitrosamine were 15%, 10% and 10%, respectively. @*Conclusion@#Steam distillation combined with DLLME may optimize the pretreatment method of N-nitrosamine compounds in ready-to-eat aquatic products and meet the measurement requirements.

Inhibitory Effect of Water Extract of Broussonetiae Fructus on Hepatocellular Carcinoma Induced by Diethylnitrosamine in Mice and Effect of PTEN/PI3K/Akt Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the inhibitory effect of water extract of Broussonetiae Fructus on hepatocellular carcinoma （HCC） induced by diethyl nitrosamine （DEN） in mice based on homologous phosphatase and tensin homolog/phosphatidylinositol 3-kinase/protein kinase B （PTEN/PI3K/Akt） signaling pathway. MethodThe primary HCC mouse model was constructed by intraperitoneal injection of DEN solution， and the HCC mice were randomly divided into model group， sorafenib group （0.01 g·kg-1·d-1）， low-dose Broussonetiae Fructus water extract group （0.9 g·kg-1·d-1）， medium-dose Broussonetiae Fructus water extract group （1.8 g·kg-1·d-1）， and high-dose Broussonetiae Fructus water extract group （3.6 g·kg-1·d-1）， with 10 mice in each group. Another 10 C57BL/6 mice were selected as a control group and intraperitoneally injected with an equal volume of normal saline. Mice were treated with different concentrations of Broussonetiae Fructus water extract when liver cancer-like white nodules appeared. sorafenib group was treated with sorafenib. The control group and model group were intraperitoneally injected with normal saline. The activities of alkaline phosphatase （ALP）， aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， and γ-glutamyl transferase （γ-GT） in the serum of mice were detected by the biochemical analyzer. The expression levels of alpha-fetoprotein （AFP） and carcinoembryonic antigen （CEA） were detected by enzyme-linked immunosorbent assay （ELISA）. The degree of hepatocyte canceration and hepatocyte injury were observed by Hematoxylin-eosin （HE） and Masson staining. The proliferation of HCC cells was observed by immunohistochemical staining. The apoptosis of HCC cells in mice was observed by erminal-deoxynucleotidyl transferase mediated nick end labelling （TUNEL） staining. The expression levels of PTEN， PI3K， Akt， and p-Akt proteins related to the PTEN/PI3K/Akt signaling pathway were detected by Western blot. ResultCompared with the control group， the activities of ALP， AST， ALT， and γ-GT， as well as the expression levels of AFP and CEA in the model group were significantly increased （P<0.01）. Carcinogenesis and inflammatory cell infiltration were obvious in liver tissue of mice， and a large number of blue collagen fiber hyperplasia was found. The number of Ki67 positive cells was significantly increased （P<0.01）， and the expression level of PTEN protein was significantly decreased， while PI3K and p-Akt protein expression was increased （P<0.01）. Compared with the model group， the activities of ALP， AST， ALT， and γ-GT， as well as the expression levels of AFP and CEA in the medium-dose and high-dose Broussonetiae Fructus water extract groups were significantly decreased （P<0.05， P<0.01）. The degree of carcinogenesis and inflammatory cell infiltration in liver tissue were reduced， and the collagen fiber hyperplasia was significantly reduced. The number of Ki67 positive cells was significantly decreased， and the number of TUNEL positive apoptotic cells was significantly increased （P<0.05， P<0.01）. PTEN protein expression was increased， while p-Akt protein expression was significantly decreased （P<0.05， P<0.01）. ConclusionThe water extract of Broussonetiae Fructus has a significant inhibitory effect on DEN-induced primary HCC in mice， and its mechanism may be related to the regulation of key protein expressions in the PTEN/PI3K/Akt signaling pathway.

Metformin and nitrosamine impurities

Metformin is considered as gold standard anti-diabetic drug and is the preferred initial pharmacologic agent for most of the patients with type 2 diabetes mellitus. Metformin is cheap, widely available and safe, backed by pharmaco-epidemiological evidence of more than 60 years regular use in clinical practice. Due to its durable efficacy, once initiated, metformin will be continued as long as it is tolerated and not contraindicated.  It has got additional benefits on cholesterol, liver, cardio vascular system and cancer. Recent evidence and recall of metformin extended release formulation due to detection of excess amount of cancer-causing nitrosamine impurities has created concern among health care providers and patients. Adherence to regulatory guidelines and use of approved technologies in manufacturing and quality control may help in solving the issue.

Regulation of toxic contents of smokeless tobacco products

Effective regulation of contents of tobacco products is one of the primary milestones to reduce negative health effects associated with the use of smokeless tobacco (SLT) products. As per the available sources, testing of some SLT products has been done on ad hoc basis, but there is a lack of comprehensive and periodic analysis of these products. In addition, the available results indicate huge variations among the levels of pH, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, N-nitrosonornicotine, benzo[a]pyrene, heavy metals and nicotine within different products as well as within different brands of the same product. This review was aimed to throw light on the variations and gaps in testing of SLT products and emphasize the need for strong policy regulation for monitoring the chemical constituents of these products.

Effects of Safflower Extract on Liver Tissue Cell Mitochondria of Hepatic Fibrosis in Rats / 医学研究杂志

Objective To investigate the protective effect of safflower extract on hepatocyte mitochondria in hepatic fibrosis rats.Methods Rat model of hepatic fibrosis was induced by diethyl nitrosamine.Oral safflower extract was taken orally in the experimental group.Hepatic mitochondria were isolated from each group.Morphological changes of hepatocyte mitochondria were observed and MDA and SOD were detected in the level hepatocyte mitochondria.Observe changes in mitochondrial membrane potential and mitochondrial ATP levels and observe changes in mitochondrial respiratory function to investigate whether safflower extract has protective effects on hepatocyte mitochondria in rats with hepatic fibrosis or not.Results In the normal group,the rat mitochondria were arranged neatly and the morphology was normal.In the control group,the mitochondria of the rats were swollen with irregular morphology,the borders were unclear,and the sizes were inconsistent.In the experimental group,the mitochondria were swollen,the structures were not clear,and the sizes were different.The above condition is significantly improved;liver fibrosis in rats' mitochondria MDA levels increased significantly,while SOD content decreased significantly,safflower extract treatment can significantly reduce mitochondrial MDA levels,and it can increase SOD content.Compared with the normal group,the mitochondrial membrane potential of liver fibrosis rats was significantly reduced,while the safflower extract treatment can increase the mitochondrial membrane potential;due to liver fibrosis,rats can increase the consumption of ATP in liver mitochondria and safflower extract can significantly reduce ATP consumption in rat mitochondria.Consistent with this,the phosphor-to-oxygen ratio in the rat mitochondria of the experimental group was significantly higher than that of the control group.Conclusion The safflower extract can significantly reduce the hepatocyte mitochondrial oxidative stress induced by hepatic fibrosis in rats.It can maintain mitochondrial ATP level,respiratory control rate and phosphorus-oxygen ratio to protect mitochondrial function.

Development of Urinary Bladder Pre-Neoplasia by Schistosoma haematobium Eggs and Chemical Carcinogen in Mice

Schistosoma haematobium is a biocarcinogen of human urinary bladder (UB). The present study investigated developing UB cancer mouse model by injecting S. haematobium eggs into the bladder wall and introduction of chemical carcinogens. Histopathological findings showed mild hyperplasia to epithelial vacuolar change, and high grade dysplasia. Squamous metaplasia was observed in the S. haematobium eggs+NDMA group at week 12 but not in other groups. Immunohistochemistry revealed significantly high expression of Ki-67 in urothelial epithelial cells of the S. haematobium eggs+BBN group at week 20. The qRT-PCR showed high expression of p53 gene in S. haematobium eggs group at week 4 and S. haematobium eggs+BBN group at week 20. E-cadherin and vimentin showed contrasting expression in S. haematobium eggs+BBN group. Such inverse expression of E-cadherin and vimentin may indicate epithelial mesenchymal transition in the UB tissue. In conclusion, S. haematobium eggs and nitrosamines may transform UB cells into squamous metaplasia and dysplasia in correlation with increased expression of Ki-67. Marked decrease in E-cadherin and increase in p53 and vimentin expressions may support the transformation. The present study introduces a promising modified animal model for UB cancer study using S. haematobium eggs.

Bee Honey Modulates the Oxidant-Antioxidant Imbalance in Diethyl nitrosamine-Initiated Rat Hepatocellular Carcinoma.

Oxidative stress reflects the mechanism that contributes to initiation and progression of hepatic injury in a variety of liver disturbance. From here, there is a great demand for the expansion of agents with a potent antioxidant effect. The aim of this work is to approximate the efficiency of bee honey as a hepatoprotective and an antioxidant agent versus diethyl nitrosamine (DEN) motivate hepatocellular damage. The single intrapritoneal (IP) management of diethyl nitrosamine (50mg/kg followed by 2ml/kg CCl4) to rats, referred for the histopathological examination of liver sections of rats after induction and before treatment with honey showed that many well differentiated tumor cells were formed in the liver of rats also, the examined sections showed disorganization of hepatic lobular architecture and obvious cellular damage. A significant lift in the enzymatic activity of liver functions (AST, ALT, ALP), and gamma glutamyltransferase (GGT) which is a signal of hepatocellular damage. DEN stimulates oxidative stress, which was assured by increase lipid peroxidation level and hindrance in antioxidant enzymes (SOD, CAT, GPx, and GST) activities in the liver. The position of non-enzymatic antioxidants comparable reduced glutathione (GSH) was likewise set up to be slimmed down significantly in DEN inoculated rats. Also, we have studied the underlying mechanism and /or (s) of the therapeutic role of bee honey as hepatocarcinogenesis remediation through investigation the inflammatory biomarkers; α-fetoprotein (AFP) and α-fucosidase (AFU). The current results clearly showed that bee honey demonstrates good ameliorative and antioxidant capacity toward diethyl nitrosamine induced hepatocellular damage in rats.

Enzymatic and Non Enzymatic Antioxidant Activity of Tabernaemontana divaricate R.Br. against DEN and Fe-NTA Induced Renal Damage in Wistar Albino Rats.

Plant derived drugs have been a part of the evolution of human healthcare for thousands of years. The present study was carried out to evaluate the antioxidant potential of ethanolic extract of Tabernaemontana divaricata on DEN initiated and Fe-NTA promoted renal damage in rats. Fe-NTA was induced after 10 days of DEN (200mg/kg body weight) initiation at a dose level of 9mg Fe-NTA/kg body weight twice a week for one month. The biochemical parameters were analyzed in serum and the antioxidant assays were carried out in kidney. Lipid peroxidation level was increased due to the administration of Fe-NTA, which caused the reduction of enzymatic antioxidant such as SOD, GPx, Catalase, G6PD, and also the non-enzymatic antioxidants vitamin C and GSH. The levels of urea, uric acid, creatinine, blood urea nitrogen were increased and protein level decreased on Fe- NTA intoxication. The secondary metabolites present in the plant increased the synthesis of antioxidant enzymes and its free radical scavenging properties helped to scavenge all free radicals thereby decreasing lipid peroxidation. Thus, the present study indicates that the plant may clinically valuable agent in the prevention of renal failure caused by DEN and Fe-NTA intoxication.

Selection and characterization of probiotic lactic acid bacteria with heterocyclic amine binding and nitrosamine degradation properties.

Potential probiotic strain for being health protectant especially intestinal illness is strain specific. This study investigated the selection of a new strain of probiotic of non-human origin and of human origin with the properties of intestinal protection against cancer. From the primary screening results, the human feces origin strains showed more bile salt tolerance than the fermented food origin strains. Whereas none of the human feces origin isolates could grow well in the acid condition. Lactobacillus plantarum CM4 was the new probiotic of non-human origin strain for this study. CM4 cells is said to tolerate and grow in 0.3% bile salt after 5 hours of incubation, at pH3 after 6 hours of incubation. This is in agreement with in vivo study for intestinal adherence ability of probiotic, a live CM4 cells was able to persist in mice small intestine and colon for 5 days. Live CM4 cells showed most effectiveness to bind 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) mutagen after 24 hours of incubation with 46.32% of binding ability while 144 hours of incubation with 85.34% of binding ability was the most effective for 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) mutagen. The significant difference (p<0.05) was found at all those time points. Moreover, the CM4 strain could degrade diphenylnitrosamine (DPN) better than 1-nitrosopyrrolidine (NPR) with dose response relationship activity. These imply that the CM4 strain could be the value added for the consuming pharmaceutical probiotic product based on scientific proof of its role in intestinal survival properties and cancer prevention through binding PhIP and IQ mutagen as well as degrading nitrosamine.

Study on effect of transforming growth factor-β1 vaccine on insulin-like growth factor binding protein in liver tissues of rats with liver fibrosis / 中华传染病杂志

Objective To observe the effect of transforming growth factor-β1 (TGF-β1) vaccine on the degree of hepatic fibrosis in rats ,and to explore the effect of TGF-β1 vaccine on the insulin-like growth factor binding protein (IGFBP)3 and IGFBP7 .Methods The hepatic fibrosis rat model was set up by injecting N-nitrosodimethylamine . Among them , 10 rats were injected with TGF-β1 vaccine , and additional 10 rats were set up as healthy control group .Changes in hepatic pathology were observe and the expressions of IGFBP3 and IGFBP7 were detected by the methods of immunohistochemistry , reverse transcription polymerase chain reaction (RT-PCR) and Western blot in rat fibrosis tissues after 6 weeks . Normality test and analysis of variance were conducted .LSD test was conducted if variances were tested homogeneity .Categorical data were analyzed using Fisher exact test . Results Changes in hepatic histology and serum levels of hyaluronic acid and laminin suggested that TGF-β1 vaccine interventions could reduce the extent of hepatic fibrosis in rats .The expressions of IGFBP3 mRNA in control group ,hepatic fibrosis model group and vaccine intervention group were 1 .735 ± 0 .097 ,1 .165 ± 0 .096 and 1 .491 ± 0 .046 ,respectively (t= 4 .575 ,6 .285 and 8 .489 ,respectively ,all P< 0 .05) .The expressions of IGFBP7 in the above three groups were 0 .497 ± 0 .021 ,1 .250 ± 0 .064 and 0 .885 ± 0 .149 ,respectively (t= 5 .161 ,30 .101 and 7 .250 , respectively ,all P < 0 .05 ) . Immunohistochemistry proved that the expressions of IGFBP7 in fibrosis model group and TGF-β1 vaccine group were all significantly higher than control group ;and the expressions of IGFBP3 in fibrosis model group and TGF-β1 vaccine group were all significantly lower than control group .The expressions of IGFBP3 protein in control group , hepatic fibrosis model group and vaccine intervention group were 7 .508 ± 0 .357 ,5 .200 ± 0 .210 and 5 .751 ± 0 .178 ,respectively (t = 7 .622 ,6 .180 and 29 .156 , respectively ,all P < 0 .05) . The expressions of IGFBP7 were 1 .176 ± 0 .051 ,1 .735 ± 0 .115 and 1 .428 ± 0 .056 ,respectively (t = 7 .188 ,4 .827 and 8 .649 ,respectively ,all P< 0 .05) .Conclusion TGF-β1 vaccine can affect the expressions of IGFBP3 and IGFBP7 ,which plays an important role in the formation and development of hepatic fibrosis .

Study on inhibition of anthraquinone in Polygonum Cillinerve (Nakai) Ohwi to Nitrosation / 国际中医中药杂志

Objective To study inhibitory activity of anthraquinone in Polygonum Cillinerve(Nakai) Ohwi (anthraquinone in PCO) to nitrosation. Methods Anthraquinone was extracted from Polygonum Cillinerve(Nakai) Ohwi with sulfuric acid and chloroform as solvents. The capability of scavenging sodium nitrite and disconnecting nitrosamine synthesis with anthraquinone in PCO were determined by spectrophotometry under simulated human gastric juice. Results Both the sodium nitrite scavenging rate and the nitrosamine synthesis disconnecting rate showed the positive correlation with the concentration of anthraquinone in PCO. The strongest capability of scavenging sodium nitrite was 53.5%, and the strongest capability of disconnecting nitrosamine synthesis was 71.3%. Conclusion Anthraquinone in PCO had strong capability of scavenging sodium nitrite and disconnecting nitrosamine synthesis.

Effect of the Magnetized Water Supplementation on Lymphocyte DNA Damage in Mice Treated with Diethylnitrosamine

Water gets magnetically charged when it is contacted with a magnet. Although magnetic water products have been promoted since the 1930's, they have received very little recognition due to questionable effectiveness. Diethylnitrosamine (DEN) is a widely occurring nitrosamine that is one of the most important environmental carcinogens primarily inducing tumors of liver. In this study, the effect of magnetized water supplementation on lymphocyte DNA damage in ICR mice treated with DEN was evaluated using the Comet assay. Mice were divided into 3 groups: control, DEN, and DEN + magnetized water group. Fifteen mice were maintained in each group for the entire experimental period of 6, 12 and 18 weeks. Five mice in each group were sacrificed at 6, 12, and 18th weeks, followed by the Comet assay using the blood obtained from heart puncture of the mice. The level of lymphocyte DNA damage reflected by tail moment and other DNA damage indices of tail DNA (%) or tail length of the magnetized water group were significantly decreased after the 6th, 12th and 18th weeks of supplementation compared with the positive control, the DEN group. The relative DNA damage of the magnetized water groups compared to the DEN control group after 6th, 12th, and 18th weeks of supplementation were 42.2%, 40.8%, and 32.9% for DNA in tail, 31.2%, 32.6%, and 21.3% for tail length, and 33.8%, 33.8%, and 24.6% for tail moment, respectively. This is the first report demonstrating that magnetized water may be involved in the lowering effect of the DNA damage in DEN-treated ICR mice. This result suggests that the magnetized water might have minimized the DNA damage by improving the antioxidant status of the mice. However, further studies are needed to characterize the condition of the magnetization and examine the long-term effect of the water product.

Effects of Extremely Low Frequency Elecromagnetic Fields on Thyroid Carcinogenesis Induced by N-bis(2-hydroxypropyl)nitrosamine and Sulfadimethoxine

PURPOSE: Long-term exposure to extremely low-frequency (60 Hz) electromagnetic fields (ELF-EMF) raises the questions of the induction of biological effects including tumorigenesis. One mechanism through which ELF-MFS could influence neoplastic development is the imbalance of cellular proliferation and cell apoptosis. The present study investigated the effect of ELF-EMF on chemically-induced thyroid carcinogenesis in a rat. METHODS: We examined cellular proliferation index measured by anti-Ki-67 antigen, apoptosis, apoptosis related proteins such as caspase 3 and p53, and cell cycle-related proteins (cyclin D1 and p21(WAF1/Cip1)). Forty Male F344 rats received a subcutaneous N-bis(2-hydroxypropyl)nitrosamine (DHPN, 2,800 mg/kg) injection, and 1 week later were allowed free access to drinking water containing sulfadimethoxine (0.1%) for 12 weeks. Twenty rats were exposed by ELF-EMF. During the carcinogenesis, sequential histological changes from hyperplasia, adenoma, and ultimately to overt carcinomas were noted. RESULTS: The exposure group of ELF-EMF, significantly increases the number size of carcinomas. Also, the proliferative and apoptotic indices were significantly increased in the ELF-EMF exposure group than in the control group. The caspase 3 protein expression did not show any significant changes between ELF-EMF group and control group. The p53 protein was not detected in both ELF-EMF exposure and control group. Among the cell cycle related proteins, cyclin D1, not p21(WAF1/Cip1), was significantly increased in adenomas and carcinomas in ELF-EMF exposure group compared with the control group. CONCLUSION: Exposure of ELF-EMF effects on chemically-induced rat thyroid carcinogenesis as results of altered increase of cellular proliferation, apoptosis, and cyclin D1 expression.

Anticancer Effect of Magnolia officinalis' Extract on N-butyl-N-(4-hydroxybutyl) Nitrosamine Induced Bladder Cancer in Mice / 대한비뇨기과학회지

PURPOSE: Magnolia officinalis has been used in traditional Chinese medicine to treat a variety of diseases. The main constituents of Magnolia officinalis are honokiol and magnolol, which have a variety of pharmacological effects, such as antitumor, antioxidant, antimicrobial, anti-inflammatory etc. This study examined the anticancer effect of a Magnolia officinalis' extract on urinary bladder cancer in vivo. MATERIALS AND METHODS: Male mice C3H/He were used as the experimental animals. The mice were divided into ten groups. Normal drinking water was provided to group 1(5 mice) for 20 weeks and 0.05% N-butyl- N-(4-hydroxybutyl) nitrosamine(BBN) was added to in the drinking water of group 2(5 mice). 0.1, 0.3, 1.0 and 3.0% Magnolia officinalis' extract was added to groups 3, 4, 5 and 6, respectively(5 mice each), and 0.1, 0.3, 1.0 and 3.0% Magnolia officinalis' extract plus 0.05% BBN was added to groups 7, 8, 9 and 10, respectively(10 mice each) for the same period. All surviving mice were sacrificed at week 20 to investigate the occurrence of bladder cancer, stage and grade. RESULTS: Bladder cancer was not observed in groups 1, 3, 4, 5 and 6 mice. The rates of bladder cancer occurrence were 57.1, 66.7, 44.4 and 20.0% in groups 7, 8, 9 and 10, respectively. The incidence decreased with increasing concentration of Magnolia officinalis (p=0.005). However, the stage and grade were not associated with the concentration of Magnolia officinalis(each p>0.05). CONCLUSIONS: This study showed that Magnolia officinalis has some protective effect against bladder cancer. In the future, Magnolia officinalis may be expected to play an important role as a chemo-preventive and therapeutic agent or as a complementary agent in bladder cancer.

Expression of Transforming Growth Factor-beta1 and Its Receptors in N-butyl-N-(4-hydroxybutyl) Nitrosamine-induced Rat Bladder Carcinogenesis / 대한비뇨기과학회지

PURPOSE: To determine the role of TGF-beta1, and its receptors, in bladder tumor, their expressions at various stages of chemically-induced rat bladder carcinogenesis were investigated. MATERIALS AND METHODS: Forty female Sprague-Dawley rats (200-250g) were given drinking water containing 0.05% N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN), and twenty rats, used as a control group, were given tap water. After 10, 20, 25 and 30 weeks of the administration, the bladders of the rats were harvested. The control and BBN-treated rat bladders were analyzed for the expressions of TGF-beta1, and its receptors (RI, RII and RIII), in the mRNA by a real-time quantitative polymerase chain reaction. The protein expression was determined by immunohistochemistry. RESULTS: The expressions of the TGF-beta1 increased in the mRNA with the BBN treatment, while those of the TGF-beta receptors decreased. The up-regulation of TGF-beta1 was statistically significant after 25 weeks of BBN treatment, but down-regulations of RI, RII, and RIII were significant after 20, 25, and 30 weeks of BBN treatment, respectively (p<0.05). The immunohistochemical analysis demonstrated that the TGF-beta1, and its receptors, were localized in the tumor cytoplasm, and their intensities reflected the expression in the mRNA of these tissues. CONCLUSIONS: These data suggest that the enhanced expression of TGF-beta1, as well as the loss of the expressions of RI, RII, and RIII, at the various stages, contributes to the carcinogenesis of the bladder and tumor progression.

Expression of Cyclins (D1, A, E, and B1) in N-butyl-N-(4-hydroxybutyl)nitrosamine-induced Rat Bladder Carcinogenesis

BACKGROUND: Cell cycle deregulation plays a major role in chemical multistage carcinogenesis.Therefore, the evaluation of cell cycle proteins is important. METHODS: In order to induce carcinogenesis in the rat urinary bladder, 0.05% N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN)was administered to male Sprague-Dawley rats for 30 weeks. Expressions of cyclin D1, A, E, and B1 were examined by immunohistochemical stainings. RESULTS: Urothelial cell hyperplasia appeared at 5 weeks, followed by papilloma at 10 weeks. Superficial carcinoma was observed at 20 weeks, and invasive carcinoma developed in 40% (4/10) of the rats at 30 weeks. Expressions of cyclin D1 and A increased sequentially from normal mucosa throughhyperplasia, papilloma, and carcinoma (p<0.01). Expressions of cyclin D1, B1 and cyclin Ewere higher in invasive carcinomas than in superficial carcinomas (p<0.01). In contrast, therewas no significant difference in the expression of cyclin B1 between hyperplasia, papillomaand superficial carcinoma. CONCLUSIONS: The present results indicate the important roles of cyclin D1 and A in the development of BBN-induced urothelial carcinoma of rats. Aberrantexpression of cyclin B1 and E may contribute to the progression from superficial to invasivebladder cancer rather than tumorigenesis.

Effects of carcinogenicity and growth ragulatory factors in human epithelial cells exposed with tobacco-specific N-nitrosamine

Since NNK is one of the most abundant tobacco-specific alkaloids and a strong carcinogenic nitrosamine, it has been used for evaluating a potential of carcinogenicity in the animal models. The present study has attempted to examine the potential of carcinogenicity of NNK in human epithelial cells, from which the cell type the most of cancers including oral cancer and nasal cavity cancer are originated. The cellular model used for the study is a human keratinocyte cell system immortalized by Ad12-SV40 hybrid virus. The cellular system has successfully been used for the carcinogenicity studies because of its limitless life span, epithelial morphology and non-tumorigenicity. When cells were treated with a variety of NNK concentrations, levels of saturation density and soft agar colony formation were increased in a dose-dependent fashion. Colonies of large cell aggregates were above 5 at the higher doses. The results indicate that exposure of human cells with NNK induced loss of contact inhibition and increases of anchorage independence and cellular adhesion, which are typical characteristics of the neoplatically transformed cells. When cells were exposed with 100uM NNK for 2hr, mRNA levels of IL-1 and PAI-2 were increased in a dose-dependent manner, but expression of TGF- 1 was not affected. While expression of growth regulatory factors were altered with a short-term exposure, there was no alteration of these factors in the NNK-transformed cells. However, mRNA levels of fibronectin were increased both in the short-term treatment and in the transformation. The results suggest that altered expression of extracellular matrix such as fibronectin following short-term exposure might be fixed in the genome and these altered properties be continuously transfered throughout the cell division. Western blot analysis showed a translocation of PKC- from cytosolic fraction to the particulate fraction, indicating a possible role of NNK in the signal transduction pathway. The present study provided an evidence that NNK in the smoking may be associated with epithelial origin cancer such as oral and nasal cavity cancers. In addition, this study suggested that altered expression of extracellular matrix and PKC may play an important role in the carcinogenic mechanism of NNK.

The Effects of Intravesical BCG Instillation on Bladder Cancer Induced by N-butyl-N-(4-hydroxybutyl) Nitrosamine / 대한비뇨기과학회지

PURPOSE: We investigate the effects of intravesical BCG therapy on the occurance of superficial bladder cancer induced by N-butyl-N-(4-hydroxybutyl) nitrosamine(BBN) in Fisher 344 rats in vivo. We also examine whether NO mediated the antitumor activity of BCG against superficial bladder cancer in Fisher 344 rats. MATERIALS AND METHODS: BBN(0.1%) is orally administered for 20 weeks and it is combined with BCG(0.27mg) or NG-monomethyl-L-arginine (NG MMA, 10mg). once every week from 8th week to 19th week. The rats are sacrified at 20th week. NO secretion in urine for 24 ours is significantly increased in the BCG treated rats compared to the animals treated with saline or NGMMA. RESULTS: Pathologic findings demonstrate that the incidence of carcinoma is not statistically different in saline, BCG, NGMMA(p>0.05). However the size and number of tumor is decreased in the BCG treated rats compared with saline or NGMMA treated rats bearing bladder cancer induced by BBN(p<0.05). Inducible NO synthase(iNOS) is strongly induced in bladder tissue of rats treated with BCG and NGMMA but not in saline. CONCLUSIONS: Intravesical instillation of BCG on bladder cancer induced by BBN does not decrease the cancer occurrence but reduces the number and size of bladder cancer. Our results suggest that nitric oxide induced by intravesical instillation of BCG may mediate antitumor activity against the occupance of superficial bladder cancer.

Carcinogenicity and Its Inhibition of 4-(N-nitrosomethylamino)-1-(3-pyridyl)-1-butanone, a Tobacco-Specific N-nitrosamine / 环境与健康杂志

The advancement of research work on the carcinogenicity and its inhibition of 4-(N-nitrosomethylamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco-specific N-nitrosamine, was reviewed in the present paper. Carbonyl reduction and ?-hydroxylation were the major routes of NNK metabolic activation, the main target organ of NNK carcinogenicity was the lung and the carcinogenic probability related to the individual, organic isothiocyanates were the most potent inhibitors for NNK carcinogenicity. Eating more cruciferae vegetables and fruit and drinking more green and black tea would be helpful for smokers and passive smokers. Research about the nosogenesis and carcinogenicity and its inhibition of the harmful components in cigarette smoke should be further reinforced.