Norcantharidin-Encapsulated C60-Modified Nanomicelles: A Potential Approach to Mitigate Cytotoxicity in Renal Cells and Simultaneously Enhance Anti-Tumor Activity in Hepatocellular Carcinoma Cells.

OBJECTIVE: The objective of this study was to examine the preparation process of DSPE-PEG-C60/NCTD micelles and assess the impact of fullerenol (C60)-modified micelles on the nephrotoxicity and antitumor activity of NCTD. METHOD: The micelles containing NCTD were prepared using the ultrasonic method and subsequently optimized and characterized. The cytotoxicity of micelles loaded with NCTD was assessed using the CCK-8 method on human hepatoma cell lines HepG2 and BEL-7402, as well as normal cell lines HK-2 and L02. Acridine orange/ethidium bromide (AO/EB) double staining and flow cytometry were employed to assess the impact of NCTD-loaded micelles on the apoptosis of the HK-2 cells and the HepG2 cells. Additionally, JC-1 fluorescence was utilized to quantify the alterations in mitochondrial membrane potential. The generation of reactive oxygen species (ROS) following micelle treatment was determined through 2',7'-dichlorofluorescein diacetate (DCFDA) staining. RESULTS: The particle size distribution of the DSPE-PEG-C60/NCTD micelles was determined to be 91.57 nm (PDI = 0.231). The zeta potential of the micelles was found to be -13.8 mV. The encapsulation efficiency was measured to be 91.9%. The in vitro release behavior of the micelles followed the Higuchi equation. Cellular experiments demonstrated a notable decrease in the toxicity of the C60-modified micelles against the HK-2 cells, accompanied by an augmented inhibitory effect on cancer cells. Compared to the free NCTD group, the DSPE-PEG-C60 micelles exhibited a decreased apoptosis rate (12%) for the HK-2 cell line, lower than the apoptosis rate observed in the NCTD group (36%) at an NCTD concentration of 75 µM. The rate of apoptosis in the HepG2 cells exhibited a significant increase (49%), surpassing the apoptosis rate observed in the NCTD group (24%) at a concentration of 150 µM NCTD. The HK-2 cells exhibited a reduction in intracellular ROS and an increase in mitochondrial membrane potential (ΔψM) upon exposure to C60-modified micelles compared to the NCTD group. CONCLUSIONS: The DSPE-PEG-C60/NCTD micelles, as prepared in this study, demonstrated the ability to decrease cytotoxicity and ROS levels in normal renal cells (HK-2) in vitro. Additionally, these micelles showed an enhanced antitumor activity against human hepatocellular carcinoma cells (HepG2, BEL-7402).

Computational, Experimental, and Clinical Evidence of a Specific but Peculiar Evolutionary Nature of (COVID-19) SARS-CoV-2.

The shell disorder models have predicted that SARS-CoV-2 is of a specific but peculiar evolutionary nature. All coronaviruses (CoVs) closely related to SARS-CoV-2 have been found to have the hardest outer shells (M protein) among CoVs. This hard shell (low M percentage of intrinsic disorder (PID)) is associated with burrowing animals, for example, pangolins, and is believed to be responsible for the high contagiousness of SARS-CoV-2 because it will be more resistant to antimicrobial enzymes found in saliva/mucus. Incoming clinical and experimental data do support this along with a prediction based on another aspect of the shell (N, inner shell) disorder models that SARS-CoV-1 is more virulent than SARS-CoV-2 because SARS-CoV-2 produces fewer virus copies in vital organs even if large amounts of infections particles are shed orally and nasally. A phylogenetic study using M reveals a closer relationship of SARS-CoV to pangolin-CoVs than the bat-RaTG13 found in Yunnan, China. Previous studies may have been confused by recombinations that were poorly handled. The shell disorder models suggest that a pangolin-CoV strain may have entered the human population in 2017 or before as an attenuated virus, which could explain why SARS-CoV is found to be highly adapted to humans.

Shell Disorder Analysis Suggests That Pangolins Offered a Window for a Silent Spread of an Attenuated SARS-CoV-2 Precursor among Humans.

A model to predict the relative levels of respiratory and fecal-oral transmission potentials of coronaviruses (CoVs) by measuring the percentage of protein intrinsic disorder (PID) of the M (Membrane) and N (Nucleoprotein) proteins in their outer and inner shells, respectively, was built before the MERS-CoV outbreak. With MPID = 8.6% and NPID = 50.2%, the 2003 SARS-CoV falls into group B, which consists of CoVs with intermediate levels of both fecal-oral and respiratory transmission potentials. Further validation of the model came with MERS-CoV (MPID = 9%, NPID = 44%) and SARS-CoV-2 (MPID = 5.5%, NPID = 48%) falling into the groups C and B, respectively. Group C contains CoVs with higher fecal-oral but lower respiratory transmission potentials. Unlike SARS-CoV, SARS-CoV-2 with MPID = 5.5% has one of the hardest outer shells among CoVs. Because the hard shell is able to resist the antimicrobial enzymes in body fluids, the infected person is able to shed large quantities of viral particles via saliva and mucus, which could account for the higher contagiousness of SARS-COV-2. Further searches have found that high rigidity of the outer shell is characteristic for the CoVs of burrowing animals, such as rabbits (MPID = 5.6%) and pangolins (MPID = 5-6%), which are in contact with the buried feces. A closer inspection of pangolin-CoVs from 2017 to 2019 reveals that pangolins provided a unique window of opportunity for the entry of an attenuated SARS-CoV-2 precursor into the human population in 2017 or earlier, with the subsequent slow and silent spread as a mild cold that followed by its mutations into the current more virulent form. Evidence of this lies in both the genetic proximity of the pangolin-CoVs to SARS-CoV-2 (∼90%) and differences in N disorder. A 2017 pangolin-CoV strain shows evidence of higher levels of attenuation and higher fecal-oral transmission associated with lower human infectivity via having lower NPID (44.8%). Our shell disorder model predicts this to be a SARS-CoV-2 vaccine strain, as lower inner shell disorder is associated with the lesser virulence in a variety of viruses.

A Novel Strategy for the Development of Vaccines for SARS-CoV-2 (COVID-19) and Other Viruses Using AI and Viral Shell Disorder.

A model that predicts levels of coronavirus (CoV) respiratory and fecal-oral transmission potentials based on the shell disorder has been built using neural network (artificial intelligence, AI) analysis of the percentage of disorder (PID) in the nucleocapsid, N, and membrane, M, proteins of the inner and outer viral shells, respectively. Using primarily the PID of N, SARS-CoV-2 is grouped as having intermediate levels of both respiratory and fecal-oral transmission potentials. Related studies, using similar methodologies, have found strong positive correlations between virulence and inner shell disorder among numerous viruses, including Nipah, Ebola, and Dengue viruses. There is some evidence that this is also true for SARS-CoV-2 and SARS-CoV, which have N PIDs of 48% and 50%, and case-fatality rates of 0.5-5% and 10.9%, respectively. The underlying relationship between virulence and respiratory potentials has to do with the viral loads of vital organs and body fluids, respectively. Viruses can spread by respiratory means only if the viral loads in saliva and mucus exceed certain minima. Similarly, a patient is likelier to die when the viral load overwhelms vital organs. Greater disorder in inner shell proteins has been known to play important roles in the rapid replication of viruses by enhancing the efficiency pertaining to protein-protein/DNA/RNA/lipid bindings. This paper suggests a novel strategy in attenuating viruses involving comparison of disorder patterns of inner shells (N) of related viruses to identify residues and regions that could be ideal for mutation. The M protein of SARS-CoV-2 has one of the lowest M PID values (6%) in its family, and therefore, this virus has one of the hardest outer shells, which makes it resistant to antimicrobial enzymes in body fluid. While this is likely responsible for its greater contagiousness, the risks of creating an attenuated virus with a more disordered M are discussed.

Cryptotanshinone reverses the epithelial-mesenchymal transformation process and attenuates bleomycin-induced pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fibrotic interstitial pneumonia that causes pulmonary tissue damage and functional impairment. To investigate the effects of cryptotanshinone on pulmonary fibrosis, the expression of NIH/3T3, HPF, and rat primary pulmonary fibroblasts was measured and found to be inhibited by CPT in a time- and concentration-dependent manner, and the upregulation of α-SMA expression in NIH/3T3 and HPF cells, which had been stimulated by TGFß-1, was decreased after CPT administration. We observed that CPT could reverse the increase in α-SMA expression and vimentin and the decrease in E-cad expression in A549 cells, which had been induced by 5 ng/mL TGFß-1, indicating that CPT has inhibitory effects in the EMT process. A BLM-induced pulmonary fibrosis model was established in C57BL/6 mice. The lung coefficient and hydroxyproline content increased significantly in the BLM-induced group and were decreased in the CPT-treated group. The expression levels of collagen-I and α-SMA and the phosphorylation level of Stat3 were significantly increased, and CPT treatment decreased these levels. Furthermore, the results from the flow cytometry analysis indicated that, in lung tissues, the frequencies of MDSCs, macrophages, DCs and T cells were considerably increased in the BLM-induced group, while CPT treatment reduced these immunocyte populations.

Comparative toxicity of Paraquat herbicide and some plant extracts in Lymnaea natalensis snails.

Paraquat has been shown to be a highly toxic compound for humans and animals, and many cases of acute poisoning and death have been reported over the past few decades. The present study was undertaken to evaluate comprehensively herbicides (Paraquat) and some plant extracts to biochemical aspects of Lymnaea natalensis snails. It was found that the exposure of L. natalensis to Paraquat and plant extracts led to a significant reduction in the infectivity of Fasciola gigantica miracidia to the snail. The glucose level in hemolymph of exposed snails was elevated, while the glycogen showed a decrease in soft tissues when compared with the control group. In addition, the activity level of some enzymes representing glycolytic enzymes as hexokinase (HK), pyruvate kinase (PK), phosphofructokinase (PFK), lactate dehydrogenase (LDH), and glucose phosphate isomerase (GPI) in snail's tissues were reduced in response to the treatment. It was concluded that the pollution of the aquatic environment by herbicide would adversely affect the metabolism of the L. natalensis snails. Snails treated with Agave attenuate, Ammi visnaga, and Canna iridiflora plant had less toxic effect compared to snails treated with Paraquat.

Shifting Nicotiana attenuata's diurnal rhythm does not alter its resistance to the specialist herbivore Manduca sexta.

Arabidopsis thaliana plants are less resistant to attack by the generalist lepidopteran herbivore Trichoplusia ni when plants and herbivores are entrained to opposite, versus identical diurnal cycles and tested under constant conditions. This effect is associated with circadian fluctuations in levels of jasmonic acid, the transcription factor MYC2, and glucosinolate contents in leaves. We tested whether a similar effect could be observed in a different plant-herbivore system: the wild tobacco Nicotiana attenuata and its co-evolved specialist herbivore, Manduca sexta. We measured larval growth on plants under both constant and diurnal conditions following identical or opposite entrainment, profiled the metabolome of attacked leaf tissue, quantified specific metabolites known to reduce M. sexta growth, and monitored M. sexta feeding activity under all experimental conditions. Entrainment did not consistently affect M. sexta growth or plant defense induction. However, both were reduced under constant dark conditions, as was M. sexta feeding activity. Our data indicate that the response induced by M. sexta in N. attenuata is robust to diurnal cues and independent of plant or herbivore entrainment. We propose that while the patterns of constitutive or general damage-induced defense may undergo circadian fluctuation, the orchestration of specific induced responses is more complex.

Glycogen synthase kinase 3 inhibitor attenuates endotoxin-induced liver injury.

BACKGROUND/AIMS: Endotoxin (lipopolysaccharide, LPS)-induced acute liver injury was attenuated by endotoxin tolerance (ET), which is characterized by phosphatidylinositol 3-kinase pathway/Akt signaling. Glycogen synthase kinase 3 (GSK-3) acts downstream of phosphatidylinositol 3-kinase pathway/Akt and GSK-3 inhibitor protects against organic injury. This study evaluates the hypothesis that ET attenuated LPS-induced liver injury through inhibiting GSK-3 functional activity and downstream signaling. METHODS: Sprague-Dawley rats with or without low-dose LPS pretreatment were challenged with or without large dose of LPS and subsequently received studies. Serum tumor necrosis factor-alpha, interleukin-10, alanine aminotransferase, lactate dehydrogenase, and total bilirubin levels were analyzed, morphology of liver tissue was performed, glycogen content, myeloperoxidase content, phagocytosis activity of Kupffer cells, and the expression and inhibitory phosphorylation as well as kinase activity of GSK-3 were examined. Survival after LPS administration was also determined. RESULTS: LPS induced significant increases of serum TNF-α, alanine aminotransferase, lactate dehydrogenase, and total bilirubin (P < 0.05), which were companied by obvious alterations in liver: the injury of liver tissue, the decrease of glycogen, the infiltration of neutrophils, and the enhancement of phagocytosis of Kupffer cells (P < 0.05). LPS pretreatment significantly attenuated these alterations, promoted the inhibitory phosphorylation of GSK-3 and inhibited its kinase activity, and improved the survival rate (P < 0.05). CONCLUSIONS: ET attenuated LPS-induced acute liver injury through inhibiting GSK-3 functional activity and its downstream signaling.

A novel visual marker to distinguish haploids from doubled haploids in rice (Oryza sativa, L) at early growth stages.

Doubled haploid technology, which enables the generation of homozygous lines in a single step, is one of the modern tools being employed for accelerating breeding processes in different crops. In rice, a globally important staple food crop, doubled haploid production through androgenesis is increasingly being employed in breeding programs. Amongst the androgenic rice lines, doubled haploids are formed spontaneously at about 50-60%, while the remaining 40-50% of plants remain as haploids. As haploids cannot be easily identified, it is routine to grow all the rice androgenic lines till maturity and harvest the seeds from the fertile doubled haploids. Therefore, the methods that facilitate easy identification of haploids at an early developmental stage in rice would enable treatment of such haploid lines with colchicine, to increase the efficiency of doubled haploid production. Further, it would also help in eliminating the operational cost involved in maintaining them till maturity. In the above context, a systematic study to identify easily observable physiological and morphological differences between haploid and doubled haploid rice lines was undertaken. Rice haploids were found to be noticeably different from doubled haploids in photosynthetic rate, transpiration rate, stomatal conductance, and morphology of lodicules, stigma and style, features which have not been reported before. Most importantly, rice haploids invariably have acute leaf apex which is easily distinguishable from the doubled haploids that have attenuated leaf apex shape. Very high per cent accuracy in the prediction of ploidy level was observed when haploids were identified at an early developmental stage based on leaf apex shape, and the results verified with flow cytometry perfectly matches with leaf apex shape. The study establishes 'acute leaf apex' shape as an accurate visual marker to rapidly identify haploid rice lines at an early developmental stage in a cost-effective manner.

Methods to mitigate Escherichia coli blooms in human ex vivo colon model experiments using the high throughput micro-Matrix bioreactor fermentation system.

Ex vivo colon model experiments are frequently employed as a means to assess the gut microbiome modulating potential of different foods, food ingredients and dietary supplements. A number of useful models already exist; however, they tend to be relatively low in terms of throughput (3-4 samples per experiment) with a long experiment duration of one to a number of weeks. Therefore, a need for a high-throughput system with a short duration time is required to enable screening of large numbers of samples. Therefore, we report here on the development of a system based on the Applikon micro-Matrix bioreactor which has the capacity to run 24 samples with an experiment duration of 48 h. However, Escherichia coli blooms are a common problem encountered in this model. Here, we describe the factors that contribute to such blooms and provide approaches to address them, providing:•Step by step optimisation of processes involved in conducting ex vivo distal colon experiments using the micro-Matrix bioreactor fermentation platform•Recommended steps for users on how to attenuate E. coli blooms in such ex vivo colon model experiments.

Vaccines to Prevent Coccidioidomycosis: A Gene-Deletion Mutant of Coccidioides Posadasii as a Viable Candidate for Human Trials.

Coccidioidomycosis is an endemic fungal infection that is reported in up to 20,000 persons per year and has an economic impact close to $1.5 billion. Natural infection virtually always confers protection from future exposure, and this suggests that a preventative vaccine strategy is likely to succeed. We here review progress toward that objective. There has been ongoing research to discover a coccidioidal vaccine over the past seven decades, including one phase III clinical trial, but for reasons of either efficacy or feasibility, a safe and effective vaccine has not yet been developed. This review first summarizes the past research to develop a coccidioidal vaccine. It then details the evidence that supports a live, gene-deletion vaccine candidate as suitable for further development as both a veterinary and a human clinical product. Finally, a plausible vaccine development plan is described which would be applicable to this vaccine candidate and also useful to other future candidates. The public health and economic impact of coccidioidomycosis fully justifies a public private partnership for vaccine development, and the development of a vaccine for this orphan disease will likely require some degree of public funding.

The regulation of Alternaria alternata resistance by LRR-RK4 through ERF109, defensin19 and phytoalexin scopoletin in Nicotiana attenuata.

Leucine-rich repeat receptor-like kinases (LRR-RKs), belonging to the largest subfamily of transmembrane receptor-like kinases in plants, are proposed to be involved in pathogen resistance. However, it is currently unknown whether LRR-RKs regulate Nicotiana attenuata resistance to Alternaria alternata, a notorious fungal pathogen causing tobacco brown disease. During transcriptome analysis, we identified a highly induced receptor kinase (NaLRR-RK4) in N. attenuata leaves after A. alternata inoculation. We speculated that this NaLRR-RK4 might be the resistance gene of tobacco to brown spot disease, and if so, what is its function and mechanism of action? Silencing of NaLRR-RK4 via virus-induced gene silencing (VIGS) lead to plants highly susceptible to A. alternata, and this result was further confirmed by two stable transformation lines (NaLRR-RK4-RNAi lines) generated by RNA interference technology. The susceptible of NaLRR-RK4-RNAi lines to A. alternata was associated with reduced levels of phytoalexin scopoletin and its key synthesis gene NaF6'H1. Further transcriptome analysis of leaves of WT and NaLRR-RK4-RNAi line after A. alternata inoculation revealed that NaLRR-RK4 regulated NaERF109 and NaDEF19. Silencing NaERF109 or NaDEF19 by VIGS lead to plants more susceptible to A.alternata, demonstrating their role in pathogen resistance. Interestingly, A.alternata-induced expression of NaF6'H1 and NaDEF19 were dramatically reduced in NaERF109-silenced VIGS plants. Taken all together, we identified LRR-RK4 as the first Leucine-rich repeat receptor-like kinases involved in A.alternata resistance in tobacco species, by regulating NaERF109, and subsequently NaDEF19 and NaF6'H1.

Developing a screening test for toxicity studies of prenatal development with the use of Hydra attenuata and embryos of zebrafish.

11 active substances used in pesticides were selected. Substances were divided into three groups depending the effect on embryos or fetuses of laboratory animals: 1 - damaging effect on embryos or fetuses (embryotoxic, fetotoxic or teratogenic), 2 - damaging effect on embryos or fetuses, but only at dose toxic for mother (maternal toxicity), 3 - no damaging effect. Changes for hydra in acute toxicity tests and recovery tests were assessed on an change scale from 0 to 10. The index of the effect on development (TI) for hydras was calculated for every compound. Changes in zebrafish embryos were assessed using a descriptive method. Pearson correlation coefficient showed the correlation between the concentration and the toxic effect in the zebrafish embryos for the substances of the first group. The study showed that substances having a strong damaging effect on fetuses cause changes that are apparent and easy to evaluate both in hydras and zebrafish embryos. A scoring system was introduced to evaluate the changes of hydras and zebrafish embryos. The point system of evaluation of changes allows quick classification of a substance as potentially embryotoxic, fetotoxic or teratogenic. It allows developing a cheap and fast method alternative to prenatal developmental toxicity studies, a screening method that enables substances of great teratogenic potential to be excluded from studies on laboratory animals.

The development of analytical methods for the purity determination of fucoidan extracted from brown seaweed species.

To determine the purity of extracted fucoidan from brown seaweeds, analytical methods were developed, including spectroscopy (i.e., Attenuate total reflectance (ATR) - Fourier-transform infrared (FT-IR) and Raman) combined with chemometrics; and the results were compared with those of high performance liquid chromatography (HPLC) and other two chemistry methods (i.e., fucoidan estimation based on fucose content and a cationic dye method based on sulphated polysaccharide estimation). Quantitative models (i.e., partial least squares regression (PLSR)) were developed and cross-validated using FT-IR spectroscopic methods (R2CV ~ 0.998, RMSECV ~1.7%). The models were also validated using other four commercial fucoidan products. On the other hand, the same commercial samples were used to validate the two chemistry methods and the HPLC method. Estimation results of these analytical methods were discussed based on the potential of these analytical methods for fucoidan purity determination. The results demonstrated FT-IR spectroscopy with chemometrics potentially could be used for non-destructive and real time determination.

Live Attenuated African Swine Fever Viruses as Ideal Tools to Dissect the Mechanisms Involved in Cross-Protection.

African swine fever (ASF) has become the major threat for the global swine industry. Furthermore, the epidemiological situation of African swine fever virus (ASFV) in some endemic regions of Sub-Saharan Africa is worse than ever, with multiple virus strains and genotypes currently circulating in a given area. Despite the recent advances on ASF vaccine development, there are no commercial vaccines yet, and most of the promising vaccine prototypes available today have been specifically designed to fight the genotype II strains currently circulating in Europe, Asia, and Oceania. Previous results from our laboratory have demonstrated the ability of BA71∆CD2, a recombinant LAV lacking CD2v, to confer protection against homologous (BA71) and heterologous genotype I (E75) and genotype II (Georgia2007/01) ASFV strains, both belonging to same clade (clade C). Here, we extend these results using BA71∆CD2 as a tool trying to understand ASFV cross-protection, using phylogenetically distant ASFV strains. We first observed that five out of six (83.3%) of the pigs immunized once with 106 PFU of BA71∆CD2 survived the tick-bite challenge using Ornithodoros sp. soft ticks naturally infected with RSA/11/2017 strain (genotype XIX, clade D). Second, only two out of six (33.3%) survived the challenge with Ken06.Bus (genotype IX, clade A), which is phylogenetically more distant to BA71∆CD2 than the RSA/11/2017 strain. On the other hand, homologous prime-boosting with BA71∆CD2 only improved the survival rate to 50% after Ken06.Bus challenge, all suffering mild ASF-compatible clinical signs, while 100% of the pigs immunized with BA71∆CD2 and boosted with the parental BA71 virulent strain survived the lethal challenge with Ken06.Bus, without almost no clinical signs of the disease. Our results confirm that cross-protection is a multifactorial phenomenon that not only depends on sequence similarity. We believe that understanding this complex phenomenon will be useful for designing future vaccines for ASF-endemic areas.

DHX9 interacts with APOBEC3B and attenuates the anti-HBV effect of APOBEC3B.

Hepatitis B virus (HBV) is a partially double-stranded DNA virus that replicates by reverse transcription. We previously demonstrated that the host restriction factor-APOBEC3B (A3B) inhibited HBV replication which was dependent on its deaminase activity during reverse transcription. However, the host factors involved in the process of regulating the anti-HBV function of A3B are less known. In this research, to obtain a comprehensive understanding of the interaction networks of A3B, we conducted coimmunoprecipitation and mass spectrometry to identify A3B-interacting proteins in the presence of HBV. By this approach, we determined that DExD/H-box helicase 9 (DHX9) suppressed the anti-HBV effect of A3B, and this suppression was dependent on their interaction. Although DHX9 did not affect the deamination activity of A3B in vitro assay or the viral DNA editing of A3B in HepG2-NTCP cells that support HBV infection, it inhibited the binding of A3B with pgRNA. These data suggest that DHX9 can interact with A3B and attenuate the anti-HBV efficacy of A3B.Abbreviations: 3D-PCR: differential DNA denaturation PCR; APOBEC3: apolipoprotein B mRNA-editing catalytic polypeptide 3; cccDNA: covalently closed circular DNA; co-IP: coimmunoprecipitation; DDX: DExD-box RNA helicases; HBc: HBV core protein; HBV: hepatitis B virus; HepAD38: HepG2 cell line stably transfected with HBV DNA; HepG2-NTCP: HepG2 cell line stably transfected with Na+/taurocholate cotransporter polypeptide; Huh7: human hepatoma cell line; pgRNA: pregenomic RNA; PPI: protein-protein interactions; RC DNA: relaxed circular DNA.

A Practicable Method of Tuning the Noise Intensity at Protein Level.

The expression of genes is inevitably subject to intracellular noise. Noise, for some regulatory networks, is constructive but detrimental to many others. The intensity of the noise is a determinant factor and the method of tuning it is of great value. In this study, we illustrated that the transcriptional delay in an incoherent feedforward loop (FFL) grants the target protein modulation the intensity of noise. Remarkably, for a wide range, the coefficient of variation (COV) of the target protein appeared to be about linear to the time span of the transcriptional delay. Without a noise-buffering method, the COV of the target protein is 0.455. While applying incoherent FFL, the COV reduced to 0.236. Then, it changed from 0.236 to 0.630 as the transcriptional delay raised from 0 to 1000 seconds. If we further increased the delay out of the linear range, the COV finally reached 0.779. In addition, we incorporated the distribution of the transcriptional delay in the delay stochastic simulation algorithm. This distribution is based on the experimental observation in the literature. The outcome suggested that the distributed delay slightly improved the ability of tuning noise. In conclusion, we demonstrated a noise-tuning method that altered only the intensity of noise without changing the deterministic steady-state behaviors. It is ready to be applied to various systems in the field of synthetic biology.

Immunogenicity generated by mRNA vaccine encoding VZV gE antigen is comparable to adjuvanted subunit vaccine and better than live attenuated vaccine in nonhuman primates.

Shingles is a painful, blistering rash caused by reactivation of latent varicella-zoster virus (VZV) and most frequently occurs in elderly and immunocompromised individuals. Currently, two approved vaccines for the prevention of shingles are on the market, a live attenuated virus vaccine ZOSTAVAX® (Merck & Co., Inc., Kenilworth, NJ, USA) and an AS01B adjuvanted subunit protein vaccine Shingrix™ (Glaxo Smith Kline, Rockville, MD, USA). Human clinical immunogenicity and vaccine efficacy data is available for these two benchmark vaccines, offering a unique opportunity for comparative analyses with novel vaccine platforms and animal model translatability studies. The studies presented here utilized non-human primates (NHP) to evaluate humoral and cellular immune response by three vaccine modalities: the new platform of lipid nanoparticle (LNP) formulated mRNA encoding VZV gE antigen (VZV gE mRNA/LNP) as compared with well-established platforms of live attenuated VZV (VZV LAV) and adjuvanted VZV gE subunit protein (VZV gE protein/adjuvant). The magnitude of response to vaccination with a single 100-200 µg mRNA dose or two 50 µg mRNA doses of VZV gE mRNA/LNP were comparable to two 50 µg protein doses of VZV gE protein/adjuvant, suggesting the VZV gE mRNA/LNP platform has the potential to elicit a robust immune response, and both modalities generated markedly higher responses than VZV LAV. Additionally, the slopes of decay for VZV-specific antibody titers were roughly similar across all three vaccines, indicating the magnitude of peak immunogenicity was the driving force in determining immune response longevity. Finally, vaccine-induced immunogenicity with VZV LAV and VZV gE protein/adjuvant in NHP closely resembled human clinical trials immune response data for ZOSTAVAX® and Shingrix™, helping to validate NHP as an appropriate preclinical model for evaluating these vaccines.

Vitexin attenuates epithelial ovarian cancer cell viability and motility in vitro and carcinogenesis in vivo via p38 and ERK1/2 pathways related VEGFA.

BACKGROUND: Epithelial ovarian cancer (EOC) is the most common type of ovarian tumor, however, effective treatment does not currently exist for this condition. This study evaluated the role of vitexin in mitigating EOC both in vitro and in vivo. METHOD: SKOV-3 cells were used for in vitro experimentation. Xenotransplantation mouse models were set up by subcutaneously injecting mice with SKOV-3 cells. CCK8 was used to screen the optimal dose in vitro. Cell proliferation, invasion, number of microtubule nodules and apoptosis were respectively detected by colony formation assay, transwell assay, microtubule formation assay and flow cytometry. TUNEL and immunohistochemistry were used to detect tissues apoptosis and VEGF content. Western blot assay was used to detect the expression of Ki67, caspase-3, VEGFA, VEGFR2, ERK1/2 and p38. RESULTS: In vitro experiment, compared with the control group, 10 µL of vitexin significantly reduced Ki67 levels and enhanced tumor cell apoptosis rate. Additionally, the colony forming rate, invasive cells per field, and number of nodes/HPF in vitexin treated group decreased dramatically. The result of western blot showed that levels of p-p38/p38 and p-ERK1/2/ERK1/2 also noticeably decreased. In vivo experiment, 40 mg/kg of vitexin significantly inhibited tumor growth. In addition, vitexin significantly enhanced the percentage of tissues apoptosis, which was accompanied by a decrease in the percentage of VEGF-positive cells. CONCLUSIONS: Vitexin decreased the proliferation and invasion of SKOV-3 cells and noticeably reduced tumor growth. These findings suggest that vitexin could be a promising therapy for EOC.

Partial least Squares- least squares- Support Vector Machine Modeling of ATR-IR as a Spectrophotometric Method for Detection and Determination of Iron in Pharmaceutical Formulations.

Iron is an essential element used as supplement in different dosage-forms. Different time and expenditure-consuming methods introduced for detection and determination of elemental ions such as Atomic Absorption Spectroscopy. In this research, two different and routine methods containing ATR-IR and atomic absorption were applied to define the amount of iron in 198 samples containing different concentrations of commercial iron drops and syrups and the output data of the methods was transferred to chemometric model to compare the accuracy and robustness of the methods. By applying this mathematical model, in addition to the confirmation of ATR-IR (a time and energy-saving method) as a replacement of Atomic Absorption Spectroscopy to produce the same results, chemometrical model was used to evaluate the output data in a faster and easier method. At first, ATR-IR spectra data converted to normal matrix by SNV preprocessing approach. Then, a relationship between iron concentrations achieved by AAS and ATR-IR data was established using PLS-LS-SVM. Consequently, model was able to predict ~99% of the samples with low error-values (root mean square-error of cross-validation equal to 0.98). Y-permutation test performed to confirm that the model was not assessed accidentally. Although, chemometric methods for detection of some heavy metals have been reported in the literature, combination of PLS-LS-SVM with ATR-IR was not cited. In this study a fast and robust method for iron assay was suggested.As a result, ATR-IR can be a suitable method in detection and qualification of iron-content in pharmaceutical dosage forms with less energy-consumption but similar accuracy.