Challenge accepted: Evolutionary lineages versus taxonomic classification of North American shrub willows (Salix).

PREMISE: The huge diversity of Salix subgenus Chamaetia/Vetrix clade in North America and the lack of phylogenetic resolution within this clade has presented a difficult but fascinating challenge for taxonomists to resolve. Here we tested the existing taxonomic classification with molecular tools. METHODS: In this study, 132 samples representing 46 species from 22 described sections of shrub willows from the United States and Canada were analyzed and combined with 67 samples from Eurasia. The ploidy levels of the samples were determined using flow cytometry and nQuire. Sequences were produced using a RAD sequencing approach and subsequently analyzed with ipyrad, then used for phylogenetic reconstructions (RAxML, SplitsTree), dating analyses (BEAST, SNAPPER), and character evolution analyses of 14 selected morphological traits (Mesquite). RESULTS: The RAD sequencing approach allowed the production of a well-resolved phylogeny of shrub willows. The resulting tree showed an exclusively North American (NA) clade in sister position to a Eurasian clade, which included some North American endemics. The NA clade began to diversify in the Miocene. Polyploid species appeared in each observed clade. Character evolution analyses revealed that adaptive traits such as habit and adaxial nectaries evolved multiple times independently. CONCLUSIONS: The diversity in shrub willows was shaped by an evolutionary radiation in North America. Most species were monophyletic, but the existing sectional classification could not be supported by molecular data. Nevertheless, monophyletic lineages share several morphological characters, which might be useful in the revision of the taxonomic classification of shrub willows.

Integrated physiological, transcriptomic and metabolomic analyses provide insights into phosphorus-mediated cadmium detoxification in Salix caprea roots.

Phosphorus (P) plays a crucial role in facilitating plant adaptation to cadmium (Cd) stress. However, the molecular mechanisms underlying P-mediated responses to Cd stress in roots remain elusive. This study investigates the effects of P on the growth, physiology, transcriptome, and metabolome of Salix caprea under Cd stress. The results indicate that Cd significantly inhibits plant growth, while sufficient P alleviates this inhibition. Under Cd exposure, P sufficiency resulted in increased Cd accumulation in roots, along with reduced oxidative stress levels (superoxide anion and hydrogen peroxide contents were reduced by 16.8% and 30.1%, respectively). This phenomenon can be attributed to the enhanced activities of antioxidant enzymes such as superoxide dismutase (SOD) and catalase (CAT), as well as increased levels of antioxidants including ascorbic acid (AsA) and flavonoids under sufficient P conditions. A total of 4208 differentially expressed genes (DEGs) and 552 differentially accumulated metabolites (DAMs) were identified in the transcriptomic and metabolomic analyses, with 2596 DEGs and 113 DAMs identified among treatments with different P levels under Cd stress, respectively. Further combined analyses reveal the potential roles of several pathways in P-mediated Cd detoxification, including flavonoid biosynthesis, ascorbate biosynthesis, and plant hormone signal transduction pathways. Notably, sufficient P upregulates the expression of genes including HMA, ZIP, NRAMP and CAX, all predicted to localize to the cell membrane. This may elucidate the heightened Cd accumulation under sufficient P conditions. These findings provide insights into the roles of P in enhancing plant resistance to Cd stress and improving of phytoremediation.

Divergent responses of rhizosphere soil phosphorus fractions and biological features of Salix psammophila to fertilization strategies under cadmium contamination.

Soil oligotrophy in areas heavily contaminated with heavy metals poses a significant challenge to vegetation establishment and phytoremediation processes. Phosphorus (P) cycling plays a critical role in global biogeochemical cycles, but there is limited understanding of its response to varying fertilization strategies and its correlation with phytoremediation effectiveness. This study primarily investigated the effects of various fertilization strategies, including nitrogen (N, 300 mg·kg-1), P (100 mg·kg-1), NP (combined N and P at 300 mg·kg-1 and 100 mg·kg-1, respectively), and HP (high P, 300 mg·kg-1) application, on rhizosphere soil P fractions and P-solubilizing microbial community (harboring phoD and phoC genes, respectively) of Salix psammophila under cadmium contamination. Application of NP significantly enhanced plant growth and cadmium accumulation, whereas HP inhibited cadmium bioaccumulation but promoted its translocation. Compared to untreated soil, N application promoted P cycling, leading to increases of 141.9 %, 60.4 %, and 10.3 % in Resin-Pi, diluted HCl-Pi, and conc.HCl-Pi, respectively. P application decreased organic phosphorus (Po) fractions by 24.4 % - 225.8 %, but N incorporation mitigated the declining trend in Po and augmented alkaline phosphatase activity. Fertilization strategies significantly regulated phoC- or phoD-harboring bacterial community structure, but their differential nutrient demands resulted in distinct responses. The phoD-harboring bacteria exhibited higher diversity and network complexity, with numerous biomarkers and fertilizer-sensitive OTUs discovered across treatments. Structural equation modeling (SEM) analysis indicated that phytoremediation efficiency was directly affected by Pi fractions, and phoD-harboring bacteria exhibited stronger associations with Pi fractions than phoC-harboring bacteria. In conclusion, our results reveal potential pathways through which fertilization strategies influence phytoremediation by affecting the structure of P-solubilizing microbial community. Furthermore, our study emphasizes the importance of combined N and P application in promoting Cd accumulation in plants, with high P levels appearing as an ideal fertilization strategy for phytoremediation targeting the harvest of aboveground biomass.

Exploring the combined effect of heavy metals on accumulation efficiency of Salix alba raised on lead and cadmium contaminated soils.

The present study illustrated that Salix alba can accumulate high level of Pb and Cd in different plant parts, with maximum accumulation in roots followed by stem and leaves in the order Cd > Pb > Cd + Pb. The phytoremediation evaluation factors such as bioconcentration factor (BCF) and translocation factor (TF) was higher for Cd over Pb in all plant parts, further the BCF for both Pb and Cd was maximum in root (BCF > 1) followed by stem and leaves. Higher accumulation of Cd over the Pb was observed inside the plant tissues due to Cd mimics with other elements and gets transported through respective transporters. The combined treatment of Pb and Cd affected the bioaccumulation at every treatment level suggesting the negative effect among both elements. Higher survival rate (>85%) was recorded up to 200mgPb/kg and 15mgCd/kg, while further increase in metal concentration reduced the plant efficiency to remediate contaminated soils, hence results in declined survival rate. The FTIR analysis revealed that Pb and Cd accumulation in plants induced changes in carboxy, amino, hydroxyl and phosphate groups that ultimately caused alteration in physiological and biochemical processes of plant and thus provided an insight to the interaction, binding and accumulation of heavy metals.

The present study conferred that Salix alba is a heavy metal (Pb and Cd) excluder plant on the basis of phytoremediation efficiency evaluation factors such BCF >1 (root) and TF <1. The correlation studies suggested the negative correlation among Pb and Cd accumulation and morphological traits. Physiological studies indicated that Pb and Cd accumulation negatively affect chlorophyll concentration and the antioxidant mechanism of plants gets activated, further these results are confirmed with FTIR studies, which reported the alteration in functional groups and associated compounds in plant tissues under Pb and Cd stress.

Phosphorus deficiency-induced cell wall pectin demethylesterification enhances cadmium accumulation in roots of Salix caprea.

Regions affected by heavy metal contamination frequently encounter phosphorus (P) deficiency. Numerous studies highlight crucial role of P in facilitating cadmium (Cd) accumulation in woody plants. However, the regulatory mechanism by which P affects Cd accumulation in roots remains ambiguous. This study aims to investigate the effects of phosphorus (P) deficiency on Cd accumulation, Cd subcellular distribution, and cell wall components in the roots of Salix caprea under Cd stress. The results revealed that under P deficiency conditions, there was a 35.4% elevation in Cd content in roots, coupled with a 60.1% reduction in Cd content in shoots, compared to the P sufficiency conditions. Under deficient P conditions, the predominant response of roots to Cd exposure was the increased sequestration of Cd in root cell walls. The sequestration of Cd in root cell walls increased from 37.1% under sufficient P conditions to 66.7% under P deficiency, with pectin identified as the primary Cd binding site under both P conditions. Among cell wall components, P deficiency led to a significant 31.7% increase in Cd content within pectin compared to P sufficiency conditions, but did not change the pectin content. Notably, P deficiency significantly increased pectin methylesterase (PME) activity by regulating the expression of PME and PMEI genes, leading to a 10.4% reduction in the degree of pectin methylesterification. This may elucidate the absence of significant changes in pectin content under P deficiency conditions and the concurrent increase in Cd accumulation in pectin. Fourier transform infrared spectroscopy (FTIR) results indicated an increase in carboxyl groups in the root cell walls under P deficiency compared to sufficient P treatment. The results provide deep insights into the mechanisms of higher Cd accumulation in root mediated by P deficiency.

Flooding regimes alleviate lead toxicity and enhance phytostabilization of salix: Evidence from physiological responses and iron-plaque formation.

Aggravated metal pollution in wetland and riparian zones has become a global environmental issue, necessitating the identification of sustainable remediation approaches. Salix exhibits great potential as a viable candidate for metal(loid) remediation. However, the underlying mechanisms for its effectiveness in different flooding regimes with Pb pollution have not been extensively studied. In this study, fast-growing Salix×jiangsuensis 'J172' was selected and planted in different Pb polluted soils (control, 400 and 800 mg ∙ kg-1) under non-flooded and flooded (CF: continuous flooding and IF: intermittent flooding) conditions for 60 days. This study aimed to explore the effects of flooding on Salix growth performance, physiological traits, and the relationship between Pb uptake/translocation and root Fe plaques. Salix×jiangsuensis 'J172' exhibited excellent tolerance and adaptation to Pb pollution with a tolerance index (TI) exceeding 0.6, even at the highest Pb levels. Moreover, the TIs under flooded conditions were higher than that under non-flooded conditions, suggesting that flooding could alleviate Pb toxicity under co-exposure to Pb and flooding. Leaf malondialdehyde (MDA) exhibited a dose-dependent response to Pb exposure; however, CF or IF mitigated the oxidative damage induced by Pb toxicity with decreased MDA content (2.2-11.9%). The superoxide dismutase and peroxidase activities were generally enhanced by flooding, but combined stress (flooding and Pb) significantly decreased catalase activity. Pb was predominantly accumulated in Salix roots, and flooding markedly increased root Pb accumulation by 19.2-173.0% compared to non-flooded condition. Additionally, a significant positive correlation was observed between the iron (Fe) content of the root plaque and root Pb accumulation, indicating that the formation of Fe plaque on the root surface could enhance the phytostabilization of Pb in Salix. The current findings highlight that fast-growing woody plants are suitable for phyto-management of metal-polluted wetlands and can potentially minimize the risk of metal mobility in soils.

Contrasting effect of pristine, ball-milled and Fe-Mn modified bone biochars on dendroremediation potential of Salix jiangsuensis "172" for cadmium- and zinc-contaminated soil.

Bone biochar (BC) has a high capacity for the immobilization of potentially toxic elements (PTEs); however, its effect on dendroremediation efficiency remains unclear. Therefore, this study aimed to determine the effects of various concentrations (0, 0.5, 1, and 2 wt%) of BC, ball-milled BC (MBC), and Fe-Mn oxide-modified BC (FMBC) on soil properties, plant growth, and metal accumulation in Salix jiangsuensis "172" (SJ-172) grown in cadmium (Cd)- and zinc (Zn)-contaminated soil. BC and MBC promoted the photosynthetic rate, mineral element absorption, and plant growth of SJ-172, whereas FMBC inhibited the growth of SJ-172. Different biochars greatly influenced the concentrations of Cd and Zn in tissues of SJ-172. BC and MBC elevated the Cd levels, whereas FMBC decreased the Cd content in the leaves, stems, and cuttings of SJ-172. Unlikely, BC, MBC and FMBC show no evident change to the Zn concentration in the aboveground tissues of SJ-172, while decreased root Cd and Zn content compared with the control. MBC, at a 2.0% application rate, significantly increased the translocation factors of Cd (55.0%) and Zn (40.87%), whereas BC and FMBC demonstrated no significant effects compared with the control (P > 0.05). Moreover, 2.0% BC and MBC increased Cd and Zn accumulation in SJ-172 by 28.40 and 41.14, and 25.89 and 36.16%, respectively, whereas 2.0% FMBC reduced Cd and Zn accumulation by 53.20% and 13.18 %, respectively, compared with the control. The phytoremediation potential of SJ-172 for Cd- and Zn-contaminated soils was enhanced by MBC and BC, whereas it was lowered by FMBC compared to the control. These results provide novel insights for the application of fast-growing trees assisted by biochar amendments in the dendroremediation of severely PTEs-contaminated soil.

Cell number regulator 8 from Salix linearistipularis enhances cadmium tolerance in poplar by reducing cadmium uptake and accumulation.

Trace metals have relatively high density and high toxicity at low concentrations. Willow (Salix genus) is an excellent phytoremediation species for soil contaminated by trace metal ions. This study identified a cell number regulator (CNR) gene family member in Salix linearistipularis exhibiting strong metal ion resistance: SlCNR8. SlCNR8 expression was affected by various metal ions, including cadmium (Cd), zinc (Zn), copper (Cu), iron (Fe), and manganese (Mn). SlCNR8 overexpression enhanced Cd, Zn, Cu, and Fe resistance in transgenic poplar seedlings (84K) compared with the wild-type (WT). Moreover, transgenic poplar seedlings showed lower root Cd uptake and less Cd accumulation than WT under Cd stress. SlCNR8 was primarily localized to the nucleus and the plasma membrane-like cell periphery. Furthermore, SlCNR8 had transcriptional activation activity in yeast. The transcript levels of multiple metal ion transporters were altered in the roots of transgenic poplar seedlings compared to WT roots under Cd stress. These results suggest that SlCNR8 may enhance Cd resistance in transgenic poplar by reducing Cd uptake and accumulation. This may be related to altered transcription levels of other transporters or to itself. Our study suggests that SlCNR8 can be used as a candidate gene for genetic improvement of phytostabilisation of trace metals by genetic engineering.

In Vitro Anti-Tumor and Hypoglycemic Effects of Total Flavonoids from Willow Buds.

Salix babylonica L. is a species of willow tree that is widely cultivated worldwide as an ornamental plant, but its medicinal resources have not yet been reasonably developed or utilized. Herein, we extracted and purified the total flavonoids from willow buds (PTFW) for component analysis in order to evaluate their in vitro anti-tumor and hypoglycemic activities. Through Q-Orbitrap LC-MS/MS analysis, a total of 10 flavonoid compounds were identified (including flavones, flavan-3-ols, and flavonols). The inhibitory effects of PTFW on the proliferation of cervical cancer HeLa cells, colon cancer HT-29 cells, and breast cancer MCF7 cells were evaluated using an MTT assay. Moreover, the hypoglycemic activity of PTFW was determined by investigating the inhibitory effects of PTFW on α-amylase and α-glucosidase. The results indicated that PTFW significantly suppressed the proliferation of HeLa cells, HT-29 cells, and MCF7 cells, with IC50 values of 1.432, 0.3476, and 2.297 mg/mL, respectively. PTFW, at different concentrations, had certain inhibitory effects on α-amylase and α-glucosidase, with IC50 values of 2.94 mg/mL and 1.87 mg/mL, respectively. In conclusion, PTFW at different doses exhibits anti-proliferation effects on all three types of cancer cells, particularly on HT-29 cells, and also shows significant hypoglycemic effects. Willow buds have the potential to be used in functional food and pharmaceutical industries.

A Comparative Analysis of Phytochemicals, Metal Ions, Volatile Metabolites in Heart Wood, Stem Bark and Leaves of Salix alba L. along with in Vitro Antioxidant, Antacid, Antimicrobial Activities for Sake of Environment Conservation by Substitution of Stem Bark With Leaf.

The genus of Salix is used in food, medicine and nutraceuticals, and standardized by using the single marker compound Salicin only. Stem bark is the official part used for the preparation of various drugs, nutraceuticals and food products, which may lead to overexploitation and damage of tree. There is need to search substitution of the stem bark with leaf of Salix alba L. (SA), which is yet not reported. Comparative phytochemicals viz. Salicin, Procyanidin B1 and Catechin were quantified in the various parts of SA viz. heart wood (SA-HW), stem bark (SA-SB) and leaves (SA-L) of Salix alba L.by using newly developed HPLC method. It was observed that SA-HW and SA-L contained far better amount of Salicin, Procyanidin B and Catechin as compared to SA-SB (SA-HW~SA-L≫SA-SB). Essential and toxic metal ions of all three parts were analysed using newly developed ICP-OES method, where SA-L were founded as a rich source of micronutrients and essential metal ions as compared to SA-SB and SA-HW. GC-MS analysis has shown the presence of fatty acids and volatile compounds. The observed TPC and TFC values for all three parts were ranged from 2.69 to 32.30 mg GAE/g of wt. and 37.57 to 220.76 mg QCE/g of wt. respectively. In DPPH assay the IC50 values of SA-SB, SA-HW, and SA-L were 1.09 (±0.02), 5.42 (±0.08), and 8.82 (±0.10) mg/mL, respectively. The order of antibacterial activities against E. coli, S. aureus, P. aeruginosa, and B. subtilis strains was SA-L>SA-HW>SA-SB with strong antibacterial activities against S. aureus, and B. subtilis strains. The antacid activities order was SA-L>SA-SB>SA-HW. The leaves of SA have shown significant source of nutrients, phytochemicals and medicinal properties than SA-HW and SA-SB. The leaves of SA may be considered as substitute of stem bark to save the environment or to avoid over exploitation, but after the complete pharmacological and toxicological studies.

Manifestation of Triploid Heterosis in the Root System after Crossing Diploid and Autotetraploid Energy Willow Plants.

Successful use of woody species in reducing climatic and environmental risks of energy shortage and spreading pollution requires deeper understanding of the physiological functions controlling biomass productivity and phytoremediation efficiency. Targets in the breeding of energy willow include the size and the functionality of the root system. For the combination of polyploidy and heterosis, we have generated triploid hybrids (THs) of energy willow by crossing autotetraploid willow plants with leading cultivars (Tordis and Inger). These novel Salix genotypes (TH3/12, TH17/17, TH21/2) have provided a unique experimental material for characterization of Mid-Parent Heterosis (MPH) in various root traits. Using a root phenotyping platform, we detected heterosis (TH3/12: MPH 43.99%; TH21/2: MPH 26.93%) in the size of the root system in soil. Triploid heterosis was also recorded in the fresh root weights, but it was less pronounced (MPH%: 9.63-19.31). In agreement with root growth characteristics in soil, the TH3/12 hybrids showed considerable heterosis (MPH: 70.08%) under in vitro conditions. Confocal microscopy-based imaging and quantitative analysis of root parenchyma cells at the division-elongation transition zone showed increased average cell diameter as a sign of cellular heterosis in plants from TH17/17 and TH21/2 triploid lines. Analysis of the hormonal background revealed that the auxin level was seven times higher than the total cytokinin contents in root tips of parental Tordis plants. In triploid hybrids, the auxin-cytokinin ratios were considerably reduced in TH3/12 and TH17/17 roots. In particular, the contents of cytokinin precursor, such as isopentenyl adenosine monophosphate, were elevated in all three triploid hybrids. Heterosis was also recorded in the amounts of active gibberellin precursor, GA19, in roots of TH3/12 plants. The presented experimental findings highlight the physiological basics of triploid heterosis in energy willow roots.

Phytochemical Screening of Ultrasonic Extracts of Salix Species and Molecular Docking Study of Salix-Derived Bioactive Compounds Targeting Pro-Inflammatory Cytokines.

Willow bark (Salix spp., Salicaceae) is a traditional analgesic and antirheumatic herbal medicine. The aim of this study was to evaluate and compare the phytochemical and antioxidant profiles of leaf and bark extracts of six species of the genus Salix obtained by ultrasound-assisted extraction (UAE) and to examine the inhibitory potential of target bioactive compounds against two inflammatory mediators, tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), through in silico molecular docking. The total phenolic and flavonoid content of the extracts was estimated using spectrophotometric methods and the antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and hydroxyl radical (•OH) scavenging assays. Chemical profiling of extracts was carried out using high-performance liquid chromatography coupled with a diode array detector (HPLC-DAD). Principal component analysis (PCA) was performed to differentiate the sample extracts based on their phytochemical profiles and amounts of target bioactive compounds. Chemical composition varied among the analyzed willow species and also among the plant organs of the same species. The major bioactive compounds of the extracts were salicin, chlorogenic acid, rutin and epicatechin. The extracts exhibited significant DPPH● and ●OH scavenging activities. Results of molecular docking revealed that chlorogenic acid had the highest binding affinity toward TNF-α and IL-6. UAE extracts represent valuable sources of antioxidant and anti-inflammatory compounds.

Phylogenomics reveals patterns of ancient hybridization and differential diversification that contribute to phylogenetic conflict in willows, poplars, and close relatives.

Despite the economic, ecological, and scientific importance of the genera Salix L. (willows) and Populus L. (poplars, cottonwoods, and aspens) Salicaceae, we know little about the sources of differences in species diversity between the genera and of the phylogenetic conflict that often confounds estimating phylogenetic trees. Salix subgenera and sections, in particular, have been difficult to classify, with one recent attempt termed a "spectacular failure" due to a speculated radiation of the subgenera Vetrix and Chamaetia. Here, we use targeted sequence capture to understand the evolutionary history of this portion of the Salicaceae plant family. Our phylogenetic hypothesis was based on 787 gene regions and identified extensive phylogenetic conflict among genes. Our analysis supported some previously described subgeneric relationships and confirmed the polyphyly of others. Using an fbranch analysis, we identified several cases of hybridization in deep branches of the phylogeny, which likely contributed to discordance among gene trees. In addition, we identified a rapid increase in diversification rate near the origination of the Vetrix-Chamaetia clade in Salix. This region of the tree coincided with several nodes that lacked strong statistical support, indicating a possible increase in incomplete lineage sorting due to rapid diversification. The extraordinary level of both recent and ancient hybridization in both Salix and Populus have played important roles in the diversification and diversity in these two genera.

Nutritional, biological, and structural properties of bioactive peptides from bellflower, Persian-willow, and bitter-orange pollens.

In this study, the composition of amino acids, nutritional characteristics, degree of hydrolysis (DH), antioxidant properties, and antibacterial activity of proteins and hydrolysates of bellflower (Campanula latifolia), Persian willow (Salix aegyptiaca), and bitter orange (Citrus aurantium L.) were investigated under the influence of different proteases (Alcalase: Al, trypsin: Tr, pancreatin: Pa, and pepsin: Pe). Evaluation of the structural features of the proteins showed amide regions (amide A, B, I-III) and secondary structures. Hydrophobic amino acids (∼38%), antioxidants (∼21%), and essential types (∼46%) form a significant part of the structure of flower pollen. The digestibility and nutritional quality (PER) of the hydrolyzed samples (CP: 1.67; CA: 1.89, and PW: 1.93) were more than the original protein. Among proteins and peptides, the highest degree of hydrolysis (34.6%: Al-PWH), inhibition of free radicals DPPH (84.2%: Al-CPH), ABTS (95.2%: Pa-CPH), OH (86.7%: Tr-CAH), NO (57.8%: Al-CPH), reducing power (1.31: Pa-CPH), total antioxidant activity (1.46: Pa-CPH), and chelation of iron ions (80%: Al-CPH and Al-CAH) and copper (50.3%: Pa-CAH) were affected by protein type, enzyme type, and amino acid composition. Also, the highest inhibition of the growth of Escherichia coli (25 mm) and Bacillus cereus (24 mm) were related to CP and PW hydrolysates, respectively. The results of this research showed that hydrolyzed flower pollens can be used as a rich source of essential amino acids as well as natural antioxidants and antibacterial in food and dietary products. PRACTICAL APPLICATION: Enzymatic hydrolysis of Campanula latifolia, Persian willow, and Citrus aurantium pollen proteins was performed. The hydrolyzed ones had high nutritional quality and digestibility (essential amino acids and PER index). Antioxidant properties and chelation of metal ions of peptides were affected by the type of protein and enzyme. The hydrolysates showed inhibitory activity against the growth of Esherichia coli and Bacillus cereus.

Assessment of the capability of cadmium accumulation and translocation among 31 willows: four patterns of willow biomass variation response to cadmium.

Cadmium (Cd) pollution threatens food security and the environment. Willow species (Salix, Salicaceae) exhibit a remarkable potential to restore Cd-polluted sites due to their high biomass production and high Cd accumulation capacities. This study examined the Cd accumulation and tolerance in 31 genotypes of shrub willow in hydroponic conditions at varying Cd levels (0 µM Cd, 5 µM Cd, and 20 µM Cd). The root, stem, and leaf biomass of 31 shrub willow genotypes showed significant differences to Cd exposure. Among 31 willow genotypes, four patterns of biomass variation response to Cd were identified: insensitive to Cd; growth inhibition due to excessive Cd supply (high Cd inhibition); low Cd causing inhibited growth, whereas high Cd leading to increased biomass (U-shape); and growth increment with excessive Cd exposure (high Cd induction). The genotypes belonging to the "insensitive to Cd" and/or "high Cd induction" were candidates for the utilization of phytoremediation. Based on the analysis of Cd accumulation of 31 shrub willow genotypes at high and low Cd levels, genotypes 2372, 51-3, and 1052 obtained from a cross between S. albertii and S. argyracea grew well and accumulated relatively more Cd levels than other genotypes. In addition, for Cd-treated seedlings, root Cd accumulation was positively correlated with shoot Cd accumulation and total Cd uptake, demonstrating that Cd accumulation in roots could serve as a biomarker for evaluating the Cd extraction capacity of willows, especially in hydroponics screening. The results of this study screened out willow genotypes with high Cd uptake and translocation capacities, which will provide valuable approaches for restoring Cd-contaminated soils with willows.

BuOH fraction of Salix Babylonica L. extract increases pancreatic beta-cell tumor death at lower doses without harming their function.

Salix babylonica L. is a species of the willow tree. Insulinoma is a tumor originating from pancreatic beta cells. This study aims to research the effect of different fractions of Salix babylonica L. leaf extract on INS-1 cells for treating pancreatic tumors. Cell death occurred at lower doses in the EtOAc fraction. The cells are functional in the BuOH fraction but not in EtOAc and H2O fractions. The EtOAc fraction has a higher percentage of necrosis and ROS. INS1, INS2, and AKT gene expressions in the H2O fraction, GLUT2, IR, HSP70 gene expressions, and WNT4 protein levels increased in the BuOH fraction. HSP90 gene expression, Beta-actin, GAPDH, insulin, HSP70, HSP90, HSF1, Beta-Catenin, and WNT7A protein levels were decreased, while IR immunolabelling intensity increased in both fractions. Ca+2, K+, Na+, and CA-19-9 in the cell, Ca+2 and K+ in secretion increased. The secondary metabolites in the EtOAc fraction cause more damage in INS-1 cells. Since the water fraction also causes the cells to die in high doses, cell function is damaged. The secondary metabolites in the BuOH fraction kill INS-1 cells with less damage. This makes the BuOH fraction of Salix babylonica L. more valuable.

Dipeptidyl peptidase-1 inhibition with brensocatib reduces the activity of all major neutrophil serine proteases in patients with bronchiectasis: results from the WILLOW trial.

BACKGROUND: Brensocatib is an oral, selective, reversible inhibitor of dipeptidyl peptidase-1 (DPP-1), responsible for activating neutrophil serine proteases (NSPs) including neutrophil elastase (NE), proteinase 3 (PR3), and cathepsin G (CatG). In chronic inflammatory lung diseases such as non-cystic fibrosis bronchiectasis (NCFBE), neutrophils accumulate in the airways resulting in excess active NSPs that cause damaging inflammation and lung destruction. METHODS: The 24-week WILLOW trial (NCT03218917) was a randomized, double-blind, placebo-controlled, parallel-group trial in patients with NCFBE conducted at 116 sites across 14 countries. In this trial, treatment with brensocatib was associated with improvements in clinical outcomes including time to first exacerbation, reduction in exacerbation frequency and a reduction in NE activity in sputum. An exploratory analysis of NE activity in white blood cell (WBC) extracts and NE, PR3 and CatG activity in sputum was conducted to further characterize brensocatib's effect and identify potential correlated effects. RESULTS: NE, PR3 and CatG activities were reduced in sputum and NE activity was reduced in WBC extracts in a dose-dependent manner after four weeks of brensocatib treatment, with a return to baseline four weeks after the end of treatment. Brensocatib produced the greatest reduction in the sputum activity of CatG, followed by NE and then PR3. Positive correlations among the sputum NSPs were observed both at baseline and in response to treatment, with the strongest correlation among the sputum NSPs for NE and CatG. CONCLUSIONS: These results suggest a broad anti-inflammatory effect of brensocatib underlying its clinical efficacy observed in NCFBE patients. TRIAL REGISTRATION: The study was approved by the corresponding ethical review boards of all participating centers. The trial was approved by the Food and Drug Administration and registered at clinicaltrials.gov (NCT03218917) on July 17, 2017 and approved by the European Medicines Agency and registered at the European Union Clinical trials Register (EudraCT No. 2017-002533-32). An independent, external data and safety monitoring committee (comprising physicians with pulmonary expertise, a statistician experienced in the evaluation of clinical safety, and experts in periodontal disease and dermatology) reviewed all adverse events.

Salix babylonica L. mitigates pancreatic damage by regulating the Beclin-P62/SQSTM1 autophagy pathway in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Salix babylonica L. belongs to the genus Salix, family Salicaceae. It is traditionally used as an antipyretic, antirheumatic, antidiabetic and for the treatment of ulcers and parasite skin diseases. It also has a range of pharmacological effects, such as anti-inflammatory, anti-tumor, antioxidant, and antibacterial effects. However, there are no reports on the phytochemical profile and efficacy of its leaves extract to modulate dexamethasone induced pancreatic damage. AIM OF THE STUDY: The present study was performed to annotate the phytoconstituents of Salix babylonica leaf extract and explore whether and how it could modulate dexamethasone-induced pancreatic damage and the role of oxidative stress and autophagy in mediating its protective effects. MATERIALS AND METHODS: Wistar rats were used for this study. Salix babylonica in two dose levels (100 and 200 mg/kg) or metformin (50 mg/kg) was given by oral gavage concurrently with dexamethasone which was injected SC in a dose of 10 mg/kg for 4 consecutive days. RESULTS: LC-MS analysis furnished 84 secondary metabolites belonging to phenolic acids, salicinoids, proanthocyanidins, flavonoids, cyclohexanediol glycosides, and hydroxy fatty acids. S. babylonica at both dose levels and metformin decreased the elevated pancreatic beclin while elevated the decreased pancreatic P62/SQSTM1 content compared to dexamethasone. These effects were associated with improved histopathological changes, glycemic and lipid parameters indicating that there might be a connection between autophagy and dexamethasone-induced pancreatic damage. Given that the level of GSH was negatively correlated with the levels of beclin and positively correlated with P62/SQSTM1, while both MDA and NO levels were positively correlated with beclin and negatively correlated with P62/SQSTM1, it seems that dexamethasone induced autophagy may be attributed to dexamethasone induced pancreatic oxidative stress. CONCLUSION: Our results indicate that S. babylonica protects pancreatic tissues against dexamethasone-induced damage by decreasing oxidative stress and its associated autophagy. Our study reveals a new mechanism for dexamethasone effects on pancreas and shows the potential therapeutic role of S. babylonica in mitigating dexamethasone adverse effects on pancreas and establishes the groundwork for future clinical applications.

Potential anti-proliferative activity of Salix mucronata and Triticum spelta plant extracts on liver and colorectal cancer cell lines.

Cancer's etiology is linked to oxidative stress. As a result, it's vital to find effective natural antioxidant remedies. Salix mucronata and Triticum spelta plant extracts were prepared using five different solvents and examined for their cytotoxicity against liver HepG2 cancer cell line. It was found that Salix mucronata ethanolic extract is high in antioxidant mediated anti-cancer activity. The functional constituents (phenolic and flavonoids) as well as preparation of different ethanolic concentrations used to study their properties that include DPPH, oxygen, hydroxyl, nitrogen radical scavenging activities, ferric reducing power and metal chelating activities. The MTT assay was used to determine antioxidant-mediated anti-cancer activity against human liver (HepG2) and colorectal (Caco-2) cancer cells to calculate the half-maximal growth inhibitory concentration (IC50). Moreover, flow cytometry analysis was used to quantify the apoptotic effect on the treated cancer cells. Additionally, qRTPCR of p53, BCL2, Cyclin D, MMP9 and VEGF were measured. Furthermore, HPLC was used to assess the most effective ingredients of the plant extract. Salix mucronata 50% ethanol extract had the highest polyphenolic content, anti-oxidant, and anti-proliferative activity. Salix mucronata increased the number of total apoptotic cells, and caused an upregulation of p53 gene expression by more than five folds and a downregulation of gene expression level of BCL2, Cyclin D, MMP9 and VEGF by more than five folds. Consequently, that could modulate oxidative stress and improve the effectiveness of cancer therapy. Results, also, showed that Triticum spelta ethanolic extract was less effective than Salix mucronata. Therefore, Salix mucronata ethanolic extract represents promising surrogate natural therapy for apoptosis-mediated cancer and recommended for further investigation using animal model.

Exogenous hydrogen sulfide and methylglyoxal alleviate cadmium-induced oxidative stress in Salix matsudana Koidz by regulating glutathione metabolism.

BACKGROUND: Cadmium (Cd) is a highly toxic element for plant growth. In plants, hydrogen sulfide (H2S) and methylglyoxal (MG) have emerged as vital signaling molecules that regulate plant growth processes under Cd stress. However, the effects of sodium hydrosulfide (NaHS, a donor of H2S) and MG on Cd uptake, physiological responses, and gene expression patterns of Salix to Cd toxicity have been poorly understood. Here, Salix matsudana Koidz. seedlings were planted in plastic pot with applications of MG (108 mg kg- 1) and NaHS (50 mg kg- 1) under Cd (150 mg kg- 1) stress. RESULTS: Cd treatment significantly increased the reactive oxygen species (ROS) levels and malondialdehyde (MDA) content, but decreased the growth parameters in S. matsudana. However, NaHS and MG supplementation significantly decreased Cd concentration, ROS levels, and MDA content, and finally enhanced the growth parameters. Cd stress accelerated the activities of antioxidative enzymes and the relative expression levels of stress-related genes, which were further improved by NaHS and MG supplementation. However, the activities of monodehydroascorbate reductase (MDHAR), and dehydroascorbate reductase (DHAR) were sharply decreased under Cd stress. Conversely, NaHS and MG applications restored the MDHAR and DHAR activities compared with Cd-treated seedlings. Furthermore, Cd stress decreased the ratios of GSH/GSSG and AsA/DHA but considerably increased the H2S and MG levels and glyoxalase I-II system in S. matsudana, while the applications of MG and NaHS restored the redox status of AsA and GSH and further improved glyoxalase II activity. In addition, compared with AsA, GSH showed a more sensitive response to exogenous applications of MG and NaHS and plays more important role in the detoxification of Cd. CONCLUSIONS: The present study illustrated the crucial roles of H2S and MG in reducing ROS-mediated oxidative damage to S. matsudana and revealed the vital role of GSH metabolism in regulating Cd-induced stress.