Biology subjects

Hao, H.

Publications and source records attributed to Hao, H..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

B cell c-Maf signaling promotes tumor progression in animal models of pancreatic cancer and melanoma

BackgroundThe role of B cells in anti-tumor immunity remains controversial, with studies suggesting the pro-tumor and anti-tumor activity. This controversy may be due to the heterogeneity in B cell populations, as the balance among the subtypes may impact tumor progression. The immunosuppressive regulatory B cells (Breg) release IL-10 but only represent a minor population. Additionally, tumor-specific antibodies (Ab) also exhibit anti-tumor and pro-tumor function dependent on the Ab isotype. Transcription factor c-Maf has been suggested to contribute to the regulation of IL-10 in Breg, but the role of B cell c-Maf signaling in anti-tumor immunity and regulating antibody responses remain unknown. MethodsConditional B cell c-Maf knockout (KO) and control mice were used to establish a KPC pancreatic cancer model and B16.F10 melanoma model. Tumor progression was evaluated. B cell and T cell phenotypes were determined by flow cytometry, mass cytometry, and cytokine/chemokine profiling. Differentially expressed genes in B cells were examined by using RNA-seq. Peripheral blood samples were collected from healthy donors and melanoma patients for B cell phenotyping. ResultsCompared to B cells from spleen and lymph nodes, B cells in the pancreas exhibited significantly less follicular phenotype and higher IL-10 production in naive mice. c-Maf deficiency resulted in a significant reduction of CD9+ IL-10-producing Breg in the pancreas. PDAC progression resulted in accumulation of circulating B cells with follicular phenotype and less IL-10 production in the pancreas. Notably, B cell c-Maf deficiency delayed PDAC tumor progression and resulted in pro-inflammatory B cells. Further, tumor volume reduction and increased effective T cells in the tumor-draining lymph node (TDLN) were observed in B cell c-Maf KO mice in the B16.F10 melanoma model. RNA-seq analysis of isolated B cells revealed that B cell c-Maf signaling modulates immunoglobulin (Ig)-associated genes and tumor specific antibody production. We furthermore demonstrated c-Maf-positive B cell subsets and increase of IL-10-producing B cells after incubation with IL-4 and CD40L in the peripheral blood of melanoma patients. ConclusionOur study highlights that B cell c-Maf signaling drives tumor progression through the modulation of Breg, inflammatory responses, and tumor-specific Ab responses. What is already known on this topicThe net effect of B cells on tumor immunity depends on the balance of various B cell subtypes. c-Maf has been suggested to contribute to the regulation of IL-10 in regulatory B cells (Breg), but the role of B cell c-Maf signaling in anti-tumor immunity remains unknown. What this study addsThis study shown that B cell c-Maf signaling drives tumor progression in pancreatic cancer and melanoma. We defined different anti-tumor mechanisms of B cell c-Maf deficiency in two tumor models. Specifically, c-Maf signaling modulates the pro-inflammatory phenotype of B cells in the KPC tumor-bearing pancreas and tumor-specific antibody responses in tumor draining lymph nodes (TDLN) of melanoma. How this study might affect research, practice or policyThese studies indicate that inhibition of c-Maf signaling is a novel and promising approach for immunotherapy in pancreatic cancer and melanoma.

immunology↗

bioRxiv2024

Genetic code expansion reveals site-specific lactylation in living cells reshapes protein function

Still in its infancy, the functions of lactylation remain elusive. To address this, we established a comprehensive workflow for lactylation studies that integrates the discovery of lactylation sites with proteomics, the expression of site-specifically lactylated proteins in living cells via genetic code expansion (GCE), and the evaluation of the resulting biological consequences. Specifically, we developed a wet-and-dry-lab combined proteomics strategy, and identified highly conserved lactylation at ALDOA-K147. Driven by its potential biological significance, we site-specifically expressed this lactylated ALDOA in mammalian cells and interrogated the biological changes. We discovered that it not only inhibited enzyme activity but also elicited gain-of-function effects----it dramatically reshaped the functionality of ALDOA by improving stability, enhancing nuclear translocation and affecting gene expression. Further, we demonstrated broad applicability of this workflow to study distinct histone lactylation sites. Together, we anticipate its wide uses in elucidating causative links between site-specific lactylation and target-centric or cell-wide changes.

molecular biology↗

bioRxiv2024

Targeting protein tyrosine phosphatase non-receptor type 2 with a novel inhibitor for AML therapy

Acute myeloid leukemia (AML) is a fatal disease characterized by a bleak prognosis. For over four decades, treatment options for AML have been constrained to administering high-dose cytotoxic chemotherapy. However, the emergence of drug resistance and the resultant toxic side effects have created an urgent necessity for identifying novel therapeutic targets. In this study, non-receptor protein tyrosine phosphatase type 2 (PTPN2) is highly expressed in AML. Remarkably, knock down of PTPN2 expression markedly mitigated AML burden both in vitro and in vivo. Additionally, we unveiled the direct interaction between PTPN2 and C-MYC, establishing C-MYC as a direct acting substrate of PTPN2. Then, we performed a small-molecule compound screening to identify K73, a selective inhibitor of PTPN2. At the same time, K73 showed a good safety profile and exhibited strong activity against AML in vivo. In conclusion, we have pinpointed a considerable therapeutic potential in targeting PTPN2 for AML treatment and discovered a novel class of selective PTPN2 inhibitors suitable for AML therapy.

cancer biology↗

bioRxiv2024

SLC13A2-transported citrate is a metabolic signal for PKM2 acetylation and degradation to suppress tumor growth

Background and aimsMetabolic reprogramming represents as a pivotal hallmark for cancer, but TCA cycle in tumorigenesis and progression has long been neglected. Solute carrier (SLC) transporters mediate the transport of TCA cycle intermediates across membrane, but their functions in cancer pathogenesis remains unclear. Approaches & ResultsUsing integrated analysis of solute carrier (SLC) transporters for TCA cycle intermediates, we found that SLC13A2 was consistently downregulated in hepatocellular carcinoma (HCC) cells and liver tissues from human patients and heterogeneous mouse models. Adeno-associated virus (AAV)-transduced liver-specific knockout or overexpression of SLC13A2 promoted or ameliorated HCC progression in the primary mouse model, demonstrating that SLC13A2 served as a protective factor during HCC pathogenesis. SLC13A2 inhibited HCC cell proliferation by decreasing mitochondrial function via suppressed oxygen consumption and ATP production. Combined with metabolic flux analysis, we found that SLC13A2 imported citrate, which secreted acetyl-CoA as a precursor for the acetylation of pyruvate kinase muscle isozyme M2 (PKM2), which led to its protein degradation. Decreased activity of pyruvate kinase depleted pyruvate for the TCA cycle, thus inhibiting amino acid synthesis and nucleotide metabolism. Additionally, a decrease in nuclear PKM2 protein transduced to reprogrammed gene transcription for cell proliferation and metabolism which is required for tumor growth. ConclusionsThis study revealed that citrate transported by SLC13A2 acts as a signal to disrupt metabolic homeostasis for tumor growth and suggests potential drug targets for HCC therapy. graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/591017v2_ufig1.gif" ALT="Figure 1"> View larger version (64K): org.highwire.dtl.DTLVardef@80c95eorg.highwire.dtl.DTLVardef@124750corg.highwire.dtl.DTLVardef@809ab4org.highwire.dtl.DTLVardef@12cbe74_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

PARP14 is pro- and anti-viral host factor that promotes IFN production and affects the replication of multiple viruses

PARP14 is a 203 kDa multi-domain protein that is primarily known as an ADP-ribosyltransferase, and is involved in a variety of cellular functions including DNA damage, microglial activation, inflammation, and cancer progression. In addition, PARP14 is upregulated by interferon (IFN), indicating a role in the antiviral response. Furthermore, PARP14 has evolved under positive selection, again indicating that it is involved in host-pathogen conflict. We found that PARP14 is required for increased IFN-I production in response to coronavirus infection lacking ADP-ribosylhydrolase (ARH) activity and poly(I:C), however, whether it has direct antiviral function remains unclear. Here we demonstrate that the catalytic activity of PARP14 enhances IFN-{beta} and IFN-{lambda} responses and restricts ARH-deficient murine hepatitis virus (MHV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication. To determine if PARP14s antiviral functions extended beyond CoVs, we tested the ability of herpes simplex virus 1 (HSV-1), a DNA virus, vesicular stomatitis virus (VSV), a negative-sense RNA virus, and lymphocytic choriomeningitis virus (LCMV), an ambisense RNA virus, to infect A549 PARP14 knockout (KO) cells. While LCMV infection was unaffected, HSV-1 replication was increased in PARP14 KO cells and VSV replication was decreased. These results indicate that PARP14 restricts HSV-1 replication but enhances the replication of VSV. A PARP14 active site inhibitor had no impact on HSV-1 or VSV replication, indicating that its effect on these viruses was independent of its catalytic activity. These data demonstrate that PARP14 promotes IFN production and has both proviral and antiviral functions targeting multiple viruses. IMPORTANCEThe antiviral response is largely regulated by post-translation modifications (PTM), including ADP-ribosylation. PARP14 is an ADP-ribosyltransferase that is upregulated by interferon and is under positive selection, indicating that it is involved in host-pathogen conflict. However, no anti-viral function has been described for PARP14. Here, we found that PARP14 represses both coronavirus and HSV-1 replication, demonstrating that PARP14 has antiviral functions. Surprisingly, we also found that PARP14 also has pro-viral functions, as it was critical for the efficient replication of VSV. These data indicate that PARP14 has both proviral and antiviral functions. Defining the mechanisms used by PARP14 to both repress and promote virus replication will provide new insights into how PARPs regulate virus infection.

microbiology↗

bioRxiv2024

Heparan sulfate selectively inhibits the collagenase activity of cathepsin K

Cathepsin K (CtsK) is a cysteine protease with potent collagenase activity. CtsK is highly expressed by bone-resorbing osteoclasts and plays an essential role in bone remodeling. Although CtsK is known to bind heparan sulfate (HS), the structural details of the interaction, and how HS ultimately regulates the biological functions of CtsK, remains largely unknown. In this report, we determined that CtsK preferably binds to larger HS oligosaccharides, such as dodecasaccharides (12mer), and that the12mer can induce monomeric CtsK to form a stable dimer in solution. Interestingly, while HS has no effect on the peptidase activity of CtsK, it greatly inhibits the collagenase activity of CtsK in a manner dependent on sulfation level. By forming a complex with CtsK, HS was able to preserve the full peptidase activity of CtsK for prolonged periods, likely by stabilizing its active conformation. Crystal structures of Ctsk with a bound 12mer, alone and in the presence of the endogenous inhibitor cystatin-C reveal the location of HS binding is remote from the active site. Mutagenesis based on these complex structures identified 6 basic residues of Ctsk that play essential roles in mediating HS-binding. At last, we show that HS 12mers can effectively block osteoclast resorption of bone in vitro. Combined, we have shown that HS can function as a multifaceted regulator of CtsK and that HS-based oligosaccharide might be explored as a new class of selective CtsK inhibitor in many diseases that involve exaggerated bone resorption.

biochemistry↗

bioRxiv2023

Three-dimensional dipole orientation mapping with high temporal-spatial resolution using polarization modulation

Fluorescence polarization microscopy is widely used in biology for molecular orientation properties. However, due to the limited temporal resolution of single-molecule orientation localization microscopy and the limited orientation dimension of polarization modulation techniques, achieving simultaneous high temporal-spatial resolution mapping of the three-dimensional (3D) orientation of fluorescent dipoles remains an outstanding problem. Here, we present a super-resolution 3D orientation mapping (3DOM) microscope that resolves 3D orientation by extracting phase information of the six polarization modulation components in reciprocal space. 3DOM achieves an azimuthal precision of 2{degrees} and a polar precision of 3{degrees} with spatial resolution of 128 nm in the experiments. We validate that 3DOM not only reveals the heterogeneity of the milk fat globule membrane, but also elucidates the 3D structure of biological filaments, including the 3D spatial conformation of {lambda}-DNA and the structural disorder of actin filaments. Furthermore, 3DOM images the dipole dynamics of microtubules labeled with green fluorescent protein in live U2OS cells, reporting dynamic 3D orientation variations. Given its easy integration into existing wide-field microscopes, we expect the 3DOM microscope to provide a multi-view versatile strategy for investigating molecular structure and dynamics in biological macromolecules across multiple spatial and temporal scales.

molecular biology↗

bioRxiv2023

Neurophysiological Features of STN LFP underlying Sleep Fragmentation in Parkinson's Disease

BackgroundSleep fragmentation is a persistent problem throughout the course of Parkinsons disease (PD). However, the related neurophysiological patterns and the underlying mechanisms remained unclear. MethodWe recorded subthalamic nucleus (STN) local field potentials (LFPs) using DBS with real-time wireless recording capacity from thirteen PD patients undergoing a one-night polysomnography recording, one month after DBS surgery before initial programming and when the patients were Off-Medication. The STN LFP features that characterized different sleep stages, correlated with arousal and sleep fragmentation index, and preceded stage transitions during N2 and REM sleep were analyzed. ResultsBoth beta and low gamma oscillations in NREM sleep increased with the severity of sleep disturbance (arousal index (ArI)-betaNREM: r=0.9,p=0.0001) and sleep fragmentation index (SFI)-betaNREM: r=0.6,p=0.0301; SFI-gammaNREM: r=0.6,p=0.0324). We next examined the low-to-high power ratio, which was the power ratio of theta oscillations to beta and low gamma oscillations, and found it to be an indicator of sleep fragmentation (ArI-LHPRNREM: r=-0.8,p=0.0053; ArI-LHPRREM: r=-0.6,p=0.0373; SFI-LHPRNREM: r=-0.7,p=0.0204; SFI-LHPRREM: r=-0.6,p=0.0428). In addition, long beta bursts (>0.25s) during NREM stage 2 were found preceding the completion of transition to stages with more cortical activities (towards Wake/N1/REM compared with towards N3 (p<0.01)) and negatively correlated with STN spindles, which were detected in STN LFPs with peak frequency distinguishable from long beta bursts (STN spindle: 11.5Hz, STN long beta bursts: 23.8Hz), in occupation during NREM sleep ({beta} = -0.24,p<0.001). ConclusionFeatures of STN LFPs help explain neurophysiological mechanisms underlying sleep fragmentations in PD, which can inform new intervention for sleep dysfunction. What is already known on this topicBeta oscillation, which is a biomarker for rigidity and bradykinesia during awake, significantly reduced during NREM sleep compared to REM or awake. Researches from MPTP non-human primate model suggested increased beta oscillation in basal ganglia contributed to insomnia in PD, however evidence in human patients is still lacking. What this study addsBeta and low gamma band activities in STN LFPs during sleep recorded from human PD patients correlated with severity of sleep impairment. The low-high power ratio can serve as a biomarker for sleep fragmentation. Besides, pathological beta bursts and physiological sleep spindles can be detected in STN LFP during NREM sleep and distinguishable from each other. How this study might affect research, practice or policyThese findings enhance our understanding of the electrophysiological mechanisms that underlie sleep fragmentation in Parkinsons disease. The study also has implications on the design of closed-loop DBS: we may need to take multiple frequency band activities into consideration and differentiate pathological beta oscillation from the sleep spindles.

physiology↗

bioRxiv2023

A resource for probing influences of perinatal factors on neurodevelopment in 3-10-years-old Chinese children

Adverse perinatal factors can disrupt the normal development of the brain, with potential long-term impacts on childrens overall development. Currently, the neuropathological mechanisms by which these factors lead to various neurodevelopmental disorders (NDDs) remain largely unknown. An open resource that integrate perinatal factors with brain and mental health development is essential for investigating NDD-related aetiology. In this Data Descriptor, we introduce a multicentre database containing information on perinatal factors that influence childrens brain-mind development, namely, periCBD, that combines neuroimaging and behavioural phenotypes with perinatal factors associated with a high incidence of NDDs at county/region/central district hospitals. PeriCBD was designed to establish a platform for the investigation of individual differences in brain-mind development among children aged 3-10 years are associated with perinatal factors. Ultimately, our goal was to develop an early prediction and screening model for NDDs that leverages normative data to facilitate NDD aetiology research. Herein, we provide a systematic overview of the data acquisition/cleaning/quality control/sharing, processes of periCBD and present preliminary brain-mind associations.

neuroscience↗

bioRxiv2023

Heparan sulfate promotes TRAIL-induced tumor cell apoptosis

TRAIL (TNF-related apoptosis-inducing ligand) is a potent inducer of tumor cell apoptosis through TRAIL receptors. While it has been previously pursued as a potential anti-tumor therapy, the enthusiasm subsided due to unsuccessful clinical trials and the fact that many tumors are resistant to TRAIL. In this report we identified heparan sulfate (HS) as an important regulator of TRAIL-induced apoptosis. TRAIL binds HS with high affinity (KD = 73 nM) and HS induces TRAIL to form higher-order oligomers. The HS-binding site of TRAIL is located at the N-terminus of soluble TRAIL, which includes three basic residues. Binding to cell surface HS plays an essential role in promoting the apoptotic activity of TRAIL in both breast cancer and myeloma cells, and this promoting effect can be blocked by heparin, which is commonly administered to cancer patients. We also quantified HS content in several lines of myeloma cells and found that the cell line showing the most resistance to TRAIL has the least expression of HS, which suggests that HS expression in tumor cells could play a role in regulating sensitivity towards TRAIL. We also discovered that death receptor 5 (DR5), TRAIL and HS can form a ternary complex and that cell surface HS plays an active role in promoting TRAIL-induced cellular internalization of DR5. Combined, our study suggests that TRAIL-HS interactions could play multiple roles in regulating the apoptotic potency of TRAIL and might be an important point of consideration when designing future TRAIL-based anti-tumor therapy.

cell biology↗

bioRxiv2023

A Gpr35 tuned gut-brain metabolic axis regulates depressive-like behavior

Gene-environment interactions shape animal behavior and the susceptibility to neurobehavioral symptoms such as depression. However, little is known about the signaling pathway that integrates genetic and environmental inputs with neurobehavioral outcomes, preventing the development of targeted therapies. Here we report that Gpr35 engages a gut microbe-to-brain metabolic pathway to modulate neuronal plasticity and depressive behavior in mice. Chronic stress decreases gut epithelial Gpr35, the genetic deletion of which induces despair and social impairment in a microbiome-dependent manner. We identify a dominant role for the imbalance of microbe-derived indole-3-carboxaldehyde (IAld) and indole-3-lactate (ILA) in the behavioral symptoms with Gpr35 deficiency. Mechanistically, these bacterial metabolites counteractively modulate dendritic spine density and synaptic transmission in the nucleus accumbens. Supplementation of IAld, which is similarly decreased in depressive patients, produce anti-depressant effects in mice with stress or gut epithelial Gpr35 deficiency. Together, these findings identify a genetics-shaped gut-brain connection underlying the susceptibility to depression and suggest a microbial metabolite-based therapeutic strategy to genetic predisposition.

animal behavior and cognition↗

bioRxiv2022

Ultra-sensitive platelet proteome maps the O-glycosylation landscape and charts the response to thrombin dosage

Platelet activation induces the secretion of proteins that promote platelet aggregation and inflammation. However, detailed analysis of the released platelet proteome is hampered by platelets tendency to pre-activate during their isolation and a lack of sensitive protocols for low abundance releasate analysis. Here we detail the most sensitive analysis to date of the platelet releasate proteome with the detection of >1,300 proteins. Unbiased scanning for post-translational modifications within releasate proteins highlighted O-glycosylation as being a major component. For the first time, we detected O-fucosylation on previously uncharacterised sites including multimerin-1 (MMRN1), a major alpha granule protein that supports platelet adhesion to collagen and is a carrier for platelet factor V. The N-terminal EMI domain of MMRN1, a key site for protein-protein interaction, was O-fucosylated at a conserved threonine within a new consensus sequence. Our data suggest that Protein O-fucosyltransferase 1 (POFUT1) is responsible for this modification. Secretion of MMRN1 was reduced in cells lacking POFUT1, supporting a key role of O-fucosylation in MMRN1 function. By comparing releasates from resting and thrombin-treated platelets, 202 proteins were found to be significantly released after high-dose thrombin stimulation. Complementary quantification of the platelet lysates identified >3,800 proteins, which confirmed the platelet origin of releasate proteins by anti-correlation analysis. Low-dose thrombin treatment yielded a smaller subset of significantly regulated proteins with fewer secretory pathway enzymes. The comprehensive platelet proteome resource provided here (larancelab.com/platelet-proteome) allows identification of novel regulatory mechanisms for drug targeting to address platelet dysfunction and thrombosis. Key PointsO_LIHigh-sensitivity proteome mapping of human platelets identifies O-glycosylation of platelet proteins at key functional sites C_LIO_LIPlatelet O-fucosyltransferase POFUT1 regulates the secretion of adhesive protein multimerin-1 (MMRN1) C_LI

biochemistry↗

bioRxiv2022

Defining specific cell states of MPTP-induced Parkinson's disease by single-nucleus RNA sequencing

Parkinsons disease (PD), a neurodegenerative disease with the impairment of movement execution that is related to age, genetic and environmental factors. 1-methyl-4-phenyl-1,2,3,6-tetrahydropyri-dine (MPTP) is a neurotoxin widely used to induce PD models, but the effect of MPTP on cell-gene of PD has not been fully elucidated. By single-nucleus RNA sequencing, we uncovered the PD-specific cells and revealed remarkable changes in their cellular states, including astrocytosis, endothelial cells absence, as well as a cluster of PD-exclusive medium spiny neuron cells. Furthermore, trajectory analysis of astrocyte and endothelial cells populations predicted candidate target gene sets that might be associated with PD. Notably, the detailed regulatory roles of astrocyte-specific transcription factors Dbx2 and Sox13 in PD were first revealed in our work. Finally, we characterized the cell-cell communications of PD-specific cells and found that the overall communication strength was enhanced in PD compared with matched control, especially the signaling pathways of NRXN and NEGR. Our work provides comprehensive overview on the changes of cellular states of the MPTP-induced mouse brain.

neuroscience↗

bioRxiv2022

Bile Acid Restrained T Cell Activation Explains Cholestasis Aggravated Hepatitis B Virus Infection

Cholestasis is a common complication of Hepatitis B Virus (HBV) infection, characterized by increased intrahepatic and plasma bile acid levels. Cholestasis was found negatively associated with hepatitis outcome, however; the exact mechanism by which cholestasis impact on anti-viral immunity and impede HBV clearance remains elusive. Here, we found that cholestatic mice are featured with dysfunctional T cell response, and bile acids inhibit the activation and metabolic reprogramming of CD4+ T cells. Mechanistically, bile acids disrupt intracellular calcium homeostasis via inhibiting mitochondria calcium uptake and elevating cytoplasmic Ca2+ concentration of CD4+ T cells, leading to STIM1 and ORAIL1 decoupling and impaired store-operated Ca2+ entry which is essential for NFAT signaling and T cell activation. Moreover, in a transgenic mouse model of HBV infection, it was confirmed that cholestasis compromised T cells activation resulting in poor viral clearance. Collectively, our results suggest that bile acids play pivotal roles in anti-HBV infection via controlling T cells activation and metabolism, and that targeting regulation of bile acids may be a therapeutic strategy for host virus defense.

immunology↗

bioRxiv2022

Metagenomic and Metabolomic Analyses Reveal Synergistic Effects of Fecal Microbiota Transplantation and Anti-PD-1 Therapy on Treating Colorectal Cancer

Anti-PD-1 immunotherapy has saved numerous lives of cancer patients; however, it only exerts efficacy in 10-15% of patients with colorectal cancer. Fecal microbiota transplantation (FMT) is a potential approach to improving the efficacy of anti-PD-1 therapy, whereas the detailed mechanisms and the applicability of this combination therapy remain unclear. In this study, we evaluated the synergistic effect of FMT with anti-PD-1 in curing colorectal tumor-bearing mice using a multi-omics approach. Mice treated with the combination therapy showed superior survival rate and tumor control, compared to the mice received anti-PD-1 therapy or FMT alone. Metagenomic analysis showed that composition of gut microbiota in tumor-bearing mice treated with anti-PD-1 therapy was remarkably altered through receiving FMT. Particularly, Bacteroides genus, including FMT-increased B. thetaiotaomicron, B. fragilis, and FMT-decreased B. ovatus might contribute to the enhanced efficacy of anti-PD-1 therapy. Furthermore, metabolomic analysis upon mouse plasma revealed several potential metabolites that upregulated after FMT, including punicic acid and aspirin, might promote the response to anti-PD-1 therapy via their immunomodulatory functions. This work broadens our understanding of the mechanism by which FMT improves the efficacy of anti-PD-1 therapy, which may contribute to the development of novel microbiota-based anti-cancer therapies.

microbiology↗

bioRxiv2021

The evolution of fluoroquinolone resistance in Salmonella under exposure to sub-inhibitory concentration of enrofloxacin

The evolution of resistance in Salmonella to fluoroquinolones (FQs) under a broad range of sub-inhibitory concentrations (sub-MICs) has not been systematically studied. This study investigated the mechanism of resistance development in Salmonella enterica serovar Enteritidis (S. Enteritidis) under sub-MICs of 1/128xMIC to 1/2xMIC of enrofloxacin (ENR), a widely used veterinary FQ. It was shown that the resistance rate and resistance level of S. Enteritidis varied with the increase of ENR concentration and duration of selection. qRT-PCR results demonstrated that the expression of outer membrane porin (OMP) genes, ompF, ompC and ompD, were down-regulated first to rapidly adapt and develop resistance of [≤] 4xMIC, and as the resistance level increased ([≥]8xMIC), the up-regulated expression of efflux pump genes, acrB, emrB amd mdfA, along with mutations in quinolone resistance-determining region (QRDR) gradually played a decisive role. Cytohubba analysis based on transcriptomic profiles demonstrated that purB, purC, purD, purF, purH, purK, purL, purM, purN and purT were the hub genes for the FQs resistance. de novo IMP biosynthetic process, purine ribonucleoside monophosphate biosynthetic process and purine ribonucleotide biosynthetic process were the top three biological processes screened by MCODE. This study first described the dynamics of FQ resistance evolution in Salmonella under a long-term selection of sub-MICs of ENR in vitro. In addition, this work offers greater insight into the transcriptome changes of S. Enteritidis under the selection of ENR and provides a framework for FQs resistance of Salmonella for further studies.

microbiology↗

bioRxiv2021

Exploration of clinical breakpoint of Danofloxacin for Glaesserella parasuis in plasma and in PELF

BackgroundTo establish the clinical breakpoint (CBP) of danofloxacin to G. parasuis, three cutoff values, including epidemiological cutoff value (ECV), pharmacodynamic cutoff value (COPD) and clinical cutoff value (COCL), was obtained in the present study. MethodsThe ECV was calculated using ECOFFinder base on MIC distribution of 347 G. parasuis collected from disease pigs. The COPD was established base on in vivo and ex vivo pharmacokinetic (PK) - pharmacodynamic (PD) modeling of danofloxacin both in plasma and pulmonary epithelial lining fluid (PELF) using Hill formula and Monte Carlo analysis. The COCL was established based on the relationship between possibility of cure (POC) and MIC in the clinical trials using "WindoW" approach, nonlinear regression and CART analysis. ResultsThe MIC50 and MIC90 of danofloxacin against 347 G. parasuis were 2 g/mL and 8 g/mL, respectively. The ECV value was set up as 8 g/mL using ECOFFinder. Concentration-time curve of danofloxacin indicated a two-compartment model for PK analysis. The PK parameters of the maximum concentration (Cmax) and area under concentration-time curve (AUC) in PELF were 3.67 {+/-} 0.25 g/mL and 24.28 {+/-} 2.70 h{middle dot}g/mL, higher than those in plasma (0.67 {+/-} 0.01g/mL and 4.47 {+/-} 0.51 h{middle dot}g/mL). The peak time (Tmax) in plasma was 0.23 {+/-} 0.07 h, shorter than that in PELF (1.61 {+/-} 0.15 h). The COPD in plasma and PELF were 0.125 g/mL and 0.5 g/mL, respectively. The COCL calculated by WindoW approach, nonlinear regression and CART analysis were 0.125[~]4 g/mL, 0.428 g/mL and 0.56 g/mL, respectively. The 0.5 g/mL was selected as eligible COCL. The ECV is much higher than the COPD and COCL, and the clinical breakpoint based on data in plasma was large different with that of in PELF. ConclusionsOur study firstly established three cutoff values of danofloxacin against G. parasuis. It suggested that epidemiological danofloxacin-resistant G. parasuis may lead to the ineffective treatment by danofloxacin. ImportanceG. parasuis, a gram-negative respiratory pathogen, can colonize in the upper respiratory tract in swine and cause Glassers disease. As the abuse of antibiotics, antimicrobial resistant G. parasuis emerged in different degrees, which brought serious threat to global economy and public health. Danofloxacin in quinolones are one of the best choices for treatment of G. parasuis infection, because of their strong bactericidal activity and good absorption into blood and great distribution in the lung. However, the clinical breakpoint (CBP) for danofloxacin against G. parasuis had not yet been established by clinical laboratory of standard Institute (CLSI) and European Commission of antimicrobial susceptibility testing (EUCAST). Our study firstly established three cutoff values of danofloxacin against G. parasuis. It suggested that epidemiological danofloxacin-resistant G. parasuis may lead to the ineffective treatment by danofloxacin.

microbiology↗

bioRxiv2021

Genetic Diversity of Dianthus Plants Revealed by SRAP Markers

Dianthus is a valuable genetic resource for flower breeding. However, study on the genetic similarity of Dianthus plants is rare. In this work, 20 pairs of SRAP markers were used to analyze genetic diversity of 44 Dianthus plants, including 13 lines of wild Dianthus chinensis L., 7 lines of wild Dianthus superbus L. and 24 commercial varieties including Dianthus caryophyllus L., Dianthus plumarius L. and Dianthus barbatus L.. Results showed that precise interspecific and intraspecific genetic diversity was provided in Dianthus plants by using 20 pairs of SRAP molecular markers. The interspecific genetic diversity of Dianthus plants was much abundant and the intraspecific genetic difference of wild Dianthus species was related to their geographical distribution and habitats. In this work, theoretical basis and technical support were provided for crossbreeding and molecular mechanism research of Dianthus plants.

genetics↗