A zebrafish NLRX1 isoform downregulates fish IFN responses by targeting the adaptor STING.

In mammals, NLRX1 is a unique member of the nucleotide-binding domain and leucine-rich repeat (NLR) family showing an ability to negatively regulate IFN antiviral immunity. Intron-containing genes, including NLRX1, have more than one transcript due to alternative splicing; however, little is known about the function of its splicing variants. Here, we identified a transcript variant of NLRX1 in zebrafish (Danio rerio), termed NLRX1-tv4, as a negative regulator of fish IFN response. Zebrafish NLRX1-tv4 was slightly induced by viral infection, with an expression pattern similar to the full-length NLRX1. Despite the lack of an N-terminal domain that exists in the full-length NLRX1, overexpression of NLRX1-tv4 still impaired fish IFN antiviral response and promoted viral replication in fish cells, similar to the full-length NLRX1. Mechanistically, NLRX1-tv4 targeted STING for proteasome-dependent protein degradation by recruiting an E3 ubiquitin ligase RNF5 to drive the K48-linked ubiquitination, eventually downregulating the IFN antiviral response. Mapping of NLRX1-tv4 domains showed that its N-terminal and C-terminal regions exhibited a similar potential to inhibit STING-mediated IFN antiviral response. Our findings reveal that like the full-length NLRX1, zebrafish NLRX-tv4 functions as an inhibitor to shape fish IFN antiviral response.IMPORTANCEIn this study, we demonstrate that a transcript variant of zebrafish NLRX1, termed NLRX1-tv4, downregulates fish IFN response and promotes virus replication by targeting STING for protein degradation and impairing the interaction of STING and TBK1 and that its N- and C-terminus exhibit a similar inhibitory potential. Our results are helpful in clarifying the current contradictory understanding of structure and function of vertebrate NLRX1s.

A finTRIM Family Protein Acquires RNA-Binding Activity and E3 Ligase Activity to Shape the IFN Response in Fish.

Tripartite motif (TRIM) family proteins have come forth as important modulators of innate signaling dependent on of E3 ligase activity. Recently, several human TRIM proteins have been identified as unorthodox RNA-binding proteins by RNA interactome analyses; however, their targets and functions remain largely unknown. FTRCA1 is a crucian carp (Carassius auratus)-specific finTRIM (fish novel TRIM) member and negatively regulates the IFN antiviral response by targeting two retinoic acid-inducible gene-I (RIG-I)-like receptor (RLR) pathway molecules, that is, TANK-binding kinase 1 (TBK1) and IFN regulatory factor 7 (IRF7). In this study, we identify FTRCA1 as an RNA-binding E3 ligase and characterize the contribution of its RNA-binding activity and E3 ligase activity to fish IFN response. Besides targeting TBK1 and IRF7, FTRCA1 downregulates fish IFN response also by targeting stimulator of IFN response cGAMP interactor 1 (STING1). E3 ligase activity is required for full inhibition on the TBK1- and IRF7-mediated IFN response, but partial inhibition on the STING1-mediated IFN response. However, FTRCA1 has a general binding potential to mRNAs in vitro, it selectively binds STING1 and IRF7 mRNAs in vivo to attenuate mRNA levels, and it directly interacts with TBK1 protein to target protein degradation for downregulating the IFN response. Our results present an interesting example of a fish species-specific finTRIM protein that has acquired RNA-binding activity and E3 ligase activity to fine-tune fish IFN response.

A Novel Non-Mammalian-Specific HERC7 Negatively Regulates IFN Response through Degrading RLR Signaling Factors.

The small HERC family currently comprises four members (HERC3-6) involved in the regulation of various physiological activities. Little is known about the role of HERCs in IFN response. In this study, we identify a novel fish HERC member, named crucian carp HERC7, as a negative regulator of fish IFN response. Genome-wide search of homologs and comprehensive phylogenetic analyses reveal that the small HERC family, apart from HERC3-6 that have been well-characterized in mammals, contains a novel HERC7 subfamily exclusively in nonmammalian vertebrates. Lineage-specific and even species-specific expansion of HERC7 subfamily in fish indicates that crucian carp HERC7 might be species-specific. In virally infected fish cells, HERC7 is induced by IFN and selectively targets three retinoic acid-inducible gene-I-like receptor signaling factors for degradation to attenuate IFN response by two distinct strategies. Mechanistically, HERC7 delivers mediator of IFN regulatory factor 3 activator and mitochondrial antiviral signaling protein for proteasome-dependent degradation at the protein level and facilitates IFN regulatory factor 7 transcript decay at the mRNA level, thus abrogating cellular IFN induction to promote virus replication. Whereas HERC7 is a putative E3 ligase, the E3 ligase activity is not required for its negative regulatory function. These results demonstrate that the ongoing expansion of the small HERC family generates a novel HERC7 to fine-tune fish IFN antiviral response.

Sequential CD19 and CD22 chimeric antigen receptor T-cell therapy for childhood refractory or relapsed B-cell acute lymphocytic leukaemia: a single-arm, phase 2 study.

BACKGROUND: Relapses frequently occur following CD19-directed chimeric antigen receptor (CAR) T-cell treatment for relapsed or refractory B-cell acute lymphocytic leukaemia in children. We aimed to assess the activity and safety of sequential CD19-directed and CD22-directed CAR T-cell treatments. METHODS: This single-centre, single-arm, phase 2 trial, done at Beijing GoBroad Boren Hospital, Beijing, China, included patients aged 1-18 years who had relapsed or refractory B-cell acute lymphocytic leukaemia with CD19 and CD22 positivity greater than 95% and an Eastern Cooperative Oncology Group performance status of 0-2. Patients were initially infused with CD19-directed CAR T cells intravenously, followed by CD22-directed CAR T-cell infusion after minimal residual disease-negative complete remission (or complete remission with incomplete haematological recovery) was reached and all adverse events (except haematological adverse events) were grade 2 or better. The target dose for each infusion was 0·5 × 106 to 5·0 × 106 cells per kg. The primary endpoint was objective response rate at 3 months after the first infusion. Secondary endpoints were duration of remission, event-free survival, disease-free survival, overall survival, safety, pharmacokinetics, and B-cell quantification. The prespecified activity analysis included patients who received the target dose and the safety analysis included all treated patients. This study is registered with ClinicalTrials.gov, NCT04340154, and enrolment has ended. FINDINGS: Between May 28, 2020, and Aug 16, 2022, 81 participants were enrolled, of whom 31 (38%) were female and 50 (62%) were male. Median age was 8 years (IQR 6-10), all patients were Asian. All 81 patients received the first infusion and 79 (98%) patients received sequential infusions, CD19-directed CAR T cells at a median dose of 2·7 × 106 per kg (IQR 1·1 × 106 to 3·7 × 106) and CD22-directed CAR T cells at a median dose of 2·2 × 106 per kg (1·1 × 106 to 3·7 × 106), with a median interval of 39 days (37-41) between the two infusions. 62 (77%) patients received the target dose, including two patients who did not receive CD22 CAR T cells. At 3 months, 60 (97%, 95% CI 89-100) of the 62 patients who received the target dose had an objective response. Median follow-up was 17·7 months (IQR 11·4-20·9). 18-month event-free survival for patients who received the target dose was 79% (95% CI 66-91), duration of remission was 80% (68-92), and disease-free survival was 80% (68-92) with transplantation censoring; overall survival was 96% (91-100). Common adverse events of grade 3 or 4 between CD19-directed CAR T-cell infusion and 30 days after CD22-directed CAR T-cell infusion included cytopenias (64 [79%] of 81 patients), cytokine release syndrome (15 [19%]), neurotoxicity (four [5%]), and infections (five [6%]). Non-haematological adverse events of grade 3 or worse more than 30 days after CD22-directed CAR T-cell infusion occurred in six (8%) of 79 patients. No treatment-related deaths occurred. CAR T-cell expansion was observed in all patients, with a median peak at 9 days (IQR 7-14) after CD19-directed and 12 days (10-15) after CD22-directed CAR T-cell infusion. At data cutoff, 35 (45%) of 77 evaluable patients had CAR transgenes and 59 (77%) had B-cell aplasia. INTERPRETATION: This sequential strategy induced deep and sustained responses with an acceptable toxicity profile, and thus potentially provides long-term benefits for children with this condition. FUNDING: The National Key Research & Development Program of China, the CAMS Innovation Fund for Medical Sciences (CIFMS), and the Non-Profit Central Research Institute Fund of Chinese Academy of Medical Sciences. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

Interferon regulatory factor 1 (IRF1) and anti-pathogen innate immune responses.

The eponymous member of the interferon regulatory factor (IRF) family, IRF1, was originally identified as a nuclear factor that binds and activates the promoters of type I interferon genes. However, subsequent studies using genetic knockouts or RNAi-mediated depletion of IRF1 provide a much broader view, linking IRF1 to a wide range of functions in protection against invading pathogens. Conserved throughout vertebrate evolution, IRF1 has been shown in recent years to mediate constitutive as well as inducible host defenses against a variety of viruses. Fine-tuning of these ancient IRF1-mediated host defenses, and countering strategies by pathogens to disarm IRF1, play crucial roles in pathogenesis and determining the outcome of infection.

KHDRBS3 accelerates glycolysis and promotes malignancy of hepatocellular carcinoma via upregulating 14-3-3ζ.

BACKGROUND: Primary hepatocellular carcinoma (HCC) is a malignancy with high morbidity and mortality. KH domain-containing, RNA-binding signal transduction-associated protein 3 (KHDRBS3) is an RNA-binding protein that is aberrantly expressed in multiple tumors; however, its expression and biological function in HCC have not been reported. METHODS: KHDRBS3 knockdown and overexpression were performed using the lentiviral vector system to investigate the effects of KHDRBS3 on cell proliferation, apoptosis, chemoresistance, and glycolysis. Murine xenograft tumor models were constructed to study the role of KHDRBS3 on tumor growth in vivo. Furthermore, RNA-Pull Down and RNA immunoprecipitation were utilized to explore the interaction between KHDRBS3 and 14-3-3ζ, a phosphopeptide-binding molecule encoded by YWHAZ. RESULTS: KHDRBS3 was highly expressed in human HCC tissues and predicted the poor prognosis of patients with HCC. Knockdown of KHDRBS3 exhibited a carcinostatic effect in HCC and impeded proliferation and tumor growth, reduced glycolysis, enhanced cell sensitivity to doxorubicin, and induced apoptosis. On the contrary, forced expression of KHDRBS3 expedited the malignant biological behaviors of HCC cells. The expression of KHDRBS3 was positively correlated with the expression of 14-3-3ζ. RNA immunoprecipitation and RNA pull-down assays demonstrated that KHDRBS3 bound to YWHAZ. We further confirmed that 14-3-3ζ silencing significantly reversed the promotion of proliferation and glycolysis and the inhibition of apoptosis caused by KHDRBS3 overexpression. CONCLUSIONS: Our findings suggest that KHDRBS3 promotes glycolysis and malignant progression of HCC through upregulating 14-3-3ζ expression, providing a possible target for HCC therapy.

Metabolic Syndrome and Clinical Outcomes of Patients with Gastric Cancer: A Meta-Analysis.

Metabolic syndrome (MetS) is suggested to participate in the pathogenesis and progress of some cancers via inducing low-grade systemic inflammation. However, the influence of MetS on patients with gastric cancer (GC) remains not fully determined. A systematic review and meta-analysis was therefore performed to evaluate the influence of MetS on clinical outcomes of patients with GC. A search of PubMed, Embase, Web of Science, Wanfang, and CNKI retrieved relevant cohort studies from the inception of the databases to October 11, 2022. We pooled the results using a random-effects model that incorporates heterogeneity. In the meta-analysis, 6649 patients with GC were included, and all of them received gastrectomy. A total of 1248 (18.8%) patients had MetS at baseline. Pooled results showed that MetS was associated with higher risks of postoperative complications [risk ratio (RR): 2.41, 95% confidence interval (CI): 1.85 to 3.14, p<0.001; I2=55%], overall mortality (RR: 1.73, 95% CI: 1.85 to 3.14, p<0.001; I2=77%), and recurrence of GC (RR: 2.00, 95% CI: 1.10 to 3.63, p=0.02; I2=39%). Subgroup analyses showed similar results in prospective and retrospective cohort studies and in studies with MetS diagnosed with the Chinese Diabetes Society criteria and the National Cholesterol Education Program Adult Treatment Panel III criteria (p for subgroup difference all>0.05). In patients with GC after gastrectomy, MetS may be a predictor of high incidence of postoperative complications, cancer recurrence, and overall mortality.

MiR-494-3p aggravates pirarubicin-induced cardiomyocyte injury by regulating MDM4/p53 signaling pathway.

OBJECTIVE: Pirarubicin (THP) is a widely used antitumor drug in clinical practice, but its cardiotoxicity limits its use. There is an urgent need to find drugs to alleviate the cardiotoxicity of THP. This study aimed to investigate the effect and mechanism of miR-494-3p on THP-induced cardiomyocytes. METHODS: THP induced immortalized mouse cardiomyocytes HL-1, silenced or overexpressed miR-494-3p. The effects of miR-494-3p on HL-1 contained in THP were investigated by CCK8, flow cytometry, ROS detection, JC-1 mitochondrial membrane potential detection, TUNEL cell apoptosis detection, RT-qPCR, and Western blot. RESULTS: miR-494-3p could reduce cell viability, increase oxidative damage, and promote cell apoptosis; at the same time, it inhibited the expression of MDM4, promoted the activation of p53, and promoted the expression of apoptosis-related proteins. MiR-494-3p inhibitors have the opposite effect. CONCLUSION: miR-494-3p can aggravate THP damage to HL-1, which may be achieved by downregulating MDM4 and promoting p53. miR-494-3p is one of the important miRNAs in THP-induced cardiotoxicity, which provides theoretical support for its possible use as a therapeutic target for THP-induced cardiovascular disease.

Risk factor analysis and prediction of postoperative clinically relevant pancreatic fistula after distal pancreatectomy.

OBJECTIVE: Postoperative pancreatic fistula (POPF) following distal pancreatectomy (DP) is a serious complication. In the present study, we aimed to identify the risk factors associated with clinically relevant postoperative pancreatic fistula (CR-POPF) and establish a nomogram model for predicting CR-POPF after DP. METHODS: In total, 115 patients who underwent DP at the General Hospital of Northern Theater Command between January 2005 and December 2020 were retrospectively studied. Univariate and multivariable logistic regression analyses were used to identify the independent risk factors associated with CR-POPF. Then, a nomogram was formulated based on the results of multivariable logistic regression analysis. The predictive performance was evaluated with receiver operating characteristic (ROC) curves. Decision curve and clinical impact curve analyses were used to validate the clinical application value of the model. RESULTS: The incidence of CR-POPF was 33.0% (38/115) in the present study. Multivariate logistic regression analysis identified the following variables as independent risk factors for POPF: body mass index (BMI) (OR 4.658, P = 0.004), preoperative albumin level (OR 7.934, P = 0.001), pancreatic thickness (OR 1.256, P = 0.003) and pancreatic texture (OR 3.143, P = 0.021). We created a nomogram by incorporating the above mentioned risk factors. The nomogram model showed better predictive value, with a concordance index of 0.842, sensitivity of 0.710, and specificity of 0.870 when compared to each risk factor. Decision curve and clinical impact curve analyses also indicated that the nomogram conferred a high clinical net benefit. CONCLUSION: Our nomogram could accurately and objectively predict the risk of postoperative CR-POPF in individuals who underwent DP, which could help clinicians with early identification of patients who might develop CR-POPF and early development of a suitable fistula mitigation strategy and postoperative management.

Zebrafish HERC7c Acts as an Inhibitor of Fish IFN Response.

In humans, four small HERCs (HERC3-6) exhibit differential degrees of antiviral activity toward HIV-1. Recently we revealed a novel member HERC7 of small HERCs exclusively in non-mammalian vertebrates and varied copies of herc7 genes in distinct fish species, raising a question of what is the exact role for a certain fish herc7 gene. Here, a total of four herc7 genes (named HERC7a-d sequentially) are identified in the zebrafish genome. They are transcriptionally induced by a viral infection, and detailed promoter analyses indicate that zebrafish herc7c is a typical interferon (IFN)-stimulated gene. Overexpression of zebrafish HERC7c promotes SVCV (spring viremia of carp virus) replication in fish cells and concomitantly downregulates cellular IFN response. Mechanistically, zebrafish HERC7c targets STING, MAVS, and IRF7 for protein degradation, thus impairing cellular IFN response. Whereas the recently-identified crucian carp HERC7 has an E3 ligase activity for the conjugation of both ubiquitin and ISG15, zebrafish HERC7c only displays the potential to transfer ubiquitin. Considering the necessity for timely regulation of IFN expression during viral infection, these results together suggest that zebrafish HERC7c is a negative regulator of fish IFN antiviral response.

Identification of novel risk loci with shared effects on alcoholism, heroin, and methamphetamine dependence.

Different substance dependences have common effects on reward pathway and molecular adaptations, however little is known regarding their shared genetic factors. We aimed to identify the risk genetic variants that are shared for substance dependence (SD). First, promising genome-wide significant loci were identified from 3296 patients (521 alcoholic/1026 heroin/1749 methamphetamine) vs 2859 healthy controls and independently replicated using 1954 patients vs 1904 controls. Second, the functional effects of promising variants on gene expression, addiction characteristics, brain structure (gray and white matter), and addiction behaviors in addiction animal models (chronic administration and self-administration) were assessed. In addition, we assessed the genetic correlation among the three SDs using LD score regression. We identified and replicated three novel loci that were associated with the common risk of heroin, methamphetamine addiction, and alcoholism: ANKS1B rs2133896 (Pmeta = 3.60 × 10-9), AGBL4 rs147247472 (Pmeta = 3.40 × 10-12), and CTNNA2 rs10196867 (Pmeta = 4.73 × 10-9). Rs2133896 in ANKS1B was associated with ANKS1B gene expression and had effects on gray matter of the left calcarine and white matter of the right superior longitudinal fasciculus in heroin dependence. Overexpression of anks1b gene in the ventral tegmental area decreased addiction vulnerability for heroin and methamphetamine in self-administration rat models. Our findings could shed light on the root cause for substance dependence and will be helpful for the development of cost-effective prevention strategies for general addiction disorders.

Characterization of DNA Binding and Nuclear Retention Identifies Zebrafish IRF11 as a Positive Regulator of IFN Antiviral Response.

In mammals, transcription factors of IFN-regulatory factors (IRFs) family translate viral recognition into IFN antiviral responses through translocating to nucleus and subsequently binding to the promoters of IFN and IFN-stimulated genes (ISGs). In addition to IRF1-9 conserved across vertebrates and IRF10 in teleost fish and bird, teleost fish has another novel member, IRF11; however, little is known about its role in IFN response. In this study, we provide evidence that IRF11 is present only in Osteichthyes (bony fish) but lost in tetrapods and subsequently characterize the stimulatory potential of zebrafish IRF11 to IFN antiviral response relevant to its subcellular localization and promoter binding. Overexpression of zebrafish IRF11 restricts virus replication through induction of IFN and ISGs. Zebrafish IRF11 is constitutively localized to nucleus, which is driven by a tripartite NLS motif, consisting of three interdependent basic clusters, two in DNA binding domain (DBD) and one in the region immediately C-terminal to DBD. Nuclear IRF11 binds to the IRF-binding element/IFN-stimulated response element motifs of zebrafish IFN promoters depending on the two conserved amino acids (K78, R82) within DBD helix α3. K78 and R82 also benefit zebrafish IRF11 nuclear import as two key residues positioned at the first basic cluster of the tripartite NLS motif. Such features enable zebrafish IRF11 to function as a positive transcription factor for fish IFN antiviral response. Our results identify a unique tripartite NLS motif that integrates DNA-binding activity and nuclear import ability, allowing zebrafish IRF11 to initiate IFN and ISG expression.

Understanding motivations behind medical student involvement in COVID-19 pandemic relief efforts.

BACKGROUND: Although students were removed from patient-facing settings at the beginning of the COVID-19 pandemic due to concerns of burdening teaching physicians and depleting personal protective equipment, some leaders suggest students can be effectively utilized when personnel resources may be scarce. There have been narrative discussions surrounding medical student involvement, but no studies exploring the attitudes of these students. The authors aim to quantify the degree to which factors influenced a medical student's decision to or to not volunteer during the pandemic and to characterize medical students' attitudes towards medical professionals' duty to serve in a pandemic. METHODS: The authors developed and tested a secure web-based survey before distribution to students at 23 different US allopathic medical schools that did not graduate medical students early to aid in pandemic efforts between April and June 2020. Of the 599 students who completed the survey, 65.5% self-identified as female and were on average 25.94 years old (SD = 2.5). Multiple comparisons were made based on volunteer status. Ordinal scale questions were compared with the Mann Whitney U test, and the Chi-Squared test was used for categorical variables using R version 3.62. RESULTS: 67.6% of students volunteered in pandemic relief activities and a majority of those students volunteered in non-patient-facing roles. Community service, new skills, and time commitment were top 3 influencing factors for students who volunteered, while risk to other, time commitment, and risk to self were top 3 influencing factors for students who chose not to volunteer. Compared to other specialties, students interested in primary care specialties agreed to a greater degree that physicians have a duty to serve in pandemic relief efforts. CONCLUSIONS: Medical students who volunteered cited self-serving factors and altruistic values as significant motivators. Students who did not volunteer were significantly more concerned with risks of COVID-19 exposure. However, medical students in general agreed that students should be allowed to volunteer in COVID-19 related relief efforts. As large areas of the United States continue to experience increases in COVID-19 cases, institutions should involve medical students in balancing the level of acceptable risk with the educational benefits.

Noncovalent microarrays from synthetic amino-terminating glycans: Implications in expanding glycan microarray diversity and platform comparison.

Glycan microarrays have played important roles in detection and specificity assignment of glycan recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent and noncovalent immobilization are the two types of methods used, but a direct comparison of results from the two platforms is required. Here we develop a chemical strategy to prepare lipid-linked probes from both naturally derived aldehyde-terminating and synthetic amino-terminating glycans that addresses the two aspects: expansion of sequence-defined glycan libraries and comparison of the two platforms. We demonstrate the specific recognition by plant and mammalian lectins, carbohydrate-binding modules and antibodies and the overall similarities from the two platforms. Our results provide new knowledge on unique glycan-binding specificities for the immune receptor Dectin-1 toward ß-glucans and the interaction of rotavirus P[19] adhesive protein with mucin O-glycan cores.

Transcription of Il17 and Il17f is controlled by conserved noncoding sequence 2.

T helper 17 (Th17) cells specifically transcribe the Il17 and Il17f genes, which are localized in the same chromosome region, but the underlying mechanism is unclear. Here, we report a cis element that we previously named conserved noncoding sequence 2 (CNS2) physically interacted with both Il17 and Il17f gene promoters and was sufficient for regulating their selective transcription in Th17 cells. Targeted deletion of CNS2 resulted in impaired retinoic acid-related orphan receptor gammat (RORγt)-driven IL-17 expression in vitro. CNS2-deficient T cells also produced substantially decreased amounts of IL-17F. These cytokine defects were associated with defective chromatin remodeling in the Il17-Il17f gene locus, possibly because of effects on CNS2-mediated recruitment of histone-modifying enzymes p300 and JmjC domain-containing protein 3 (JMJD3). CNS2-deficient animals were also shown to be resistant to experimental autoimmune encephalomyelitis (EAE). Our results thus suggest that CNS2 is sufficient and necessary for Il17 and optimal Il17f gene transcription in Th17 cells.

FTRCA1, a Species-Specific Member of finTRIM Family, Negatively Regulates Fish IFN Response through Autophage-Lysosomal Degradation of TBK1.

In mammals, tripartite motif (TRIM) proteins have emerged as pivotal players endowed with, directly, antiviral effects and, indirectly, modulatory capacity of the innate immune response. An unprecedented expansion of TRIM family has occurred in fish; however, the functional role of fish TRIM family members remains largely unknown. In this study, we identify a species-specific TRIM gene from crucian carp Carassius auratus, named FTRCA1, phylogenetically similar to the members of finTRIM, a subfamily of TRIM exclusively in teleost fish. FTRCA1 is induced by IFN and IFN stimuli as a typical IFN-stimulated gene. Overexpression of FTRCA1 negatively regulates IFN antiviral response by inhibition of IRF3 phosphorylation; consistently, knockdown of FTRCA1 results in enhanced levels of IRF3 phosphorylation and also IFN expression following poly(I:C) transfection. Whereas FTRCA1 is associated with several pivotal signaling molecules of RIG-I-like receptor pathway, its association with TBK1 results in autophage-lysosomal degradation of TBK1, thus abrogating the downstream IFN induction. Interestingly, FTRCA1 is phosphorylated by TBK1, but this phosphorylation is not required for downregulation of TBK1 protein. Transfection assays indicate that FTRCA1 is likely an E3 ligase with the requirement of RING finger domain, and deletion of N-terminal RING domain or mutation of seven conservative sites abolishes the negative regulatory function of FTRCA1. Collectively, these results illuminate a novel finTRIM-mediated innate immune modulatory pathway, thus providing insights into species-specific regulation of fish IFN response.

Glycan Markers of Human Stem Cells Assigned with Beam Search Arrays.

Glycan antigens recognized by monoclonal antibodies have served as stem cell markers. To understand regulation of their biosynthesis and their roles in stem cell behavior precise assignments are required. We have applied state-of-the-art glycan array technologies to compare the glycans bound by five antibodies that recognize carbohydrates on human stem cells. These are: FC10.2, TRA-1-60, TRA-1-81, anti-i and R-10G. Microarray analyses with a panel of sequence-defined glycans corroborate that FC10.2, TRA-1-60, TRA-1-81 recognize the type 1-(Galß-3GlcNAc)-terminating backbone sequence, Galß-3GlcNAcß-3Galß-4GlcNAcß-3Galß-4GlcNAc, and anti-i, the type 2-(Galß-4GlcNAc) analog, Galß-4GlcNAcß-3Galß-4GlcNAcß-3Galß-4GlcNAc, and we determine substituents they can accommodate. They differ from R-10G, which requires sulfate. By Beam Search approach, starting with an antigen-positive keratan sulfate polysaccharide, followed by targeted iterative microarray analyses of glycan populations released with keratanases and mass spectrometric monitoring, R-10G is assigned as a mono-sulfated type 2 chain with 6-sulfation at the penultimate N-acetylglucosamine, Galß-4GlcNAc(6S)ß-3Galß-4GlcNAcß-3Galß-4GlcNAc. Microarray analyses using newly synthesized glycans corroborate the assignment of this unique determinant raising questions regarding involvement as a ligand in the stem cell niche.

Gender differences in refraction prediction error of five formulas for cataract surgery.

OBJECTIVES: To evaluate gender differences in optical biometry measurements and lens power calculations. METHODS: Eight thousand four hundred thirty-one eyes of five thousand five hundred nineteen patients who underwent cataract surgery at University of Michigan's Kellogg Eye Center were included in this retrospective study. Data including age, gender, optical biometry, postoperative refraction, implanted intraocular lens (IOL) power, and IOL formula refraction predictions were gathered and/or calculated utilizing the Sight Outcomes Research Collaborative (SOURCE) database and analyzed. RESULTS: There was a statistical difference between every optical biometry measure between genders. Despite lens constant optimization, mean signed prediction errors (SPEs) of modern IOL formulas differed significantly between genders, with predictions skewed more hyperopic for males and myopic for females for all 5 of the modern IOL formulas tested. Optimization of lens constants by gender significantly decreased prediction error for 2 of the 5 modern IOL formulas tested. CONCLUSIONS: Gender was found to be an independent predictor of refraction prediction error for all 5 formulas studied. Optimization of lens constants by gender can decrease refraction prediction error for certain modern IOL formulas.

Visual Outcomes in Surgically Treated Intracranial Meningiomas.

BACKGROUND: Intracranial meningiomas that arise from the medial sphenoid ridge, anterior clinoid process, tuberculum sellae, or planum sphenoidale often impair vision by compressing the optic nerves and optic chiasm. Although many studies have reported visual outcome following surgery for these tumors, documentation has often been incomplete and not validated by patient self-report. METHODS: Retrospective study of 40 patients drawn from a single, academic, medical center. We used a unique method of assessing visual outcome based on whether the change in visual function affected the preoperatively better-sighted or worse-sighted eye in the belief that this method would correlate with effects on activities of daily living (ADL). To elicit patient self-reports of those effects, we conducted telephone interviews of 25 patients with a standard questionnaire. We also assessed putative ophthalmic, imaging, and surgical predictors of visual outcome. RESULTS: Visual improvement occurred in 61% of patients with preoperative monocular visual dysfunction, but only 22% of patients reported improvement in their ability to conduct ADL, and 17% lost vision. Visual outcomes were better in patients with preoperative binocular visual dysfunction, where visual improvement occurred in 73% and no patient lost vision in the preoperatively better-sighted eye. However, only 27% of patients with preoperative binocular visual dysfunction reported improvement in their ability to conduct ADL. Long duration of vision impairment, presence of optic disc pallor, large tumor size, and imaging-based preoperative optic canal involvement did not preclude a favorable visual outcome. Aggressive surgical reduction in displacement of the optic nerves was not necessary to obtain a favorable visual outcome and sometimes led to an unfavorable visual outcome. CONCLUSIONS: In this study, surgery often improved vision, especially in patients with preoperative binocular visual dysfunction. But patients indicated that the effect on their ability to perform ADL was more modest. Moreover, 17% of patients with preoperative monocular visual dysfunction lost vision in the only affected eye, often to a considerable degree. In those patients, surgery would be justified primarily to relieve the concern of having a large brain tumor and to prevent tumor growth. Preoperative ophthalmic and imaging features poorly predicted visual outcomes. Favorable visual outcomes occurred without aggressive surgical debulking of the tumors.

14-3-3ζ binds to and stabilizes phospho-beclin 1S295 and induces autophagy in hepatocellular carcinoma cells.

Data from The Cancer Genome Atlas (TCGA) indicate that the expression levels of 14-3-3ζ and beclin 1 (a key molecule involved in cellular autophagy) are up-regulated and positively correlated with each other (R = .5, P < .05) in HCC tissues. Chemoresistance developed in hepatoma cancer cells is associated with autophagy initiation. This study aimed to explore 14-3-3ζ's role in regulating autophagy in HCC cells, with a focus on beclin 1. The co-localization of 14-3-3ζ and beclin 1 was detectable in primary HCC tissues. To simulate in vivo tumour microenvironment (hypoxia), CSQT-2 and HCC-LM3 cells were exposed to 2% oxygen for 24 hours. The protein levels of 14-3-3ζ and phospho-beclin 1S295 peaked at 12 hours following hypoxia. Meanwhile, the strongest autophagy flux occurred: LC3II was increased, and p62 was decreased significantly. By sequencing the coding area of BECN 1 gene of CSQT-2 and HCC-LM3 cells, we found that the predicted translational products of BECN 1 gene contained RLPS295 VP (R, arginine; L, leucine; P, proline; S, serine; V, valine), a classic 14-3-3ζ binding motif. CO-IP results confirmed that 14-3-3ζ bound to beclin 1, and this connection was markedly weakened when S295 was mutated into A295 (alanine). Further, 14-3-3ζ overexpression prevented phospho-beclin 1S295 from degradation and enhanced its binding to VPS34, whilst its knockdown accelerated the degradation. Additionally, 14-3-3ζ enhanced the chemoresistance of HCC cells to cis-diammined dichloridoplatium by activating autophagy. Our work reveals that 14-3-3ζ binds to and stabilizes phospho-beclin 1S295 and induces autophagy in HCC cells to resist chemotherapy.