A pair of transporters controls mitochondrial Zn2+ levels to maintain mitochondrial homeostasis

Zn2+ is required for the activity of many mitochondrial proteins, which regulate mitochondrial dynamics, apoptosis and mitophagy. However, it is not understood how the proper mitochondrial Zn2+ level is achieved to maintain mitochondrial homeostasis. Using Caenorhabditis elegans, we reveal here that a pair of mitochondrion-localized transporters controls the mitochondrial level of Zn2+. We demonstrate that SLC-30A9/ZnT9 is a mitochondrial Zn2+ exporter. Loss of SLC-30A9 leads to mitochondrial Zn2+ accumulation, which damages mitochondria, impairs animal development and shortens the life span. We further identify SLC-25A25/SCaMC-2 as an important regulator of mitochondrial Zn2+ import. Loss of SLC-25A25 suppresses the abnormal mitochondrial Zn2+ accumulation and defective mitochondrial structure and functions caused by loss of SLC-30A9. Moreover, we reveal that the endoplasmic reticulum contains the Zn2+ pool from which mitochondrial Zn2+ is imported. These findings establish the molecular basis for controlling the correct mitochondrial Zn2+ levels for normal mitochondrial structure and functions.

Ameliorative effect of Argyreia boseana Sant. & Pat. on stress in C. elegans

Background: Pathyashadangam kwath, a classical ayurvedic polyherbal formulation is used for thetreatment of cluster head ache, migraine, upper respiratory diseases, ear ache and night blindness. Review of literature suggested that characterization parameters of Pathyashadangam kwath are notreported.Objective: To report characteristic parameters of Pathyashadangam kwath to confirm quality and purity.Materials and methods: The fruit pericarps of Haritaki, Bibhitaki and Amalaki, aerial parts of Bhunimba,rhizome of Haridra, stem bark of Nimba and stem of Guduchi were the ingredients of Pathyashadangamkwath. Three batches of the kwath were prepared as per standard procedures. The kwath was evaluatedfor organoleptic, physical, phytochemical and chromatographic parameters as per standard methods.Results: HPTLC analysis revealed that Toluene: Ethyl Acetate: Formic acid (2.5: 2.0: 0.5) was a suitablemobile phase for characterization of the kwath. HPLC analysis revealed that andrographolide was asuitable marker for standardization of the kwath.Conclusion: The characterization parameters presented in this paper may serve as standard reference forquality control analysis of Pathyashadangam kwath.

Rapid detection of sepsis using cesda: the caenorhabditis elegans sepsis detection assay

Abstract INTRODUCTION The nematode Caenorhabditis elegans was used as a biological sensor to detect the urine of sepsis patients (CESDA assay). METHODS C. elegans was aliquoted onto the center of assay plates and allowed to migrate towards sepsis (T) or control (C) urine samples spotted on the same plate. The number of worms found in either (T) or (C) was scored at 10-minute intervals over a 60-minute period. RESULTS The worms were able to identify the urine (<48 hours) of sepsis patients rapidly within 20 minutes (AUROC=0.67, p=0.012) and infection within 40 minutes (AUROC=0.80, p=0.016). CONCLUSIONS CESDA could be further explored for sepsis diagnosis.

Physicochemical characterization and evaluation of in vitro and in vivo toxicity of goldenberry extract nanoemulsion / Caracterização físico-química e avaliação da toxicidade in vitro e in vivo de nanoemulsão contendo extrato de goldenberry

ABSTRACT: Oil-in-water (O/W) nanoemulsion containing goldenberry extract was elaborated using a high-energy ultrasonic bath method. Physicochemical characterization of the formulation was carried out by determining pH, mean droplet diameter, polydispersity index (PDI) and zeta potential. Nanoemulsion toxicity was assessed using in vitro assays with tumor and non-tumor cell lines, and in vivo using Caenorhabditis elegans. The pH of the nanoemulsion was 3.84, the mean droplet diameter was 268 ± 7 nm, PDI 0.113 and zeta potential -13.94 mV. Results of the cytotoxicity assays employing non-tumor cells indicated that the extract associated or not with nanoemulsion maintained cell viability at different concentrations tested. In the assays using tumor lineage, it is observed that the nanoemulsion containing the extract had higher antitumor activity than the free extract. As for the in vivo tests, there was no change in the survival rate of the worms.

RESUMO: Nanoemulsão óleo/água (O/A) contendo extrato de goldenberry foi elaborada utilizando método de banho ultrassônico de alta energia. A caracterização físico-química da formulação foi realizada pela determinação do pH, diâmetro médio de gotas, índice de polidispersão (PDI) e potencial zeta. A toxicidade das nanoemulsões foi avaliada utilizando ensaios in vitro com linhas celulares tumorais e não tumorais e in vivo utilizando Caenorhabditis elegans. O pH da nanoemulsão foi de 3,84, o diâmetro médio das gotículas foi de 268 ± 7 nm, PDI 0,113 e o potencial zeta -13,94 mV. Os resultados dos ensaios de citotoxicidade empregando células não tumorais indicaram que o extrato associado ou não à nanoemulsão manteve a viabilidade celular em diferentes concentrações testadas. Nos ensaios, utilizando linhagem tumoral, observou-se que a nanoemulsão contendo o extrato apresentou maior atividade antitumoral do que o extrato livre. Quanto aos testes in vivo, não houve mudança na taxa de sobrevivência dos vermes.

Caracterización fenotípica de la cepa N2 de Caenorhabditis elegans como un modelo en enfermedades neurodegenerativas / Phenotypic characterization of the N2 strain of Caenorhabditis elegans as a model in neurodegenerative diseases

Resumen El nematodo C. elegans se estableció desde I960, gracias al biólogo sudafricano Sydney Brenner, como un organismo modelo en investigación. Sus cualidades biológicas permiten mejorar la visión y comprensión de patologías en los seres humanos y otros seres pluricelulares; además, sus fenotipos claros y observables lo convierten en un organismo adecuado para el estudio básico de enfermedades neurodegenerativas, inmunológicas y procesos cancerígenos. Objetivo. Analizar las características fenotípicas de la cepa silvestre N2 de C. elegans para su posterior uso como modelo de tamizaje en el laboratorio de Biotecnología y Genética (Universidad Colegio Mayor de Cundinamarca). Materiales y Métodos. El nematodo fue cultivado y crecido en el medio NGM con la cepa E. coli OP50. La cepa N2 fue sincronizada para obtener huevos y posteriormente larvas L1. Se estandarizaron los ensayos de longevidad, reproducción, longitud y estrés térmico. Resultados. La caracterización fenotípica de la cepa N2 de C. elegans presentó: una longevidad de 16 a 22 días, una reproducción promedio de 225 crías, la longitud del nematodo fue de 1100±50 μm y la supervivencia bajo estrés térmico evaluada en las dos etapas de desarrollo del nematodo es muy reducida a 37°C en comparación de 35°C; además, los nematodos fueron más resistentes al primer día de adulto joven en comparación con el sexto día de adulto. Conclusiones. Los resultados aportados por este estudio permiten sugerir que las características fenotípicas del nematodo analizadas se encuentran dentro de lo reportado en la literatura, por lo cual es viable usarlo como como modelo biológico en diferentes ensayos tal como lo reportan otros estudios.

Abstract The nematode C. elegans was established since I960, thanks to the South African biologist Sydney Brenner, as a model organism in research. Their biological qualities allow to improve the vision and understanding of pathologies in human and other multicellular beings; In addition, its clear and observable phenotypes make it a suitable organism for the basic study of neurodegenerative, immunological diseases and carcinogenic processes. Objective. To analyze the phenotypic characteristics of the C. elegans N2 wild strain for later use as a screening model in the Biotechnology and Genetics Laboratory (Colegio Mayor de Cundinamarca University). Materials and Methods. The nematode was grown and grown in the NGM medium with the strain E. coli OP50. The N2 strain was synchronized to obtain eggs and later L1 larvae. The tests of longevity, reproduction, length and thermal stress were standardized. Results. The phenotypic characterization of the N2 strain of C. elegans presented a longevity of 16 to 22 days, an average reproduction of 225 offspring, the length of the nematode was 1100 ± 50 μm and the survival under thermal stress evaluated in the two Development stages of the nematode is greatly reduced at 37 ° C compared to 35 ° C; In addition, nematodes were more resistant to the first day of young adult compared to the sixth day of adulthood. Conclusions. The results of this study suggest that the phenotypic characteristics of the nematode analyzed are within the literature, so it is feasible to use it as a biological model in different trials as reported in other studies.

Selenocysteine modulates resistance to environmental stress and confers anti-aging effects in C. elegans

OBJECTIVE: The free radical theory of aging suggests that cellular oxidative damage caused by free radicals is a leading cause of aging. In the present study, we examined the effects of a well-known anti-oxidant amino acid derivative, selenocysteine, in response to environmental stress and aging using Caenorhabditis elegans as a model system. METHOD: The response to oxidative stress induced by H2O2 or ultraviolet irradiation was compared between the untreated control and selenocysteine-treated groups. The effect of selenocysteine on lifespan and fertility was then determined. To examine the effect of selenocysteine on muscle aging, we monitored the change in motility with aging in both the untreated control and selenocysteine-treated groups. RESULTS: Dietary supplementation with selenocysteine significantly increased resistance to oxidative stress. Survival after ultraviolet irradiation was also increased by supplementation with selenocysteine. Treatment with selenocysteine confers a longevity phenotype without an accompanying reduction in fertility, which is frequently observed in lifespan-extending interventions as a trade-off in C. elegans. In addition, the age-related decline in motility was significantly delayed by supplementation of selenocysteine. CONCLUSION: These findings suggest that dietary supplementation of selenocysteine can modulate response to stressors and lead to lifespan extension, thus supporting the free radical theory of aging.

Supplementation of S-allyl cysteine improves health span in Caenorhabditis elegans / A suplementação de S-alil cisteína melhora a duração da saúde em Caenorhabditis elegans

Previous studies show that nutritional interventions with anti-oxidants have various healthpromoting effects in several model organisms. Here, we examine the effects of S-allyl cysteine on resistance to environmental stressors and age-related physiological changes using C. elegans as a model system. S-allyl cysteine is a modified amino acid found in aged garlic extracts and known to have strong anti-oxidant activity. The survival of worms under oxidative-stress conditions significantly increased with supplementation of S-allyl cysteine. In addition, pretreatment of S-allyl cysteine significantly increased resistance to both heat stress and ultraviolet irradiation. However, lifespan was not affected by S-allyl cysteine treatment. We also examined the effect of S-allyl cysteine on motility of C. elegans and found that S-allyl cysteine can retard the age-related decline of muscle tissue locomotive activity. S-allyl cysteine also significantly suppressed amyloid -induced paralysis in Alzheimer's disease model animals. Taken together, our study indicates that dietary supplementation of S-allyl cysteine can improve health span and suggests that S-allyl cysteine can be used to develop novel health-promoting pharmaceuticals.

Estudos anteriores mostram que intervenções nutricionais com antioxidantes têm vários efeitos promotores da saúde em vários organismos-modelo. Aqui, examinamos os efeitos da S-alil cisteína sobre a resistência a estressores ambientais e alterações fisiológicas relacionadas com a idade usando C. elegans como um sistema modelo. Salil cisteína é um aminoácido modificado encontrado em extratos de alho envelhecido e conhecido por ter forte atividade antioxidante. A sobrevivência de vermes sob condições de estresse oxidativo aumentou significativamente com a suplementação de S-alil cisteína. Além disso, o pré-tratamento com S-alil cisteína aumentou significativamente a resistência tanto ao estresse térmico como à irradiação ultravioleta. No entanto, o tempo de vida não foi afetado pelo tratamento com S-alil cisteína. Nós também examinamos o efeito da S-alil cisteína na motilidade de C. elegans e descobrimos que a S-alil cisteína pode retardar o declínio relacionado à idade da atividade locomotora do tecido muscular. A S-alil cisteína também suprimiu significativamente a paralisia induzida por amilóide em animais-modelo da doença de Alzheimer. Tomados em conjunto, o nosso estudo indica que a suplementação dietética de S-alil cisteína pode melhorar a duração da saúde e sugere que S-alil cisteína pode ser usada para desenvolver novos produtos farmacêuticos de promoção da saúde.

Cell-to-cell Transmission of Polyglutamine Aggregates in C. elegans

Huntington disease (HD) is an inherited neurodegenerative disorder characterized by motor and cognitive dysfunction caused by expansion of polyglutamine (polyQ) repeat in exon 1 of huntingtin (HTT). In patients, the number of glutamine residues in polyQ tracts are over 35, and it is correlated with age of onset, severity, and disease progression. Expansion of polyQ increases the propensity for HTT protein aggregation, process known to be implicated in neurodegeneration. These pathological aggregates can be transmitted from neuron to another neuron, and this process may explain the pathological spreading of polyQ aggregates. Here, we developed an in vivo model for studying transmission of polyQ aggregates in a highly quantitative manner in real time. HTT exon 1 with expanded polyQ was fused with either N-terminal or C-terminal fragments of Venus fluorescence protein and expressed in pharyngeal muscles and associated neurons, respectively, of C. elegans. Transmission of polyQ proteins was detected using bimolecular fluorescence complementation (BiFC). Mutant polyQ (Q97) was transmitted much more efficiently than wild type polyQ (Q25) and forms numerous inclusion bodies as well. The transmission of Q97 was gradually increased with aging of animal. The animals with polyQ transmission exhibited degenerative phenotypes, such as nerve degeneration, impaired pharyngeal pumping behavior, and reduced life span. The C. elegans model presented here would be a useful in vivo model system for the study of polyQ aggregate propagation and might be applied to the screening of genetic and chemical modifiers of the propagation.

Riboflavin transporter-2 (rft-2) of Caenorhabditis elegans: Adaptive and developmental regulation.

Riboflavin transporters (rft-1 and rft-2), orthologous to human riboflavin transporter-3 (hRVFT-3), are identified and characterized in Caenorhabditis elegans. However, studies pertaining to functional contribution of rft-2 in maintaining body homeostatic riboflavin levels and its regulation are very limited. In this study, the expression pattern of rft-2 at different life stages of C. elegans was studied through real-time PCR, and found to be consistent from larval to adult stages that demonstrate its involvement in maintaining the body homeostatic riboflavin levels at whole animal level all through its life. A possible regulation of rft-2 expression at mRNA levels at whole animal was studied after adaptation to low and high concentrations of riboflavin. Abundance of rft-2 transcript was upregulated in riboflavin-deficient conditions (10 nM), while it was downregulated with riboflavin-supplemented conditions (2 mM) as compared with control (10 μM). Further, the 5 -regulatory region of the rft-2 gene was cloned, and transgenic nematodes expressing transcriptional rft-2 promoter::GFP fusion constructs were generated. The expression of rft-2 was found to be adaptively regulated in vivo when transgenic worms were maintained under different extracellular riboflavin levels, which was also mediated partly via changes in the rft-2 levels that directs towards the possible involvement of transcriptional regulatory events.

Lifespan extension and increased resistance to environmental stressors by N-Acetyl-L-Cysteine in Caenorhabditis elegans

OBJECTIVE: This study was performed to determine the effect of N-acetyl-L-cysteine, a modified sulfur-containing amino acid that acts as a strong cellular antioxidant, on the response to environmental stressors and on aging in C. elegans. METHOD: The survival of worms under oxidative stress conditions induced by paraquat was evaluated with and without in vivo N-acetyl-L-cysteine treatment. The effect of N-acetyl-L-cysteine on the response to other environmental stressors, including heat stress and ultraviolet irradiation (UV), was also monitored. To investigate the effect on aging, we examined changes in lifespan, fertility, and expression of age-related biomarkers in C. elegans after N-acetyl-L-cysteine treatment. RESULTS: Dietary N-acetyl-L-cysteine supplementation significantly increased resistance to oxidative stress, heat stress, and UV irradiation in C. elegans. In addition, N-acetyl-L-cysteine supplementation significantly extended both the mean and maximum lifespan of C. elegans. The mean lifespan was extended by up to 30.5% with 5 mM N-acetyl-L-cysteine treatment, and the maximum lifespan was increased by 8 days. N-acetyl-L-cysteine supplementation also increased the total number of progeny produced and extended the gravid period of C. elegans. The green fluorescent protein reporter assay revealed that expression of the stress-responsive genes, sod-3 and hsp-16.2, increased significantly following N-acetyl-L-cysteine treatment. CONCLUSION: N-acetyl-L-cysteine supplementation confers a longevity phenotype in C. elegans, possibly through increased resistance to environmental stressors. .

Effect of ginseng polysaccharides on polyQ accumulation toxicity and lifes pan of Caenorhabditis elegans / 中国生化药物杂志

Objective To study the ginseng polysaccharides for prolonging the life span of the C.elegans and inhibit the toxic effects of polyQ accumulation.Methods Caenorhabditis elegans of HA759 and AM141 were divided into control group and Ginseng group, seprately.Control group didn’ t do any special treatment, Ginseng group were given 10 mg/mL polysaccharide and OP50-1 in the proportion of 1:4 mixed volume to 50 mL.C.elegans of glutamine ( polyQ) polymer HA759 neurotoxicity model test of glutamine protein polymer toxicity experiment were done.The ASH neuron survival condition were tested.After sampling statistics gathered nematodes in the whole fluorescent points every day, study ginseng polysaccharide on polymers glutamine aggregation inhibition.Finally the solid life of two C.elegans were studied.Results The survival rate of ASH neurons in Caenorhabditis elegans HA759 of control group after 3 days culture was 53%.which of Ginseng group was 64% (P<0.05).The fluorescence of 48~96h aggregation points in Caenorhabditis elegans AM141 of control group were(6 ±1), (27 ±2), (56 ±4), which of Ginseng group were (4 ±1) in 48 h, (20 ±3) in 72 h and (45 ±2) in 96 h, the differences between two groups were all significant(P <0.05).The average survival time of Caenorhabditis elegans AM141 of control group was (23 ±2)days, which of Ginseng group was (27 ±2)days;average survival time of Caenorhabditis elegans HA759 of control group was (24 ±2)days, which of Ginseng group was (27 ±2)days,the difterences were all signiyicant(P<0.05).Conclusion Ginseng polysaccharides can not only prolong the lifespan of the C.elegans, but also can restrain polyQ gathered and ease the polyQ neurotoxicity associated with aging.

Influence of clove oil on certain quorum-sensing-regulated functions and biofilm of Pseudomonas aeruginosa and Aeromonas hydrophila.

Quorum sensing (QS) plays an important role in virulence, biofilm formation and survival of many pathogenic bacteria including Pseudomonas aeruginosa. This signalling pathway is considered as novel and promising target for anti-infective agents. In the present investigation, effect of the Sub-MICs of clove oil on QS regulated virulence factors and biofilm formation was evaluated against P. aeruginosa PAO1 and Aeromonas hydrophila WAF-38 strain. Sub-inhibitory concentrations of the clove oil demonstrated statistically significant reduction of las- and rhl-regulated virulence factors such as LasB, total protease, chitinase and pyocyanin production, swimming motility and exopolysaccharide production. The biofilm forming capability of PAO1 and A. hydrophila WAF-38 was also reduced in a concentration-dependent manner at all tested sub-MIC values. Further, the PAO1-preinfected Caenorhabditis elegans displayed an enhanced survival when treated with 1.6% v/v of clove oil. The above findings highlight the promising anti-QS-dependent therapeutic function of clove oil against P. aeruginosa.

Effect of the tocotrienol-rich fraction on the lifespan and oxidative biomarkers in Caenorhabditis elegans under oxidative stress

OBJECTIVE: This study was performed to determine the effect of the tocotrienol-rich fraction on the lifespan and oxidative status of C. elegans under oxidative stress. METHOD: Lifespan was determined by counting the number of surviving nematodes daily under a dissecting microscope after treatment with hydrogen peroxide and the tocotrienol-rich fraction. The evaluated oxidative markers included lipofuscin, which was measured using a fluorescent microscope, and protein carbonyl and 8-hydroxy-2′-deoxyguanosine, which were measured using commercially available kits. RESULTS: Hydrogen peroxide-induced oxidative stress significantly decreased the mean lifespan of C. elegans, which was restored to that of the control by the tocotrienol-rich fraction when administered before or both before and after the hydrogen peroxide. The accumulation of the age marker lipofuscin, which increased with hydrogen peroxide exposure, was decreased with upon treatment with the tocotrienol-rich fraction (p<0.05). The level of 8-hydroxy-2′-deoxyguanosine significantly increased in the hydrogen peroxide-induced group relative to the control. Treatment with the tocotrienol-rich fraction before or after hydrogen peroxide induction also increased the level of 8-hydroxy-2′-deoxyguanosine relative to the control. However, neither hydrogen peroxide nor the tocotrienol-rich fraction treatment affected the protein carbonyl content of the nematodes. CONCLUSION: The tocotrienol-rich fraction restored the lifespan of oxidative stress-induced C. elegans and reduced the accumulation of lipofuscin but did not affect protein damage. In addition, DNA oxidation was increased. .

HSP12.1, A Small Heat Shock Protein in C.elegans, Has Chaperone-like Activity / 生物化学与生物物理进展

Many kinds of small heat shock proteins (sHSPs) are able to prevent protein aggregation in stress, which show the ATP independent chaperone-like activity. The smallest protein HSP12.1 in sHSP family of the nematode Caenorhabditis elegans exhibits chaperone-like activities in vitro. It prevents protein aggregation in a certain extent when use insulin, ADH and lysozyme as the substrates, though it is not as efficient as the typical chaperones (such as HSP16.1 in C. elegans). By contrast, the other three sHSP12s (HSP12.2, HSP12.3 and HSP12.6), which have similar molecular masses and primary structure, appear devoid of in vitro chaperone-like activities. In addition, overexpressing HSP12.1 enhances cell thermotolerance of Escherichia coli. The survival rate of the HSP12.1 overexpressed cells is 4-fold higher than the control, yet whether it does the same function in C. elegans is still unknown. Results indicate that C-terminal region is not necessary for the chaperone-like activity of sHSPs, for HSP12.1 terminates a short C-terminal tail. N-terminal domain may play a relatively important role in the exhibition of chaperone-like activities, while ?-crystalline domain may also involve in this function.