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Therapeutic vaccines based on monocyte-derived dendritic cells have been shown to be promising strategies and may act as complementary treatments for viral infections, cancers, and, more recently, autoimmune diseases. Alpha-type-1-polarized dendritic cells (aDC1s) have been shown to induce type-1 immunity with a high capacity to produce interleukin-12p70 (IL-12p70). In the clinical use of cell-based therapeutics, injectable solutions can affect the morphology, immunophenotypic profile, and viability of cells before delivery and their survival after injection. In this sense, preparing a cell suspension that maintains the quality of aDC1s is essential to ensure effective immunotherapy. In the present study, monocytes were differentiated into aDC1s in the presence of IL-4 and GM-CSF. On day 5, the cells were matured by the addition of a cytokine cocktail consisting of IFN-α, IFN-γ, IL-1β, TNF-α, and Poly I:C. After 48 hr, mature aDC1s were harvested and suspended in two different solutions: normal saline and Ringer’s lactate. The maintenance of cells in suspension was evaluated after 4, 6, and 8 hr of storage. Cell viability, immunophenotyping, and apoptosis analyses were performed by flow cytometry. Cellular morphology was observed by electron microscopy, and the production of IL-12p70 by aDC1s was evaluated by ELISA. Compared with normal saline, Ringer’s lactate solution was more effective at maintaining DC viability for up to 8 hr of incubation at 4 or 22°C.

Abstract We investigated the anti-SARS-CoV-2 post-vaccine response through serum and salivary antibodies, serum antibody neutralizing activity and cellular immune response in samples from health care workers who were immunized with two doses of an inactivated virus-based vaccine (CoronaVac) who had or did not have COVID-19 previously. IgA and IgG antibodies directed at the spike protein were analysed in samples of saliva and/or serum by ELISA and/or chemiluminescence assays; the neutralizing activity of serum antibodies against reference strain B, Gamma and Delta SARS-CoV-2 variants were evaluated using a virus neutralization test and SARS-CoV-2 reactive interferon-gamma T-cell were analysed by flow cytometry. CoronaVac was able to induce serum and salivary IgG anti-spike antibodies and IFN-γ producing T cells in most individuals who had recovered from COVID-19 and/or were vaccinated. Virus neutralizing activity was observed against the ancestral strain, with a reduced response against the variants. Vaccinated individuals who had previous COVID-19 presented higher responses than vaccinated individuals for all variables analysed. Our study provides evidence that the CoronaVac vaccine was able to induce the production of specific serum and saliva antibodies, serum virus neutralizing activity and cellular immune response, which were increased in previously COVID-19-infected individuals compared to uninfected individuals.

IntroductionConsidering the likely need for the development of novel effective vaccines adapted to emerging relevant CoV-2 variants, the increasing knowledge of epitope recognition profile among convalescents and afterwards vaccinated with identification of immunodominant regions may provide important information.MethodsWe used an RBD peptide microarray to identify IgG and IgA binding regions in serum of 71 COVID-19 convalescents and 18 vaccinated individuals. ResultsWe found a set of immunodominant RBD antibody epitopes, each recognized by more than 30% of the tested cohort, that differ among the two different groups and are within conserved regions among betacoronavirus. Of those, only one peptide, P44 (S415-429), recognized by 68% of convalescents, presented IgG and IgA antibody reactivity that positively correlated with nAb titers, suggesting that this is a relevant RBD region and a potential target of IgG/IgA neutralizing activity.DiscussionThis peptide is localized within the area of contact with ACE-2 and harbors the mutation hotspot site K417 present in gamma (K417T), beta (K417N), and omicron (K417N) variants of concern. The epitope profile of vaccinated individuals differed from convalescents, with a more diverse repertoire of immunodominant peptides, recognized by more than 30% of the cohort. Noteworthy, immunodominant regions of recognition by vaccinated coincide with mutation sites at Omicron BA.1, an important variant emerging after massive vaccination. Together, our data show that immune pressure induced by dominant antibody responses may favor hotspot mutation sites and the selection of variants capable of evading humoral response.

Therapeutic vaccines based on monocyte‐derived dendritic cells have been shown to be promising strategies and may act as complementary treatments for viral infections, cancers, and, more recently, autoimmune diseases. Alpha‐type‐1‐polarized dendritic cells (aDC1s) have been shown to induce type‐1 immunity with a high capacity to produce interleukin‐12p70 (IL‐12p70). In the clinical use of cell‐based therapeutics, injectable solutions can affect the morphology, immunophenotypic profile, and viability of cells before delivery and their survival after injection. In this sense, preparing a cell suspension that maintains the quality of aDC1s is essential to ensure effective immunotherapy. In the present study, monocytes were differentiated into aDC1s in the presence of IL‐4 and GM‐CSF. On day 5, the cells were matured by the addition of a cytokine cocktail consisting of IFN‐α, IFN‐γ, IL‐1β, TNF‐α, and Poly I:C. After 48 hr, mature aDC1s were harvested and suspended in two different solutions: normal saline and Ringer's lactate. The maintenance of cells in suspension was evaluated after 4, 6, and 8 hr of storage. Cell viability, immunophenotyping, and apoptosis analyses were performed by flow cytometry. Cellular morphology was observed by electron microscopy, and the production of IL‐12p70 by aDC1s was evaluated by ELISA. Compared with normal saline, Ringer's lactate solution was more effective at maintaining DC viability for up to 8 hr of incubation at 4 or 22°C. [ABSTRACT FROM AUTHOR]

Several Prunus species are among the most important cultivated stone fruits in the Mediterranean region, and there is an urgent need to obtain rootstocks with specific adaptations to challenging environmental conditions. The development of adventitious roots (ARs) is an evolutionary mechanism of high relevance for stress tolerance, which has led to the development of environmentally resilient plants. As a first step towards understanding the genetic determinants involved in AR formation in Prunus sp., we evaluated the rooting of hardwood cuttings from five Prunus rootstocks (Adafuel, Adarcias, Cadaman, Garnem, and GF 677) grown in hydroponics. We found that auxin-induced callus and rooting responses were strongly genotype-dependent. To investigate the molecular mechanisms involved in these differential responses, we performed a time-series study of AR formation in two rootstocks with contrasting rooting performance, Garnem and GF 677, by culturing in vitro microcuttings with and without auxin treatment (0.9 mg/L of indole-3-butyric acid [IBA]). Despite showing a similar histological structure, Garnem and GF677 rootstocks displayed dynamic changes in endogenous hormone homeostasis involving metabolites such as indole-3-acetic acid (IAA) conjugated to aspartic acid (IAA-Asp), and these changes could explain the differences observed during rooting.

Root systems can display variable architectures that contribute to nutrient foraging or to increase the tolerance of abiotic stress conditions. Root tip excision promotes the developmental progression of previously specified lateral root (LR) founder cells, which allows to easily measuring the branching capacity of a given root as regards its genotype and/or growth conditions. Here, we describe the natural variation among 120 Arabidopsis thaliana accessions in root system architecture (RSA) after root tip excision. Wound-induced changes in RSA were associated with 19 genomic loci using genome-wide association mapping. Three candidate loci associated with wound-induced LR formation were investigated. Sequence variation in the hypothetical protein encoded by the At4g01090 gene affected wound-induced LR development and its loss-of-function mutants displayed a reduced number of LRs after root tip excision. Changes in a histidine phosphotransfer protein putatively involved in cytokinin signaling were significantly associated with LR number variation after root tip excision. Our results provide a better understanding of some of the genetic components involved in LR capacity variation among accessions.

Commercial carnation (Dianthus caryophyllus) cultivars are vegetatively propagated from axillary stem cuttings through adventitious rooting; a process which is affected by complex interactions between nutrient and hormone levels and is strongly genotype-dependent. To deepen our understanding of the regulatory events controlling this process, we performed a comparative study of adventitious root (AR) formation in two carnation cultivars with contrasting rooting performance, “2101–02 MFR” and “2003 R 8”, as well as in the reference cultivar “Master”. We provided molecular evidence that localized auxin response in the stem cutting base was required for efficient adventitious rooting in this species, which was dynamically established by polar auxin transport from the leaves. In turn, the bad-rooting behavior of the “2003 R 8” cultivar was correlated with enhanced synthesis of indole-3-acetic acid conjugated to aspartic acid by GH3 proteins in the stem cutting base. Treatment of stem cuttings with a competitive inhibitor of GH3 enzyme activity significantly improved rooting of “2003 R 8”. Our results allowed us to propose a working model where endogenous auxin homeostasis regulated by GH3 proteins accounts for the cultivar dependency of AR formation in carnation stem cuttings.

The rooting of stem cuttings is a highly efficient procedure for the vegetative propagation of ornamental plants. In cultivated carnations, an increased auxin level in the stem cutting base produced by active auxin transport from the leaves triggers adventitious root (AR) formation from the cambium. To provide additional insight into the physiological and genetic basis of this complex trait, we studied AR formation in a collection of 159 F1 lines derived from a cross between two hybrid cultivars (2003 R 8 and 2101-02 MFR) showing contrasting rooting performances. In three different experiments, time-series for several stem and root architectural traits were quantified in detail in a subset of these double-cross hybrid lines displaying extreme rooting phenotypes and their parental genotypes. Our results indicate that the water content and area of the AR system directly contributed to the shoot water content and shoot growth. Moreover, morphometric data and rooting quality parameters were found to be associated with some stress-related metabolites such as 1-aminocyclopropane-1-carboxylic acid (ACC), the ethylene precursor, and the conjugated auxin indol-3-acetic acid-aspartic acid (IAA-Asp).

Introduction: Considering the likely need for the development of novel effective vaccines adapted to emerging relevant CoV-2 variants, the increasing knowledge of epitope recognition profile among convalescents and afterwards vaccinated with identification of immunodominant regions may provide important information. Methods: We used an RBD peptide microarray to identify IgG and IgA binding regions in serum of 71 COVID-19 convalescents and 18 vaccinated individuals. Results: We found a set of immunodominant RBD antibody epitopes, each recognized by more than 30% of the tested cohort, that differ among the two different groups and are within conserved regions among betacoronavirus. Of those, only one peptide, P44 (S415-429), recognized by 68% of convalescents, presented IgG and IgA antibody reactivity that positively correlated with nAb titers, suggesting that this is a relevant RBD region and a potential target of IgG/ IgA neutralizing activity. Discussion: This peptide is localized within the area of contact with ACE-2 and harbors the mutation hotspot site K417 present in gamma (K417T), beta (K417N), and omicron (K417N) variants of concern. The epitope profile of vaccinated individuals differed from convalescents, with a more diverse repertoire of immunodominant peptides, recognized by more than 30% of the cohort. Noteworthy, immunodominant regions of recognition by vaccinated coincide with mutation sites at Omicron BA.1, an important variant emerging after massive vaccination. Together, our data show that immune pressure induced by dominant antibody responses may favor hotspot mutation sites and the selection of variants capable of evading humoral response. [ABSTRACT FROM AUTHOR]

Several Prunus species are among the most important cultivated stone fruits in the Mediterranean region, and there is an urgent need to obtain rootstocks with specific adaptations to challenging environmental conditions. The development of adventitious roots (ARs) is an evolutionary mechanism of high relevance for stress tolerance, which has led to the development of environmentally resilient plants. As a first step towards understanding the genetic determinants involved in AR formation in Prunus sp., we evaluated the rooting of hardwood cuttings from five Prunus rootstocks (Adafuel, Adarcias, Cadaman, Garnem, and GF 677) grown in hydroponics. We found that auxin-induced callus and rooting responses were strongly genotype-dependent. To investigate the molecular mechanisms involved in these differential responses, we performed a time-series study of AR formation in two rootstocks with contrasting rooting performance, Garnem and GF 677, by culturing in vitro microcuttings with and without auxin treatment (0.9 mg/L of indole-3-butyric acid [IBA]). Despite showing a similar histological structure, Garnem and GF677 rootstocks displayed dynamic changes in endogenous hormone homeostasis involving metabolites such as indole-3-acetic acid (IAA) conjugated to aspartic acid (IAA-Asp), and these changes could explain the differences observed during rooting. [ABSTRACT FROM AUTHOR]

Root systems can display variable architectures that contribute to nutrient foraging or to increase the tolerance of abiotic stress conditions. Root tip excision promotes the developmental progression of previously specified lateral root (LR) founder cells, which allows to easily measuring the branching capacity of a given root as regards its genotype and/or growth conditions. Here, we describe the natural variation among 120 Arabidopsis thaliana accessions in root system architecture (RSA) after root tip excision. Wound-induced changes in RSA were associated with 19 genomic loci using genome-wide association mapping. Three candidate loci associated with wound-induced LR formation were investigated. Sequence variation in the hypothetical protein encoded by the At4g01090 gene affected wound-induced LR development and its loss-of-function mutants displayed a reduced number of LRs after root tip excision. Changes in a histidine phosphotransfer protein putatively involved in cytokinin signaling were significantly associated with LR number variation after root tip excision. Our results provide a better understanding of some of the genetic components involved in LR capacity variation among accessions. [ABSTRACT FROM AUTHOR]

Commercial carnation (Dianthus caryophyllus) cultivars are vegetatively propagated from axillary stem cuttings through adventitious rooting; a process which is affected by complex interactions between nutrient and hormone levels and is strongly genotypedependent. To deepen our understanding of the regulatory events controlling this process, we performed a comparative study of adventitious root (AR) formation in two carnation cultivars with contrasting rooting performance, "2101-02 MFR" and "2003 R 8", as well as in the reference cultivar "Master". We provided molecular evidence that localized auxin response in the stem cutting base was required for efficient adventitious rooting in this species, which was dynamically established by polar auxin transport from the leaves. In turn, the bad-rooting behavior of the "2003 R 8" cultivar was correlated with enhanced synthesis of indole-3-acetic acid conjugated to aspartic acid by GH3 proteins in the stem cutting base. Treatment of stem cuttings with a competitive inhibitor of GH3 enzyme activity significantly improved rooting of "2003 R 8". Our results allowed us to propose a working model where endogenous auxin homeostasis regulated by GH3 proteins accounts for the cultivar dependency of AR formation in carnation stem cuttings. [ABSTRACT FROM AUTHOR]

Although cassava is of great importance as a starch source for industrial purposes or food consumption, the evaluation of different varieties is an under-investigated topic. Thus, in order to contribute to the identification of cassava starches more suitable for use and application in the food industry, this study aimed to analyze and compare the physicochemical properties of starches from ten different cassava varieties of household consumption, industrial use, and mixed use. In order to accomplish this goal, analyses were performed into the thermal and pasting properties, granule size, and amylopectin branch-chain-length distribution and amylose and phosphorus contents, as well as scanning electron microscopy (SEM) images of the starches. In addition, a principal component analysis (PCA) biplot was generated, as one of the most useful methods to analyze multidimensional datasets with quantitative variables. Significant differences in phosphorus and amylose contents, branch-chain-length distribution of amylopectin, starch granule diameter, gelatinization and retrogradation temperatures, and pasting properties were obtained. Therefore, this study adds to the literature regarding the physicochemical properties of starches of these cassava varieties, contributing to improving their uses in the food industry by novel applications these starches, and adding and corroborating uses of some varieties studied. [ABSTRACT FROM AUTHOR]

Pycnodysostosis is a rare, autosomal recessive syndrome characterized by osteosclerosis, brittle bones, stunting, and significant craniofacial changes. The objective of this study was to report a case of a 6-year-old patient with pycnodysostosis orthodontically treated and followed up until age 10 years and to discuss the risk factors, options for orthodontic treatment, and limitations involving this type of treatment, which has not yet been performed. Prevention through counseling and periodic follow-up visits is essential in eliminating factors that predispose patients to infections and fractures. New studies are necessary to establish safe and efficient orthodontic treatment plans. [ABSTRACT FROM AUTHOR]

Background/Aims: There are many controversies regarding side effects on craniofacial and extremity growth due to growth hormone (GH) treatment. Our aim was to estimate GH action on craniofacial development and extremity growth in GH-deficient patients. Methods: Twenty patients with GH deficiency with a chronological age ranging from 4.6 to 24.3 years (bone age from 1.5 to 13 years) were divided in 2 groups: group 1 (n = 6), naive to GH treatment, and group 2 (n = 14), ongoing GH treatment for 2–11 years. GH doses (0.1–0.15 U/kg/day) were adjusted to maintain insulin-like growth factor 1 and insulin-like growth factor binding protein 3 levels within the normal range. Anthropometric measurements, cephalometric analyses and facial photographs to verify profile and harmony were performed annually for at least 3 years. Results: Two patients with a disharmonious profile due to mandibular growth attained harmony, and none of them developed facial disharmony. Increased hand or foot size (>P97) was observed in 2 female patients and in 4 patients (1 female), respectively, both not correlated with GH treatment duration and increased levels of insulin-like growth factor 1. Conclusions: GH treatment with standard doses in GH-deficient patients can improve the facial profile in retrognathic patients and does not lead to facial disharmony although extremity growth, mainly involving the feet, can occur. Copyright © 2009 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Plants have a remarkable regenerative capacity, which allows them to survive tissue damage after biotic and abiotic stresses. In this study, we use Solanum lycopersicum 'Micro-Tom' explants as a model to investigate wound-induced de novo organ formation, as these explants can regenerate the missing structures without the exogenous application of plant hormones. Here, we performed simultaneous targeted profiling of 22 phytohormone-related metabolites during de novo organ formation and found that endogenous hormone levels dynamically changed after root and shoot excision, according to region-specific patterns. Our results indicate that a defined temporal window of high auxin-to-cytokinin accumulation in the basal region of the explants was required for adventitious root formation and that was dependent on a concerted regulation of polar auxin transport through the hypocotyl, of local induction of auxin biosynthesis, and of local inhibition of auxin degradation. In the apical region, though, a minimum of auxin-to-cytokinin ratio is established shortly after wounding both by decreasing active auxin levels and by draining auxin via its basipetal transport and internalization. Cross-validation with transcriptomic data highlighted the main hormonal gradients involved in wound-induced de novo organ formation in tomato hypocotyl explants. [ABSTRACT FROM AUTHOR]

The rooting of stem cuttings is a highly efficient procedure for the vegetative propagation of ornamental plants. In cultivated carnations, an increased auxin level in the stem cutting base produced by active auxin transport from the leaves triggers adventitious root (AR) formation from the cambium. To provide additional insight into the physiological and genetic basis of this complex trait, we studied AR formation in a collection of 159 F1 lines derived from a cross between two hybrid cultivars (2003 R 8 and 2101-02 MFR) showing contrasting rooting performances. In three different experiments, time-series for several stem and root architectural traits were quantified in detail in a subset of these double-cross hybrid lines displaying extreme rooting phenotypes and their parental genotypes. Our results indicate that the water content and area of the AR system directly contributed to the shoot water content and shoot growth. Moreover, morphometric data and rooting quality parameters were found to be associated with some stress-related metabolites such as 1-aminocyclopropane-1-carboxylic acid (ACC), the ethylene precursor, and the conjugated auxin indol-3-acetic acid-aspartic acid (IAA-Asp). [ABSTRACT FROM AUTHOR]