Salivary factor LTRIN from Aedes aegypti facilitates the transmission of Zika virus by interfering with the lymphotoxin-ß receptor.

Pathogens have co-evolved with mosquitoes to optimize transmission to hosts. Mosquito salivary-gland extract is known to modulate host immune responses and facilitate pathogen transmission, but the underlying molecular mechanisms of this have remained unknown. In this study, we identified and characterized a prominent 15-kilodalton protein, LTRIN, obtained from the salivary glands of the mosquito Aedes aegypti. LTRIN expression was upregulated in blood-fed mosquitoes, and LTRIN facilitated the transmission of Zika virus (ZIKV) and exacerbated its pathogenicity by interfering with signaling through the lymphotoxin-ß receptor (LTßR). Mechanically, LTRIN bound to LTßR and 'preferentially' inhibited signaling via the transcription factor NF-κB and the production of inflammatory cytokines by interfering with the dimerization of LTßR during infection with ZIKV. Furthermore, treatment with antibody to LTRIN inhibited mosquito-mediated infection with ZIKV, and abolishing LTßR potentiated the infectivity of ZIKV both in vitro and in vivo. This study provides deeper insight into the transmission of mosquito-borne diseases in nature and supports the therapeutic potential of inhibiting the action of LTRIN to disrupt ZIKV transmission.

Isoliquiritigenin limits inflammasome activation of macrophage via docking into Syk to alleviate murine non-alcoholic fatty liver disease.

Isoliquiritigenin (ISL) is a chalcone-type flavonoid derived from the root of licorice with antioxidant, anti-inflammatory, anti-tumour and neuroprotective properties. ISL has been proven to downregulate the productions of IL-1ß, TNF-α and IL-6 by macrophages. However, detailed molecular mechanisms of this modulation remain elusive. Here, ISL suppressed Syk phosphorylation and CD80, CD86, IL-1ß, TNF-α and IL-6 expressions in lipopolysaccharide-stimulated macrophages ex vivo. ApoC3-transgenic (ApoC3TG) mice had more activated macrophages. ISL was also able to downregulate the inflammatory activities of macrophages from ApoC3TG mice. Administration of ISL inhibited Syk activation and inflammatory activities of macrophages in ApoC3TG mice in vivo. The treatment of ISL further alleviated MCD-induced non-alcoholic fatty liver disease (NAFLD) in wild-type and ApoC3TG mice, accompanied by less recruitment and activation of liver macrophages. Due to the inhibition of Syk phosphorylation, ISL-treated macrophages displayed less production of cytoplasmic ROS, NLRP3, cleaved-GSDMD and cleaved-IL-1ß, suggesting less inflammasome activation. Finally, the molecular docking study demonstrated that ISL bound to Syk directly with the Kd of 1.273 × 10-8 M. When the Syk expression was knocked down by its shRNA, the inhibitory effects of ISL on activated macrophages disappeared, indicating that Syk was at least one of key docking-molecules of ISL. Collectively, ISL could alleviate MCD-induced NAFLD in mice involved with the inhibition of macrophage inflammatory activity by the blockade of Syk-induced inflammasome activation.

Macrophage-Targeted Isoniazid-Selenium Nanoparticles Promote Antimicrobial Immunity and Synergize Bactericidal Destruction of Tuberculosis Bacilli.

Pathogenesis hallmarks for tuberculosis (TB) are the Mycobacterium tuberculosis (Mtb) escape from phagolysosomal destruction and limited drug delivery into infected cells. Several nanomaterials can be entrapped in lysosomes, but the development of functional nanomaterials to promote phagolysosomal Mtb clearance remains a big challenge. Here, we report on the bactericidal effects of selenium nanoparticles (Se NPs) against Mtb and further introduce a novel nanomaterial-assisted anti-TB strategy manipulating Ison@Man-Se NPs for synergistic drug-induced and phagolysosomal destruction of Mtb. Ison@Man-Se NPs preferentially entered macrophages and accumulated in lysosomes releasing Isoniazid. Surprisingly, Ison@Man-Se/Man-Se NPs further promoted the fusion of Mtb into lysosomes for synergistic lysosomal and Isoniazid destruction of Mtb. Concurrently, Ison@Man-Se/Man-Se NPs also induced autophagy sequestration of Mtb, evolving into lysosome-associated autophagosomal Mtb degradation linked to ROS-mitochondrial and PI3K/Akt/mTOR signaling pathways. This novel nanomaterial-assisted anti-TB strategy manipulating antimicrobial immunity and Mtb clearance may potentially serve in more effective therapeutics against TB and drug-resistant TB.

PD-L1 expression and prognosis in definitive radiotherapy patients with neuroendocrine cervical carcinoma.

Background: Neuroendocrine carcinoma of the cervix (NECC) is more prone to lymphatic infiltration, lymph node involvement, local recurrence, and distant metastasis. Using concurrent chemoradiotherapy (CCRT) with or without adjuvant chemotherapy as the standard treatment for locally advanced NECCs and CCRT for patients with early lesions confined to the cervix. However, the prognosis of NECC patients treated with definitive radiotherapy (RT) is unknown. Immune checkpoint inhibitors are a promising therapeutic strategy for locally advanced cervical cancer. Some reports suggest that the expression of PD-L1 in solid tumors correlates with prognosis. Aim: This study investigates prognostic factors for survival in patients with neuroendocrine cervical carcinoma (NECC) treated with definitive RT and the relationship between PD-L1 expression and prognosis in these patients. Methods: This retrospective study included 66 patients with histologically confirmed NECC who received RT with or without chemotherapy. From January 2015 to December 2020, patients received routine extended-field irradiation (EFI), and PD-L1 expression was assessed by immunohistochemistry. The most commonly used chemotherapy agents were etoposide-platinum and paclitaxel-platinum. Results: PD-L1 expression was positive in 17 of 45 (37.8%) patients. There were 52 cases of pure NECC and 14 cases of mixed carcinoma. Sixty stage IB-III patients received definitive RT. The 3- and 5-year progression-free survival (PFS) was 39.8% and 34.1%, and 3- and 5-year overall survival (OS) was 48.0% and 40.2%, respectively. There was no significant difference in 3 and 5-year PFS and 3 and 5-year OS between patients with pure and mixed carcinoma. Positive PD-L1 expression was associated with higher 3-year PFS in patients with mixed histology. Univariate analysis showed that lymph node metastasis (LNM) and the International Federation of Gynecology and Obstetrics stages predicted 3- and 5-year PFS in patients who received definitive RT. The median OS in patients receiving less than four cycles and at least four cycles of chemotherapy (CT) was 26.0 and 44.0 months, respectively (P = 0.038); moreover, 3- and 5-year PFS was 34.1% and 25.7% in the former and 46.4% and 40.4% in the latter. There were no significant differences in OS and PFS between pelvic irradiation and prophylactic EFI in patients treated with definitive RT. There were no significant differences in para-aortic failure rate after concurrent chemoradiotherapy between patients who underwent pelvic irradiation or prophylactic EFI (P = 0.147). Conclusion: In patients with mixed NECC, positive PD-L1 expression is correlated with higher 3-year PFS. Chemoradiotherapy was effective for NECCs. The LNM and stage predicted PFS. Four or more cycles of chemotherapy improve prognosis. Prophylactic EFI did not significantly improve PFS and OS. Relevance for Patients: This study is relevant to patients as it confirms that chemoradiotherapy is effective for both early and locally advanced NECC and that four or more cycles of chemotherapy improved prognosis. The regimen should be carefully evaluated to ensure that patients receive the most effective radiation therapy for the prophylactic of para-aortic LNM. Potential risk factors for the recurrence of radical radiotherapy should be fully understood to minimize these risks. This study observed that PD-L1 expression positive in patients with mixed NECC types is correlated with higher 3-year PFS.

Advances of MnO2 nanomaterials as novel agonists for the development of cGAS-STING-mediated therapeutics.

As an essential micronutrient, manganese plays an important role in the physiological process and immune process. In recent decades, cGAS-STING pathway, which can congenitally recognize exogenous and endogenous DNA for activation, has been widely reported to play critical roles in the innate immunity against some important diseases, such as infections and tumor. Manganese ion (Mn2+) has been recently proved to specifically bind with cGAS and activate cGAS-STING pathway as a potential cGAS agonist, however, is significantly restricted by the low stability of Mn2+ for further medical application. As one of the most stable forms of manganese, manganese dioxide (MnO2) nanomaterials have been reported to show multiple promising functions, such as drug delivery, anti-tumor and anti-infection activities. More importantly, MnO2 nanomaterials are also found to be a potential candidate as cGAS agonist by transforming into Mn2+, which indicates their potential for cGAS-STING regulations in different diseased conditions. In this review, we introduced the methods for the preparation of MnO2 nanomaterials as well as their biological activities. Moreover, we emphatically introduced the cGAS-STING pathway and discussed the detailed mechanisms of MnO2 nanomaterials for cGAS activation by converting into Mn2+. And we also discussed the application of MnO2 nanomaterials for disease treatment by regulating cGAS-STING pathway, which might benefit the future development of novel cGAS-STING targeted treatments based on MnO2 nanoplatforms.

Potentiality of Exosomal Proteins as Novel Cancer Biomarkers for Liquid Biopsy.

Liquid biopsy has been rapidly developed in recent years due to its advantages of non-invasiveness and real-time sampling in cancer prognosis and diagnosis. Exosomes are nanosized extracellular vesicles secreted by all types of cells and abundantly distributed in all types of body fluid, carrying diverse cargos including proteins, DNA, and RNA, which transmit regulatory signals to recipient cells. Among the cargos, exosomal proteins have always been used as immunoaffinity binding targets for exosome isolation. Increasing evidence about the function of tumor-derived exosomes and their proteins is found to be massively associated with tumor initiation, progression, and metastasis in recent years. Therefore, exosomal proteins and some nucleic acids, such as miRNA, can be used not only as targets for exosome isolation but also as potential diagnostic markers in cancer research, especially for liquid biopsy. This review will discuss the existing protein-based methods for exosome isolation and characterization that are more appropriate for clinical use based on current knowledge of the exosomal biogenesis and function. Additionally, the recent studies for the use of exosomal proteins as cancer biomarkers are also discussed and summarized, which might contribute to the development of exosomal proteins as novel diagnostic tools for liquid biopsy.

Application Value of Magnetic Resonance Radiomics and Clinical Nomograms in Evaluating the Sensitivity of Neoadjuvant Chemotherapy for Nasopharyngeal Carcinoma.

OBJECTIVE: To predict the sensitivity of nasopharyngeal carcinoma (NPC) to neoadjuvant chemotherapy (NACT) based on magnetic resonance (MR) radiomics and clinical nomograms prior to NACT. MATERIALS AND METHODS: From January 2014 to July 2015, 284 consecutive patients with pathologically confirmed NPC underwent 3.0 T MR imaging (MRI) before initiating NACT. The patients' data were randomly assigned to a training set (n = 200) or a test set (n = 84) at a ratio of 7:3. The clinical data included sex, tumor (T) stage, lymph node (N) stage, American Joint Committee on Cancer (AJCC) stage, and the plasma concentration of Epstein-Barr virus (EBV) DNA. The regions of interest (ROI) were manually segmented on the axial T2-weighted imaging (T2WI) and enhanced T1-weighted imaging (T1WI) sequences using ITK-SNAP software. The radiomics data were post-processed using AK software. Moreover, the Maximum Relevance Minimum Redundancy (mRMR) algorithm and the Least Absolute Shrinkage and Selection Operator (LASSO) were adopted for dimensionality reduction to screen for the features that best predicted the treatment efficacy, and clinical risk factors were used in combination with radiomics scores (Rad-scores) to construct the clinical radiomics-based nomogram. DeLong's test was utilized to compare the area under the curve (AUC) values of the clinical radiomics-based nomogram, radiomics model, and clinical nomogram. Decision curve analysis (DCA) was employed to evaluate each model's net benefit. RESULTS: The clinical nomogram was constructed based on data from patients who were randomly assigned according to T2WI and enhanced T1WI sequences. In the training set, the T2WI sequence-based clinical radiomics nomogram and the radiomics model outperformed the clinical nomogram in predicting the NACT efficacy (AUC, 0.81 vs. 0.60, p = 0.001279 and 0.76 vs. 0.60, p = 0.03026). These findings were well-verified in the test set. The enhanced T1WI sequence-based clinical radiomics nomogram exhibited better performance in predicting treatment efficacy than the clinical nomogram (AUC, 0.79 vs. 0.62, respectively; p = 0.0000834). The DCA revealed that the T2WI and clinical radiomics-based nomograms resulted in a net benefit in predicting the NACT efficacy. CONCLUSION: The clinical radiomics-based nomogram improved the prediction of NACT efficacy, with the T2WI sequence-based clinical radiomics achieving the best effect.

Image quality stability of whole-body diffusion weighted imaging.

OBJECTIVE: To assess the reproducibility of whole-body diffusion weighted imaging (WB-DWI) technique in healthy volunteers under normal breathing with background body signal suppression. METHODS: WB-DWI was performed on 32 healthy volunteers twice within two-week period using short TI inversion-recovery diffusion-weighted echo-planar imaging sequence and built-in body coil. The volunteers were scanned across six stations continuously covering the entire body from the head to the feet under normal breathing. The bone apparent diffusion coefficient (ADC) and exponential ADC (eADC) of regions of interest (ROIs) were measured. We analyzed correlation of the results using paired-t-test to assess the reproducibility of the WB-DWI technique. RESULTS: We were successful in collecting and analyzing data of 64 WB-DWI images. There was no significant difference in bone ADC and eADC of 824 ROIs between the paired observers and paired scans (P>0.05). Most of the images from all stations were of diagnostic quality. CONCLUSION: The measurements of bone ADC and eADC have good reproducibility. WB-DWI technique under normal breathing with background body signal suppression is adequate.

Mannosylated graphene oxide as macrophage-targeted delivery system for enhanced intracellular M.tuberculosis killing efficiency.

Tuberculosis (TB), caused by M.tuberculosis (Mtb), has become a top killer among infectious diseases. Enhancing the ability of anti-TB drugs to kill intracellular Mtb in host cells remains a big challenge. Here, an innovative nano-system was developed to increase drug delivery and Mtb-killing efficacy in Mtb-infected macrophages. We employed mannose surface decoration to develop mannosylated and PEGylated graphene oxide (GO-PEG-MAN). Such nano-platform exhibited increased uptake by macrophages via mannose receptor-mediated endocytosis in vitro. Interestingly, drug-loaded GO-PEG-MAN was preferentially up-taken by mannose receptor-expressing mucosal CD14+ macrophages isolated from Mtb-infected rhesus macaques than drug-loaded GO-PEG. Consistently, the drug concentration was also significantly higher in macrophages than that in T and B cells expressing no or low mannose receptor, implicating a useful macrophage/mannose receptor-targeted drug-delivery system relevant to the in vivo settings. Concurrently, rifampicin-loaded GO-PEG-MAN (Rif@GO-PEG-MAN) significantly increased rifampicin uptake, inducing long-lasting higher concentration of rifampicin in macrophages. Such innovative Rif@GO-PEG-MAN could readily get into the lysosomes of the Mtb host cells, where rifampicin underwent an accelerated release in acidic lysosomic condition, leading to explosive rifampicin release after cell entry for more effective killing of intracellular Mtb. Most importantly, Rif@GO-PEG-MAN-enhanced intracellular rifampicin delivery and pharmacokinetics significantly increased the efficacy of rifampicin-driven killing of intracellular BCG and Mtb bacilli in infected macrophages both in vitro and ex vivo. Such innovative nanocarrier approach may potentially enhance anti-TB drug efficacy and reduce drug side effects.

Type I IFN operates pyroptosis and necroptosis during multidrug-resistant A. baumannii infection.

Multidrug-resistant Acinetobacter baumannii, a common pathogen responsible for nosocomial infections, is the main cause for outbreaks of infectious diseases, such as pneumonia, meningitis, and bacteremia, especially among critically ill patients. Epidemic A. baumannii is a growing public health concern as it is resistant to all existing antimicrobial agents, thereby necessitating the development of new therapeutic approaches to mount an effective immune response against this bacterial pathogen. In this study, we identified a critical role for type I interferon (IFN) in epigenetic regulation during A. baumannii infection and established a central role for it in multiple cell death pathways. A. baumannii infection induced mixed cell death constituted of apoptosis, pyroptosis, and necroptosis. Mechanically, A. baumannii triggered TRIF-dependent type I IFN production, which in turn induced the expression of genes Zbp1, Mlkl, caspase-11, and Gsdmd via KAT2B-mediated and P300-mediated H3K27ac modification, leading to NLRP3 inflammasome activation, and potentially contributed to GSDMD-mediated pyroptosis and MLKL-dependent necroptosis. Our study offers novel insights into the mechanisms of type I IFN and provides potential therapeutic targets for infectious and inflammatory diseases.

HECTD3 mediates TRAF3 polyubiquitination and type I interferon induction during bacterial infection.

Lysine-63-linked (K63-linked) polyubiquitination of TRAF3 coordinates the engagement of pattern-recognition receptors with recruited adaptor proteins and downstream activator TBK1 in pathways that induce type I IFN. Whether autoubiquitination or other E3 ligases mediate K63-linked TRAF3 polyubiquitination remains unclear. We demonstrated that mice deficient in the E3 ligase gene Hectd3 remarkably increased host defense against infection by intracellular bacteria Francisella novicida, Mycobacterium, and Listeria by limiting bacterial dissemination. In the absence of HECTD3, type I IFN response was impaired during bacterial infection both in vivo and in vitro. HECTD3 regulated type I IFN production by mediating K63-linked polyubiquitination of TRAF3 at residue K138. The catalytic domain of HECTD3 regulated TRAF3 K63 polyubiquitination, which enabled TRAF3-TBK1 complex formation. Our study offers insights into mechanisms of TRAF3 modulation and provides potential therapeutic targets against infections by intracellular bacteria and inflammatory diseases.

Cell-Based High-Throughput Screening Assay Identifies 2',2'-Difluoro-2'-deoxycytidine Gemcitabine as a Potential Antipoliovirus Agent.

As we approach the global eradication of circulating wild-type polioviruses (PV), vaccination with oral poliovirus vaccine (OPV) has led to the emergence of circulating vaccine-derived poliovirus (cVDPV) and vaccine-associated paralytic poliomyelitis (VAPP). Complete cessation of all poliovirus infections may require stopping use of OPV and formulating improved vaccines and new antiviral drugs. Currently, no licensed drugs are available to treat chronically infected poliovirus excretors. Here, we created a modified PV expressing Gaussia Luciferase (Sb-Gluc) and developed a cell-based high-throughput screening (HTS) antiviral assay. Using the validated HTS assay, we screened the FDA-approved drug library of compounds and identified candidate agents capable of inhibiting PV replication. We then characterized antipoliovirus activity for the best hit, gemcitabine, a nucleoside analogue used in tumor chemotherapy. We found that gemcitabine inhibited PV Mahoney replication with an IC50 of 0.3 µM. It completely protected HeLa cells from PV-induced cytopathic effects at 25 µM, without detectable toxicity for cell viability. Furthermore, a gemcitabine metabolite directly inhibited the ability of PV RNA polymerase to synthesize or elongate PV RNA. Because PV RNA polymerase is somehow conserved among species in the Picornaviridae family, gemcitabine may be further developed as an attractive broad-spectrum antiviral for PV and others.

Optimization of magnetic resonance sequences in lymph node staging of nasopharyngeal carcinoma.

BACKGROUND: Detection rate of retropharyngeal lymph node metastasis in patients with nasopharyngeal carcinoma (NPC) needs to be improved. The purpose of this study was to compare three magnetic resonance (MR) sequences for detecting lymph nodes in patients with NPC. METHODS: Between July 2007 and March 2008, MR staging of pre-treated tumor was conducted on 120 patients with pathologically confirmed NPC. The outcome of three different sequences for MR NPC staging were compared: coronal short TI inversion recovery (STIR), axial proton density fat-suppressed (PDWI fs), and coronal contrast enhanced fast spin echo T1 weighted fat-suppressed (CE FSE T1WI fs). Nodal classification method (1999) was applied to count the number of retropharyngeal and cervical lymph nodes discovered by each MR sequence. Paired t tests were used for statistical analysis. RESULTS: A total of 2575 lymph nodes were found using coronal STIR sequence; 1816 lymph nodes for coronal CE FSE T1WI fs sequence and 2638 lymph nodes for axial PDWI fs sequence. Significant differences existed in the number of lymph nodes detected by axial PDWI fs and coronal CE FSE T1WI fs sequence (paired t test, P < 0.05), with the former sequence getting higher numbers. Statistical differences also existed between coronal STIR and coronal CE FSE T1WI fs sequence (paired t test, P < 0.05), with the former sequence getting higher numbers. No significant difference was found between coronal STIR sequence and axial PDWI fs sequence (paired t test, P > 0.05). CONCLUSIONS: For the detection of retropharyngeal and cervical lymph nodes, coronal STIR sequence and axial PDWI fs sequence have similar performance and both sequences showed better detection than CE FSE T1WI fs sequence. Furthermore, by combining coronal STIR sequence and axial PDWI fs sequence, we can improve the detection of lymph nodes in NPC N-staging before treatment, especially for lymph nodes located in the thoracic entrance.