New insights into the all-testis differentiation in zebrafish with compromised endogenous androgen and estrogen synthesis.

The regulatory mechanism of gonadal sex differentiation, which is complex and regulated by multiple factors, remains poorly understood in teleosts. Recently, we have shown that compromised androgen and estrogen synthesis with increased progestin leads to all-male differentiation with proper testis development and spermatogenesis in cytochrome P450 17a1 (cyp17a1)-/- zebrafish. In the present study, the phenotypes of female-biased sex ratio were positively correlated with higher Fanconi anemia complementation group L (fancl) expression in the gonads of doublesex and mab-3 related transcription factor 1 (dmrt1)-/- and cyp17a1-/-;dmrt1-/- fish. The additional depletion of fancl in cyp17a1-/-;dmrt1-/- zebrafish reversed the gonadal sex differentiation from all-ovary to all-testis (in cyp17a1-/-;dmrt1-/-;fancl-/- fish). Luciferase assay revealed a synergistic inhibitory effect of Dmrt1 and androgen signaling on fancl transcription. Furthermore, an interaction between Fancl and the apoptotic factor Tumour protein p53 (Tp53) was found in vitro. The interaction between Fancl and Tp53 was observed via the WD repeat domain (WDR) and C-terminal domain (CTD) of Fancl and the DNA binding domain (DBD) of Tp53, leading to the K48-linked polyubiquitination degradation of Tp53 activated by the ubiquitin ligase, Fancl. Our results show that testis fate in cyp17a1-/- fish is determined by Dmrt1, which is thought to stabilize Tp53 by inhibiting fancl transcription during the critical stage of sexual fate determination in zebrafish.

Biosynthetic deficiency of docosahexaenoic acid causes nonalcoholic fatty liver disease and ferroptosis-mediated hepatocyte injury.

Exogenous omega-3 fatty acids, particularly docosahexaenoic acid (DHA), have shown to exert beneficial effects on nonalcoholic fatty liver disease (NAFLD), which is characterized by the excessive accumulation of lipids and chronic injury in the liver. However, the effect of endogenous DHA biosynthesis on the lipid homeostasis of liver is poorly understood. In this study, we used a DHA biosynthesis-deficient zebrafish model, elovl2 mutant, to explore the effect of endogenously biosynthesized DHA on hepatic lipid homeostasis. We found the pathways of lipogenesis and lipid uptake were strongly activated, while the pathways of lipid oxidation and lipid transport were inhibited in the liver of elovl2 mutants, leading to lipid droplet accumulation in the mutant hepatocytes and NAFLD. Furthermore, the elovl2 mutant hepatocytes exhibited disrupted mitochondrial structure and function, activated endoplasmic reticulum stress, and hepatic injury. We further unveiled that the hepatic cell death and injury was mainly mediated by ferroptosis, rather than apoptosis, in elovl2 mutants. Elevating DHA content in elovl2 mutants, either by the introduction of an omega-3 desaturase (fat1) transgene or by feeding with a DHA-rich diet, could strongly alleviate NAFLD features and ferroptosis-mediated hepatic injury. Together, our study elucidates the essential role of endogenous DHA biosynthesis in maintaining hepatic lipid homeostasis and liver health, highlighting that DHA deficiency can lead to NAFLD and ferroptosis-mediated hepatic injury.

Multi-omics reveals the role of MCM2 and hnRNP K phosphorylation in mouse renal aging through genomic instability.

The process of aging is characterized by structural degeneration and functional decline, as well as diminished adaptability and resistance. The aging kidney exhibits a variety of structural and functional impairments. In aging mice, thinning and graying of fur were observed, along with a significant increase in kidney indices compared to young mice. Biochemical indicators revealed elevated levels of creatinine, urea nitrogen and serum uric acid, suggesting impaired kidney function. Histological analysis unveiled glomerular enlargement and sclerosis, severe hyaline degeneration, capillary occlusion, lymphocyte infiltration, tubular and glomerular fibrosis, and increased collagen deposition. Observations under electron microscopy showed thickened basement membranes, altered foot processes, and increased mesangium and mesangial matrix. Molecular marker analysis indicated upregulation of aging-related ß-galactosidase, p16-INK4A, and the DNA damage marker γH2AX in the kidneys of aged mice. In metabolomics, a total of 62 significantly different metabolites were identified, and 10 pathways were enriched. We propose that citrulline, dopamine, and indoxyl sulfate have the potential to serve as markers of kidney damage related to aging in the future. Phosphoproteomics analysis identified 6656 phosphosites across 1555 proteins, annotated to 62 pathways, and indicated increased phosphorylation at the Ser27 site of Minichromosome maintenance complex component 2 (Mcm2) and decreased at the Ser284 site of heterogeneous nuclear ribonucleoprotein K (hnRNP K), with these modifications being confirmed by western blotting. The phosphorylation changes in these molecules may contribute to aging by affecting genome stability. Eleven common pathways were detected in both omics, including arginine biosynthesis, purine metabolism and biosynthesis of unsaturated fatty acids, etc., which are closely associated with aging and renal insufficiency.

S100A8/A9-activated IFNγ+ NK cells trigger ß-cell necroptosis in hepatitis B virus-associated liver cirrhosis.

BACKGROUND AND AIMS: Hepatitis B virus (HBV)-associated liver cirrhosis (LC), a common condition with high incidence and mortality rates, is often associated with diabetes mellitus (DM). However, the molecular mechanisms underlying impaired glucose regulation during HBV-associated LC remain unclear. METHODS: Data from 63 patients with LC and 62 patients with LC-associated DM were analysed. Co-culture of NK cells and islet ß cell lines were used to study the glucose regulation mechanism. A mouse model of LC was used to verify the effect of S100A8/A9 on the glucose regulation. RESULTS: Higher levels of interferon (IFN)-γ derived from natural killer (NK) cells and lower levels of insulin emerged in the peripheral blood of patients with both LC and DM compared with those from patients with LC only. IFN-γ derived from NK cells facilitated ß cell necroptosis and impaired insulin production. Furthermore, S100A8/A9 elevation in patients with both LC and DM was found to upregulate IFN-γ production in NK cells. Consistently, in the mouse model for LC, mice treated with carbon tetrachloride (CCL4) and S100A8/A9 exhibited increased blood glucose, impaired insulin production, increased IFN-γ, and increased ß cells necroptosis compared with those treated with CCL4. Mechanistically, S100A8/A9 activated the p38 MAPK pathway to increase IFN-γ production in NK cells. These effects were diminished after blocking RAGE. CONCLUSION: Together, the data indicate that IFN-γ produced by NK cells induces ß cell necroptosis via the S100A8/A9-RAGE-p38 MAPK axis in patients with LC and DM. Reduced levels of S100A8/A9, NK cells, and IFN-γ could be valuable for the treatment of LC with DM. Accumulation of S100A8/A9 in patients with LC may indicate the emergence of DM.

Proteomics and ß-hydroxybutyrylation Modification Characterization in the Hearts of Naturally Senescent Mice.

Aging is widely accepted as an independent risk factor for cardiovascular disease (CVD), which contributes to increasing morbidity and mortality in the elderly population. Lysine ß-hydroxybutyrylation (Kbhb) is a novel post-translational modification (PTM), wherein ß-hydroxybutyrate is covalently attached to lysine ε-amino groups. Recent studies have revealed that histone Kbhb contributes to tumor progression, diabetic cardiomyopathy progression, and postnatal heart development. However, no studies have yet reported a global analysis of Kbhb proteins in aging hearts or elucidated the mechanisms underlying this modification in the process. Herein, we conducted quantitative proteomics and Kbhb PTM omics to comprehensively elucidate the alterations of global proteome and Kbhb modification in the hearts of aged mice. The results revealed a decline in grip strength and cardiac diastolic function in 22-month-old aged mice compared to 3-month-old young mice. High-throughput liquid chromatogram-mass spectrometry analysis identified 1710 ß-hydroxybutyrylated lysine sites in 641 proteins in the cardiac tissue of young and aged mice. Additionally, 183 Kbhb sites identified in 134 proteins exhibited significant differential modification in aged hearts (fold change (FC) > 1.5 or <1/1.5, p < 0.05). Notably, the Kbhb-modified proteins were primarily detected in energy metabolism pathways, such as fatty acid elongation, glyoxylate and dicarboxylate metabolism, tricarboxylic acid cycle, and oxidative phosphorylation. Furthermore, these Kbhb-modified proteins were predominantly localized in the mitochondria. The present study, for the first time, provides a global proteomic profile and Kbhb modification landscape of cardiomyocytes in aged hearts. These findings put forth novel possibilities for treating cardiac aging and aging-related CVDs by reversing abnormal Kbhb modifications.

Quantification and Proteomic Characterization of ß-Hydroxybutyrylation Modification in the Hearts of AMPKα2 Knockout Mice.

AMP-activated protein kinase alpha 2 (AMPKα2) regulates energy metabolism, protein synthesis, and glucolipid metabolism myocardial cells. Ketone bodies produced by fatty acid ß-oxidation, especially ß-hydroxybutyrate, are fatty energy-supplying substances for the heart, brain, and other organs during fasting and long-term exercise. They also regulate metabolic signaling for multiple cellular functions. Lysine ß-hydroxybutyrylation (Kbhb) is a ß-hydroxybutyrate-mediated protein posttranslational modification. Histone Kbhb has been identified in yeast, mouse, and human cells. However, whether AMPK regulates protein Kbhb is yet unclear. Hence, the present study explored the changes in proteomics and Kbhb modification omics in the hearts of AMPKα2 knockout mice using a comprehensive quantitative proteomic analysis. Based on mass spectrometry (LC-MS/MS) analysis, the number of 1181 Kbhb modified sites in 455 proteins were quantified between AMPKα2 knockout mice and wildtype mice; 244 Kbhb sites in 142 proteins decreased or increased after AMPKα2 knockout (fold change >1.5 or <1/1.5, p < 0.05). The regulation of Kbhb sites in 26 key enzymes of fatty acid degradation and tricarboxylic acid cycle was noted in AMPKα2 knockout mouse cardiomyocytes. These findings, for the first time, identified proteomic features and Kbhb modification of cardiomyocytes after AMPKα2 knockout, suggesting that AMPKα2 regulates energy metabolism by modifying protein Kbhb.

A novel strategy to map a locus associated with flowering time in canola (Brassica napus L.).

Flowering time is an important agronomic trait for canola breeders, as it provides growers with options for minimizing exposure to heat stress during flowering and to more effectively utilize soil moisture. Plants have evolved various systems to control seasonal rhythms in reproductive phenology including an internal circadian clock that responds to environmental signals. In this study, we used canola cultivar 'Westar' as a recurrent parent and canola cultivar 'Surpass 400' as the donor parent to generate a chromosome segment substitution line (CSSL) and to map a flowering time locus on chromosome A10 using molecular marker-assisted selection. This CSSL contains an introgressed 4.6 mega-bases (Mb) segment (between 13 and 17.6 Mb) of Surpass 400, which substantially delayed flowering compared with Westar. To map flowering time gene(s) within this locus, eight introgression lines (ILs) were developed carrying a series of different lengths of introgressed chromosome A10 segments using five co-dominant polymorphic markers located at 13.5, 14.0, 14.5, 15.0, 15.5, and 16.0 Mb. Eight ILs were crossed with Westar reciprocally and flowering time of resultant 16 F1 hybrids and parents were evaluated in a greenhouse (2021 and 2022). Four ILs (IL005, IL017, IL035, and IL013) showed delayed flowering compared to Westar (P < 0.0001), and their reciprocal crosses displayed a phenotype intermediate in flowering time of both homozygote parents. These results indicated that flowering time is partial or incomplete dominance, and the flowering time locus mapped within a 1 Mb region between two co-dominant polymorphic markers at 14.5-15.5 Mb on chromosome A10. The flowering time locus was delineated to be between 14.60 and 15.5 Mb based on genotypic data at the crossover site, and candidate genes within this region are associated with flowering time in canola and/or Arabidopsis. The co-dominant markers identified on chromosome A10 should be useful for marker assisted selection in breeding programs but will need to be validated to other breeding populations or germplasm accessions of canola.

Association mapping of tan spot and septoria nodorum blotch resistance in cultivated emmer wheat.

KEY MESSAGE: A total of 65 SNPs associated with resistance to tan spot and septoria nodorum blotch were identified in a panel of 180 cultivated emmer accessions through association mapping Tan spot and septoria nodorum blotch (SNB) are foliar diseases caused by the respective fungal pathogens Pyrenophora tritici-repentis and Parastagonospora nodorum that affect global wheat production. To find new sources of resistance, we evaluated a panel of 180 cultivated emmer wheat (Triticum turgidum ssp. dicoccum) accessions for reactions to four P. tritici-repentis isolates Pti2, 86-124, 331-9 and DW5, two P. nodorum isolate, Sn4 and Sn2000, and four necrotrophic effectors (NEs) produced by the pathogens. About 8-36% of the accessions exhibited resistance to the four P. tritici-repentis isolates, with five accessions demonstrating resistance to all isolates. For SNB, 64% accessions showed resistance to Sn4, 43% to Sn2000 and 36% to both isolates, with Spain (11% accessions) as the most common origin of resistance. To understand the genetic basis of resistance, association mapping was performed using SNP (single nucleotide polymorphism) markers generated by genotype-by-sequencing and the 9 K SNP Infinium array. A total of 46 SNPs were significantly associated with tan spot and 19 SNPs with SNB resistance or susceptibility. Six trait loci on chromosome arms 1BL, 3BL, 4AL (2), 6BL and 7AL conferred resistance to two or more isolates. Known NE sensitivity genes for disease development were undetected except Snn5 for Sn2000, suggesting novel genetic factors are controlling host-pathogen interaction in cultivated emmer. The emmer accessions with the highest levels of resistance to the six pathogen isolates (e.g., CItr 14133-1, PI 94634-1 and PI 377672) could serve as donors for tan spot and SNB resistance in wheat breeding programs.

Integrated metabolomics and phosphoproteomics reveal the protective role of exosomes from human umbilical cord mesenchymal stem cells in naturally aging mouse livers.

BACKGROUND: Aging is characterized by a general decline in cellular function, which ultimately affects whole body homeostasis. This study aimed to investigate the effects and underlying mechanisms of exosomes derived from human umbilical cord mesenchymal stem cells (hUCMSC-exos) on the livers of naturally aging mice. METHOD: Twenty-two-month-old C57BL6 mice were used as a natural aging animal model, divided into a saline-treated wild-type aged control group (WT-AC) and a hUCMSC-exo-treated group (WT-AEX), and then detected by morphology, metabolomics and phosphoproteomics. RESULTS: Morphological analysis showed that hUCMSC-exos ameliorated structural disorder and decreased markers of senescence and genome instability in aging livers. Metabolomics showed that hUCMSC-exos decreased the contents of saturated glycerophospholipids, palmitoyl-glycerols and eicosanoid derivatives associated with lipotoxicity and inflammation, consistent with the decreased phosphorylation of metabolic enzymes, such as propionate-CoA ligase (Acss2), at S267 detected by phosphoproteomics. Moreover, phosphoproteomics indicated that hUCMSC-exos reduced the phosphorylation of proteins participating in nuclear transport and cancer signaling, such as heat shock protein HSP90-beta (Hsp90ab1) at S226 and nucleoprotein TPR (Tpr) at S453 and S379, while increasing those involved in intracellular communication, such as calnexin (Canx) at S563 and PDZ domain-containing protein 8 (Pdzd8). Finally, phosphorylated HSP90ß and Tpr were verified predominantly in hepatocytes. CONCLUSION: HUCMSC-exos improved metabolic reprogramming and genome stability mainly associated with phosphorylated HSP90ß in hepatocytes in natural aging livers. This work provides a comprehensive resource of biological data by omics to support future investigations of hUCMSC-exos in aging.

Cytoskeletal gene alterations linked to sorafenib resistance in hepatocellular carcinoma.

BACKGROUND: Although sorafenib has been consistently used as a first-line treatment for advanced hepatocellular carcinoma (HCC), most patients will develop resistance, and the mechanism of resistance to sorafenib needs further study. METHODS: Using KAS-seq technology, we obtained the ssDNA profiles within the whole genome range of SMMC-7721 cells treated with sorafenib for differential analysis. We then intersected the differential genes obtained from the analysis of hepatocellular carcinoma patients in GSE109211 who were ineffective and effective with sorafenib treatment, constructed a PPI network, and obtained hub genes. We then analyzed the relationship between the expression of these genes and the prognosis of hepatocellular carcinoma patients. RESULTS: In this study, we identified 7 hub ERGs (ACTB, CFL1, ACTG1, ACTN1, WDR1, TAGLN2, HSPA8) related to drug resistance, and these genes are associated with the cytoskeleton. CONCLUSIONS: The cytoskeleton is associated with sorafenib resistance in hepatocellular carcinoma. Using KAS-seq to analyze the early changes in tumor cells treated with drugs is feasible for studying the drug resistance of tumors, which provides reference significance for future research.

Estrogen Signaling Inhibits the Expression of anti-Müllerian hormone (amh) and gonadal-soma-derived factor (gsdf) during the Critical Time of Sexual Fate Determination in Zebrafish.

The mechanism of fish gonadal sex differentiation is complex and regulated by multiple factors. It has been widely known that proper steroidogenesis in Leydig cells and sex-related genes in Sertoli cells play important roles in gonadal sex differentiation. In teleosts, the precise interaction of these signals during the sexual fate determination remains elusive, especially their effect on the bi-potential gonad during the critical stage of sexual fate determination. Recently, all-testis phenotypes have been observed in the cyp17a1-deficient zebrafish and common carp, as well as in cyp19a1a-deficient zebrafish. By mating cyp17a1-deficient fish with transgenic zebrafish Tg(piwil1:EGFP-nanos3UTR), germ cells in the gonads were labelled with enhanced green fluorescent protein (EGFP). We classified the cyp17a1-deficient zebrafish and their control siblings into primordial germ cell (PGC)-rich and -less groups according to the fluorescence area of the EGFP labelling. Intriguingly, the EGFP-labelled bi-potential gonads in cyp17a1+/+ fish from the PGC-rich group were significantly larger than those of the cyp17a1-/- fish at 23 days post-fertilization (dpf). Based on the transcriptome analysis, we observed that the cyp17a1-deficient fish of the PGC-rich group displayed a significantly upregulated expression of amh and gsdf compared to that of control fish. Likewise, the upregulated expressions of amh and gsdf were observed in cyp19a1a-deficient fish as examined at 23 dpf. This upregulation of amh and gsdf could be repressed by treatment with an exogenous supplement of estradiol. Moreover, tamoxifen, an effective antagonist of both estrogen receptor α and ß (ERα and Erß), upregulates the expression of amh and gsdf in wild-type (WT) fish. Using the cyp17a1- and cyp19a1a-deficient zebrafish, we provide evidence to show that the upregulated expression of amh and gsdf due to the compromised estrogen signaling probably determines their sexual fate towards testis differentiation. Collectively, our data suggest that estrogen signaling inhibits the expression of amh and gsdf during the critical time of sexual fate determination, which may broaden the scope of sex steroid hormones in regulating gonadal sex differentiation in fish.

Structural Insight into Molecular Inhibitory Mechanism of InsP6 on African Swine Fever Virus mRNA-Decapping Enzyme g5Rp.

Removal of 5' cap on cellular mRNAs by the African swine fever virus (ASFV) decapping enzyme g5R protein (g5Rp) is beneficial to viral gene expression during the early stages of infection. As the only nucleoside diphosphate-linked moiety X (Nudix) decapping enzyme encoded in the ASFV genome, g5Rp works in both the degradation of cellular mRNA and the hydrolyzation of the diphosphoinositol polyphosphates. Here, we report the structures of dimeric g5Rp and its complex with inositol hexakisphosphate (InsP6). The two g5Rp protomers interact head to head to form a dimer, and the dimeric interface is formed by extensive polar and nonpolar interactions. Each protomer is composed of a unique N-terminal helical domain and a C-terminal classic Nudix domain. As g5Rp is an mRNA-decapping enzyme, we identified key residues, including K8, K94, K95, K98, K175, R221, and K243 located on the substrate RNA binding interfaces of g5Rp which are important to RNA binding and decapping enzyme activity. Furthermore, the g5Rp-mediated mRNA decapping was inhibited by InsP6. The g5Rp-InsP6 complex structure showed that the InsP6 molecules occupy the same regions that primarily mediate g5Rp-RNA interaction, elucidating the roles of InsP6 in the regulation of the viral decapping activity of g5Rp in mRNA degradation. Collectively, these results provide the structural basis of interaction between RNA and g5Rp and highlight the inhibitory mechanism of InsP6 on mRNA decapping by g5Rp. IMPORTANCE ASF is a highly contagious hemorrhagic viral disease in domestic pigs which causes high mortality. Currently, there are still no effective vaccines or specific drugs available against this particular virus. The protein g5Rp is the only viral mRNA-decapping enzyme, playing an essential role in the machinery assembly of mRNA regulation and translation initiation. In this study, we solved the crystal structures of g5Rp dimer and complex with InsP6. Structure-based mutagenesis studies revealed critical residues involved in a candidate RNA binding region, which also play pivotal roles in complex with InsP6. Notably, InsP6 can inhibit g5Rp activity by competitively blocking the binding of substrate mRNA to the enzyme. Our structure-function studies provide the basis for potential anti-ASFV inhibitor designs targeting the critical enzyme.

Nocardia rubra cell wall skeleton regulates tumour-associated macrophage polarization by reprogramming M2 macrophages into M1 macrophages via STAT1/STAT6 pathways.

Targeted therapy with tumour-associated macrophages (TAMs) has emerged as a new paradigm for immunotherapy of cervical cancer. Nocardia rubra cell wall skeleton (Nr-CWS) for external use is an immunotherapeutic agent. In this study, we aimed to explore the effects of Nr-CWS on TAMs and the potential mechanisms. Cervical tissue samples were collected before and after Nr-CWS treatment from patients with high-risk HPV infection and cervical intraepithelial neoplasia (CIN). The effect of Nr-CWS on macrophages in vivo was examined by immunohistochemistry and double-labeling immunofluorescence histochemistry. In vitro experiments were performed using a TAM model established by THP-1 cells under Nr-CWS treatment. We found that Nr-CWS treatment significantly reduced the numbers of total macrophages and M2 macrophages, increased the proportion of M1 macrophages and decreased the proportion of M2 macrophages in cervical tissues. After Nr-CWS treatment in vitro, the expression levels of the M1 macrophage markers were increased, while the expression levels of the M2 macrophage markers were decreased. Nr-CWS treatment also activated STAT1 pathways but inhibited STAT6 pathways. These results indicated that Nr-CWS may improve local immune response and reverse immunosuppression by regulating the M2 to M1 polarization of TAMs via STAT1/STAT6 pathways.

Lamina propria interleukin 17 A aggravates natural killer T-cell activation in autoimmune hepatitis.

Autoimmune hepatitis is an interface hepatitis characterized by the progressive destruction of the liver parenchyma, the cause of which is still obscure. Interleukin (IL)-17A is a major driver of autoimmunity, which can be produced by innate immune cells against several intracellular pathogens. Here, we investigated the involvement of IL-17A in a mice model of immune-mediated hepatitis with the intestine exposed to Salmonella typhimurium. Our results showed more severe Concanavalin (Con) A-induced liver injury and gut microbiome dysbiosis when the mice were treated with a gavage of S. typhimurium. Then, the natural killer (NK) T cells were overactivated by the accumulated IL-17A in the liver in the Con A and S. typhimurium administration group. IL-17A could activate NKT cells by inducing CD178 expression via IL-4/STAT6 signaling. Furthermore, via the portal tract, the laminae propria mucosal-associated invariant T (MAIT)-cell-derived IL-17A could be the original driver of NKT cell overactivation in intragastric administration of S. typhimurium and Con A injection. In IL-17A-deficient mice, Con A-induced liver injury and NKT cell activation were alleviated. However, when AAV-sh-mIL-17a was used to specifically knock down IL-17A in liver, it seemed that hepatic IL-17a knock down did not significantly influence the liver injury. Our results suggested that, under Con A-induction, laminae propria MAIT-derived IL-17A activated hepatic NKT, and this axis could be a therapeutic target in autoimmune liver disease.

Genome-wide association analyses of leaf rust resistance in cultivated emmer wheat.

KEY MESSAGE: Fifteen and eleven loci, with most loci being novel, were identified to associate with seedling and adult resistances, respectively, to the durum-specific races of leaf rust pathogen in cultivated emmer. Leaf rust, caused by Puccinia triticina (Pt), constantly threatens durum (Triticum turgidum ssp. durum) and bread wheat (Triticum aestivum) production worldwide. A Pt race BBBQD detected in California in 2009 poses a potential threat to durum production in North America because resistance source to this race is rare in durum germplasm. To find new resistance sources, we assessed a panel of 180 cultivated emmer wheat (Triticum turgidum ssp. dicoccum) accessions for seedling resistance to BBBQD and for adult resistance to a mixture of durum-specific races BBBQJ, CCMSS, and MCDSS in the field, and genotyped the panel using genotype-by-sequencing (GBS) and the 9 K SNP (Single Nucleotide Polymorphism) Infinium array. The results showed 24 and nine accessions consistently exhibited seedling and adult resistance, respectively, with two accessions providing resistance at both stages. We performed genome-wide association studies using 46,383 GBS and 4,331 9 K SNP markers and identified 15 quantitative trait loci (QTL) for seedling resistance located mostly on chromosomes 2B and 6B, and 11 QTL for adult resistance on 2B, 3B and 6A. Of these QTL, one might be associated with leaf rust resistance (Lr) gene Lr53, and two with the QTL previously reported in durum or hexaploid wheat. The remaining QTL are potentially associated with new Lr genes. Further linkage analysis and gene cloning are necessary to identify the causal genes underlying these QTL. The emmer accessions with high levels of resistance will be useful for developing mapping populations and adapted durum germplasm and varieties with resistance to the durum-specific races.

Effect of the Radial Velocity Distribution on the Loss Generation of a Contra-Rotating Fan in a Ventilation System.

Quantification of the loss generation of ducted contra-rotating fan (CRF) blades is difficult to achieve, since there are no guide vanes between rotors. A blade design program was established to investigate the relationship between radial velocity distribution and incurred loss. Numerical and experimental techniques were used to confirm the optimal configuration's overall performance. The relationship between loss and velocity distribution under the impact of spanwise load distribution was confirmed by the entropy contour from various perspectives. The appropriate radial velocity distribution can improve the operating efficiency of a CRF by reducing the entropy around the annulus under design and near-stall conditions. This regularity could provide some strategies in the design of contra-rotating blades.

Structural insights into multifunctionality of human FACT complex subunit hSSRP1.

Human structure-specific recognition protein 1 (hSSRP1) is an essential component of the facilitates chromatin transcription complex, which participates in nucleosome disassembly and reassembly during gene transcription and DNA replication and repair. Many functions, including nuclear localization, histone chaperone activity, DNA binding, and interaction with cellular proteins, are attributed to hSSRP1, which contains multiple well-defined domains, including four pleckstrin homology (PH) domains and a high-mobility group domain with two flanking disordered regions. However, little is known about the mechanisms by which these domains cooperate to carry out hSSRP1's functions. Here, we report the biochemical characterization and structure of each functional domain of hSSRP1, including the N-terminal PH1, PH2, PH3/4 tandem PH, and DNA-binding high-mobility group domains. Furthermore, two casein kinase II binding sites in hSSRP1 were identified in the PH3/4 domain and in a disordered region (Gly617-Glu709) located in the C-terminus of hSSRP1. In addition, a histone H2A-H2B binding motif and a nuclear localization signal (Lys677‒Asp687) of hSSRP1 are reported for the first time. Taken together, these studies provide novel insights into the structural basis for hSSRP1 functionality.

zVAD alleviates experimental autoimmune hepatitis in mice by increasing the sensitivity of macrophage to TNFR1-dependent necroptosis.

BACKGROUND & AIMS: Autoimmune hepatitis (AIH) is characterized by hepatocyte destruction, leading to lymphocyte and macrophage accumulation in the liver. Macrophages are key drivers of AIH. A membrane-permeable pan-caspase inhibitor, Z-Val-Ala-DL-Asp-fluoromethylketone (zVAD), induces macrophage necroptosis in response to certain stimuli. However, the function of zVAD in the pathogenesis of autoimmune hepatitis remains elusive. In this study, we aimed to evaluate the effect and explore the underlying mechanisms of zVAD against AIH. METHODS: Murine acute autoimmune liver injury was established by concanavalin A (ConA) injection. Bone marrow-derived macrophages (BMDMs) were used in adoptive cell transfer experiments. The mechanism of action of zVAD was examined using macrophage cell lines and BMDMs. Phosphorylation of mixed lineage kinase domain-like proteins was used as a marker of necroptosis. RESULTS: Treatment with zVAD increased necroptosis, reduced inflammatory cytokine production, and alleviated liver injury in a ConA-induced liver injury mouse model. Regardless of zVAD treatment, macrophage deletion resulted in reduced neutrophil accumulation and relieved ConA-induced liver injury. In vitro studies have shown that zVAD pretreatment promotes lipopolysaccharide-induced macrophage necroptosis and leads to reduced pro-inflammatory cytokine and chemokine secretion. Transferring zVAD-pretreated BMDMs in mice notably reduced ConA-associated liver inflammation and injury, resulting in lower mortality than that observed after transferring normal BMDMs. Mechanistically, zVAD treatment increased the expression of tumour necrosis factor receptor (TNFR)-1 and interleukin (IL)-10 in macrophages. TNFR1 expression decreased upon transfection with IL-10-specific small interfering RNAs and blocking of TNFR1 decreased macrophage necroptosis. CONCLUSIONS: We found that zVAD alleviated ConA-induced liver injury by increasing the sensitivity of macrophages to necroptosis via IL-10-induced TNFR1 expression. This study provides new insights into the treatment of autoimmune hepatitis via zVAD-induced macrophage necroptosis.

Protection of Exogenous Phosphocreatine for Myocardium in Percutaneous Coronary Intervention Related to Inflammation.

OBJECTIVES: Although injury of myocardium after percutaneous coronary intervention (PCI) has been reported, the mechanism and effect of exogenous phosphocreatine (PCr) supplementation on the injury are yet to be elucidated. Biomarkers, such as interleukin-6 (IL-6) and variations in white blood cells for inflammation, and serum cardiac troponin I (cTnI) for myocardial injury are examined. METHODS: A total of 105 patients undergoing PCI were included and randomly divided into two groups: control (treated with routine hydration therapy) and PCr (treated with additional intravenous infusion of exogenous PCr). The serum levels of biomarkers were detected at administration and 4, 12, 24, and 48 h after PCI, with natural logarithmic (loge) transformation of data when modeling assumptions were not fulfilled. RESULTS: The level of loge-transformed IL-6 increased in both groups, especially at 12 and 24 h after the operation, and that of PCr group was less than the control group at 48 h. The content of loge-transformed cTnI was significantly increased in both groups, while that of the PCr group was markedly lower than the control group at all time points after PCI. Moreover, the ratio of neutrophils was elevated at all time points after PCI, while that of the PCr group was lower at 48 h, and the variations in the ratio of lymphocytes showed opposite results. CONCLUSIONS: Exogenous phosphocreatine reduces stent implantation, triggers inflammation manifested as decreased serum levels of IL-6 and the aggregation of neutrophils, and protects the myocardium of the patients undergoing PCI. These findings provided the potential mechanism and treatment for myocardial injury associated with PCI.

Exosomes from human umbilical cord mesenchymal stem cells protect aortas in Db/db mice characterized by combination of metabolomics and proteomics.

Diabetic cardiovascular complication is a common systemic disease with high morbidity and mortality worldwide. We hypothesise that exosomes derived from human umbilical cord mesenchymal stem cells (hUCMSCs-exos) can rescue these disorders and alleviate vascular remodeling in diabetes. Morphological, non-targeted metabolomics and 4D label-free proteomics techniques were used to analyze the aortas of db/m mice as normal control group (NCA), saline treated db/db mice (DMA), and hUCMSCs-exos treated db/db mice (DMTA), and to clarify the molecular mechanism of the protection of hUCMSCs-exos in vascular remodeling from a new point of view. The results showed that 74 metabolites were changed significantly in diabetic aortas, of which 15 were almost restored by hUCMSCs-exos. In proteomics, 30 potential targets such as Stromal cell-derived factor 2-like protein 1, Leukemia inhibitory factor receptor, Peroxisomal membrane protein and E3 ubiquitin-protein ligase MYCBP2 were detected. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway-based analysis showed that Central carbon metabolism in cancer and Galactose metabolism pathway were up-regulated to near normal by hUCMSCs-exos in metabolomics, with janus associated kinase-signal transducer and activator of transcription (JAK-STAT) pathway displayed in proteomics. According to bioinformatics and integrated analysis, these targeted molecules of hUCMSCs-exos to attenuate the vascular remodeling were mainly associated with regulation of energy metabolism, oxidative stress, inflammation, and cellular communications. This study provided a reference for the therapy of diabetes-induced cardiovascular complications.