Search

Hydrogen sulfide (H(2)S) is widely recognized as the third endogenous gas signaling molecule and may play a key role in cancer biological processes. ADT-OH (5-(4-hydroxyphenyl)−3H-1,2-dithiocyclopentene-3-thione) is one of the most widely used organic donors for the slow release of H(2)S and considered to be a potential anticancer compound. In this study, we investigated the antimetastatic effects of ADT-OH in highly metastatic melanoma cells. A tail-vein-metastasis model was established by injecting B16F10 and A375 cells into the tail veins of mice, whereas a mouse footpad-injection model was established by injecting B16F10 cells into mouse footpads. We showed that administration of ADT-OH significantly inhibited the migration and invasion of melanoma cells in the three different animal models. We further showed that ADT-OH dose-dependently inhibited the migration and invasion of B16F10, B16F1 and A375 melanoma cells as evaluated by wound healing and Transwell assays in vitro. LC-MS/MS and bioinformatics analyses revealed that ADT-OH treatment inhibited the EMT process in B16F10 and A375 cells by reducing the expression of FAK and the downstream response protein Paxillin. Overexpression of FAK reversed the inhibitory effects of ADT-OH on melanoma cell migration. Moreover, after ADT-OH treatment, melanoma cells showed abnormal expression of the H(2)S-producing enzymes CSE/CBS and the AKT signaling pathways. In addition, ADT-OH significantly suppressed the proliferation of melanoma cells. Collectively, these results demonstrate that ADT-OH inhibits the EMT process in melanoma cells by suppressing the CSE/CBS and FAK signaling pathways, thereby exerting its antimetastatic activity. ADT-OH may be used as an antimetastatic agent in the future.

Background: Melanoma is a highly aggressive cancer, and its high metastasis results in a high lethality. Hydrogen sulfide (H2S) is now widely recognized as the third endogenous gas delivery substance and may play a key role in cancer biological processes. The present study was designed to evaluate the anti-metastatic effect of H2S-donor ADT-OH on melanoma cells and the underlying mechanism.Methods: Firstly, the effect of ADT-OH on the migration of melanoma cells was explored in vivo by the mouse footpad injection model and the mouse tail vein metastasis model. Tumor xenograft growth and tumor tissue H&E analyses were also measured in vivo. Then, the migration inhibitory effect and the underlying mechanism of ADT-OH on B16F10, B16F1 and A375 melanoma cell lines were evaluated by wound healing, transwell, western blot and immunofluorescence analyses. Results: Our data showed that ADT-OH inhibited the migration and invasion of melanoma cells significantly in vivo in three different animal models. Further research showed that ADT-OH significantly suppressed the migratory, invasive and adhesive properties of A375 and B16F10 cells as measured by the wound-healing and transwell assays. Mechanistically, ADT-OH treatment suppressed the EMT processes and reduced the enzymatic activity of FAK and Paxillin. Moreover, abnormal CSE/CBS and AKT signaling pathways in A375 and B16F10 cells were notably observed following ADT-OH treatment. Additionally, ADT-OH at higher concentration significantly inhibited the proliferation of highly metastatic melanoma A375 and B16F10 cells.Conclusions: Our results suggest that ADT-OH exerts anti-metastasis activity in melanoma cells by suppressing the EMT process through the CSE/CBS and FAK signaling pathways, and it might be used as an agent against metastatic melanoma in future.

Lipid accumulation of Schizochytrium sp. can be induced by stress condition, but this stress-induction usually reduce cell growth and cause oxidative damage, which can eventually lower the lipid yield. Here, adaptive laboratory evolution (ALE) combined high salinity was performed to enhance the antioxidant system and lipid accumulation. The final strain ALE150, which was obtained after 150 days, showed a maximal cell dry weight (CDW) of 134.5 g/L and lipid yield of 80.14 g/L, representing a 32.7 and 53.31% increase over the starting strain, respectively. Moreover, ALE150 exhibited an overall higher total antioxidant capacity (T-AOC) and lower reactive oxygen species (ROS) levels than the starting strain. Furthermore, the regulatory mechanisms responsible for the improved performance of ALE150 were analyzed by transcriptomic analysis. Genes related to the antioxidant enzymes and central carbon metabolism were up-regulation. Moreover, the metabolic fluxes towards the fatty acid synthase (FAS) and polyketide synthase (PKS) pathways were also changed.

Background Schizochytrium sp. is a marine microalga with great potential as a promising sustainable source of lipids rich in docosahexaenoic acid (DHA). This organism’s lipid accumulation machinery can be induced by various stress conditions, but this stress induction usually comes at the expense of lower biomass in industrial fermentations. Moreover, oxidative damage induced by various environmental stresses can result in the peroxidation of lipids, and especially polyunsaturated fatty acids, which causes unstable DHA production, but is often ignored in fermentation processes. Therefore, it is urgent to develop new production strains that not only have a high DHA production capacity, but also possess strong antioxidant defenses. Results Adaptive laboratory evolution (ALE) is an effective method for the development of beneficial phenotypes in industrial microorganisms. Here, a novel cooperative two-factor ALE strategy based on concomitant low temperature and high salinity was applied to improve the production capacity of Schizochytrium sp. Low-temperature conditions were used to improve the DHA content, and high salinity was applied to stimulate lipid accumulation and enhance the antioxidative defense systems of Schizochytrium sp. After 30 adaptation cycles, a maximal cell dry weight of 126.4 g/L and DHA yield of 38.12 g/L were obtained in the endpoint strain ALE-TF30, which was 27.42 and 57.52% higher than parental strain, respectively. Moreover, the fact that ALE-TF30 had the lowest concentrations of reactive oxygen species and malondialdehyde among all strains indicated that lipid peroxidation was greatly suppressed by the evolutionary process. Accordingly, the ALE-TF30 strain exhibited an overall increase of gene expression levels of antioxidant enzymes and polyketide synthases compared to the parental strain. Conclusion This study provides important clues on how to overcome the negative effects of lipid peroxidation on DHA production in Schizochytrium sp. Taken together, the cooperative two-factor ALE process can not only increase the accumulation of lipids rich in DHA, but also prevent the loss of produced lipid caused by lipid peroxidation. The strategy proposed here may provide a new and alternative direction for the industrial cultivation of oil-producing microalgae.

Temperature is an important factor that not only affects cell growth, but also changes fatty acid composition in oleaginous microorganisms. Here, adaptive laboratory evolution (ALE) was performed in Schizochytrium sp. to enhance strain thermotolerance in docosahexaeonic acid (DHA) production. The adaptive strain showed higher growth rate and lower temperature sensitivity. Cell growth rate of the adaptive strain after 70 cycles at high temperature stimulation (ALE70) at 28 °C and 34 °C were 46.93% and 60.55% higher than that of original strain. Meanwhile, the reduction ratio of DHA yield from 28 °C to 34 °C of ALE70 decreased by 61.71%. Finally, lipid and DHA concentration at 34 °C reached 4.31 and 4.33 times of the starting strain. Further physiological analysis showed ALE70 generated less ROS, synthesized less HSP proteins, and produced more pigments, which might be related to the improved performance. This work increased the fermentation temperature by 5 degrees without affecting DHA production, which significantly reduced the energy consumption cost of the industrialization.

Background Schizochytrium sp. is a marine microalga with great potential as a promising sustainable source of lipids rich in docosahexaenoic acid (DHA). This organism’s lipid accumulation machinery can be induced by various stress conditions, but this stress induction usually comes at the expense of lower biomass in industrial fermentations. Moreover, oxidative damage induced by various environmental stresses can result in the peroxidation of lipids, and especially polyunsaturated fatty acids, which causes unstable DHA production, but is often ignored in fermentation processes. Therefore, it is urgent to develop new production strains that not only have a high DHA production capacity, but also possess strong antioxidant defenses. Results Adaptive laboratory evolution (ALE) is an effective method for the development of beneficial phenotypes in industrial microorganisms. Here, a novel cooperative two-factor ALE strategy based on concomitant low temperature and high salinity was applied to improve the production capacity of Schizochytrium sp. Low-temperature conditions were used to improve the DHA content, and high salinity was applied to stimulate lipid accumulation and enhance the antioxidative defense systems of Schizochytrium sp. After 30 adaptation cycles, a maximal cell dry weight of 126.4 g/L and DHA yield of 38.12 g/L were obtained in the endpoint strain ALE-TF30, which was 27.42 and 57.52% higher than parental strain, respectively. Moreover, the fact that ALE-TF30 had the lowest concentrations of reactive oxygen species and malondialdehyde among all strains indicated that lipid peroxidation was greatly suppressed by the evolutionary process. Accordingly, the ALE-TF30 strain exhibited an overall increase of gene expression levels of antioxidant enzymes and polyketide synthases compared to the parental strain. Conclusion This study provides important clues on how to overcome the negative effects of lipid peroxidation on DHA production in Schizochytrium sp. Taken together, the cooperative two-factor ALE process can not only increase the accumulation of lipids rich in DHA, but also prevent the loss of produced lipid caused by lipid peroxidation. The strategy proposed here may provide a new and alternative direction for the industrial cultivation of oil-producing microalgae. Electronic supplementary material The online version of this article (10.1186/s13068-018-1065-4) contains supplementary material, which is available to authorized users.