Seed functional ecology in Brazilian rock outcrop vegetation: an integrative synthesis.

BACKGROUND AND AIMS: Rock outcrop vegetation is distributed worldwide and hosts a diverse and unique flora that evolved under harsh environmental conditions. Unfortunately, seed ecology in such ecosystems has received little attention, especially regarding seed traits, germination responses to abiotic factors and the potential role of phylogenetic relatedness on such features Here, we provide the first quantitative and phylogenetically-informed synthesis of the seed functional ecology of Brazilian rock outcrop vegetation, with a particular focus on quartzitic and ironstone campo rupestre. METHODS: Using a database of functional trait data, we calculated the phylogenetic signal of seven seed traits for 371 taxa and tested whether they varied among growth forms, geographic distribution, and microhabitats. We also conducted meta-analyses that included 4,252 germination records for 102 taxa to assess the effects of light, temperature, and fire-related cues on the germination of campo rupestre species and explored how the aforementioned ecological groups and seed traits modulate germination responses. KEY RESULTS: All traits and germination responses showed a moderate-to-strong phylogenetic signal. Campo rupestre species responded positively to light and had maximum germination between 20-25 ºC. The effect of temperatures beyond this range was moderated by growth form, species geographic distribution, and microhabitat. Seeds exposed to heat shocks above 80 °C lost viability, but smoke accelerated germination. We found a moderating effect of seed mass for in responses to light and heat shocks, with larger, dormant seeds tolerating heat better but less sensitive to light. Species from xeric habitats evolved phenological strategies to synchronise germination during periods of increased soil water availability. CONCLUSIONS: Phylogenetic relatedness plays a major role in shaping seed ecology of Brazilian rock outcrop vegetation. Nevertheless, seed traits and germination responses varied significantly between growth forms, species geographic distribution and microhabitats, providing support to the regeneration niche hypothesis and the role of functional traits in shaping germination in these ecosystems.

Encapsulation of Saccharomyces spp. for Use as Probiotic in Food and Feed: Systematic Review and Meta-analysis.

Probiotics, particularly yeasts from the genus Saccharomyces, are valuable for their health benefits and potential as antibiotic alternatives. To be effective, these microorganisms must withstand harsh environmental conditions, necessitating advanced protective technologies such as encapsulation to maintain probiotic viability during processing, storage, and passage through the digestive system. This review and meta-analysis aims to describe and compare methods and agents used for encapsulating Saccharomyces spp., examining operating conditions, yeast origins, and species. It provides an overview of the literature on the health benefits of nutritional yeast consumption. A bibliographic survey was conducted following the Preferred Reporting Items for Systematic Review and Meta-Analysis (PRISMA) guidelines. The meta-analysis compared encapsulation methods regarding their viability after encapsulation and exposure to the gastrointestinal tract. Nineteen studies were selected after applying inclusion/exclusion criteria. Freeze drying was found to be the most efficient for cell survival, while ionic gelation was best for maintaining viability after exposure to the gastrointestinal tract. Consequently, the combination of freeze drying and ionic gelation proved most effective in maintaining high cell viability during encapsulation, storage, and consumption. Research on probiotics for human food and animal feed indicates that combining freeze drying and ionic gelation effectively protects Saccharomyces spp.; however, industrial scalability must be considered. Reports on yeast encapsulation using agro-industrial residues as encapsulants offer promising strategies for preserving potential probiotic yeasts, contributing to the environmental sustainability of industrial processes.

Chondroitin Sulfate for Cartilage Regeneration, Administered Topically Using a Nanostructured Formulation.

In the pharmaceutical sector, solid lipid nanoparticles (SLN) are vital for drug delivery incorporating a lipid core. Chondroitin sulfate (CHON) is crucial for cartilage health. It is often used in osteoarthritis (OA) treatment. Due to conflicting results from clinical trials on CHON's efficacy in OA treatment, there has been a shift toward exploring effective topical systems utilizing nanotechnology. This study aimed to optimize a solid lipid nanoparticle formulation aiming to enhance CHON permeation for OA therapy. A 3 × 3 × 2 Design of these experiments determined the ideal parameters: a CHON concentration of 0.4 mg/mL, operating at 20,000 rpm speed, and processing for 10 min for SLN production. Transmission electron microscopy analysis confirmed the nanoparticles' spherical morphology, ensuring crucial uniformity for efficient drug delivery. Cell viability assessments showed no significant cytotoxicity within the tested parameters, indicating a safe profile for potential clinical application. The cell internalization assay indicates successful internalization at 1.5 h and 24 h post-treatment. Biopharmaceutical studies supported SLNs, indicating them to be effective CHON carriers through the skin, showcasing improved skin permeation and CHON retention compared to conventional methods. In summary, this study successfully optimized SLN formulation for efficient CHON transport through pig ear skin with no cellular toxicity, highlighting SLNs' potential as promising carriers to enhance CHON delivery in OA treatment and advance nanotechnology-based therapeutic strategies in pharmaceutical formulations.

Effect of iontophoresis on dacarbazine cutaneous delivery for melanoma topical treatment.

Dacarbazine (DTIC) is the drug of choice for melanoma treatment, but its systemic administration is related to several adverse effects. Here, DTIC topical delivery stimulated by iontophoresis is proposed to overcome such drawbacks. Hence, this work analyzed the impact of anodal iontophoresis on DTIC cutaneous delivery to provide an innovative topical alternative for melanoma treatment. The electrical stability of the drug was evaluated prior to the iontophoretic experiments, which demonstrated the need to add an antioxidant to the drug formulation. DTIC cutaneous permeation was evaluated in vitro for 6 h using three current densities (0.10, 0.25, and 0.50 mA/cm2). In addition, the effect of DTIC against skin cancer cells (MeWo and WM164) was investigated for 72 h of exposure to the drug. Iontophoresis stimulated skin drug permeation compared to the passive control. However, the antioxidant presence reduced DTIC permeation under the lower currents of 0.10 and 0.25 mA/cm2, which was compensated by increasing the current density to 0.50 mA/cm2. At 0.50 mA/cm2, iontophoresis enhanced topical cutaneous drug permeation 7-fold (p < 0.05) compared to the passive control. DTIC showed a concentration-dependent antiproliferative effect on melanoma cell lines. Thus, iontophoresis intensifies DTIC skin penetration in concentrations that can reduce cell viability and induce cell death. In conclusion, DTIC cutaneous delivery mediated by iontophoresis is a promising approach for treating melanomas and other skin tumors.

In-vitro study on type I collagen synthesis in low-level laser therapy on the early ligament fibroblasts' healing process.

BACKGROUND: Low-level Laser Therapy (LLLT) has demonstrated its potential in promoting fiber matrix maturation, collagen synthesis, and fibroblast proliferation, contributing to tissue regeneration. Our study aimed to investigate the impact of LLLT on collagen type I synthesis, cell proliferation, and viability in human ligament fibroblasts derived from the Anterior Cruciate Ligament (ACL). METHODS: Tissue samples were obtained from individuals undergoing arthroscopic ACL reconstruction surgery. Primary human fibroblasts were isolated, and immunohistochemical assays confirmed their characteristics. LLLT at 850 nm was administered in three groups: Low dose (1.0 J/cm²), High dose (5.0 J/cm²), and Control (0.0 J/cm²). Cell viability was calculated using a membrane integrity assay, proliferation was determined by automated counting, and collagen type I concentration in cell culture was measured using an immunoassay. RESULTS: Fibroblasts showed decreased viability after low and high doses of LLLT, increased proliferation at the low dose, and increased collagen synthesis at the high dose on day 10 for both sexes after treatment. CONCLUSION: Our study demonstrated that LLLT may improve the early ligament healing process by increasing cell proliferation at the low dose and enhancing collagen type I synthesis at the high dose in human ligament fibroblasts.

Behavioral dynamics of medicinal signaling cells from porcine bone marrow in long-term culture.

Medicinal signaling cells (MSC) hold promise for regenerative medicine due to their ability to repair damaged tissues. However, their effectiveness can be affected by how long they are cultured in the lab. This study investigated how passage number influences key properties for regenerative medicine of pig bone marrow MSC. The medicinal signiling cells derived from pig bone marrow (BM-MSC) were cultured in D-MEM High Glucose supplemented with 15% foetal bovine serum until the 25th passage and assessed their growth, viability, ability to differentiate into different cell types (plasticity), and cell cycle activity. Our findings showed that while the cells remained viable until the 25th passage, their ability to grow and differentiate declined after the 5th passage. Additionally, cells in later passages spent more time in a resting phase, suggesting reduced activity. In conclusion, the number of passages is a critical factor for maintaining ideal MSC characteristics. From the 9th passage BM-MSC exhibit decline in proliferation, differentiation potential, and cell cycle activity. Given this, it is possible to suggest that the use of 5th passage cells is the most suitable for therapeutic applications.

Shotgun proteomics profiling of chia seeds (Salvia hispanica L.) reveals genotypic differential responses to viability loss.

Introduction: Exposure to elevated temperatures and relative humidity expedites the seed aging process, finally leading to seed viability loss. In this context, certain proteins play a pivotal role in safeguarding the longevity of seeds. However, the seedproteomic response to loss viability in Salvia hispanica L., commonly known as chia, remains incompletely understood. Methods: This work explores the application of proteomics as a potent tool for uncovering molecular responses to viability loss caused by artificial aging in two chia genotypes, WN and MN. Results: By using a quantitative label-free proteomics analysis (LC-MS/MS), 1787 proteins wereidentified in chia seeds at a 95% confidence level, including storage proteins, heat shock proteins (HSPs), late embryogenesis abundant proteins (LEA),oleosins, reactive oxygen species (ROS)-related enzymes, and ribosomal proteins. A relatively low percentage of exclusive proteins were identified in viable and non-viable seeds. However, proteins exhibiting differential abundancebetween samples indicated variations in the genotype and physiological status. Specifically, the WN genotype showed 130 proteins with differential abundancecomparing viable and non-viable seeds, while MN displayed changes in the abundance of 174 proteins. While both showed a significant decrease in keyproteins responsible for maintaining seed functionality, longevity, and vigor withhigh-temperature and humidity conditions, such as LEA proteins or HSPs, ROS, and oleosins, distinct responses between genotypes were noted, particularly in ribosomal proteins that were accumulated in MN and diminished in WN seeds. Discussion: Overall, the results emphasize the importance of evaluating changes in proteins of viable and non-viable seeds as they offer valuable insights into the underlying biological mechanisms responsible for the maintenance of chia seed integrity throughout high-temperature and humidity exposure.

Impact of Heat Stress on Oocyte Developmental Competence and Pre-Implantation Embryo Viability in Cattle.

Rectal and vaginal temperatures are utilised in both in vivo and in vitro models to study the effects of heat stress on oocyte competence and embryo viability in cattle. However, uterine temperature increases by only 0.5 °C in heat-stressed cows, significantly lower than simulated increases in in vitro models. Temperature variations within oviducts and ovarian follicles during heat stress are poorly understood or unavailable, and evidence is lacking that oocytes and pre-implantation embryos experience mild (40 °C) or severe (41 °C) heat stress inside the ovarian follicle and the oviduct and uterus, respectively. Gathering detailed temperature data from the reproductive tract and follicles is crucial to accurately assess oocyte competence and embryo viability under realistic heat stress conditions. Potential harm from heat stress on oocytes and embryos may result from reduced nutrient availability (e.g., diminished blood flow to the reproductive tract) or other unidentified mechanisms affecting tissue function rather than direct thermal effects. Refining in vivo stress models in cattle is essential to accurately identify animals truly experiencing heat stress, rather than assuming heat stress exposure as done in most studies. This will improve model reliability and aid in the selection of heat-tolerant animals.

Effects and action mechanism of gonadotropins on ovarian follicular cells: A novel role of Sphingosine-1-Phosphate (S1P). A review.

Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) control antral follicular growth by regulating several processes, such as the synthesis of hormones and signaling molecules, proliferation, survival, apoptosis, luteinization, and ovulation. To exert these effects, gonadotropins bind to their respective Gs protein-coupled receptors, activating the protein kinase A (PKA) pathway or recruiting Gq proteins to activate protein kinase C (PKC) signaling. Although the action mechanism of FSH and LH is clear, recently, it has been shown that both gonadotropins promote the synthesis of sphingosine-1-phosphate (S1P) in granulosa and theca cells through the activation of sphingosine kinase 1. Moreover, the inhibition of SPHKs reduces S1P synthesis, cell viability, and the proliferation of follicular cells in response to gonadotropins, and the addition of S1P to the culture medium increases the proliferation of granulosa and theca cells without apparent effects on sexual steroid synthesis. Therefore, we consider that S1P is a crucial signaling molecule that complements the canonical gonadotropin pathway to promote the proliferation and viability of granulosa and theca cells.

Ascorbic acid and ascorbyl palmitate-loaded liposomes: Development, characterization, stability evaluation, in vitro security profile, antimicrobial and antioxidant activities.

The objective of this work was to prepare and characterize liposomes containing co-encapsulated ascorbic acid (AA) and ascorbyl palmitate (AP), as well as to evaluate their stability, cytotoxicity, antioxidant, and antimicrobial activity. Through the pre-formulation studies, it was possible to improve the formulation, as leaving it more stable and with a greater antioxidant activity, resulting in a formulation designated LIP-AAP, with 161 nm vesicle size, 0.215 polydispersity index, -31.7 mV zeta potential, and pH of 3.34. Encapsulation efficiencies were 37% for AA and 79% for AP, and the content was 1 mg/mL for each compound. The optimized liposomes demonstrated stability under refrigeration for 60 days, significant antioxidant activity (31.4 µMol of TE/mL), and non-toxicity, but no antimicrobial effects against bacteria and fungi were observed. These findings confirm that the co-encapsulated liposomes are potent, stable antioxidants that maintain their physical and chemical properties under optimal storage conditions.

Impact of CAD/CAM burs lifetime on surface properties of dental zirconia: Implications for biocompatibility of custom abutments.

OBJECTIVE: To evaluate the effect of the deterioration of computer aided design/computer aided manufacturing (CAD/CAM) burs during zirconia milling, on surface roughness, contact angle, and fibroblast viability. MATERIALS AND METHODS: Ceramic blocks were milled and 75 ceramic disks (8 × 1.5 mm) made and allocated into three groups (n = 25): G1-brand new 2L and 1L burs, G2-2L bur at the end of lifetime and brand new 1L bur and G3-both burs at the end of their lifetimes. Roughness (Ra, Rq, and Rz) was evaluated using a 3D optical profilometer, the contact angle by the sessile drop method and the cell viability of the mouse NIH/3T3 fibroblast, using the Alamar Blue assay at intervals of 24, 48, and 72 h (ISO 10993-5). Data were analyzed by one-way ANOVA and Kruskal-Wallis tests (p ≤ 0.05). RESULTS: Roughness increased as the burs deteriorated and G3 (0.27 ± 0.04) presented a higher value for Ra (p < 0.001). The highest contact angle was observed in G3 (86.2 ± 2.66) when compared with G1 (63.7 ± 12.49) and G2 (75.3 ± 6.36) (p < 0.001). Alamar Blue indicated an increase in cell proliferation, with no significant differences among the groups at 24 and 72 h (p > 0.05). CONCLUSIONS: The deterioration of the burs increased the surface roughness and decreased the wettability, but did not interfere in cell viability and proliferation. CLINICAL SIGNIFICANCE: The use of custom zirconia abutments represents an effective strategy for single crowns restorations. Our findings suggest that these abutments can be efficiently milled using CAD/CAM burs within their recommended lifetime.

Efficiency of the photodynamic therapy on viability of Streptococcus mutans in the oral cavity using chitosan nanoparticles: an in vitro study.

PURPOSE: This study aimed to investigate the efficiency of antimicrobial photodynamic therapy (aPDT) on Streptococcus mutans biofilm in the oral cavity using the photosensitizer chloroaluminum phthalocyanine encapsulated in chitosan nanoparticles (ClAlPc/Ch) at three preirradiation times. METHODS: Biofilms of Streptococcus mutans strains (ATCC 25,175) were cultivated on bovine tooth blocks and exposed to a 10% sucrose solution three times a day for 1 min over three consecutive days. The samples were randomly distributed into five treatment groups (n = 5): (I) aPDT with ClAlPc/Ch with a preirradiation time of 5 min (F5), (II) aPDT with ClAlPc/Ch with a preirradiation time of 15 min (F15), (III) aPDT with ClAlPc/Ch with a preirradiation time of 30 min (F30), (IV) 0.12% chlorhexidine digluconate (CHX), and (V) 0.9% saline solution (NaCl). After treatment, the S. mutans biofilms formed on each specimen were collected to determine the number of viable bacteria (colony-forming units (CFU)/mL). Data were analyzed for normality using the Shapiro-Wilk test and the analysis of variance (ANOVA) and Tukey HSD tests to analyze the number of viable bacteria (α = 0.05). RESULTS: The one-way ANOVA showed a difference between the groups (p = 0.0003), and the Tukey HSD posttest showed that CHX had the highest microbial reduction of S. mutans, not statistically different from the F5 and F15 groups, whereas the NaCl group had the lowest microbial reduction statistically similar to the F30 group. CONCLUSION: The results demonstrate that aPDT mediated by ClAlPc/Ch when used at preirradiation times of 5-15 min can be an effective approach in controlling cariogenic biofilm of S. mutans, being an alternative to 0.12% CHX.

Transverse and vertical incisions affect the viability of in vitro-produced embryos submitted to a simplified microsurgery approach.

Integrating in vitro embryo production with embryonic microsurgery facilitates the generation of monozygotic twins. However, despite their potential benefits, these methods have not been widely adopted in commercial settings because of their substantial costs. Hence, there is a need to streamline the bisection procedure while ensuring efficient production of viable demi-embryos. In this study, we investigated the impact of different orientations of microsurgical incisions in relation to inner cell mass on embryonic development, morphology, viability, and expression of cell fate protein markers using a simplified microsurgery approach. Ovaries were transported from the slaughterhouse to the laboratory and aspirated to obtain oocytes that were selected and subjected to in vitro embryo production. The selected expanded blastocysts (n = 204) underwent microsurgery. The blastocysts were immobilized to facilitate incision using an adapted microblade, yielding demi-embryos (vertical incision) and viable embryonic fragments (transverse incision). The structures were then re-cultured for 12 h. Viability was assessed by measuring the re-expansion rate after re-culture, followed by immunofluorescence analysis of proteins (CDX2 and NANOG) and apoptosis analysis using terminal deoxynucleotyl transferase dUTP nick end-labeling (TUNEL). Microsurgically derived embryos exhibited remarkable plasticity, as evidenced by a slight reduction (P < 0.05) in the re-expansion rate (transverse 64.2 % and vertical 57.2 %) compared to that of the control group (blastocysts without microsurgery) (86.7 %). They also demonstrated the ability of morphological reconstitution after culturing. Despite the anticipated decrease (P < 0.05) in the total number of cells and embryo volume, microsurgery did not result in a significant increase (P > 0.05) in the number of apoptotic cells. Furthermore, microsurgery led to higher (P < 0.05) expression of markers associated with pluripotency, indicating its efficiency in preserving regenerative capacity. Moreover, microsurgery, whether followed by immunosurgery or not, made the isolation of embryonic cells easier. In conclusion, both transverse and vertical microsurgery incisions enabled the production of identical demi-embryos and served as tools for isolating embryonic cells without compromising the resumption of development and the apoptotic index.

Adenosine A2A receptor as a potential regulator of Mycobacterium leprae survival mechanisms: new insights into leprosy neural damage.

Background: Leprosy is a chronic infectious disease caused by Mycobacterium leprae, which can lead to a disabling neurodegenerative condition. M. leprae preferentially infects skin macrophages and Schwann cells-glial cells of the peripheral nervous system. The infection modifies the host cell lipid metabolism, subverting it in favor of the formation of cholesterol-rich lipid droplets (LD) that are essential for bacterial survival. Although researchers have made progress in understanding leprosy pathogenesis, many aspects of the molecular and cellular mechanisms of host-pathogen interaction still require clarification. The purinergic system utilizes extracellular ATP and adenosine as critical signaling molecules and plays several roles in pathophysiological processes. Furthermore, nucleoside surface receptors such as the adenosine receptor A2AR involved in neuroimmune response, lipid metabolism, and neuron-glia interaction are targets for the treatment of different diseases. Despite the importance of this system, nothing has been described about its role in leprosy, particularly adenosinergic signaling (AdoS) during M. leprae-Schwann cell interaction. Methods: M. leprae was purified from the hind footpad of athymic nu/nu mice. ST88-14 human cells were infected with M. leprae in the presence or absence of specific agonists or antagonists of AdoS. Enzymatic activity assays, fluorescence microscopy, Western blotting, and RT-qPCR analysis were performed. M. leprae viability was investigated by RT-qPCR, and cytokines were evaluated by enzyme-linked immunosorbent assay. Results: We demonstrated that M. leprae-infected Schwann cells upregulated CD73 and ADA and downregulated A2AR expression and the phosphorylation of the transcription factor CREB (p-CREB). On the other hand, activation of A2AR with its selective agonist, CGS21680, resulted in: 1) reduced lipid droplets accumulation and pro-lipogenic gene expression; 2) reduced production of IL-6 and IL-8; 3) reduced intracellular M. leprae viability; 4) increased levels of p-CREB. Conclusion: These findings suggest the involvement of the AdoS in leprosy neuropathogenesis and support the idea that M. leprae, by downmodulating the expression and activity of A2AR in Schwann cells, decreases A2AR downstream signaling, contributing to the maintenance of LD accumulation and intracellular viability of the bacillus.

Flavonoids and their relationship with the physiological quality of seeds from different soybean genotypes.

Flavonoids are compounds that result from the secondary metabolism of plants and play a crucial role in plant development and mitigating biotic and abiotic stresses. The highest levels of flavonoids are found in legumes such as soybean. Breeding programs aim to increase desirable traits, such as higher flavonoid contents and vigorous seeds. Soybeans are one of the richest sources of protein in the plant kingdom and the main source of flavonoid derivatives for human health. In view of this, the hypothesis of this study is based on the possibility that the concentration of isoflavones in soybean seeds contributes to the physiological quality of the seeds. The aim of this study was to analyze the content of flavonoids in soybean genotypes and their influence on the physiological quality of the seeds. Seeds from thirty-two soybean genotypes were obtained by carrying out a field experiment during the 2021/22 crop season. The experimental design was randomized blocks with four replications and thirty-two F3 soybean populations. The seeds obtained were subjected to germination, first germination counting, electrical conductivity and tetrazolium vigor and viability tests. After drying and milling the material from each genotype, liquid chromatography analysis was carried out to obtain flavonoids, performed at UPLC level. Data were submitted to analysis of variance and, when significant, the means were compared using the Scott-Knott test at 5% probability. The results found here show the occurrence of genotypes with higher amounts of flavonoids when compared to their peers. The flavonoid FLVD_G2 had the highest concentration and differed from the others. Thus, we can assume that the type and concentration of flavonoids does not influence the physiological quality of seeds from different soybean genotypes, but it does indirectly contribute to viability and vigor, since the genotypes with the highest FLVD_G2 levels had better FGC values. The findings indicate that there is a difference between the content of flavonoids in soybean genotypes, with a higher content of genistein. The content of flavonoids does not influence the physiological quality of seeds, but contributes to increasing viability and vigor.

Adulterated dietary supplements commercialized in Brazil: development of a screening method and a preliminary study of cytotoxicity.

The high consumption of dietary supplements was a fundamental driver for the creation of the regulatory framework by the Brazilian governmental authorities. However, the regulatory agencies lack official low-cost methodologies to evaluate the quality of food supplements. A preliminary screening method by HPLC-DAD was proposed and validated for screening and quantification of adulterants in dietary supplements. The limits of detection and quantification were <0.11 and 0.37 µg.g-1, respectively. The method was applied for the investigation of ten unauthorized substances (spironolactone, hydrochlorothiazide, furosemide, clenbuterol, testosterone, testosterone propionate, yohimbine, vardenafil, tadalafil, and sildenafil) with a time of analysis of <5 min. Sixteen percent of the 44 samples analyzed had at least one adulterant at or above therapeutic concentrations. Subsequently, in vitro evaluations were performed of the potential cytotoxicity to evaluate the cell viability, DNA damage, determination of nitric oxide levels, and quantification of reactive oxygen species. Despite the necessity of further studies, the results indicate a relationship between the presence of adulterants in food supplements and a potential cytotoxic effect.

Reproductive biology and pollen-pistil compatibility relationships in an Argentinian collection of Stevia rebaudiana Bertoni / Biología reproductiva y relaciones de compatibilidad polen-pistilo en una colección de Stevia rebaudiana Bertoni en Argentina

ABSTRACT Stevia rebaudiana Bertoni (Asteraceae) is a diploid species (2n=2x=22) with sexual and asexual reproduction. The sexual propagules are seeds produced by cross-pollination (allogamy) whereas the asexual propagules are either vegetative shoots or apomictic seeds. Various authors have reported that allogamy in this species is promoted by the expression of a sporophytic self-incompatibility (SSI) system. To introduce the cultivation of S. rebaudiana as a production alternative in Tucumán, Argentina, a germplasm collection of this species was established with accessions from four Argentinian provinces in the Famaillá Agropecuarian Experimental Station (EEA Famaillá), National Institute of Agropecuarian Technology (INTA). The reproductive biology of the collection was studied between 2014 and 2021 to develop strategies for breeding and conservation of these genetic resources. Fifty-six genotypes were analyzed, all of them were 2n=2x=22. Pollen viability was high (69.4 to 99.6%) in all the genotypes except in four of them, which exhibited low viability (36.0 to 51.5%) in 2015 and 2017. Forty-eight genotypic combinations were obtained by manual controlled crosses. In 12 of these combinations, one pollen tube was observed in the style zone and, in four of them, one pollen tube was observed in the embryo sac; these observations indicate, respectively, incompatible and compatible pollen-pistil relationships. Normal plump seeds were obtained in all compatible genotypic combinations. The observed incompatibility might be due to the functioning of the sporophytic homomorphic system and/or a cross-incompatibility system. The observed compatibility will allow the planification of controlled crosses within and between accessions of different geographical origins to generate genetically variable progenies for breeding purposes.

RESUMEN Stevia rebaudiana Bertoni (Asteraceae) es una especie diploide (2n=2x=22) con reproducción sexual y asexual. Los propágulos sexuales son semillas producidas por polinización cruzada (alogamia), mientras que los propágulos asexuales son brotes vegetativos y semillas apomםcticas. Varios autores han seסalado que la alogamia en esta especie se ve favorecida por la expresión de un sistema de autoincompatibilidad esporofםtica (SSI). Para introducir el cultivo de S. rebaudiana como alternativa productiva en Tucumán, Argentina, se estableció una colección de germoplasma de esta especie proveniente de cuatro provincias de la Argentina en la Estación Experimental Agropecuaria (EEA) Famaillá, Instituto Nacional de Tecnologםa Agropecuaria (INTA). Se estudió la biología reproductiva de la colección entre 2014 y 2021 para desarrollar estrategias de mejoramiento y conservación de estos recursos genéticos. Se analizaron 56 genotipos, que fueron 2n=2x=22. La viabilidad del polen fue alta (69,4 a 99,6%) excepto en cuatro de ellos que exhibieron baja viabilidad (36,0 a 51,5%) en 2015 y 2017. Se obtuvieron 48 combinaciones genotópicas mediante cruzamientos controlados manuales. En 12 de estas combinaciones, se observó un tubo polםnico en la zona estilar y, en cuatro de ellas, un tubo polםnico en el saco embrionario; estas observaciones indican, respectivamente, relaciones polen-pistilo incompatibles y compatibles. Se obtuvieron semillas rellenas normales en todas las combinaciones genotópicas compatibles. La incompatibilidad observada podría deberse al funcionamiento del sistema de autoincompatibilidad homomórfica esporofítica, un sistema de incompatibilidad cruzada, o ambos. La compatibilidad observada permitirá la planificación de cruzamientos controlados dentro y entre introducciones de diferentes orígenes geográficos para generar progenies genéticamente variables con fines de mejoramiento genético.

Effect of Rapid High-Intensity Light-Curing on Increasing Transdentinal Temperature and Cell Viability: An In Vitro Study.

BACKGROUND: This study investigated effects of rapid high-intensity light-curing (3 s) on increasing transdentinal temperature and cell viability. METHODS: A total of 40 dentin discs (0.5 mm) obtained from human molars were prepared, included in artificial pulp chambers (4.5 × 5 mm), and subjected to four light-curing protocols (n = 5), with a Valo Grand light curing unit: (i) 10 s protocol with a moderate intensity of 1000 mW/cm2 (Valo-10 s); (ii) 3 s protocol with a high intensity of 3200 mW/cm2 (Valo-3 s); (iii) adhesive system + Filtek Bulk-Fill Flow bulk-fill composite resin in 10 s (FBF-10 s); (iv) adhesive system + Tetric PowerFlow bulk-fill composite resin in 3 s (TPF-3 s). Transdentinal temperature changes were recorded with a type K thermocouple. Cell viability was assessed using the MTT assay. Data were analyzed using one-way ANOVA and Tukey tests for comparison between experimental groups (p < 0.05). RESULTS: The 3 s high-intensity light-curing protocol generated a higher temperature than the 10 s moderate-intensity standard (p < 0.001). The Valo-10 s and Valo-3 s groups demonstrated greater cell viability than the FBF-10s and TPF-3 s groups and statistical differences were observed between the Valo-3 s and FBF-10 s groups (p = 0.023) and Valo-3 s and TPF-3 s (p = 0.025), with a potential cytotoxic effect for the FBF-10 s and TPF-3 s groups. CONCLUSIONS: The 3 s rapid high-intensity light-curing protocol of bulk-fill composite resins caused a temperature increase greater than 10 s and showed cell viability similar to and comparable to the standard protocol.

Supercritical Carbon Dioxide Extraction of Coumarins from the Aerial Parts of Pterocaulon polystachyum.

Pterocaulon polystachyum is a species of pharmacological interest for providing volatile and non-volatile extracts with antifungal and amebicidal properties. The biological activities of non-volatile extracts may be related to the presence of coumarins, a promising group of secondary metabolites. In the present study, leaves and inflorescences previously used for the extraction of essential oils instead of being disposed of were subjected to extraction with supercritical CO2 after pretreatment with microwaves. An experimental design was followed to seek the best extraction condition with the objective function being the maximum total extract. Pressure and temperature were statistically significant factors, and the optimal extraction condition was 240 bar, 60 °C, and pretreatment at 30 °C. The applied mathematical models showed good adherence to the experimental data. The extracts obtained by supercritical CO2 were analyzed and the presence of coumarins was confirmed. The extract investigated for cytotoxicity against bladder tumor cells (T24) exhibited significant reduction in cell viability at concentrations between 6 and 12 µg/mL. The introduction of green technology, supercritical extraction, in the exploration of P. polystachyum as a source of coumarins represents a paradigm shift with regard to previous studies carried out with this species, which used organic solvents. Furthermore, the concept of circular bioeconomy was applied, i.e., the raw material used was the residue of a steam-distillation process. Therefore, the approach used here is in line with the sustainable exploitation of native plants to obtain extracts rich in coumarins with cytotoxic potential against cancer cells.

Amantadine mitigates the cytotoxic and genotoxic effects of doxorubicin in SH-SY5Y cells and reduces its mutagenicity.

Amantadine (AMA) is a useful drug in neuronal disorders, but few studies have been performed to access its toxicological profile. Conversely, doxorubicin (Dox) is a well-known antineoplastic drug that has shown neurotoxic effects leading to cognitive impairment. The aims of this study are to evaluate the cytotoxic, genotoxic, and mutagenic effects of AMA, as well as its possible protective actions against deleterious effects of Dox. The Salmonella/microsome assay was performed to assess mutagenicity while cytotoxicity and genotoxicity were evaluated in SH-SY5Y cells using MTT and comet assays. Possible modulating effects of AMA on the cytotoxicity, genotoxicity, and mutagenicity induced by Dox were evaluated through cotreatment procedures. Amantadine did not induce mutations in the Salmonella/microsome assay and decreased Dox-induced mutagenicity in the TA98 strain. AMA reduced cell viability and induced DNA damage in SH-SY5Y cells. In cotreatment with Dox, AMA attenuated the cytotoxicity of Dox and showed an antigenotoxic effect. In conclusion, AMA does not induce gene mutations, although it has shown a genotoxic effect. Furthermore, AMA decreases frameshift mutations induced by Dox as well as the cytotoxic and genotoxic effects of Dox in SH-SY5Y cells, suggesting that AMA can interfere with Dox mutagenic activity and attenuate its neurotoxic effects.