During the development of yellow-fleshed kiwifruit (Actinidia chinensis), the flesh appeared light pink at the initial stage, the pink faded at the fastest growth stage, and gradually changed into green. At the maturity stage, it showed bright yellow. In order to analyze the mechanism of flesh color change at the metabolic and gene transcription level, the relationship between color and changes of metabolites and key enzyme genes was studied. In this study, five time points (20 d, 58 d, 97 d, 136 d, and 175 d) of yellow-fleshed kiwifruit were used for flavonoid metabolites detection and transcriptome, and four time points (20 d, 97 d, 136 d, and 175 d) were used for targeted detection of carotenoids. Through the analysis of the content changes of flavonoid metabolites, it was found that the accumulation of pelargonidin and cyanidin and their respective anthocyanin derivatives was related to the pink flesh of young fruit, but not to delphinidin and its derivative anthocyanins. A total of 140 flavonoid compounds were detected in the flesh, among which anthocyanin and 76% of the flavonoid compounds had the highest content at 20 d, and began to decrease significantly at 58 d until 175 d, resulting in the pale-pink fading of the flesh. At the mature stage of fruit development (175 d), the degradation of chlorophyll and the increase of carotenoids jointly led to the change of flesh color from green to yellow, in addition to chlorophyll degradation. In kiwifruit flesh, 10 carotenoids were detected, with none of them being linear carotenoids. During the whole development process of kiwifruit, the content of β-carotene was always higher than that of α-carotene. In addition, β-cryptoxanthin was the most-accumulated pigment in the kiwifruit at 175 d. Through transcriptome analysis of kiwifruit flesh, seven key transcription factors for flavonoid biosynthesis and ten key transcription factors for carotenoid synthesis were screened. This study was the first to analyze the effect of flavonoid accumulation on the pink color of yellow-fleshed kiwifruit. The high proportion of β-cryptoxanthin in yellow-fleshed kiwifruit was preliminarily found. This provides information on metabolite accumulation for further revealing the pink color of yellow-fleshed kiwifruit, and also provides a new direction for the study of carotenoid biosynthesis and regulation in yellow-fleshed kiwifruit. Full article

Osteoprotegerin (OPG) is a new member of the tumor necrosis factor (TNF) receptor superfamily, which can inhibit the differentiation and activity of osteoclasts by binding to nuclear factor kappa B receptor activator (RANK) competitively with nuclear factor kappa B receptor activator ligand (RANKL). The previous experiments found that OPG can induce apoptosis of mature osteoclasts in vitro, which can inhibit the activity of mature osteoclasts, thereby exerting its role in protecting bone tissue. In addition, pyroptosis is a new type of cell death that is different from apoptosis. It is unclear whether OPG can induce mature osteoclast pyroptosis and thereby play its role in protecting bone tissue. In this study, the results showed that compared with the control group, the survival rate of osteoclasts in the OPG group was significantly reduced, and the contents of IL-1β, IL-18, and LDH in the supernatant both increased. Many osteoclast plasma membranes were observed to rupture in bright fields, and OPG induced loss of their morphology. Flow cytometry was used to analyze the pyroptosis rate; OPG significantly increased the osteoclast pyroptosis rate. To further reveal the mechanism of OPG-induced osteoclast pyroptosis, we examined the expression level of pyroptosis-related genes and proteins, and the results found that OPG increased the expression of NLRP3, ASC, caspase-1, and GSDMD-N compared with the control group. In summary, OPG can induce osteoclast pyroptosis, and its mechanism is related to the expression levels of ASC, NLRP3, caspase 1 and GSDMD, which were included in the classical pathway of pyroptosis. Full article

Salmonella Typhimurium is a Gram-negative intestinal pathogen that can infect humans and a variety of animals, causing gastroenteritis or serious systemic infection. Replication within host macrophages is essential for S. Typhimurium to cause systemic infection. By analyzing transcriptome data, the expression of yhjC gene, which encodes a putative regulator in S. Typhimurium, was found to be significantly up-regulated after the internalization of Salmonella by macrophages. Whether yhjC gene is involved in S. Typhimurium systemic infection and the related mechanisms were investigated in this study. The deletion of yhjC reduced the replication ability of S. Typhimurium in macrophages and decreased the colonization of S. Typhimurium in mouse systemic organs (liver and spleen), while increasing the survival rate of the infected mice, suggesting that YhjC protein promotes systemic infection by S. Typhimurium. Furthermore, by using transcriptome sequencing and RT-qPCR assay, the transcription of several virulence genes, including spvD, iroCDE and zraP, was found to be down-regulated after the deletion of yhjC. Electrophoretic mobility shift assay showed that YhjC protein can directly bind to the promoter region of spvD and zraP to promote their transcription. These findings suggest that YhjC contributes to the systemic virulence of S. Typhimurium via the regulation of multiple virulence genes and YhjC could represent a promising target to control S. Typhimurium infection. Full article

We previously reported that Korean red ginseng (KRG) exerts an anti-inflammatory role through inhibiting caspase-11 non-canonical inflammasome in macrophages; however, the components responsible for the anti-inflammatory role remained unclear. This study explored the anti-inflammatory activity of the KRG saponin fraction (KRGSF) in caspase-11 non-canonical inflammasome-activated macrophages. KRGSF inhibited pyroptosis, pro-inflammatory cytokine secretion, and inflammatory mediator production in caspase-11 non-canonical inflammasome-activated J774A.1 cells. A mechanism study revealed that KRGSF-induced anti-inflammatory action was mediated via suppressing the proteolytic activation of caspase-11 and gasdermin D (GSDMD) in caspase-11 non-canonical inflammasome-activated J774A.1 cells. Moreover, KRGSF increased the survival of lethal septic mice. Taken together, these results reveal KRGSF-mediated anti-inflammatory action with a novel mechanism, by inhibiting caspase-11 non-canonical inflammasome in macrophages. Full article

Swim-up selected human sperm were incubated with 7 ng F₄-neuroprostanes (F₄-NeuroPs) for 2 and 4 h. Sperm motility and membrane mitochondrial potential (MMP) were evaluated. The percentage of reacted acrosome was assessed by pisum sativum agglutinin (PSA). Chromatin integrity was detected using the acridine orange (AO) assay and localization of the ryanodine receptor was performed by immunofluorescence analysis. Sperm progressive motility (p = 0.02) and the percentage of sperm showing a strong MMP signal (p = 0.012) significantly increased after 2 h F₄-NeuroP incubation compared to control samples. The AO assay did not show differences in the percentage of sperm with dsDNA between treated or control samples. Meanwhile, a significantly higher number of sperm with reacted acrosomes was highlighted by PSA localization after 4 h F₄-NeuroP incubation. Finally, using an anti-ryanodine antibody, the immunofluorescence signal was differentially distributed at 2 and 4 h: a strong signal was evident in the midpiece and postacrosomal sheath (70% of sperm) at 2 h, whereas a dotted one appeared at 4 h (53% of sperm). A defined concentration of F₄-NeuroPs in seminal fluid may induce sperm capacitation via channel ions present in sperm cells, representing an aid during in vitro sperm preparation that may increase the positive outcome of assisted fertilization. Full article

Host regulatory immune response is involved in the hepatic inflammatory process caused by the hepatitis C virus (HCV). We aimed to determine if HCV clearance with direct-acting antivirals (DAAs) changes the hepatic fibrosis stage, biochemical parameters of liver injury, and inflammatory/immune responses. Sample: 329 chronic hepatitis C (CHC) patients, 134 of them treated with DAAs. Liver fibrosis was evaluated by transient elastography (FibroScan), biochemical and cellular parameters were determined by standard methods, cytokine concentration by enzyme-linked immunoabsorbent assay (ELISA), and genetic polymorphisms by polymerase chain reaction—restriction fragment length polymorphism (PCR-RFLP) or endpoint genotyping. Before DAA treatment, severe fibrosis or cirrhosis (F3/4) was associated with higher values of tumor necrosis factor-alpha (TNF-α) and genotypes transforming growth factor-beta-509 C/T_CC (TGF-β-509 C/T_CC), interleukine-10-1082 T/C_CC (IL-10-1082 T/C_CC), and IL-10-592 G/T_GT. After DAA treatment, fewer F3/4 patients and lower values of TNF-α were found. Patients with TNF-α-308 G/A_GG and IL-10-592 G/T_GT were at risk for F3/4. Lack of improvement of liver fibrosis was associated with lower baseline values of platelet count for genotypes TNF-α-308 G/A_GG and haplotype TT/GG of IL-10-1082 T/C and IL-10-592 G/T. Our study showed decreased liver fibrosis/inflammation and normalization of liver injury biomarkers after DAA treatment. It also points to the importance of suppressing the pro-inflammatory response by DAAs in the resolution of hepatitis C, contributing to the improvement of liver damage evaluated by transient elastography. Full article

S-layers are involved in the adaptation of bacteria to the outside environment and in pathogenesis, often representing special virulence factors. Vegetative cells of the entomopathogenic bacterium Brevibacillus laterosporus are characterized by an overproduction of extracellular surface layers that are released in the medium during growth. The purpose of this study was to characterize cell wall proteins of this bacterium and to investigate their involvement in pathogenesis. Electron microscopy observations documented the presence of multiple S-layers, including an outermost (OW) and a middle (MW) layer, in addition to the peptidoglycan layer covering the plasma membrane. After identifying these proteins (OWP and MWP) by mass spectrometry analyses, and determining their gene sequences, the cell wall multilayer-released fraction was successfully isolated and used in insect bioassays alone and in combination with bacterial spores. This study confirmed a central role of spores in bacterial pathogenicity to insects but also detected a significant virulence associated with fractions containing released cell wall multilayer proteins. Taken together, S-layer proteins appear to be part of the toxins and virulence factors complex of this microbial control agent of invertebrate pests. Full article

In this research, we investigated the structural and biological properties of phosphate glasses (PGs) after the addition of V₂O₅. A xV₂O₅∙(100 − x)[CaF₂∙3P₂O₅∙CaO] glass system with 0 ≤ x ≤ 16 mol% was synthesized via a conventional melt-quenching technique. Several analysis techniques (dissolution tests, pH, SEM-EDS, FT-IR, and EPR) were used to obtain new experimental data regarding the structural behavior of the system. In vitro tests were conducted to assess the antitumor character of V₂O₅-doped glass (x = 16 mol%) compared to the matrix (x = 0 mol%) and control (CTRL-) using several tumoral cell lines (A375, A2780, and Caco-2). The characterization of PGs showed an overall dissolution rate of over 90% for all vitreous samples (M and V1–V7) and the high reactivity of this system. EPR revealed a well-resolved hyperfine structure (hfs) typical of vanadyl ions in a C_4v symmetry. FT-IR spectra showed the presence of all structural units expected for P₂O₅, as well as very clear depolymerization of the vitreous network induced by V₂O₅. The MTT assay indicated that the viability of tumor cells treated with V7-glass extract was reduced to 50% when the highest concentration was used (10 µg/mL) compared to the matrix treatment (which showed no cytotoxic effect at any concentration). Moreover, the matrix treatment (without V₂O₅) provided an optimal environment for tumor cell attachment and proliferation. In conclusion, the two types of treatment investigated herein were proven to be very different from a statistical point of view (p < 0.01), and the in vitro studies clearly underline the cytotoxic potential of vanadium ions from phosphate glass (V7) as an antitumor agent. Full article

SNAI2 (Snai2) is a zinc-finger transcriptional repressor that belongs to the Snail family. The accumulated evidence suggests that SNAI2 exhibits biphasic effects on regulating a stem-like phenotype in various types of cells, both normal and malignant. In this study, by exogenously expressing SNAI2 in SiHa cells, SNAI2 exhibited the capacity to inhibit a stem-like phenotype in cervical cancer cells. The SNAI2-overexpressing cells inhibited cell growth, tumorsphere formation, tumor growth, enhanced sensitivity to cisplatin, reduced stem cell-related factors’ expression, and lowered tumor initiating frequency. In addition, the EPCAM^high cells sorted from SiHa cells exhibited an enhanced capacity to maintain a stem-like phenotype. Further study demonstrated that the trans-suppression of EPCAM expression by SNAI2 led to blockage of the nuclear translocation of β-catenin, as well as reduction in SOX2 and c-Myc expression in SiHa and HeLa cells, but induction in SNAI2 knockdown cells (CaSki), which would be responsible for the attenuation of the stem-like phenotype in cervical cancer cells mediated by SNAI2. All of these results demonstrated that SNAI2 could attenuate the stem-like phenotype in cervical cancer cells through the EPCAM/β-catenin axis. Full article

Sorafenib is currently a targeted agent widely used in the treatment of advanced hepatocellular carcinoma (aHCC). However, to date there is still a lack of a reliable marker capable of predicting sorafenib therapeutic responses. Here, we conducted a genome-wide association study (GWAS) to identify candidate single-nucleotide polymorphism outcome predictors in aHCC patients. A total of 74 real-world sorafenib-treated aHCC patients were enrolled for GWAS and outcome analysis. GWAS showed that rs1010816 (p = 2.2 × 10⁻⁷) was associated with sorafenib therapeutic response in aHCC patients. Kaplan–Meier analysis indicated that the “TT” genotype was significantly associated with a favorable therapeutic response but not significantly associated with overall survival (OS). Univariate followed by multivariate Cox proportional hazard analysis showed that ascites, main portal vein thrombosis, lower platelet count, lower total sorafenib doses, higher PALBI score in model A and higher ALBI grade in model B were significantly associated with a shorter OS. Subgroup analysis showed that only in alcoholic aHCC patients treated by sorafenib, rs1010816 “TT” genotype was significantly associated with longer OS (p = 0.021). Sorafenib had a favorable therapeutic outcome in alcoholic aHCC patients carrying rs1010816 “TT” genotype. Full article

Cellular effects of hypergravity have been described in many studies. We investigated the transcriptional dynamics in Jurkat T cells between 20 s and 60 min of 9 g hypergravity and characterized a highly dynamic biphasic time course of gene expression response with a transition point between rapid adaptation and long-term response at approximately 7 min. Upregulated genes were shifted towards the center of the nuclei, whereby downregulated genes were shifted towards the periphery. Upregulated gene expression was mostly located on chromosomes 16–22. Protein-coding transcripts formed the majority with more than 90% of all differentially expressed genes and followed a continuous trend of downregulation, whereas retained introns demonstrated a biphasic time-course. The gene expression pattern of hypergravity response was not comparable with other stress factors such as oxidative stress, heat shock or inflammation. Furthermore, we tested a routine centrifugation protocol that is widely used to harvest cells for subsequent RNA analysis and detected a huge impact on the transcriptome compared to non-centrifuged samples, which did not return to baseline within 15 min. Thus, we recommend carefully studying the response of any cell types used for any experiments regarding the hypergravity time and levels applied during cell culture procedures and analysis. Full article

Tick-borne encephalitis (TBE) is an emerging zoonosis that may cause long-term neurological sequelae or even death. Thus, there is a growing interest in understanding the factors of TBE pathogenesis. Viral genetic determinants may greatly affect the severity and consequences of TBE. In this study, nonstructural protein 1 (NS1) of the tick-borne encephalitis virus (TBEV) was tested as such a determinant. NS1s of three strains with similar neuroinvasiveness belonging to the European, Siberian and Far-Eastern subtypes of TBEV were studied. Transfection of mouse cells with plasmids encoding NS1 of the three TBEV subtypes led to different levels of NS1 protein accumulation in and secretion from the cells. NS1s of TBEV were able to trigger cytokine production either in isolated mouse splenocytes or in mice after delivery of NS1 encoding plasmids. The profile and dynamics of TNF-α, IL-6, IL-10 and IFN-γ differed between the strains. These results demonstrated the involvement of TBEV NS1 in triggering an immune response and indicated the diversity of NS1 as one of the genetic factors of TBEV pathogenicity. Full article

Antarctic expeditions have a high risk of participant depression owing to long stays and isolated environments. By quantifying the stress state and changes in biomolecules over time before the onset of depressive symptoms, predictive markers of depression can be explored. Here, we evaluated the psychological changes in 30 participants in the Japanese Antarctic Research Expedition using the Patient Health Questionnaire-9 (PHQ-9). Urinary samples were collected every three months for a year, and comprehensive urinary metabolomic profiles were quantified using liquid chromatography time-of-flight mass spectrometry. Five participants showed major depressive episodes (PHQ-9 ≥ 10) at 12 months. The urinary metabolites between these participants and the 25 unaffected participants were compared at individual metabolite and pathway levels. The individual comparisons showed the most significant differences at 12 months in 14 metabolites, including ornithine and beta-alanine. Data from shorter stays showed less significant differences. In contrast, pathway and enrichment analyses showed the most significant difference at three months and a less significant difference at longer stays. These time transitions of urinary metabolites could help in the development of urinary biomarkers to detect subjects with depressive episodes at an early stage. Full article

Insects use a powerful and complex olfactory recognition system to sense odor molecules in the external environment to guide behavior. A large family of odorant receptors (ORs) mediates the detection of pheromone compounds. Anoplophora glabripennis is a destructive pest that harms broad-leaved tree species. Although olfactory sensation is an important factor affecting the information exchange of A. glabripennis, little is known about the key ORs involved. Here, we identified ninety-eight AglaORs in the Agla2.0 genome and found that the AglaOR gene family had expanded with structural and functional diversity. RT-qPCR was used to analyze the expression of AglaORs in sex tissues and in adults at different developmental stages. Twenty-three AglaORs with antennal-biased expression were identified. Among these, eleven were male-biased and two were female-biased and were more significantly expressed in the sexual maturation stage than in the post-mating stage, suggesting that these genes play a role in sexual communication. Relatively, two female-biased AglaORs were overexpressed in females seeking spawning grounds after mating, indicating that these genes might be involved in the recognition of host plant volatiles that may regulate the selection of spawning grounds. Our study provides a theoretical basis for further studies into the molecular mechanism of A. glabripennis olfaction. Full article

Cell division regulators play a vital role in neural progenitor cell (NPC) proliferation and differentiation. Cell division cycle 25C (CDC25C) is a member of the CDC25 family of phosphatases which positively regulate cell division by activating cyclin-dependent protein kinases (CDKs). However, mice with the Cdc25c gene knocked out were shown to be viable and lacked the apparent phenotype due to genetic compensation by Cdc25a and/or Cdc25b. Here, we investigate the function of Cdc25c in developing rat brains by knocking down Cdc25c in NPCs using in utero electroporation. Our results indicate that Cdc25c plays an essential role in maintaining the proliferative state of NPCs during cortical development. The knockdown of Cdc25c causes early cell cycle exit and the premature differentiation of NPCs. Our study uncovers a novel role of CDC25C in NPC division and cell fate determination. In addition, our study presents a functional approach to studying the role of genes, which elicit genetic compensation with knockout, in cortical neurogenesis by knocking down in vivo. Full article