Carbon Monoxide Alleviates Salt-Induced Oxidative Damage in Sorghum bicolor by Inducing the Expression of Proline Biosynthesis and Antioxidant Genes.

Crop growth and yield are affected by salinity, which causes oxidative damage to plant cells. Plants respond to salinity by maintaining cellular osmotic balance, regulating ion transport, and enhancing the expression of stress-responsive genes, thereby inducing tolerance. As a byproduct of heme oxygenase (HO)-mediated degradation of heme, carbon monoxide (CO) regulates plant responses to salinity. This study investigated a CO-mediated salt stress tolerance mechanism in sorghum seedlings during germination. Sorghum seeds were germinated in the presence of 250 mM NaCl only, or in combination with a CO donor (1 and 1.5 µM hematin), HO inhibitor (5 and 10 µM zinc protoporphyrin IX; ZnPPIX), and hemoglobin (0.1 g/L Hb). Salt stress decreased the germination index (47.73%) and root length (74.31%), while hydrogen peroxide (H2O2) (193.5%), and proline (475%) contents increased. This increase correlated with induced HO (137.68%) activity and transcripts of ion-exchanger and antioxidant genes. Salt stress modified vascular bundle structure, increased metaxylem pit size (42.2%) and the Na+/K+ ratio (2.06) and altered primary and secondary metabolites. However, exogenous CO (1 µM hematin) increased the germination index (63.01%) and root length (150.59%), while H2O2 (21.94%) content decreased under salt stress. Carbon monoxide further increased proline (147.62%), restored the vascular bundle structure, decreased the metaxylem pit size (31.2%) and Na+/K+ ratio (1.46), and attenuated changes observed on primary and secondary metabolites under salt stress. Carbon monoxide increased HO activity (30.49%), protein content, and antioxidant gene transcripts. The alleviatory role of CO was abolished by Hb, whereas HO activity was slightly inhibited by ZnPPIX under salt stress. These results suggest that CO elicited salt stress tolerance by reducing oxidative damage through osmotic adjustment and by regulating the expression of HO1 and the ion exchanger and antioxidant transcripts.

Novel Cytochrome P450-3A4 Enzymatic Nanobiosensor for Lapatinib (a Breast Cancer Drug) Developed on a Poly(anilino-co-4-aminobenzoic Acid-Green-Synthesised Indium Nanoparticle) Platform.

Breast cancer (BC) is one of the most common types of cancer disease worldwide and it accounts for thousands of deaths annually. Lapatinib is among the preferred drugs for the treatment of breast cancer. Possible drug toxicity effects of lapatinib can be controlled by real-time determination of the appropriate dose for a patient at the point of care. In this study, a novel highly sensitive polymeric nanobiosensor for lapatinib is presented. A composite of poly(anilino-co-4-aminobenzoic acid) co-polymer {poly(ANI-co-4-ABA)} and coffee extract-based green-synthesized indium nanoparticles (InNPs) was used to develop the sensor platform on a screen-printed carbon electrode (SPCE), i.e., SPCE||poly(ANI-co-4-ABA-InNPs). Cytochrome P450-3A4 (CYP3A4) enzyme and polyethylene glycol (PEG) were incorporated on the modified platform to produce the SPCE||poly(ANI-co-4-ABA-InNPs)|CYP3A4|PEG lapatinib nanobiosensor. Experiments for the determination of the electrochemical response characteristics of the nanobiosensor were performed with cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The nanobiosensor calibration for 0-100 ng/mL lapatinib was linear and gave limit of detection (LOD) values of 13.21 ng/mL lapatinib and 18.6 ng/mL lapatinib in physiological buffer and human serum, respectively. The LOD values are much lower than the peak plasma concentration (Cmax) of lapatinib (2.43 µg/mL), which is attained 4 h after the administration of a daily dose of 1250 mg lapatinib. The electrochemical nanobiosensor also exhibited excellent anti-interference performance and stability.

Hen egg white bovine colostrum supplement reduces symptoms of mild/moderate COVID-19: a randomized control trial.

Aim: The ability of a hen egg white bovine colostrum supplement to prevent severe COVID-19 was tested in a double-blind randomized control study. Methods: Adults with mild/moderate COVID-19, risk factors for severe disease, and within 5 days of symptom onset were assigned to the intervention (n = 77) or placebo (n = 79) arms. Symptoms were documented until day 42 post-enrollment and viral clearance was assessed at 11-13 days post-symptom onset. Results: One participant developed severe COVID-19. The severe-type symptom score was lower in the active arm at 11-13 days post-symptom onset (p = 0.049). Chest pain, fever/chills, joint pain/malaise, and sore throat were significantly less frequent in the active arm. No differences in viral clearance were observed. Conclusion: The intervention reduced symptoms of mild/moderate COVID-19. Clinical Trial Registration: DOH-27-062021-9191 (South African National Clinical Trials Register).

Natural proteins found in milk (lactoferrin) and egg white (ovotransferrin and lysozyme) could have therapeutic value in COVID-19 through their effects on the immune system. We identified bovine colostrum and hen egg white powders containing adequate quantities of these proteins. We investigated whether short-term daily consumption of a hen egg white and bovine colostrum mixture (reconstituted with glycerin and water) could reduce the risk of progression to severe disease and assist in the recovery of patients with mild or moderate COVID-19. Adults with mild or moderate COVID-19 who were within 5 days of symptom onset and had risk factors for severe disease were enrolled, and randomly assigned to take a hen egg white and bovine colostrum mixture or placebo mixture twice daily for 5 days, and then followed up telephonically for 6 weeks. The main findings were that consumption of the hen egg white and bovine colostrum mixture was associated with fewer protocol-defined severe-type symptoms overall, and in particular lower frequencies of joint pain/malaise, chest pain, fever/chills, and sore throat. Only one individual developed severe COVID-19 and therefore the effect of the intervention on reducing the risk of progression to severe disease could not be assessed. The results of this study suggest that consumption of the hen egg white bovine colostrum mixture within a few days of symptom onset lessens symptoms in people with mild or moderate COVID-19.

Methylglyoxal improves zirconium stress tolerance in Raphanus sativus seedling shoots by restricting zirconium uptake, reducing oxidative damage, and upregulating glyoxalase I.

Raphanus sativus also known as radish is a member of the Brassicaceae family which is mainly cultivated for human and animal consumption. R. sativus growth and development is negatively affected by heavy metal stress. The metal zirconium (Zr) have toxic effects on plants and tolerance to the metal could be regulated by known signaling molecules such as methylglyoxal (MG). Therefore, in this study we investigated whether the application of the signaling molecule MG could improve the Zr tolerance of R. sativus at the seedling stage. We measured the following: seed germination, dry weight, cotyledon abscission (%), cell viability, chlorophyll content, malondialdehyde (MDA) content, conjugated diene (CD) content, hydrogen peroxide (H2O2) content, superoxide (O2•-) content, MG content, hydroxyl radical (·OH) concentration, ascorbate peroxidase (APX) activity, superoxide dismutase (SOD) activity, glyoxalase I (Gly I) activity, Zr content and translocation factor. Under Zr stress, exogenous MG increased the seed germination percentage, shoot dry weight, cotyledon abscission, cell viability and chlorophyll content. Exogenous MG also led to a decrease in MDA, CD, H2O2, O2•-, MG and ·OH, under Zr stress in the shoots. Furthermore, MG application led to an increase in the enzymatic activities of APX, SOD and Gly I as well as in the complete blocking of cotyledon abscission under Zr stress. MG treatment decreased the uptake of Zr in the roots and shoots. Zr treatment decreased the translocation factor of the Zr from roots to shoots and MG treatment decreased the translocation factor of Zr even more significantly compared to the Zr only treatment. Our results indicate that MG treatment can improve R. sativus seedling growth under Zr stress through the activation of antioxidant enzymes and Gly I through reactive oxygen species and MG signaling, inhibiting cotyledon abscission through H2O2 signaling and immobilizing Zr translocation.

Review on Surface-Modified Electrodes for the Enhanced Electrochemical Detection of Selective Serotonin Reuptake Inhibitors (SSRIs).

Selective serotonin re-uptake inhibitors (SSRIs) are one of the most commonly prescribed classes of antidepressants used for the treatment of moderate to severe depressive disorder, personality disorders and various phobias. This class of antidepressants was created with improved margins of safety. However, genetic polymorphism may be responsible for the high variability in patients' responses to treatment, ranging from failure to delayed therapeutic responses to severe adverse effects of treatment. It is crucial that the appropriate amount of SSRI drugs is administered to ensure the optimum therapeutic efficacy and intervention to minimise severe and toxic effects in patients, which may be the result of accidental and deliberate cases of poisoning. Determining SSRI concentration in human fluids and the environment with high sensitivity, specificity and reproducibility, and at a low cost and real-time monitoring, is imperative. Electrochemical sensors with advanced functional materials have drawn the attention of researchers as a result of these advantages over conventional techniques. This review article aims to present functional materials such as polymers, carbon nanomaterials, metal nanomaterials as well as composites for surface modification of electrodes for sensitive detection and quantification of SSRIs, including fluoxetine, citalopram, paroxetine, fluvoxamine and sertraline. Sensor fabrication, sensor/analyte interactions, design rationale and properties of functional material and the electrocatalytic effect of the modified electrode on SSRI detection are discussed.

Seed Priming with MeJa Prevents Salt-Induced Growth Inhibition and Oxidative Damage in Sorghum bicolor by Inducing the Expression of Jasmonic Acid Biosynthesis Genes.

Salinity is one of the major detrimental abiotic stresses at the forefront of deterring crop productivity globally. Although the exogenous application of phytohormones has formerly proven efficacious to plants, their effect on the moderately stress-tolerant crop "Sorghum bicolor" remains elusive. To investigate this, S. bicolor seeds primed with methyl jasmonate (0; 10 and 15 µM MeJa) were exposed to salt (200 mM NaCl) stress, and their morpho-physiological, biochemical, and molecular attributes were measured. Salt stress significantly decreased shoot length and fresh weight by 50%, whereas dry weight and chlorophyll content were decreased by more than 40%. Furthermore, salt-stress-induced oxidative damage was evident by the formation of brown formazan spots (indicative of H2O2 production) on sorghum leaves and a more than 30% increase in MDA content. However, priming with MeJa improved growth, increased chlorophyll content, and prevented oxidative damage under salt stress. While 15 µM MeJa maintained proline content to the same level as the salt-stressed samples, total soluble sugars were maintained under 10 µM MeJa, indicating a high degree of osmotic adjustment. Shriveling and thinning of the epidermis and xylem tissues due to salt stress was prevented by MeJa, followed by a more than 70% decrease in the Na+/K+ ratio. MeJa also reversed the FTIR spectral shifts observed for salt-stressed plants. Furthermore, salt stress induced the expression of the jasmonic acid biosynthesis genes; linoleate 92-lipoxygenase 3, allene oxide synthase 1, allene oxide cyclase, and 12-oxophytodienoate reductase 1. In MeJa-primed plants, their expression was reduced, except for the 12-oxophytodienoate reductase 1 transcript, which further increased by 67%. These findings suggest that MeJa conferred salt-stress tolerance to S. bicolor through osmoregulation and synthesis of JA-related metabolites.

Green Synthesis of Iron Oxide (Hematite) Nanoparticles and Their Influence on Sorghum bicolor Growth under Drought Stress.

Drought is a major abiotic stress that confronts plant growth and productivity, thus compromising food security. Plants use physiological and biochemical mechanisms to cope with drought stress, but at the expense of growth. Green-synthesized nanoparticles (NPs) have gained great attention in agriculture due to their environmental friendliness and affordability while serving as potential biofertilizers. This study investigates the role of hematite (αFe2O3) NPs, synthesized from Aspalathus linearis (rooibos), to improve Sorghum bicolor growth under drought stress. About 18 nm, spherical, and highly agglomerated hematite (αFe2O3) NPs were obtained. Sorghum seeds were primed with 5, 10, and 15 mg/L αFe2O3 NPs, and, after seven days of germination, the seedlings were transferred into potting soil, cultivated for fourteen days, and were subsequently water deprived (WD) for a further seven days. A reduction in plant height (78%), fresh (FW; 35%) and dry (DW; 36%) weights, and chlorophyll (chl) content ((total chl (81%), chla (135%), and chlb (1827%)) was observed in WD plants, and this correlated with low nutrients (Mg, Si, P, and K) and alteration in the anatomic structure (epidermis and vascular bundle tissues). Oxidative damage was observed as deep blue (O2●-) and brown (H2O2) spots on the leaves of WD plants, in addition to a 25% and 40% increase in oxidative stress markers (H2O2 and MDA) and osmolytes (proline and total soluble sugars), respectively. Seed priming with 10 mg/L αFe2O3 NPs improved plant height (70%), FW (56%), DW (34%), total Chl (104%), chla (160%) and chlb (1936%), anatomic structure, and nutrient distribution. Priming with 10 mg/L αFe2O3 NPs also protected sorghum plants from drought-induced oxidative damage by reducing ROS formation and osmolytes accumulation and prevented biomolecule degradation. The study concludes that green synthesized hematite NPs positively influenced sorghum growth and prevented oxidative damage of biomolecules by improving nutrient uptake and osmoregulation under drought stress.

Partial compartmentalisation of HIV-1 subtype C between lymph nodes, peripheral blood mononuclear cells and plasma.

HIV-1 compartmentalisation is likely to have important implications for a preventative vaccine as well as eradication strategies. We genetically characterised HIV-1 subtype C variants in lymph nodes, peripheral blood mononuclear cells and plasma of six antiretroviral (ART) naïve individuals and four individuals on ART. Full-length env (n = 171) and gag (n = 250) sequences were generated from participants using single genome amplification. Phylogenetic relatedness of sequences was assessed, and compartmentalisation was determined using both distance and tree-based methods implemented in HyPhy. Additionally, potential associations between compartmentalisation and immune escape mutations were assessed. Partial viral compartmentalisation was present in nine of the ten participants. Broadly neutralising antibody (bnAb) escape was found to be associated with partial env compartmentalisation in some individuals, while cytotoxic T lymphocyte escape mutations in Gag were limited and did not differ between compartments. Viral compartmentalisation may be an important consideration for bnAb use in viral eradication.

HIV-1 subtype C Nef-mediated SERINC5 down-regulation significantly contributes to overall Nef activity.

BACKGROUND: Nef performs multiple cellular activities that enhance HIV-1 pathogenesis. The role of Nef-mediated down-regulation of the host restriction factor SERINC5 in HIV-1 pathogenesis is not well-defined. We aimed to investigate if SERINC5 down-regulation activity contributes to HIV-1 subtype C disease progression, to assess the relative contribution of this activity to overall Nef function, and to identify amino acids required for optimal activity. We measured the SERINC5 down-regulation activity of 106 subtype C Nef clones, isolated from individuals in early infection, for which the Nef activities of CD4 and HLA-I down-regulation as well as alteration of TCR signalling were previously measured. The relationship between SERINC5 down-regulation and markers of disease progression, and the relative contribution of SERINC5 down-regulation to a Nef fitness model-derived E value (a proxy for overall Nef fitness in vivo), were assessed. RESULTS: No overall relationship was found between SERINC5 down-regulation and viral load set point (p = 0.28) or rate of CD4+ T cell decline (p = 0.45). CD4 down-regulation (p = 0.02) and SERINC5 down-regulation (p = 0.003) were significant determinants of E values in univariate analyses, with the greatest relative contribution for SERINC5 down-regulation, and only SERINC5 down-regulation remained significant in the multivariate analysis (p = 0.003). Using a codon-by-codon analysis, several amino acids were significantly associated with increased (10I, 11V, 38D, 51T, 65D, 101V, 188H and, 191H) or decreased (10K, 38E, 65E, 135F, 173T, 176T and, 191R) SERINC5 down-regulation activity. Site-directed mutagenesis experiments of selected mutants confirmed a substantial reduction in SERINC5 down-regulation activity associated with the mutation 173T, while mutations 10K, 135F, and 176T were associated with more modest reductions in activity that were not statistically significant. CONCLUSIONS: These results suggest that SERINC5 down-regulation is a significant contributor to overall Nef function and identify potential genetic determinants of this Nef function that may have relevance for vaccines or therapeutics.

On the initiation of jasmonate biosynthesis in wounded leaves.

The basal level of the plant defense hormone jasmonate (JA) in unstressed leaves is low, but wounding causes its near instantaneous increase. How JA biosynthesis is initiated is uncertain, but the lipolysis step that generates fatty acid precursors is generally considered to be the first step. Here, we used a series of physiological, pharmacological, genetic, and kinetic analyses of gene expression and hormone profiling to demonstrate that the early spiking of JA upon wounding does not depend on the expression of JA biosynthetic genes in Arabidopsis (Arabidopsis thaliana). Using a transgenic system, we showed how decoupling the responses to wounding and JA prevents the perpetual synthesis of JA in wounded leaves. We then used DEFECTIVE IN ANTHER DEHISCENCE1 (DAD1) as a model wound-responsive lipase to demonstrate that although its transient expression in leaves can elicit JA biosynthesis to a low level, an additional level of activation is triggered by wounding, which causes massive accumulation of JA. This wound-triggered boosting effect of DAD1-mediated JA synthesis can happen directly in damaged leaves or indirectly in undamaged remote leaves by the systemically transmitted wound signal. Finally, protein stability of DAD1 was influenced by wounding, α-linolenic acid, and mutation in its catalytic site. Together, the data support mechanisms that are independent of gene transcription and translation to initiate the rapid JA burst in wounded leaves and demonstrate how transient expression of the lipase can be used to reveal changes occurring at the level of activity and stability of the key lipolytic step.

Modification and optimization of the inhibition of HIV-1 cell-to-cell transmission assay.

HIV-1 infection is caused by cell-free and cell-associated viruses. Currently most of the assays used to screen potential HIV-1 entry inhibitors focus on the inhibition of cell-free viruses. One assay that is widely employed is the TZM-bl neutralization assay that uses pseudotyped viruses. However, a study by Abela et al. showed that many inhibitors that potently inhibit cell-free HIV-1 in this assay can be less effective against the cell-to-cell transmission of the virus. These researchers then designed a method to screen entry inhibitors for activity against cell-associated HIV-1, using pseudotyped viruses. The main limitation of this method, however, was that it can only be reliably employed against viruses that cannot infect target cells as cell-free virion in the absence of a polycation supplement such as DEAE (diethylaminoethyl). Thus, in the current study we provide modifications to this method that solves the problem and makes it possible to study entry inhibitors against cell-to-cell infection of both polycation depend and independent viruses. The main modification involves the introduction of the relative light unit (RLU) vs. virus producing 293-T cells / corresponding supernatants graph. This graph is used to select a virus input that only allows for the detection of cell-associated viruses infection.•The method is a modification of the cell-to-cell transmission assay published by Abela et al.•The method allows for the study of the inhibition of cell-to-cell transmission of both polycation dependent and independent HIV-1 pseudoviruses.

Calcium Improves Germination and Growth of Sorghum bicolor Seedlings under Salt Stress.

Salinity is a major constraint limiting plant growth and productivity worldwide. Thus, understanding the mechanism underlying plant stress response is of importance to developing new approaches that will increase salt tolerance in crops. This study reports the effects of salt stress on Sorghum bicolor during germination and the role of calcium (Ca2+) to ameliorate some of the effects of salt. To this end, sorghum seeds were germinated in the presence and absence of different NaCl (200 and 300 mM) and Ca2+ (5, 15, or 35 mM) concentrations. Salt stress delayed germination, reduced growth, increased proline, and hydrogen peroxide (H2O2) contents. Salt also induced the expression of key antioxidant (ascorbate peroxidase and catalase) and the Salt Overlay Sensitive1 genes, whereas in the presence of Ca2+ their expression was reduced except for the vacuolar Na+/H+ exchanger antiporter2 gene, which increased by 65-fold compared to the control. Ca2+ reversed the salt-induced delayed germination and promoted seedling growth, which was concomitant with reduced H2O2 and Na+/K+ ratio, indicating a protective effect. Ca2+ also effectively protected the sorghum epidermis and xylem layers from severe damage caused by salt stress. Taken together, our findings suggest that sorghum on its own responds to high salt stress through modulation of osmoprotectants and regulation of stress-responsive genes. Finally, 5 mM exogenously applied Ca2+ was most effective in enhancing salt stress tolerance by counteracting oxidative stress and improving Na+/K+ ratio, which in turn improved germination efficiency and root growth in seedlings stressed by high NaCl.

Comparison of the antiviral activity of the microbicide candidate griffithsin and its tandemers derivatives against different modes of HIV-1 transmission.

Tandemers 2MG, 2MG3, 3MG and 4MG are derivatives of the potent anti-HIV-1 microbicide candidate griffithsin (GRFT). We compared these compounds anti-HIV-1 activity to GRFT using the viruses CAP206.08 and CAAN5342.A2 that have decreased sensitivity to this lectin. The 2MG and 2MG3 tandemers had similar activity to GRFT against cell-free and cell-associated viruses, while 3MG and 4MG were significantly more potent. Furthermore, the restoration of the 234N or 295N glycan in these viruses, known to increase sensitivity to GRFT, also increased sensitivity to 2MG and 2MG3, and not to 3MG and 4MG. In addition, GRFT resistant viruses generated in-vitro were equally resistant to 2MG and 2MG3 while they had considerably low resistance to 3MG and 4MG. Lastly, all five compounds showed increased inhibitory activity in seminal and vaginal simulants although the effect was more pronounced in the former. These data support further studies of tandemers as potential microbicides.

Solution structure of RING finger-like domain of retinoblastoma-binding protein-6 (RBBP6) suggests it functions as a U-box.

Retinoblastoma-binding protein-6 (RBBP6) plays a facilitating role, through its RING finger-like domain, in the ubiquitination of p53 by Hdm2 that is suggestive of E4-like activity. Although the presence of eight conserved cysteine residues makes it highly probable that the RING finger-like domain coordinates two zinc ions, analysis of the primary sequence suggests an alternative classification as a member of the U-box family, the members of which do not bind zinc ions. We show here that despite binding two zinc ions, the domain adopts a homodimeric structure highly similar to those of a number of U-boxes. Zinc ions could be replaced by cadmium ions without significantly disrupting the structure or the stability of the domain, although the rate of substitution was an order of magnitude slower than any previous measurement, suggesting that the structure is particularly stable, a conclusion supported by the high thermal stability of the domain. A hallmark of U-box-containing proteins is their association with chaperones, with which they cooperate in eliminating irretrievably unfolded proteins by tagging them for degradation by the proteasome. Using a yeast two-hybrid screen, we show that RBBP6 interacts with chaperones Hsp70 and Hsp40 through its N-terminal ubiquitin-like domain. Taken together with the structural similarities to U-box-containing proteins, our data suggest that RBBP6 plays a role in chaperone-mediated ubiquitination and possibly in protein quality control.

Identification of a novel Arabidopsis thaliana nitric oxide-binding molecule with guanylate cyclase activity in vitro.

While there is evidence of nitric oxide (NO)-dependent signalling via the second messenger cyclic guanosine 3',5'-monophosphate (cGMP) in plants, guanylate cyclases (GCs), enzymes that catalyse the formation of cGMP from guanosine 5'-triphosphate (GTP) have until recently remained elusive and none of the candidates identified to-date are NO-dependent. Using both a GC and heme-binding domain specific (H-NOX) search motif, we have identified an Arabidopsis flavin monooxygenase (At1g62580) and shown electrochemically that it binds NO, has a higher affinity for NO than for O(2) and that this molecule can generate cGMP from GTP in vitro in an NO-dependent manner.