Optimizing the Vanillin-Acid Sulfuric Method to Total Saponin Content in Leaves of Yerba Mate Clones.

Yerba mate (Ilex paraguariensis) is a forest species consumed in the form of non-alcoholic beverages in South America, with applications in foods, cosmetics, and pharmaceutical industries. The species leaves are globally recognized for their important bioactive compounds, including, saponins. We adjusted the vanillin-acid sulfuric method for determining spectrophotometrically the total saponin in yerba mate leaves. Seeking to maximize the extraction of saponins from leaves, a Doehlert design combined with Response Surface Methodology (RSM) was used, considering ethanol:water ratios and ultrasound times. In addition, the same methodology was used for the analysis of times and temperatures in the vanillin-sulfuric acid reaction heating. The contents of total saponin in mature leaves were compared in four yerba mate clones. The extraction was maximized using 40 % ethanol:60 % water and 60 minutes of ultrasound assisted extraction (UAE) without heating. For the reaction conditions, 70 °C for 10 minutes heating is recommended, and UV/Vis reading from 460 to 680 nm. Using the optimized methodology, total saponin contents ranged from 28.43 to 53.09 mg g-1 in the four yerba mate clones. The significant difference in saponin contents between clones indicate great genetic diversity and potential for clones' selection and extraction of these compounds from yerba mate leaves.

Cross-reactivity of anti-human programmed cell death ligand 1 (PD-L1) monoclonal antibody, clone 28-8 against feline PD-L1.

Immunotherapy is a breakthrough in human cancer therapy and has become a major concern in veterinary oncology. However, in cats, many unclear points of the tumor microenvironment exist, including immune checkpoint molecules. A reason is that very few monoclonal antibodies have been proven to react with feline molecules. Therefore, this study investigated whether anti-human programmed cell death ligand 1 (PD-L1) monoclonal antibody, clone 28-8, which is currently commercially available, can also recognize feline PD-L1 by flow cytometry, immunoprecipitation, and immunohistochemical (IHC) staining. We confirmed that the antibody's specificity by flow cytometry and immunoprecipitation using NIH3T3 cells transfected with feline PD-L1. Additionally, we revealed that PD-L1 was expressed on the surface of some feline cell lines by flow cytometry and clone 28-8 antibody unbound to the cells where feline PD-L1 was knocked out. Furthermore, IHC analysis revealed that PD-L1 was expressed in macrophages in the spleen and lymph nodes from healthy cats and mast cell tumor cells. Therefore, we indicated that the clone 28-8 antibody is a valuable tool in detecting feline PD-L1, and further analysis of tumor tissues is expected in the future.

Long noncoding RNA FAM157C contributes to clonal proliferation in paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare clonal disease of hematopoietic stem cells (HSCs). Long noncoding RNAs (lncRNAs) perform a wide range of biological functions, including the regulation of gene expression, cell differentiation, and proliferation, but their role in PNH remains unclear.CD59- and CD59+ granulocytes and monocytes from 35 PNH patients were sorted. High-throughput sequencing was analyzed in 5 PNH patients, and differentially expressed lncRNAs and mRNAs were identified. The mRNAs with fragments per kilobase of exon model per million mapped fragments (FPKM) > 10 in at least 3 patients were selected, and experiments were performed to identify their upstream regulatory lncRNAs. The expression of selected mRNAs and lncRNAs was verified by qRT‒PCR, and the correlation of these expression patterns with clinical data from other 30 PNH patients was analyzed. Then, the functions of the lncRNAs were studied in the PIGA-KO-THP-1 cell line.Transcription analysis revealed 742 upregulated and 1376 downregulated lncRNAs and 3276 upregulated and 213 downregulated mRNAs. After deep screening, 8 highly expressed mRNAs that were related to the NF-κB pathway were analyzed to determine coexpression patterns. LINC01002, FAM157C, CTD-2530H12.2, XLOC-064331 and XLOC-106677 were correlated with the 8 mRNAs. After measuring the expression of these molecules in 30 PNH patients by qRT‒PCR, lncRNA FAM157C was verified to be upregulated in the PNH clone, and its expression levels were positively correlated with the LDH levels and CD59- granulated and monocyte cell ratios. After knockdown of the FAM157C gene in the PIGA-KO-THP-1 cell line, we found that the cells were arrested in the G0/G1 phase and S phase, the apoptosis rate increased, and the cell proliferation decreased.LncRNA FAM157C was proven to promote PNH clone proliferation, and this is the first study to explore the role of lncRNAs in PNH.

Characterization of antibody clones that bind exclusively to insoluble fibrin.

Previously, we established an antibody, termed 102-10, which recognizes insoluble fibrin exclusively, unlike the previously established anti-insoluble fibrin antibodies that also cross-reacted with fibrinogen. We established that the epitope of this antibody is on the ß chain that lines an indented structure that becomes exposed only when insoluble fibrin is formed. The amino acid sequence of the epitope is completely conserved from mouse to humans. This study attempted to determine the most suitable insoluble fibrin clone for future diagnostic and therapeutic development. Binding kinetics and properties of antibodies were evaluated by the surface plasmon resonance analysis (SPR) and ELISA among 1101, 99, 443, and 102-10. Immunohistochemical staining for mouse and human pancreatic cancer tissues were also performed. For frozen sections, visually appropriate staining results were observed at an antibody concentration of 1-10 µg/ml, while for paraffin sections, 10 µg/ml was required. From immunohistochemistry and ELISA analyses, clone 99 and clone 1101 showed almost no nonspecific binding in normal pancreatic tissues. Hybridoma production for 1101 yielded more antibodies than that of 99 and demonstrated good long-term stability. It was, therefore, concluded that clone 1101 would be useful for future clinical development as well as basic research.

Metal tolerance capacity and antioxidant responses of new Salix spp. clones in a combined Cd-Pb polluted system.

To investigate the physiochemical characteristics of two new clones, Salix matsudana 'J172' (A7) and Salix matsudana 'Yankang1' (A64) in combined Cd-Pb contaminated systems, a hydroponic experiment was designed. The plant biomass, photosynthesis, antioxidant responses and the accumulation of metals in different plant parts (leaf, stem, and root) were measured after 35-day treatments with Cd (15, 30 µM) and Pb (250, 500 µM). The results showed that exposure to Cd-Pb decreased the biomass but increased the net photosynthetic rate for both A7 and A64, demonstrating that photosynthesis may be one of the metabolic processes used to resist Cd-Pb stress. Compared with control, roots exposed to Cd-Pb had higher activity of superoxide dismutase and more malondialdehyde concentrations, which indicated the roots of both clones were apt to be damaged. The concentrations of soluble protein were obviously higher in the roots of A64 than A7, indicating the roles of the antioxidative substance were different between two willow clones. Soluble protein also had significant relationship with translocation factors from accumulation in roots of A64, which illustrated it played important roles in the tolerance of A64 roots to heavy metals. The roots could accumulate more Pb rather than transport to the shoots compared with Cd. The tolerance index was more than 85% on average for both clones under all the treatments, indicating their tolerance capacities to the combined stress of Cd and Pb are strong under the tested metal levels. Both clones are the good candidates for phytoremediation of Cd and Pb by the root filtration in the combined contamination environment.

Immunohistochemistry with 3 different clones in anaplastic lymphoma kinase fluorescence in situ hybridization positive non-small-cell lung cancer with thymidylate synthase expression analysis: a multicentre, retrospective, Italian study.

Introduction: ALK rearrangement is the only druggable oncogenic driver detectable by immunohistochemistry (IHC) not requiring further confirmation of positivity in accessing first-line specific inhibitors. ALK-positive patients experience clinical benefit from pemetrexed-based chemotherapy possibly due to lower thymidylate synthase (TS) levels. This study assesses agreement with three different ALK IHC clones in 37 FISH-positive NSCLC. TS expression by real time (RT)-PCR was compared with ALK FISH-negative cases. Materials and methods: 37 ALK FISH-positive NSCLC cases diagnosed between 2010 and 2015 in 7 Italian centres were investigated with ICH using three different anti-ALK antibodies (ALK1, 5A4 and D5F3). Staining for ALK1 and 5A4 was graded as 0+,1+,2+, and 3+, while the scoring for D5F3 was recorded as negative or positive. Proportion agreement analysis was done using Cohen's unweighted kappa (k). TS and ß-actin expression levels were analysed by quantitative RT-PCR. Comparison between TS expression in ALK FISH-positive specimens and a control cohort of ALK FISH-negative ones was performed with the Mann-Whitney and Kruskal-Wallis tests. Results: Considering 2+ and 3+ as positive, the proportion of IHC agreement was 0.1691 (95% CI 0-0.4595) for ALK1/5A4, 0.1691 (95% CI 0-0.4595) for ALK1/D5F3, and 1 for D5F3/5A4. Considering 3+ as positive, it was 0.1543 (95% CI 0-0.4665) for ALK1/ 5A4, 0.0212 (95% CI 0-0.1736) for ALK1/D5F3, and 0.2269 (95% CI 0-0.5462) for 5A4/D5F3. Median TS expression was 6.07 (1.28-14.94) and ALK-positive cases had a significant lower TS expression than ALK-negative tumours (p = 0.002). Conclusions: IHC proved to be a reliable tool for the diagnosis of ALK-rearranged NSCLC. D5F3 and 5A4 clones have the highest percentage of agreement. TS levels are significantly lower in FISH-positive patients.

Is the harvest of Salix and Populus clones in the growing season truly advantageous for the phytoextraction of metals from a long-term perspective?

The phytoextraction potential of short-rotation field coppice plantations was investigated on soil historically contaminated with risk elements (REs), especially Cd, Pb and Zn. The main objective of the study was to assess the effect of biomass harvest time (summer harvest versus traditional winter one) on biomass yield, REs accumulation and removal in the long-term study. The precise field experiment with two Salix clones and two Populus clones was established in April 2009. Shoots of all clones were harvested in February 2012 for the first time, and then identical branches and leaves were harvested every two years in September (summer harvest = SH) and the branches every two years in February (winter harvest = WH). The first summer harvest seemed to be more promising compared to the winter one, but the yields in the second and third harvests were inconsistent. The total phytoextraction efficiency of the SH declined in second and third harvests due to a decrease of leaves/aboveground biomass ratio, and the RE concentrations in leaves. Clonal Salix smithiana was the most productive one in the SH, with a dry matter yield of 15.1 t ha-1 year-1 and showed promising extraction ability for Cd (11.65 %) and Zn (4.59 %) over a 6-year field experiment. A lower portion of Cd (6.97 %) and Zn (2.38 %) was removed by this clone in the WH (calculated from the total soil content of REs). SH was more reasonable for phytoextraction compared to WH. Higher RE concentrations were found in leaves of Salix compared to Populus. Populus accumulated the highest Pb content in the branches; unfortunately, the Pb extraction was low, due to extremely high soil Pb contamination. Locally bred willows and poplars performed substantially better than internationally recognised ones, indicating the importance of phytoremediation optimisation, including clone selection, for suitable climatic conditions.

Variability of Constitutive Stilbenoid Levels and Profiles in Grape Cane (Vitis vinifera L.) Depending upon Variety and Clone, Location in the Vineyard, Pruning Time, and Vintage.

Stilbenoids in grape cane (Vitis vinifera L.) are bioactive compounds relevant for plant defense and the potential valorization of this byproduct. Our screening of grape cane from 102 varieties showed constitutive stilbenoid levels in a wide range (557-7748 mg/kg of dry weight). Analyses of genetically distinct clones of selected cultivars unraveled that intravarietal variability (e.g., cv. Riesling, 3236-6541 mg/kg) was higher than that across samples from a single clone but different vineyard positions (3017-3710 mg/kg). Furthermore, stilbenoid levels in samples obtained in October, December, and February (3 years, 2017-2019) showed pronounced quantitative and qualitative variability and the highest yields upon December pruning. For instance, vitisin B and ε-viniferin in cv. Pinot Noir and Accent were predominant in 2017 and 2019 (both >90% of total stilbenoids) but not in 2018 (both <55%) when temperatures were high and precipitation low. In brief, we report the variability of stilbenoid levels in grape cane depending upon genetic and environmental factors.

C3 glomerulopathy associated with monoclonal gammopathy: impact of chronic histologic lesions and beneficial effects of clone-targeted therapies.

BACKGROUND: C3 glomerulopathy associated with monoclonal gammopathy (C3G-MIg) is a rare entity. Herein we analysed the clinical and histologic features of a cohort of C3G-MIg patients. METHODS: We conducted a retrospective, multicentre, observational study. Patients diagnosed with C3G-MIg between 1995 and 2021 were enrolled. All had genetic studies of the alternative complement pathway. The degree of disease activity and chronicity were analysed using the C3G histologic index. Descriptive statistics and propensity score matching (PSM) analysis were used to evaluate the main outcome of the study [kidney failure (KF)]. RESULTS: The study group included 23 patients with a median age 63 of years [interquartile range (IQR) 48-70], and 57% were males. Immunoglobulin G kappa was the most frequent MIg (65%). The diagnosis of C3G-MIg was made in transplanted kidneys in seven patients (30%). Five (22%) patients had C3 nephritic factor and five (22%) had anti-factor H antibodies. One patient carried a pathogenic variant in the CFH gene. During a follow-up of 40 months (IQR 14-69), nine patients (39%) reached KF and these patients had a significantly higher total chronicity score on kidney biopsy. Patients who received clone-targeted therapy had a significantly higher survival compared with other management. Those who achieved haematological response had a significantly higher kidney survival. Outcome was remarkably poor in kidney transplant recipients, with five of them (71%) reaching KF. By PSM (adjusting for age, kidney function, proteinuria and chronicity score), no significant differences were observed in kidney survival between C3G patients with/without MIg. CONCLUSIONS: The C3G histologic index can be used in patients with C3G-MIg to predict kidney prognosis, with higher chronicity scores being associated with worse outcomes. Clone-targeted therapies and the development of a haematological response are associated with better kidney prognosis.

Pyrrolizidine alkaloids cause cell cycle and DNA damage repair defects as analyzed by transcriptomics in cytochrome P450 3A4-overexpressing HepG2 clone 9 cells.

Pyrrolizidine alkaloids (PAs) are a large group of highly toxic chemical compounds, which are found as cross-contaminants in numerous food products (e.g., honey), dietary supplements, herbal teas, and pharmaceutical herbal medicines. PA contaminations are responsible for serious hepatotoxicity and hepatocarcinogenesis. Health authorities have to set legal limit values to guarantee the safe consumption of plant-based nutritional and medical products without harmful health. Toxicological and chemical analytical methods are conventionally applied to determine legally permitted limit values for PAs. In the present investigation, we applied a highly sensitive transcriptomic approach to investigate the effect of low concentrations of five PAs (lasiocarpine, riddelliine, lycopsamine, echimidine, and monocrotaline) on human cytochrome P450 3A4-overexpressing HepG2 clone 9 hepatocytes. The transcriptomic profiling of deregulated gene expression indicated that the PAs disrupted important signaling pathways related to cell cycle regulation and DNA damage repair in the transfected hepatocytes, which may explain the carcinogenic PA effects. As PAs affected the expression of genes that involved in cell cycle regulation, we applied flow cytometric cell cycle analyses to verify the transcriptomic data. Interestingly, PA treatment led to an arrest in the S phase of the cell cycle, and this effect was more pronounced with more toxic PAs (i.e., lasiocarpine and riddelliine) than with the less toxic monocrotaline. Using immunofluorescence, high fractions of cells were detected with chromosome congression defects upon PA treatment, indicating mitotic failure. In conclusion, the tested PAs revealed threshold concentrations, above which crucial signaling pathways were deregulated resulting in cell damage and carcinogenesis. Cell cycle arrest and DNA damage repair point to the mutagenicity of PAs. The disturbance of chromosome congression is a novel mechanism of Pas, which may also contribute to PA-mediated carcinogenesis. Transcriptomic, cell cycle, and immunofluorescence analyses should supplement the standard techniques in toxicology to unravel the biological effects of PA exposure in liver cells as the primary target during metabolization of PAs.

Clinical utility of chromosomal microarray in establishing clonality and high risk features in patients with Richter transformation.

Richter transformation (RT) refers to the development of an aggressive lymphoma in patients with pre-existing chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL). It carries a poor prognosis secondary to poor response to therapy or rapid disease relapse. Currently there are no randomized trials to guide treatment. Therapeutic decisions are often influenced by the presence or absence of a clonal relationship between the underlying CLL/SLL and the new lymphoma given the poor prognosis of patients with clonally related RT. Chromosomal microarray analysis (CMA) can help to establish clonality while also detecting genomic complexity and clinically relevant genetic variants such as loss of CDKN2A and/or TP53. As a result, CMA has potential prognostic and therapeutic implications. For this study, CMA results from patients with Richter transformation were evaluated in paired CLL/SLL and transformed lymphoma samples. CMA revealed that 86% of patients had common aberrations in the two samples indicating evidence of common clonality. CMA was also useful in detecting aberrations associated with a poor prognosis in 71% of patients with RT. This study highlights the potential clinical utility of CMA to investigate the clonal relationship between CLL/SLL and RT, provide prognostic information, and possibly guide therapeutic decision making for patients with Richter transformation.

Phytoremediation potential evaluation of multiple Salix clones for heavy metals (Cd, Zn and Pb) in flooded soils.

Climate-induced flooding makes soil more vulnerable to heavy metal contamination, posing challenges for soil remediation. Salix has the potential to cope with flooding stress and environmental contamination, but its effectiveness in flooded soils with multiple heavy metals has not yet been well assessed. Thus, the present work tested fifteen Salix clones grown in multimetal (Cd, Zn and Pb) contaminated soils under non-flooded versus flooded conditions. The results indicated that all tested Salix clones withstood long-term (90 d) flooding. Compared to the non-flooded condition, the flooded condition reduced the Cd (11.7-90.1%) contents in all organs but dramatically increased the Zn and Pb contents in the roots. The bioconcentration factor values of heavy metals in the aboveground organs were in the order of Cd > Zn > Pb. The tested Salix clones were characterized by high phytoextraction capacity for Cd and Zn under non-flooded condition and phytostabilization trait for Pb under flooded condition. To assess the overall performance of phytoremediation potentials, we attempted to use an analytic hierarchy process-entropy weight (AHP-EW) model, which considered the growth performance, photosynthetic parameters, accumulation, and mobility of toxic metals. Three Salix clones (J1010, P54 and P667) exhibited significant potential for multimetal remediation capacities. The current study provided valuable insights into the phytomanagement of woody plants, and the AHP-EW model is helpful for screening suitable trees for the phytoremediation of heavily multimetal contaminated wetlands.

Evaluation of some nutritional quality criteria of seventeen Moroccan dates varieties and clones, fruits of date palm (Phoenix dactylifera L. ) / Avaliação de alguns critérios de qualidade nutricional de 17 variedades e clones marroquinos de datas, frutos de tamareira (Phoenix dactylifera L. )

Date fruit is known to be the staple food in the Arab countries. It provides a lot of potential health benefits and can be the essential source of nutrients. The majority of Moroccan varieties are not characterized for their chemical, biochemical and quality properties. The aim of this work was to assess the chemical composition of 17 varieties of Moroccan date fruits (Phoenix dactylifera L.) and to determine their nutritive components. The analysis showed that the dates are rich in sugars (51.80-87.98%), they contain low concentration of proteins (1.09-2.80%) and lipids (0.16-0.39%). The predominant mineral is potassium (1055.26-1604.10 mg/100 g DW). Moreover, they contain high concentrations of malic acid (69.48-495.58 mg/100 g (DW)), oxalic acid (18.47-233.35 mg/100 g DW) and tartaric acid (115.70-484.168 mg/100 g DW). These results suggest that the date fruit are nutritious and can be an excellent source for human nutrition and health benefits.

A fruta da tâmara é conhecida por ser o alimento básico nos países árabes. Oferece muitos benefícios potenciais à saúde e pode ser a fonte essencial de nutrientes. A maioria das variedades marroquinas não se caracteriza por suas propriedades químicas, bioquímicas nem de qualidade. O objetivo deste trabalho foi avaliar a composição química de 17 variedades de frutos de tâmara marroquina (Phoenix dactylifera L.) e determinar seu valor nutritivo. A análise mostrou que as tâmaras são ricas em açúcares (51,80-87,98%) e contêm baixa concentração de proteínas (1,09-2,80%) e lipídios (0,16-0,39%). O mineral predominante é o potássio (1.055,26-1.604,10 mg/100 g DW). Além disso, contêm altas concentrações de ácido málico (69,48-495,58 mg/100 g DW), ácido oxálico (18,47-233,35 mg/100 g DW) e ácido tartárico (115,70-484,168 mg/100 g DW). Esses resultados sugerem que o fruto da tamareira é nutritivo e pode ser uma excelente fonte de nutrição humana e conferir benefícios à saúde.

Evaluation of some nutritional quality criteria of seventeen Moroccan dates varieties and clones, fruits of date palm (Phoenix dactylifera L.).

Date fruit is known to be the staple food in the Arab countries. It provides a lot of potential health benefits and can be the essential source of nutrients. The majority of Moroccan varieties are not characterized for their chemical, biochemical and quality properties. The aim of this work was to assess the chemical composition of 17 varieties of Moroccan date fruits (Phoenix dactylifera L.) and to determine their nutritive components. The analysis showed that the dates are rich in sugars (51.80-87.98%), they contain low concentration of proteins (1.09-2.80%) and lipids (0.16-0.39%). The predominant mineral is potassium (1055.26-1604.10 mg/100 g DW). Moreover, they contain high concentrations of malic acid (69.48-495.58 mg/100 g (DW)), oxalic acid (18.47-233.35 mg/100 g DW) and tartaric acid (115.70-484.168 mg/100 g DW). These results suggest that the date fruit are nutritious and can be an excellent source for human nutrition and health benefits.

Long-term effects of phytoextraction by a poplar clone on the concentration, fractionation, and transportation of heavy metals in mine tailings.

Mine tailings are one of main causes of diffused heavy metal pollution since the heavy metals in there may acquire mobility. The current knowledge of the processes at work in long-term phytoremediation by woody species remains insufficient. Through a 4-year field study, we evaluated the phytoextraction efficiency of Populus deltoides CL. 'Xianglin 90' grown on a mine tailing co-polluted by Cd, Cu, Cr, Ni, Pb, and Zn. The concentrations of Cd, Cu, Ni, Pb, and Zn in the rhizospheric soil were reduced by amounts ranging from 12.86 to 42.19% during the study period. Bioconcentration factors and translocation factors showed that the accumulation of Cd and Zn occurring in the shoots was the most effective. Combined with the considerable biomass produced by poplar, the extracted amounts of Cd and Zn could reach 0.61 g and 10.66 g plant-1, respectively, in which the shoots account for 77.3% (Cd) and 89.0% (Zn) of the overall extraction amounts. Acid-soluble Cd and Zn increased by 5.49% and 4.29%, respectively, in the rhizosphere compared to the bulk soil, indicating that poplar enhanced the mobility of Cd and Zn in the rhizosphere, which explained its ability for bioaccumulation and root-shoot translocation. Moreover, calculated time required to address the issue of Cd and Zn pollution was theoretically shortened by more than half from 2015 to 2019. This study brings new insights into the long-term effects of phytoextraction on the concentration, fractionation, and transportation of heavy metals and confirms the potential of poplar as a Cd and Zn remediation species.

Reprogramming enriches for somatic cell clones with small-scale mutations in cancer-associated genes.

Cellular therapies based on induced pluripotent stem cells (iPSCs) come out of age and an increasing number of clinical trials applying iPSC-based transplants are ongoing or in preparation. Recent studies, however, demonstrated a high number of small-scale mutations in iPSCs. Although the mutational load in iPSCs seems to be largely derived from their parental cells, it is still unknown whether reprogramming may enrich for individual mutations that could lead to loss of functionality and tumor formation from iPSC derivatives. 30 hiPSC lines were analyzed by whole exome sequencing. High accuracy amplicon sequencing showed that all analyzed small-scale variants pre-existed in their parental cells and that individual mutations present in small subpopulations of parental cells become enriched among hiPSC clones during reprogramming. Among those, putatively actionable driver mutations affect genes related to cell-cycle control, cell death, and pluripotency and may confer a selective advantage during reprogramming. Finally, a short hairpin RNA (shRNA)-based experimental approach was applied to provide additional evidence for the individual impact of such genes on the reprogramming efficiency. In conclusion, we show that enriched mutations in curated onco- and tumor suppressor genes may account for an increased tumor risk and impact the clinical value of patient-derived hiPSCs.

Fluctuation analysis to select for Samarium bio-uptaking microalgae clones the repurposing of a classical evolution experiment.

Rare Earth Elements (REE) increasing demand prompts the research of biotechnological approaches to exploit secondary resources. We made use of the adapted Fluctuation analyses experiment to obtain Chlamydomonas reinhardtii ChlA strains resistant to Samarium (Sm) as the reference REE. The starting hypothesis was that adaptation to metal-containing media leads to an enhanced metal uptake. ChlA was able to adapt to 1.33·10-4 Sm M and pH~3 by pre-existing genetic variability, allowing the evolutionary rescue of 13 of the 99 populations studied. The rescuing resistant genotypes presented a mutation rate of 8.65·10-7 resistant cells per division. The resulting resistant population contradicted the expected fitness cost associated with the adaptation to Sm, selection resulted in larger and faster-growing resistant cells. Among the three isolated strains studied for Sm uptake, only one presented uplifted performance compared to the control population (46.64 µg Sm g-¹ of wet biomass and 3.26·10-7 ng Sm per cell, mainly bioaccumulated within the cells). The selection of microalgae strains with improved tolerance to REEs by this methodology could be a promising solution for REES sequestration. However, increased tolerance can be independent or have negative effects on uptake performance and cellular features studied are not directly correlated with the metal uptake. SUMMARY SENTENCE: Repurposing a classic laboratory evolution experiment to select for microalgae Samarium adapted strains for metals recovery and biotechnology approaches. DATA AVAILABILITY STATEMENT: All data generated or analyzed during this study are included in this published article (and its raw files).

Extracting cadmium in the presence of salt: a study on three poplar clones under controlled conditions.

This study aimed at determining the cadmium phytoextraction potential of three Populus alba L. clones cultivated in the presence of increasing sodium chloride concentrations. Plantlets of a commercial and two autochthonous poplar clones were grown in perlite with nutrient solution enriched in CdSO4 (50 and 100 µM) and NaCl (25 and 50 mM), administered either alone or in combination. The three clones showed significant variation not only in cadmium and salt tolerance, accumulation and content, but also in the effect of the interaction between the two elements on these parameters. The toxic effect of Cd and salt excess on plants was mutually exacerbated by the presence of both. Even though the outcome of the joint treatment was always a decrease in shoot Cd or Na accumulation, the three clones showed variation in the extent of such reduction. Evaluating the total element content per plant shoot, the fast-growing commercial clone displayed the highest phytoextraction potential for Cd and Na, either alone or in mixture. Our results demonstrated for the first time that the Cd response in presence of salt can vary in the different clones.

Panax quinquefolium L. Ginsenosides from Hairy Root Cultures and Their Clones Exert Cytotoxic, Genotoxic and Pro-Apoptotic Activity towards Human Colon Adenocarcinoma Cell Line Caco-2.

American ginseng, Panax quinquefolium (L.), is traditionally used in folk medicine. It exhibits a range of anti-inflammatory, hepatoprotective, anti-diabetic, anti-obesity, anti-hyperlipidemic and anti-carcinogenic effects. Its main components are ginsenosides, also known as panaxosides or triterpene saponins. In order to obtain high yields of ginsenosides, different methods of controlled production are involved, i.e., with hairy root cultures. However, they are still employed under in vitro conditions. Our studies revealed that hairy root cultures subjected to an elicitation process can be considered as a potent source of ginsenosides. The present study examines the biological activity of ginseng hairy root cultures against the Caco-2 human adenocarcinoma cell line. Among our six different clones of P. quinquefolium hairy roots, extracts B and Be (treated with elicitor) were the strongest inhibitors of the cellular metabolic activity. While all extracts induced DNA damage, B and Be also generated reactive oxygen species (ROS) in a concentration-dependent manner, which was correlated with the depletion of the mitochondrial membrane potential and induction of apoptosis. These findings indicate that further research concerning P. quinquefolium hairy root cultures should focus on the activity of rare ginsenosides and other biologically active compound profiles (i.e., phenolic compounds).