Proper RPA acetylation promotes accurate DNA replication and repair.

The single-stranded DNA (ssDNA) binding protein complex RPA plays a critical role in promoting DNA replication and multiple DNA repair pathways. However, how RPA is regulated to achieve its functions precisely in these processes remains elusive. Here, we found that proper acetylation and deacetylation of RPA are required to regulate RPA function in promoting high-fidelity DNA replication and repair. We show that yeast RPA is acetylated on multiple conserved lysines by the acetyltransferase NuA4 upon DNA damage. Mimicking constitutive RPA acetylation or blocking its acetylation causes spontaneous mutations with the signature of micro-homology-mediated large deletions or insertions. In parallel, improper RPA acetylation/deacetylation impairs DNA double-strand break (DSB) repair by the accurate gene conversion or break-induced replication while increasing the error-prone repair by single-strand annealing or alternative end joining. Mechanistically, we show that proper acetylation and deacetylation of RPA ensure its normal nuclear localization and ssDNA binding ability. Importantly, mutation of the equivalent residues in human RPA1 also impairs RPA binding on ssDNA, leading to attenuated RAD51 loading and homologous recombination repair. Thus, timely RPA acetylation and deacetylation likely represent a conserved mechanism promoting high-fidelity replication and repair while discriminating the error-prone repair mechanisms in eukaryotes.

Serum cytokine profiles during engraftment syndrome and acute graft-versus-host disease in adult patients after hematopoietic stem cell transplantation.

BACKGROUND: The underlying biology of engraftment syndrome (ES) following allogeneic hematopoietic stem cell transplantation (HSCT) is not fully elucidated, and the extent of its overlap with acute graft-versus-host disease (aGvHD) remains unclear. In order to establish potential indicator to distinguish ES more accurately, we conducted a retrospective analysis of cytokine levels during HSCT. METHODS: A total of 121 consecutive adult patients who underwent HSCT were enrolled in this study. Blood samples for interleukin (IL)-2, IL-2R, IL-4, IL-5, IL-6, IL-8, IL-10, IL-1ß, IL-12p70, interferon (IFN)-γ, IFN-α, tumor necrosis factor alpha (TNF-α) and C-reactive protein CRP were regularly assessed after transplantation and during transplantation related adverse events. Additionally, the balance of naïve, central memory and effector memory of CD4+ and CD8+ was analyzed around 30 and 60 days after stem cell infusion, respectively. RESULTS: Thirty (24.79 %) and 33 (27.27 %) patients were diagnosed with ES and aGvHD, respectively. ES was characterized by a significant increase in level of IL-5, IL-6, IL-8 and sIL-2R, while aGvHD was associated with a significant upregulation of IL-6, IL-5, IL-10 and sIL-2R in the patients from grade I to grade IV. Notably, patients got much higher levels of IL-6, IL-5 and sIL-2R when developed to ES than to aGvHD. Moreover, a pronounced shift from naïve to memory cells, both in CD4+ and CD8+ subsets, was found in ES patients. CONCLUSIONS: These findings suggest that cytokine profiles could serve as potential indicators for detecting and differentiating ES and aGvHD, enabling timely clinical intervention. Prospective clinical trials involving larger, independent patient cohorts are required to validate these observations.

Computed Diffusion-Weighted Images of Rectal Cancer: Image Quality, Restaging, and Treatment Response after Neoadjuvant Therapy.

BACKGROUND: Computed diffusion-weighted images (cDWI) of random b value could be derived from acquired DWI (aDWI) with at least two different b values. However, its comparison between aDWI and cDWI images in locally advanced rectal cancer (LARC) patients after neoadjuvant therapy (NT) is needed. PURPOSE: To compare the cDWI and aDWI in image quality, restaging, and treatment response of LARC after NT. STUDY TYPE: Retrospective. POPULATION: Eighty-seven consecutive patients. FIELD STRENGTH/SEQUENCE: 3.0 T/DWI. ASSESSMENT: All patients underwent two DWI sequences, including conventional acquisition with b = 0 and 1000 s/mm2 (aDWIb1000 ) and another with b = 0 and 700 s/mm2 on a 3.0-T MR scanner. The images of the latter were used to compute the diffusion images with b = 1000 s/mm2 (cDWIb1000 ). Four radiologists with 3, 4, 14, and 25 years of experience evaluated the images to compare the image quality, TN restaging performance, and treatment response between aDWIb1000 and cDWIb1000 . STATISTICAL TESTS: Interclass correlation coefficients, weighted κ coefficient, paired Wilcoxon, and McNemar or Fisher test were used. A significance level of 0.05 was used. RESULTS: The cDWIb1000 images were superior to the aDWIb1000 ones in both subjective and objective image quality. In T restaging, the overall diagnostic accuracy of cDWIb1000 images was higher than that of aDWIb1000 images (57.47% vs. 49.43%, P = 0.289 for the inexperienced radiologist; 77.01% vs. 63.22%, significant for the experienced radiologist), with better sensitivity in determining ypT0-Tis tumors. Additionally, it increased the sensitivity in detecting ypT2 tumors for the inexperienced radiologist and ypT3 tumors for the experienced radiologist. N restaging and treatment response were found to be similar between two sequences for both radiologists. DATA CONCLUSION: Compared to aDWIb1000 images, the computed ones might serve as a wise approach, providing comparable or better image quality, restaging performance, and treatment response assessment for LARC after NT. LEVEL OF EVIDENCE: 3 TECHNICAL EFFICACY STAGE: 2.

Rtt105 promotes high-fidelity DNA replication and repair by regulating the single-stranded DNA-binding factor RPA.

Single-stranded DNA (ssDNA) covered with the heterotrimeric Replication Protein A (RPA) complex is a central intermediate of DNA replication and repair. How RPA is regulated to ensure the fidelity of DNA replication and repair remains poorly understood. Yeast Rtt105 is an RPA-interacting protein required for RPA nuclear import and efficient ssDNA binding. Here, we describe an important role of Rtt105 in high-fidelity DNA replication and recombination and demonstrate that these functions of Rtt105 primarily depend on its regulation of RPA. The deletion of RTT105 causes elevated spontaneous DNA mutations with large duplications or deletions mediated by microhomologies. Rtt105 is recruited to DNA double-stranded break (DSB) ends where it promotes RPA assembly and homologous recombination repair by gene conversion or break-induced replication. In contrast, Rtt105 attenuates DSB repair by the mutagenic single-strand annealing or alternative end joining pathway. Thus, Rtt105-mediated regulation of RPA promotes high-fidelity replication and recombination while suppressing repair by deleterious pathways. Finally, we show that the human RPA-interacting protein hRIP-α, a putative functional homolog of Rtt105, also stimulates RPA assembly on ssDNA, suggesting the conservation of an Rtt105-mediated mechanism.

c-Met is a chimeric antigen receptor T-cell target for treating recurrent nasopharyngeal carcinoma.

BACKGROUND AIMS: Radiation therapy is the standard treatment for patients with nasopharyngeal carcinoma (NPC), but relapse occurs in 10% to 20% of patients. The treatment of recurrent nasopharyngeal carcinoma (rNPC) remains challenging. Chimeric antigen receptors (CAR)-T-cell therapy has achieved good outcomes in the treatment of leukemia and seems to be a promising therapeutic strategy for solid tumors. c-Met has been found to be highly expressed in multiple cancer types, and the activation of c-Met leads to the proliferation and metastasis of cancer cells. However, the expression of c-Met in rNPC tissues and whether it can be used as a target for CAR-T therapy in rNPC remain to be investigated. METHODS: We detected the expression of c-Met in 24 primary human rNPC tissues and three NPC cell lines and constructed two different antibody-derived anti-c-Met CARs, namely, Ab928z and Ab1028z. To estimate the function of these two different c-Met-targeted CAR-T cells, CD69 expression, cytotoxicity and cytokine secretion of CAR-T cells were assessed after coculture with target cells. A cell line-derived xenograft mouse model also was used to evaluate these two anti-c-Met CAR-T cells. Furthermore, we determined whether combination with an anti-EGFR antibody could promote the antitumor effect of CAR-T cells in a patient-derived xenograft mouse model. RESULTS: High c-Met expression was detected in 23 of 24 primary human rNPC tissues by immunohistochemistry staining and in three NPC cell lines by flow cytometry. Ab928z-T cells and Ab1028z-T cells showed significantly upregulated expression of CD69 after coculture with targeted cells. However, Ab1028z-T cells showed superior cytokine secretion and antitumor activity. Furthermore, Ab1028z-T cells effectively suppressed tumor growth compared with control CAR-T cells, and the combination with nimotuzumab further enhanced the tumor-clearing ability of Ab1028z-T cells. CONCLUSIONS: We found that c-Met is highly expressed in rNPC tissues and confirmed its potential as a CAR-T target for rNPC. Our study provides a new idea for the clinical treatment of rNPC.

IMNI PRECISION trial protocol: a phase II, open-label, non-inferior randomized controlled trial of tailoring omission of internal mammary node irradiation for early-stage breast cancer.

BACKGROUND: Since the publication of MA-20 and EORTC-22922 trials, chest wall (CW)/ whole breast (WB) irradiation + comprehensive regional nodal irradiation (RNI) with internal mammary node irradiation (IMNI) has been the standard adjuvant treatment for early-stage breast cancer (BC). However, one size does not fit all BC, and the risk of recurrence significantly varies among this patient population. In addition, whether all BC patients presented with one to three positive lymph nodes (pN1) could benefit from IMNI remains controversial. Thus, the optimal adjuvant RNI volume for early-stage BC with T1-2N1 remains undetermined. METHODS: The IMNI PRECISION trial is a single institute, open-labeled, non-inferior, randomized controlled trial. A total of 214 clinically "high risk" BC patients which is characterized as having at least two of the five clinically adverse factors (age ≤ 40, three positive LN, T2 stage, grade 3 and Ki-67 index ≥ 14%), but genomic score "low risk" (the genomic score ≤ 44) N1 breast cancers are randomly assigned to omitting IMNI group (experimental group) or with IMNI (control group) with a 1:1 ratio. The primary endpoint of this trial is event-free survival, and secondary endpoints include overall survival and locoregional recurrence-free survival. DISCUSSION: The IMNI PRECISION design allows promising clinical-genomic model to stratify the individualized risk of developing recurrence and guides the optimal RNI treatment for early-stage (pT1-2N1) BC patients. We anticipate that our results would provide high-level evidence to tailor IMNI according to individualized recurrence risk of BC. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT04517266 . Date of registration: August 18, 2020. Status: Recruiting.

Lactobacillus plantarum-derived extracellular vesicles protect against ischemic brain injury via the microRNA-101a-3p/c-Fos/TGF-ß axis.

Currently, the reported source of extracellular vesicles (EVs) for the treatment of ischemic stroke（IS）is limited to mammals. Moreover, these EVs are restricted to clinical translation by the high cost of cell culture. In this respect, Lactobacillus plantarum culture is advantaged by low cost and high yield. However, it is poorly understood whether Lactobacillus plantarum-derived EVs (LEVs) are applicable for the treatment of IS. Here, our results demonstrated that LEVs reduced apoptosis in ischemic neuron both in vivo and in vitro. As revealed by high-throughput sequencing, miR-101a-3p expression was significantly elevated by LEV treatment in OGD/R-induced neurons, as confirmed in the tMCAO mice treated with LEVs. Mechanistically, c-Fos was directly targeted by miR-101a-3p. In addition, c-Fos determined ischemia-induced neuron apoptosis in vivo and in vitro through the TGF-ß1 pathway, miR-101a-3p inhibition aggravated ischemia-induced neuron apoptosis in vitro and in vivo, and miR-101a-3p overexpression produced the opposite results. Hsa-miR-101-3p was downregulated in the plasma of patients with IS but upregulated in the patients with neurological recovery after rt-PA intravenous thrombolysis. In conclusion, Our results demonstrated for the first time that LEVs might inhibit neuron apoptosis via the miR-101a-3p/c-Fos/TGF-ß axis, and has-miR-101-3p is a potential marker of neurological recovery in IS patients.

Surfactant protein-A inhibits thymic stromal lymphopoietin-mediated T follicular helper cell differentiation and IgE production in asthma.

Lung surfactant protein A (SP-A) is critical for immunomodulation. Thymic stromal lymphopoietin (TSLP)-activated dendritic cells (DCs) drive T follicular helper (Tfh) cells differentiation in allergic asthma. We employed wild-type (WT) and SP-A-/- mice injected with TSLP and ovalbumin (OVA)-activated DCs and challenged with OVA. Compared with WT mice, we showed that allergic inflammation was dramatically increased in SP-A-/- mice. In parallel, both IL-4-producing CD45RA-CXCR5+PD-1+CD4+ cells (Tfh2) and IgE were markedly increased in SP-A-/- mice. Further study showed that SP-A prohibited TSLP activated-DCs from expressing OX40L. When we blocked OX40L-OX40 and IL-4R signaling, the differentiation of Tfh2 and IgE responses in SP-A-/- mice was significantly inhibited. In severe asthma patients, SP-A is dysfunctional in modulating the TSLP-DCs-mediated differentiation of Tfh cells. This study suggests that SP-A acts as a modulator of Tfh differentiation and IgE generation in asthma.

Inflammation accelerates copper-mediated cytotoxicity through induction of six-transmembrane epithelial antigens of prostate 4 expression.

Copper is an essential trace metal, but imbalance in copper homeostasis can induce oxidative damage. Inflammation is a fundamental element of various pulmonary diseases. Although a positive relationship between copper and chronic pulmonary diseases has been reported, the underlying reasons are still not clear. The copper level in the sputum of patients with various pulmonary diseases was measured. An inflammatory model was established to evaluate the impact of inflammation on copper uptake in the lung. We found that the level of sputum copper was increased in patients with various pulmonary diseases, especially chronic obstructive pulmonary disease and asthma. Then, we confirmed that mice with pulmonary inflammation were susceptible to copper-mediated oxidative damage because of copper overload in lung tissue. Further investigation demonstrated that interleukin (IL)-17 and tumor necrosis factor (TNF)-α exerted synergistic effects in airway epithelial cells by upregulating the expression of six-transmembrane epithelial antigens of prostate 4 (STEAP4), a metalloreductase that reduces extracellular copper ions from the cupric state to the cuprous state and facilitates copper uptake. Inhibition of STEAP4 decreased the copper uptake of cells and inhibited copper-mediated oxidative damage. Moreover, we demonstrated that the upregulation of STEAP4 by IL-17 and TNF-α was largely dependent on TNF receptor-associated factor 4 (TRAF4). Traf4-/- mice were resistant to copper-mediated oxidative damage. Our data suggest a novel IL-17/TNF-α-TRAF4-STEAP4 axis that regulates copper homeostasis.

How Does the Interval Between Completion of Adjuvant Chemotherapy and Initiation of Radiotherapy Impact Clinical Outcomes in Operable Breast Cancer Patients?

PURPOSE: The aim of this study was to evaluate the impact of time to radiotherapy (TTR) after completion of chemotherapy (CT), and TTR after surgery, in breast cancer (BC) patients. PATIENTS AND METHODS: Continuous breast cancer patients treated with surgery and CT followed by radiotherapy (RT) from 2009 through 2015 were retrospectively reviewed. Patients were categorized into four groups with respect to TTR after CT, i.e. <4, 4-8, 8-12, and >12 weeks, and TTR after surgery, i.e. <147, 147-180, 180-202, and >202 days. The Cox proportional hazards model was used to identify the independent effect of TTRs. RESULTS: Overall, 989 patients were enrolled. Patients with a TTR of >12 weeks after CT showed significantly worse breast cancer-specific survival (BCSS) and overall survival (OS) compared with those who had a TTR of <4 weeks (BCSS: hazard ratio [HR] 0.28, 95% confidence interval [CI] 0.1-0.76; OS: HR 0.33, 95% CI 0.13-0.88), 4-8 weeks (BCSS: HR 0.23, 95% CI 0.08-0.66; OS: HR 0.29, 95% CI 0.11-0.8), and 8-12 weeks (BCSS: HR 0.22, 95% CI 0.05-0.96; OS: HR 0.23, 95% CI 0.06-0.99). TTR after surgery showed no significant association with survival outcomes in the entire cohort, except in patients with hormone receptor (HR)-positive disease and those receiving mastectomy. In HR-positive tumors, a TTR after CT of >12 weeks remained an independent predictor for adverse BCSS and OS. CONCLUSION: Initiation of RT beyond 12 weeks after CT might compromise survival outcomes. Efforts should be made to avoid delaying RT, especially after completion of CT and in patients with HR-positive tumors, positive lymph nodes, and those receiving mastectomy.

IL-6 alters migration capacity of CD4+Foxp3+ regulatory T cells in systemic lupus erythematosus.

Regulatory T cells (Tregs) are impaired in human systemic lupus erythematosus (SLE) and involved in disease pathogenesis. However, the mechanisms responsible for the Treg dysfunction in SLE remain unclear. In this study, we aimed to investigate the chemotaxis of Treg response to inflammatory stimulation. Sixty two patients were enrolled, and chemokine receptors, including CCR4, CCR5, CCR6, CCR8 and CXCR3 on CD4+Foxp3+Tregs and non-Treg CD4 T cells, were analysed using FACS. The expression of CCR4 and CCR6 on Tregs of SLE patients decreased, while the expression of CCR4 on non-Treg CD4 T cells increased, as compared with those of age- and sex-matched healthy donors. In parallel, in SLE, the chemotactic capacity of non-Treg CD4 T cells response to CCR4 and CCR6 ligands dramatically increased, while that of Tregs significantly decreased. Moreover, we found that cytokines IL-6 and IL-10 positively and negatively modulate the expression of those receptors respectively. IL-6, the significantly increased cytokine in active SLE, dramatically elevated CCR4 and CCR6 expression on non-Treg CD4 T cells. However, as for Tregs, these cells produced more IL-10 than non-Treg CD4 T cells upon IL-6 stimulation, and these IL-10 led to the inhibition of CCR4 and CCR6. In sum, our data provided new evidence suggesting a functional deficiency of Tregs in SLE. It may suggest that those dysfunctional Tregs have less access to the inflammation locus to exert inhibitory capacity.

TRAF4 binds to the juxtamembrane region of EGFR directly and promotes kinase activation.

The activation of the epidermal growth factor receptor (EGFR) is crucial for triggering diverse cellular functions, including cell proliferation, migration, and differentiation, and up-regulation of EGFR expression or activity is a key factor in triggering the development of cancer. Here we show that overexpression of a scaffold protein, tumor necrosis factor receptor (TNF-R)-associated factor 4 (TRAF4), promotes EGF-induced autophosphorylation of EGFR (activation) and downstream signaling, whereas TRAF4 deficiency attenuates EGFR activation and EGF-driven cell proliferation. Using structure-based sequence alignment and NMR spectroscopy, we identified a TRAF4 binding site in the C-terminal half of the juxtamembrane (JM) segment of EGFR, a region known to promote asymmetric dimerization and subsequent activation. Deletion of the TRAF4 binding site led to dramatic defects in EGFR activation and EGF-driven cell proliferation. Specific point mutations in the TRAF4 binding site also resulted in significant attenuation of EGFR activation. Detailed structural examination of the inactive versus active forms of EGFR suggests that TRAF4 binding probably induces a conformational rearrangement of the JM region to promote EGFR dimerization. These results identify a novel mechanism of TRAF4-mediated EGFR activation and signaling.

[Relationship of Expression Between SOX11 and PAX5 in Pathological Tissue Specimens of Patients with Mantle Cell Lymphoma and Its Clinical Significance].

OBJECTIVE: Investigate the expression of SRY-related HMG box 11 (SOX11) and paired box domain 5 (PAX5) in patients with mantle cell lymphoma (MCL) and analyze the relationship between them and their clinical significance. METHODS: Seventy-six formalin-fixed paraffin-embedded (FFPE) samples of patients who were diagnosed with MCL from January 2012 to August 2017 were collected．Fifty-six FFPE samples from patients with diffuse large B cell lymphoma (DLBCL), thirty-eight FFPE samples from patients with follicular lymphoma (FL) and nine FFPE samples from patients with Burkitt's lymphoma (BL) were used as control groups. Real-time quantitative PCR (qRT-PCR) and immunohistochemistry were used to detect the mRNA and protein expressions of SOX11 and PAX5. The association between expressions of SOX11 and PAX5 in patients with MCL was analyzed. On the basis of the median H score of SOX11 and PAX5 protein expressions in patients with MCL, they were divided into high and low expression group, and the relationship between the different groups and patients' clinical characteristics and prognosis were analyzed. RESULTS: The different mRNA expression levels of SOX11 and PAX5 in different lymphoma tissues were statistically significant ( P<0.01). The mRNA expression levels of SOX11 and PAX5 in MCL group were higher than those of the control groups, and the differences of those between MCL and DLBCL or FL were statistically significant ( P<0.01). However, the differences of those between MCL and BL were not significant ( P>0.05). The expression level of SOX11 protein was also higher than those of the control groups ( P<0.000 1). However, there was no significant difference in PAX5 protein expression level between the MCL group and the control group, nor the expression levels of SOX11 and PAX5 genes and proteins among the control groups ( P>0.05). By analyzing the samples from patients with MCL, we observed a positive relevance between SOX11 and PAX5 both in mRNA expression level ( r s=0.714, P<0.000 1) and protein expression level ( G=0.407, P=0.01). There was no difference in clinical characteristics and overall survival between the high and low expression group. CONCLUSION: In MCL, there was a positive relevance between the expressions of SOX11 and PAX5. The expression of SOX11 or PAX5 alone has no significant effect on the prognostic stratification of MCL patients.

Reprograming of peripheral Foxp3+ regulatory T cell towards Th17-like cell in patients with active systemic lupus erythematosus.

Treg is essential to limit the extend and duration of the immune response, but its stability is still under debate. Here we demonstrate that IL-17-producing Treg cells (Th17-like cells) increased in peripheral blood of patients with Systemic Lupus Erythematosus (SLE). Notably, the Th17-like cells from patient with active SLE were characterized with some phenotype and function of Th17 cells. Upon stimulation, Helios-Foxp3 + CD4+ T cells decrease Foxp3 expression but increase expression of IL-17 and RORγt. Damage associated molecule pattern and inflammatory cytokines are important for induction of IL-17 expression in Treg cells. The Th17-like cells from patients with active SLE lose suppressive function and have robust response to stimulation of autoantigens. We also observed that the level of Th17-like cells in peripheral blood is closely associated with the clinical index of SLE. These findings suggest that instability of Treg plays a critical role in pathogenesis of autoimmune diseases.

The Role of the Neo-Bioscore Staging System in Guiding the Optimal Strategies for Regional Nodal Irradiation Following Neoadjuvant Treatment in Breast Cancer Patients with cN1 and ypN0-1.

BACKGROUND: The role of regional nodal irradiation (RNI) in patients with cN1 breast cancer following neoadjuvant treatment (NAT) is still controversial. The Neo-Bioscore staging system has shown promising prospect in assessing individual prognosis after NAT, and we sought to evaluate the role of Neo-Bioscore in guiding RNI following NAT. METHODS: Medical records of 163 women with cN1 and ypN0-1 disease treated with NAT between 2009 and 2014 were retrospectively reviewed and a Neo-Bioscore was assigned to each patient. Survivals were calculated using the Kaplan-Meier method and compared with the log-rank test. Multivariate analysis was used to identify independent predictors by using Cox proportional hazards models. RESULTS: The median follow-up after surgery was 59.4 months. Of all 163 patients, 119 received RNI. At surgery, 36 patients (22.1%) had pathological complete response (pCR), while 89 patients (54.6%) achieved ypN0. In the whole cohort, RNI significantly improved distant metastasis-free survival (DMFS) on multivariable analysis. In the subgroup of patients with a Neo-Bioscore of 1-3, RNI significantly improved the 5-year DMFS rate of 97.0% versus 76.9% (p = 0.002), 5-year regional node recurrence-free survival rate of 95.5% versus 76.9% (p = 0.007), and 5-year overall survival rate of 100% versus 89.2% (p = 0.005). No significant difference in outcomes was found between the RNI and non-RNI groups in patients with a score of 4-6. CONCLUSIONS: In patients with cN1 and ypN0-1, RNI was found to significantly improve DMFS following NAT. Patients with a Neo-Bioscore of 1-3 are more likely to benefit from RNI, however a large prospective study is needed to confirm this finding.

PLOD2 contributes to drug resistance in laryngeal cancer by promoting cancer stem cell-like characteristics.

BACKGROUND: Advanced stage laryngeal squamous cell carcinoma (LSCC) presents a poor prognosis; thus, there is a great need to identify novel prognostic molecular markers. Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) is thought to be a novel prognostic factor in several cancers, but its role in LSCC remains unknown. Cancer stem cells (CSCs) are responsible for most instances of tumor recurrence and the development of drug resistance and have been proven to be present in head and neck cancers. Our preliminary study indicated that PLOD2 was elevated in LSCC tissues; therefore, we hypothesized that PLOD2 is related to the prognosis of LSCC patients and aimed to explore the role and underlying mechanism of PLOD2 in LSCC. METHODS: We validated the prognostic role of PLOD2 in 114 LSCC patients by immunohistochemistry. Stable PLOD2-overexpressing Hep-2 and FaDu cells were established and assessed by molecular biology and biochemistry methods both in vitro and in vivo. RESULTS: We confirmed that PLOD2 overexpression was correlated with poor prognosis in LSCC patients. PLOD2 overexpression strengthened the CSC-like properties of Hep-2 and FaDu cells, activated the Wnt signaling pathway and conferred drug resistance in LSCC in vitro and in vivo. CONCLUSIONS: We found that PLOD2 could serve as a prognostic marker in patients with LSCC and confer drug resistance in LSCC by increasing CSC-like traits; in addition, a Wnt-responsive CSC pathway was identified.

Structural basis for receptor recognition and pore formation of a zebrafish aerolysin-like protein.

Various aerolysin-like pore-forming proteins have been identified from bacteria to vertebrates. However, the mechanism of receptor recognition and/or pore formation of the eukaryotic members remains unknown. Here, we present the first crystal and electron microscopy structures of a vertebrate aerolysin-like protein from Danio rerio, termed Dln1, before and after pore formation. Each subunit of Dln1 dimer comprises a ß-prism lectin module followed by an aerolysin module. Specific binding of the lectin module toward high-mannose glycans triggers drastic conformational changes of the aerolysin module in a pH-dependent manner, ultimately resulting in the formation of a membrane-bound octameric pore. Structural analyses combined with computational simulations and biochemical assays suggest a pore-forming process with an activation mechanism distinct from the previously characterized bacterial members. Moreover, Dln1 and its homologs are ubiquitously distributed in bony fishes and lamprey, suggesting a novel fish-specific defense molecule.

[The Value of Circulating Exsomal miR-451a to Monitor Therapy Response in Diffuse Large B Cell Lymphoma].

OBJECTIVE: To investigate the expression level of circulating exsomal miR-451a and its significances in therapy monitoring in diffuse large B cell patients. METHODS: We isolated exsomal RNAs fractions from serum of 56 DLBCL patients before treatment,during treatment and after treatment. The serum of 56 healthy controls was collected at the same time. Quantitative real time polymerase chain reaction (qRT-PCR) were performed to detected the expression level of circulating exsomal miR-451a. Receive operater characteristic (ROC) curve was performed to comfirm the diagnostic efficiency of miR-451a. Chemotherapy effect corresponding with miR-451a was analyzed. RESULTS: Circulating exsomal miR-451a was down-expression in DLBCL compared with healthy controls (P<0.000 1), and the area under the ROC curve (AUC) was 0.737 (95%CI0.645-0.816) . In 43 patients who had complete follow-up information,the patients who obtained remission,including complete remission (CR) and partial remission (PR) ,had the levels of circulating exsomal miR-451a gradually increased. While in patients who did not get remission, including stable disease (SD) and progression disease (PD) ,had no significant changes of circulating exsomal miR-451a. CONCLUSION: Circulating exsomal miR-451a may be an potential indicator for therapy response monitoring in DLBCL.

Hepatic expansion of virus-specific CD8+BTLA+ T cells with regulatory properties in chronic hepatitis B virus infection.

Similar to programmed death-1 (PD-1), B and T lymphocyte attenuator (BTLA) is a co-inhibitory molecule of the CD28 family. PD-1 is involved in T cell exhaustion during chronic viral infection. However, the role of BTLA in virus-specific T cells is poorly defined. Here we investigated the expression and function of BTLA in T cells from patients with chronic hepatitis B virus (HBV) infection. The phenotype of peripheral and intrahepatic HBV-specific T cells from 43 patients with chronic HBV infection was assessed by flow cytometry. Functional evaluation was analyzed by T cell expansion and cytokine secretion after different treatments. In chronic HBV patients, a subset of inefficient interferon-γ producing antigen-specific CD8+ T cells recruited to the liver expressed high BTLA levels. The BTLA+ HBV-specific CD8+ T cell suppressive function was antigen-specific, at least in the induction phase, because they were only activated by a pool of HBV peptides but not with a pool of unrelated peptides. Suppression of T cell responses was restored by a BTLA signaling blockade and neutralizing IL-10, indicating that BTLA signaling-mediated IL-10 secretion plays a key role in suppression. This study provides important evidence that there is a subset of liver infiltrated virus-specific CD8+BTLA+ regulatory T cells in patients with chronic HBV infection. This subset of cells plays a pivotal role in controlling hepatic effector CD8+ T cell responses through BTLA signaling mediated regulatory factor IL-10 production.

AI-2 quorum sensing negatively regulates rbf expression and biofilm formation in Staphylococcus aureus.

Staphylococcus aureus is an important pathogen that is capable of forming biofilms on biomaterial surfaces to cause biofilm-associated infections. Autoinducer 2 (AI-2), a universal language for interspecies communication, is involved in a variety of physiological activities, although its exact role in Gram-positive bacteria, especially in S. aureus, is not yet thoroughly characterized. Herein we demonstrate that inactivation of luxS, which encodes AI-2 synthase, resulted in increased biofilm formation and higher polysaccharide intercellular adhesion (PIA) production compared with the wild-type strain in S. aureus NCTC8325. The transcript level of rbf, a positive regulator of biofilm formation, was significantly increased in the luxS mutant. All of the parental phenotypes could be restored by genetic complementation and chemically synthesized 4,5-dihydroxy-2,3-pentanedione, the AI-2 precursor molecule, suggesting that AI-2 has a signaling function to regulate rbf transcription and biofilm formation in S. aureus. Phenotypic analysis revealed that the luxS rbf double mutant produced approximately the same amount of biofilms and PIA as the rbf mutant. In addition, real-time quantitative reverse transcription-PCR analysis showed that the icaA transcript level of the rbf mutant was similar to that of the luxS rbf double mutant. These findings demonstrate that the LuxS/AI-2 system regulates PIA-dependent biofilm formation via repression of rbf expression in S. aureus. Furthermore, we demonstrated that Rbf could bind to the sarX and rbf promoters to upregulate their expression.