Unraveling the interplay of ferroptosis and immune dysregulation in diabetic kidney disease: a comprehensive molecular analysis.

BACKGROUND: Diabetic kidney disease (DKD) is a primary microvascular complication of diabetes with limited therapeutic effects. Delving into the pathogenic mechanisms of DKD and identifying new therapeutic targets is crucial. Emerging studies reveal the implication of ferroptosis and immune dysregulation in the pathogenesis of DKD, however, the precise relationship between them remains not fully elucidated. Investigating their interplay is pivotal to unraveling the pathogenesis of diabetic kidney disease, offering insights crucial for targeted interventions and improved patient outcomes. METHODS: Integrated analysis, Consensus clustering, Machine learning including Generalized Linear Models (GLM), RandomForest (RF), Support Vector Machine (SVM) and Extreme Gradient Boosting (xGB), Artificial neural network (ANN) methods of DKD glomerular mRNA sequencing were performed to screen DKD-related ferroptosis genes.CIBERSORT, ESTIMATE and ssGSEA algorithm were used to assess the infiltration of immune cells between DKD and control groups and in two distinct ferroptosis phenotypes. The ferroptosis hub genes were verified in patients with DKD and in the db/db spontaneous type 2 diabetes mouse model via immunohistochemical and Western blotting analyses in mouse podocyte MPC5 and mesangial SV40-MES-13 cells under high-glucose (HG) conditions. RESULTS: We obtained 16 differentially expressed ferroptosis related genes and patients with DKD were clustered into two subgroups by consensus clustering. Five ferroptosis genes (DUSP1,ZFP36,PDK4,CD44 and RGS4) were identified to construct a diagnostic model with a good diagnosis performance in external validation. Analysis of immune infiltration revealed immune heterogeneity between DKD patients and controls.Moreover, a notable differentiation in immune landscape, comprised of Immune cells, ESTIMATE Score, Immune Score and Stromal Score was observed between two FRG clusters. GSVA analysis indicated that autophagy, apoptosis and complement activation can participate in the regulation of ferroptosis phenotypes. Experiment results showed that ZFP36 was significantly overexpressed in both tissue and cells while CD44 was on the contrary.Meanwhile,spearman analysis showed both ZFP36 and CD44 has a strong correlation with different immune cells,especially macrophage. CONCLUSION: The regulation of the immune landscape in DKD is significantly influenced by the focal point on ferroptosis. Newly identified ferroptosis markers, CD44 and ZFP36, are poised to play essential roles through their interactions with macrophages, adding substantial value to this regulatory landscape.

Unraveling DDIT4 in the VDR-mTOR pathway: a novel target for drug discovery in diabetic kidney disease.

Introduction: Diabetic kidney disease (DKD) necessitates innovative therapeutic strategies. This study delves into the role of DNA damage-inducing transcription factor 4 (DDIT4) within the VDR-mTOR pathway, aiming to identify a novel target for DKD drug discovery. Methods: Transcriptome data from the Gene Expression Omnibus Database were analyzed to assess the expression of mTOR and VDR expression in human renal tissues. Clinical samples from DKD patients and minimal change disease (MCD) controls were examined, and a DKD animal model using 20-week-old db/db mice was established. DDIT4 plasmid transfection was employed to modulate the VDR-mTOR pathway, with its components evaluated using immunohistochemistry, real-time quantitative PCR (qRT-PCR), Western blotting, and enzyme-linked immunosorbent assay (ELISA). Results: Changes in the expression of the VDR-mTOR pathway were observed in both DKD patients and the animal model. Overexpression of DDIT4 increased VDR expression and decreased levels of mTOR, p70s6k, and 4E-BP1. Furthermore, DDIT4 treatment regulated autophagy by upregulating LC3I expression and downregulating LC3II expression. Notably, DDIT4 alleviated oxidative stress by reducing the levels of lipid peroxidation product MDA, while simultaneously increasing the levels of superoxide dismutase (SOD) and glutathione (GSH), underscoring the role of DDIT4 in the pathological process of DKD and its potential as a therapeutic target. Conclusion: Unraveling DDIT4's involvement in the VDR-mTOR pathway provides insights for innovative DKD drug discovery, emphasizing its potential as a therapeutic target for future interventions.

BCL11B and the NuRD complex cooperatively guard T-cell fate and inhibit OPA1-mediated mitochondrial fusion in T cells.

The nucleosome remodeling and histone deacetylase (NuRD) complex physically associates with BCL11B to regulate murine T-cell development. However, the function of NuRD complex in mature T cells remains unclear. Here, we characterize the fate and metabolism of human T cells in which key subunits of the NuRD complex or BCL11B are ablated. BCL11B and the NuRD complex bind to each other and repress natural killer (NK)-cell fate in T cells. In addition, T cells upregulate the NK cell-associated receptors and transcription factors, lyse NK-cell targets, and are reprogrammed into NK-like cells (ITNKs) upon deletion of MTA2, MBD2, CHD4, or BCL11B. ITNKs increase OPA1 expression and exhibit characteristically elongated mitochondria with augmented oxidative phosphorylation (OXPHOS) activity. OPA1-mediated elevated OXPHOS enhances cellular acetyl-CoA levels, thereby promoting the reprogramming efficiency and antitumor effects of ITNKs via regulating H3K27 acetylation at specific targets. In conclusion, our findings demonstrate that the NuRD complex and BCL11B cooperatively maintain T-cell fate directly by repressing NK cell-associated transcription and indirectly through a metabolic-epigenetic axis, providing strategies to improve the reprogramming efficiency and antitumor effects of ITNKs.

DAP10 integration in CAR-T cells enhances the killing of heterogeneous tumors by harnessing endogenous NKG2D.

Although chimeric antigen receptor T (CAR-T) cells have achieved remarkable successes in hematological malignancies, the efficacies of CAR-T cells against solid tumors remains unsatisfactory. Heterogeneous antigen expression is one of the obstacles on its effective elimination of solid cancer cells. DNAX-activating protein 10 (DAP10) interacts with natural killer group 2D (NKG2D), acting as an adaptor that targets various malignant cells for surveillance. Here, we designed a DAP10 chimeric receptor that utilized native NKG2D on T cells to target NKG2D ligand-expressing cancer cells. We then tandemly incorporated it with anti-glypican 3 (GPC3) single-chain variable fragment (scFv) to construct a dual-antigen-targeting system. T cells expressing DAP10 chimeric receptor (DAP10-T cells) displayed with an enhancement on both cytotoxicity and cytokine secretion against solid cancer cell lines, and its tandem connection with anti-GPC3 scFv (CAR GPC3-DAP10-T cells) exhibited a dual-antigen-targeting capacity on eliminating heterogeneous cancer cells in vitro and suppressing the growth of heterogeneous cancer in vivo. Thus, this novel dual-targeting system enabled a high efficacy on killing cancer cells and extended the recognition profile of CAR-T cells toward tumors, which providing a potential strategy on treatment of solid cancer clinically.

The Chemokine Receptor CCR8 Is a Target of Chimeric Antigen T Cells for Treating T Cell Malignancies.

Chimeric antigen receptor (CAR) T cells have been successfully used in the therapy of B cell leukemia and lymphoma, but still have many challenges in their use for treating T cell malignancies, such as the lack of unique tumor antigens, their limitation of T cell expansion, and the need for third party donors or genome editing. Therefore, we need to find novel targets for CAR T cell therapy to overcome these challenges. Here, we found that both adult T-cell leukemia/lymphoma (ATLL) patients and ATLL cells had increased CCR8 expression but did not express CD7. Moreover, targeting CCR8 in T cells did not impair T cell expansion in vitro. Importantly, anti-CCR8 CAR T cells exhibited antitumor effects on ATLL- and other CCR8-expressing T-ALL cells in vitro and in vivo, and prolonged the survival of ATLL and Jurkat tumor-bearing mouse models. In conclusion, these collective results show that anti-CCR8 CAR T cells possess strong antitumor activity and represent a promising therapeutic approach for ATLL and CCR8+ tumors.

Assessment of chimeric antigen receptor T cytotoxicity by droplet microfluidics in vitro.

Chimeric antigen receptor T (CAR-T) cells are cytotoxic T cells engineered to specifically kill cancer cells expressing specific target receptor(s). Prior CAR-T efficacy tests include CAR expression analysis by qPCR or ELISA, in vitro measurement of interferon-γ (IFNγ) or interleukin-2 (IL-2), and xenograft models. However, the in vitro measurements did not reflect CAR-T cytotoxicity, whereas xenograft models are low throughput and costly. Here, we presented a robust in vitro droplet microfluidic assay for CAR-T cytotoxicity assessment. This method not only enabled assessment of CAR-T cytotoxic activity under different fluid viscosity conditions, but also facilitated measurement of CAR-T expansion and dissection of mechanism of action via phenotype analysis in vitro. Furthermore, our data suggested that label-free cytotoxicity analysis is feasible by acquiring data before and after treatment. Hence, this study presented a novel in vitro method for assessment of cellular cytotoxicity that could potentially be applied to any cytotoxicity experiment with varying solvent composition.

Human induced-T-to-natural killer cells have potent anti-tumour activities.

BACKGROUND: Adoptive cell therapy (ACT) is a particularly promising area of cancer immunotherapy, engineered T and NK cells that express chimeric antigen receptors (CAR) are being explored for treating hematopoietic malignancies but exhibit limited clinical benefits for solid tumour patients, successful cellular immunotherapy of solid tumors demands new strategies. METHODS: Inactivation of BCL11B were performed by CRISPR/Cas9 in human T cells. Immunophenotypic and transcriptional profiles of sgBCL11B T cells were characterized by cytometer and transcriptomics, respectively. sgBCL11B T cells are further engineered with chimeric antigen receptor. Anti-tumor activity of ITNK or CAR-ITNK cells were evaluated in preclinical and clinical studies. RESULTS: We report that inactivation of BCL11B in human CD8+ and CD4+ T cells induced their reprogramming into induced T-to-natural killer cells (ITNKs). ITNKs contained a diverse TCR repertoire; downregulated T cell-associated genes such as TCF7 and LEF1; and expressed high levels of NK cell lineage-associated genes. ITNKs and chimeric antigen receptor (CAR)-transduced ITNKs selectively lysed a variety of cancer cells in culture and suppressed the growth of solid tumors in xenograft models. In a preliminary clinical study, autologous administration of ITNKs in patients with advanced solid tumors was well tolerated, and tumor stabilization was seen in six out nine patients, with one partial remission. CONCLUSIONS: The novel ITNKs thus may be a promising novel cell source for cancer immunotherapy. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03882840 . Registered 20 March 2019-Retrospectively registered.

PRRSV Non-Structural Proteins Orchestrate Porcine E3 Ubiquitin Ligase RNF122 to Promote PRRSV Proliferation.

Ubiquitination plays a major role in immune regulation after viral infection. An alternatively spliced porcine E3 ubiquitin ligase RNF122 promoted PRRSV infection and upregulated in PRRSV-infected PAM cells was identified. We characterized the core promoter of RNF122, located between -550 to -470 bp upstream of the transcription start site (TSS), which displayed significant differential transcriptional activities in regulating the transcription and expression of RNF122. The transcription factor HLTF was inhibited by nsp1α and nsp7 of PRRSV, and the transcription factor E2F complex regulated by nsp9. Together, they modulated the transcription and expression of RNF122. RNF122 could mediate K63-linked ubiquitination to raise stability of PRRSV nsp4 protein and thus promote virus replication. Moreover, RNF122 also performed K27-linked and K48-linked ubiquitination of MDA5 to degrade MDA5 and inhibit IFN production, ultimately promoted virus proliferation. In this study, we illustrate a new immune escape mechanism of PRRSV that enhances self-stability and function of viral nsp4, thus, regulating RNF122 expression to antagonize IFNα/ß production. The present study broadens our knowledge of PRRSV-coding protein modulating transcription, expression and modification of host protein to counteract innate immune signaling, and may provide novel insights for the development of antiviral drugs.

IL-7 and CCL19-secreting CAR-T cell therapy for tumors with positive glypican-3 or mesothelin.

Although chimeric antigen receptor (CAR)-engineered T cells have shown great success in the treatment of B cell malignancies, this strategy has limited efficacy in patients with solid tumors. In mouse CAR-T cells, IL-7 and CCL19 expression have been demonstrated to improve T cell infiltration and CAR-T cell survival in mouse tumors. Therefore, in the current study, we engineered human CAR-T cells to secrete human IL-7 and CCL19 (7 × 19) and found that these 7 × 19 CAR-T cells showed enhanced capacities of expansion and migration in vitro. Furthermore, 7 × 19 CAR-T cells showed superior tumor suppression ability compared to conventional CAR-T cells in xenografts of hepatocellular carcinoma (HCC) cell lines, primary HCC tissue samples and pancreatic carcinoma (PC) cell lines. We then initiated a phase 1 clinical trial in advanced HCC/PC/ovarian carcinoma (OC) patients with glypican-3 (GPC3) or mesothelin (MSLN) expression. In a patient with advanced HCC, anti-GPC3-7 × 19 CAR-T treatment resulted in complete tumor disappearance 30 days post intratumor injection. In a patient with advanced PC, anti-MSLN-7 × 19 CAR-T treatment resulted in almost complete tumor disappearance 240 days post-intravenous infusion. Our results demonstrated that the incorporation of 7 × 19 into CAR-T cells significantly enhanced the antitumor activity against human solid tumor. Trial registration: NCT03198546. Registered 26 June 2017, https://clinicaltrials.gov/ct2/show/NCT03198546?term=NCT03198546&draw=2&rank=1.

Development of IMBs-qPCR detection method for Yersinia enterocolitica based on the foxA gene.

Yersinia enterocolitica is an important zoonotic pathogen, which seriously endangers food-safety risk. In this study, the recombinant outer membrane protein OmpF and its antibody were prepared and coupled with immunomagnetic beads (IMBs) to capture Y. enterocolitica in food samples, combining the quantitative PCR detection with primers of virulence factor gene foxA for Yersinia enterocolitica contamination. The results showed that the capture efficiency of approximately 80% using anti-OmpF antibody-immunomagnetic beads and linearly dependent capture under 101-105 CFU/mL Y. enterocolitica compared with less than 10% capture of other bacteria. The detection limit of 64 CFU/mL was obtained based on foxA gene PCR detection combined with capture of the anti-OmpF antibody-immunomagnetic beads to detect Yersinia enterocolitica in artificially contaminated milk and pork samples. Compared to the culture method, the developed IMBs-qPCR method has higher consistency, was less time consuming, which taken together provides an effective alternative method for rapid detection of Y. enterocolitica in food.

The superposition anti-viral activity of porcine tri-subtype interferon expressed by Saccharomyces cerevisiae.

Interferon (IFN)-mediated antiviral responses are central to host defense against viral infection. Porcine viral infection has emerged as a serious hazard for the pig industry. The construction of an engineered Saccharomyces cerevisiae strain that efficiently produces porcine IFN has demonstrated several advantages. It can be easily fed to pigs, which helps in reducing antibiotic residues in pork and improve meat quality. In this study, the stable expression of several porcine IFN molecules (pIFN-α1, pIFN-ß, pIFN-λ1, pIFN-λ1-ß, pIFN-λ1-ß-α1) were determined using an engineered S. cerevisiae system. With the YeastFab assembly method, the complete transcriptional units containing promoter (GPD), secretory peptide (α-mating factor), target gene (IFN) and terminator (ADH1) were successfully constructed using the characteristics of type II restriction endonuclease, and then integrated into the chromosomes Ⅳ and XVI of ST1814 yeast host strain, respectively. The expression kinetics of recombinant pIFNs were further analyzed. Synergism in the expression level of IFN receptor, antiviral protein, and viral loading was observed in viral-cell infection model treated with different porcine IFN subtypes. The porcine reproductive and respiratory syndrome viral load and antibody titer in serum decreased significantly after oral administration of IFN expression yeast fermentation broth. These findings indicate the potential efficacy of multi-valent pIFNs expressing S. cerevisiae as a potent feed material to prevent viral infections of pigs.

RBM39 Alters Phosphorylation of c-Jun and Binds to Viral RNA to Promote PRRSV Proliferation.

As transcriptional co-activator of AP-1/Jun, estrogen receptors and NF-κB, nuclear protein RBM39 also involves precursor mRNA (pre-mRNA) splicing. Porcine reproductive and respiratory syndrome virus (PRRSV) causes sow reproductive disorders and piglet respiratory diseases, which resulted in serious economic losses worldwide. In this study, the up-regulated expression of RBM39 and down-regulated of inflammatory cytokines (IFN-ß, TNFα, NF-κB, IL-1ß, IL-6) were determined in PRRSV-infected 3D4/21 cells, and accompanied with the PRRSV proliferation. The roles of RBM39 altering phosphorylation of c-Jun to inhibit the AP-1 pathway to promote PRRSV proliferation were further verified. In addition, the nucleocytoplasmic translocation of RBM39 and c-Jun from the nucleus to cytoplasm was enhanced in PRRSV-infected cells. The three RRM domain of RBM39 are crucial to support the proliferation of PRRSV. Several PRRSV RNA (nsp4, nsp5, nsp7, nsp10-12, M and N) binding with RBM39 were determined, which may also contribute to the PRRSV proliferation. Our results revealed a complex mechanism of RBM39 by altering c-Jun phosphorylation and nucleocytoplasmic translocation, and regulating binding of RBM39 with viral RNA to prompt PRRSV proliferation. The results provide new viewpoints to understand the immune escape mechanism of PRRSV infection.

Role of H+, HF, SO42- and kaolin in fixing Hg of coal fire sponge.

Coal fire sponges (CFS) are common in coal-fire areas. Due to the enrichment of Hg in CFS, large amounts of Hg are released by CFS into the atmosphere via natural weathering or solar radiation. Therefore, CFS should be of concern in Hg pollution management and control globally. In addition, CFS changes the Hg cycle path by capturing Hg from coal fires that would have entered the atmosphere. In this study, the concentration, distribution, species, and enrichment mechanism of CFS Hg were investigated. The results showed that the Hg concentration in CFS ranged from 1008 to 35,310 ng/g, with an average of 8932 ng/g (CFS number, n = 153). The Hg concentration of CFS in different types of land was found to be significantly inhomogeneous. To determine the status of subterranean spontaneous combustion, the Hg concentration was added, which can improve the effect of coal-fire monitoring. Compared to the background area topsoil, CFS was enriched in Hg, acid, SO42-, and total fluoride. The Hg species in CFS was primarily HgSO4, followed by HgO. However, the primary Hg species in the surrounding topsoil were HgCl2 and HgO. By the simulation experiment, it was determined that hydrofluoric acid (HF) was beneficial to activate the stable species in the coal-fire areas. HgCl2, HgO, or Hg0 were ionized by acid liquor or HF, which can promote Hg migration and increase the adsorbed ratio; in the presence of SO42-, the primary Hg species was HgSO4. Ultimately, Hg was absorbed by clay minerals and organic matter. The high-efficiency activation of steady Hg species by the coal-fire HF should be studied further.

Controlling Factors and Prediction of Lead Uptake and Accumulation in Various Soil-Pepper Systems.

Lead (Pb) is a typical toxic heavy metal element in soils and plants, which has a potential threat to human health through the food chain. Uptake of Pb in the soil-vegetable system has attracted broad attention, whereas reports on the main controlling factors of Pb uptake and accumulation in different soil-vegetable systems are limited. The effect of soil properties on Pb uptake and accumulation in pepper (Capsicum annuum L.) was studied by a pot experiment with 16 typical soils in China. The results showed that the Pb bioavailability was lower in alkaline soils, and that soil cation exchange capacity (CEC), CaCO3 , and total phosphorus contents might influence the uptake and transfer of Pb by peppers. Soil pH and CEC were the most significant factors affecting Pb accumulation in pepper fruits. Soil pH was negatively correlated with Pb uptake and accumulation due to its influence on Pb mobility and bioavailability. The accumulation of Pb decreased as soil CEC increased, which might inhibit the absorption and transfer of Pb in peppers. The multiple linear regression function based on soil Pb content, pH, and CEC could provide enough information for a good prediction of the accumulation of Pb in soil-pepper systems (R2 = 0.733). The results are in favor of developing a Pb threshold for vegetables in agricultural soils in China, thus improving the food safety of crops. Environ Toxicol Chem 2021;40:1443-1451. © 2021 SETAC.

Porcine Immunoglobulin Fc Fused P30/P54 Protein of African Swine Fever Virus Displaying on Surface of S. cerevisiae Elicit Strong Antibody Production in Swine.

African swine fever virus (ASFV) infects domestic pigs and European wild boars with strong, hemorrhagic and high mortality. The primary cellular targets of ASFV is the porcine macrophages. Up to now, no commercial vaccine or effective treatment available to control the disease. In this study, three recombinant Saccharomyces cerevisiae (S. cerevisiae) strains expressing fused ASFV proteins-porcine Ig heavy chains were constructed and the immunogenicity of the S. cerevisiae-vectored cocktail ASFV feeding vaccine was further evaluated. To be specific, the P30-Fcγ and P54-Fcα fusion proteins displaying on surface of S. cerevisiae cells were produced by fusing the Fc fragment of porcine immunoglobulin IgG1 or IgA1 with p30 or p54 gene of ASFV respectively. The recombinant P30-Fcγ and P54-Fcα fusion proteins expressed by S. cerevisiae were verified by Western blotting, flow cytometry and immunofluorescence assay. Porcine immunoglobulin Fc fragment fused P30/P54 proteins elicited P30/P54-specific antibody production and induced higher mucosal immunity in swine. The absorption and phagocytosis of recombinant S. cerevisiae strains in IPEC-J2 cells or porcine alveolar macrophage (PAM) cells were significantly enhanced, too. Here, we introduce a kind of cheap and safe oral S. cerevisiae-vectored vaccine, which could activate the specific mucosal immunity for controlling ASFV infection.

Visual Detection of Clostridium perfringens Alpha Toxin by Combining Nanometer Microspheres with Smart Phones.

Clostridium perfringens α toxin (CPA) is an important virulence factor that causes livestock hemorrhagic enteritis and food poisoning by contaminated meat products. In this study, the nano-silica microspheres combined with smartphone image processing technology was developed to realize real-time CPA detection. First, the N-terminal and C-terminal domain of the CPA toxin (CPAC3 and CPAN) and their anti-sera were prepared. The silica microspheres coupled with the antibody of CPAC3 was prepared to capture the toxin that existed in the detection sample and the fluorescent-labeled antibody of CPAN was incubated. Moreover, the fluorescent pictures of gray value were performed in a cell phone app, corresponding to toxin concentration. The new assay takes 90 min to perform and can detect CPA as little as 32.8 ng/mL. Our results showed a sensitive, stable, and convenient CPA detection system, which provides a novel detection method of native CPA in foods.

The antiviral protein viperin interacts with the viral N protein to inhibit proliferation of porcine epidemic diarrhea virus.

In the early stage of virus infection, the pattern recognition receptor (PRR) signaling pathway of the host cell is activated to induce interferon production, activating interferon-stimulated genes (ISGs) that encode antiviral proteins that exert antiviral effects. Viperin is one of the innate antiviral proteins that exert broad-spectrum antiviral effects by various mechanisms. Porcine epidemic diarrhea virus (PEDV) is a coronavirus that causes huge losses to the pig industry. Research on early antiviral responses in the gastrointestinal tract is essential for developing strategies to prevent the spread of PEDV. In this study, we investigated the mechanisms of viperin in PEDV-infected IPEJ-C2 cells. Increased expression of interferon and viperin and decreased replication of PEDV with a clear reduction in the viral load were observed in PEDV-infected IPEC-J2 cells. Amino acids 1-50 of porcine viperin contain an endoplasmic reticulum signal sequence that allows viperin to be anchored to the endoplasmic reticulum and are necessary for its function in inhibiting PEDV proliferation. The interaction of the viperin S-adenosylmethionine domain with the N protein of PEDV was confirmed via confocal laser scanning microscopy and co-immunoprecipitation. This interaction might interfere with viral replication or assembly to reduce virus proliferation. Our results highlight a potential mechanism whereby viperin is able to inhibit PEDV replication and play an antiviral role in innate immunity.

E3 ubiquitin ligase ASB8 negatively regulates interferon via regulating TBK1/IKKi homeostasis.

Cells recognize virus nucleic acid by pattern recognition receptors (PRRs), virus involve in the activation of signaling cascade of variable adaptor proteins, TANK-binding kinase1(TBK1)/ inhibitor of nuclear factor kappa-B kinase subunit epsilon(IKKi) complex, IκB kinase(IKKs) to trigger activation of transcription factor, interferon regulatory factor 3/7(IRF3/7), ultimately, leading to the production of type I interferon and exert anti-viral effects. In this study, E3 ubiquitin ligase ankyrin repeat and SOCS box-containing 8(ASB8) interacted with TBK1/IKKi and phosphorylation modification of ASB8 at site of Ser17 to further strengthen its ubiquitination activity were verified. Conversely, phosphorylated ASB8 accelerate K48-linked ubiquitination and degradation of TBK1/IKKi, which further reduces phosphorylation level of IRF3 and inhibits production of IFN-ß. At the same time, a new bridge molecule Leucine-rich repeat containing protein 10B(LRRC10B) upregulated after viral infection are involved in the formation and interaction with ASB8-TBK1/IKKi complex was reported. Our study reveals a new mechanism of ubiquitin ligase ASB8 modulating antiviral innate immunity by altering stability of TBK1/IKKi kinase complex.

PSCA is a target of chimeric antigen receptor T cells in gastric cancer.

BACKGROUND: Gastric cancer is a deadly malignancy and is a prognostically unfavorable entity with restricted therapeutic strategies available. Prostate stem cell antigen (PSCA) is a glycosylphosphatidylinositol (GPI)-anchored cell surface protein widely expressed in bladder, prostate, and pancreatic cancers. Existing studies have thoroughly recognized the availability of utilizing anti-PSCA CAR-T cells in the treatment of metastatic prostate cancer and non-small-cell lung cancer. However, no previous study has investigated the feasibility of using anti-PSCA CAR-T cells to treat gastric cancer, irrespective of the proven expression of PSCA on the gastric cancer cell surface. METHODS: We determined the expression of PSCA in several primary tumor tissues and constructed third-generation anti-PSCA CAR-T cells. We then incubated anti-PSCA CAR-T cells and GFP-T cells with target tumor cell lines at E:T ratios of 2:1, 1:1, 1:2, and 1:4 to evaluate the therapeutic efficacy of anti-PSCA CAR-T cells in vitro. We also assayed canonical T cell activation markers after coculturing anti-PSCA CAR-T cells with target cell lines by flow cytometry. The detection of a functional cytokine profile was carried out via enzyme-linked immunosorbent assays. We then evaluated the antitumor activity of anti-PSCA CAR-T cells in vivo by establishing two different xenograft GC mouse models. RESULTS: Anti-PSCA CAR-T cells exhibited upregulated activation markers and increased cytokine production profiles related to T cell cytotoxicity in an antigen-dependent manner. Moreover, anti-PSCA CAR-T cells exhibited robust anti-tumor cytotoxicity in vitro. Importantly, we demonstrated that anti-PSCA CAR-T cells delivered by peritumoral injection successfully stunted tumor progression in vivo. However, intravenous administration of anti-PSCA CAR-T cells failed to reveal any therapeutic improvements. CONCLUSIONS: Our findings corroborated the feasibility of anti-PSCA CAR-T cells and their efficacy against gastric cancer, implicating the potential of applying anti-PSCA CAR-T cells to treat GC patients in the clinic.

Ca2+ and SO42- accelerate the reduction of Cr(VI) by Penicillium oxalicum SL2.

Some ions in soils may affect the growth and metabolism of microorganisms and subsequently alter the remediation efficiency of Cr(VI). Here, the effects of different Ca2+ and SO42- levels on the reduction of Cr(VI) by Penicillium oxalicum SL2 were investigated. The results showed that Cr(VI) reduction by P. oxalicum SL2 in potato dextrose liquid (PDL) medium was accelerated by the presence of exogenous Ca2+ and SO42-. The Cr(VI) reduction rates were increased by 52.5% (200 mg L-1 Ca2+ treated) and 55.9% (2000 mg L-1 SO42- treated), respectively. High concentration of Ca2+ in medium resulted in the production of calcium oxalate crystals, which was contributed to the adsorption of chromium. In addition, X-ray absorption near-edge spectroscopy (XANES) analysis showed that P. oxalicum SL2 could reduce the toxicity of Cr(VI) by synthesizing cysteine (Cys) and reduced glutathione (GSH). The decrease of thiol compounds (Cys and GSH) in P. oxalicum SL2 mycelia treated with SO42- proved the alleviation of oxidative stress. In conclusion, exogenous Ca2+ could reduce the damage of Cr(VI) to P. oxalicum SL2 by maintaining the integrity of cell wall, and the addition of SO42- alleviated the Cr(VI) toxicity to P. oxalicum SL2, thus accelerating the reduction of Cr(VI).