Xianzhu Xia’s research while affiliated with Shantou University and other places

Objectives Formyl peptide receptor 1 (FPR1) is a member of G protein-coupled receptor (GPCR) family that detects potentially danger signals characterized by the appearance of N-formylated peptides which originate from either bacteria or host mitochondria during organ injury, including sepsis. Mitochondrial-encoded NADH dehydrogenase 6 (MT-ND6) and Annexin A1 (ANXA1), as mitochondrial damage-associated molecular patterns (mtDAMPs) agonist and endogenous agonist of FPR1 respectively, interact with FPR1 regulating polymorphonuclear leukocytes (PMNs) function and inflammatory response during sepsis. However, there is no direct evidence of MT-ND6 or ANXA1 in the circulation of patients with sepsis and their potential role in clinical significance, including diagnosis and mortality prediction during sepsis. Methods A prospective cohort study was conducted in ICU within a large academic hospital. We measured serum MT-ND6 or ANXA1 in a cohort of patients with sepsis in ICU (n=180) and patients with non-sepsis in ICU (n=60) by Enzyme-linked immunosorbent assays (ELISA). The ROC curve and Kaplan Meier analysis was used to evaluate the diagnostic and prognostic ability of two biomarkers for patients with sepsis. Results The concentration of MT-ND6 and ANXA1 were significantly elevated in the patients with sepsis, and the diagnostic values of MT-ND6 (0.789) for sepsis patients was second only to SOFA scores (AUC = 0.870). Higher serum concentrations of MT-ND6 (>1.41 ng/ml) and lower concentrations of ANXA1 (< 8.09 ng/mL) were closely related to the higher mortality in patients with sepsis, with the predictive values were 0.705 and 0.694, respectively. When patients with sepsis classified based on four pro-inflammation and two anti-inflammation cytokines, it was shown that combination of MT-ND6 and ANXA1 obviously improved the predictive values in the septic patients with mixed hyperinflammation or immunosuppression phenotypes. Conclusion Our findings provide valuable models testing patient risk prediction and strengthen the evidence for agonists of FPR1, MT-ND6 and ANXA1, as novel biomarker for patient selection for novel therapeutic agents to target mtDAMPs and regulator of GPCRs in sepsis.

Objective This study aimed to investigate the proportion of neutrophils among leukocytes, in various tissues following intraperitoneal injection of Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) in mice. Methods Twelve specific-pathogen free (SPF) male mice, aged eight weeks, were segregated into three groups, each containing four mice. Two of these groups were subjected to intraperitoneal injections of E. coli and S. aureus, both in high concentrations, to establish mouse models of inflammation. The remaining group, which received an intraperitoneal injection of phosphate buffered saline (PBS), served as the control group. Observe the mice every half hour. Then mice were anesthetized, and samples from peripheral blood, liver, and brain tissues were carefully collected nearing death. These samples underwent a digestion process to produce single-cell suspensions. Subsequently, these suspensions were stained with fluorescent antibodies targeting CD45, Ly6G, and CD11b. A flow cytometric analyzer was then employed to enumerate and compare the neutrophil alterations across each group (Fig. 1). Results The results indicated a significant variation in the ratio of CD11b⁺ Ly6G⁺ neutrophils to CD45⁺ leukocytes among the groups. In peripheral blood, the control group showed a neutrophil proportion of approximately 1.44 %, while the E. coli and S. aureus groups exhibited increased proportions of 6.53 % and 3.82 %, respectively. In liver tissue, a marked elevation was observed in the experimental groups, with ratios of 19.20 % and 20.40 % for E. coli and S. aureus, respectively, compared to 1.64 % in the control. In brain tissue, the increments were more modest but noticeable, with the experimental groups showing 2.40 % and 1.11 % in contrast to 0.13 % in the control group. Conclusions These findings suggest neutrophils are involved in the response after intraperitoneal injection of E. coli and S. aureus, with marked differences in neutrophil responses in different tissues. This study enhances our understanding of the acute inflammatory response to bacterial infection.

Background Understanding the mechanism behind sepsis-associated encephalopathy (SAE) remains a formidable task. This study endeavors to shed light on the complex cellular and molecular alterations that occur in the brains of a mouse model with SAE, ultimately unraveling the underlying mechanisms of this condition. Methods We established a murine model using intraperitoneal injection of lipopolysaccharide (LPS) in wild type and Anxa1−/− mice and collected brain tissues for analysis at 0-hour, 12-hour, 24-hour, and 72-hour post-injection. Utilizing advanced techniques such as single-nucleus RNA sequencing (snRNA-seq) and Stereo-seq, we conducted a comprehensive characterization of the cellular responses and molecular patterns within the brain. Results Our study uncovered notable temporal differences in the response to LPS challenge between Anxa1−/− (annexin A1 knockout) and wild type mice, specifically at the 12-hour and 24-hour time points following injection. We observed a significant increase in the proportion of Astro-2 and Micro-2 cells in these mice. These cells exhibited a colocalization pattern with the vascular subtype Vas-1, forming a distinct region known as V1A2M2, where Astro-2 and Micro-2 cells surrounded Vas-1. Moreover, through further analysis, we discovered significant upregulation of ligands and receptors such as Timp1-Cd63, Timp1-Itgb1, Timp1-Lrp1, as well as Ccl2-Ackr1 and Cxcl2-Ackr1 within this region. In addition, we observed a notable increase in the expression of Cd14-Itgb1, Cd14-Tlr2, and Cd14-C3ar1 in regions enriched with Micro-2 cells. Additionally, Cxcl10-Sdc4 showed broad upregulation in brain regions containing both Micro-2 and Astro-2 cells. Notably, upon LPS challenge, there was an observed increase in Anxa1 expression in the mouse brain. Furthermore, our study revealed a noteworthy increase in mortality rates following Anxa1 knockdown. However, we did not observe substantial differences in the types, numbers, or distribution of other brain cells between Anxa1−/− and wildtype mice over time. Nevertheless, when comparing the 24-hour post LPS injection time point, we observed a significant decrease in the proportion and distribution of Micro-2 and Astro-2 cells in the vicinity of blood vessels in Anxa1−/− mice. Additionally, we noted reduced expression levels of several ligand-receptor pairs including Cd14-Tlr2, Cd14-C3ar1, Cd14-Itgb1, Cxcl10-Sdc4, Ccl2-Ackr1, and Cxcl2-Ackr1. Conclusions By combining snRNA-seq and Stereo-seq techniques, our study successfully identified a distinctive cellular colocalization, referred to as a special pathological niche, comprising Astro-2, Micro-2, and Vas-1 cells. Furthermore, we observed an upregulation of ligand-receptor pairs within this niche. These findings suggest a potential association between this cellular arrangement and the underlying mechanisms contributing to SAE or the increased mortality observed in Anxa1 knockdown mice.

Introduction. Inappropriate use of antibiotics and inadequate therapeutic regimens for early-stage pulmonary infections are major contributors to increased prevalence of complications and mortality. Moreover, due to the limitations in sensitivity of conventional testing, there is an urgent need for more diagnostically efficient methods for the detection and characterization of pathogens in pulmonary infections. Hypothesis/Gap Statement. Metagenomic next-generation sequencing (mNGS) can contribute to the diagnosis and management of pulmonary infections. Aim. This study aimed to evaluate the clinical application and value of mNGS in the diagnosis of clinically suspected pulmonary infections by comparing with conventional testing. Methodology. In this study, the diagnosis performance of mNGS was evaluated using bronchoalveolar lavage fluid (BALF) samples from 143 patients with suspected lung infections. First, we conducted a prospective study on 31 patients admitted to Yuebei People’s Hospital Affiliated to Shantou University Medical College to investigate the clinical value. Then a retrospective analysis was performed by including more patients ( n =112) to reduce the random error. Pathogens were detected by mNGS and conventional methods (culture and PCR). Then, the types and cases of detected pathogens, as well as the specificity and sensitivity, were compared between the two methods. We evaluated the performance of mNGS in detecting bacterial, fungal, viral and mixed infections in BALF. The effect of disease severity in pulmonary infections on the integrity of mNGS pathogen detection was also explored. Results. The mNGS provided an earlier and more comprehensive pathogen profile than conventional testing, which in turn prompted a change in clinical medication, which led to improvement in eight patients (8/31=25.81 %) in the presence of other serious comorbidities. In a retrospective analysis, mNGS was much more sensitive than conventional testing in the diagnosis of pulmonary infections (95.33 % vs. 55.56 %; P <0.001), with a 39.77 % increase in sensitivity. The detection rate of mNGS for mixed infections was significantly higher than that of conventional testing methods for both common and severe pneumonia (48/67=71.64 % vs. 12/52=23.08 %, P <0.001; 44/59=74.58 % vs. 11/59=18.64 %, P <0.0001). Conclusion. The sensitivity of mNGS in the diagnosis of pathogenic microorganisms in pulmonary infections far exceeds that of conventional culture tests. As a complementary method to conventional methods, mNGS can help improve the diagnosis of pulmonary infections. In addition, mNGS pathogen integrity detection rate was similar in common and severe pneumonia. We recommend the prompt use of mNGS when mixed or rare pathogen infections are suspected, especially in immunocompromised individuals and/or critically ill individuals.

Three or four intramuscular doses of the inactivated human rabies virus vaccines are needed for pre- or post- exposure prophylaxis in humans. This procedure has made a great contribution to prevent human rabies deaths, which bring huge economic burdens in developing countries. Herein, a recombinant adeno-associated virus serotype 9, AAV9-RABVG, harboring a RABV G gene, was generated to serve as a single dose rabies vaccine candidate. The RABV G protein was stably expressed in the 293T cells infected with AAV9-RABVG. A single dose of 2×1011 v.p. of AAV9-RABVG induced robust and long term positive seroconversions in BALB/c mice with a 100% survival from a lethal RABV challenge. In Cynomolgus Macaques vaccinated with a single dose of 1×1013 v.p. of AAV9-RABVG, the titers of rabies VNAs increased remarkably from 2 weeks after immunity, and maintained over 31.525 IU/ml at 52 weeks. More DCs were activated significantly for efficient antigen presentations of RABV G protein, and more B cells were activated to responsible for antibody responses. Significantly more RABV G specific IFN-γ-secreting CD4+ and CD8+ T cells, and IL-4-secreting CD4+ T cells were activated, and significantly higher levels of IL-2, IFN-γ, IL-4 and IL-10 were secreted to aid immune responses. Overall, the AAV9-RABVG was a single dose rabies vaccine candidate with great promising by inducing robust, long term humoral responses and both Th1 and Th2 cell-mediated immune responses in mice and non-human primates.

Ebola virus (EBOV), one of the deadliest viruses, is the cause of fatal Ebola virus disease (EVD). The underlying mechanism of viral replication and EBOV-related hemorrhage is not fully understood. Here, we show that EBOV VP35, a cofactor of viral RNA-dependent RNA polymerase, binds human A kinase interacting protein (AKIP1), which consequently activates protein kinase A (PKA) and the PKA-downstream transcription factor CREB1. During EBOV infection, CREB1 is recruited into EBOV ribonucleoprotein complexes in viral inclusion bodies (VIBs) and employed for viral replication. AKIP1 depletion or PKA-CREB1 inhibition dramatically impairs EBOV replication. Meanwhile, the transcription of several coagulation-related genes, including THBD and SERPINB2, is substantially upregulated by VP35-dependent CREB1 activation, which may contribute to EBOV-related hemorrhage. The finding that EBOV VP35 hijacks the host PKA-CREB1 signal axis for viral replication and pathogenesis provides novel potential therapeutic approaches against EVD.

Coronaviruses are commonly characterized by a unique discontinuous RNA transcriptional synthesis strategy guided by transcription-regulating sequences (TRSs). However, the details of RNA synthesis in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have not been fully elucidated. Here, we present a time-scaled, gene-comparable transcriptome of SARS-CoV-2, demonstrating that ACGAAC functions as a core TRS guiding the discontinuous RNA synthesis of SARS-CoV-2 from a holistic perspective. During infection, viral transcription, rather than genome replication, dominates all viral RNA synthesis activities. The most highly expressed viral gene is the nucleocapsid gene, followed by ORF7 and ORF3 genes, while the envelope gene shows the lowest expression. Host transcription dysregulation keeps exacerbating after viral RNA synthesis achieves to the top. The most enriched host pathways are metabolism-related. Two of them (cholesterol and valine metabolism) affect viral replication in reverse. Furthermore, the activation of numerous cytokines emerges before the large-scale viral RNA synthesis. Importance SARS-CoV-2 is responsible for the current severe global health emergency that began at the end of 2019. Although the universal transcriptional strategies of coronaviruses are preliminarily understood, the details of RNA synthesis, especially the time-matched transcription level of each SARS-CoV-2 gene and the principles of subgenomic mRNA synthesis, are not clear. The coterminal subgenomic mRNAs of SARS-CoV-2 present obstacles in identifying the expression of most genes by PCR-based methods, which is exacerbated by the lack of related antibodies. Moreover, SARS-CoV-2-related metabolic imbalance and cytokine storm are receiving increasing attention from both clinical and mechanistic perspectives. Our transcriptomic research provides information on both viral RNA synthesis and host responses, in which the transcription-regulating sequences and transcription levels of viral genes are demonstrated, and the metabolic dysregulation and cytokine levels identified at the host cellular level support the development of novel medical treatment strategies.

Cystic echinococcosis (CE), caused by Echinococcus granulosus (E.granulosus), is a zoonosis with a worldwide distribution, resulting in heavy impact to public health and social economics. In this study, we generated a recombinant RABV expressing EG95 protein of E.granulosus (LBNSE-EG95) as a bivalent candidate vaccine for use in sheep and cattle against CE and rabies, which is another severe health threat in CE-endemic areas. It was found that EG95 was successfully expressed without altering the pathogenicity of parent LBNSE vector. Further study showed that LBNSE-EG95 immunization in mice elicited activation of dendric cells (DCs) and B cells, and induced Th1-/Th2-mediated cellular immune responses, leading to robust production of RABV neutralizing antibodies and high level of EG95-sepecific antibodies with more than 90% protection against CE. In addition, single dose of LBNSE-EG95 conferred full protection against lethal RABV challenge in mice. Collectively, these results suggest that the recombinant LBNSE-EG95 has potential to be developed as an efficient bivalent vaccine for sheep and cattle use in endemic areas of CE and rabies. This article is protected by copyright. All rights reserved