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Abstract Background Glechoma longituba, recognized as a medicinal plant, provides valuable pharmaceutical raw materials for treating various diseases. Saline-alkali stress may effectively enhance the medicinal quality of G. longituba by promoting the synthesis of secondary metabolites. To investigate the changes in the primary medicinal components of G. longituba under saline-alkali stress and improve the quality of medicinal materials, Na2CO3 was applied to induce short-term stress under different conditions and the biomass, physiologically active substances and primary medicinal components of G. longituba were measured in this study. Results Under alkaline salt stress, the activities of catalase (CAT), superoxide dismutase (SOD), peroxidase (POD), and ascorbate peroxidase (APX) were elevated in G. longituba, accompanied by increased accumulation of proline (Pro) and malondialdehyde (MDA). Furthermore, analysis of the medicinal constituents revealed that G. longituba produced the highest levels of soluble sugars, flavonoids, ursolic acid, and oleanolic acid under 0.6% Na2CO3 stress for 48 h, 0.2% Na2CO3 stress for 72 h, 0.4% Na2CO3 stress for 12 h, and 0.4% Na2CO3 stress for 8 h, respectively. Conclusions Short-term Na2CO3 stress enhances the synthesis of medicinal components in G. longituba. By manipulating stress conditions, the production of various medicinal substances could be optimized. This approach may serve as a basis for the targeted cultivation of G. longituba, offering potential applications in the treatment of diverse diseases.

Abstract Background Gastric cancer is a significant global malignancy with poor prognosis. Although the emergence of immune checkpoint inhibitors (ICIs) prolonged the duration of survival, resistance and progression are inevitable. We aim to evaluate the effectiveness of programmed death-1 (PD-1) inhibitors in immunotherapy beyond progression (IBP). Method We divided the advanced gastric cancer patients who received two lines immunotherapy into same regimen group (with same PD-1 inhibitor regime after IBP) and different regimen group (with different PD-1 inhibitor regime after IBP). Statistical analysis conducted to compare patient characteristics and evaluate survival differences between groups. Result The clinical outcome analysis showed that the same PD-1 inhibitor regime seemed to exhibit a higher disease control rate (DCR) (51.8% vs. 29.2%, P = 0.062), significantly prolonged progression-free survival 2 (PFS2) (162 vs. 75 days, P = 0.001) and overall survival (OS) (312 vs. 166 days, P = 0.022) when compared with those of cross line. In the multivariate analysis, when using different regimen group as reference, the same regimen group was found to be independently associated with improved PFS2 [hazard ratio (HR) = 0.467, 95% confidence interval (CI): 0.267–0.816, P = 0.008] and OS (HR = 0.508, 95%CI: 0.278–0.927, P = 0.027). Conclusion Continuation of the same type of PD-1 inhibitor regime in IBP shows clinical benefits and represents a promising therapeutic approach.

Abstract Background Obesity is known as a risk factor for cardiovascular disease (CVD). However, there is an absence of preoperative cardiac risk assessment in bariatric surgery candidates and the incidence of CVD among these high-risk patients is still unknown. Methods A consecutive series of bariatric surgery candidates at two Chinese tertiary hospitals received coronary CT angiography or coronary angiography from 2017 to 2023. Patients were categorized as metabolically unhealthy obesity (MUO) and metabolically healthy obesity (MHO) based on the presence or absence of MetS. CVD was diagnosed based on the maximum intraluminal stenosis > 1% in any of the segments of the major epicardial coronary arteries. Obstructive CVD was defined as coronary stenosis ≥ 50%. Binary multivariable logistic regression was performed to analyze the association between CVD and metabolic status. The number of principal MetS components was categorized into zero (without glycemic, lipid, and BP components), one (with one of the components), two (with any two components), and three (with all components) to explore their association with CVD. Results A total of 1446 patients were included in the study. The incidence of CVD and obstructive CVD were 31.7% and 9.6%. Compared with MHO patients, MUO patients had a significantly higher incidence of mild (13.7% vs. 6.1%, P

Abstract Background The ricefield eel Monopterus albus undergoes a natural sex change from female to male during its life cycle, and previous studies have shown the potential mechanisms of this transition at the transcriptional and protein levels. However, the changes in protein levels have not been fully explored, especially in the intersexual stage. Results In the present study, the protein expression patterns in the gonadal tissues from five different periods, the ovary (OV), early intersexual stage gonad (IE), middle intersexual stage gonad (IM), late intersexual stage gonad (IL), and testis (TE), were determined by untargeted proteomics sequencing. A total of 5125 proteins and 394 differentially expressed proteins (DEPs) were detected in the gonadal tissues. Of the 394 DEPs, there were 136 between the OV and IE groups, 20 between the IM and IE groups, 179 between the IL and IM groups, and 59 between the TE and IL groups. Three candidate proteins, insulin-like growth factor 2 mRNA-binding protein 3 isoform X1 (Igf2bp3), triosephosphate isomerase (Tpi), and Cu-Zn superoxide dismutase isoform X1 [(Cu-Zn) Sod1], were validated by western blotting to verify the reliability of the data. Furthermore, metal metabolite-related proteins were enriched in the IL vs. IM groups and TE vs. IL groups, which had close relationships with sex change, including Cu2+-, Ca2+-, Zn2+- and Fe2+/Fe3+-related proteins. Analysis of the combined transcriptome data revealed consistent protein/mRNA expression trends for two metal metabolite-related proteins/genes [LOC109953912 and calcium Binding Protein 39 Like (cab39l)]. Notably, we detected significantly higher levels of Cu2+ during the sex change process, suggesting that Cu2+ is a male-related metal metabolite that may have an important function in male reproductive development. Conclusions In summary, we analyzed the protein profiles of ricefield eel gonadal tissues in five sexual stages (OV, IE, IM, IL, and TE) and verified the plausibility of the data. After preforming the functional enrichment of metal metabolite-related DEPs, we detected the contents of the metal metabolites Zn2+, Cu2+, Ca2+, and Fe2+/Fe3+ at these five stages and screened for (Cu-Zn) Sod1 and Mmp-9 as possible key proteins in the sex reversal process.

Abstract Purpose To compare the difference in analgesic effect between femoral triangle block (FTB) and adductor canal block (ACB) during arthroscopic knee surgery. Methods Patients who underwent arthroscopic knee surgery were randomized preoperatively to FTB group or ACB group. For each group, 20 mL of 0.1% ropivacaine was injected. Primary outcomes: The numeric rating score (NRS) at 12 h after surgery at rest and during movement. Secondary outcome: (1) The NRS at post anesthesia care unit (PACU) and 2, 24 h after surgery at rest and during movement; (2) The quadriceps muscle strength at PACU and 2, 12, 24 h after surgery; (3) Consumption of Rescue analgesia; (4) Incidence of adverse reactions. Results The NRS at 12 h after surgery at rest and during movement of ACB group were higher than FTB group. Among secondary outcomes, the NRS at PACU at rest and during movement, 2 h after surgery during movement of FTB group lower than ACB group; the quadriceps muscle strength at 2 h after surgery of FTB group stronger than ACB group. After multiple linear regression model analysis, the data showed additional statistically significant reduction NRS at 24 h after surgery at rest (0.757, p = 0.037) in FTB group. Other outcomes were similar between two groups. Conclusions The FTB appears to provide superior pain control after knee arthroscopy than ACB, the FTB is superior to the ACB in quadriceps muscle strength at 2 h after surgery. Trial registration The trial was registered in the Chinese Clinical Trial Registry (ChiCTR2300068765). Registration date: 28/02/2023.

Abstract Accurate preoperative differentiation of the chronic rhinosinusitis (CRS) endotype between eosinophilic CRS (eCRS) and non-eosinophilic CRS (non-eCRS) is an important topic in predicting postoperative outcomes and administering personalized treatment. To this end, we have constructed a sinus CT dataset, which comprises CT scan data and pathological biopsy results from 192 patients of chronic rhinosinusitis with nasal polyps (CRSwNP), treated at the Second Affiliated Hospital of Shantou University Medical College between 2020 and 2022. To differentiate CRSwNP endotype on preoperative CT and improve efficiency at the same time, we developed a multi-view fusion model that contains a mini-architecture with each network of 10 layers by modifying the deep residual neural network. The proposed model is trained on a training set and evaluated on a test set. The multi-view deep learning fusion model achieved the area under the receiver-operating characteristics curve (AUC) of 0.991, accuracy of 0.965 and F1-Score of 0.970 in test set. We compared the performance of the mini-architecture with other lightweight networks on the same Sinus CT dataset. The experimental results demonstrate that the developed ResMini architecture contribute to competitive CRSwNP endotype identification modeling in terms of accuracy and parameter number.

Abstract Background Atresia and degeneration, a follicular developmental fate that reduces female fertility and is triggered by granulosa cell (GC) apoptosis, have been induced by dozens of miRNAs. Here, we report a miRNA, miR-423, that inhibits the initiation of follicular atresia (FA), and early apoptosis of GCs. Results We showed that miR-423 was down-regulated during sow FA, and its levels in follicles were negatively correlated with the GC density and the P4/E2 ratio in the follicular fluid in vivo. The in vitro gain-of-function experiments revealed that miR-423 suppresses cell apoptosis, especially early apoptosis in GCs. Mechanically speaking, the miR-423 targets and interacts with the 3'-UTR of the porcine SMAD7 gene, which encodes an apoptosis-inducing factor in GCs, and represses its expression and pro-apoptotic function. Interestingly, FA and the GC apoptosis-related lncRNA NORHA was demonstrated as a ceRNA of miR-423. Additionally, we showed that a single base deletion/insertion in the miR-423 promoter is significantly associated with the number of stillbirths (NSB) trait of sows. Conclusion These results demonstrate that miR-423 is a small molecule for inhibiting FA initiation and GC early apoptosis, suggesting that treating with miR-423 may be a novel approach for inhibiting FA initiation and improving female fertility.

Abstract Aims To evaluate the effect of antibiotic prophylaxis(AP) in the prognosis of Post-neurosurgical meningitis(PNM) patients. Methods A cohort analysis was performed using the clinical database in Beijing Tiantan Hospital and Capital Medical University. Data were collected on patients with the diagnosis of PNM (n = 3931) during 2012.01 to 2022.04. The microbial distribution, types of AP, and 42 and 90 days survival analysis of AP patients were evaluated using probable statistical methods. Independent risk factors for mortality were established by constructing a logistic regression analysis. Result A total of 1,190 patients were included in this study, Klebsiella pneumoniae, Acinetobacter baumannii, and Staphylococcus aureus occupied the highest proportion. Of them, 929 cases received AP, cefuroxime and ceftriaxone are the most frequent used antibiotics. In addition, We found that PNM patients without AP significantly increased the 42 days and 90 days all-cause mortality rates. The use of different levels of AP did not improve patient outcomes, and ICU admission and assisted mechanical ventilation (AMV) were identified as independent mortality risk factors for PNM patient received AP. Conclusions AP plays an important role in the prognosis of PNM patients and has a significant function in improving prognosis. The prevention of PNM with antibiotics prior to neurosurgery should be emphasized in clinical practice, and appropriate selection of antibiotics is necessary to prevent the occurrence of infection and inhibit the emergence of antibiotic-resistant bacteria.

Abstract The WUSCHEL-related homeobox (WOX) family members are plant-specific transcriptional factors, which function in meristem maintenance, embryogenesis, lateral organ development, as well as abiotic stress tolerance. In this study, 14 MsWOX transcription factors were identified and comprehensively analyzed in the cultivated alfalfa cv. Zhongmu No.1. Overall, 14 putative MsWOX members containing conserved structural regions were clustered into three clades according to phylogenetic analysis. Specific expression patterns of MsWOXs in different tissues at different levels indicated that the MsWOX genes play various roles in alfalfa. MsWUS, MsWOX3, MsWOX9, and MsWOX13-1 from the three subclades were localized in the nucleus, among which, MsWUS and MsWOX13-1 exhibited strong self-activations in yeast. In addition, various cis-acting elements related to hormone responses, plant growth, and stress responses were identified in the 3.0 kb promoter regions of MsWOXs. Expression detection of separated shoots and roots under hormones including auxin, cytokinin, GA, and ABA, as well as drought and cold stresses, showed that MsWOX genes respond to different hormones and abiotic stress treatments. Furthermore, transcript abundance of MsWOX3, and MsWOX13-2 were significantly increased after rhizobia inoculation. This study presented comprehensive data on MsWOX transcription factors and provided valuable insights into further studies of their roles in developmental processes and abiotic stress responses in alfalfa.

Abstract Background The fat attenuation index (FAI) is a radiological parameter that represents pericoronary adipose tissue (PCAT) inflammation, along with myocardial bridging (MB), which leads to pathological shear stress in the coronary vessels; both are associated with coronary atherosclerosis. In the present study, we assessed the predictive value of FAI values and MB parameters through coronary computed tomography angiography (CCTA) for predicting the risk of coronary atherosclerosis and vulnerable plaque in patients with MB. Methods We included 428 patients who underwent CCTA and were diagnosed with MB. FAI values, MB parameters, and high-risk coronary plaque (HRP) characteristics were recorded. The subjects were classified into two groups (A and B) according to the absence or presence of coronary plaque in the segment proximal to the MB. Group B was further divided into Groups B1 (HRP-positive) and B2 (HRP-negative) according to the HRP characteristic classification method. The differences among the groups were analysed. Multiple logistic regression analysis was performed to determine the independent correlation between FAI values and MB parameters and coronary atherosclerosis and vulnerable plaque risk. Results Compared to the subjects in Group A, those in Group B presented greater MB lengths, MB depths and muscle index values, more severe MB systolic stenosis and higher FAIlesion values (all P

Abstract Background Pregnancy-related intracranial hemorrhage (ICH) is a rare but potentially life-threatening event with complex and varied cause, such as HELLP syndrome and hemophagocytic syndrome. Case presentation A 33-year-old patient underwent a cesarean section with a preliminary diagnosis of "severe preeclampsia and class3 HELLP syndrome ". The patient had poor response to language before surgery, and the catheter drainage fluid was hematuria. Later, the surgeon reported severe bleeding in the operation. Following thromboelastography (TEG) result and postoperative laboratory tests confirmed class1 HELLP syndrome and ICH occurred on the second day after the surgery, and hemophagocytic syndrome was diagnosed during subsequent treatments. Conclusion For patients with HELLP syndrome, we should pay attention to their coagulation condition. The coagulation tests and platelet counts should be repeated if their clinical presentation changed. Those with neurological alarm signs should receive CT or MRI scan. If a pregnant woman had prolonged hemocytopenia and thrombocytopenia, not only the HELLP but also the hemophagocytic syndrome should be considered.

Abstract Background Transmission-blocking vaccines (TBVs) target the sexual stages of malaria parasites to reduce or interrupt the transmission cycle in human and mosquito populations. The genetic diversity of TBVs candidate antigens, Pvs25 and Pvs28, in Plasmodium vivax could provide evidence for the development of TBVs. Methods Dry blood spots from P. vivax patients were collected from Dandong, Suining, Hainan, Nyingchi, Tengchong, and Yingjiang in China. The pvs25 and pvs28 genes were amplified and sequenced. The genetic diversity of pvs25 and pvs28 were analyzed using DNASTAR, MEGA6, and DnaSP 5.0 programs. Results A total of 377 samples were collected, among which 324 and 272 samples were successfully amplified in the pvs25 and pvs28 genes, respectively. Eight haplotypes were identified in Pvs25, for which the predominant mutation was I130T with 100% prevalence. A variety of 22 haplotypes in Pvs28 were identified. The number of GSGGE/D repeats of Pvs28 was a range of 4–8, among which, high (7–8) and low (4–5) copy numbers of tandem repeats were found in haplotypes H2 and H17, respectively. The nucleotide diversity of pvs28 (π = 0.00305 ± 0.00061) was slightly higher than that of pvs25 (π = 0.00146 ± 0.00007), thus they were not significantly different (P > 0.05). The Tajima's D value of pvs25 was positive whereas pvs28 was negative, which indicated that both genes were affected by natural selection. Conclusion The genetic diversity of pvs25 and pvs28 genes in China was relatively limited, which provided valuable information for TBVs design and optimization.

Abstract Background Continuous tilling and the lateral growth of rhizomes confer rhizomatous grasses with the unique ability to laterally expand, migrate and resist disturbances. They play key roles especially in degraded grasslands, deserts, sand dunes, and other fragile ecological system. The rhizomatous plant Leymus secalinus has both rhizome buds and tiller buds that grow horizontally and upward at the ends of rhizome differentiation and elongation, respectively. The mechanisms of rhizome formation and differentiation in L. secalinus have not yet been clarified. Results In this study, we found that the content of gibberellin A3 (GA3) and indole-3-acetic acid (IAA) were significantly higher in upward rhizome tips than in horizontal rhizome tips; by contrast, the content of methyl jasmonate and brassinolide were significantly higher in horizontal rhizome tips than in upward rhizome tips. GA3 and IAA could stimulate the formation and turning of rhizomes. An auxin efflux carrier gene, LsPIN1, was identified from L. secalinus based on previous transcriptome data. The conserved domains of LsPIN1 and the relationship of LsPIN1 with PIN1 genes from other plants were analyzed. Subcellular localization analysis revealed that LsPIN1 was localized to the plasma membrane. The length of the primary roots (PRs) and the number of lateral roots (LRs) were higher in Arabidopsis thaliana plants overexpressing LsPIN1 than in wild-type (Col-0) plants. Auxin transport was altered and the gravitropic response and phototropic response were stronger in 35S:LsPIN1 transgenic plants compared with Col-0 plants. It also promoted auxin accumulation in root tips. Conclusion Our findings indicated that LsPIN1 plays key roles in auxin transport and root development. Generally, our results provide new insights into the regulatory mechanisms underlying rhizome development in L. secalinus.

Abstract Background Cold stress is one of the main abiotic stresses limiting cucumber (Cucumis sativus L.) growth and production. C-repeat binding factor/Dehydration responsive element-binding 1 protein (CBF/DREB1), containing conserved APETALA2 (AP2) DNA binding domains and two characteristic sequences, are key signaling genes that can be rapidly induced and play vital roles in plant response to low temperature. However, the CBF family has not been systematically elucidated in cucumber, and the expression pattern of this family genes under cold stress remains unclear. Results In this study, three CsCBF family genes were identified in cucumber genome and their protein conserved domain, protein physicochemical properties, gene structure and phylogenetic analysis were further comprehensively analyzed. Subcellular localization showed that all three CsCBFs were localized in the nucleus. Cis-element analysis of the promoters indicated that CsCBFs might be involved in plant hormone response and abiotic stress response. Expression analysis showed that the three CsCBFs could be significantly induced by cold stress, salt and ABA. The overexpression of CsCBFs in cucumber seedlings enhanced the tolerance to cold stress, and importantly, the transcript levels of CsCOR genes were significantly upregulated in 35S:CsCBFs transgenic plants after cold stress treatment. Biochemical analyses ascertained that CsCBFs directly activated CsCOR genes expression by binding to its promoter, thereby enhancing plant resistance to cold stress. Conclusion This study provided a foundation for further research on the function of CsCBF genes in cold stress resistance and elucidating its mechanism.

Abstract Traumatic brain injury (TBI) is a major public health problem worldwide which causes high mortality and disability. Functioning as microRNA (miRNA) sponges, long non-coding RNA (lncRNA) regulates the expression of protein-coding genes in a competing endogenous RNA (ceRNA) network. However, the lncRNA-associated ceRNA in TBI remains unclear. In this study, we processed the raw SRR files of mice cortex samples of sham injury (n = 3) and TBI groups (n = 3) to count files. Then, the expression profiles of lncRNAs and mRNAs were identified, and 86 differentially expressed (DE) lncRNAs and 1201 DEmRNAs between sham and TBI groups were identified. The DEmRNAs were used to perform enrichment analyses. Next, a lncRNA-miRNA-mRNA regulatory ceRNA network was constructed. The network consisted of 23 mRNAs, 5 miRNAs and 2 lncRNAs. The expression alternations of the 5 miRNAs were validated via qRT-PCR. The subnetwork of hub lncRNA Neat1 was extracted. We identified a potential inflammatory associated regulatory axis: Neat1/miR-31-5p/Myd88 axis. The PPI network based on DEmRNA involved in ceRNA network was constructed PPI networks to identify the hub genes. Finally, DElncRNAs and DEmRNAs were selected randomly and validated by qRT-PCR. In conclusion, with the lncRNA-miRNA-mRNA ceRNA network provided above, we can improve our understanding of the regulatory mechanisms and interaction among lncRNAs, miRNAs and mRNAs in TBI process.

Abstract Background The unripe fruits of Rubus chingii Hu. (“Fu-peng-zi” in Chinese) is a well-known herbal tonic in traditional Chinese medicine (TCM) for tonifying liver and kidney. However, little is known regarding its therapeutic efficacy against liver fibrosis and the underlying mechanism. Methods The current research aims to explore the potential of Rubus chingii Hu. unripe fruits extract (RF) in the treatment of liver fibrosis and explore the underlying mechanism. RF was administered (450 and 900 mg·kg− 1 of body weight per day) orally to male C57BL/6 mice with CCl4-induced liver fibrosis for 3 weeks. The histopathological changes and fibrosis stage in liver tissue were assessed using hematoxylin and eosin (H&E) and Sirius red staining. The distribution of α-SMA and Col1A1 in the liver was analyzed to determine the hepatic stellate cells (HSCs) activation using immunohistochemistry and immunofluorescent analysis. Various biochemical markers in serum (ALT, AST) and liver (Hyp, IL1-β, IL6, TNF-α and MCP-1) were observed to assess the liver’s injury, fibrosis, and inflammation. In liver tissue, fibrosis-associated proteins including α-SMA, TGF-β1, Smad2/3, p-Smad2/3, and Smad4 were detected through a Western blot assay. Pyrosequencing-based analysis of bacterial 16 S ribosomal RNA from variable regions V3–V4 of fecal samples characterized the gut microbiota. Spearman’s rank correlation analysis was performed for the association between altered bacterial genera by RF and pharmacodynamics parameters. Results Three weeks of RF treatment can significantly lower liver inflammatory levels, pathological abnormalities, and collagen fibrous deposition in mice with CCl4-induced liver fibrosis. The expressions of α-SMA and Col1A1 were lowered by RF, while the expression levels of TGF-β/Smads signaling pathway-related proteins, including TGF-β1, p-Smad2/3, and Smad4, were dramatically decreased by RF. The RF treatment significantly increased or reduced 18 different bacterial species, restoring the CCl4-induced gut microbiota imbalance to the normal group’s levels. According to correlation analysis, the bacterial genera Bifidobacterium and Turicibacter were the most significant in restoring CCl4-induced liver fibrosis. Conclusions RF can reduce liver damage and delay the onset of liver fibrosis through modulating TGF-β/Smads signaling pathway. Furthermore, RF’s anti-liver fibrosis effect was related to balancing the gut microbial community, partly attained by increasing Bifidobacterium and Turicibacter in liver fibrosis.

Abstract Blockade of programmed cell death 1 ligand (PD-L1) has been used to treat triple-negative breast cancer (TNBC), and various strategies are under investigation to improve the treatment response rate. Inhibition of glutamine metabolism can reduce the massive consumption of glutamine by tumor cells and meet the demand for glutamine by lymphocytes in tumors, thereby improving the anti-tumor effect on the PD-L1 blockade therapy. Here, molybdenum disulfide (MoS2) was employed to simultaneously deliver anti-PDL1 antibody (aPDL1) and V9302 to boost the anti-tumor immune response in TNBC cells. The characterization results show that MoS2 has a dispersed lamellar structure with a size of about 181 nm and a size of 232 nm after poly (l-lysine) (PLL) modification, with high stability and biocompatibility. The loading capacity of aPDL1 and V9302 are 3.84% and 24.76%, respectively. V9302 loaded MoS2 (MoS2-V9302) can effectively kill 4T1 cells and significantly reduce glutamine uptake of tumor cells. It slightly increases CD8+ cells in the tumor and promotes CD8+ cells from the tumor edge into the tumor core. In vivo studies demonstrate that the combination of aPDL1 and V9302 (MoS2-aPDL1-V9302) can strongly inhibit the growth of TNBC 4T1 tumors. Interestingly, after the treatment of MoS2-aPDL1-V9302, glutamine levels in tumor interstitial fluid increased. Subsequently, subtypes of cytotoxic T cells (CD8+) in the tumors were analyzed according to two markers of T cell activation, CD69, and CD25, and the results reveal a marked increase in the proportion of activated T cells. The levels of cytokines in the corresponding tumor interstitial fluid are also significantly increased. Additionally, during the treatment, the body weights of the mice remain stable, the main indicators of liver and kidney function in the blood do not increase significantly, and there are no obvious lesions in the main organs, indicating low systemic toxicity. In conclusion, our study provides new insights into glutamine metabolism in the tumor microenvironment affects immune checkpoint blockade therapy in TNBC, and highlights the potential clinical implications of combining glutamine metabolism inhibition with immune checkpoint blockade in the treatment of TNBC. Graphical Abstract

Abstract Background AdpA is a global regulator of morphological differentiation and secondary metabolism in Streptomyces, but the regulatory roles of the Streptomyces AdpA family on the biosynthesis of the natural product ε-poly-l-lysine (ε-PL) remain unidentified, and few studies have focused on increasing the production of ε-PL by manipulating transcription factors in Streptomyces. Results In this study, we revealed the regulatory roles of different AdpA homologs in ε-PL biosynthesis and morphological differentiation and effectively promoted ε-PL production and sporulation in Streptomyces albulus NK660 by heterologously expressing adpA from S. neyagawaensis NRRLB-3092 (adpA Sn ). First, we identified a novel AdpA homolog named AdpA Sa in S. albulus NK660 and characterized its function as an activator of ε-PL biosynthesis and morphological differentiation. Subsequently, four heterologous AdpA homologs were selected to investigate their phylogenetic relationships and regulatory roles in S. albulus, and AdpA Sn was demonstrated to have the strongest ability to promote both ε-PL production and sporulation among these five AdpA proteins. The ε-PL yield of S. albulus heterologously expressing adpA Sn was approximately 3.6-fold higher than that of the control strain. Finally, we clarified the mechanism of AdpA Sn in enhancing ε-PL biosynthesis and its effect on ε-PL polymerization degree using real-time quantitative PCR, microscale thermophoresis and MALDI-TOF–MS. AdpA Sn was purified, and its seven direct targets, zwf, tal, pyk2, pta, ack, pepc and a transketolase gene (DC74_2409), were identified, suggesting that AdpA Sn may cause the redistribution of metabolic flux in central metabolism pathways, which subsequently provides more carbon skeletons and ATP for ε-PL biosynthesis in S. albulus. Conclusions Here, we characterized the positive regulatory roles of Streptomyces AdpA homologs in ε-PL biosynthesis and their effects on morphological differentiation and reported for the first time that AdpA Sn promotes ε-PL biosynthesis by affecting the transcription of its target genes in central metabolism pathways. These findings supply valuable insights into the regulatory roles of the Streptomyces AdpA family on ε-PL biosynthesis and morphological differentiation and suggest that AdpA Sn may be an effective global regulator for enhanced production of ε-PL and other valuable secondary metabolites in Streptomyces.

Abstract Background Avian hepatitis E virus (HEV) is the pathogenic agent of big liver and spleen disease (BLS) and of hepatitis-splenomegaly syndrome (HSS) in chickens, which have caused economic losses to the poultry industry in China. In this study, 18 samples of BLS chickens were collected to reveal the molecular epidemiological characteristics of avian HEV in the province of Shandong, China. Results Gross and microscopic lesions of clinical samples were observed; then, virology detection and genetic analysis of avian HEV were performed. The results showed that there was significant swelling and rupture in the liver and that the spleen was enlarged. Microscopic lesions demonstrated obvious hemorrhage in the liver, with infiltration of heterophilic granulocytes, lymphocytes, and macrophages, as well as the reduction of lymphocytes in the spleen. Eleven of the 18 samples were positive for avian HEV, with a positive rate of 61.11%. More importantly, all avian HEV-positive samples were mixed infections: among these, the mixed infections of avian HEV and chicken infectious anemia virus (CIAV) and avian HEV and fowl adenovirus (FAdV) were the most common. Furthermore, the genetic evolution analysis showed that all avian HEV strains obtained here did not belong to the reported 4 genotypes, thus constituting a potential novel genotype. Conclusions These results of this study further enrich the epidemiological data on avian HEV in Shandong, prove the genetic diversity of avian HEV in China, and uncover the complex mixed infections of avian HEV clinical samples.

Abstract Background Due to the lack of enough interaction data among compositions, targets and diseases, it is difficult to construct a complete network of Traditional Chinese Medicine (TCM) that comprehensively reflects active compositions and their synergistic network in terms of specific diseases. Therefore, mapping of the full spectrum of interaction between compounds and their targets is of central importance when we use network pharmacology approach to explore the therapeutic potential of the TCM. Methods To address this challenge, we developed a large-scale simultaneous interaction prediction approach (SiPA) integrated one interaction network based simple inference model (SIM), focusing on ‘logical relevance’ between compounds, proteins or diseases, and another compound-target correlation space based interaction prediction model (CTCS-IPM) that was built on the basis of the canonical correlation analysis (CCA) to estimate the position of compounds (or targets) in compound-protein correlated space. Then SiPA was applied to discover reliable multiple interactions for interaction network expansion of a TCM, compound Salvia miltiorrhiza. By means of network analysis, potential active compounds and their related network synergy underlying cardiovascular diseases were evaluated between expanded and original interaction networks. Part of new interactions were validated with existing experimental evidence and molecular docking. Results As evaluated with known test dataset, the established combination approach was proved to make highly accurate prediction, showing a well prediction performance for the SIM and a high recall rate of 85.2% for the CTCS-IPM. Then 710 pairs of new compound-target interactions, 24 pairs of new compound-cardiovascular disease interactions and 294 pairs of new cardiovascular disease-protein interactions were predicted for compound Salvia miltiorrhiza. Results of network analysis suggested the network expansion could dramatically improve the completeness and effectiveness of the network. Validation results of literature and molecular docking manifested that inferred interactions had good reliability. Conclusions We provided a practical and efficient way for large-scale inference of multiple interactions of TCM ingredients, which was not limited by the lack of negative samples, sample size and target 3D structures. SiPA could help researchers more accurately prioritize the effective compounds and more completely explore network synergy of TCM for treating specific diseases, indicating a potential way for effectively identifying candidate compound (or target) in drug discovery.

Abstract Background Air pollution increases the morbidity and mortality of cardiovascular disease (CVD). Atherosclerosis (AS) is the pathological basis of most CVD, and the progression of atherosclerosis and the increase of fragile plaque rupture are the mechanism basis of the relationship between atmospheric particulate pollution and CVD. The aim of the present study was to investigate the effects of coal-fired fine particulate matter (PM2.5) on the expression levels of atherosclerosis-related proteins (von Willebrand factor (vWF), Endothelin-1 (ET-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin, and to explore the role and mechanism of the progression of atherosclerosis induced by coal-fired PM2.5 via the mitogen-activated protein kinase (MAPK) signaling pathways. Methods Different concentrations of PM2.5 were given to apolipoprotein-E knockout (ApoE−/−) mice via intratracheal instillation for 8 weeks. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of vWF, ET-1 in serum of mice. Immunohistochemistry was used to observe the expression and distribution of ICAM-1 and E-selectin in the aorta of mice. Western blot was used to investigate the phosphoylation of proteins relevant to MAPK signaling pathways. Results Coal-fired PM2.5 exacerbated atherosclerosis induced by a high-fat diet. Fibrous cap formation, foam cells accumulation, and atherosclerotic lesions were observed in the aortas of PM2.5-treated mice. Coal-fired PM2.5 increased the protein levels of ET-1, ICAM-1, and E-selectin, but there was no significant difference in the vWF levels between the PM2.5-treatment mice and the HFD control mice. Coal-fired PM2.5 promoted the phosphorylation of p38, c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) in aortic tissues of mice. Conclusion Coal-derived PM2.5 exacerbated the formation of atherosclerosis in mice, increased the expression levels of atherosclerosis-related proteins in mice serum, and promoted the phosphorylation of proteins relevant to MAPK signaling pathway. Thus, MAPK signaling pathway may play a role in the atherosclerosis pathogenesis induced by Coal-derived PM2.5.

Abstract Background Endoglucanase has been extensively employed in industrial processes as a key biocatalyst for lignocellulosic biomass degradation. Thermostable endoglucanases with high catalytic activity at elevated temperatures are preferred in industrial use. To improve the activity and thermostability, site-directed mutagenesis was conducted to modify the N-glycosylation sites of the thermostable β-1,4-endoglucanase CTendo45 from Chaetomium thermophilum. Results In this study, structure-based rational design was performed based on the modification of N-glycosylation sites in CTendo45. Eight single mutants and one double mutant were constructed and successfully expressed in Pichia pastoris. When the unique N-glycosylation site of N88 was eliminated, a T90A variant was active, and its specific activity towards CMC-Na and β-d-glucan was increased 1.85- and 1.64-fold, respectively. The mutant R67S with an additional N-glycosylation site of N65 showed a distinct enhancement in catalytic efficiency. Moreover, T90A and R67S were endowed with extraordinary heat endurance after 200 min of incubation at different temperatures ranging from 30 to 90 °C. Likewise, the half-lives (t 1/2) indicated that T90A and R67S exhibited improved enzyme thermostability at 80 °C and 90 °C. Notably, the double-mutant T90A/R67S possessed better hydrolysis activity and thermal stability than its single-mutant counterparts and the wild type. Conclusions This study provides initial insight into the biochemical function of N-glycosylation in thermostable endoglucanases. Moreover, the design approach to the optimization of N-glycosylation sites presents an effective and feasible strategy to improve enzymatic activity and thermostability.

Abstract Background The mechanism underlying sorafenib resistance in hepatocellular carcinoma (HCC) remains unclear. Accumulating evidence suggests that tumor-initiating cells (TICs) are a pivotal driving force. Both CD44 and Hedgehog signaling play crucial roles in TIC properties in HCC. In this study, we explored the roles of CD44 and Hedgehog signaling in sorafenib resistance and evaluated the therapeutic effect of cotreatment with sorafenib and Hedgehog signaling inhibitors in HCC patient-derived organoid (PDO) models to improve treatment efficacy. Methods We collected HCC specimens to establish PDO models. Cell viability and malignant transformation properties were investigated after treatment with different TIC-related inhibitors alone or in combination with sorafenib to evaluate the therapeutic effect in PDOs and cell lines by in vitro and in vivo experiments. Expression levels of Hedgehog signaling proteins and CD44 were monitored to reveal potential relationships. Results We demonstrated that our HCC PDO models strongly maintained the histological features of the corresponding tumors and responded to drug treatment. Furthermore, CD44-positive HCC PDOs were obviously resistant to sorafenib, and sorafenib increased CD44 levels. A drug screen showed that compared with Notch, Hippo and Wnt signaling inhibitors, a Hedgehog signaling inhibitor (GANT61) potently suppressed HCC PDO cell viability. In addition, there was a highly synergistic effect in vitro and in vivo on the suppression of cell viability and malignant properties when sorafenib and GANT61 were added to CD44-positive HCC PDOs and cell lines, respectively. Furthermore, the upregulation of CD44 and Hedgehog signaling induced by sorafenib was reversed by GANT61. Conclusions GANT61 significantly suppressed Hedgehog signaling to reverse sorafenib resistance in CD44-positive HCC. The combination of sorafenib and Hedgehog signaling inhibitors might be effective in HCC patients with high CD44 levels as a personalized-medicine approach.

Abstract Background Wheat straw is a rich resource worldwide. Straw return is an effective strategy to alleviate soil-borne diseases on monoculture watermelon. Previous studies focus on soil structure, physical and chemical properties; however, little is known about the molecular responses on host plant. Results No significant difference on the population of Fusarium oxysporum f.sp. niveum race 1(Fon1) in rhizosphere soil was found between control (no addition of wheat straw) and the treated groups (addition of 1% (T1) or 2% (T2) wheat straw). RNA-Seq analysis showed that 3419 differentially expressed genes were clustered into 8 profiles. KEGG analysis revealed that phenylpropanoid biosynthesis and plant hormone signal transduction were involved in wheat straw induced response in monoculture watermelon. Genes in lignin biosynthesis were found to be upregulated, and the lignin and auxin contents were higher in T1 and T2 compared to the control. Lignin was also enriched and the Fon1 population decreased in watermelon roots treated with wheat straw. The enzyme activities of phenylalanine ammonia-lyase and peroxidase were increased. Conclusions Our data suggest that the addition of wheat straw enhances the defense response to Fon1 infection in watermelon through increasing lignin and auxin biosynthesis.

Abstract Background Glioblastomas have a high degree of malignancy, high recurrence rate, high mortality rate, and low cure rate. Searching for new markers of glioblastomas is of great significance for improving the diagnosis, prognosis and treatment of glioma. Methods Using the GEO public database, we combined 34 glioma microarray datasets containing 1893 glioma samples and conducted genetic data mining through statistical analysis, bioclustering, and pathway analysis. The results were validated in TCGA, CGGA, and internal cohorts. We further selected a gene for subsequent experiments and conducted cell proliferation and cell cycle analyses to verify the biological function of this gene. Results Eight glioblastoma-specific differentially expressed genes were screened using GEO. In the TCGA and CGGA cohorts, patients with high CBX3, BARD1, EGFR, or IFRD1 expression had significantly shorter survival but patients with high GUCY1A3 or MOBP expression had significantly longer survival than patients with lower expression of these genes. After reviewing the literature, we selected the CBX3 gene for further experiments. We confirmed that CBX3 was overexpressed in glioblastoma by immunohistochemical analysis of tissue microarrays and qPCR analysis of surgical specimens. The functional assay results showed that silencing CBX3 arrests the cell cycle in the G2/M phase, thereby weakening the cell proliferation ability. Conclusions We used a multidisciplinary approach to analyze glioblastoma samples in 34 microarray datasets, revealing novel diagnostic and prognostic biomarkers in patients with glioblastoma and providing a new direction for screening tumor markers.

Abstract Background Erythritol is a four-carbon sugar alcohol with sweetening properties that is used by the agro-food industry as a food additive. In the yeast Yarrowia lipolytica, the last step of erythritol synthesis involves the reduction of erythrose by specific erythrose reductase(s). In the earlier report, an erythrose reductase gene (YALI0F18590g) from erythritol-producing yeast Y. lipolytica MK1 was identified (Janek et al. in Microb Cell Fact 16:118, 2017). However, deletion of the gene in Y. lipolytica MK1 only resulted in some lower erythritol production but the erythritol synthesis process was still maintained, indicating that other erythrose reductase gene(s) might exist in the genome of Y. lipolytica. Results In this study, we have isolated genes g141.t1 (YALI0D07634g) and g3023.t1 (YALI0C13508g) encoding two novel erythrose reductases (ER). The biochemical characterization of the purified enzymes showed that they have a strong affinity for erythrose. Deletion of the two ER genes plus g801.t1 (YALI0F18590g) did not prevent erythritol synthesis, suggesting that other ER or ER-like enzymes remain to be discovered in this yeast. Overexpression of the newly isolated two genes (ER10 or ER25) led to an average 14.7% higher erythritol yield and 31.2% higher productivity compared to the wild-type strain. Finally, engineering NADPH cofactor metabolism by overexpression of genes ZWF1 and GND1 encoding glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, respectively, allowed a 23.5% higher erythritol yield and 50% higher productivity compared to the wild-type strain. The best of our constructed strains produced an erythritol titer of 190 g/L in baffled flasks using glucose as main carbon source. Conclusions Our results highlight that in the Y. lipolytica genome several genes encode enzymes able to reduce erythrose into erythritol. The catalytic properties of these enzymes and their cofactor dependency are different from that of already known erythrose reductase of Y. lipolytica. Constitutive expression of the newly isolated genes and engineering of NADPH cofactor metabolism led to an increase in erythritol titer. Development of fermentation strategies will allow further improvement of this productivity in the future.