In-vitro antioxidant and anti-inflammatory potential along with p.o. pharmacokinetic profile of key bioactive phytocompounds of Snow Mountain Garlic: a comparative analysis vis-à-vis normal garlic.

Snow mountain garlic (SMG) is a trans-Himalayan medicinal plant used in the traditional medicine system for several ailments, including inflammatory arthritis. Research studies are insufficient to validate its folk medicinal applications. In the present study, the comparative abundance of its key bioactive phytocompounds, viz., S-allyl-L-cysteine (SAC), alliin, and S-methyl-L-cysteine (SMC) against normal garlic were assessed using the LC-MS/MS-MRM method. In addition, the study also explored the antioxidant and anti-inflammatory potency of crude extract of SMG and purified signature phytocompounds (i.e., SMC, SAC, and alliin) in comparison with normal garlic and dexamethasone in LPS-stimulated RAW264.7 macrophage cells. The LC-MS/MS-MRM study revealed significant differences among SMG and normal garlic, viz., alliin 22.8-fold higher in SMG, and SMC could be detected only in SMG. In the bioassays, SMG extract and purified signature phytocompounds significantly downregulated oxidative damage in activated macrophages, boosting endogenous antioxidants' activity. SMG extract-treated macrophages significantly suppressed NF-κB expression and related inflammatory indicators such as cytokines, COX-2, iNOS, and NO. Notably, the observed anti-inflammatory and antioxidant bioactivities of SMG extract were comparable to signature phytocompounds and dexamethasone. In addition, SAC being uniformly found in SMG and normal garlic, its comparative pharmacokinetics was studied to validate the pharmacodynamic superiority of SMG over normal garlic. Significantly higher plasma concentrations (Cmax), half-life (t1/2), and area under curve (AUC) of SAC following SMG extract administration than normal garlic validated the proposed hypothesis. Thus, the abundance of bioactive phytocompounds and their better pharmacokinetics in SMG extract might be underlying its medicinal merits over normal garlic.

Ligustilide covalently binds to Cys703 in the pre-S1 helix of TRPA1, blocking the opening of channel and relieving pain in rats with acute soft tissue injury.

ETHNOPHARMACOLOGICAL RELEVANCE: The natural anodyne Ligustilide (Lig), derived from Angelica sinensis (Oliv.) Diels and Ligusticum chuanxiong Hort., has been traditionally employed for its analgesic properties in the treatment of dysmenorrhea and migraine, and rheumatoid arthritis pain. Despite the existing reports on the correlation between TRP channels and the analgesic effects of Lig, a comprehensive understanding of their underlying mechanisms of action remains elusive. AIM OF THE STUDY: The objective of this study is to elucidate the mechanism of action of Lig on the analgesic target TRPA1 channel. METHODS: The therapeutic effect of Lig was evaluated in a rat acute soft tissue injury model. The analgesic target was identified through competitive inhibition of TRP channel agonists at the animal level, followed by Fluo-4/Ca2+ imaging on live cells overexpressing TRP proteins. The potential target was verified through in-gel imaging, colocalization using a Lig-derived molecular probe, and a drug affinity response target stability assay. The binding site of Lig was identified through protein spectrometry and further analyzed using molecular docking, site-specific mutation, and multidisciplinary approaches. RESULTS: The administration of Lig effectively ameliorated pain and attenuated oxidative stress and inflammatory responses in rats with soft tissue injuries. Moreover, the analgesic effects of Lig were specifically attributed to TRPA1. Mechanistic studies have revealed that Lig directly activates TRPA1 by interacting with the linker domain in the pre-S1 region of TRPA1. Through metabolic transformation, 6,7-epoxyligustilide (EM-Lig) forms a covalent bond with Cys703 of TRPA1 at high concentrations and prolonged exposure time. This irreversible binding prevents endogenous electrophilic products from entering the cysteine active center of ligand-binding pocket of TRPA1, thereby inhibiting Ca2+ influx through the channel opening and ultimately relieving pain. CONCLUSIONS: Lig selectively modulates the TRPA1 channel in a bimodal manner via non-electrophilic/electrophilic metabolic conversion. The epoxidized metabolic intermediate EM-Lig exerts analgesic effects by irreversibly inhibiting the activation of TRPA1 on sensory neurons. These findings not only highlight the analgesic mechanism of Lig but also offer a novel nucleophilic attack site for the development of TRPA1 antagonists in the pre-S1 region.

Different methionine to cysteine supplementation ratios altered bone quality of broilers with or without Eimeria challenge assessed by dual energy X-ray absorptiometry and microtomography.

Despite the acknowledged significance of nutrition in bone development, effects of methionine (Met) and cysteine (Cys) on bone quality remain under-researched, particularly during Eimeria challenge. We investigated the effects of different supplemental Met to Cys ratios (MCR) on bone quality of broilers under Eimeria challenge. A total of 720 fourteen-day old Cobb500 broilers were allocated into a 5 × 2 factorial arrangement. Five diets with Met and Cys supplemented at MCR of 100:0, 75:25, 50:50, 25:75, and 0:100 were fed to the birds with or without Eimeria challenge. Body composition was measured by dual energy x-ray absorptiometry, and the femur bone characteristics were assessed by microtomography. Data were analyzed by two-way ANOVA and orthogonal polynomial contrast. The results reaffirmed the detrimental effects of Eimeria challenge on bone quality. On 9 d post inoculation (DPI), significant interaction effects were found for whole body bone mineral content (BMC), lean tissue weight, and body weight (P < 0.05); in the nonchallenged group (NCG), these parameters linearly decreased as MCR decreased (P < 0.05). In the challenged group (CG), body weight and lean tissue weight were unaffected by MCR, and BMC linearly increased as MCR decreased (P < 0.05). For the cortical bone of femoral metaphysis on 6 DPI, bone mineral density (BMD) linearly increased as MCR decreased (P < 0.05). Bone volume to tissue volume ratio (BV/TV) in the CG linearly increased as MCR decreased (P < 0.05). On 9 DPI, BMC and TV linearly increased as MCR decreased (P < 0.05) in the NCG. BMD and BV/TV changed quadratically as MCR decreased (P < 0.05). For the trabecular bone of femoral metaphysis on 9 DPI, BV/TV, and trabecular number linearly increased as MCR decreased (P < 0.05) in the NCG. For the femoral diaphysis, BV, TV, BMC on 6 DPI, and BMD on 9 DPI linearly increased as MCR decreased (P < 0.05). In conclusion, this study showed that both Eimeria challenge and varying supplemental MCR could influence bone quality of broilers.

Effects of taurine, cysteine and melatonin as antioxidant supplements to the freezing medium of Prochilodus brevis sperm.

Cryopreservation consist of a set of methods to preserve cells and tissues by drastically reducing the temperature. Among some undesired effects, cryopreservation might generate reactive oxygen species that lead to an increase of oxidative stress, causing damage to cells. This study aimed to test taurine, cysteine, and melatonin on the freezing of Prochilodus brevis sperm and assess its effects on post-thawed sperm quality. Sperm was collected and seven pools were formed (n = 7). They were diluted (1:9) in standard medium (5% glucose, 10% dimethyl sulfoxide and 5% egg yolk) supplemented or not (control) with taurine (0.3, 1.0, 3.16 or 10.0 mM), cysteine (0.3, 1.0, 3.16 or 10.0 mM) or melatonin (0.6, 1.12, 2.0 or 3.56 mM). Post-thawed sperm was evaluated for kinetic (total motility, velocities, and percentage of rapid cells), morphology and membrane and DNA integrity. Differences were found when melatonin was used as an antioxidant. For the variables rapid sperm and sperm velocities, 3.56 mM melatonin presented higher results than the control (melatonin 0 mM). Melatonin 2 mM was similar to 3.56 mM on rapid sperm, average path velocity (VAP) and curvilinear velocity (VCL). No difference was found between concentration 0 mM (control) and taurine treatments. As for cysteine, 0.3 mM presented the best results for rapid sperm than 10 mM, and higher VCL and VAP than 1 mM. Melatonin 3.56 mM presented higher results on kinetic parameters (rapid motility, VCL, VSL and VAP) than other tested antioxidants. Therefore, melatonin 3.56 mM is recommended to be added to the sperm freezing medium of P. brevis.

Cysteine supplementation pre-freeze and post-thaw improves integrity and reduces oxidative stress in cryopreserved ram spermatozoa.

Cryopreserved ram sperm is highly sensitive to oxidative stress by reactive oxygen species (ROS) which impair sperm function and integrity. Antioxidants such as cysteine can mitigate the effect of ROS, although the optimal concentration or timing of supplementation is unknown. This study aimed to determine the effect of concentration and timing of cysteine supplementation on the integrity and function of cryopreserved ram spermatozoa. Nine ejaculates were collected from three Texel rams then cryopreserved and supplemented with cysteine (0, 0.5, or 1.0 mg/mL) added pre-freeze (PF), post-thaw (PT) or pre-freeze and post-thaw (PF + PT) generating seven treatments: 1) control 0 mg/mL, 2) PF 0.5 mg/mL, 3) PF 1 mg/mL, 4) PT 0.5 mg/mL, 5), PT 1.0 mg/mL, 6) PF + PT 0.5 mg/mL and 7) PF + PT 1.0 mg/mL. Sperm motility, viability, acrosome integrity, ROS production and penetrability through artificial cervical mucus were assessed post-thaw. Cysteine supplementation reduced ROS production which thereby improved spermatozoa motility, viability, acrosome integrity and penetrability (p < 0.001) Sperm integrity for all parameters was greatest in spermatozoa treated PF + PT with 1.0 mg/mL cysteine, although treatment pre-freeze or post-thaw also improved integrity beyond the control. This study has identified that 1.0 mg/mL cysteine is most beneficial and has highlighted the importance of preventing oxidative stress in spermatozoa post-thaw. These finding can help to mitigate the detrimental effect of cryopreservation on spermatozoa and aid the development of cryopreservation protocols in sheep.

The nonessential amino acid cysteine is required to prevent ferroptosis in acute myeloid leukemia.

ABSTRACT: Cysteine is a nonessential amino acid required for protein synthesis, the generation of the antioxidant glutathione, and for synthesizing the nonproteinogenic amino acid taurine. Here, we highlight the broad sensitivity of leukemic stem and progenitor cells to cysteine depletion. By CRISPR/CRISPR-associated protein 9-mediated knockout of cystathionine-γ-lyase, the cystathionine-to-cysteine converting enzyme, and by metabolite supplementation studies upstream of cysteine, we functionally prove that cysteine is not synthesized from methionine in acute myeloid leukemia (AML) cells. Therefore, although perhaps nutritionally nonessential, cysteine must be imported for survival of these specific cell types. Depletion of cyst(e)ine increased reactive oxygen species (ROS) levels, and cell death was induced predominantly as a consequence of glutathione deprivation. nicotinamide adenine dinucleotide phosphate hydrogen oxidase inhibition strongly rescued viability after cysteine depletion, highlighting this as an important source of ROS in AML. ROS-induced cell death was mediated via ferroptosis, and inhibition of glutathione peroxidase 4 (GPX4), which functions in reducing lipid peroxides, was also highly toxic. We therefore propose that GPX4 is likely key in mediating the antioxidant activity of glutathione. In line, inhibition of the ROS scavenger thioredoxin reductase with auranofin also impaired cell viability, whereby we find that oxidative phosphorylation-driven AML subtypes, in particular, are highly dependent on thioredoxin-mediated protection against ferroptosis. Although inhibition of the cystine-glutamine antiporter by sulfasalazine was ineffective as a monotherapy, its combination with L-buthionine-sulfoximine (BSO) further improved AML ferroptosis induction. We propose the combination of either sulfasalazine or antioxidant machinery inhibitors along with ROS inducers such as BSO or chemotherapy for further preclinical testing.

L-cysteine improves boar semen motility at 5 ºC but does not affect the oxidative status.

Hypothermic storage has been proposed as a method to reduce bacterial loads and promoting prudent use of antibiotics. Reducing temperature, however, can lead to cold shock damage and oxidative stress in boar semen. This study verified the effect of L-cysteine on the quality of semen stored at 5 °C for 120 h. Twenty-one normospermic ejaculates were diluted in Beltsville Thawing Solution into five treatments: Positive control (Pos_Cont, storage at 17 °C without L-cysteine) and groups with 0, 0.5, 1, and 2 mmol/L of L-cysteine supplementation stored at 5 °C. Variables were analyzed as repeated measures, considering treatment, storage time, and interaction as main factors. The effects of different L-cysteine concentrations were also evaluated using polynomial orthogonal contrasts. Sperm motility and pH were higher in the Pos_Cont compared to the groups stored at 5 °C (P < 0.05). In polynomial orthogonal contrast models, total motility was affected by the interaction between L-cysteine and storage time (P = 0.04), with a linear increase in motility when increasing the amount of L-cysteine at 72 and 120 h. Progressive motility increased quadratically as the L-cysteine reached 1 mmol/L (P < 0.01). In the thermoresistance test at 120 h, sperm motility increased quadratically up to an L-cysteine dose of 1 mmol/L (P < 0.05). Sulfhydryl content linearly increased with L-cysteine supplementation (P = 0.01), with no effect on intracellular ROS and sperm lipid peroxidation (P ≥ 0.06) in 5ºC-stored doses. In conclusion, L-cysteine supplementation has a positive effect on sperm motility up to 120 h of storage at 5 °C.

Exploring the physiological plasticity of giant grouper (Epinephelus lanceolatus) to dietary sulfur amino acids and taurine to measure dietary requirements and essentiality.

Giant grouper (Epinephelus lanceolatus) is an economically important yet under-researched species, still reliant on 'trash fish' or generic aquafeeds. The transition toward sustainable formulations is contingent on establishing requirements of target species for limiting nutrients, among which the sulfur amino acids (methionine and cysteine) commonly limit fish growth. Further, there remains significant conjecture around the role of the sulfonic acid taurine in marine aquafeed formulation and its relationship to sulfur amino acids. To develop a species-specific feed formulation for giant grouper, dietary methionine was modulated in a dose-response experiment to achieve five graded levels from 9.5 to 21.5 g/kg, including an additional diet with methionine at 18.6 g/kg supplemented with 8 g/kg taurine. The mean (±SD) cysteine level of the diets was 4.5 ± 0.3 g/kg. Each diet was randomly allocated to triplicate tanks of 14 fish (83.9 ± 8.4 g). The best-fit regression for growth showed that the optimal dietary methionine content was 15.8 g/kg and the total sulfur amino acid content was 20.3 g/kg. Inadequate dietary methionine content triggered physiological responses, including hepatic hyperplasia and hypoplasia at 9.5 and 21.5 g/kg, respectively, and high aspartate transaminase levels at 18.9 g/kg. Moreover, inadequate dietary methionine contents resulted in higher densities of mixed goblet cell mucin and reduced absorptive surface area of posterior intestinal villi. Our results suggest that adequate levels of methionine, but not taurine, improved posterior intestinal conditions and liver homeostasis. These findings may aid in formulating aquafeeds to optimize gastrointestinal and liver functions in juvenile giant grouper.

Inhibition of Cysteine Proteases via Thiol-Michael Addition Explains the Anti-SARS-CoV-2 and Bioactive Properties of Arteannuin B.

Artemisia annua is the plant that produces artemisinin, an endoperoxide-containing sesquiterpenoid used for the treatment of malaria. A. annua extracts, which contain other bioactive compounds, have been used to treat other diseases, including cancer and COVID-19, the disease caused by the virus SARS-CoV-2. In this study, a methyl ester derivative of arteannuin B was isolated when A. annua leaves were extracted with a 1:1 mixture of methanol and dichloromethane. This methyl ester was thought to be formed from the reaction between arteannuin B and the extracting solvent, which was supported by the fact that arteannuin B underwent 1,2-addition when it was dissolved in deuteromethanol. In contrast, in the presence of N-acetylcysteine methyl ester, a 1,4-addition (thiol-Michael reaction) occurred. Arteannuin B hindered the activity of the SARS CoV-2 main protease (nonstructural protein 5, NSP5), a cysteine protease, through time-dependent inhibition. The active site cysteine residue of NSP5 (cysteine-145) formed a covalent bond with arteannuin B as determined by mass spectrometry. In order to determine whether cysteine adduction by arteannuin B can inhibit the development of cancer cells, similar experiments were performed with caspase-8, the cysteine protease enzyme overexpressed in glioblastoma. Time-dependent inhibition and cysteine adduction assays suggested arteannuin B inhibits caspase-8 and adducts to the active site cysteine residue (cysteine-360), respectively. Overall, these results enhance our understanding of how A. annua possesses antiviral and cytotoxic activities.

Effects of supplementation of cryopreservation media with cysteine on the post-thaw quality and fertility of brown-marbled grouper (Epinephelus fuscoguttatus) spermatozoa.

The cryopreservation process is associated with the generation of excessive reactive oxygen species, which causes a series of cellular damage to spermatozoa. The objective of the current study was to investigate the effect of different concentrations of cysteine on post-thaw sperm quality of brown-marbled grouper sperm. Semen samples were frozen with cysteine supplemented at 0.5, 1, 2, 5, 10 mM and the control group (no additive). After thawing, sperm quality parameters were analyzed. In comparison to the control, cysteine treatment groups yielded relatively higher sperm total motility, progressive motility, and curvilinear velocity. Different concentrations of cysteine had no effect on average path velocity, straight linear velocity and viability (P > 0.05), while an increase in the concentration of cysteine resulted in a significant improvement in the mitochondrial membrane potential, SOD activity, and ATP content (P < 0.05). As for lipid peroxidation, the extent of which in cysteine treated spermatozoa was less than the control, although the differences were not statistically significant (P > 0.05). In terms of fertilizing capacity, a greater hatching rate (91.7 ± 1.2%) was obtained in thawed sperm treated with 2 mM cysteine, compared to the control (84.3 ± 4.2%; P < 0.05). Overall, it is concluded that the addition of cysteine is helpful in maintaining the function of frozen-thawed brown-marbled grouper sperm, which can be recommended as an effective antioxidant to improve the semen cryopreservation efficiency.

Shikonin Causes Non-apoptotic Cell Death in B16F10 Melanoma.

BACKGROUND: Melanoma treatment is highly resistant to current chemotherapeutic agents. Due to its resistance towards apoptotic cell death, non-apoptotic cell death pathways are sought after. OBJECTIVE: We investigated a Chinese herbal medicine, shikonin, and its effect on B16F10 melanoma cells in vitro. METHODS: Cell growth of B16F10 melanoma cells treated with shikonin was analyzed using an MTT assay. Shikonin was combined with necrostatin, an inhibitor of necroptosis; caspase inhibitor; 3-methyladenine, an inhibitor of autophagy; or N-acetyl cysteine, an inhibitor of reactive oxygen species. Flow cytometry was used to assess types of cell death resulting from treatment with shikonin. Cell proliferation was also analyzed utilizing a BrdU labeling assay. Monodansylcadaverine staining was performed on live cells to gauge levels of autophagy. Western blot analysis was conducted to identify specific protein markers of necroptosis including CHOP, RIP1, and pRIP1. MitoTracker staining was utilized to identify differences in mitochondrial density in cells treated with shikonin. RESULTS: Analysis of MTT assays revealed a large decrease in cellular growth with increasing shikonin concentrations. The MTT assays with necrostatin, 3-methyladenine, and N-acetyl cysteine involvement, suggested that necroptosis, autophagy, and reactive oxygen species are a part of shikonin's mechanism of action. Cellular proliferation with shikonin treatment was also decreased. Western blotting confirmed that shikonin-treated melanoma cells increase levels of stress-related proteins, e.g., CHOP, RIP, pRIP. CONCLUSION: Our findings suggest that mainly necroptosis is induced by the shikonin treatment of B16F10 melanoma cells. Induction of ROS production and autophagy are also involved.

Establishment and optimization of an in vitro guinea pig oocyte maturation system.

Guinea pigs are a valuable animal model for studying various diseases, including reproductive diseases. However, techniques for generating embryos via embryo engineering in guinea pigs are limited; for instance, in vitro maturation (IVM) technique is preliminary for guinea pig oocytes. In this study, we aimed to establish and optimize an IVM method for guinea pig oocytes by investigating various factors, such as superovulation induced by different hormones, culture supplementation (e.g., amino acids, hormone, and inhibitors), culture conditions (e.g., oocyte type, culture medium type, and treatment time), and in vivo hCG stimulation. We found that oocytes collected from guinea pigs with superovulation induced by hMG have a higher IVM rate compared to those collected from natural cycling individuals. Moreover, we found that addition of L-cysteine, cystine, and ROS in the culture medium can increase the IVM rate. In addition, we demonstrated that in vivo stimulation with hCG for 3-8 h can further increase the IVM rate. As a result, the overall IVM rate of guinea pig oocytes under our optimized conditions can reach ~69%, and the mature oocytes have high GSH levels and normal morphology. In summary, we established an effective IVM method for guinea pig oocytes by optimizing various factors and conditions, which provides a basis for embryo engineering using guinea pigs as a model.

Improving endothelial health with food-derived H2S donors: an in vitro study with S-allyl cysteine and with a black-garlic extract enriched in sulfur-containing compounds.

A healthy vascular endothelium plays an essential role in modulating vascular tone by producing and releasing vasoactive factors such as nitric oxide (NO). Endothelial dysfunction (ED), the loss of the endothelium physiological functions, results in the inability to properly regulate vascular tone, leading to hypertension and other cardiovascular risk factors. Alongside NO, the gasotransmitter hydrogen sulfide (H2S) has emerged as a key molecule with vasodilatory and antioxidant activities. Since a reduction in H2S bioavailability is related to ED pathogenesis, natural H2S donors are very attractive. In particular, we focused on the sulfur-containing amino acid S-allyl cysteine (SAC), a bioactive metabolite, of which black garlic is particularly rich, with antioxidant activity and, among others, anti-diabetic and anti-hypertensive properties. In this study, we analyzed the protective effect of SAC against ED by evaluating reactive oxygen species level, H2S release, eNOS phosphorylation, and NO production (by fluorescence imaging and western blot analysis) in Bovine Aortic Endothelial cells (BAE-1). Furthermore, we chemically characterized a Black Garlic Extract (BGE) for its content in SAC and other sulfur-containing amino acids. BGE was used to carry out an analysis on H2S release on BAE-1 cells. Our results show that both SAC and BGE significantly increase H2S release. Moreover, SAC reduces ROS production and enhances eNOS phosphorylation and the consequent NO release in our cellular model. In this scenario, a natural extract enriched in SAC could represent a novel therapeutic approach to prevent the onset of ED-related diseases.

Effects of dietary sulfur amino acid levels on growth performance and intestinal immunity in broilers vaccinated and subsequently infected with coccidiosis.

Coccidia vaccination is a common practice in the poultry industry. However, research is lacking regarding the optimal nutritional support for coccidia vaccinated broilers. In this study, broilers were vaccinated with coccidia oocyst at hatch and were fed with a common starter diet from 1 to 10 d. On d 11, the broilers were randomly assigned to groups in a 4 × 2 factorial arrangement. Briefly, the broilers were fed one of four diets containing 0.6, 0.8, 0.9, and 1.0% of standardized ileal digestible methionine plus cysteine (SID M+C), respectively, from 11 to 21 d. On d 14, the broilers from each diet group were orally gavaged with either PBS (Mock challenge) or Eimeria oocysts. Compared to PBS-gavaged broilers and regardless of dietary SID M+C levels, the Eimeria-gavaged broilers had 1) decreased gain-to-feed ratio (15-21 d, P = 0.002; 11-21 d, P = 0.011); 2) increased fecal oocysts (P < 0.001); 3) increased plasma anti-Eimeria IgY (P = 0.033); and 4) increased intestinal luminal interleukin-10 (IL-10; duodenum, P = 0.039; jejunum, P = 0.018) and gamma interferon (IFN-γ; duodenum, P < 0.001; jejunum, P = 0.017). Regardless of Eimeria gavage, broilers fed 0.6% SID M+C had decreased (P<0.001) body weight gain (15-21 and 11-21 d) and gain-to-feed ratio (11-14, 15-21, and 11-21 d) when compared to those fed ≥ 0.8% SID M+C. Eimeria challenge increased (P < 0.001) duodenum lesions when the broilers were fed with 0.6, 0.8, and 1.0% SID M+C, and increased (P = 0.014) mid-intestine lesions when the broilers were fed with 0.6 and 1.0% SID M+C. An interaction between the two experimental factors was detected on plasma anti-Eimeria IgY titers (P = 0.022), as coccidiosis challenge increased plasma anti-Eimeria IgY titers only when the broilers were fed with 0.9% SID M+C. In summary, the dietary SID M+C requirement for grower (11-21 d) broilers vaccinated with coccidiosis was ranged from 0.8 to 1.0% for optimal growth performance and intestinal immunity, regardless of coccidiosis challenge.

Effect of L-cysteine treatment to induce postharvest disease resistance of Monilinia fructicola in plum fruits and the possible mechanisms involved.

Plum is an important stone fruit in China, but the fruit is easily perishable and susceptible to infection by pathogens. Traditionally, synthetic fungicides are used to control diseases. However, the side effects of fungicides should not be ignored. Cysteine, generally recognized as safe (GRAS) amino acid, has been reported to play roles in the plant abiotic stress response, but little is known about the role of cysteine to control postharvest diseases in fruits. Therefore, this study was designed to investigate the effect of L-cysteine treatment on control of postharvest brown rot in artificially inoculated plum fruits and the possible biocontrol mechanisms involved. Postharvest plum fruits were inoculated with 1, 10, 100 and 1000 mg L-1 L-cysteine. 100 mg L-1 L-cysteine treatment effectively controlled brown rot in artificially inoculated plum fruits by inducing resistance. Furthermore, 100 mg L-1 L-cysteine treatment increased the activities of glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH), enhanced the content of NADPH of the pentose phosphate pathway, as well as improved the contents of H2O2 and some amino acids in the artificially inoculated plum fruits. 100 mg L-1 L-cysteine treatment also elevated the antioxidant content (AsA, GSH) and the antioxidant enzymes activities (APX, GR, MDAR, DHAR) of the ascorbate-glutathione (AsA-GSH) pathway. The protective effects of L-cysteine treatment on postharvest plum fruits likely be due to activating some defense-related responses of the fruit against infection. L-cysteine treatment is a safe promising method for controlling postharvest brown rot in plum fruits.

Protection Against Drug-Induced Liver Injuries Through Nutraceuticals via Amelioration of Nrf-2 Signaling.

Hepatotoxicity caused by the overdose of various medications is a leading cause of drug-induced liver injury. Overdose of drugs causes hepatocellular necrosis. Nutraceuticals are reported to prevent drug-induced liver failure. The present article aims to review the protection provided by various medicinal plants against hepatotoxic drugs. Ayurveda is considered a conventional restorative arrangement in India. It is consistently used for ages and is still used today to cure drug-induced hepatotoxicity by focusing on antioxidant stress response pathways such as the nuclear factor erythroid-2 (Nrf-2) antioxidant response element signaling pathway. Nrf-2 is a key transcription factor that entangles Kelch-like ECH-associating protein 1, a protein found in the cell cytoplasm. Some antioxidant enzymes, such as gamma glycine cysteine ligase (γ-GCL) and heme oxygenase-1 (HO-1), are expressed in Nrf-2 targeted genes. Their expression, in turn, decreases the stimulation of hepatic macrophages and induces the messenger RNA (mRNA) articulation of proinflammatory factors including tumor necrosis factor α. This review will cover various medicinal plants from a mechanistic view and how they stimulate and interact with Nrf-2, the master regulator of the antioxidant response to counterbalance oxidative stress. Interestingly, therapeutic plants have become popular in the medical sector due to safer yet effective supplementation for the prevention and treatment of new human diseases. The contemporary study is expected to collect information on a variety of therapeutic traditional herbs that have been studied in the context of drug-induced liver toxicity, as nutraceuticals are the most effective treatments for oxidative stress-induced hepatotoxicity. They are less genotoxic, have a lower cost, and are readily available. Together, nutraceuticals exert protective effects against drug-induced hepatotoxicity through the inhibition of oxidative stress, inflammation, and apoptosis. Its mechanism(s) are considered to be associated with the γ-GCL/HO-1 and Nrf-2 signaling pathways.KEY TEACHING POINTSThe liver is the most significant vital organ that carries out metabolic activities of the body such as the synthesis of glycogen, the formation of triglycerides and cholesterol, as well as the formation of bile.Acute liver failure is caused by the consumption of certain drugs; drug-induced liver injury is the major condition.The chemopreventive activity of nutraceuticals may be related to oxidative stress reduction and attenuation of biosynthetic processes involved in hepatic injury via amelioration of the nuclear factor erythroid-2 (Nrf-2) signaling pathway.Nrf-2 is a key transcription factor that is found in the cell cytoplasm resulting in the expression of various genes such as gamma glycine cysteine ligase and heme oxygenase-1.Nutraceutical-rich phytochemicals possess high antioxidant activity, which helps in the prevention of hepatic injury.

Comparative effects of dietary methionine and cysteine supplementation on redox status and intestinal integrity in immunologically challenged-weaned pigs.

Sulfur-containing amino acids such as methionine and cysteine play critical roles in immune system and redox status. A body of evidence shows that metabolic aspects of supplemented Met and Cys may differ in the body. Therefore, the study aimed to investigate the effects of dietary Met and Cys supplementation in immunologically challenged weaned pigs. Forty weaned piglets (6.5 ± 0.3 kg) were randomly allocated to five treatment groups. The treatment included: (1) sham-challenged control (SCC), (2) challenged control (CC), (3) MET (CC + 0.1% DL-Met), (4) CYS (CC + 0.1% L-Cys), and (5) MET + CYS (CC + 0.1% DL-Met + 0.1% L-Cys). On day 7, all pigs were intramuscularly injected with either Escherichia coli O55:B5 lipopolysaccharides (LPS) or phosphate-buffered saline. Blood, liver, and jejunum samples were analyzed for immune response and redox status. The CC group had lower (P < 0.05) villus surface area and higher (P < 0.05) flux of 4-kDa fluorescein isothiocyanate dextran (FD4) than the SCC group. A lower (P < 0.05) glutathione (GSH) concentration was observed in the jejunum of pigs in the CC group than those in the SCC group. Dietary Cys supplementation increased (P < 0.05) villus surface area, GSH levels, and reduced (P < 0.05) the flux of FD4 in the jejunum of LPS-challenged pigs. Dietary Met supplementation enhanced (P < 0.05) hepatic GSH content. Pigs challenged with LPS in the MET group had lower serum IL-8 concentration than those in the CC group. There was a Met × Cys interaction (P < 0.05) in serum IL-4 and IL-8 concentrations, and Trolox equivalent antioxidant capacity. Dietary L-Cys supplementation restored intestinal integrity and GSH levels that were damaged by lipopolysaccharides administration. Dietary DL-Met supplementation improved hepatic GSH and reduced systemic inflammatory response, but antagonistic interaction with dietary L-Cys supplementation was observed in the inflammatory response and redox status.

Use of Purslane (Portulaca oleracea) Extracts and Cysteine in Diluents to Improve the Cryopreserved Quality of Goat Sperm.

Highlights Using cysteine and purslane extracts in extenders improved significantly the post-thaw sperm characteristics. Sperm viability, DNA integrity, and mitochondrial activity demonstrate an improvement in post-thaw sperm. Malondialdehyde production was decreased based on the positive effects of treated extenders. The obtained results demonstrate that supplementation of 50 µg/mL of purslane methanolic extract with cysteine to freezing extenders was significantly superior compared with other treatments.

Effects of cysteine addition to low-fishmeal diets on the growth, anti-oxidative stress, intestine immunity, and Streptococcus agalactiae resistance in juvenile golden pompano (Trachinotus ovatus).

As the precursor of taurine, cysteine serves physiological functions, such as anti-oxidative stress and immune improvement. Investigation of cysteine and its derivatives has made positive progress in avian and mammalian species, yet the study and application of cysteine in aquatic animals are relatively rare. Therefore, we evaluated the effects of supplementing a low-fishmeal diet with various levels of cysteine on the growth, antioxidant capacity, intestine immunity, and resistance against Streptococcus agalactiae of the juvenile golden pompano (Trachinotus ovatus). According to our study, exogenous supplementation with 0.6-1.2% cysteine greatly increased the final body weight (FBW) and specific growth rate (SGR) of golden pompano compared to the control group. Under the present conditions, the optimum dietary cysteine supplementation level for golden pompano was 0.91% based on the polynomial regression analysis of SGR. Meanwhile, we found that the Nrf2/Keap1/HO-1 signaling pathway was notably upregulated with the increase of exogenous cysteine, which increased antioxidant enzyme activity in serum and gene expression in the intestine and reduced the level of reactive oxygen species (ROS) in the serum of golden pompano. In addition, morphological analysis of the midgut demonstrated that exogenous cysteine improved muscle thickness and villi length, which suggested that the physical barrier of the intestine was greatly strengthened by cysteine. Moreover, cysteine increased the diversity and relative abundance of the intestinal flora of golden pompano. Cysteine suppressed intestinal NF-κB/IKK/IκB signaling and pro-inflammatory cytokine mRNA levels. Conversely, intestinal anti-inflammatory cytokine gene expression and serum immune parameters were upregulated with the supplementary volume of cysteine and improved intestine immunity. Further, exogenous cysteine supplementation greatly reduced the mortality rate of golden pompano challenged with S. agalactiae. In general, our findings provide more valuable information and new insights into the rational use of cysteine in the culture of healthy aquatic animals.

The Stimulation of Indigenous Bacterial Antagonists by γ-Glutamyl-S-Allyl-l-Cysteine Increases Soil Suppressiveness to Fusarium Wilt.

The development of suppressive soil is an ideal strategy to sustainably combat soilborne diseases. Previously, the cultivation of Allium plants increased antagonistic bacteria populations in soil, alleviating Fusarium wilt of different crops. This study aimed to identify a compound produced by Allium plants that can induce bacteria-mediated soil suppressiveness toward Fusarium wilt. The amendment of soils with γ-glutamyl-S-allyl-l-cysteine (GSAC), a unique dipeptide abundantly detected in the root extract of Welsh onion (Allium fistulosum), significantly suppressed Fusarium wilt diseases, whereas three other commercial dipeptides had no such effects. GSAC application did not suppress the disease in sterilized soil. Furthermore, the suppressiveness of soil amended with GSAC could be transferred to sterilized soil via soil microflora transplantation. This suppressiveness was eliminated by pretreating GSAC-amended soil microflora with antibacterial antibiotics, indicating that the suppressiveness of GSAC-amended soil is generated by the activity of antagonistic bacteria. Amplicon sequencing of the 16S rRNA gene revealed that GSAC application significantly increased the relative abundance of Pseudomonas (OTU224), Burkholderia-Caballeronia-Paraburkholderia (OTU387), and Bdellovibrio (OTU1259) in soils. Surprisingly, the relative abundance of OTU224 was significantly greater in Welsh onion rhizospheres than in noncultivated soil. Pseudomonas strains corresponding to OTU224, isolated from Welsh onion rhizospheres, displayed a remarkable suppressive effect against cucumber Fusarium wilt, implying that OTU224 was involved in GSAC-mediated suppressiveness. This is the first study on the potential of GSAC as a soil microflora-manipulating agent that can enhance soil suppressiveness to Fusarium wilt. IMPORTANCE Methods for increasing soil suppressiveness via soil microflora manipulation have long been explored as an ideal strategy to protect plants from soilborne pathogens. However, viable methods offering consistent disease control effects have not yet been developed. Previously, the cultivation of Allium plants was demonstrated to induce bacteria-mediated soil suppressiveness to Fusarium wilt of different crop plants. This study discovered that the application of γ-glutamyl-S-allyl-l-cysteine, a unique dipeptide synthesized by Welsh onion, to soil enhances Fusarium wilt suppressiveness by increasing the relative abundance of indigenous antagonistic bacteria irrespective of the soil type. This finding will facilitate research supporting the development of environmentally friendly control measures for soilborne diseases.