Retinal proteomics of experimental glaucoma model reveal intraocular pressure-induced mediators of neurodegenerative changes

Current evidence suggests that exposure to chronically induced intraocular pressure (IOP) leads to neurodegenerative changes in the inner retina. This study aimed to determine retinal proteomic alterations in a rat model of glaucoma and compared findings with human retinal proteomics changes in glaucoma reported previously. We developed an experimental glaucoma rat model by subjecting the rats to increased IOP (9.3 ± 0.1 vs 20.8 ± 1.6 mm Hg) by weekly microbead injections into the eye (8 weeks). The retinal tissues were harvested from control and glaucomatous eyes and protein expression changes analysed using a multiplexed quantitative proteomics approach (TMT-MS3). Immunofluorescence was performed for selected protein markers for data validation. Our study identified 4304 proteins in the rat retinas. Out of these, 139 proteins were downregulated (≤0.83) while the expression of 109 proteins was upregulated (≥1.2-fold change) under glaucoma conditions (P ≤ .05). Computational analysis revealed reduced expression of proteins associated with glutathione metabolism, mitochondrial dysfunction/oxidative phosphorylation, cytoskeleton, and actin filament organisation, along with increased expression of proteins in coagulation cascade, apoptosis, oxidative stress, and RNA processing. Further functional network analysis highlighted the differential modulation of nuclear receptor signalling, cellular survival, protein synthesis, transport, and cellular assembly pathways. Alterations in crystallin family, glutathione metabolism, and mitochondrial dysfunction associated proteins shared similarities between the animal model of glaucoma and the human disease condition. In contrast, the activation of the classical complement pathway and upregulation of cholesterol transport proteins were exclusive to human glaucoma. These findings provide insights into the neurodegenerative mechanisms that are specifically affected in the retina in response to chronically elevated IOP.     

New heat shock protein (Hsp90) inhibitors,designed by pharmacophore modeling and virtual screening: synthesis,biological evaluation and molecular dynamics studies

Abstract

Inhibition of heat shock protein 90 (Hsp90) is known to be a significantly effective strategy in cancer therapy. Here, pyrazolopyranopyrimidine derivatives were characterized as new Hsp90 inhibitors. The molecules’ key structure (ZINC02819805) was determined by utilizing a pharmacophore model virtual screening workflow. Structural optimization was then carried out on compound ZINC02819805, pyrazolopyranopyrimidine derivatives were designed and six chosen derivatives were synthesized. The inhibition of Hsp90 ATPase activity of synthesized compounds revealed that para methylphenyl derivative of pyrazolopyranopyrimidine (HM3) was the most potent inhibitor (IC₅₀ = 5.5?µM). The anti-proliferative activity of this compound was evaluated against a panel of cell lines including MCF-7, HeLa and HUVEC (IC₅₀ = 1.28?µM, IC₅₀ = 1.74?µM and IC₅₀ = 61.48?µM respectively) by MTT method. The western blot analysis of treated MCF-7 cells with compound HM3 showed that the expression level of Hsp70 and Her2 proteins changed. The high level of Hsp70 expression and low level of Her2 expression suggest that compound HM3 exhibits inhibitory effect on Hsp90. Finally, the key interactions between HM3 and Hsp90 protein were studied by molecular dynamics simulation and showed that compound HM3 was stable in Hsp90 active cite during 200?ns simulation. Abbreviations Hsp90 Heat shock protein 90

MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

ATP adenosine triphosphate

MD molecular dynamics simulation

RMSD root-mean-square deviation

RMSF root-mean-square fluctuation

Rg gyration radius

m-SABNPs boehmite nanoparticles-supported sulfamic acid

Communicated by Ramaswamy H. Sarma     

Ethnobotanical appraisal and cultural values of medicinally important wild edible vegetables of Lesser Himalayas-Pakistan

Background

The association among food and health is momentous as consumers now demand healthy, tasty and natural functional foods. Knowledge of such food is mainly transmitted through the contribution of individuals of households. Throughout the world the traditions of using wild edible plants as food and medicine are at risk of disappearing, hence present appraisal was conducted to explore ethnomedicinal and cultural importance of wild edible vegetables used by the populace of Lesser Himalayas-Pakistan.

Methods

Data was collected through informed consent semi-structured interviews, questionnaires, market survey and focus group conversation with key respondents of the study sites including 45 female, 30 children and 25 males. Cultural significance of each species was calculated based on use report.

Results

A total of 45 wild edible vegetables belonging to 38 genera and 24 families were used for the treatment of various diseases and consumed. Asteraceae and Papilionoideae were found dominating families with (6 spp. each), followed by Amaranthaceae and Polygonaceae. Vegetables were cooked in water (51%) followed by diluted milk (42%) and both in water and diluted milk (7%). Leaves were among highly utilized plant parts (70%) in medicines followed by seeds (10%), roots (6%), latex (4%), bark, bulb, flowers, tubers and rhizomes (2% each). Modes of preparation fall into seven categories like paste (29%), decoction (24%), powder (14%), eaten fresh (12%), extract (10%), cooked vegetable (8%) and juice (4%). Ficus carica was found most cited species with in top ten vegetables followed by Ficus palmata, Bauhinia variegata, Solanum nigrum, Amaranthus viridis, Medicago polymorpha, Chenopodium album, Cichorium intybus, Amaranthus hybridus and Vicia faba.

Conclusions

Patterns of wild edible plant usage depend mainly on socio-economic factors compare to climatic conditions or wealth of flora but during past few decades have harshly eroded due to change in the life style of the inhabitants. Use reports verified common cultural heritage and cultural worth of quoted taxa is analogous. Phytochemical analysis, antioxidant activities, pharmacological applications; skill training in farming and biotechnological techniques to improve the yield are important feature prospective regarding of wild edible vegetables.

     

Botanical ethnoveterinary therapies in three districts of the Lesser Himalayas of Pakistan

Background

Ethnoveterinary knowledge is highly significant for persistence of traditional community-based approaches to veterinary care. This is of particular importance in the context of developing and emerging countries, where animal health (that of livestock, especially) is crucial to local economies and food security. The current survey documents the traditional veterinary uses of medicinal plants in the Lesser Himalayas-Pakistan.

Methods

Data were collected through interviews, focus groups, participant observation, and by administering questionnaires. A total of 105 informants aged between 20–75 years old who were familiar with livestock health issues (i.e. farmers, shepherds, housewives and herbalists) participated in the study.

Results

A total of 89 botanical taxa, belonging to 46 families, were reported to have ethnoveterinary applications. The most quoted families were Poaceae (6 taxa), Fabaceae (6), Asteraceae (5), and Polygonaceae (5). Adhatoda vasica was the most cited species (43%), followed by Trachyspermum ammi (37%), and Zanthoxylum armatum var. armatum (36%). About 126 medications were recorded against more than 50 veterinary conditions grouped into seven categories. The highest cultural index values were recorded for Trachyspermum ammi, Curcuma longa, Melia azedarach, Zanthoxylum armatum var. armatum and Adhatoda vasica. The highest informant consensus factor was found for pathologies related to respiratory and reproductive disorders. Comparison with the local plant-based remedies used in human folk medicine revealed that many of remedies were used in similar ways in local human phytotherapy. Comparison with other field surveys conducted in surrounding areas demonstrated that approximately one-half of the recorded plants uses are novel to the ethnoveterinary literature of the Himalayas.

Conclusion

The current survey shows a remarkable resilience of ethnoveterinary botanical knowledge in the study area. Most of the species reported for ethnoveterinary applications are wild and under threat. Thus, not only is it imperative to conserve traditional local knowledge of folk veterinary therapies for bio-cultural conservation motives, but also to assist with in-situ and ex-situ environmental conservation initiatives, which are urgently needed. Future studies that focus on the validation of efficacy of these ethnoveterinary remedies can help to substantiate emic concepts regarding the management of animal health care and for rural development programs.

     

THY1 as a reliable marker for enrichment of undifferentiated spermatogonia in the goat

Spermatogonial stem cells are unique cells of testes that can restore fertility upon transplantation into recipient testes. However, use of suitable markers for enrichment of these cells have important potential application. THY1, is an established conserved marker of spermatogonial stem cells in bovine, rodents, and primates, but there is no information available in goats. After three rounds of enzymatic digestion of prepubertal goat testicular tissues, undifferentiated spermatogonia positive for THY1 were isolated by magnetic-activated cell sorting and were used for immunocytochemistry, real-time polymerase chain reaction analysis for gene expression, protein expression, and transplantation into recipient mice. Immunocytochemical analyses showed that significantly higher percentage of THY1⁺ cells were positive for PLZF and VASA when compared with unselected population. This result for PLZF was further confirmed at the protein level. Real-time polymerase chain reaction analysis revealed that expression of THY1, PLZF, VASA, BCL6B, and UCHL1 as SCCs characteristic genes in THY1⁺ cells was significantly higher than in the initial population. Finally, transplantation of PKH26-labeled cells revealed that THY1⁺ cells had higher capacity for colony formation when compared with unselected cells. In conclusion, the results provide indications that THY1 surface marker can be reliably used for enrichment of undifferentiated spermatogonial in the goats.     

Effect of bark extract of different plant species on the hatching of Meloidogyne incognita

An in vitro experiment was conducted to evaluate the effect of bark extract of different plant species viz., Azadirachta indica, Acacia nilotica, Emblica officinalis, Eucalyptus citriodora, Mangifera indica and Terminalia arjuna at varying concentration levels S, S/2, S/10 and S/100 on the hatching of Meloidogyne incognita. One hundred per cent inhibition was observed with Azadirachta indica under both un-decomposed and decomposed conditions. The effect was decreased with Acacia nilotica, Emblica officinalis, Eucalyptus citriodora, Mangifera indica and Terminalia arjuna, respectively. Effect of decomposed bark extract on hatching was more pronounced as compared to un-decomposed bark extract.     

Assessment of the effect of biosurfactant produced by Pseudomonas aeruginosa in lethality of Bacillus thuringiensis Berl. against 3rd instars larvae of white cabbage butterfly (Pieris brassicae L.)

Surfactant that is produced from cheap sources like oil sludge by biological agents such as bacteria can be used in various industrial processes. For example, it can be used in environmental processes such as bioremediation and elimination of environmental pollutants, and acts as synergistic agents and distributor pesticides on waxy leaves in agriculture. In this study, biosurfactant which is produced by Pseudomonas aeruginosa (isolated from petroleum sludge) at the intervals of 24, 48, 72 and 96?h, along with chemical surfactant Tween 80 and the biological control agent, Bacillus thuringiensis, in a pilot project for controlling one important cabbage pest (Pieris brassicae), their synergistic properties were evaluated. Statistical analysis of the results showed that B. thuringiensis in combination with biosurfactant produced at different times and B. thuringiensis in combination with chemical surfactant Tween 80 when compared with control treatments like B. thuringiensis alone and B. thuringiensis plus tween 80 as positive controls and distilled water as negative control have significant differences (p?<?0.05). This research showed that surfactant treatment produced at the intervals of 24 and 48?h in combination with B. thuringiensis has the greatest synergistic effect when compared to chemical surfactant treatment. This study concluded that biosurfactant can be used as a distributor and synergistic agent against plant pests and in addition to this, their biological roles in bioremediation can be used as a viable alternative to non-economical chemical surfactants that annually enter millions of tonnes of harmful chemical substances into the fields and underground water.     

Anti-fungal properties of 43 plant species against Alternaria solani and Botrytis cinerea

Investigation of plants containing natural anti-microbial metabolites for plant protection has been identified as a desirable method of disease control. Crude methanolic extracts of 43 plant species belonging to 27 families, which most of them are medicinal plants, mostly collected from the west of Iran were screened for anti-fungal activity against two economically important phytopathogenic fungi, Alternaria solani and Botrytis cinerea during 2010–2012. Bioassay of the extracts was conducted by agar diffusion method on agar plate cultures with five replications. Among all the 43 plant methanolic extracts, mycelia growth of A. solani and B. cinerea was reduced by 28 (65%) and 30 (70%) plant extracts when compared to the control, respectively. The strongest extracts with more than 50% inhibition against A. solani were Elaeagnus angustifolia, Dodonaea viscosa, Haplophyllum perforatum and inflorescence of Allium hirtifolium, respectively. Leaves of A. hirtifolium, H. perforatum, inflorescence of A. hirtifolium and D. viscosa showed highest inhibitory effect (≥50%) against B. cinerea. Moreover, complete inhibition of leaves of A. hirtifolium against B. cinerea was due to their fungistatic activity. The results of this experiment and high number of plants with anti-fungal activity showed that the flora in the west of Iran could be regarded as a rich source of plants with anti-fungal activity. Therefore, further screening of other plant species, identifying active fractions or metabolites and in vivo application of active extracts are warranted.