Genistein promotes M1 macrophage apoptosis and reduces inflammatory response by disrupting miR-21/TIPE2 pathway.

Cardiovascular diseases are a major cause of mortality, and vascular injury, a common pathological basis of cardiovascular disease, is deeply correlated with macrophage apoptosis and inflammatory response. Genistein, a type of phytoestrogen, exerts cardiovascular protective activities, but the underlying mechanism has not been fully elucidated. In this study, RAW264.7 cells were treated with genistein, lipopolysaccharide (LPS), nuclear factor-kappa B (NF-κB) inhibitor, and/or protein kinase B (AKT) agonist to determine the role of genistein in apoptosis and inflammation in LPS-stimulated cells. Simultaneously, high fat diet-fed C57BL/6 mice were administered genistein to evaluate the function of genistein on LPS-induced cardiovascular injury mouse model. Here, we demonstrated that LPS obviously increased apoptosis resistance and inflammatory response of macrophages by promoting miR-21 expression, and miR-21 downregulated tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) expression by targeting the coding region. Genistein reduced miR-21 expression by inhibiting NF-κB, then blocked toll-like receptor 4 (TLR4) pathway and AKT phosphorylation dependent on TIPE2, resulting in inhibition of LPS. Our research suggests that miR-21/TIPE2 pathway is involved in M1 macrophage apoptosis and inflammatory response, and genistein inhibits the progression of LPS-induced cardiovascular injury at the epigenetic level via regulating the promoter region of Vmp1 by NF-κB.

Comparative analysis of chlorambucil-induced DNA lesion formation and repair in a spectrum of different human cell systems.

Chlorambucil (CLB) belongs to the class of nitrogen mustards (NMs), which are highly reactive bifunctional alkylating agents and were the first chemotherapeutic agents developed. They form DNA interstrand crosslinks (ICLs), which cause a blockage of DNA strand separation, inhibiting essential processes in DNA metabolism like replication and transcription. In fast replicating cells, e.g., tumor cells, this can induce cell death. The upregulation of ICL repair is thought to be a key factor for the resistance of tumor cells to ICL-inducing cytostatic agents including NMs. To monitor induction and repair of CLB-induced ICLs, we adjusted the automated reversed fluorometric analysis of alkaline DNA unwinding assay (rFADU) for the detection of ICLs in adherent cells. For the detection of monoalkylated DNA bases we established an LC-MS/MS method. We performed a comparative analysis of adduct formation and removal in five human cell lines and in peripheral blood mononuclear cells (PBMCs) after treatment with CLB. Dose-dependent increases in adduct formation were observed, and suitable treatment concentrations were identified for each cell line, which were then used for monitoring the kinetics of adduct formation. We observed significant differences in the repair kinetics of the cell lines tested. For example, in A2780 cells, hTERT immortalized VH10 cells, and in PBMCs a time-dependent repair of the two main monoalkylated DNA-adducts was confirmed. Regarding ICLs, repair was observed in all cell systems except for PBMCs. In conclusion, LC-MS/MS analyses combined with the rFADU technique are powerful tools to study the molecular mechanisms of NM-induced DNA damage and repair. By applying these methods to a spectrum of human cell systems of different origin and transformation status, we obtained insight into the cell-type specific repair of different CLB-induced DNA lesions, which may help identify novel resistance mechanisms of tumors and define molecular targets for therapeutic interventions.

Understanding the Role of GLUT2 in Dysglycemia Associated with Fanconi-Bickel Syndrome.

Fanconi−Bickel Syndrome (FBS) is a rare disorder of carbohydrate metabolism that is characterized by the accumulation of glycogen mainly in the liver. It is inherited in an autosomal recessive manner due to mutations in the SLC2A2 gene. SLC2A2 encodes for the glucose transporter GLUT2 and is expressed in tissues that are involved in glucose homeostasis. The molecular mechanisms of dysglycemia in FBS are still not clearly understood. In this study, we report two cases of FBS with classical phenotypes of FBS associated with dysglycemia. Genomic DNA was extracted and analyzed by whole-genome and Sanger sequencing, and patient PBMCs were used for molecular analysis. One patient had an exonic SLC2A2 mutation (c.1093C>T in exon 9, R365X), while the other patient had a novel intronic SLC2A2 mutation (c.613-7T>G). Surprisingly, the exonic mutation resulted in the overexpression of dysfunctional GLUT2, resulting in the dysregulated expression of other glucose transporters. The intronic mutation did not affect the coding sequence of GLUT2, its expression, or glucose transport activity. However, it was associated with the expression of miRNAs correlated with type 1 diabetes mellitus, with a particular significant overexpression of hsa-miR-29a-3p implicated in insulin production and secretion. Our findings suggest that SLC2A2 mutations cause dysglycemia in FBS either by a direct effect on GLUT2 expression and/or activity or, indirectly, by the dysregulated expression of miRNAs implicated in glucose homeostasis.

A Path-Based Analysis of Infected Cell Line and COVID-19 Patient Transcriptome Reveals Novel Potential Targets and Drugs Against SARS-CoV-2.

Most transcriptomic studies of SARS-CoV-2 infection have focused on differentially expressed genes, which do not necessarily reveal the genes mediating the transcriptomic changes. In contrast, exploiting curated biological network, our PathExt tool identifies central genes from the differentially active paths mediating global transcriptomic response. Here we apply PathExt to multiple cell line infection models of SARS-CoV-2 and other viruses, as well as to COVID-19 patient-derived PBMCs. The central genes mediating SARS-CoV-2 response in cell lines were uniquely enriched for ATP metabolic process, G1/S transition, leukocyte activation and migration. In contrast, PBMC response reveals dysregulated cell-cycle processes. In PBMC, the most frequently central genes are associated with COVID-19 severity. Importantly, relative to differential genes, PathExt-identified genes show greater concordance with several benchmark anti-COVID-19 target gene sets. We propose six novel anti-SARS-CoV-2 targets ADCY2, ADSL, OCRL, TIAM1, PBK, and BUB1, and potential drugs targeting these genes, such as Bemcentinib, Phthalocyanine, and Conivaptan.

T Lymphocyte Serotonin 5-HT7 Receptor Is Dysregulated in Natalizumab-Treated Multiple Sclerosis Patients.

Serotonin (5-HT) is known as a potent immune cell modulator in autoimmune diseases and should be protective in the pathogenesis of multiple sclerosis (MS). Nevertheless, there is limited knowledge about receptors involved in 5-HT effects as well as induced mechanisms. Among 5-HT receptors, the 5-HT7 receptor is able to activate naïve T cells and influence the inflammatory response; however, its involvement in the disease has never been studied so far. In this study, we collected blood sample from three groups: acute relapsing MS patients (ARMS), natalizumab-treated MS patients (NTZ), and control subjects. We investigated the 5-HT7 expression on circulating lymphocytes and evaluated the effects of its activation on cytokine production with peripheral blood mononuclear cell (PBMC) cultures. We found a significant increase in the 5-HT7 surface expression on T lymphocytes and on the different CD4+ T cell subsets exclusively in NTZ-treated patients. We also showed that the selective agonist 5-carboxamidotryptamine (5-CT)-induced 5-HT7R activation significantly promotes the production of IL-10, a potent immunosuppressive cytokine in PBMCs. This study provides for the first time a dysregulation of 5-HT7 expression in NTZ-MS patients and its ability to promote IL-10 release, suggesting its protective role. These findings strengthen the evidence that 5-HT7 may play a role in the immuno-protective mechanisms of NTZ in MS disease and could be considered as an interesting therapeutic target in MS.

Serum levels of vitamin D and immune system function in patients with COVID-19 admitted to intensive care unit.

OBJECTIVE: Vitamin D is believed to affect the functionality of the immune system for the prevention of coronavirus disease. To investigate the role of this vitamin against the Coronavirus, this study analyzed the serum levels of vitamin D, the transcription pattern of inflammatory cytokines, and the frequency of total lymphocytes, TCD4+, TCD8+, and NK cells in 50 COVID-19-affected subjects in comparison to 50 healthy participants. MATERIALS AND METHODS: This study diagnosed and evaluated 100 patients. Frequency of lymphocytes was determined using flow cytometry. Cytokine expression levels were measured using Real-Time PCR. Serum levels of vitamin D and cytokines levels in cultured cell supernatant were measured by ELISA. RESULTS: Patients with COVID-19 exhibited decreased serum levels of vitamin D versus the healthy participants (p = 0.0024). The total number of lymphocytes, TCD4+, TCD8+, and NK cells was significantly reduced in patients with COVID-19 (p < 0.0001). Considerable upregulation of IL-12, IFN-γ, and TNF-α was seen in COVID-19 patients compared to the control group, whereas IFN-α was downregulated in COVID-19 patients. ELISA results also had increased levels of IL-12, TNF-α, and IFN-γ (p = 0.0014, 0.0012, and p < 0.0001, respectively), and decreased level of IFN-α (p = 0.0021) in patients with COVID-19 compared to the control group. CONCLUSION: These findings suggest a probable association among vitamin D concentrations, immune system function, and risk of COVID-19 infection. As a result, it is recommended that vitamin D be considered as a candidate for handling and controlling COVID-19 because of its ability to target the cytokine storm and its antiviral effects.

The effects of acute exercise and inflammation on immune function in early-stage prostate cancer.

Background: The immune system plays a vital role in cancer development and progression. Strategies mobilizing cytotoxic cells of the immune system to combat immunosuppression in cancer may help to improve the treatment response of patients. To this end, we aimed to characterize the anti-cancer effect of acute exercise, including the involvement of inflammatory signals. Patients and methods: Twenty patients with early-stage prostate cancer (PCa) scheduled to undergo prostatectomy performed one bout of acute exercise consisting of a watt-max test and four high-intensity intervals. Natural Killer (NK), NKT-like and T cell phenotype, NK cell cytotoxic activity (NKCA), and NKCA per-cell against cell lines of leukemia (K562) and prostate cancer origin (LNCaP and PC-3) were assessed. Inflammatory markers (TNF-α, IL-6, and CRP) were measured in plasma. Results: Exercise increased NK, NKT-like, and CD8 T cell concentration in the circulation. Furthermore, exercise shifted immune cells towards a mature and cytotoxic phenotype e.g., NK cells exhibited higher CD57 as well as lower NKG2A expression. NKT-like and CD8 cells exhibited elevated CD57, TIGIT and Granzyme-B expression. Exercise significantly improved NKCA against K562 (+16% [5%; 27%]; p = 0.002) and LNCaP (+24% [14%; 34%]; p < 0.001) but not PC-3. NKCA per NK cell decreased during exercise and increased 1-h post exercise compared to baseline in K562, LNCap, and PC-3 cell lines. Baseline IL-6 correlated with lymphocyte, monocyte and T cell concentration pre-exercise and inversely correlated with the fold-change of mobilized lymphocytes and CD8 T cells during exercise. Furthermore, baseline IL-6 and TNF-α inversely correlated with NKCA against PC-3 cells during exercise. Conclusions: Acute exercise mobilized cytotoxic immune cells and improved NKCA in patients with PCa whereas low-grade inflammation might impair the response. Whether the observed improvements impact long-term outcomes warrant further investigation. Clinical trial number: NCT03675529.

Surfactant protein D inhibits growth, alters cell surface polysaccharide exposure and immune activation potential of Aspergillus fumigatus.

Humoral immunity plays a defensive role against invading microbes. However, it has been largely overlooked with respect to Aspergillus fumigatus, an airborne fungal pathogen. Previously, we have demonstrated that surfactant protein D (SP-D), a major humoral component in human lung-alveoli, recognizes A. fumigatus conidial surface exposed melanin pigment. Through binding to melanin, SP-D opsonizes conidia, facilitates conidial phagocytosis, and induces the expression of protective pro-inflammatory cytokines in the phagocytic cells. In addition to melanin, SP-D also interacts with galactomannan (GM) and galactosaminogalactan (GAG), the cell wall polysaccharides exposed on germinating conidial surfaces. Therefore, we aimed at unravelling the biological significance of SP-D during the germination process. Here, we demonstrate that SP-D exerts direct fungistatic activity by restricting A. fumigatus hyphal growth. Conidial germination in the presence of SP-D significantly increased the exposure of cell wall polysaccharides chitin, α-1,3-glucan and GAG, and decreased ß-1,3-glucan exposure on hyphae, but that of GM was unaltered. Hyphae grown in presence of SP-D showed positive immunolabelling for SP-D. Additionally, SP-D treated hyphae induced lower levels of pro-inflammatory cytokine, but increased IL-10 (anti-inflammatory cytokine) and IL-8 (a chemokine) secretion by human peripheral blood mononuclear cells (PBMCs), compared to control hyphae. Moreover, germ tube surface modifications due to SP-D treatment resulted in an increased hyphal susceptibility to voriconazole, an antifungal drug. It appears that SP-D exerts its anti-A. fumigatus functions via a range of mechanisms including hyphal growth-restriction, hyphal surface modification, masking of hyphal surface polysaccharides and thus altering hyphal immunostimulatory properties.

Fucose-containing bacterial exopolysaccharides: Sources, biological activities, and food applications.

Bacterial exopolysaccharides are high molecular weight polysaccharides that are secreted by a wide range of bacteria, with diverse structures and easy preparation. Fucose, fucose-containing oligosaccharides (FCOs), and fucose-containing polysaccharides (FCPs) have important applications in the food and medicine fields, including applications in products for removing Helicobacter pylori and infant formula powder. Fucose-containing bacterial exopolysaccharide (FcEPS) is a prospective source of fucose, FCOs, and FCPs. This review systematically summarizes the common sources and applications of FCPs and FCOs and the bacterial strains capable of producing FcEPS reported in recent years. The repeated-unit structures, synthesis pathways, and factors affecting the production of FcEPS are reviewed, as well as the degradation methods of FcEPS for preparing FCOs. Finally, the bioactivities of FcEPS, including anti-oxidant, prebiotic, anti-cancer, anti-inflammatory, anti-viral, and anti-microbial activities, are discussed and may serve as a reference strategy for further applications of FcEPS in the functional food and medicine industries.

Immunopeptidome of hepatocytes isolated from patients with HBV infection and hepatocellular carcinoma.

Background & Aims: Antigen-specific immunotherapy is a promising strategy to treat HBV infection and hepatocellular carcinoma (HCC). To facilitate killing of malignant and/or infected hepatocytes, it is vital to know which T cell targets are presented by human leucocyte antigen (HLA)-I complexes on patient-derived hepatocytes. Here, we aimed to reveal the hepatocyte-specific HLA-I peptidome with emphasis on peptides derived from HBV proteins and tumour-associated antigens (TAA) to guide development of antigen-specific immunotherapy. Methods: Primary human hepatocytes were isolated with high purity from (HBV-infected) non-tumour and HCC tissues using a newly designed perfusion-free procedure. Hepatocyte-derived HLA-bound peptides were identified by unbiased mass spectrometry (MS), after which source proteins were subjected to Gene Ontology and pathway analysis. HBV antigen and TAA-derived HLA peptides were searched for using targeted MS, and a selection of peptides was tested for immunogenicity. Results: Using unbiased data-dependent acquisition (DDA), we acquired a high-quality HLA-I peptidome of 2 × 105 peptides that contained 8 HBV-derived peptides and 14 peptides from 8 known HCC-associated TAA that were exclusive to tumours. Of these, 3 HBV- and 12 TAA-derived HLA peptides were detected by targeted MS in the sample they were originally identified in by DDA. Moreover, 2 HBV- and 2 TAA-derived HLA peptides were detected in samples in which no identification was made using unbiased MS. Finally, immunogenicity was demonstrated for 5 HBV-derived and 3 TAA-derived peptides. Conclusions: We present a first HLA-I immunopeptidome of isolated primary human hepatocytes, devoid of immune cells. Identified HBV-derived and TAA-derived peptides directly aid development of antigen-specific immunotherapy for chronic HBV infection and HCC. The described methodology can also be applied to personalise immunotherapeutic treatment of liver diseases in general. Lay summary: Immunotherapy that aims to induce immune responses against a virus or tumour is a promising novel treatment option to treat chronic HBV infection and liver cancer. For the design of successful therapy, it is essential to know which fragments (i.e. peptides) of virus-derived and tumour-specific proteins are presented to the T cells of the immune system by diseased liver cells and are thus good targets for immunotherapy. Here, we have isolated liver cells from patients who have chronic HBV infection and/or liver cancer, analysed what peptides are presented by these cells, and assessed which peptides are able to drive immune responses.

Differentiating between liver diseases by applying multiclass machine learning approaches to transcriptomics of liver tissue or blood-based samples.

Background & Aims: Liver disease carries significant healthcare burden and frequently requires a combination of blood tests, imaging, and invasive liver biopsy to diagnose. Distinguishing between inflammatory liver diseases, which may have similar clinical presentations, is particularly challenging. In this study, we implemented a machine learning pipeline for the identification of diagnostic gene expression biomarkers across several alcohol-associated and non-alcohol-associated liver diseases, using either liver tissue or blood-based samples. Methods: We collected peripheral blood mononuclear cells (PBMCs) and liver tissue samples from participants with alcohol-associated hepatitis (AH), alcohol-associated cirrhosis (AC), non-alcohol-associated fatty liver disease, chronic HCV infection, and healthy controls. We performed RNA sequencing (RNA-seq) on 137 PBMC samples and 67 liver tissue samples. Using gene expression data, we implemented a machine learning feature selection and classification pipeline to identify diagnostic biomarkers which distinguish between the liver disease groups. The liver tissue results were validated using a public independent RNA-seq dataset. The biomarkers were computationally validated for biological relevance using pathway analysis tools. Results: Utilizing liver tissue RNA-seq data, we distinguished between AH, AC, and healthy conditions with overall accuracies of 90% in our dataset, and 82% in the independent dataset, with 33 genes. Distinguishing 4 liver conditions and healthy controls yielded 91% overall accuracy in our liver tissue dataset with 39 genes, and 75% overall accuracy in our PBMC dataset with 75 genes. Conclusions: Our machine learning pipeline was effective at identifying a small set of diagnostic gene biomarkers and classifying several liver diseases using RNA-seq data from liver tissue and PBMCs. The methodologies implemented and genes identified in this study may facilitate future efforts toward a liquid biopsy diagnostic for liver diseases. Lay summary: Distinguishing between inflammatory liver diseases without multiple tests can be challenging due to their clinically similar characteristics. To lay the groundwork for the development of a non-invasive blood-based diagnostic across a range of liver diseases, we compared samples from participants with alcohol-associated hepatitis, alcohol-associated cirrhosis, chronic hepatitis C infection, and non-alcohol-associated fatty liver disease. We used a machine learning computational approach to demonstrate that gene expression data generated from either liver tissue or blood samples can be used to discover a small set of gene biomarkers for effective diagnosis of these liver diseases.

Effect of adjusted cut-offs of interferon-γ release assays on diagnosis of tuberculosis in patients with fever of unknown origin.

BACKGROUND: Tuberculosis (TB) is a leading cause of fever of unknown origin (FUO). In recent years, interferon-γ release assays (IGRAs) have been widely utilized and the cut-off values given by the manufacturers are set in countries where rates of TB are not as high. METHODS: A prospective cohort study was conducted in a Chinese general hospital to evaluate the diagnostic performance of T-SPOT.TB (T-SPOT) and QuantiFERON-TB Gold (QFT) in detecting active TB (ATB) in a high TB endemic area. Test results were compared with the culture and clinically confirmed diagnosis. Further, we explored an alternative method of interpreting IGRAs by increasing the cut-off values. RESULTS: The sensitivity and specificity of T-SPOT in detecting ATB were 85.3% (95% CI 81.6-94.0%) and 71.8% (95% CI 67.3-76.0%), respectively. The sensitivity and specificity of QFT were 72.3% (95% CI 62.8-80.1%) and 77.0% (95% CI 72.7-80.8%), respectively. Receiver operating characteristic analysis was used for evaluation of different cut-off values. When the cut-off values were adjusted as 125 spot-forming cells (SFCs)/ 2.5*105 cells for T-SPOT and 4.0 IU/ml for QFT, the specificity could be improved to > 90.0% (90.3% and 94.1%, respectively), and the sensitivity were 43.1% and 41.6%, respectively. The new adjusted cut-off values were validated in another independent validation cohort. CONCLUSION: The adjusted cut-off values of the two assays considerably improved the diagnostic value when applied to FUO patients in clinical settings.

XBP1-mediated activation of the STING signalling pathway in macrophages contributes to liver fibrosis progression.

Background & Aims: XBP1 modulates the macrophage proinflammatory response, but its function in macrophage stimulator of interferon genes (STING) activation and liver fibrosis is unknown. X-box binding protein 1 (XBP1) has been shown to promote macrophage nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) activation in steatohepatitis. Herein, we aimed to explore the underlying mechanism of XBP1 in the regulation of STING signalling and the subsequent NLRP3 activation during liver fibrosis. Methods: XBP1 expression was measured in the human fibrotic liver tissue samples. Liver fibrosis was induced in myeloid-specific Xbp1-, STING-, and Nlrp3-deficient mice by carbon tetrachloride injection, bile duct ligation, or a methionine/choline-deficient diet. Results: Although increased XBP1 expression was observed in the fibrotic liver macrophages of mice and clinical patients, myeloid-specific Xbp1 deficiency or pharmacological inhibition of XBP1 protected the liver against fibrosis. Furthermore, it inhibited macrophage NLPR3 activation in a STING/IRF3-dependent manner. Oxidative mitochondrial injury facilitated cytosolic leakage of macrophage self-mtDNA and cGAS/STING/NLRP3 signalling activation to promote liver fibrosis. Mechanistically, RNA sequencing analysis indicated a decreased mtDNA expression and an increased BCL2/adenovirus E1B interacting protein 3 (BNIP3)-mediated mitophagy activation in Xbp1-deficient macrophages. Chromatin immunoprecipitation (ChIP) assays further suggested that spliced XBP1 bound directly to the Bnip3 promoter and inhibited the transcription of Bnip3 in macrophages. Xbp1 deficiency decreased the mtDNA cytosolic release and STING/NLRP3 activation by promoting BNIP3-mediated mitophagy activation in macrophages, which was abrogated by Bnip3 knockdown. Moreover, macrophage XBP1/STING signalling contributed to the activation of hepatic stellate cells. Conclusions: Our findings demonstrate that XBP1 controls macrophage cGAS/STING/NLRP3 activation by regulating macrophage self-mtDNA cytosolic leakage via BNIP3-mediated mitophagy modulation, thus providing a novel target against liver fibrosis. Lay summary: Liver fibrosis is a typical progressive process of chronic liver disease, driven by inflammatory and immune responses, and is characterised by an excess of extracellular matrix in the liver. Currently, there is no effective therapeutic strategy for the treatment of liver fibrosis, resulting in high mortality worldwide. In this study, we found that myeloid-specific Xbp1 deficiency protected the liver against fibrosis in mice, while XBP1 inhibition ameliorated liver fibrosis in mice. This study concluded that targeting XBP1 signalling in macrophages may provide a novel strategy for protecting the liver against fibrosis.

Circulating Mir-140 and leptin improve the accuracy of the differential diagnosis between psoriatic arthritis and rheumatoid arthritis: a case-control study.

The differential diagnosis of psoriatic arthritis (PsA) and rheumatoid arthritis (RA) is difficult because of the lack of diagnostic clinical signs and reliable biomarkers. This study investigated microRNAs (miRNA) and adipokines as potential additional markers to discriminate PsA from RA. The expression profile of miRNA (miR-21, miR-140, miR-146a, miR-155, miR-181b, miR-223, miR-let-7e) and inflammatory cytokines (IL-1ß, IL-6, IL-17a, IL-23a, TNF-α) from peripheral blood mononuclear cells of PsA and RA patients compared to healthy controls (HC) were evaluated by real-time PCR, and serum adipokines (adiponectin, chemerin, leptin, resistin, visfatin) and cytokines by ELISA assay. Univariable binary logistic regression was used to find the association between PsA and potential predictors. The gene expression of miRNA and cytokines and the serum levels of adipokines were found significantly different in PsA and RA patients compared to HC, as well as in PsA versus RA. MiR-140 gene expression resulted up-regulated in PsA patients and reduced in RA in comparison to HC, and, for the first time, significantly higher in PsA compared with RA. Serum levels of IL-23a and leptin were significantly increased in PsA and RA populations than in HC, as well as in PsA versus RA. Furthermore, circulating TNF-α was up-regulated in PsA and RA in comparison to controls, while resulted higher in RA than in PsA. Univariable binary logistic regression analysis found the above-mentioned markers associated to PsA versus RA. Our results first demonstrated an increased expression of circulating miR-140 and serum leptin in PsA patients compared to RA, which were identified as potential additional biomarkers to discriminate PsA from RA. Since the differential diagnosis of PsA and RA poses challenges in clinical practice, our data may help to enhance the diagnostic performance of PsA in daily practice.

Therapeutic vaccine BRII-179 restores HBV-specific immune responses in patients with chronic HBV in a phase Ib/IIa study.

BACKGROUND & AIMS: Functional cure of chronic HBV infection (CHB) without life-long treatment requires the restoration of defective HBV-specific humoral and cellular immunity. Therapeutic vaccines based on the major structural and non-structural proteins have been tested in patients with CHB but have shown scarce immunogenicity. BRII-179, also known as VBI-2601, is a novel formulation comprised of all 3 HBV surface envelope proteins (Pre-S1, Pre-S2, and S). Safety, antiviral activity, and immunogenicity of BRII-179 admixed with co-adjuvant interferon (IFN)-α were assessed in patients with CHB. METHOD: This randomized, open-label, controlled phase Ib/IIa study included 2 dose levels, 20 µg BRII-179 (Part 1, n = 25) and 40 µg BRII-179 (Part 2, n = 24). Patients, virally suppressed under nucleos(t)ide analogue (NA) therapy were randomized 1:2:2 into 3 cohorts in Part 1 and 1:1 into 2 cohorts in Part 2 to receive 4 monthly intramuscular injections of BRII-179 admixed with/without 3 MIU IFN-α. Antibody and cellular responses to HBsAg, as well as evolution of circulating HBsAg were monitored. RESULTS: Both 20 µg and 40 µg BRII-179 with/without IFN-α were well tolerated with no severe adverse events. BRII-179 induced anti-HBs responses in >30% patients in all treatment cohorts, however, moderate anti-Pre-S1 or anti-Pre-S2 antibody responses were only observed in patients receiving BRII-179 with IFN-α. BRII-179 also restored S-, Pre-S1-, Pre-S2-specific IFN-γ-producing T-cells in the majority of treated patients. Overall, no notable reduction of HBsAg was observed after BRII-179 treatment. CONCLUSION: In patients with CHB under NA therapy, BRII-179 with/without IFN-α exhibited a good safety profile and induced HBV-specific B- and T-cell immune responses. These data support further clinical evaluation of BRII-179 in combination with other therapies. CLINICAL TRIAL NUMBER: ACTRN12619001210167. LAY SUMMARY: BRII-179 is a therapeutic vaccine designed to improve the immune response in patients with chronic hepatitis B. In this study, BRII-179 alone or with a low dose of interferon-α was safe, well tolerated, and induced enhanced HBV-specific antibody and T-cell responses in patients with chronic hepatitis B. However, BRII-179 treatment alone had minimal effect on patient's virological status. The potential of BRII-179 to achieve a functional cure in conjunction with other agents is being evaluated in the clinic.

Irisin supports integrin-mediated cell adhesion of lymphocytes.

Irisin, a myokine released from skeletal muscle, has recently been found to act as a ligand for the integrins αVß5, αVß1, and α5ß1 expressed on mesenchymal cells, thereby playing an important role in the metabolic remodeling of the bone, skeletal muscle and adipose tissues. Although the immune-modulatory effects of irisin in chronic inflammation have been documented, its interactions with lymphocytic integrins have yet to be elucidated. Here, we show that irisin supports the cell adhesion of human and mouse lymphocytes. Cell adhesion assays using a panel of inhibitory antibodies to integrins have shown that irisin-mediated lymphocyte adhesion involves multiple integrins including not only α4ß1 and α5ß1, but also leukocyte-specific αLß2 and α4ß7. Importantly, mouse lymphocytic TK-1 cells that lack the expression of ß1 integrins have exhibited αLß2- and α4ß7-mediated cell adhesion to irisin. Irisin has also been demonstrated to bind to purified recombinant integrin αLß2 and α4ß7 proteins. Thus, irisin represents a novel ligand for integrin αLß2 and α4ß7, capable of supporting lymphocyte cell adhesion independently of ß1 integrins. These results suggest that irisin may play an important role in regulating lymphocyte adhesion and migration in the inflamed vasculature.

Differential blood transcriptome modules predict response to corticosteroid therapy in alcoholic hepatitis.

BACKGROUND & AIMS: In patients with severe alcoholic hepatitis (SAH), little is known about the profile of peripheral blood mononuclear cells (PBMCs) at baseline and during corticosteroid therapy, among those who can be treated successfully with steroids (steroid-responders [R] and those who cannot (steroid-non-responders [NR]); 2 groups with different outcomes. METHODS: We performed RNA-seq analysis in PBMCs from 32 patients with definite SAH, at baseline and after 7 days of corticosteroids. The data were sorted into R and NR (n = 16, each group) using the Lille model and 346 blood transcription modules (BTMs) were identified. BTMs are predefined modules of highly co-expressed PBMC genes, which can determine specific immune cell types and cellular functions. The activity of each BTM was taken as the mean value of its member genes. RESULTS: At baseline, 345 BTMs had higher activity (i.e. were upregulated) in NR relative to R. The 100 most upregulated BTMs in NR, included several modules related to lymphoid lineage (T, B, and natural killer [NK] cells), modules for cell division and mitochondrial respiratory electron transport chain (ETC, relating to energy production), but only a few modules of myeloid cells. Correlation studies of BTM activities found features of significantly greater activation/proliferation and differentiation for T and B cells in NR relative to R. After 7 days of corticosteroids, NR had no significant changes in BTM activities relative to baseline, whereas R had downregulation of BTMs related to innate and adaptive immunity. CONCLUSIONS: At baseline and during corticosteroid therapy, increased activity in the PBMCs of gene modules related to activation/proliferation and differentiation of T and B cells, NK cells, and mitochondrial ETC, is a hallmark of SAH patients who are steroid-non-responders. LAY SUMMARY: Patients with severe alcoholic hepatitis receive steroid therapy as the main line of treatment; however, this treatment is ineffective in some patients. This only becomes apparent after 7 days of steroid therapy. We have developed an approach where it can be estimated if a patient is going to respond or not to steroid therapy using the gene expression information of blood cells. This method will allow clinicians to assess the response of patients to steroids earlier, and will help them in adopting alternate strategies if the treatment is found to be ineffective in a particular patient.

Human hepatitis D virus-specific T cell epitopes.

HDV is a small, defective RNA virus that requires the HBsAg of HBV for its assembly, release, and transmission. Chronic HBV/HDV infection often has a severe clinical outcome and is difficult to treat. The important role of a robust virus-specific T cell response for natural viral control has been established for many other chronic viral infections, but the exact role of the T cell response in the control and progression of chronic HDV infection is far less clear. Several recent studies have characterised HDV-specific CD4+ and CD8+ T cell responses on a peptide level. This review comprehensively summarises all HDV-specific T cell epitopes described to date and describes our current knowledge of the role of T cells in HDV infection. While we now have better tools to study the adaptive anti-HDV-specific T cell response, further efforts are needed to define the HLA restriction of additional HDV-specific T cell epitopes, establish additional HDV-specific MHC tetramers, understand the degree of cross HDV genotype reactivity of individual epitopes and understand the correlation of the HBV- and HDV-specific T cell response, as well as the breadth and specificity of the intrahepatic HDV-specific T cell response.

De novo expression and antibacterial potential of four lactoferricin peptides in cell-free protein synthesis system.

For the first time, we produced four lactoferricin (LFcin) peptides by a cell-free (in vitro) method. These short antimicrobial peptides were expressed in an E. coli cell-free protein synthesis (CFPS) system and the bioactivity of the produced peptides was demonstrated. Additionally, we designed a novel synthetic consensus peptide (ConLFcin). The genes of bovine Lfcin (bLFcin), human Lfcin (hLFcin), camel Lfcin (cLFcin), and ConLFcin were cloned into pET101/D-TOPO vector then peptides were synthesized in vitro by E. coli CFPS system. The antibacterial activity of these synthesized peptides was evaluated against Escherichia coli, Salmonella typhi, Pseudomonas aeruginosa, Staphylococcus aureus, and methicillin-resistant Staphylococcus aureus (MRSA). The four cell-free synthesized peptides showed significant antibacterial potency at minimum inhibitory concentration (MIC) values between 1.25 and 10 µg/mL. cLFcin and ConLFcin showed higher antibacterial effects than bLFcin and hLFcin. Thus, cell-free expression system is an ideal system for rapid expression of functionally active short bioactive peptides.

Profiling circulating microRNAs in patients with cirrhosis and acute-on-chronic liver failure.

BACKGROUND & AIMS: MicroRNAs (miRNAs) circulate in several body fluids and can be useful biomarkers. The aim of this study was to identify blood-circulating miRNAs associated with cirrhosis progression and acute-on-chronic liver failure (ACLF). METHODS: Using high-throughput screening of 754 miRNAs, serum samples from 45 patients with compensated cirrhosis, decompensated cirrhosis, or ACLF were compared with those from healthy individuals (n = 15). miRNA levels were correlated with clinical parameters, organ failure, and disease progression and outcome. Dysregulated miRNAs were evaluated in portal and hepatic vein samples (n = 33), liver tissues (n = 17), and peripheral blood mononuclear cells (PBMCs) (n = 16). RESULTS: miRNA screening analysis revealed that circulating miRNAs are dysregulated in cirrhosis progression, with 51 miRNAs being differentially expressed among all groups of patients. Unsupervised clustering and principal component analysis indicated that the main differences in miRNA expression occurred at decompensation, showing similar levels in patients with decompensated cirrhosis and those with ACLF. Of 43 selected miRNAs examined for differences among groups, 10 were differentially expressed according to disease progression. Moreover, 20 circulating miRNAs were correlated with model for end-stage liver disease and Child-Pugh scores. Notably, 11 dysregulated miRNAs were associated with kidney or liver failure, encephalopathy, bacterial infection, and poor outcomes. The most severely dysregulated miRNAs (i.e. miR-146a-5p, miR-26a-5p, and miR-191-5p) were further evaluated in portal and hepatic vein blood and liver tissue, but showed no differences. However, PBMCs from patients with cirrhosis showed significant downregulation of miR-26 and miR-146a, suggesting a extrahepatic origin of some circulating miRNAs. CONCLUSIONS: This study is a repository of circulating miRNA data following cirrhosis progression and ACLF. Circulating miRNAs were profoundly dysregulated during the progression of chronic liver disease, were associated with failure of several organs and could have prognostic utility. LAY SUMMARY: Circulating miRNAs are small molecules in the blood that can be used to identify or predict a clinical condition. Our study aimed to identify miRNAs for use as biomarkers in patients with cirrhosis or acute-on-chronic liver failure. Several miRNAs were found to be dysregulated during the progression of disease, and some were also related to organ failure and disease-related outcomes.