A single-chain antibody construct with specificity of a natural IgM antibody reduces hepatic ischemia reperfusion injury in mice.

Natural immunoglobulin M (IgM) antibodies have been shown to recognize post-ischemic neoepitopes following reperfusion of tissues and to activate complement. Specifically, IgM antibodies and complement have been shown to drive hepatic ischemia reperfusion injury (IRI). Herein, we investigate the therapeutic effect of C2 scFv (single-chain antibody construct with specificity of a natural IgM antibody) on hepatic IRI in C57BL/6 mice. Compared with PBS-treated mice, C2 scFv-treated mice displayed almost no necrotic areas, significant reduction in serum ALT, AST and LDH levels, and significantly reduced in the number of TUNEL positive cells. Moreover, C2 scFv-treated mice exhibited a notable reduction in inflammatory cells after hepatic IRI than PBS-treated mice. The serum IL-6, IL-1ß, TNF-α and MPC-1 levels were also severely suppressed by C2 scFv. Interestingly, C2 scFv reconstituted hepatic inflammation and IRI in Rag1-/- mice. We found that C2 scFv promoted hepatic cell death and increased inflammatory cytokines and infiltration of inflammatory cells after hepatic IRI in Rag1-/- mice. In addition, IgM and complement 3d (C3d) were deposited in WT mice and in Rag1-/- mice reconstituted with C2 scFv, indicating that C2 scFv can affect IgM binding and complement activation and reconstitute hepatic IRI. C3d expression was significantly lower in C57BL/6 mice treated with C2 scFv compared to PBS, indicating that excessive exogenous C2 scFv inhibited complement activation. These data suggest that C2 scFv alleviates hepatic IRI by blocking complement activation, and treatment with C2 scFv may be a promising therapy for hepatic IRI.

Ceramide kinase-mediated C1P metabolism attenuates acute liver injury by inhibiting the interaction between KEAP1 and NRF2.

Acute liver injury is the basis of the pathogenesis of diverse liver diseases. However, the mechanism underlying liver injury is complex and not completely understood. In our study, we revealed that CERK, which phosphorylates ceramide to produce ceramide-1-phosphate (C1P), was the sphingolipid pathway-related protein that had the most significantly upregulated expression during acute liver injury. A functional study confirmed that CERK and C1P attenuate hepatic injury both in vitro and in vivo through antioxidant effects. Mechanistic studies have shown that CERK and C1P positively regulate the protein expression of NRF2, which is a crucial protein that helps maintain redox homeostasis. Furthermore, our results indicated that C1P disrupted the interaction between NRF2 and KEAP1 by competitively binding to KEAP1, which allowed for the nuclear translocation of NRF2. In addition, pull-down assays and molecular docking analyses revealed that C1P binds to the DGR domain of KEAP1, which allows it to maintain its interaction with NRF2. Importantly, these findings were verified in human primary hepatocytes and a mouse model of hepatic ischemia‒reperfusion injury. Taken together, our findings demonstrated that CERK-mediated C1P metabolism attenuates acute liver injury via the binding of C1P to the DGR domain of KEAP1 and subsequently the release and nuclear translocation of NRF2, which activates the transcription of cytoprotective and antioxidant genes. Our study suggested that the upregulation of CERK and C1P expression may serve as a potential antioxidant strategy to alleviate acute liver injury.

Pulmonary interleukin 1 beta/serum amyloid A3 axis promotes lung metastasis of hepatocellular carcinoma by facilitating the pre-metastatic niche formation.

BACKGROUND: Increasing evidence suggests a vital role of the pre-metastatic niche in the formation of distant metastasis of many cancers. However, how the pre-metastatic niche is formed and promotes pulmonary metastasis of hepatocellular carcinoma (HCC) remains unknown. METHODS: Orthotopic liver tumor models and RNA-Seq were used to identify dysregulated genes in the pre-metastatic lung. Il1b knockout (Il1b-/-) mice and lentivirus-mediated gene knockdown/overexpression were utilized to demonstrate the role of interleukin 1 beta (IL-1ß)/serum amyloid A3 (SAA3) in the pre-metastatic niche formation and pulmonary metastasis. The potential molecular mechanisms were investigated by RNA-Seq, real-time quantitative PCR (qPCR), western blotting, immunohistochemistry (IHC), flow cytometry, luciferase reporter assay, double immunofluorescent staining and H&E staining. RESULTS: Accumulation of myeloid cells and upregulation of IL-1ß were observed in the pre-metastatic lung of orthotopic liver tumor models. Myeloid cells accumulation and pulmonary metastasis were suppressed in Il1b-/- mice and Il1r1-silencing mice. Mechanistically, SAA3 and matrix metallopeptidase 9 (MMP9) were identified as potential downstream targets of IL-1ß. Overexpression of SAA3 in the lungs of Il1b-/- mice restored myeloid cells accumulation and pulmonary metastasis of the orthotopic HCC xenografts. Moreover, alveolar macrophages-derived IL-1ß dramatically enhanced SAA3 expression in alveolar epithelial cells in an NF-κB dependent manner and increased MMP9 levels in an autocrine manner. Furthermore, SAA3 recruited myeloid cells to the lung without affecting the expression of MMP9 in myeloid cells. CONCLUSIONS: Our study suggests a key role of pulmonary IL-1ß and SAA3 in creating a permissive lung pre-metastatic niche by enhancing MMP9 expression and recruiting myeloid cells, respectively, thus promoting pulmonary metastasis of HCC.

Proteomic analysis of small extracellular vesicles from the plasma of patients with hepatocellular carcinoma.

PURPOSE: Liver cancer is one of the most common tumors with the seventh-highest incidence and the third-highest mortality. Many studies have shown that small extracellular vesicles (sEVs) play an important role in liver cancer. Here, we report comprehensive signatures for sEV proteins from plasma obtained from patients with hepatocellular carcinoma (HCC), which might be valuable for the evaluation and diagnosis of HCC. METHODS: We extracted sEVs from the plasma of controls and patients with HCC. Differentially expressed proteins in the sEVs were analyzed using label-free quantification and bioinformatic analyses. Western blotting (WB) was used to validate the abovementioned sEV proteins. RESULTS: Proteomic analysis was performed for plasma sEVs from 21 patients with HCC and 15 controls. Among the 335 identified proteins in our study, 27 were significantly dysregulated, including 13 upregulated proteins that were involved predominantly in the complement cascade (complement C1Q subcomponent subunit B (C1QB), complement C1Q subcomponent subunit C (C1QC), C4B-binding protein alpha chain (C4BPA), and C4B-binding protein beta chain (C4BPB)) and the coagulation cascade (F13B, fibrinogen alpha chain (FGA), fibrinogen beta chain (FGB), and fibrinogen gamma chain (FGG)). We verified increased levels of the C1QB, C1QC, C4BPA, and C4BPB proteins in the plasma sEVs from patients with HCC in both the discovery cohort and validation cohort. CONCLUSIONS: The complement cascade in sEVs was significantly involved in HCC progression. C1QB, C1QC, C4BPA, and C4BPB were highly abundant in the plasma sEVs from patients with HCC and might represent molecular signatures.

S1PR1 induces metabolic reprogramming of ceramide in vascular endothelial cells, affecting hepatocellular carcinoma angiogenesis and progression.

Angiogenesis is a fundamental process underlying the occurrence, growth and metastasis of hepatocellular carcinoma (HCC), a prevalent tumour type with an extremely poor prognosis due to abundant vasculature. However, the underlying mechanism of angiogenesis in HCC remains largely unknown. Herein, we found that sphingosine-1-phosphate receptor 1 (S1PR1) plays an important role in HCC angiogenesis. S1PR1 was found to be selectively and highly expressed in the blood vessels of HCC tissues compared with those of paratumour tissues. Functionally, high expression of S1PR1 in endothelial cells (ECs) promoted angiogenesis and progression of HCC in vitro and in vivo. Mechanistically, proangiogenic factors (S1P, IL-6, VEGFA) in conditioned medium from HCC cells induced the upregulation of S1PR1 in ECs via the phosphorylation of STAT3 at Y705. Further study also revealed that S1PR1 promotes angiogenesis by decreasing ceramide levels via CerS3 downregulation. Interestingly, we demonstrated that S1PR1 downregulates CerS3 by inducing CerS6 translocation into the nucleus to inhibit CerS3 at the transcriptional level in ECs. In addition, we found that a high concentration of Lenvatinib significantly downregulated the expression of S1PR1 and obviously enhanced S1PR1 knockdown-mediated angiogenesis inhibition, indicating that S1PR1 may be a target by which Lenvatinib combats angiogenesis in HCC. Thus, S1PR1 may be an important target for suppressing angiogenesis in HCC, and inhibiting S1PR1 is a promising approach to antitumor therapy in HCC.

C1orf74 positively regulates the EGFR/AKT/mTORC1 signaling in lung adenocarcinoma cells.

Background: Lung adenocarcinoma (LUAD) is a major type of lung cancer with poor prognosis and low 5-year survival rate, which urgently needs further investigation in order to elucidate its mechanisms completely and discover novel therapeutic targets. C1orf74 is a novel protein with unknown function either in normal cells or cancer cells. The aim of this study is to investigate the expression and function of C1orf74 in LUAD cells. Methods: The expression of C1orf74 in LUAD was analyzed using the LUAD datasets from public databases. The prognostic value of C1orf74 in LUAD was analyzed using Kaplan-Meier Plotter. C1orf74 expression in LUAD cell line A549, H1993 and HCC827 was silenced using small interfering RNA, and then the effects of C1orf74 knockdown on proliferation, migration and invasion of LUAD cells were detected by colony formation assay and Transwell assay, the role of C1orf74 in EGFR/AKT/mTORC1 signaling pathway was examined by Western blot, and the function of C1orf74 in cell cycle was detected by flow cytometry. Results: The results of LUAD clinical data showed that C1orf74 was upregulated in LUAD tissues, and its high expression was associated with poor prognosis. The results from cultured LUAD cells demonstrated that C1orf74 knockdown inhibited cell proliferation, migration and invasion, but induced cell cycle arrest and autophagy. Moreover, C1orf74 knockdown suppressed EGFR/AKT/mTORC1 signaling in LUAD cells. In conclusion, the present study revealed that C1orf74 is upregulated in LUAD tissues and plays an oncogenic role in LUAD, and that C1orf74 positively regulates cell proliferation and mobility through the EGFR/AKT/mTORC1 signaling pathway in LUAD cells.

p38 MAPK Is a Major Regulator of Amyloid Beta-Induced IL-6 Expression in Human Microglia.

The accumulation of amyloid beta (Aß) plaques in the brain is a hallmark of Alzheimer's disease (AD) pathology. Microglial activation-mediated neuroinflammation has been implicated in the pathogenesis of AD and the expression levels of interleukin-6 (IL-6) were increased in the brains of AD patients. However, the mechanisms by which IL-6 expression is regulated in human microglia are incompletely understood. Here, we show that Aß1-40 oligomers (Aß40) dose-dependently stimulate IL-6 expression in HMC3 human microglial cells. Treatment with Aß40 promotes the transcription of IL-6 and tumor necrosis factor α (TNFα) mRNAs in both HMC3 and THP-1 cells. Mechanistic studies reveal that Aß40-induced increase of IL-6 secretion is associated with the activation of p38 mitogen-activated protein kinase (p38 MAPK). Inhibition of p38 MAPK by BIRB 796 or SB202190 abrogates Aß40-induced increase of IL-6 production. Through analyzing brain specimens, we found that the immunoreactivity for IL-6 and phosphorylated (the activated form) p38 MAPK was markedly higher in microglia of AD patients than in age-matched control subjects. Moreover, our studies identified the co-localization of IL-6 with phosphorylated p38 MAPK in microglia in the cortices of AD patients. Taken together, these results indicate that p38 MAPK is a major regulator of Aß-induced IL-6 production in human microglia, which suggests that targeting p38 MAPK may represent a new approach to ameliorate Aß accumulation-induced neuroinflammation in AD.

GBA1-dependent membrane glucosylceramide reprogramming promotes liver cancer metastasis via activation of the Wnt/ß-catenin signalling pathway.

The effect of glucosylceramide (GlcCer) reprogramming on liver cancer metastasis remains poorly understood. In this study, we demonstrated that the protein expression of GBA1, which catalyses the conversion of GlcCer to ceramide, was downregulated in liver cancer tissue. A clinical relevance analysis revealed that low expression of GBA1 was associated with the metastatic potential of liver cancer cells. Furthermore, loss- and gain-of-function studies confirmed that low expression of GBA1 promoted metastasis of liver cancer both in vitro and in vivo. Mechanistic studies indicated that low expression of GBA1 enhanced the metastatic ability of liver cancer by promoting the epithelial-mesenchymal transition (EMT), in which Wnt signalling pathway is involved. In the plasma membrane (PM), GBA1-dependent GlcCer reprogramming increased LRP6 location in the PM leading to an interaction between GlcCer and LRP6, subsequently promoting LRP6 phosphorylation at Ser1490, and finally activating the Wnt/ß-catenin signalling pathway. To our knowledge, this is the first time to be found that GlcCer interacted with a protein. In addition, the results of mass spectrometry indicated that GlcCer d18:1/18:0 was the most notably increased studied species in the PM when GBA1 was downregulated, suggesting that GlcCer d18:1/18:0 may be the major functional lipid that promotes GBA1-dependent liver cancer metastasis. Thus, GBA1-mediated GlcCer reprogramming in the PM promotes metastasis of liver cancer via activation of the Wnt/ß-catenin signalling pathway, upregulation of GBA1 may be a potential therapeutic strategy to combat liver cancer metastasis.

Characterization of Novel P-Selectin Targeted Complement Inhibitors in Murine Models of Hindlimb Injury and Transplantation.

The complement system has long been recognized as a potential druggable target for a variety of inflammatory conditions. Very few complement inhibitors have been approved for clinical use, but a great number are in clinical development, nearly all of which systemically inhibit complement. There are benefits of targeting complement inhibition to sites of activation/disease in terms of efficacy and safety, and here we describe P-selectin targeted complement inhibitors, with and without a dual function of directly blocking P-selectin-mediated cell-adhesion. The constructs are characterized in vitro and in murine models of hindlimb ischemia/reperfusion injury and hindlimb transplantation. Both constructs specifically targeted to reperfused hindlimb and provided protection in the hindlimb ischemia/reperfusion injury model. The P-selectin blocking construct was the more efficacious, which correlated with less myeloid cell infiltration, but with similarly reduced levels of complement deposition. The blocking construct also improved tissue perfusion and, unlike the nonblocking construct, inhibited coagulation, raising the possibility of differential application of each construct, such as in thrombotic vs. hemorrhagic conditions. Similar outcomes were obtained with the blocking construct following vascularized composite graft transplantation, and treatment also significantly increased graft survival. This is outcome may be particularly pertinent in the context of vascularized composite allograft transplantation, since reduced ischemia reperfusion injury is linked to a less rigorous alloimmune response that may translate to the requirement of a less aggressive immunosuppressive regime for this normally nonlife-threatening procedure. In summary, we describe a new generation of targeted complement inhibitor with multi-functionality that includes targeting to vascular injury, P-selectin blockade, complement inhibition and anti-thrombotic activity. The constructs described also bound to both mouse and human P-selectin which may facilitate potential translation.

Biomimetic Hydrogels Loaded with Nanofibers Mediate Sustained Release of pDNA and Promote In Situ Bone Regeneration.

Polymer hydrogels are generally insufficient biomechanics, strong resistance to cell adhesion, and weak bioactivity which limits their application in bone tissue engineering considerably. In order to develop a bone tissue engineering material with both good mechanical properties, osteogenic and angiogenic activity. Nanofibers carrying DNA plasmid (pNF) are introduced to gelatin methacryloyl (GelMA) and thiolated chitosan (TCS) system for preparing a novel GelMA/TCS/pNF composite hydrogel with dual network structure. By characterization of the compressive measurements, the resulting composite scaffold shows greatly enhanced mechanical strength (0.53 MPa) and is not damaged after 20 cycles of compression. And the fabricated composite scaffold displays sustained release of bone morphogenetic protein-2 that can induce osteogenic differentiation and angiopoietin-1 that promotes vascularization. The cell experiment shows that this system can significantly promote MC3T3-E1 cell attachment, proliferation, as well as osteogenic-related and angiogenic-related genes expression of MC3T3-E1 cells. Moreover, the in vivo results show that the composite scaffold with activated gene fibers can significantly promote osteogenesis and vascularization leading to favorable capacity of bone regeneration, meaning that the resulting biomimetic composite hydrogel scaffolds are excellent candidates for bone repair materials.

Mogroside IIE Inhibits Digestive Enzymes via Suppression of Interleukin 9/Interleukin 9 Receptor Signalling in Acute Pancreatitis.

The incidence of pancreatitis (AP) is increasing and there is no specific treatment available. Intracellular digestive enzyme activation is a key event in the pathogenesis of AP downstream of cytosolic calcium overload and impaired autophagy. Siraitia grosvenorii (Swingle) was used in Traditional Chinese Medicine to reduce inflammation and facilitate bowel movement. The bioactive components of this plant show hypolipedimic, antidiabetic, antifibrotic activity and have been used against pancreatic cancer. Here, we examined whether mogroside IIE, a major bioactive component of unripe S. grosvenorii fruit, can protect against AP. We found that mogroside IIE decreased the activity of trypsin and cathepsin B induced by cerulein plus lipopolysaccharide (LPS) in the pancreatic acinar cell line AR42J and primary acinar cells in a dose- and time-dependent manner. Mogroside IIE treatment decreased the levels of serum lipase and serum amylase in mice injected with cerulein plus LPS without influencing inflammation significantly. A multi-cytokine array revealed that mogroside IIE decreased the level of interleukin 9 (IL-9) in AP mice. Exogenous IL-9 eliminated the mogroside IIE induced reduction of trypsin and cathepsin B activity and reversed the inhibition of cytosolic calcium and modulation of autophagy mediated by mogroside IIE. An IL-9 receptor antibody neutralized the effect of IL-9, restoring mogroside IIE activity. The mogroside IIE targeted IL-9 may partially arise from Th9 cells. Taken together, we provide experimental evidence that mogroside IIE ameliorates AP in cell models and mice through downregulation of the IL-9/IL-9 receptor pathway.

Human alkaline ceramidase 2 promotes the growth, invasion, and migration of hepatocellular carcinoma cells via sphingomyelin phosphodiesterase acid-like 3B.

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. It has a poor prognosis because it is often diagnosed at the advanced stage when treatments are limited. In addition, HCC pathogenesis is not fully understood, and this has affected early diagnosis and treatment of this disease. Human alkaline ceramidase 2 (ACER2), a key enzyme that regulates hydrolysis of cellular ceramides, affects cancer cell survival, however its role in HCC has not been well characterized. Our results showed that ACER2 is overexpressed in HCC tissues and cell lines. In addition, high ACER2 protein expression was associated with tumor growth; ACER2 knockdown resulted in decreased cell growth and migration. Sphingomyelin phosphodiesterase acid-like 3B (SMPDL3B) promoted HCC cell growth, invasion, and migration; SMPDL3B knockdown had a significant inhibitory effect on HCC tumor growth in vivo. Moreover, ACER2 positively regulated the protein level of SMPDL3B. Of note, ACER2/SMPDL3B promoted ceramide hydrolysis and S1P production. This axis induced HCC survival and could be blocked by inhibition of S1P formation. In conclusion, ACER2 promoted HCC cell survival and migration, possibly via SMPDL3B. Thus, inhibition of ACER2/SMPDL3B may be a novel therapeutic target for HCC treatment.

Targeting CLK3 inhibits the progression of cholangiocarcinoma by reprogramming nucleotide metabolism.

CDC-like kinase 3 (CLK3) is a dual specificity kinase that functions on substrates containing serine/threonine and tyrosine. But its role in human cancer remains unknown. Herein, we demonstrated that CLK3 was significantly up-regulated in cholangiocarcinoma (CCA) and identified a recurrent Q607R somatic substitution that represented a gain-of-function mutation in the CLK3 kinase domain. Gene ontology term enrichment suggested that high CLK3 expression in CCA patients mainly was associated with nucleotide metabolism reprogramming, which was further confirmed by comparing metabolic profiling of CCA cells. CLK3 directly phosphorylated USP13 at Y708, which promoted its binding to c-Myc, thereby preventing Fbxl14-mediated c-Myc ubiquitination and activating the transcription of purine metabolic genes. Notably, the CCA-associated CLK3-Q607R mutant induced USP13-Y708 phosphorylation and enhanced the activity of c-Myc. In turn, c-Myc transcriptionally up-regulated CLK3. Finally, we identified tacrine hydrochloride as a potential drug to inhibit aberrant CLK3-induced CCA. These findings demonstrate that CLK3 plays a crucial role in CCA purine metabolism, suggesting a potential therapeutic utility.

A completeness-independent method for pre-selection of closely related genomes for species delineation in prokaryotes.

BACKGROUND: Whole-genome approaches are widely preferred for species delineation in prokaryotes. However, these methods require pairwise alignments and calculations at the whole-genome level and thus are computationally intensive. To address this problem, a strategy consisting of sieving (pre-selecting closely related genomes) followed by alignment and calculation has been proposed. RESULTS: Here, we initially test a published approach called "genome-wide tetranucleotide frequency correlation coefficient" (TETRA), which is specially tailored for sieving. Our results show that sieving by TETRA requires > 40% completeness for both genomes of a pair to yield > 95% sensitivity, indicating that TETRA is completeness-dependent. Accordingly, we develop a novel algorithm called "fragment tetranucleotide frequency correlation coefficient" (FRAGTE), which uses fragments rather than whole genomes for sieving. Our results show that FRAGTE achieves ~ 100% sensitivity and high specificity on simulated genomes, real genomes and metagenome-assembled genomes, demonstrating that FRAGTE is completeness-independent. Additionally, FRAGTE sieved a reduced number of total genomes for subsequent alignment and calculation to greatly improve computational efficiency for the process after sieving. Aside from this computational improvement, FRAGTE also reduces the computational cost for the sieving process. Consequently, FRAGTE extremely improves run efficiency for both the processes of sieving and after sieving (subsequent alignment and calculation) to together accelerate genome-wide species delineation. CONCLUSIONS: FRAGTE is a completeness-independent algorithm for sieving. Due to its high sensitivity, high specificity, highly reduced number of sieved genomes and highly improved runtime, FRAGTE will be helpful for whole-genome approaches to facilitate taxonomic studies in prokaryotes.

Anticancer Activity of Platinum (II) Complex with 2-Benzoylpyridine by Induction of DNA Damage, S-Phase Arrest, and Apoptosis.

OBJECTIVE: To overcome the disadvantages of cisplatin, numerous platinum (Pt) complexes have been prepared. However, the anticancer activity and mechanism of Pt(II) complexed with 2-benzoylpyridine [Pt(II)- Bpy]: [PtCl2(DMSO)L] (DMSO = dimethyl sulfoxide, L = 2-benzoylpyridine) in cancer cells remain unknown. METHODS: Pt(II)-Bpy was synthesized and characterized by spectrum analysis. Its anticancer activity and underlying mechanisms were demonstrated at the cellular, molecular, and in vivo levels. RESULTS: Pt(II)-Bpy inhibited tumor cell growth, especially HepG2 human liver cancer cells, with a halfmaximal inhibitory concentration of 9.8±0.5µM, but with low toxicity in HL-7702 normal liver cells. Pt(II)- Bpy induced DNA damage, which was demonstrated through a marked increase in the expression of cleavedpoly (ADP ribose) polymerase (PARP) and gamma-H2A histone family member X and a decrease in PARP expression. The interaction of Pt(II)-Bpy with DNA at the molecular level was most likely through an intercalation mechanism, which might be evidence of DNA damage. Pt(II)-Bpy initiated cell cycle arrest at the S phase in HepG2 cells. It also caused severe loss of the mitochondrial membrane potential; a decrease in the expression of caspase-9 and caspase-3; an increase in reactive oxygen species levels; the release of cytochrome c and apoptotic protease activation factor; and the activation of caspase-9 and caspase-3 in HepG2 cells, which in turn resulted in apoptosis. Meanwhile, changes in p53 and related proteins were observed including the upregulation of p53, the phosphorylation of p53, p21, B-cell lymphoma-2-associated X protein, and NOXA; and the downregulation of B-cell lymphoma 2. Moreover, Pt(II)-Bpy displayed marked inhibitory effects on tumor growth in the HepG2 nude mouse model. CONCLUSION: Pt(II)-Bpy is a potential candidate for cancer chemotherapy.

Serum sphingosine negatively correlates with albumin predicting the risk of hepatocellular carcinoma.

BACKGROUND: The roles of sphingosine in various cancers have not been fully investigated. Our aim was to identify the relationship between serum sphingosine and the risk of hepatocellular carcinoma (HCC). METHODS: Serum sphingosine in 34 normal people and 73 HCC patients were reviewed retrospectively. Receiver operating characteristic curve analysis was performed to determine the cut-off values of sphingosine in the serum. Chi-square test, t test and regression analysis were used to test the association between serum sphingosine and individual clinicopathologic parameters. RESULTS: Serum sphingosine was higher in HCC patients (155.91±331.5 ng/mL) with normal persons as the control (30.92±29.4 ng/mL). The sphingosine threshold according to ROC curve was set at 22.5 ng/mL with a sensitivity of 74%, and a specificity of 55.9%. Meanwhile, sphingosine in HCC patients with abnormal albumin was significantly higher than that in patients with normal albumin (t=2.452, P=0.019). When HCC patients were divided into two groups serum sphingosine was negatively associated with albumin in HCC patients (χ2=4.469, P=0.035). Moreover, the logistic regression analysis showed that large tumor size (P=0.018, OR=0.13) and a low albumin (P=0.005, OR=8.856) were two independent risk factors for serum sphingosine upregulation. High AFP coupled with high serum sphingosine, high sphingosine and high AFP respectively were found in 91.2%, 75.4%, 73% of the HCC patients. CONCLUSIONS: These results suggest that serum sphingosine could be treated as a marker for the risk of HCC. AFP and sphingosine in the serum could be used together for HCC diagnosis.

Serum AKR1B10 predicts the risk of hepatocellular carcinoma - A retrospective single-center study / El AKR1B10 sérico predice el riesgo de carcinoma hepatocelular. Un estudio retrospectivo unicéntrico

Objectives: AKR1B10, first cloned from liver cancer tissues, has recently been reported to be up-regulated significantly in hepatocellular carcinoma (HCC) tissues, but the relationship between serum level of AKR1B10 and the risk of HCC is not understood. Methods: 170 HCC patients and 120 health donors from October 2014 to March 2017 were recruited in the affiliated hospital of Guilin Medical University. Serum AKR1B10 in all cases were detected and in 30 HCC patients were analyzed preoperatively and postoperatively by Time-resolved fluoroimmunoassay. Results: The level of serum AKR1B10 was significantly higher in HCC patients (1800.24±2793.79) than in health donors (129.34±194.129), and downregulation of serum AKR1B10 in HCC patients was observed after hepatectomy. When samples were grouped according to the serum level of AKR1B10 (≥232.7pg/ml), serum AKR1B10 positively correlated to serum AFP (χ2=6.295, P=0.012), ALT (χ2=18.803, P=0.000), AST (χ2=33.421, P=0.000), tumor nodule number (χ2=6.777, P=0.009), cirrhosis (χ2=43.458, P=0.000), and tumor size (χ2=6.042, P=0.014) in the Chi-square test. Conclusions: Diagnosis of HCC could be improved using the both predictors of serum AKR1B10 and AFP. AKR1B10 was thus considered to be a new serological biomarker for HCC

Objetivos: Recientemente se ha notificado que el AKR1B10, clonado por primera vez a partir de tejidos hepáticos cancerosos, se encuentra aumentado de forma significativa en tejidos afectados por carcinoma hepatocelular (CHC), aunque no se comprende la relación entre la concentración sérica de AKR1B10 y el riesgo de CHC. Métodos: Se incluyeron 170 pacientes con CHC y 120 donantes sanos desde octubre de 2014 a marzo de 2017 en el hospital afiliado a la Guilin Medical University. Se analizó el AKR1B10 en todos los casos y en 30 pacientes con CHC antes y después de la cirugía, mediante fluoroinmunoensayo a tiempo resuelto. Resultados: La concentración sérica de AKR1B10 fue significativamente mayor en los pacientes con CHC (1.800,24±2.793,79) que en los donantes sanos (129,34±194,129), y se observó una reducción del AKR1B10 sérico en los pacientes con CHC tras la hepatectomía. Cuando se agruparon las muestras en función de la concentración sérica de AKR1B10 (≥ 232,7pg/ml), el AKR1B10 sérico se correlacionó positivamente con la AFP sérica (χ2=6,295; p=0,012), la ALT (χ2=18,803; p=0,000), la AST (χ2=33,421; p=0,000), la cifra de nódulos tumorales (χ2=6,777; p=0,009), la presencia de cirrosis (χ2=43,458; p=0,000) y el tamaño tumoral (χ2=6,042; p=0,014) en la prueba de χ2. Conclusiones: Podría mejorarse el diagnóstico del CHC usando los 2 factores pronósticos de AKR1B10 sérico y AFP. Por lo tanto, el AKR1B10 se consideró un nuevo biomarcador serológico del CHC

A high-resolution genomic composition-based method with the ability to distinguish similar bacterial organisms.

BACKGROUND: Genomic composition has been found to be species specific and is used to differentiate bacterial species. To date, almost no published composition-based approaches are able to distinguish between most closely related organisms, including intra-genus species and intra-species strains. Thus, it is necessary to develop a novel approach to address this problem. RESULTS: Here, we initially determine that the "tetranucleotide-derived z-value Pearson correlation coefficient" (TETRA) approach is representative of other published statistical methods. Then, we devise a novel method called "Tetranucleotide-derived Z-value Manhattan Distance" (TZMD) and compare it with the TETRA approach. Our results show that TZMD reflects the maximal genome difference, while TETRA does not in most conditions, demonstrating in theory that TZMD provides improved resolution. Additionally, our analysis of real data shows that TZMD improves species differentiation and clearly differentiates similar organisms, including similar species belonging to the same genospecies, subspecies and intraspecific strains, most of which cannot be distinguished by TETRA. Furthermore, TZMD is able to determine clonal strains with the TZMD = 0 criterion, which intrinsically encompasses identical composition, high average nucleotide identity and high percentage of shared genomes. CONCLUSIONS: Our extensive assessment demonstrates that TZMD has high resolution. This study is the first to propose a composition-based method for differentiating bacteria at the strain level and to demonstrate that composition is also strain specific. TZMD is a powerful tool and the first easy-to-use approach for differentiating clonal and non-clonal strains. Therefore, as the first composition-based algorithm for strain typing, TZMD will facilitate bacterial studies in the future.

Lipopolysaccharide enhances DNA-induced IFN-ß expression and autophagy by upregulating cGAS expression in A549 cells.

Cyclic guanosine monophosphate-adenosine monophosphate synthase (cGAS) is a newly identified cytosolic DNA sensor, but its function in lung epithelial cells is relatively unknown. In the present study, the effects of lipopolysaccharide (LPS) on the expression and function of cGAS in the A549 lung epithelial cell line was investigated. The cells were treated with LPS at different concentrations (e.g., 100, 200, 400 and 800 ng/ml), and the cGAS expression levels were examined via western blot analysis and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The cells were pretreated with LPS, followed by E. coli DNA transfection using Lipofectamine® 3000. After 24 h, interferon (IFN)-ß production was measured using ELISA and the expression of the autophagic markers, microtubule-associated proteins 1A/1B light chain 3 and sequestosome-1, were determined using western blot analysis. The cells were also pretreated with either a toll-like receptor (TLR) 4 inhibitor, a serine/threonine-protein kinase TBK1 (TBK1) inhibitor or an nuclear factor (NF)-κB inhibitor, followed by LPS treatment, and the cGAS expression levels were examined via western blot analysis and RT-qPCR. The result showed that LPS treatment upregulated cGAS expression in a dose-dependent manner. E. coli DNA treatment could induce IFN-ß production and autophagy via cGAS, which was enhanced by LPS pretreatment. The effect of LPS on cGAS expression was suppressed by treatment with a TLR4 inhibitor, a TBK1 inhibitor and an NF-κB inhibitor. In conclusion, LPS enhances DNA-induced IFN-ß production and autophagy by upregulating cGAS expression through the myeloid differentiation primary response protein MyD88-independent TLR4 signaling pathway in A549 cells.