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Aim of the study The anticancer activity of red cabbage (RC) and the underlying mechanisms against human cervical and hepatocarcinoma cancer cells were explored in this study. Materials and methods The cytotoxic activity of RC extract was assessed using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) cytotoxicity assay. ELISA was used to detect the level of anticancer cytokines, TNFα and IFNβ, in the culture supernatants of cancer cells. Apoptotic cells were investigated by flow cytometry. The levels of apoptotic genes (Bax, BCL-2, Capsases 7–9), cell cycle regulatory genes (cyclin D, E, and A) and tumor suppressor proteins (p27, p21, and p53) were assessed by real time RT-PCR. Results The cytotoxic effect of the extract on normal human cells was significantly different from its effects on HeLa and HepG2 cells, 251.28 ± 4.3, 23.38 ± 1.87, and 28.66 ± 2.85 mg/ml, respectively. The selectivity index (SI) was 10.88 ± 0.82 for HeLa and 8.93 ± 0.81 for HepG2 cells. Increased levels of TNFα, but not IFNβ were observed in the treated HeLa and HepG2 culture supernatants when compared with untreated cells. RC extract induced G0/G1 phase arrest in the cancer cells. The extract induced apoptosis, in a dose and time dependent manner, in treated HeLa and HepG2 cells while no observed apoptosis was found in untreated cells. Moreover, RC IC50 showed remarkable influence on the expression of the apoptosis-related genes in a positive and negative manner on both HeLa and HepG2 cells. Discussion/conclusion RC extract could be considered as a novel anticancer agent providing new prospects for anticancer therapy using a natural product.

Association Between Telomere Shortening and Ageing During Occupational ExposureTelomere length is considered as a biomarker of ageing, resulting in shortening during the process. The present investigation was an attempt to determine the relative telomere length in mechanical workshop workers. Telomere length shortening in cells during occupational exposure causes accelerated ageing. Genomic DNA was isolated from buccal epithelial cells collected from 240 individuals, comprising two groups of 120 exposed workers and 120 unexposed controls. Telomere length was measured by using real time PCR. Both telomere (T) and single copy gene (S) specific primers were used to compute the relative T/S ratio and expressed as the relative telomere length. Telomere length differed significantly between the workers and controls (pp

The continuous escalation of resistant bacteria against a wide range of antibiotics necessitates discovering novel unconventional sources of antibiotics. B. oleracea L (red cabbage) is health-promoting food with proven anticancer and anti-inflammatory activities. However, it has not been researched adequately for its antimicrobial activity on potential resistant pathogens. The methanol crude extract of B. oleracea L. was investigated for a possible anti-microbial activity. The screening method was conducted using disc diffusion assay against 22 pathogenic bacteria and fungi. It was followed by evaluation of the minimum inhibitory concentration (MIC). Moreover, the antibacterial and the antifungal activities were confirmed using the minimum bactericidal concentration (MBC) and the minimum fungicidal concentration (MFC), respectively. Remarkable, antibacterial activity was evident particularly against highly infectious microorganisms such as Methicillin-resistant Staphylococcus aureus, Escherichia coli O157:H7, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus, and Salmonella enterica serovar Typhimurium as well as against human fungal pathogens, Trichophyton rubrum and Aspergillus terreus. Red cabbage is a rich source of phenolic compounds, anthocyanins being the most abundant class, which might explain its potent antimicrobial action. This extract is potentially novel for future antimicrobials, inexpensive, and readily available at a large scale for pharmaceutical companies for further investigation and processing.

Different biological methods are gaining recognition for the production of silver nanoparticles (Ag-NPs) due to their multiple applications. One of the most important applications of Ag-NPs is their use as an anti-bacterial agent. The use of plants in the synthesis of nanoparticles emerges as a cost effective and eco-friendly approach. In this study the biosynthesis of silver nanoparticles using Vitex negundo L. extract and its antimicrobial properties has been reported. The resulting silver particles are characterized using transmission electron microscopy (TEM), X-ray diffraction (XRD) and UV–Visible (UV-Vis) spectroscopic techniques. The TEM study showed the formation of silver nanoparticles in the 10–30 nm range and average 18.2 nm in size. The XRD study showed that the particles are crystalline in nature, with a face centered cubic (fcc) structure. The silver nanoparticles showed the antimicrobial activity against Gram positive and Gram negative bacteria. Vitex negundo L. was found to display strong potential for the synthesis of silver nanoparticles as antimicrobial agents by rapid reduction of silver ions (Ag+ to Ag0).

Despite all efforts, there is still no effective vaccine available against the human respiratory syncytial virus (HRSV) that is a major cause of severe lower respiratory tract disease in infants and the elderly. In this review, we examined the potential of the conserved immunogenic region (residues 122-226) of the HRSV glycoprotein G alone as the inducer of neutralizing antibodies against this virus. The Escherichia coli produced recombinant conserved region of G (designated as G domain), which was used for rabbit immunization. Although rabbit is semipermissive host for HRSV, our result showed that the polyclonal antibodies against the G domain protein could strongly neutralize the virus (69.3), suggesting that the G immunogenic region of HRSV alone has a great potential in vaccine development. To our knowledge, this is the first report in which neutralizing antibodies to respiratory syncytial virus have been evoked using bacterially expressed G immunogenic domain protein without any adjuvant. (C) 2011 International Union of Biochemistry and Molecular Biology, Inc. Volume 58, Number 2, March/April 2011, Pages 97-102. E-mail: kyusoff@biotech.upm.edu.my

The H5 gene of avian influenza virus (AIV) strain A/chicken/Malaysia/5744/2004(H5N1) was cloned into pcDNA3.1 vector, and Esat-6 gene of Mycobacterium tuberculosis was fused into downstream of the H5 gene as a genetic adjuvant for DNA vaccine candidates. The antibody level against AIV was measured using enzyme-linked immunosorbent assay (ELISA) and haemagglutination inhibition (HI) test. Sera obtained from specific-pathogen-free chickens immunized with pcDNA3.1/H5 and pcDNA3.1/H5/Esat-6 demonstrated antibody responses as early as 2 weeks after the first immunization. Furthermore, the overall HI antibody titer in chickens immunized with pcDNA3.1/H5/Esat-6 was higher compared to the chickens immunized with pcDNA3.1/H5 (p < 0.05). The results suggested that Esat-6 gene of M. tuberculosis is a potential genetic adjuvant for the development of effective H5 DNA vaccine in chickens.

Aims: A system for displaying heterologous respiratory syncytial virus (RSV) glycoproteins on the surface of Lactococcus lactis NZ9000 was developed. Methods and Results: Fusion of the USP45 signal peptide and the cA (C terminus of the peptidoglycan-binding) domains of AcmA, a major autolysin from L. lactis, to the N- and C-terminal of the target proteins, respectively, was carried out. The target protein was the major immunogenic domain of either the F (40·17-kDa) or G (11·49-kDa) glycoprotein domains of the RSV. Whole-cell ELISA readings obtained after 24 h of induction showed an increase in protein expression as the cA domain repeats increased, for the G glycoprotein of RSV. On the other hand, the F glycoprotein indicated decreasing expression levels as the number of cA domain repeats increased. The difference in the expression levels of the F and G domains may be attributed to the different sizes of the antigenic domains. Conclusions: The size and properties of the target proteins are vital in determining the amount of antigenic domains being displayed on the surface of live cells. Significance and Impact of the Study: The system demonstrated here can aid in the utilization of the generally regarded as safe (GRAS) bacteria L. lactis, as a vaccine delivery vehicle to surface display the antigenic proteins of RSV.

In order to study the effect of Tryptophan (Tip) on growth performance and blood parameters of broiler chickens, an in vivo experiment was conducted. A corn-soybean meal based diet containing different levels of Trp (0, 0.10 and 0.20) for the starter, (0, 0.07 and 0.15) for the grower and (0, 0.05 and 0.13) for the finisher was used. In a completely randomized design with three treatments of five replicates each and 10 chickens per replicate, 150 Cobb500 male broiler chickens from 0-49 days of age were subjected to Trp diet. Growth performance (body weight gain, feed intake and feed, gain ratio) and blood serum (albumin, total protein, glucose, cholesterol, triglyceride, urea, uric acid, aspartate amino-transferase, alanine amino-transferase, alkalin phosphatase, lactic dehydrogenase and creatine. e kinase) parameters were measured at 27 and 49 days of age. As the result showed the increase of dietary Trp elevated (p 0.05) body weight gain, feed intake, albumin, total protein, glucose, urea and uric acid and decreased (p>0.05) feed gain ratio, aspartate amino-transferase, lactic dehydrogenase, triglyceride and cholesterol. Therefore, we conclude that dietary Tip might have positive effects on health status of the broiler chickens.

Infectious bursal disease is still a challenging issue by posing a serious threat to the commercial poultry industry especially due to the emergence of highly Infectious Bursal Disease Virus (IBDV). In the present study, we evaluated the immunomodulatory effects of Tryptophan (Tip) on innate, humoral and cellular immune responses in chickens challenged by oral administration of intermediate plus strain of IBD virus at 28 days of age. A corn-soybean meal based diet containing different levels of Tip (0, 0.10 and 0.20) for the starter, (0, 0.07 and 0.15) for the grower and (0, 0.05 and 0.13) for the finisher has been utilized. In a completely randomized design with three treatments of five replicates each and 10 chickens per replicate, 150 Cobb 500 male broiler chickens from 0-49 days of age were subjected to Tip diet. To measure the innate, cellular and humoral immunity indicators (interferon-alpha, interferon-gamma, immunoglobulin G, respectively) at 27, 35, 42 and 49 days of age, serum samples from each replicate of treatments were collected and subjected to ELISA. The result showed that Trp supplementation in the chickens basal diets significantly increased the serum levels of interferon-alpha at 35, 42 and 49 days of age (p

Infectious Bursal Disease (IBD) continues to pose potential threat to poultry industry all over the world. The disease can spell disaster not only through its infection but also by break of immunity in chickens vaccinated for other diseases. On the other hand, arginine (Arg), a ubiquitous, semi-essential amino acid has emerged as an imunostimulant from variety of human and animal studies. In the present study, we demonstrate the stimulatory effects of Arg on systemic immune response in chickens challenged by orally administration of intermediate plus strain of IBD virus at 28 days of age. A corn-soybean meal based diet containing different levels of Arg (0, 0.67, 1.37, 2.07 and 2.77) for the starter, (0, 0.53, 1.1, 1.68 and 2.25) for the grower and (0, 0.52, 1.04, 1.56 and 2.08) for the finisher was used. In a completely randomized design with five treatments of five replicates each and 10 chickens per replicate, 250 Cobb500 male broiler chickens from 0-49 days of age were used. To measure the innate, cellular and humoral immunity indicators (interferon-α, interferon-γ, immunoglobulin G) at 27, 35, 42 and 49 days of age, serum samples from each replicate of treatments were collected and subjected to ELISA. The result showed that Arg supplementation in the chickens basal diets significantly increased the serum levels of interferon-α, interferon-γ, immunoglobulin G at 35, 42 and 49 days of age (p

An experiment was conducted to determine effects of dietary Arginine (ARG) on growth performance and blood serum parameters in broiler chickens. A corn-soybean meal based diet containing different levels of ARG (0, 0.67, 1.37, 2.07 and 2.77) for the starter (0, 0.53, 1.1, 1.68 and 2.25) for the grower and (0, 0.52, 1.04, 1.56 and 2.08) for the finisher was used. In a completely randomized design with five treatments of five replicates each and 10 chickens per replicate, 250 Cobb 500 male broiler chickens from 0-49 days of age were used. Growth performance (body weight gain, feed intake and feed: gain ratio) and blood serum (albumin, total protein, glucose, cholesterol, triglyceride, urea, uric acid, aspartate amino-transferase, alanine amino-transferase, alkalin phosphatase, lactic dehydrogenase and creatine kinase) parameters were measured at 27 and 49 days of age. Increase of dietary ARG increased (p 0.05) aspartate amino-transferase and cholesterol. It was concluded that dietary ARG might have positive effects on health status of the broiler chickens.

Methicillin-resistant Staphylococcus aureus (MRSA) is a nosocomial pathogen of increasing risk on community. This study aims at determining the risk of MRSA transfer from coins as silent and underestimated reservoir in community. One hundred swabs from coins were collected from college students in Malaysia. A series of identification and differentiating tests were conducted for precise identification of MRSA bacteria. Moreover, this study compared the efficacy of the different identification tests with gold standard, polymerase chain reaction (PCR) assay. The tests used were tube coagulase, DNase agar test, antibiogram, several routine biochemical identification tests and PCR assays. PCR assay used specific primers for resistance or ID -related genes: mecA, ermA, ermB, ermC, msrA, linA, femA, and nuc genes. A total of 37 bacterial isolates were isolated from college students’ coins; non-PCR assays of identification and resistance detection revealed the presence and spread of MRSA in coins of 2 college students. PCR-amplification of the nuc gene was used as a baseline test to detect S. aureus. PCR showed only one isolate as true MRSA. False positive MRSA by disc diffusion assay might be attributed to low pH and high thymidine content of Muller Hinton agar medium. Collectively, coins proved to be possible source for the transfer of MRSA in community of college students in South East Asia. Moreover, PCR assay for identification of S. aureus resistance proved to be superior on other methods. Key words : Staphylococcus aureus , MRSA, nail, PCR, resistance.

Allium hirtifoliumBoiss. known as Persian shallot, is a spice used as a traditional medicine in Iran and, Mediterranean region. In this study, the chemical composition of the hydromethanolic extract of this plant was analyzed using GC/MS. The result showed that 9-hexadecenoic acid, 11,14-eicosadienoic acid, and n-hexadecanoic acid are the main constituents. The antibacterial activity of the shallot extract was also examined by disk diffusion and microdilution broth assays. It was demonstrated that Persian shallot hydromethanolic extract was effective against 10 different species of pathogenic bacteria including methicillin resistantStaphylococcus aureus(MRSA), methicillin sensitiveStaphylococcus aureus(MSSA),Staphylococcus aureus,Staphylococcus epidermidis,Streptococcus pneumoniae,Escherichia coli, Escherichia coliO157:H7,Salmonella typhimurium, Proteus mirabilis, andKlebsiella pneumoniae. Specifically, the minimum concentration of the extract which inhibited bacterial growth (MIC values) was 1.88 mg/mL for most of the gram-positive bacteria. This concentration was not much different from the concentration that was safe for mammalian cells (1.50 mg/mL) suggesting that the hydromethanolic extract of Persian shallot may be a safe and strong antibacterial agent.

Human respiratory syncytial virus (HRSV) is a leading pathogen causing lower respiratory tract infections in infants and young children worldwide. In line with the development of an effective vaccine against HRSV, a domain of the fusion (F) glycoprotein of HRSV was produced and its immunogenicity and antigenic properties, namely the effect of deficient glycosylation was examined. A His-tagged recombinant F (rF) protein was expressed in Escherichia coli, solubilized with 8 mol/l urea, purified by the Ni-NTA affinity chromatography and used for the raising of a polyclonal antibody in rabbits. The non-glycosylated rF protein proved to be a strong immunogen that induced a polyclonal antibody that was able to recognize also the glycosylated F1 subunit of native HRSV. The other way around, a polyclonal antibody prepared against the native HRSV was able to react with the rF protein. These results indicated that glycosylation was not necessary for the F domain aa 212-574 in order to be recognized by the specific polyclonal antibody.

Background/Aims: Newcastle disease virus (NDV) is an avian paramyxovirus that has gained a lot of interest in cancer viro-therapeutic applications because of its ability to selectively induce apoptosis in human cancer cells. However, the underlying mechanisms by which NDV induces apoptosis in human cancer cells are still not entirely understood. Methods: In this study we examined the effect of a Malaysian velogenic strain of NDV, known as AF2240, on some elements of the intrinsic pathway of apoptosis. Results: We show that NDV infection leads to conformational change of Bax protein. This is associated with the translocation of Bax from the cytoplasm to mitochondria and the release of cytochrome c into the cytoplasm. Interestingly, the level of Bcl-2 protein was not affected by NDV treatment. Conclusion: We have shown that Bax conformational change and subcellular distribution is involved in the intrinsic pathway of apoptosis induced by NDV.

Background Avian influenza viruses (AIV) cause high morbidity and mortality among the poultry worldwide. Their highly mutative nature often results in the emergence of drug resistant strains, which have the potential of causing a pandemic. The virus has two immunologically important glycoproteins, hemagglutinin (HA), neuraminidase (NA), and one ion channel protein M2 which are the most important targets for drug discovery, on its surface. In order to identify a peptide-based virus inhibitor against any of these surface proteins, a disulfide constrained heptapeptide phage display library was biopanned against purified AIV sub-type H9N2 virus particles. Results After four rounds of panning, four different fusion phages were identified. Among the four, the phage displaying the peptide NDFRSKT possessed good anti-viral properties in vitro and in ovo. Further, this peptide inhibited the hemagglutination activity of the viruses but showed very little and no effect on neuraminidase and hemolytic activities respectively. The phage-antibody competition assay proved that the peptide competed with anti-influenza H9N2 antibodies for the binding sites. Based on yeast two-hybrid assay, we observed that the peptide inhibited the viral replication by interacting with the HA protein and this observation was further confirmed by co-immunoprecipitation. Conclusion Our findings show that we have successfully identified a novel antiviral peptide against avian influenza virus H9N2 which act by binding with the hemagglutination protein of the virus. The broad spectrum activity of the peptide molecule against various subtypes of the avian and human influenza viruses and its comparative efficiency against currently available anti-influenza drugs are yet to be explored.

The yeast two-hybrid system has been used to identify domains of the Newcastle disease virus (NDV) phosphoprotein (P) involved in self-association and interaction with the nucleocapsid protein (NP). Deletion analysis was used to map the domain(s) of the P protein involved in P:P and P:NP interactions. The C-terminal 45 amino acids (residues 247-291) were shown to play a major role in both of the interactions. Comparison of these findings with other reports suggests that paramyxoviruses are different with respect to interaction domain(s) between these two essential viral proteins involved in genome replication.

The underlying mechanisms by which Newcastle disease virus (NDV) kills cancer cells are still unclear. Recent discoveries have shown that many viruses contain Bcl-2 homology-like domains which enabled their interaction with Bcl-2 family members, and thereby accounting for their virulence and pathogenicity. Alignment of the protein sequences of Malaysian strain of NDV, known as AF2240, with those from members of the human Bcl-2 family showed many similar regions; most notably we found that its matrix (AF2240-M) protein, large (AF2240-L) protein and fusion (AF2240-F) protein all contain BH3-like regions. In addition, there are BH1-like domains in these proteins, where AF2240-F and Mcl-1 share 55% identity within this region. To further investigate our hypothesis that the presence of the BH3-like domains in these proteins may convey cytotoxicity, AF2240-M and AF2240-F genes were cloned into pFLAG and pEGFP.N2 vectors and transfected into HeLa cells. The expression of these constructs promoted cell death. As shown by flow cytometry, AF2240-M protein with deleted BH3-like region showed five-fold decrease in apoptosis. Moreover, the construct containing the N-terminal of AF2240-M showed nearly the same cell death rate as to that of the full-length protein, strongly suggesting that the BH3-like domain within this protein participates in promoting cell death. Moreover, AF2240-M transfection promoted Bax redistribution to mitochondria. Therefore, to determine whether there is any direct interaction between NDV viral proteins with some members of the Bcl-2 family, various constructs were co-transfected into HeLa cells. Co-immunoprecipitation trials showed that the AF2240-M indeed directly interacted with Bax protein via its BH3-domain, as the mutant proteins failed to interact with Bax. AF2240-F failed to interact with any of the tested proteins, although Bcl-XL slowed down the rate of cell death caused by this construct by nearly five-fold. In a parallel experiment, the level of expression of endogenous Bax and Bcl-2 after infection of HeLa cells with NDV was assessed by qRT-PCR, but no statistically significant change was observed. Consequently, the Bax/Bcl-2 ratio at the mRNA level did not alter. Overall, our study has shed additional light into the mechanisms by which NDV induces apoptosis.