Optimising the recovery of phenolic compounds and antioxidant activity from orange peels through solid-state fermentation.

It is widely recognised that orange peels contain a considerable quantity of phenolics, primarily in the form of glycosides. The process of fermentation holds potential as a viable method for extracting phenolic compounds and facilitating their biotransformation into novel metabolites. The aim of this study was to assess the enhanced release of phenolic compounds through the process of solid-state fermentation of orange peels using microorganisms. Following a 6-day incubation period, the methanol extract obtained from the sample fermented with starter Banh men exhibited the highest concentration of total phenolic compounds (17.57 ± 0.34 mg GAE/g DW) and demonstrated the most significant DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity (55.03%). The Reverse Phase High Performance Liquid Chromatography (RP-HPLC) analysis revealed that the predominant phenolic compounds in all fermented samples were flavonoid aglycones, specifically naringenin, hesperetin, and nobiletin. Conversely, in the unfermented orange peels, the major compound observed was the glycoside derivative hesperidin.

Immunosuppressive Cyclotides: A Promising Approach for Treating Autoimmune Diseases.

The immune system maintains constant surveillance to prevent the infiltration of both endogenous and exogenous threats into host organisms. The process is regulated by effector immune cells that combat external pathogens and regulatory immune cells that inhibit excessive internal body inflammation, ultimately establishing a state of homeostasis within the body. Disruption to this process could lead to autoimmunity, which is often associated with the malfunction of both T cells and B cells with T cells playing a more major role. A number of therapeutic mediators for autoimmune diseases are available, from conventional disease-modifying drugs to biologic agents and small molecule inhibitors. Recently, ribosomally synthesized peptides, specifically cyclotides from plants are currently attracting more attention as potential autoimmune disease therapeutics due to their decreased toxicity compared to small molecules inhibitors as well as their remarkable stability against a number of factors. This review provides a concise overview of various cyclotides exhibiting immunomodulatory properties and their potential as therapeutic interventions for autoimmune diseases.

Identification and α-glucosidase inhibitory activity evaluation of two new coumarins derived from Mansonia gagei J. R. Drumm.

Five coumarins were isolated from the heartwood of Mansonia gagei, which included two newly discovered compounds, namely 11-hydroxypopulene E (1) and mansorin D (2), along with three previously identified compounds. The structures were determined through the utilisation of comprehensive spectroscopic data, ECD calculations, and a thorough comparison with existing literature data. The α-glucosidase inhibitory activities of all isolated compounds were assessed in yeast. Out of the compounds tested, compound 2 exhibited the most significant activity, displaying a percentage inhibition of 34.33% at a concentration of 200 µM.

Effective Strategies in Designing Chitosan-hyaluronic Acid Nanocarriers: From Synthesis to Drug Delivery Towards Chemotherapy.

The biomedical field faces an ongoing challenge in developing more effective anti-cancer medication due to the significant burden that cancer poses on human health. Extensive research has been conducted on the utilization of natural polysaccharides in nanomedicine owing to their properties of biocompatibility, biodegradability, non-immunogenicity, and non-toxicity. These characteristics make them a potent drug delivery system for cancer therapy. The chitosan hyaluronic acid nanoparticle (CSHANp) system, consisting of chitosan and hyaluronic acid nanoparticles, has exhibited considerable potential as a nanocarrier for various cancer drugs, rendering it one of the most auspicious systems presently accessible. The CSHANps demonstrate remarkable drug loading capacity, precise control over drug release, and exceptional selectivity towards cancer cells. These properties enhance the therapeutic effectiveness against cancerous cells. This article aims to provide a comprehensive analysis of CSHANp, focusing on its characteristics, production techniques, applications, and future prospects.

Cyclotides: The next generation in biopesticide development for eco-friendly agriculture.

Chemical pesticides remain the predominant method for pest management in numerous countries. Given the current landscape of agriculture, the development of biopesticides has become increasingly crucial. The strategy empowers farmers to efficiently manage pests and diseases, while prioritizing minimal adverse effects on the environment and human health, hence fostering sustainable management. In recent years, there has been a growing interest and optimism surrounding the utilization of peptide biopesticides for crop protection. These sustainable and environmentally friendly substances have been recognized as viable alternatives to synthetic pesticides due to their outstanding environmental compatibility and efficacy. Numerous studies have been conducted to synthesize and identify peptides that exhibit activity against significant plant pathogens. One of the peptide classes is cyclotides, which are cyclic cysteine-rich peptides renowned for their wide range of sequences and functions. In this review, we conducted a comprehensive analysis of cyclotides, focusing on their structural attributes, developmental history, significant biological functions in crop protection, techniques for identification and investigation, and the application of biotechnology to enhance cyclotide synthesis. The objective is to emphasize the considerable potential of cyclotides as the next generation of plant protection agents on the global scale.

Cyclotides derived from Viola dalatensis Gagnep: A novel approach for enrichment and evaluation of antimicrobial activity.

Cyclotides, plant-derived cysteine-rich peptides, exhibit a wide range of beneficial biological activities and possess exceptional structural stability. Cyclotides are commonly distributed throughout the Violaceae family. Viola dalatensis Gagnep, a Vietnamese species, has not been well studied, especially for cyclotides. This pioneering research explores cyclotides from V. dalatensis as antimicrobials. This study used a novel approach to enhance cyclotides after extraction. The approach combined 30% ammonium sulfate salt precipitation and RP-HPLC. A comprehensive analysis was performed to ascertain the overall protein content, flavonoids content, polyphenol content, and free radical scavenging capacity of compounds derived from V. dalatensis. Six known cyclotides were sequenced utilizing MS tandem. Semi-purified cyclotide mixtures (M1, M2, and M3) exhibited antibacterial efficacy against Bacillus subtilis (inhibitory diameters: 19.67-23.50 mm), Pseudomonas aeruginosa (22.17-23.50 mm), and Aspergillus flavus (14.67-21.33 mm). The enriched cyclotide precipitate from the stem extract demonstrated a minimum inhibitory concentration (MIC) of 0.08 mg/mL against P. aeruginosa, showcasing significant antibacterial effectiveness compared to the stem extract (MIC: 12.50 mg/mL). Considerable advancements have been achieved in the realm of cyclotides, specifically in their application as antimicrobial agents.

Robust preimplantation genetic testing of the common F8 Inv22 pathogenic variant of severe hemophilia A using a highly polymorphic multi-marker panel encompassing the paracentric inversion.

BACKGROUND: Hemophilia A (HEMA) is an X-linked bleeding disorder caused by reduced/absent coagulation factor VIII expression, as a result of pathogenic variants in the F8 gene. Preimplantation prevention of HEMA should ideally include direct pathogenic F8 variant detection, complemented by linkage analysis of flanking markers to identify the high-risk F8 allele. Linkage analysis is particularly indispensable when the pathogenic variant cannot be detected directly or identified. This study evaluated the suitability of a panel of F8 intragenic and extragenic short tandem repeat markers for standalone linkage-based preimplantation genetic testing for monogenic disorder (PGT-M) of the Inv22 pathogenic variant, an almost 600 kb paracentric inversion responsible for almost half of all severe HEMA globally, for which direct detection is challenging. METHODS: Thirteen markers spanning 1 Mb and encompassing both F8 and the Inv22 inversion interval were genotyped in 153 unrelated females of Viet Kinh ethnicity. RESULTS: All individuals were heterozygous for ≥ 1 marker, ~ 90% were heterozygous for ≥ 1 of the five F8 intragenic markers, and almost 98% were heterozygous for ≥ 1 upstream (telomeric) and ≥ 1 downstream (centromeric) markers. A prospective PGT-M couple at risk of transmitting F8 Inv22 were fully informative at four marker loci (2 intra-inversion, 1 centromeric, 1 telomeric) and partially informative at another five (2 intra-inversion, 3 centromeric), allowing robust phasing of low- and high-risk haplotypes. In vitro fertilization produced three embryos, all of which clearly inherited the low-risk maternal allele, enabling reliable unaffected diagnoses. A single embryo transfer produced a clinical pregnancy, which was confirmed as unaffected by amniocentesis and long-range PCR, and a healthy baby girl was delivered at term. CONCLUSION: Robust and reliable PGT-M of HEMA, including the common F8 Inv22 pathogenic variant, can be achieved with sufficient informative intragenic and flanking markers.

Length limitation of astral microtubules orients cell divisions in murine intestinal crypts.

Planar spindle orientation is critical for epithelial tissue organization and is generally instructed by the long cell-shape axis or cortical polarity domains. We introduced mouse intestinal organoids in order to study spindle orientation in a monolayered mammalian epithelium. Although spindles were planar, mitotic cells remained elongated along the apico-basal (A-B) axis, and polarity complexes were segregated to basal poles, so that spindles oriented in an unconventional manner, orthogonal to both polarity and geometric cues. Using high-resolution 3D imaging, simulations, and cell-shape and cytoskeleton manipulations, we show that planar divisions resulted from a length limitation in astral microtubules (MTs) which precludes them from interacting with basal polarity, and orient spindles from the local geometry of apical domains. Accordingly, lengthening MTs affected spindle planarity, cell positioning, and crypt arrangement. We conclude that MT length regulation may serve as a key mechanism for spindles to sense local cell shapes and tissue forces to preserve mammalian epithelial architecture.

Insights into the synthesis strategies of plant-derived cyclotides.

Cyclotides are plant peptides characterized with a head-to-tail cyclized backbone and three interlocking disulfide bonds, known as a cyclic cysteine knot. Despite the variations in cyclotides peptide sequences, this core structure is conserved, underlying their most useful feature: stability against thermal and chemical breakdown. Cyclotides are the only natural peptides known to date that are orally bioavailable and able to cross cell membranes. Cyclotides also display bioactivities that have been exploited and expanded to develop as potential therapeutic reagents for a wide range of conditions (e.g., HIV, inflammatory conditions, multiple sclerosis, etc.). As such, in vitro production of cyclotides is of the utmost importance since it could assist further research on this peptide class, specifically the structure-activity relationship and its mechanism of action. The information obtained could be utilized to assist drug development and optimization. Here, we discuss several strategies for the synthesis of cyclotides using both chemical and biological routes.

Cysteine-rich peptides: From bioactivity to bioinsecticide applications.

Greater levels of insect resistance and constraints on the use of current pesticides have recently led to increased crop losses in agricultural production. Further, the health and environmental impacts of pesticides now restrict their application. Biologics based on peptides are gaining popularity as efficient crop protection agents with low environmental toxicity. Cysteine-rich peptides (whether originated from venoms or plant defense substances) are chemically stable and effective as insecticides in agricultural applications. Cysteine-rich peptides fulfill the stability and efficacy requirements for commercial uses and provide an environmentally benign alternative to small-molecule insecticides. In this article, cysteine-rich insecticidal peptide classes identified from plants and venoms will be highlighted, focusing on their structural stability, bioactivity and production.

Extending the Range of SLIM-Labeling Applications: From Human Cell Lines in Culture to Caenorhabditis elegans Whole-Organism Labeling.

The simple light isotope metabolic-labeling technique relies on the in vivo biosynthesis of amino acids from U-[12C]-labeled molecules provided as the sole carbon source. The incorporation of the resulting U-[12C]-amino acids into proteins presents several key advantages for mass-spectrometry-based proteomics analysis, as it results in more intense monoisotopic ions, with a better signal-to-noise ratio in bottom-up analysis. In our initial studies, we developed the simple light isotope metabolic (SLIM)-labeling strategy using prototrophic eukaryotic microorganisms, the yeasts Candida albicans and Saccharomyces cerevisiae, as well as strains with genetic markers that lead to amino-acid auxotrophy. To extend the range of SLIM-labeling applications, we evaluated (i) the incorporation of U-[12C]-glucose into proteins of human cells grown in a complex RPMI-based medium containing the labeled molecule, considering that human cell lines require a large number of essential amino-acids to support their growth, and (ii) an indirect labeling strategy in which the nematode Caenorhabditis elegans grown on plates was fed U-[12C]-labeled bacteria (Escherichia coli) and the worm proteome analyzed for 12C incorporation into proteins. In both cases, we were able to demonstrate efficient incorporation of 12C into the newly synthesized proteins, opening the way for original approaches in quantitative proteomics.

The acyclotide ribe 31 from Rinorea bengalensis has selective cytotoxicity and potent insecticidal properties in Drosophila.

Cyclotides and acyclic versions of cyclotides (acyclotides) are peptides involved in plant defense. These peptides contain a cystine knot motif formed by three interlocked disulfide bonds, with the main difference between the two classes being the presence or absence of a cyclic backbone, respectively. The insecticidal activity of cyclotides is well documented, but no study to date explores the insecticidal activity of acyclotides. Here, we present the first in vivo evaluation of the insecticidal activity of acyclotides from Rinorea bengalensis on the vinegar fly Drosophila melanogaster. Of a group of structurally comparable acyclotides, ribe 31 showed the most potent toxicity when fed to D. melanogaster. We screened a range of acyclotides and cyclotides and found their toxicity toward human red blood cells was substantially lower than toward insect cells, highlighting their selectivity and potential for use as bioinsecticides. Our confocal microscopy experiments indicated their cytotoxicity is likely mediated via membrane disruption. Furthermore, our surface plasmon resonance studies suggested ribe 31 preferentially binds to membranes containing phospholipids with phosphatidyl-ethanolamine headgroups. Despite having an acyclic backbone, we determined the three-dimensional NMR solution structure of ribe 31 is similar to that of cyclotides. In summary, our results suggest that, with further optimization, ribe 31 could have applications as an insecticide due to its potent in vivo activity against D. melanogaster. More broadly, this work advances the field by demonstrating that acyclotides are more common than previously thought, have potent insecticidal activity, and have the advantage of potentially being more easily manufactured than cyclotides.

Modulation of designer biomimetic matrices for optimized differentiated intestinal epithelial cultures.

The field of intestinal biology is thirstily searching for different culture methods that complement the limitations of organoids, particularly the lack of a differentiated intestinal compartment. While being recognized as an important milestone for basic and translational biological studies, many primary cultures of intestinal epithelium (IE) rely on empirical trials using hydrogels of various stiffness, whose mechanical impact on epithelial organization remains vague until now. Here, we report the development of hydrogel scaffolds with a range of elasticities and their influence on IE expansion, organization, and differentiation. On stiff substrates (>5 kPa), mouse IE cells adopt a flat cell shape and detach in the short-term. In contrast, on soft substrates (80-500 Pa), they sustain for a long-term, pack into high density, develop columnar shape with improved apical-basal polarity and differentiation marker expression, a phenotype reminiscent of features in vivo mouse IE. We then developed a soft gel molding process to produce 3D Matrigel scaffolds of close-to-nature stiffness, which support and maintain a culture of mouse IE into crypt-villus architecture. Thus, the present work is up-to-date informative for the design of biomaterials for ex vivo intestinal models, offering self-renewal in vitro culture that emulates the mouse IE.

Adipogenesis of ear mesenchymal stem cells (EMSCs): adipose biomarker-based assessment of genetic variation, adipocyte function, and brown/brite differentiation.

Ear mesenchymal stem cells (EMSCs) have been investigated to differentiate into adipocytes, chondrocytes, and muscle cells in vitro. However, the factors controlling adipogenesis of this stem cell population in vitro, function, and type of adipocytes raised from them are still unclear. Here we found that genetics have a modest effect on adipogenic capacity of EMSCs. Adipocytes differentiated from EMSCs have a potential function in lipid metabolism as indicated by expression of lipogenic genes and this function of EMSC adipocytes is regulated by genetics. EMSCs failed to be differentiated into brite/brown adipocytes due to their lack of a thermogenic program, but adipocytes raised from EMSCs showed a fate of white adipocytes. Overall, our data suggest that EMSCs differentiate into functional white adipocytes in vitro and this is genetic-dependent.

Investigating the nature of active forces in tissues reveals how contractile cells can form extensile monolayers.

Actomyosin machinery endows cells with contractility at a single-cell level. However, within a monolayer, cells can be contractile or extensile based on the direction of pushing or pulling forces exerted by their neighbours or on the substrate. It has been shown that a monolayer of fibroblasts behaves as a contractile system while epithelial or neural progentior monolayers behave as an extensile system. Through a combination of cell culture experiments and in silico modelling, we reveal the mechanism behind this switch in extensile to contractile as the weakening of intercellular contacts. This switch promotes the build-up of tension at the cell-substrate interface through an increase in actin stress fibres and traction forces. This is accompanied by mechanotransductive changes in vinculin and YAP activation. We further show that contractile and extensile differences in cell activity sort cells in mixtures, uncovering a generic mechanism for pattern formation during cell competition, and morphogenesis.

In Planta Discovery and Chemical Synthesis of Bracelet Cystine Knot Peptides from Rinorea bengalensis.

Cyclotides are plant-derived peptides that have attracted interest as biocides and scaffolds for the development of stable peptide therapeutics. Cyclotides are characterized by their cyclic backbone and cystine knot framework, which engenders them with remarkably high stability. This study reports the cystine knot-related peptidome of Rinorea bengalensis, a small rainforest tree in the Violaceae family that is distributed from Australia westward to India. Surprisingly, many more acyclic knotted peptides (acyclotides) were discovered than cyclic counterparts (cyclotides), with 32 acyclotides and 1 cyclotide sequenced using combined transcriptome and proteomic analyses. Nine acyclotides were isolated and screened against a panel of mammalian cell lines, showing they had the cytotoxic properties normally associated with cyclotide-like peptides. NMR analysis of the acyclotide ribes 21 and 22 and the cyclotide ribe 33 confirmed that these peptides contained the cystine knot structural motif. The bracelet-subfamily cyclotide ribe 33 was amenable to chemical synthesis in reasonable yield, an achievement that has long eluded previous attempts to synthetically produce bracelet cyclotides. Accordingly, ribe 33 represents an exciting new bracelet cyclotide scaffold that can be subject to chemical modification for future molecular engineering applications.

Exploring the Sequence Diversity of Cyclotides from Vietnamese Viola Species.

Viola is the largest genus in the Violaceae plant family and is known for its ubiquitous natural production of cyclotides. Many Viola species are used as medicinal herbs across Asia and are often consumed by humans in teas for the treatment of diseases, including ulcers and asthma. Previous studies reported the isolation of cyclotides from Viola species in many countries in the hope of discovering novel compounds with anti-cancer activities; however, Viola species from Vietnam have not been investigated to date. Here, the discovery of cyclotides from three Viola species (V. arcuata, V. tonkinensis, and V. austrosinensis) collected in the northern mountainous region of Vietnam is reported. Ten cyclotides were isolated from these three Viola species: four are novel and six were previously reported to be expressed in other plants. The structures of three of the new bracelet cyclotides are similar to that of cycloviolacin O2. Because cycloviolacin O2 has previously been shown to have potent activity against a wide range of cancer cell lines including HeLa (human cervical cancer cells) and PC-3 (human prostate cancer cells), the cancer cytotoxicity of the cyclotides isolated from V. arcuata was assessed. All tested cyclotides were cytotoxic against cancer cells, albeit to varying degrees. The sequences discovered in this study significantly expand the understanding of cyclotide diversity, especially in comparison with other cyclotides found in plants from the Asian region.

Recent Progress of Stem Cell Therapy in Cancer Treatment: Molecular Mechanisms and Potential Applications.

The insufficient and unspecific target of traditional therapeutic approaches in cancer treatment often leads to therapy resistance and cancer recurrence. Over the past decades, accumulating discoveries about stem cell biology have provided new potential approaches to cure cancer patients. Stem cells possess unique biological actions, including self-renewal, directional migration, differentiation, and modulatory effects on other cells, which can be utilized as regenerative medicine, therapeutic carriers, drug targeting, and generation of immune cells. In this review, we emphasize the mechanisms underlying the use of various types of stem cells in cancer treatment. In addition, we summarize recent progress in the clinical applications of stem cells, as well as common risks of this therapy. We finally give general directions for future studies, aiming to improve overall outcomes in the fight against cancer.

An Update on the Progress of Isolation, Culture, Storage, and Clinical Application of Human Bone Marrow Mesenchymal Stem/Stromal Cells.

Bone marrow mesenchymal stem/stromal cells (BMSCs), which are known as multipotent cells, are widely used in the treatment of various diseases via their self-renewable, differentiation, and immunomodulatory properties. In-vitro and in-vivo studies have supported the understanding mechanisms, safety, and efficacy of BMSCs therapy in clinical applications. The number of clinical trials in phase I/II is accelerating; however, they are limited in the size of subjects, regulations, and standards for the preparation and transportation and administration of BMSCs, leading to inconsistency in the input and outcome of the therapy. Based on the International Society for Cellular Therapy guidelines, the characterization, isolation, cultivation, differentiation, and applications can be optimized and standardized, which are compliant with good manufacturing practice requirements to produce clinical-grade preparation of BMSCs. This review highlights and updates on the progress of production, as well as provides further challenges in the studies of BMSCs, for the approval of BMSCs widely in clinical application.