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In the realm of cancer immunotherapy, the meticulous selection of neoantigens plays a fundamental role in enhancing personalized treatments. Traditionally, this selection process has heavily relied on predicting the binding of peptides to human leukocyte antigens (pHLA). Nevertheless, this approach often overlooks the dynamic interaction between tumor cells and the immune system. In response to this limitation, we have developed an innovative prediction algorithm rooted in machine learning, integrating T cell receptor β chain (TCRβ) profiling data from colorectal cancer (CRC) patients for a more precise neoantigen prioritization. TCRβ sequencing was conducted to profile the TCR repertoire of tumor-infiltrating lymphocytes (TILs) from 28 CRC patients. The data unveiled both intra-tumor and inter-patient heterogeneity in the TCRβ repertoires of CRC patients, likely resulting from the stochastic utilization of V and J segments in response to neoantigens. Our novel combined model integrates pHLA binding information with pHLA-TCR binding to prioritize neoantigens, resulting in heightened specificity and sensitivity compared to models using individual features alone. The efficacy of our proposed model was corroborated through ELISpot assays on long peptides, performed on four CRC patients. These assays demonstrated that neoantigen candidates prioritized by our combined model outperformed predictions made by the established tool NetMHCpan. This comprehensive assessment underscores the significance of integrating pHLA binding with pHLA-TCR binding analysis for more effective immunotherapeutic strategies.

This is one part of the 4-H Volunteer Training series. It was originally written in 2005, adapted in 2014, and updated in 2023.

This is one part of the 4-H Volunteer Training series. It was originally published in 2005 and revised in 2012. It was updated in the summer of 2023 and is now published in 2024.

Sap coconut (Cocos nucifera L. var. makapuno) in Vietnam is a mutant coconut variant; coconut water is in the state of lotus glue, and coconut rice is like cream. For high-quality transcriptome, sequencing, quality, purity and concentration of RNA are the key factors. However, coconut endosperm tissue has higher stiffness and fatness than the leaf tissue, which complicates the extraction process. Moreover, RNA is much more difficult to preserve than DNA. In this study, various RNA extraction methods were examined in Vietnamese waxy coconut endosperm tissue samples. Optimum extraction and preservation of the RNA using the simplest possible chemicals was the objective of this study. The modified CTAB method with LiCl and the TRI reagent method were tested and evaluated. The purity, concentration and quality of RNA after storage were improved. The findings indicated that the TRIsure extraction method with the addition of NaCl and β-mercaptoethanol yielded optimum RNA quality. The RNA concentration was 159 ng/µL, with a purity ratio of 1.94 ± 0.04 for A260/A280 and 1.58 ± 0.02 for A260/230. RNA samples remained stable for up to 3 weeks when stored in absolute ethanol at 8°C–10°C, which significantly reduced their degradation during transportation. This study facilitated the use of simple chemicals for high-quality RNA extraction from coconut endosperm and its preservation for applications in high throughput sequencing.

During the last decade, the detection of neurotropic astroviruses has increased dramatically. The MLB genogroup of astroviruses represents a genetically distinct group of zoonotic astroviruses associated with gastroenteritis and severe neurological complications in young children, the immunocompromised, and the elderly. Using different virus evolution approaches, we identified dispensable regions in the 3' end of the capsid-coding region responsible for attenuation of MLB astroviruses in susceptible cell lines. To create recombinant viruses with identified deletions, MLB reverse genetics (RG) and replicon systems were developed. Recombinant truncated MLB viruses resulted in imbalanced RNA synthesis and strong attenuation in iPSC-derived neuronal cultures confirming the location of neurotropism determinants. This approach can be used for the development of vaccine candidates using attenuated astroviruses that infect humans, livestock animals, and poultry.

In this study, chemical components, α-glucosidase inhibitory activities, and molecular modelling studies of the essential oil extracted from the Cleistocalyx operculatus leaves were investigated. In total, thirty compounds were identified using GC/MS, representing 98.3% of the oil. Of these, the two most dominant constituents of the essential oil were determined as (Z)-β-ocimene (30.4%) and allo-ocimene (31.6%). The α-glucosidase inhibitory experiments indicated that the essential oil exhibited potent α-glucosidase inhibitory activities, with IC50 values of 61.82 ± 3.91 µg/mL. For further investigation into inhibitory mechanisms, molecular docking simulations were performed to investigate structural interactions between two dominant constituents and the α-glucosidase protein. The simulation revealed that allo-ocimene (31.6%) and (Z)-β-ocimene (30.4%) have protein binding affinities of −5.358 and −5.330 kcal/mol, respectively. Moreover, molecular dynamic simulation indicated that the complexes of two compounds and the target protein were stable over 100 ns. Overall, these findings suggest that the essential oil of C. operculatus leaves could be a natural source of potential α-glucosidase inhibitors.

Astronauts are always faced with serious health problems during prolonged spaceflights. Previous studies have shown that weightlessness significantly affects the physiological function of female astronauts, including a change in reproductive hormones and ovarian cells, such as granulosa and theca cells. However, the effects of microgravity on these cells have not been well characterized, especially in granulosa cells. This study aimed to investigate the effects of simulated microgravity (SMG) on the proliferation and morphology of porcine granulosa cells (pGCs). pGC proliferation from the SMG group was inhibited, demonstrated by the reduced O.D. value and cell density in the WST-1 assay and cell number counting. SMG-induced pGCs exhibited an increased ratio of cells in the G0/G1 phase and a decreased ratio of cells in the S and G2/M phase. Western blot analysis indicated a down-regulation of cyclin D1, cyclin-dependent kinase 4 (cdk4), and cyclin-dependent kinase 6 (cdk6), leading to the prevention of the G1-S transition and inducing the arrest phase. pGCs under the SMG condition showed an increase in nuclear area. This caused a reduction in nuclear shape value in pGCs under the SMG condition. SMG-induced pGCs exhibited different morphologies, including fibroblast-like shape, rhomboid shape, and pebble-like shape. These results revealed that SMG inhibited proliferation and induced morphological changes in pGCs.

Fucoxanthin extracted and purified from Vietnamese Sargassum oligocystum Montagne, 1845 exhibits various biological activities. In this study, the ability of fucoxanthin to inhibit acetylcholinesterase (AChE), the antioxidant activities, and the expression of antioxidant enzymes were investigated. Fucoxanthin isolated from Vietnamese S. oligocystum showed no cytotoxic effects; moreover, it exhibited AChE inhibitory activity (with an IC50 value of 130.12 ± 6.65 μg mL−1) and antioxidant activity (with an IC50 value of 3.42 ± 0.15 mg mL−1). At concentrations of 50 and 100 µg mL−1, fucoxanthin provided protection against amyloid β-protein fragment 25–35-induced neurotoxicity in a C6 neuronal cell line, and the survival of C6 cells was higher than 81.01% and 80.98%, respectively, compared to the control group (59%). Moreover, antioxidant enzyme activity and quantitative PCR analysis suggested that the neuroprotective effect of fucoxanthin resulted from regulation of the gene expression of antioxidant enzymes (CAT and GPx) and ER pathways (caspase-3 and Bax), as well as the promotion of expression of genes involved in PI3K/Akt signaling (GSK-3β), autophagy (p62 and ATG5), and the biosynthesis of ACh (VAChT and ChAT). Therefore, fucoxanthin extracted from the seaweed S. oligocystum in Vietnam is a potential feedstock source for the production of health foods that exert neuroprotective effects.

Background: Wilson disease (WD) is caused by mutations in the copper-transporting P-type adenosine triphosphatase encoded by the ATP7B gene. In this study, we screened and identified the ATP7B mutations among unrelated Vietnamese pediatric patients. Methods: One-hundred-thirteen pediatric patients with clinically diagnosed WD were recruited. DNA samples were extracted from peripheral blood. Mutations in the ATP7B gene were identified by Sanger sequencing. Results: Approximately 98% of the clinically diagnosed WD patients carried ATP7B mutations. A total of 35 different ATP7B variants were detected, including five novel mutations (L658P, L792P, T977K, IVS4 + 1G > A and IVS20 + 4A > G). Remarkably, this study revealed that S105* was the most prevalent variant (32.27%), followed by L1371P (9.09%), I1148T (7.27%), R778L (6.36%), T850I (5.45%), V176Sfs*28 and IVS14-2A > G (4.55%). Most ATP7B mutations were located in the exon 2 (37.73%), exon 16 (10.00%), exon 8 (9.55%), exon 20 (9.09%), exon 10 and exon 18 (5.45%), exon 14 (5.00%), exon 13 and intron 14 (4.55%). We developed a streamlined procedure to quickly characterize mutations in the ATP7B gene in the Vietnamese children, starting with sequencing exon 2 and subsequently to exons 8,10,13-16,18, and 20 to allow quick diagnosis of clinically suspected patients. Conclusion: The mutational spectrum and hotspots of ATP7B gene in the Vietnamese population were fairly different from other East Asian populations. A streamlined procedure was developed to screen exon 2 in ATP7B gene among suspected WD patients to reduce genetically diagnostic cost, to facilitate early detection and intervention in countries with limited resources.

Cellular deposition of protein aggregates, one of the hallmarks of neurodegeneration, disrupts cellular functions and leads to neuronal death. Mutations, posttranslational modifications, and truncations are common molecular underpinnings in the formation of aberrant protein conformations that seed aggregation. The major proteins involved in neurodegeneration include amyloid beta (Aβ) and tau in Alzheimer’s disease, α-synuclein in Parkinson’s disease, and TAR DNA-binding protein (TDP-43) in amyotrophic lateral sclerosis (ALS). These proteins are described as intrinsically disordered and possess enhanced ability to partition into biomolecular condensates. In this review, we discuss the role of protein misfolding and aggregation in neurodegenerative diseases, specifically highlighting implications of changes to the primary/secondary (mutations, posttranslational modifications, and truncations) and the quaternary/supramolecular (oligomerization and condensation) structural landscapes for the four aforementioned proteins. Understanding these aggregation mechanisms provides insights into neurodegenerative diseases and their common underlying molecular pathology.

Aurantiochytrium is a heterotrophic marine microalga that has potential industrial applications. The main objectives of this study were to isolate an Aurantiochytrium strain from Sand Cay (Son Ca) Island, Vietnam, optimize its culture conditions, determine its nutritional composition, extract polyunsaturated fatty acids (PUFAs) in the free (FFA) and the alkyl ester (FAAE) forms, and evaluate the antioxidation and neuroprotection properties of the PUFAs. Aurantiochytrium sp. SC145 can be grown stably under laboratory conditions. Its culture conditions were optimized for a dry cell weight (DCW) of 31.18 g/L, with total lipids comprising 25.29%, proteins 7.93%, carbohydrates 15.21%, and carotenoid at 143.67 µg/L of DCW. The FAAEs and FFAs extracted from Aurantiochytrium sp. SC145 were rich in omega 3–6–9 fatty acids (40.73% and 44.00% of total fatty acids, respectively). No acute or subchronic oral toxicity was determined in mice fed with the PUFAs in FFA or FAAE forms at different doses over 90 days. Furthermore, the PUFAs in the FFA or FAAE forms and their main constituents of EPA, DHA, and ALA showed antioxidant and AChE inhibitory properties and neuroprotective activities against damage caused by H2O2- and amyloid-ß protein fragment 25–35 (Aβ25-35)-induced C6 cells. These data suggest that PUFAs extracted from Aurantiochytrium sp. SC145 may be a potential therapeutic target for the treatment of neurodegenerative disorders.

Normalization of RNA-seq data has been an active area of research since the problem was first recognized a decade ago. Despite the active development of new normalizers, their performance measures have been given little attention. To evaluate normalizers, researchers have been relying on ad hoc measures, most of which are either qualitative, potentially biased, or easily confounded by parametric choices of downstream analysis. We propose a metric called condition-number based deviation, or cdev, to quantify normalization success. cdev measures how much an expression matrix differs from another. If a ground truth normalization is given, cdev can then be used to evaluate the performance of normalizers. To establish experimental ground truth, we compiled an extensive set of public RNA-seq assays with external spike-ins. This data collection, together with cdev, provides a valuable toolset for benchmarking new and existing normalization methods.

Abstract Objective To advance public health support for the U.S. Department of Housing and Urban Development’s smoke-free rule, the Centers for Disease Control and Prevention collaborated with the Georgia Institute of Technology to develop a geospatial mapping tool. The objective was to create a tool state and local public health agencies could use to tailor smoke-free educational materials and cessation interventions for specific public housing development resident populations. Results The resulting “Extinguish Tool” includes an interactive map of U.S. public housing developments (PHDs) and healthcare facilities that provides detailed information on individual PHDs, their proximity to existing healthcare facilities, and the demographic characteristics of residents. The tool also estimates the number of PHD residents who smoke cigarettes and calculates crude estimates of the potential economic benefits of providing cessation interventions to these residents. The geospatial mapping tool project serves as an example of a collaborative and innovative public health approach to protecting the health and well-being of the nation’s two million public housing residents, including 760,000 children, from the harms of tobacco smoking and secondhand smoke exposure in the places where they live, play, and gather.

Examining whether and how the rhythms of limb and breathing movements interact is highly informative about the mechanistic origin of hyperpnoea during running exercise. However, studies have failed to reveal regularities. In particular, whether breathing frequency is inherently proportional to limb velocity and imposed by a synchronization of breaths to strides is still unclear. Here, we examined respiratory changes during running in the resourceful mouse model. We show that, for a wide range of trotting speeds on a treadmill, respiratory rate increases to a fixed and stable value irrespective of trotting velocities. Respiratory rate was yet further increased during escape-like running and most particularly at gallop. However, we found no temporal coordination of breaths to strides at any speed, intensity, or gait. Our work thus highlights that exercise hyperpnoea can operate, at least in mice and in the presently examined running regimes, without phasic constraints from limb movements.

White mulberry (Morus alba L.) root was reported to contain potential tyrosinase inhibitors such as norartocarpetin, streppogenin, oxyresveratrol (5, 7, 2′, 4′-tetrahydroxy-flavavone) and 2-arylbenzofuran. This study aims at investigating Morus alba L. hairy root culture to create a source of tyrosinase inhibitors for skin-whitening application. Morus alba L. leaf (including mesophyll and petiole) and stem explants were randomly wounded and inoculated with Agrobacterium rhizogenes ATCC 15834 in 10-40 minutes, and coculturing for 4 days. After inoculation, Agrobacterium rhizogenes were removed by using MS medium adding 200mg/L cefotaxime as antibiotics. Then, fresh inducted hairy roots were grown in nonphytohormone liquid medium (MS, White, Gamborg B5 or WPM) on a shaker (80rpm) at 25 ±1°C. Inducted hairy roots were tested for the present of rolB by PCR analysis and its effect on tyrosinase activity using a 96-well plate reader. The suitable explants for hairy root induction are leaves, especially petioles in 10 minutes (root induction frequency: 83%). After induction, hairy roots on MS was grown more efficiently (increased 3.933 times in FW) than on WPM, B5 and White medium. Inducted hairy roots were successfully transgenes by the present of rolB in their genome and had the same anti-tyrosinase activity as the natural roots without significance differences (IC50: 3.860 and 3.597 μg/mL, respectively). Cultured Morus alba L. hairy roots are potential source of tyrosinase inhibitors for future skin-whitening ingredient.

Exosomes are nano-scale and closed membrane vesicles which are promising for therapeutic applications due to exosome-enclosed therapeutic molecules such as DNA, small RNAs, proteins and lipids. Recently, it has been demonstrated that mesenchymal stem cell (MSC)-derived exosomes have capacity to regulate many biological events associated with wound healing process, such as cell proliferation, cell migration and blood vessel formation. This study investigated the regenerative potentials for cutaneous tissue, in regard to growth factors associated with wound healing and skin cell proliferation and migration, by exosomes released from primary MSCs originated from bone marrow (BM), adipose tissue (AD), and umbilical cord (UC) under serum- and xeno-free condition. We found crucial wound healing-mediated growth factors, such as vascular endothelial growth factor A (VEGF-A), fibroblast growth factor 2 (FGF-2), hepatocyte growth factor (HGF), and platelet-derived growth factor BB (PDGF-BB) in exosomes derived from all three MSC sources. However, expression levels of these growth factors in exosomes were influenced by MSC origins, especially transforming growth factor beta (TGF-β) was only detected in UCMSC-derived exosomes. All exosomes released by three MSCs sources induced keratinocyte and fibroblast proliferation and migration; and, the induction of cell migration is a dependent manner with the higher dose of exosomes was used (20 μg), the faster migration rate was observed. Additionally, the influences of exosomes on cell proliferation and migration was associated with exosome origins and also target cells of exosomes that the greatest induction of primary dermal fibroblasts belongs to BMMSC-derived exosomes and keratinocytes belongs to UCMSC-derived exosomes. Data from this study indicated that BMMSCs and UCMSCs under clinical condition secreted exosomes are promising to develop into therapeutic products for wound healing treatment.

Abstract Objective To increase access to safe and appropriate exercise for people with balance and mobility limitations, community organizations have partnered with healthcare providers to deliver an evidence-based, task-oriented group exercise program in community centers in Canada. We aimed to understand challenges and solutions to implementing this program model to inform plans for expansion. Results At a 1-day meeting, 53 stakeholders (healthcare/recreation personnel, program participants/caregivers, researchers) identified challenges to program implementation that were captured by seven themes: Resources to deliver the exercise class (e.g., difficulty finding instructors with the skills to work with people with mobility limitations); Program marketing (e.g., to foster healthcare referrals); Transportation (e.g., particularly from rural areas); Program access (e.g., program full); Maintaining program integrity; Sustaining partnerships (i.e., with healthcare partners); and Funding (e.g., to deliver program or register). Stakeholders prioritized solutions to form an action plan. A survey of individuals supervising 28 programs revealed that people with stroke, acquired brain injury, multiple sclerosis, and Parkinson’s disease register at 95–100% of centers. The most prevalent issues with program fidelity across centers were not requiring a minimum level of walking ability (32%), class sizes exceeding 12 (21%), and instructor-to-participant ratios exceeding 1:4 (19%). Findings provide considerations for program expansion.

Abstract Background Quantification and identification of microbial genomes based on next-generation sequencing data is a challenging problem in metagenomics. Although current methods have mostly focused on analyzing bacteria whose genomes have been sequenced, such analyses are, however, complicated by the presence of unknown bacteria or bacteria whose genomes have not been sequence. Results We propose a method for detecting unknown bacteria in environmental samples. Our approach is unique in its utilization of short reads only from 16S rRNA genes, not from entire genomes. We show that short reads from 16S rRNA genes retain sufficient information for detecting unknown bacteria in oral microbial communities. Conclusion In our experimentation with bacterial genomes from the Human Oral Microbiome Database, we found that this method made accurate and robust predictions at different read coverages and percentages of unknown bacteria. Advantages of this approach include not only a reduction in experimental and computational costs but also a potentially high accuracy across environmental samples due to the strong conservation of the 16S rRNA gene.

Background: Oncological patients have a higher risk of prolonged SARS-CoV-2 shedding, which, in turn, can lead to evolutionary mutations and emergence of novel viral variants. The aim of this study was to analyze biological samples of a cohort of oncological patients by deep sequencing to detect any significant viral mutations. Methods: High-throughput sequencing was performed on selected samples from a SARS-CoV-2-positive oncological patient cohort. Analysis of variants and minority variants was performed using a validated bioinformatics pipeline. Results: Among 54 oncological patients, we analyzed 12 samples of 6 patients, either serial nasopharyngeal swab samples or samples from the upper and lower respiratory tracts, by high-throughput sequencing. We identified amino acid changes D614G and P4715L as well as mutations at nucleotide positions 241 and 3037 in all samples. There were no other significant mutations, but we observed intra-host evolution in some minority variants, mainly in the ORF1ab gene. There was no significant mutation identified in the spike region and no minority variants common to several hosts. Conclusions: There was no major and rapid evolution of viral strains in this oncological patient cohort, but there was minority variant evolution, reflecting a dynamic pattern of quasi-species replication.

Background: Based on recent findings, we speculated the existence of the lung, heart, and kidney axis as the main pathway for the COVID-19 disease progression. Methods: This paper reports on an observational study conducted by a team of researchers and doctors of the 118-Pre-Hospital and Emergency Department of SG Moscati of Taranto City in Italy. The study was conducted on a totality of 185 participants that were divided into three groups. The study group included COVID-19 affected patients (PP n = 80), the first control group included patients with different pathologies (non-COVID-19 NNp n = 62) of the SG Moscati Hospital, and the second control group included healthy individuals (NNh n = 43). The core of the current trial was focused on assessing the level of the vitamin D (serum 25(OH) D concentration), IL-6, and the renal glomerular filtrate (eGFR) in COVID-19 disease and non-COVID-19 patients in both groups. Results: It was observed that the majority of COVID-19-infected patients showed a progressive multi-organ involvement, especially in regard to the lung, kidney, and heart. The majority of the COVID-19 patients exhibited preexisting comorbidities which include cardiovascular, respiratory, and renal disorders accompanied by a severely low level of vitamin D, extremely high level of IL-6, and low glomerular filtration rate (eGFR). The significant overall damages exerted by the immune-mediated responses under the hyper-expression of proinflammatory cytokines and interleukins, such as IL-6, may be facilitated by either a decreased level of vitamin D or the ageing process. The reduced presence of vitamin D was often found together with a reduced functionality of renal activity, as revealed by the low eGFR, and both were seen to be concomitant with an increased mortality risk in patients with lung disorders and heart failure (HF), whether it is showed at baseline or it develops during manifestation of COVID-19. Therefore, the documentation of the modifiable risk factors related to SARS-CoV-2 and lung impairment in older patients with kidney and heart disease may help the clinician to better manage the situation. Conclusions: This paper addresses how a low level of vitamin D and older age may be indicative of systemic worsening in patients with COVID-19, with a goal of providing a broader context in which to view a better therapeutic approach.

Breast cancer (BC) a very common cancer in women worldwide. Triple negative breast cancer (TNBC) has been shown to have a poor prognosis with a high level of tumor metastatic spread. Here, the inhibitory effects of ginsenoside-Rh1 (Rh1) on BC metastasis, and its underlying signaling pathway in TNBC were investigated. Rh1-treated MDA-MB-231 cells were analyzed for metastasis using a wound healing assay, transwell migration and invasion assay, western blotting, and qRT-PCR. Rh1 treatment significantly inhibited BC metastasis by inhibiting the both protein and mRNA levels of MMP2, MMP9, and VEGF-A. Further, Rh1-mediated inhibitory effect on BC migration was associated with mitochondrial ROS generation. Rh1 treatment significantly eliminated STAT3 phosphorylation and NF-κB transactivation to downregulate metastatic factors, such as MMP2, MMP9, and VEGF-A. In addition, Mito-TEMPO treatment reversed Rh1 effects on the activation of STAT3, NF-κB, and their transcriptional targets. Rh1 further enhanced the inhibitory effects of STAT3 or NF-κB specific inhibitor, stattic or BAY 11-7082 on MMP2, MMP9, and VEGF-A expression, respectively. In summary, our results revealed the potent anticancer effect of Rh1 on TNBC migration and invasion through mtROS-mediated inhibition of STAT3 and NF-κB signaling.

Background: A novel coronavirus (SARS-CoV-2)-induced pneumonia (COVID-19) emerged in December 2019 in China, spreading worldwide. The aim of the present investigation was to evaluate the immunological response and the clinical subset of peripheral lymphocyte subset alteration in COVID-19 infection. Methods: the study was conducted on four different clinical groups (n = 4; total n = 138). Each individual was assigned to different groups based on specific criteria evaluated at the admission such as fever, dyspnea, arterial blood gas analysis (ABG), oral-nasopharyngeal swab/RT-PCR, and thoracic CT-scan. Treatment was performed only after blood samples were collected from each patient (PP and PP) at day 1. The blood samples were analyzed and tested the same day (CBC and Flowcytometry). The positive–positive group (PP n = 45; F = 18/ M = 27; median age = 62.33), comprised individuals affected by COVID-19 who showed fever, dyspnea (ABG = pO2 < 60), confirmed positive by oral-nasopharyngeal swab/RT-PCR and with CT-scan showing ground-glass opacities. The negative–positive (NP; n = 37; F = 11/M = 26; median age = 75.94) or “COVID-like” group comprised individuals with fever and dyspnea (ABG = pO2 < 60), who tested negative to nasopharyngeal swab/RT-PCR, with CT-scans showing ground-glass opacities in the lungs. The negative–affected group (NA; n = 40; F = 14/M = 26; median age = 58.5) included individuals negative to COVID-19 (RT-PCR) but affected by different chronic respiratory diseases (the CT-scans didn’t show ground-glass opacities). Finally, the negative–negative group (NN; n = 16; F = 14/M = 2) included healthy patients (NN; n = 16; median age = 42.62). Data and findings were collected and compared. Results: Lymphocytes (%) cells showed a decline in COVID-19 patients. The subsets showed a significant association with the inflammatory status in COVID-19, especially with regard to increased neutrophils, T-killer, T-active, T-suppressor, and T-CD8+CD38+ in individuals belong to the either COVID-19 and Covid-like NP group. Conclusions: Peripheral lymphocyte subset alteration was associated with the clinical characteristics and progression of COVID-19. The level of sub-set cells T-lymphocytes (either high or low) and B-lymphocytes could be used as an independent predictor for COVID-19 severity and treatment efficacy.

To date, several cases of thrombosis have been confirmed to be related to Sars-CoV-2 infection. Multiple attempts detected the prolonged occurrence of Sars-CoV-2 viral RNA (long COVID) in whole blood suggesting that virus byproducts may remain within cells and tissues well over the disease has finished. Patients may develop severe thrombocytopenia, acute anemia of inflammation and, systemic thrombosis with the fatal course of disease, which is suggestive of further interferences of Sars-CoV-2 on hematopoietic stem cells (HSCs) within the differentiation process towards erythroid and megakaryocytic cells. Therefore, we speculated whether Sars-CoV-2 propagates in or compartmentalizes with hematopoietic progenitor, erythroid, and megakaryocytic cells as the main cause of thrombotic events in either COVID-19 patients or vaccinated individuals. Results: The Sars-CoV-2 RNA replication, protein translation and infectious particle formation as the spike proteins in hematopoietic cell lines take place via the angiotensin-converting enzyme 2 (ACE2) entry pathway within primary CD34+ HSCs inducing, ex vivo, the formation of defected erythroid and megakaryocytic cells that eventually become targets of humoral and adaptive immune cells. Conclusions: Viral particles from affected CD34+ HSCs or the cellular component of RBC units and eventually platelets, present the greatest risk for sever thrombosis-transmitted Sars-CoV-2 infections.

Human astroviruses (HAstV) are understudied positive-strand RNA viruses that cause gastroenteritis mostly in children and the elderly. Three clades of astroviruses, classic, MLB-type and VA-type have been reported in humans. One limitation towards a better understanding of these viruses has been the lack of a physiologically relevant cell culture model that supports growth of all clades of HAstV. Herein, we demonstrate infection of HAstV strains belonging to all three clades in epithelium-only human intestinal enteroids (HIE) isolated from biopsy-derived intestinal crypts. A detailed investigation of infection of VA1, a member of the non-canonical HAstV-VA/HMO clade, showed robust replication in HIE derived from different patients and from different intestinal regions independent of the cellular differentiation status. Flow cytometry and immunofluorescence analysis revealed that VA1 infects several cell types, including intestinal progenitor cells and mature enterocytes, in HIE cultures. RNA profiling of VA1-infected HIE uncovered that the host response to infection is dominated by interferon (IFN)-mediated innate immune responses. A comparison of the antiviral host response in non-transformed HIE and transformed human colon carcinoma Caco-2 cells highlighted significant differences between these cells, including an increased magnitude of the response in HIE. Additional studies confirmed the sensitivity of VA1 to exogenous IFNs, and indicated that the endogenous IFN response of HIE to curtail the growth of strains from all three clades. Genotypic variation in the permissiveness of different HIE lines to HAstV could be overcome by pharmacologic inhibition of JAK/STAT signaling. Collectively, our data identify HIE as a universal infection model for HAstV and an improved model of the intestinal epithelium to investigate enteric virus-host interactions.

Simulated microgravity (SMG) induced the changes in cell proliferation and cytoskeleton organization, which plays an important factor in various cellular processes. The inhibition in cell cycle progression has been considered to be one of the main causes of proliferation inhibition in cells under SMG, but their mechanisms are still not fully understood. This study aimed to evaluate the effects of SMG on the proliferative ability and cytoskeleton changes of Chang Liver Cells (CCL-13). CCL-13 cells were induced SMG by 3D clinostat for 72 h, while the control group were treated in normal gravity at the same time. The results showed that SMG reduced CCL-13 cell proliferation by an increase in the number of CCL-13 cells in G0/G1 phase. This cell cycle phase arrest of CCL-13 cells was due to a downregulation of cell cycle-related proteins, such as cyclin A1 and A2, cyclin D1, and cyclin-dependent kinase 6 (Cdk6). SMG-exposed CCL-13 cells also exhibited a downregulation of α-tubulin 3 and β-actin which induced the cytoskeleton reorganization. These results suggested that the inhibited proliferation of SMG-exposed CCL-13 cells could be associate with the attenuation of major cell cycle regulators and main cytoskeletal proteins.

Ginsenosides have been reported to have various biological effects, such as immune regulation and anticancer activity. In this study, we investigated the anti-inflammatory role of a combination of Rg2 and Rh1, which are minor ginsenosides, in lipopolysaccharide (LPS)-stimulated inflammation. In vitro experiments were performed using the RAW264.7 cell line, and an in vivo model of inflammation was established using LPS-treated ICR mice. We employed Griess assay, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, quantitative reverse transcriptase-polymerase chain reaction, western blotting, immunofluorescence staining, and hematoxylin and eosin staining to evaluate the effect of Rg2 and Rh1. We found that Rg2 and Rh1 significantly decreased LPS-induced major inflammatory mediator production, inducible-nitric oxide synthase expression, and nitric oxide production in macrophages. Moreover, Rg2 and Rh1 combination treatment inhibited the binding of LPS to toll-like receptor 4 (TLR4) on peritoneal macrophages. Therefore, the combination of ginsenoside Rg2 and Rh1 suppressed inflammation by abolishing the binding of LPS to TLR4, thereby inhibiting the TLR4-mediated signaling pathway. The combined ginsenoside synergistically blocked LPS-mediated PKCδ translocation to the plasma membrane, resulting in p38-STAT1 activation and NF-κB translocation. In addition, mRNA levels of pro-inflammatory cytokines, including TNF-α, IL-1β, and IFN-β, were significantly decreased by combined ginsenoside treatment. Notably, the 20 mg/kg ginsenoside treatment significantly reduced LPS-induced acute tissue inflammation levels in vivo, as indicated by the tissue histological damage scores and the levels of biochemical markers for liver and kidney function from mouse serum. These results suggest that the minor ginsenosides Rg2 and Rh1 may play a key role in prevention of LPS-induced acute inflammation and tissue damage.

The increasing incidence of community- and hospital-acquired infections with multidrug-resistant (MDR) bacteria poses a critical threat to public health and the healthcare system. Although β-lactam antibiotics are effective against most bacterial infections, some bacteria are resistant to β-lactam antibiotics by producing β-lactamases. Among β-lactamases, metallo-β-lactamases (MBLs) are especially worrisome as only a few inhibitors have been developed against them. In MBLs, the metal ions play an important role as they coordinate a catalytic water molecule that hydrolyzes β-lactam rings. We determined the crystal structures of different variants of PNGM-1, an ancient MBL with additional tRNase Z activity. The variants were generated by site-directed mutagenesis targeting metal-coordinating residues. In PNGM-1, both zinc ions are coordinated by six coordination partners in an octahedral geometry, and the zinc-centered octahedrons share a common face. Structures of the PNGM-1 variants confirm that the substitution of a metal-coordinating residue causes the loss of metal binding and β-lactamase activity. Compared with PNGM-1, subclass B3 MBLs lack one metal-coordinating residue, leading to a shift in the metal-coordination geometry from an octahedral to tetrahedral geometry. Our results imply that a subtle change in the metal-binding site of MBLs can markedly change their metal-coordination geometry and catalytic activity.

Dendrobium has been widely used not only as ornamental plants but also as food and medicines. The identification and evaluation of the genetic diversity of Dendrobium species support the conservation of genetic resources of endemic Dendrobium species. Uniquely identifying Dendrobium species used as medicines helps avoid misuse of medicinal herbs. However, it is challenging to identify Dendrobium species morphologically during their immature stage. Based on the DNA barcoding method, it is now possible to efficiently identify species in a shorter time. In this study, the genetic diversity of 76 Dendrobium samples from Southern Vietnam was investigated based on the ITS (Internal transcribed spacer), ITS2, matK (Maturase_K), rbcL (ribulose-bisphosphate carboxylase large subunit) and trnH-psbA (the internal space of the gene coding histidine transfer RNA (trnH) and gene coding protein D1, a polypeptide of the photosystem I reaction center (psaB)) regions. The ITS region was found to have the best identification potential. Nineteen out of 24 Dendrobium species were identified based on phylogenetic tree and Indel information of this region. Among these, seven identified species were used as medicinal herbs. The results of this research contributed to the conservation, propagation, and hybridization of indigenous Dendrobium species in Southern Vietnam.

Tripeptide glutathione, which plays important roles in many cellular mechanisms, is also a biotechnology-oriented molecule with applications in medicine, food and cosmetic. Here, the engineering of the yeast Yarrowia lipolytica for the production of this metabolite at high titer values from various agro-industrial by-products is reported. The constitutive overexpression of the glutathione biosynthetic genes GSH1 and GSH2 encoding respectively γ-glutamylcysteine synthetase and glutathione synthetase, together with the INU1 gene from Kluyveromyces marxianus encoding inulinase yielded a glutathione titer value and a productivity of 644 nmol/mg protein and 510 µmol/gDCW, respectively. These values were obtained during bioreactor batch cultures in a medium exclusively comprising an extract of Jerusalem artichoke tuber, used as a source of inulin, and ammonium sulfate, used as a nitrogen source.

(1) Background: Dendritic cell (DC) vaccination has shown outstanding achievements in cancer treatment, although it still has some adverse side effects. Vaccination with DC-derived exosomes has been thought to overcome the side effects of the parental DCs. (2) Method: We performed the experiments to check the ability of cryopreserved umbilical cord blood mononuclear cell-derived DCs (cryo CBMDCs) and their exosomes to prime allogeneic T cell proliferation and allogeneic peripheral blood mononuclear cell (alloPBMCs) cytotoxicity against A549 lung cancer cells. (3) Results: We found that both lung tumor cell lysate-pulsed DCs and their exosomes could induce allogeneic T cell proliferation. Moreover, alloPBMCs primed with tumor cell lysate-pulsed DCs and their exosomes have a greater cytotoxic activity against A549 cells compared to unprimed cells and cells primed with unpulsed DCs and their exosomes. (4) Conclusion: Tumor cell lysate-pulsed DCs and their exosomes should be considered to develop into a novel immunotherapeutic strategy—e.g., vaccines—for patients with lung cancer. Our results also suggested that cryo umbilical cord blood mononuclear cells source, which is a readily and available source, is effective for generation of allogeneic DCs and their exosomes will be material for vaccinating against cancer.

Paphiopedilum is among the most popular ornamental orchid genera due to its unique slipper flowers and attractive leaf coloration. Most of the Paphiopedilum species are in critical danger due to over-exploitation. They were listed in Appendix I of the Convention on International Trade in Endangered Species of Wild Fauna and Flora, which prevents their being traded across borders. While most Paphiopedilum species are distinctive, owing to their respective flowers, their vegetative features are more similar and undistinguished. Hence, the conservation of these species is challenging, as most traded specimins are immature and non-flowered. An urgent need exists for effective identification methods to prevent further illegal trading of Paphiopedilum species. DNA barcoding is a rapid and sensitive method for species identification, at any developmental stage, using short DNA sequences. In this study, eight loci, i.e., ITS, LEAFY, ACO, matK, trnL, rpoB, rpoC1, and trnH-psbA, were screened for potential barcode sequences on the Vietnamese Paphiopedilum species. In total, 17 out of 22 Paphiopedilum species were well identified. The studied DNA sequences were deposited to GenBank, in which Paphiopedilum dalatense accessions were introduced for the first time. ACO, LEAFY, and trnH-psbA were limited in amplification rate for Paphiopedilum. ITS was the best single barcode. Single ITS could be used along with nucleotide polymorphism characteristics for species discrimination. The combination of ITS + matK was the most efficient identification barcode for Vietnamese Paphiopedilum species. This barcode also succeeded in recognizing misidentified or wrongly-named traded samples. Different bioinformatics programs and algorithms for establishing phylogenetic trees were also compared in the study to propose quick, simple, and effective tools for practical use. It was proved that both the Bayesian Inference method in the MRBAYES program and the neighbor-joining method in the MEGA software met the criteria. Our study provides a barcoding database of Vietnamese Paphiopedilum which may significantly contribute to the control and conservation of these valuable species.

Non-conventional yeasts are efficient cell factories for the synthesis of value-added compounds such as recombinant proteins, intracellular metabolites, and/or metabolic by-products. Most bioprocess, however, are still designed to use pure, ideal sugars, especially glucose. In the quest for the development of more sustainable processes amid concerns over the future availability of resources for the ever-growing global population, the utilization of organic wastes or industrial by-products as feedstocks to support cell growth is a crucial approach. Indeed, vast amounts of industrial and commercial waste simultaneously represent an environmental burden and an important reservoir for recyclable or reusable material. These alternative feedstocks can provide microbial cell factories with the required metabolic building blocks and energy to synthesize value-added compounds, further representing a potential means of reduction of process costs as well. This review highlights recent strategies in this regard, encompassing knowledge on catabolic pathways and metabolic engineering solutions developed to endow cells with the required metabolic capabilities, and the connection of these to the synthesis of value-added compounds. This review focuses primarily, but not exclusively, on Yarrowia lipolytica as a yeast cell factory, owing to its broad range of naturally metabolizable carbon sources, together with its popularity as a non-conventional yeast.

Neuronal subpopulations display differential vulnerabilities to disease, but the factors that determine their susceptibility are poorly understood. Toxic increases in intracellular calcium are a key factor in several neurodegenerative processes, with calcium-binding proteins providing an important first line of defense through their ability to buffer incoming calcium, allowing the neuron to quickly achieve homeostasis. Since neurons expressing different calcium-binding proteins have been reported to be differentially susceptible to degeneration, it can be hypothesized that rather than just serving as markers of different neuronal subpopulations, they might actually be a key determinant of survival. In this review, we will summarize some of the evidence that expression of the EF-hand calcium-binding proteins, calbindin, calretinin and parvalbumin, may influence the susceptibility of distinct neuronal subpopulations to disease processes.

The advancement of bioinformatics and machine learning has facilitated the discovery and validation of omics-based biomarkers. This study employed a novel approach combining multi-platform transcriptomics and cutting-edge algorithms to introduce novel signatures for accurate diagnosis of colorectal cancer (CRC). Different random forests (RF)-based feature selection methods including the area under the curve (AUC)-RF, Boruta, and Vita were used and the diagnostic performance of the proposed biosignatures was benchmarked using RF, logistic regression, naïve Bayes, and k-nearest neighbors models. All models showed satisfactory performance in which RF appeared to be the best. For instance, regarding the RF model, the following were observed: mean accuracy 0.998 (standard deviation (SD) < 0.003), mean specificity 0.999 (SD < 0.003), and mean sensitivity 0.998 (SD < 0.004). Moreover, proposed biomarker signatures were highly associated with multifaceted hallmarks in cancer. Some biomarkers were found to be enriched in epithelial cell signaling in Helicobacter pylori infection and inflammatory processes. The overexpression of TGFBI and S100A2 was associated with poor disease-free survival while the down-regulation of NR5A2, SLC4A4, and CD177 was linked to worse overall survival of the patients. In conclusion, novel transcriptome signatures to improve the diagnostic accuracy in CRC are introduced for further validations in various clinical settings.

Why does a 4-H club (for youth ages 8–18) need bylaws? For the same reason our country needs a constitution! By having fair and written rules, the rights of all members can be protected. The following is a suggested format for 4-H bylaws. The bylaws below may be adapted to meet local conditions. Be sure to send a copy of your club’s approved bylaws to your county 4-H office. Include in the bylaws the date when they were approved and most recently revised. This 3-page fact sheet was written by Keith G. Diem, Judy Levings, and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H338/4H338: Adopting 4-H Club Bylaws (ufl.edu)

The UF/IFAS Extension 4-H Youth Development Program is committed to the safety and well-being of its youth as well as the volunteers who help deliver 4-H programs under the supervision of 4-H faculty and staff. All things in life involve some sort of risk. This is especially true when working with youth, but there are many ways that volunteers can minimize this risk and lead safe 4-H activities. Proper planning and common sense certainly go a long way. Although not to be considered legal advice, here are some facts and issues to know and keep in mind while serving as a 4-H volunteer. This 3-page fact sheet was written by Keith G. Diem, and published by the UF Department of 4-H Youth Development, July 2014. 4H342/4H342: Protecting Youth and Volunteers by Planning Ahead, Reducing Risk (ufl.edu)

Working with youth, especially other people’s, can be a challenge. While some are better behaved than others, they all need acceptance and the opportunity to learn. Especially in groups of mixed ages, experience, or interests, the challenge of a youth leader is to see that disruptive youth do not prevent others from learning or having fun. Their very presence in the group, combined with your wise guidance, will help them learn what is socially acceptable and how to do what is right. This information sheet will help you to understand youth better and to provide some ideas on how to deal with the unruly kids with minimal disruption of the learning situation. This 5-page fact sheet was written by Keith G. Diem, Heidi L. Radunovich, and Georgene Bender, and published by the UF Department of 4-H Youth Development, July 2014. 4H341/4H341: Positive Discipline for Youth (ufl.edu)

Field trips can be great experiences for youth to learn project content and life skills. Field trips also contribute to the experiential learning philosophy of 4-H. This document includes some suggestions to make a field trip fun, safe, and educational. This 3-page fact sheet was written by Keith G. Diem and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H339/4H339: Planning a Successful Field Trip (ufl.edu)

As a 4-H volunteer, one of your most important roles is helping youth learn. Especially because membership in 4-H is voluntary, learning in 4-H must be fun. Even if you’re not a teacher by profession, you can successfully guide youth to learn communication, leadership, citizenship, and other life skills, as well as subject matter through their 4-H projects and club participation. This 5-page fact sheet was written by Keith G. Diem and Judy Levings, and published by the UF Department of 4-H Youth Development, July 2014. 4H340/4H340: Making Learning Fun (ufl.edu)

This information sheet is the introduction to the 4-H Volunteer Training Series (VTS). The series contains a wealth of information, from how to start a club, to planning field trips, to how to work with youth. This 5-page fact sheet was written by Keith G. Diem, Sarah Hensley, and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H335/4H335: The ABCs of 4-H: A Primer for 4-H Volunteers (ufl.edu)

The UF/IFAS Extension 4-H Youth Development Program is committed to the safety and well-being of its youth as well as the volunteers who help deliver 4-H programs under the supervision of 4-H faculty and staff. All things in life involve some sort of risk. This is especially true when working with youth, but there are many ways that volunteers can minimize this risk and lead safe 4-H activities. Proper planning and common sense certainly go a long way. Although not to be considered legal advice, here are some facts and issues to know and keep in mind while serving as a 4-H volunteer. This 3-page fact sheet was written by Keith G. Diem, and published by the UF Department of 4-H Youth Development, July 2014. 4H342/4H342: Protecting Youth and Volunteers by Planning Ahead, Reducing Risk (ufl.edu)

Working with youth, especially other people’s, can be a challenge. While some are better behaved than others, they all need acceptance and the opportunity to learn. Especially in groups of mixed ages, experience, or interests, the challenge of a youth leader is to see that disruptive youth do not prevent others from learning or having fun. Their very presence in the group, combined with your wise guidance, will help them learn what is socially acceptable and how to do what is right. This information sheet will help you to understand youth better and to provide some ideas on how to deal with the unruly kids with minimal disruption of the learning situation. This 5-page fact sheet was written by Keith G. Diem, Heidi L. Radunovich, and Georgene Bender, and published by the UF Department of 4-H Youth Development, July 2014. 4H341/4H341: Positive Discipline for Youth (ufl.edu)

Why does a 4-H club (for youth ages 8–18) need bylaws? For the same reason our country needs a constitution! By having fair and written rules, the rights of all members can be protected. The following is a suggested format for 4-H bylaws. The bylaws below may be adapted to meet local conditions. Be sure to send a copy of your club’s approved bylaws to your county 4-H office. Include in the bylaws the date when they were approved and most recently revised. This 3-page fact sheet was written by Keith G. Diem, Judy Levings, and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H338/4H338: Adopting 4-H Club Bylaws (ufl.edu)

As a 4-H volunteer, one of your most important roles is helping youth learn. Especially because membership in 4-H is voluntary, learning in 4-H must be fun. Even if you’re not a teacher by profession, you can successfully guide youth to learn communication, leadership, citizenship, and other life skills, as well as subject matter through their 4-H projects and club participation. This 5-page fact sheet was written by Keith G. Diem and Judy Levings, and published by the UF Department of 4-H Youth Development, July 2014. 4H340/4H340: Making Learning Fun (ufl.edu)

This information sheet is the introduction to the 4-H Volunteer Training Series (VTS). The series contains a wealth of information, from how to start a club, to planning field trips, to how to work with youth. This 5-page fact sheet was written by Keith G. Diem, Sarah Hensley, and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H335/4H335: The ABCs of 4-H: A Primer for 4-H Volunteers (ufl.edu)

Field trips can be great experiences for youth to learn project content and life skills. Field trips also contribute to the experiential learning philosophy of 4-H. This document includes some suggestions to make a field trip fun, safe, and educational. This 3-page fact sheet was written by Keith G. Diem and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H339/4H339: Planning a Successful Field Trip (ufl.edu)

The UF/IFAS Extension 4-H Youth Development Program is committed to the safety and well-being of its youth as well as the volunteers who help deliver 4-H programs under the supervision of 4-H faculty and staff. All things in life involve some sort of risk. This is especially true when working with youth, but there are many ways that volunteers can minimize this risk and lead safe 4-H activities. Proper planning and common sense certainly go a long way. Although not to be considered legal advice, here are some facts and issues to know and keep in mind while serving as a 4-H volunteer. This 3-page fact sheet was written by Keith G. Diem, and published by the UF Department of 4-H Youth Development, July 2014. 4H342/4H342: Protecting Youth and Volunteers by Planning Ahead, Reducing Risk (ufl.edu)

Why does a 4-H club (for youth ages 8–18) need bylaws? For the same reason our country needs a constitution! By having fair and written rules, the rights of all members can be protected. The following is a suggested format for 4-H bylaws. The bylaws below may be adapted to meet local conditions. Be sure to send a copy of your club’s approved bylaws to your county 4-H office. Include in the bylaws the date when they were approved and most recently revised. This 3-page fact sheet was written by Keith G. Diem, Judy Levings, and Ben Knowles, and published by the UF Department of 4-H Youth Development, July 2014. 4H338/4H338: Adopting 4-H Club Bylaws (ufl.edu)

Working with youth, especially other people’s, can be a challenge. While some are better behaved than others, they all need acceptance and the opportunity to learn. Especially in groups of mixed ages, experience, or interests, the challenge of a youth leader is to see that disruptive youth do not prevent others from learning or having fun. Their very presence in the group, combined with your wise guidance, will help them learn what is socially acceptable and how to do what is right. This information sheet will help you to understand youth better and to provide some ideas on how to deal with the unruly kids with minimal disruption of the learning situation. This 5-page fact sheet was written by Keith G. Diem, Heidi L. Radunovich, and Georgene Bender, and published by the UF Department of 4-H Youth Development, July 2014. 4H341/4H341: Positive Discipline for Youth (ufl.edu)

As a 4-H volunteer, one of your most important roles is helping youth learn. Especially because membership in 4-H is voluntary, learning in 4-H must be fun. Even if you’re not a teacher by profession, you can successfully guide youth to learn communication, leadership, citizenship, and other life skills, as well as subject matter through their 4-H projects and club participation. This 5-page fact sheet was written by Keith G. Diem and Judy Levings, and published by the UF Department of 4-H Youth Development, July 2014. 4H340/4H340: Making Learning Fun (ufl.edu)