Augmented efficacy of tamoxifen in rat breast tumorigenesis when gavaged along with riboflavin, niacin, and CoQ10: effects on lipid peroxidation and antioxidants in mitochondria

Reactive oxygen species (ROS) play a major role in causing mitochondrial changes linked to cancer and metastasis. Uptake of antioxidants by tissue to reduce the ROS production could be instrumental in controlling cancer. Tamoxifen (TAM), a nonsteroidal anti-estrogen drug most used in the chemotherapy and chemoprevention of breast cancer. Riboflavin, niacin and coenzyme Q10 (CoQ10) are proved to be potent antioxidants and protective agents against many diseases including cancer. The objective of this research is to determine the therapeutic efficacy of combinatorial therapy on mammary carcinoma bearing rats in terms of the mitochondrial lipid peroxidation and antioxidant status especially MnSOD. Female albino rats of Sprague-Dawley strain were selected for the investigation. Mammary carcinoma was induced with 7,12-dimethyl benz(a)anthracene (DMBA: 25 mg), and the treatment was started by the oral administration of TAM (10 mg/kg body weight/day) along with riboflavin (45 mg/kg body weight/day), niacin (100 mg/kg body weight/day) and CoQ10 (40 mg/kg body weight/day) for 28 days. The levels of lipid peroxides, activities of enzymic and non-enzymic antioxidants were measured in the mitochondria isolated from the mammary gland and liver of control and experimental rats. Rats treated with DMBA showed an increase in mitochondrial lipid peroxidation (mammary gland 52.3%; liver 25.1%) accompanied by high malondialdehyde levels along with lowered activities of mitochondrial enzymic antioxidants [superoxide dismutase (mammary gland 19.9%; liver 24.8%), catalase (mammary gland 50%; liver 19.7%), glutathione peroxidase (mammary gland 47.8%; liver 31.1%)] and non-enzymic antioxidants [reduced glutathione (mammary gland 14.3%; liver 13.3%), Vitamin C (mammary gland 6.49%; liver 21.4%) and E (mammary gland 20.3%; liver 22.2%)]. Administration of combinatorial therapy restored lipid peroxide level and the activities of enzymic and non-enzymic antioxidants to near normalcy. In addition, antitumour activity was also found to be enhanced which is evident from the increased expression of tumour suppressor gene MnSOD thereby preventing cancer cell proliferation. These results suggested that TAM treatment is the most effective during co-administration of riboflavin, niacin and CoQ10 in terms of mitochondrial antioxidant and antitumour activity.     

Alternatively spliced human genes by exon skipping--a database (ASHESdb)

Alternative splicing of mRNA allows many gene products with different functions to be produced from a single coding sequence. Exon skipping is the most commonly known alternative splicing mechanism. A comprehensive database of alternative splicing by exon skipping is made available for the human genome data. 1,229 human genes are identified to exhibit alternative splicing by exon skipping. Availability: http://sege.ntu.edu.sg/wester/ashes/.     

A major ozonation product of cholesterol, 3beta-hydroxy-5-oxo-5,6-secocholestan-6-al, induces apoptosis in H9c2 cardiomyoblasts

Cholesterol, a major neutral lipid component of biological membranes and the lung epithelial lining fluids, is susceptible to oxidation by reactive oxygen and nitrogen species including ozone. The oxidation by ozone in biological environments results in the formation of 3beta-hydroxy-5-oxo-5,6-secocholestan-6-al (cholesterol secoaldehyde or CSeco, major product) along with some other minor products. Recently, CSeco has been implicated in the pathogenesis of atherosclerosis and Alzheimer's disease. In this communication, we report that CSeco induces cytotoxicity in H9c2 cardiomyoblasts with an IC(50) of 8.9+/-1.29 microM (n=6). The observed effect of CSeco at low micromolar concentrations retained several key features of apoptosis, such as changes in nuclear morphology, phosphatidylserine externalization, DNA fragmentation, and caspase 3/7 activity. Treatment of cardiomyocytes with 5 microM CSeco for 24h, for instance, resulted in 30.8+/-3.28% apoptotic and 1.8+/-1.11% of necrotic cells as against DMSO controls that only showed 1.3+/-0.33% of apoptosis and 1.6+/-0.67% of necrosis. In general, the loss of cellular viability paralleled the increased occurrence of apoptotic cells in various CSeco treatments. This study, for the first time, demonstrates the induction of apoptotic cell death in cardiomyocytes by a cholesterol ozonation product, implying a role for ozone in myocardial injury.     

Synthesis of densely functionalised C-glycosides by a tandem oxy Michael addition-Wittig olefination pathway

Wittig olefination of 5,6-dideoxy-5,6-anhydro-6-nitro-d-glucofuranose (5) triggered a concomitant cyclisation via oxy Michael addition of the C2-hydroxyl group resulting in the formation of C-vinyl glycosides with Z-olefinic geometry.     

Molecular characterization of a truncated antigen (Wb14) of SXP-1 of Wuchereria bancrofti from four endemic regions in India

Wb14 of Wuchereria bancrofti, an orthologue of Brugia malayiSXP-1 and W. bancrofti SXP-1, was amplified from genomic DNA of W. bancrofti microfilaria collected from four distant geographical locations in India viz., Vellore, Bhubaneshwar, Pondicherry and Sevagram. The gene was sub-cloned in a prokaryotic vector pRSET and expressed in Escherichia coli as a truncated protein (∼23 kDa). The nucleotide sequence of the gene is 98% similar to that of WbSXP-1 and is found to be intron-less. However, the analysis and comparison of the derived amino acid sequence with WbSXP-1 showed that Wb14 is truncated at amino acid position 153. The distribution of the two genes in the studied four geographical locations indicated that WbSXP-1 is prevalent only in parasite samples from Sevagram while Wb14 is present in parasites from all the other locations. Only a limited polymorphism was observed in both the genes among the parasites from different geographical locations.     

Adaptive responses to low-dose/low-dose-rate gamma rays in normal human fibroblasts: the role of growth architecture and oxidative metabolism

To investigate low-dose/low-dose-rate effects of low-linear energy transfer (LET) ionizing radiation, we used gamma-irradiated cells adapted to grow in a three-dimensional architecture that mimics cell growth in vivo. We determined the cellular, molecular and biochemical changes in these cells. Quiescent normal human fibroblasts were irradiated with single acute or chronic doses (1-10 cGy) of (137)Cs gamma rays. Whereas exposure to an acute dose of 10 cGy increased micronucleus formation, protraction of the dose over 48 h reduced micronucleus frequency to a level similar to or lower than what occurs spontaneously. The protracted treatment also up-regulated the cellular content of the antioxidant glutathione. These changes correlated with modulation of phospho-TP53 (serine 15), a stress marker that was regulated by doses as low as 1 cGy. The DNA damage that occurred after exposure to an acute dose of 10 cGy was protected against in two ways: (1) up-regulation of cellular antioxidant enzyme activity by ectopic overexpression of MnSOD, catalase or glutathione peroxidase, and (2) inhibition of superoxide anion generation by flavin-containing oxidases. These results support a significant role for oxidative metabolism in mediating low-dose radiation effects and demonstrate that cell culture in three dimensions is ideal to investigate radiation-induced adaptive responses. Expression of connexin 43, a constitutive protein of gap junctions, and the G(1) checkpoint were more sensitive to regulation by gamma rays in cells maintained in a three-dimensional than in a two-dimensional configuration.     

The Human Lagging Strand DNA Polymerase δ Holoenzyme Is Distributive

Polymerase δ is widely accepted as the lagging strand replicative DNA polymerase in eukaryotic cells. It forms a replication complex in the presence of replication factor C and proliferating cell nuclear antigen to perform efficient DNA synthesis in vivo. In this study, the human lagging strand holoenzyme was reconstituted in vitro. The rate of DNA synthesis of this holoenzyme, measured with a singly primed ssM13 DNA substrate, is 4.0 ± 0.4 nucleotides. Results from adenosine 5′-(3-thiotriphosphate) tetralithium salt (ATPγS) inhibition experiments revealed the nonprocessive characteristic of the human DNA polymerase (Pol δ) holoenzyme (150 bp for one binding event), consistent with data from chase experiments with catalytically inactive mutant Pol δ^AA. The ATPase activity of replication factor C was characterized and found to be stimulated ∼10-fold in the presence of both proliferating cell nuclear antigen and DNA, but the activity was not shut down by Pol δ in accord with rapid association/dissociation of the holoenzyme to/from DNA. It is noted that high concentrations of ATP inhibit the holoenzyme DNA synthesis activity, most likely due to its inhibition of the clamp loading process.     

Myrtenal, a natural monoterpene, down-regulates TNF-α expression and suppresses carcinogen-induced hepatocellular carcinoma in rats

Hepatocellular carcinoma is one of the most common cancers and lethal diseases in the world. Recently, many researchers focused to identify novel chemotherapeutic agents from natural sources against hepatocarcinogenesis. The diverse therapeutic potential of essential oils has drawn the attention of researchers to test them for anticancer activity, taking advantage of the fact that their mechanism of action is dissimilar to that of chemotherapeutic agents. Earlier reports indicated that essential oil components, especially monoterpenes, have multiple pharmacological effects which could account for the terpene-tumor suppressive activity. In the present study, it is shown that myrtenal, a natural monoterpene, which acts as an antineoplastic agent against diethylnitrosamine induced phenobarbital promoted experimental hepatocellular carcinoma. The results revealed an elevated level of microsomal lipid peroxidation in the liver, which was found to be significantly reduced by myrtenal treatment. On the contrary, the Phase I hepatic drug metabolizing enzymes' (cytochrome P(450), cytochrome b (5), NADPH-cytochrome c reductase, NADH-cytochrome b ( 5 ) reductase) levels were decreased and the Phase II enzymes (glutathione-S-transferase, uridine 5'-diphospho-glucuronyl transferase) were increased in carcinogen-administered animals, which were reverted to near normalcy upon myrtenal administration. Our findings also showed that myrtenal restrains the liver cancer by preventing the DEN-PB induced up-regulation of TNF-α protein expression by immunoblot. Furthermore, transmission electron microscopic examination also indicated that myrtenal prevents the carcinogen-induced changes in the architecture of liver tissue and cell structure. Thus, this study shows that myrtenal has the ability to suppress the hepatocellular carcinoma in rats.     

Synthesis, study on anti-arthritic, anti-inflammatory activity and toxicity of some novel bis-oxy cyclophane diamides

A series of bis-oxy cyclophane diamides with bis(aminomethyl)m-terphenyl as spacer have been synthesized and characterized from spectral and analytical data. All the cyclophane diamides exhibit better anti-arthritic activity than the reference drug viz. diclofenac sodium. Some of the cyclophane diamides exhibit good anti-inflammatory activity. The cyclophane amide 4 and 5 do not show any evidence of mutagenicity and cytotoxicity.     

Molecular hybridization of bioactives: Synthesis and antitubercular evaluation of novel dibenzofuran embodied homoisoflavonoids via Baylis–Hillman reaction

A novel series of natural product like dibenzofuran embodied homoisoflavonoids [(E)-3-(dibenzo[b,d]furan-2-ylmethylene)chroman-4-ones] designed by molecular hybridization were synthesized in very good yields via a sequence of reactions involving base catalyzed Baylis–Hillmann (BH) reaction of 2-dibenzofuran carboxaldehyde and methyl acrylate; bromination of BH adduct; condensation of resulted allylic bromide with substituted phenols or 2-dibenzofuranol followed by cyclization. Among the all 11 new compounds screened for in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv (MTB), (E)-3-(dibenzo[b,d]furan-2-ylmethylene)-6-fluorochroman-4-one (7f) and (E)-3-(dibenzo[b,d] furan-2-ylmethylene)-6-fluorochroman-4-one (7g) were found to be active with MIC 12.5 μg/mL.