BubR1 depletion delays apoptosis in the microtubule-depolymerized cells.

We investigated the role of a spindle assembly checkpoint protein, BubR1, in determining the mechanism of cell killing of an anti-microtubule agent CXI-benzo-84. CXI-benzo-84 dampened microtubule dynamics in live MCF-7 cells. The compound arrested MCF-7 cells in mitosis and induced apoptosis in these cells. Though CXI-benzo-84 efficiently depolymerized microtubules in the BubR1-depleted MCF-7 cells, it did not arrest the BubR1-depleted cells at mitosis. Interestingly, apoptosis occurred in the BubR1-depleted MCF-7 cells in the absence of a mitotic block suggesting that the mitotic block is not a prerequisite for the induction of apoptosis by anti-microtubule agents. In the presence of CXI-Benzo-84, the level of apoptosis was initially found to be lesser in the BubR1-depleted MCF-7 cells than the control cells; however, the BubR1-depleted cells displayed a similar level of apoptosis as the control cells at 72 h of drug treatment. The depletion of BubR1 enhanced DNA damage in MCF-7 cells upon microtubule depolymerization. In addition, CXI-benzo-84 in combination with cisplatin induced more cell death in BubR1-depleted cells than the BubR1-expressing MCF-7 cells. The results indicated a possibility that the BubR1-compromised cancer patients can be treated with combination therapy.

Anticoccidial activity of fruit peel of Punica granatum L.

In the interests of food safety and public health, plants and their compounds are now re-emerging as an alternative approach to treat parasitic diseases. Here, we studied the anticoccidial effect of different solvent extracts of the fruit peel of Punica granatum-a commercial waste from pomegranate juice industries. The hope underlying these experiments was to find a sustainable natural product for controlling coccidiosis. The plant extracts were prepared using solvents of different polarity. Acute oral toxicity study was first carried out to see the safety of crude extracts. A high dose of crude extracts (300 mg/kg body weight) was tested for possession of anticoccidial activity against experimentally induced coccidial infection in broiler chicken. Activity was measured in comparison to the reference drug amprolium on the basis of oocyst output reduction, mean weight gain of birds and feed conversion ratio. Oocyst output was measured using Mc-Masters counting technique. Acute oral toxicity study showed that crude extracts of P. granatum are safe up to dosage of 2000 mg/kg body weight. LD50 was not determined as mortalities were not recorded in any of the five groups of chicken. For anticoccidial activity crude methanolic extract (CME) of the fruit peel of P. granatum showed the maximum effect as evident by oocyst output reduction (92.8 ±â€¯15.3), weight gain of birds (1403.0 ±â€¯11.9 g) and feed conversion ratio (1.66 ±â€¯0.04), thereby affirming the presence of alcohol soluble active ingredients in the plant. We also tested different doses (100-400 mg/kg body weight) of the CME of the fruit peel of P. granatum, the most active extract on E. tenella and observed a dose dependent effect. From the present study it can be concluded that alcoholic extract of the fruit peel of P. granatum has significant potential to contribute to the control of coccidian parasites of chicken.

Anticoccidial activity of Artemisia vestita (Anthemideae, Asteraceae) - a traditional herb growing in the Western Himalayas, Kashmir, India.

Coccidiosis, caused by various species of genus Eimeria, is a major parasitic disease in chicken. The increasing resistance of these parasites to currently used anticoccidial drugs has stimulated the search for new methods of control. As part of this effort, a study was designed to see the anticoccidial effect of different solvent extracts of Artemisia vestita-a traditional herb growing in Kashmir Himalayas. The plant extracts were prepared using different solvents. Preliminary toxicity study was first carried out to see the safety of crude plant extracts. A high dose of crude extracts (300 mg/kg body weight) was tested for possession of anticoccidial activity against experimentally induced coccidial infection in broiler chicken. Activity was measured in comparison to the reference drug amprolium on the basis of oocyst output reduction, mean weight gain of birds and feed conversion ratio. Oocyst output was measured using Mc-Masters counting technique. Preliminary toxicity study showed that crude extracts of A. vestita are safe up to dosage of 2000 mg/kg body weight. LD50 was not determined as mortalities were not recorded in any of the five groups of chicken. For anticoccidial activity crude methanolic extract (CME) of A. vestita showed the maximum effect as evident by oocyst output reduction (71.5 ± 12.2), weight gain of birds (1406.4 ± 12.2) and feed conversion ratio (1.58 ± 0.06), thereby affirming the presence of alcohol soluble active ingredients in the plant. We also tested different doses (100-400 mg/kg body weight) of the CME of A. vestita, the most active extract on E. tenella and observed a dose dependent effect. From the present study it can be concluded that alcoholic extract of A. vestita has the immense potential to contribute to the control of coccidian parasites of chicken. Our results corroborate the use of genus Artemisia and could justify its use in folk medicine for treatment of parasitic diseases.

Breaking the resistance of Escherichia coli: Antimicrobial activity of Berberis lycium Royle.

The antimicrobial activity of root bark of Berberis lycium and its principal component berberine was tested against a panel of microbial strains using agar well diffusion test and further analyzed using micro-broth dilution method. Preliminary analysis, on the basis of zone of Inhibition (ZOI) showed that the methanolic extract of B. lycium was highly effective against Escherichia coli (ZOI 41 ± 1 mm). Among the bacterial strains E. coli was found to be most susceptible and among fungi Candida albicans was the most susceptible for berberine as well as the crude methanolic extract of the plant. Methanolic extract of the plant was more effective for E. coli (MIC 1.7 ± 1.18; MBC 2.4 ± 1.18) than berberine (MIC 3.5 ± 0.57) (p < 0.05), whereas berberine was more effective than crude extracts for C. albicans. In addition, E. coli showed the development of resistant colonies after 72 h when tested with berberine but the development of such colonies was not observed with the methanolic extract of the plant. This could be due to the presence of resistance breaking molecules in the crude methanolic extract of B. lycium. Also the MIC index of crude methanolic extract was 1.39 for E. coli, which showed the mode of action to be bactericidal. HPLC analysis revealed the presence of berberine at highest concentration in methanolic extract of the plant, followed by aqueous extract. Potentiation of this berberine by resistance breaking molecules in the crude extract could be a possible explanation for its strong effectiveness.

Anticoccidial activity of aqueous extract of a wild mushroom (Ganoderma applanatum) during experimentally induced coccidial infection in broiler chicken.

Despite presence of anticoccidial drugs and vaccines in the market, coccidiosis continues to result in substantial economic losses to the poultry industry. There is wide-spread resistance to already known anticoccidial drugs. It is an infectious disease of poultry and rigorous management is required during vaccination. In addition there is possibility of drug residues in meat and other byproducts of such treated animals and consequently makes more imperative to explore and understand the role of natural products in livestock parasite management. Therefore a study was designed to evaluate the anticoccidial activity of aqueous extract of Ganoderma applanatum in broiler chicken. In-vivo anticoccidial activity of aqueous extract of G. applanatum was measured in comparison to the reference drug amprolium on the basis of oocysts per gram of faeces, weight gain and feed conversion ratio. Oocyst output was measured with the help of Mc-Masters counting technique. The results of this study established the virulence of coccidian oocysts and the effectiveness of both amprolium and aqueous extract of G. applanatum against coccidian oocysts, confirmed by the fact that treatment with aqueous extract of G. applanatum resulted in a noticeable reduction in coccidian oocysts output, leading to improved weight gain and better feed conversion ratio. The study highlighted the potential of G. applanatum as a natural source of bioactive components for controlling a protozoan parasite, which can be isolated and tested in a bioassay-guided manner and harnessed in the form of anticoccidial drugs.

Synergistic approach for treatment of chicken coccidiosis using berberine--A plant natural product.

Despite the advent of anticoccidial drugs and vaccines, coccidiosis continues to result in substantial economic losses to the poultry industry. Berberine, a natural alkaloid is well known in studies involving synergistic approaches, thereby reducing the dosage of principal drugs. Therefore, a study was designed to see whether a synergistic anticoccidial effect could be obtained between amprolium and berberine, in vivo using broiler chicken. Anticoccidial activity was measured in comparison to the reference drug amprolium on the basis of oocyst output reduction, mean weight gain and feed conversion ratio. Oocyst output was measured using Mc-Masters counting technique. Different combinations of berberine and amprolium were tested and out of which 1:1 ratio was the most effective for controlling these parasites. Oral gavaging of 100(50 + 50) mg/kg body weight of 1:1 ratio of amprolium and berberine caused the equivalent reduction in number of oocysts (38.85 ± 9.61) one day prior to that of standard drug amprolium (49.95 ± 16.65) as well as pure berberine (44.4 ± 9.61) used in the study. Weight gain of birds was also highest in the synergistic group (1547.43 ± 12.86) among all the infected groups. Besides feed conversion ratio in the synergistic group was also better (1.387 ± 0.026). The results of this study proved the effectiveness of both amprolium and berberine and revealed synergism between amprolium and berberine against coccidian oocysts, confirmed by significant reduction in the number of coccidian oocysts shed in the feces, leading to better weight gain and improved feed conversion ratio. The study deep-rooted the synergistic potential of berberine, a natural bioactive compound for controlling a protozoan parasite and the results of this study corroborate with its use for treatment of severe diarrhoea, amoebiasis and intestinal infections.

Seasonal impact on the prevalence of coccidian infection in broiler chicks across poultry farms in the Kashmir valley.

The information on the epidemiology and control of coccidian parasites of broilers in Kashmir valley is based on the reports available from other regions of the world. With this background, the present study was conducted to investigate the seasonal prevalence of the disease in the temperate agro-climatic conditions of Kashmir valley. A standard protocol for sampling was followed according to which five birds per 10,000 is sufficient to diagnose coccidiosis. Microscopic examination (under 10× and 40× objective lens) was used to reveal the presence of coccidial oocysts. Different species of genus Eimeria were identified on the basis of their predilection site, morphology and size. Coccidiosis was most prevalent in autumn 45.12 ± 2.55 (September 47.5 %, October 42.42 % and November 45.46 %) followed by summer 30.84 ± 6.86, spring 23.81 ± 2.81 and winter 20.29 ± 6.40. In summer, prevalence of disease was low but afterwards prevalence of disease rose up from August to October. In spite of high relative humidity in winter, disease showed low prevalence rate. Over all prevalence for the whole year was 29.87 %. Among species Eimeria tenella was the most dominant parasite showing highest prevalence of 18.13 %. Variation in incidence of coccidiosis with respect to seasons showed a strong correlation and data was found to be statistically significant with P < 0.05. The results obtained would be quite useful to devise appropriate and effective control strategies and prophylactic programs for coccidiosis in poultry unique to this climatic zone and other parts of the world with similar climatic and poultry production systems.

Morphological characterization of nematodes of the genera Capillaria, Acuaria, Amidostomum, Streptocara, Heterakis, and Ascaridia isolated from intestine and gizzard of domestic birds from different regions of the temperate Kashmir valley.

During a random survey of parasitisation of poultry birds of Kashmir, conducted for 3 years more than 300 poultry birds were randomly collected from different parts of the Kashmir valley and were examined for the helminth parasites. The most dominant group of parasites which was parasitizing the poultry birds was found to be the Cestode, whose prevalence in the infected birds was 76.9 %. Nematodes were found to be the second dominant group of parasites after cestodes with the prevalence of 70 %. Among the different species of nematodes Heterakis gallinae were found infecting almost all the birds autopsied. The highest intensity of H. gallinae might be because of the fact that it requires cockroaches, beetles and grasshoppers as intermediate hosts in addition of having a direct life cycle. Nematodes which were recovered from the intestine and gizzard of the birds belong to the genera Capillaria, Acuaria, Amidostomum, Streptocara, Heterakis, and Ascaridia.

Novel third-generation water-soluble noscapine analogs as superior microtubule-interfering agents with enhanced antiproliferative activity.

Noscapine, an opium-derived 'kinder-gentler' microtubule-modulating drug is in Phase I/II clinical trials for cancer chemotherapy. However, its limited water solubility encumbers its development into an oral anticancer drug with clinical promise. Here we report the synthesis of 9 third-generation, water-soluble noscapine analogs with negatively charged sulfonato and positively charged quaternary ammonium groups using noscapine, 9-bromonoscapine and 9-aminonoscapine as scaffolds. The predictive free energy of solvation was found to be lower for sulfonates (6a-c; 8a-c) compared to the quaternary ammonium-substituted counterparts, explaining their higher water solubility. In addition, sulfonates showed higher charge dispersability, which may effectively shield the hydrophobicity of isoquinoline nucleus as indicated by hydrophobicity mapping methods. These in silico data underscore efficient net charge balancing, which may explain higher water solubility and thus enhanced antiproliferative efficacy and improved bioavailability. We observed that 6b, 8b and 8c strongly inhibited tubulin polymerization and demonstrated significant antiproliferative activity against four cancer cell lines compared to noscapine. Molecular simulation and docking studies of tubulin-drug complexes revealed that the brominated compound with a four-carbon chain (4b, 6b, and 8b) showed optimal binding with tubulin heterodimers. Interestingly, 6b, 8b and 8c treated PC-3 cells resulted in preponderance of mitotic cells with multipolar spindle morphology, suggesting that they stall the cell cycle. Furthermore, in vivo pharmacokinetic evaluation of 6b, 8b and 8c revealed at least 1-2-fold improvement in their bioavailability compared to noscapine. To our knowledge, this is the first report to demonstrate novel water-soluble noscapine analogs that may pave the way for future pre-clinical drug development.

In vivo anticoccidial activity of berberine [18, 5,6-dihydro-9,10-dimethoxybenzo(g)-1,3-benzodioxolo(5,6-a) quinolizinium]--an isoquinoline alkaloid present in the root bark of Berberis lycium.

Coccidiosis, caused by various Eimeria species, is a major parasitic disease in chicken. However the increasing resistance of these parasites to currently used anticoccidial drugs has stimulated the search for new methods of control. As part of this effort we investigated the root bark of Berberis lycium (barberry) as a potential source of compounds with anticoccidial activity. In the present study anticoccidial activity of different solvent extracts of the root bark of B. lycium and berberine was evaluated in vivo using broiler chicken. Results of the study demonstrated equipotent efficacy of pure berberine in comparison to that of standard drug amprolium on the basis of reduction in coccidian oocyst output, body weight gain of chicken and feed conversion ratio. Among the extracts crude methanolic extract showed highest anticoccidial activity tested at 300 mg/kg body weight which could be due to the presence of alcohol-soluble active ingredients in root bark of B. lycium. Toxicological studies revealed that B. lycium extracts as well as berberine were not lethal up to dosage of 2000 mg/kg body weight. LD(50) was not determined as mortalities were not recorded in any of the five groups of chicken. From the present study it can be concluded that root bark of B. lycium has the immense potential to contribute to the control of coccidian parasites of chicken. Our results corroborate the use of berberine for treatment of severe diarrhoea, amoebiasis and intestinal infections and could justify its use in folk medicine for treatment of haemorrhagic dysentery.