Tanshinone IIA protects intestinal epithelial cells from ferroptosis through the upregulation of <i>GPX4</i> and <i>SLC7A11</i>

Background: Inflammatory bowel disease (IBD) is a chronic inflammatory disease of the gastrointestinal tract. The destruction of the intestinal epithelial barrier is one of the major pathological processes in IBD pathology. Growing evidence indicated that epithelial cell ferroptosis is linked to IBD and is considered a target process. Methods: RAS-selective lethal 3 (RSL3) was used to induce ferroptosis in intestinal epithelial cell line No. 6 (IEC-6) cells, and cell ferroptosis and the effects of tanshinone IIA (Tan IIA) were determined by cell counting kit-8 (CCK-8), reactive oxygen species (ROS) staining, Giemsa staining and transmission electron microscope (TEM). The cell viability of natural product library compounds was determined by CCK-8. The expression of ferroptosis-related genes were detected by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. Results: Treatment of IEC-6 cells results in the accumulation of ROS and typical morphological characteristics of ferroptosis. RSL3 treatment caused rapid cellular cytotoxicity which could be reversed by ferrostatin-1 (Fer-1) in IEC-6 cells. Natural product library screening revealed that Tan IIA is a potent inhibitor of IEC-6 cell ferroptosis. Tan IIA could significantly protect the RSL3-induced ferroptosis of IEC-6 cells. Furthermore, the ferroptosis suppressors, glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), and miR-17-92 were found to be early response genes in RSL3-treated cells. Treatment of IEC-6 cells with Tan IIA resulted in upregulation of GPX4, SLC7A11, and miR-17-92. Conclusion: Our study demonstrated that Tan IIA protects IEC-6 cells from ferroptosis through the upregulation of GPX4, SLC7A11, and miR-17-92. The findings might provide a theoretical grounding for the future application of Tan IIA to treat or prevent IBD.     

Phytohormonal and metabolism analysis of <i>Brassica rapa</i> L. ssp. <i>pekinensis</i> with different resistance during <i>Plasmodiophora brassicae</i> infection

Clubroot of Chinese cabbage (Brassica rapa L. ssp. pekinensis), caused by the obligate parasite Plasmodiophora brassicae, accounts for serious yield losses. The aim of our study was to explore the phytohormone levels and metabolome changes in the roots of resistant and susceptible B. rapa genotypes at a late stage of infection, i.e., 28 days post-infection. Both genotypes showed decreased auxin levels after P. brassicae infection except for indole-3-acetic acid. Overall, the susceptible genotype had higher auxin and cytokinin levels after infection, with the exception of trans-zeatin and 3- indolebutyric acid as compared to the resistant genotype. Jasmonic acid levels declined after infection regardless of the genotype. Resistance against clubroot was evident with the increased levels of salicylic acid in the resistant genotype. The susceptible genotype had a higher number of differentially accumulated metabolites (DAMs) (262) than the resistant genotype (238) after infection. Interestingly, 132 DAMs were commonly detected in both genotypes when infected with the pathogen, belonging to metabolite classes such as phenolic acids, amino acids, and derivatives, glucosinolates, organic acids, flavonoids, nucleotides and derivatives, and fatty acids. The differential metabolite analysis revealed that metabolites related to amino acid biosynthesis, fatty acid biosynthesis and elongation, glutathione metabolism, and glucosinolate metabolism were highly accumulated in the resistant genotype, suggesting their essential roles in resistance against P. brassicae infection.     

<i>Trypanosoma rangeli:</i> growth in mammalian cells <i>in vitro</i> and action of a repositioned drug (17-AAG) and a natural extract (<i>Artemisia sp</i>. essential oil)

Trypanosoma rangeli and T. cruzi are both parasitic unicellular species that infect humans. Unlike T. cruzi, the causative agent of Chagas disease, T. rangeli is an infective and non-pathogenic parasite for humans, but pathogenic for vectors from the Rhodnius genus. Because both species can coexist in different hosts and overlap their infective cycles but very little is known about the infection of T. rangeli in mammalian cells, we decided to characterize both the development of this parasite in cell culture and the effect of therapeutic agents with potential trypanocidal action on it. We found that T. rangeli exhibits a cycle of infection in Vero cells similar to that for T. cruzi and that the repurposed drug, 17-AAG, and the natural extract Artemisia sp. essential oil produce a toxic effect on epimastigotes showing a trypanocidal action from the fifth day of culture. Both treatments also affected the infection of trypomastigotes and reduced the capacity of replication of amastigotes of T. rangeli. Since T. cruzi / T. rangeli coinfection cases have been reported, the finding of drugs with potential activity against both species could be significant in the future. Furthermore, studies of susceptibility of both species to drugs could also help to know the different mechanisms of pathogenicity in humans displayed by T. cruzi that are absent in T. rangeli     

Isolation and species diversity of arbuscular mycorrhizal fungi in the rhizosphere of <i>Puccinellia tenuiflora</i> of Songnen saline-alkaline grassland,China

Salinization has led to the deterioration of the ecological environment, affected the growth of plants, and hindered the development of agriculture and forestry. Arbuscular mycorrhizal (AM) fungi, as important soil microorganisms, play significant physiological and ecological roles in promoting plant nutrient absorption and improving soil structure. Puccinellia tenuiflora (Turcz.) Scribn. et Merr. in Songnen saline-alkaline grassland was selected as the research object to observe AM fungal colonization of the roots and explore the species and diversity of AM fungi in symbiotic association with P. tenuiflora. This study showed that AM fungi colonized in P. tenuiflora roots and formed a typical Arum–type mycorrhizal structure. A significant correlation was observed between vesicular abundance and the colonization intensity of mycorrhiza. Isolation and identification revealed 40 species of AM fungi in the rhizosphere of P. tenuiflora, belonging to 14 genera, of which two species could not be identified. The richness of the genus Glomus was the highest, accounting for 30% of the total species. Funneliformis mosseae and Rhizophagus intraradices were isolated from all the samples and were the species with the widest distribution in the rhizosphere of P. tenuiflora. Correlation analysis showed that pH only had a significant impact on the distribution of a few species, such as Glomus pustulatum, Diversispora spurca, Glomus aggregatum, Rhizophagus clarum, and Acaulospora foveata. The present study provides a theoretical basis for further exploring the resources of AM fungi in saline-alkaline soil.     

<i>In vitro</i> propagation of <i>Opuntia ellisiana</i> Griff. and acclimatization to field conditions

The genus Opuntia is a valuable forage resource in arid and semiarid lands during periods of drought and shortage of herbaceous plants. However, absolute minimum temperatures in the plains of Mendoza represent a limiting factor to cultivate several species.
Opuntia ellisiana is a cold hardy species, so the goals of this study were to massively propagate it using in vitro culture techniques, and then to acclimatize plantlets obtained to field conditions.
Different sterilization protocols were tested. Areoles were isolated in laminar airflow cabinet, and cultured on Murashige-Skoog medium, supplemented with sucrose and different BAP and IBA combinations. Explants were grown at 27±2ºC, under a 16-h photoperiod. The shoots produced were used in the rooting assay using different auxin combinations. In the most efficient growth treatment, plantlets reached 100% shooting after 35 days of culture, and a mean length of 10.2 mm after 49 days of culture. A 100% rooted plantlets was obtained on a medium containing 5 mg L^-1 IBA, after 12 days of culture. Acclimatization was achieved under greenhouse conditions, showing 100% plantlet survival.
This study suggests that O. ellisiana can be successfully micropropagated by areoles, and easily acclimatizated to field conditions.     

A <i>Brachypodium distachyon</i> calcineurin B-like protein-interacting protein kinase,BdCIPK26, enhances plant adaption to drought and high salinity stress

As sessile organisms, plants possess a complex system to cope with environmental changes. Ca²⁺ functions as a vital second messenger in the stress signaling of plants, and the CBL-interacting protein kinases (CIPKs) serve as essential elements in the plant Ca²⁺ signaling pathway. In this study, calcineurin B-like protein-interacting protein kinase 26 (BdCIPK26) from Brachypodium distachyon was characterized. Overexpression of BdCIPK26 enhanced tolerance to drought and salt stress of transgenic plants. Further investigations revealed that BdCIPK26 participated in abscisic acid (ABA) signaling, conferred hypersensitivity to exogenous ABA in transgenic plants, and promoted endogenous ABA biosynthesis. Moreover, BdCIPK26 was found to maintain ROS homeostasis in plants under stress conditions. Therefore, this study indicates that BdCIPK26 functions as a positive regulator in drought and salt stress response.     

Karyological and electrophoretic differences between <i>Pomacea flagellata</i> and <i>P. patula catemacensis</i> (Caenogastropoda: Ampullariidae)

The widespread Mexican apple snail Pomacea flagellata (Say 1827) and the strictly endemic "tegogolo" P. patula catemacensis (Baker 1922) (restricted to Lake Catemaco), are the only known American Ampullariidae that have haploid complements n=13. Pomacea patula catemacensis has suffered a critical reduction in abundance due to immoderate fishing for human consumption. Chromosome slides were obtained from colchicine-injected Pomacea snails collected from nine locations along the coastal zone of the Gulf of Mexico, including Lake Catemaco, for use in principal component analysis (PCA). Total proteins in foot homogenates were analyzed through isoelectric focusing (IEF) and native-PAGE electrophoresis on polyacrylamide gels. The chromosome number 2n=26 was confirmed for snails from all locations, with a uniform 9 m + 4 sm formula. However, P. patula catemacensis showed significantly larger chromosomes (absolute size) than any population of P. flagellata. Pomacea patula catemacensis also differed from all populations of P. flagellata in a PCA with standardized data, i.e., independently of the absolute size difference between species. Proteins with an acid isoelectric point were dominant in the foot of both species. The electrophoresis analysis showed that P. flagellata has 17 protein bands, with an upper bound at IEF=7.6, while P. patula catemacensis has only 15 bands, with an upper bound at IEF=7 and a more evenly spaced band pattern. Molecular weights ranged from 40 to approximately 130 kDa in both species. Proteins with high values (>94 kDa) were the most abundant. Pomacea patula catemacensis showed a band of 93 kDa, which was absent from all specimens of P. flagellata. Samples of P. flagellata did not cluster according to any geographical pattern in the statistical analyses, nor did they show any taxonomically useful differences in their electrophoretic patterns that merit sub-specific discrimination.     

The chloroplast genome comparative characteristic of artificial breeding tree,a case about <i>Broussonetia kazinoki</i> × <i>Broussonetia papyrifera</i>

Broussonetia kazinoki × Broussonetia papyrifera (ZJGS) is a hybrid species in Moraceae family, which has a very complicated hybrid origin. The excellent characteristics of fast growth, strong soil and water conservation ability, high leaf protein content and stem fiber content in ZJGS make it both ecological benefits in the mining area and economically valuable. This study aims to further understand ZJGS and other Moraceae taxa through the ZJGS chloroplast (cp) genome structure and the comparison with 12 closely related Moraceae species. Among the 13 Moraceae species, the cp genome length of seven Broussonetia species (ranges from 160,239 bp to 162,594 bp) is larger than that of six Morus species (ranges from 158,459 bp to 159,265 bp). Among the 77 shared protein-coding genes (PCGs) in Moraceae species, the obvious positive selection of Ka/Ks ratios acted on petD and rpl16 genes of B. kazinoki and B. papyrifera, respectively. Phylogenetic analysis based on shared PCGs from 28 species shows that ZJGS is closely related to maternal B. kazinoki. These findings provide data support for the origin of ZJGS hybridization and provide genomic resources for future ZJGS resource development and molecular breeding.     

Secondary zoospores in the algal endoparasite <i>Maullinia ectocarpii</i> (Plasmodiophoromycota)

The present paper deals with the ultrastructure of zoospores produced by the plasmodiophorid Maullinia ectocarpii, living in the marine algal host Ectocarpus siliculosus. The zoospores described here are very similar to secondary zoospores of Polymyxa graminis and Phagomyxa sp. (the latter an algal endoparasite, also). Our results indicate that M. ectocarpii produces two types of plasmodia, and suggest that is a species with a complete life cycle, as it is known for all the Plasmodiophormycota that have been studied. Sporogenic and sporangial plasmodia produce, respectively, primary zoospores with parallel flagella within thick walled resting sporangia, and secondary zoospores with opposite flagella within thin walled sporangia.     

Polymorphic information and genetic diversity in <i>Brassica</i> species revealed by RAPD markers

Randomly amplified polymorphic DNA (RAPD) is a tremendously convenient approach used to discriminate between Brassica species owing to its accuracy and speed. RAPD primers generate adequate genetic information that can be used in the primer-marker system. In this work, twenty RAPD-PCR based markers were executed to generate polymorphic data, like polymorphic information content (PIC), mean resolving power (MRP), resolving power (RP), effective multiplex ratio (EMR), and marker index (MI) for the first time and genetic distance among and between six Brassica species were calculated. Our results indicated that 20 primers produced a total of 231 scored band and generated 87% polymorphic bands. Average PIC, MRP, RP, MI, and EMR values were 0.088, 0.65, 6.7, 0.78, and 8.9, respectively. PIC showed an overall negative correlation with MRP, RP, MI, and EMR, whereas MRP, RP, and EMR, were positively correlated with each other. Genetic identities ranged from 41.99% (between Brassica napus and Brassica oleracea) to 68.83% (between Brassica campestris and Brassica oleracea). Dendrogram results showed no clustering between species except between Brassica campestris and Brassica nigra. Nevertheless, these results will be helpful to acquire useful information about the markers and their use to determine the genomic structures of Brassica species. Further, based on genetic distance and polymorphic information, new hybrids can be developed for effective oilseed production.     

Application of ferrous sulfate alleviates negative impact of cadmium in rice (<i>Oryza sativa</i> L.)

Soil contamination with toxic heavy metals [such as cadmium (Cd)] is becoming a serious global problem due to rapid development of social economy. Iron (Fe), being an important element, has been found effective in enhancing plant tolerance against biotic and abiotic stresses. The present study investigated the extent to which different levels of Ferrous sulphate (FeSO₄) modulated the Cd tolerance of rice (Oryza sativa L.), when maintained in artificially Cd spiked regimes. A pot experiment was conducted under controlled conditions for 146 days, by using natural soil, mixed with different levels of CdCl₂ [0 (no Cd), 0.5 and 1 mg/kg] together with the exogenous application of FeSO₄ at [0 (no Fe), 1.5 and 3 mg/kg] levels to monitor different growth, gaseous exchange characteristics, oxidative stress, antioxidative responses, minerals accumulation, organic acid exudation patterns of O. sativa. Our results depicted that addition of Cd to the soil significantly (P < 0.05) decreased plant growth and biomass, gaseous exchange parameters, mineral uptake by the plants, sugars (soluble, reducing, and non-reducing sugar) and altered the ultrastructure of chloroplasts, plastoglobuli, mitochondria, and many other cellular organelles in Cd-stressed O. sativa compared to those plants which were grown without the addition of Cd in the soil. However, Cd toxicity boosted the production of reactive oxygen species (ROS) by increasing the contents of malondialdehyde (MDA), which is the indication of oxidative stress in O. sativa and was also manifested by hydrogen peroxide (H₂O₂) contents and electrolyte leakage to the membrane bounded organelles. Although, activities of various antioxidative enzymes like superoxidase dismutase (SOD), peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) and non-enzymatic antioxidants like phenolics, flavonoid, ascorbic acid, anthocyanin and proline contents increased up to a Cd level of 0.5 mg/kg in the soil but were significantly diminished at the highest Cd level of 1 mg/kg in the soil compared to those plants which were grown without the addition of Cd in the soil. The negative impacts of Cd injury were reduced by the application of FeSO₄ which increased plant growth and biomass, improved photosynthetic apparatus, antioxidant enzymes, minerals uptake together with diminished exudation of organic acids as well as oxidative stress indicators in roots and shoots of O. sativa by decreasing Cd retention in different plant parts. These results shed light on the effectiveness of FeSO₄ in improving the growth and upregulation of antioxidant enzyme activities of O. sativa in response to Cd stress. However, further studies at field levels are required to explore the mechanisms of FeSO₄-mediated reduction of the toxicity of not only Cd, but possibly also other heavy metals in plants.     

Cloning and characterization of 5-enopyruvylshikimate-3-phosphate synthase from <i>Phragmites australis</i>

Glyphosate is a non-selective broad-spectrum herbicide that blocks plant growth by inhibiting 5- Enolpyruvylshikimate-3-phosphate synthase (EPSPS), a key enzyme of the shikimate pathway in microorganisms and plants. The full-length epsps cDNA sequence (paepsps, Genebank: KY860582.1) was cloned and characterized for the first time from Phragmites australis. The full-length cDNA of paepsps was 1308 bp encoding a polypeptide of 435 amino acids. The bioinformatic analyses showed that PaEPSPS has highly homologous with EPSPS from other plants. RT-PCR analysis of paepsps expression indicated that the gene expressed in leaves, stems, and roots, with higher expression in leaves. The expression of the paepsps gene increased with glyphosate application. In addition, the transgenic tobacco containing the paepsps gene showed glyphosate resistance in comparison with control. The novel paepsps is a good candidate gene in transgenic crops with glyphosate tolerance in the future.     

Synergistic combination of colistin with imipenem,amikacine or ciprofloxacin against <i>Acinetobacter baumannii</i> and <i>Pseudomonas aeruginosa</i> carbapenem-resistant isolated in Annaba hospital Algeria

Objective: The aim of this study is to detect in vitro the synergetic activity of colistin in combination with imipenem, amikacin or ciprofloxacin, at sub-inhibitory concentrations, against carbapenems-resistant (CR) Acinetobacter baumannii and Pseudomonas aeruginosa strains isolated from various wards in Annaba teaching hospital in eastern Algeria.
Materials and Methods: The minimal inhibitory concentrations (MIC) were determined by broth macrodilution (BMD). Carbapenemase encoding genes were screened using polymerase chain reaction (PCR). The activity of colistin in combination with second antibiotic was evaluated by the Checkerboard Technique.
Results: 39 CR P. aeruginosa and 21 CR A. baumanni strains where collected. The MIC values ranging from (0.25 to 4 µg/ml) to colistin, ≥16 µg/ml for imipenem, ≥4 µg/ml to amikacin and ≥8 µg/ml ciprofloxacin. The PCR reveals the presence of the genes bla_OXA23 (n = 12), blaOXA24 (n = 6), blaNDM1 (n = 3) in A. baumannii and blaVIM2 (n = 12) in P. aeruginosa. The combination of colistin with imipenem showed synergistic effect on 57.14% and 46.15% of A. baumannii and P. aeruginosa isolates, respectively. For colistin and amikacin, the synergistic effect is detected in 28.6% of A. baumannii and 30.8% of P. aeruginosa. While colistin and ciprofloxacin showed synergy on 14.29% and 15.38% of A. baumannii and P. aeruginosa isolates, respectively.
Conclusion: CR A. baumannii and P. aeruginosa remain the most prevalent infection agents in patients from high-risk wards at Annaba Hospital. Colistin associated with imipenem or with amikacin at sub-inhibitory concentrations gives very encouraging results allowing better management of infections caused by this type of bacteria.     

Isolation and molecular characterization of a <i>cax</i> gene from <i>Capsella bursa-pastoris</i>

A new cation exchangers (CAXs) gene was cloned and characterized from Capsella bursapastoris by rapid amplification of cDNA ends (RACE). The full-length cDNA sequence of cax from C. bursa-pastoris (designated as Cbcax51) was 1754 bp containing a 1398 bp open reading frame encoding a polypeptide of 466 amino-acid residues with a calculated molecular mass of 50.5 kDa and an isoelectric point of 5.69. The predicted CbCAX51 contained an IMP dehydrogenase/GMP reductase domain, two Na⁺/Ca²⁺ exchanger protein domains. Comparative and bioinformatics analyses revealed that CbCAX51 showed extensive homology with CAX from other plant species. The expression analysis by different treatments indicated that Cbcax51 could be activated by cold triggering and was related to the cold acclimation process, but its expression is regulated negatively by drought and not affected by ABA or salt.     

Targeting the “undruggable” cancer driver genes: <i>Ras</i>, <i>myc</i>, and <i>tp53</i>

The term “undruggable” is to describe molecules that are not targetable or at least hard to target pharmacologically. Unfortunately, some targets with potent oncogenic activity fall into this category, and currently little is known about how to solve this problem, which largely hampered drug research on human cancers. Ras, as one of the most common oncogenes, was previously considered “undruggable”, but in recent years, a few small molecules like Sotorasib (AMG-510) have emerged and proved their targeted anti-cancer effects. Further, myc, as one of the most studied oncogenes, and tp53, being the most common tumor suppressor genes, are both considered “undruggable”. Many attempts have been made to target these “undruggable” targets, but little progress has been made yet. This article summarizes the current progress of direct and indirect targeting approaches for ras, myc, two oncogenes, and tp53, a tumor suppressor gene. These are potential therapeutic targets but are considered “undruggable”. We conclude with some emerging research approaches like proteolysis targeting chimeras (PROTACs), cancer vaccines, and artificial intelligence (AI)-based drug discovery, which might provide new cues for cancer intervention. Therefore, this review sets out to clarify the current status of targeted anti-cancer drug research, and the insights gained from this review may be of assistance to learn from experience and find new ideas in developing new chemicals that directly target such “undruggable” molecules.