The global succinylation of SARS-CoV-2-infected host cells reveals drug targets.

SARS-CoV-2, the causative agent of the COVID-19 pandemic, undergoes continuous evolution, highlighting an urgent need for development of novel antiviral therapies. Here we show a quantitative mass spectrometry-based succinylproteomics analysis of SARS-CoV-2 infection in Caco-2 cells, revealing dramatic reshape of succinylation on host and viral proteins. SARS-CoV-2 infection promotes succinylation of several key enzymes in the TCA, leading to inhibition of cellular metabolic pathways. We demonstrated that host protein succinylation is regulated by viral nonstructural protein (NSP14) through interaction with sirtuin 5 (SIRT5); overexpressed SIRT5 can effectively inhibit virus replication. We found succinylation inhibitors possess significant antiviral effects. We also found that SARS-CoV-2 nucleocapsid and membrane proteins underwent succinylation modification, which was conserved in SARS-CoV-2 and its variants. Collectively, our results uncover a regulatory mechanism of host protein posttranslational modification and cellular pathways mediated by SARS-CoV-2, which may become antiviral drug targets against COVID-19.

Targeted delivery of CEBPA-saRNA for the treatment of pancreatic ductal adenocarcinoma by transferrin receptor aptamer decorated tetrahedral framework nucleic acid.

Pancreatic cancer, predominantly pancreatic ductal adenocarcinoma (PDAC), remains a highly lethal malignancy with limited therapeutic options and a dismal prognosis. By targeting the underlying molecular abnormalities responsible for PDAC development and progression, gene therapy offers a promising strategy to overcome the challenges posed by conventional radiotherapy and chemotherapy. This study sought to explore the therapeutic potential of small activating RNAs (saRNAs) specifically targeting the CCAAT/enhancer-binding protein alpha (CEBPA) gene in PDAC. To overcome the challenges associated with saRNA delivery, tetrahedral framework nucleic acids (tFNAs) were rationally engineered as nanocarriers. These tFNAs were further functionalized with a truncated transferrin receptor aptamer (tTR14) to enhance targeting specificity for PDAC cells. The constructed tFNA-based saRNA formulation demonstrated exceptional stability, efficient saRNA release ability, substantial cellular uptake, biocompatibility, and nontoxicity. In vitro experiments revealed successful intracellular delivery of CEBPA-saRNA utilizing tTR14-decorated tFNA nanocarriers, resulting in significant activation of tumor suppressor genes, namely, CEBPA and its downstream effector P21, leading to notable inhibition of PDAC cell proliferation. Moreover, in a mouse model of PDAC, the tTR14-decorated tFNA-mediated delivery of CEBPA-saRNA effectively upregulated the expression of the CEBPA and P21 genes, consequently suppressing tumor growth. These compelling findings highlight the potential utility of saRNA delivered via a designed tFNA nanocarrier to induce the activation of tumor suppressor genes as an innovative therapeutic approach for PDAC.

Oroxylin a glucuronide as a novel class of reversible inhibitors of Sortase a, combats MRSA-induced infections.

AIMS: The main purpose of this study was to study the therapeutical effect of oroxylin A glucuronide (OAG) on methicillin-resistant Staphylococcus aureus (MRSA). METHODS AND RESULTS: By substrate peptide reaction-based fluorescence resonance energy transfer (FRET) screening, we identified that OAG was an efficient inhibitor of Sortase A (SrtA) with an IC50 of 45.61 µg mL-1, and achieved efficacy in the treatment of Staphylococcus aureus (S. aureus) infections. We further demonstrated that OAG inhibited the adhesion of the S. aureus to fibrinogen, the surface protein A anchoring and diminished biofilm formation. Results obtained from fluorescence quenching assay elucidated a direct interaction between OAG and SrtA. Employing molecular dynamics simulations, we proved that OAG binds to the binding sites of R197, G192, E105, and V168 in the SrtA. Notably, OAG exhibited a robust therapeutic effect in a MRSA-induced pneumonia model. CONCLUSIONS: We identified that OAG as a novel class of reversible inhibitors of SrtA, combats MRSA-induced Infections.

Punicalagin, an Inhibitor of Sortase A, Is a Promising Therapeutic Drug to Combat Methicillin-Resistant Staphylococcus aureus Infections.

Antimicrobial resistance (AMR) poses a major threat to human health globally. Staphylococcus aureus is recognized as a cause of disease worldwide, especially methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA). The enzyme sortase A (SrtA), present on the cell surface of S. aureus, plays a key role in bacterial virulence without affecting the bacterial viability, and SrtA-deficient S. aureus strains do not affect the growth of bacteria. Here, we found that punicalagin, a natural compound, was able to inhibit SrtA activity with a very low half maximal inhibitory concentration (IC50) value of 4.23 µg/mL, and punicalagin is a reversible inhibitor of SrtA. Moreover, punicalagin has no distinct cytotoxicity toward A549, HEK293T, or HepG2 cells at a much higher concentration than the IC50 detected by MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide] assays. In addition, punicalagin visibly attenuated the virulence-related phenotype of SrtA in vitro by decreasing adhesion of S. aureus to fibrinogen, reducing the ability of protein A (SpA) displayed on the surface of the bacteria and biofilm formation. Fluorescence quenching elucidated the interaction between punicalagin and SrtA. Molecular docking further implied that the inhibitory activity lay in the bond between punicalagin and SrtA residues LYS190, TYR187, ALA104, and GLU106. In In vivo studies, we surprisingly found that punicalagin had a more effective curative effect combined with cefotaxime when mice were infected with pneumonia caused by MRSA. Essentially, punicalagin, a therapeutic compound targeting SrtA, demonstrates great potential for combating MRSA infections.

Hinokiflavone Attenuates the Virulence of Methicillin-Resistant Staphylococcus aureus by Targeting Caseinolytic Protease P.

Drug-resistant bacteria was the third leading cause of death worldwide in 2019, which sounds like a cautionary note for global public health. Therefore, developing novel strategies to combat Methicillin-resistant Staphylococcus aureus (MRSA) infections is the need of the hour. Caseinolytic protease P (ClpP) represents pivotal microbial degradation machinery in MRSA involved in bacterial homeostasis and pathogenicity, considered an ideal target for combating S. aureus infections. Herein, we identified a natural compound, hinokiflavone, that inhibited the activity of ClpP of MRSA strain USA300 with an IC50 of 34.36 µg/mL. Further assays showed that hinokiflavone reduced the virulence of S. aureus by inhibiting multiple virulence factors expression. Results obtained from cellular thermal transfer assay (CETSA), thermal shift assay (TSA), local surface plasmon resonance (LSPR) and molecular docking (MD) assay enunciated that hinokiflavone directly bonded to ClpP with confirmed docking sites, including SER-22, LYS-26 and ARG-28. In vivo, the evaluation of anti-infective activity showed that hinokiflavone in combination with vancomycin effectively protected mice from MRSA-induced fatal pneumonia, which was more potent than vancomycin alone. As mentioned above, hinokiflavone, as an inhibitor of ClpP, could be further developed into a promising adjuvant against S. aureus infections.

Tamarixetin Attenuated the Virulence of Staphylococcus aureus by Directly Targeting Caseinolytic Protease P.

Staphylococcus aureus, especially drug-resistant S. aureus infections, is a worldwide healthcare challenge. There is a growing focus on antivirulence therapy against S. aureus. Caseinolytic protease p (ClpP) is a protein hydrolase essential for pathogenicity in S. aureus. A flavonoid compound, tamarixetin, which was screened in this work, was specifically able to inhibit the hydrolytic activity of ClpP on the fluorescent substrate Suc-LY-AMC with an IC50 of 49.73 µM, without affecting the growth of methicillin-resistant S. aureus strain USA300 and was without obvious cytotoxicity. Further assays found that tamarixetin inhibited the transcription of hla, agr, RNAIII, pvl, PSM-α, and spa genes as well as suppressed the protein expression levels of Hla and PVL. Moreover, tamarixetin was observed to dramatically inhibit the hemolytic activity of hla in S. aureus. Consistent with that of S. aureus USA300-ΔclpP, tamarixetin was shown to increase urease expression. The thermal shift and cellular thermal shift assays showed that tamarixetin markedly changed the thermal stability of ClpP. The dissociation constant (KD) value of tamarixetin with ClpP was 2.52 × 10-6 M measured by surface plasmon resonance. The molecular docking and ClpP point mutation results also demonstrated that tamarixetin had a strong interaction with ClpP. In vivo study showed that tamarixetin was effective in protecting mice from S. aureus pneumonia by increasing survival, reducing lung tissue load, and slowing down the infiltration of inflammatory factors. In addition, tamarixetin was able to enhance the antibacterial activity of cefotaxime in combination. In conclusion, tamarixetin was promising as a ClpP inhibitor for S. aureus infections.

WWP2 ameliorates oxidative stress and inflammation in atherosclerotic mice through regulation of PDCD4/HO-1 pathway.

WWP2 is a HECT-type E3 ubiquitin ligase that regulates various physiological and pathological activities by binding to different substrates, but its role in atherosclerosis (AS) remains largely unknown. The objective of the present study is to investigate the role and underlying molecular mechanisms of WWP2 in endothelial injury. We found that WWP2 expression is significantly decreased in Apolipoprotein E (ApoE) -/- mice. Overexpression of WWP2 attenuates oxidative stress and inflammation in AS mice, while knockdown of WWP2 has opposite effects. WWP2 overexpression alleviates oxidized low-density lipoprotein (ox-LDL)-induced human umbilical vein endothelial cell (HUVEC) injury, evidenced by the decreased oxidative stress levels and the secretion of inflammatory cytokines. Programmed cell death 4 (PDCD4) is identified as a potential substrate of WWP2. Co-immunoprecipitation (Co-IP) further demonstrates that WWP2 interacts with PDCD4, which is enhanced by ox-LDL treatment. Furthermore, the level of PDCD4 ubiquitination is significantly increased by WWP2 overexpression under the condition of MG132 treatment, while WWP2 knockdown shows opposite results. Subsequently, rescue experiments demonstrate that WWP2 knockdown further aggravates oxidative stress and inflammation in ox-LDL-treated HUVECs, while knockdown of PDCD4 alleviates this effect. Moreover, the use of sn-protoporphyrin (SnPP), an inhibitor of HO-1 pathway, confirms that PDCD4 enhances endothelial injury induced by ox-LDL through inhibiting HO-1 pathway. In conclusion, our results suggest that WWP2 protects against atherosclerosis progression via the PDCD4/HO-1 pathway, which may provide a novel treatment strategy for atherosclerosis.

The protection effect of rhodionin against methicillin-resistant Staphylococcus aureus-induced pneumonia through sortase A inhibition.

Methicillin-resistant Staphylococcus aureus (MRSA) is a zoonotic antibiotic-resistant pathogen that negatively impacts society from medical, veterinary, and societal standpoints. The search for alternative therapeutic strategies and innovative anti-infective agents is urgently needed. Among the pathogenic mechanisms of Staphylococcus aureus (S. aureus), sortase A is a virulence factor of great concern because it is highly linked with the ability of MRSA to invade the host. In this study, we identified that rhodionin, a natural compound of flavonoid glucosides, effectively inhibited the activity of SrtA without affecting the survival and growth of bacteria, and its half maximal inhibitory concentration (IC50) value was 22.85 µg/mL. In vitro, rhodionin prominently attenuated the virulence-related phenotype of SrtA by reducing the adhesion of S. aureus to fibrinogen, reducing the capacity of protein A (SpA) on the bacterial surface and biofilm formation. Subsequently, fluorescence quenching and molecular docking were performed to verify that rhodionin directly bonded to SrtA molecule with KA value of 6.22 × 105 L/mol. More importantly, rhodionin showed a significant protective effect on mice pneumonia model and improved the survival rate of mice. According to the above findings, rhodionin achieved efficacy in the treatment of MRSA-induced infections, which holds promising potential to be developed into a candidate used for MRSA-related infections.

Effects of high glucose and insulin on the electrophysiological properties of cardiomyocytes derived from human-induced pluripotent stem cells. / é«˜ç³–ä¸Žèƒ°å²›ç´ å¯¹äººè¯±å¯¼å¤šæ½œèƒ½å¹²ç»†èƒžæ¥æºçš„å¿ƒè‚Œç»†èƒžç”µç”Ÿç†ç‰¹æ€§çš„å½±å“.

OBJECTIVES: The risk of arrhythmia increases in diabetic patients. However, the effects of hyperglycemia and insulin therapy on the electrophysiological properties of human cardiomyocytes remain unclear. This study is to explore the effects of high glucose and insulin on the electrophysiological properties and arrhythmias of cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CMs). METHODS: Immunofluorescent staining and flow cytometry were used to analyze the purity of hiPSC-CMs generated from human skin fibroblasts of a healthy donor. The hiPSC-CMs were divided into 3 group (treated with normal medium, high glucose and insulin for 4 days): a control group (NM group, containing 5 mmol/L glucose), a high glucose group (HG group, containing 15 mmol/L glucose), and a high glucose combined with insulin (HG+INS group, containing 15 mmol/L glucose+100 mg/L insulin). Electrophysiological changes of hiPSC-CMs were detected by microelectrode array (MEA) before or after treatment with glucose and insulin, including beating rate (BR), field potential duration (FPD) (similar to QT interval in ECG), FPDc (FPD corrected by BR), spike amplitude and conduction velocity (CV). Effects of sotalol on electrophysiological properties and arrhythmias of hiPSC-CMs were also evaluated. RESULTS: The expression of cardiac-specific marker of cardiac troponin T was high in the hiPSC-CMs. The purity of hiPSC-CMs was 99.06%. Compared with the NM group, BR was increased by (9.14±0.8)% in the HG group (P<0.01). After treatment with high glucose, FPD was prolonged from (460.4±9.0) ms to (587.6±23.7) ms in the HG group, while it was prolonged from (462.5±14.5) ms to (512.6±17.6) ms in the NM group. Compared with the NM group, FPD of hiPSC-CMs was prolonged by (16.8±1.4)% in the HG group (P<0.01). The FPDc of hiPSC-CMs was prolonged from (389.1±13.7) ms to (478.3±31.5) ms in the HG group, and that was prolonged from (387.7±21.6) ms to (422.6±32.9) ms in the NM group. Compared with the NM group, the FPDc of hiPSC-CMs was prolonged by (13.9±1.3)% in HG group (P<0.01). The spike amplitude and CV remained unchanged between the HG group and the NM group (P>0.05). Ten µmol/L of sotalol can induce significant arrhythmias from all wells in the HG group. After treatment with insulin and high glucose, compared with the HG group, BR was increased by (8.3±0.5)% in the HG+INS group (P<0.05). The FPD was prolonged from (463.4±9.7) ms to (532.6±12.8) ms in the HG+INS group, while it was prolonged from (460.4±9.0) ms to (587.6±23.7) ms in the HG group. Compared with the HG group, the FPD of hiPSC-CMs was shortened by (12.7±1.9)% in the HG+INS group (P<0.01). The FPDc of hiPSC-CMs was prolonged from (387.4±4.1) ms to (422.4±10.0) ms in the HG+INS group, and that was prolonged from (384.8±4.0) ms to (476.3±11.5) ms in HG group. Compared with the HG group, the FPDc of hiPSC-CMs was shortened by (14.7±1.1)% in HG group (P<0.01). After the insulin treatment, the spike amplitude of hiPSC-CMs was increased from (3.12±0.46) mV to (4.35±0.64) mV in the HG+INS group, while it was enhanced from (3.06±0.35) mV to (3.33±0.41) mV in the HG group. The spike amplitude of hiPSC-CMs was increased by (30.8±3.7)% in the HG+INS group compared with that in the HG group (P<0.05). The CV in the HG+INS group was increased from (0.23±0.08) mm/ms to (0.32±0.08) mm/ms after insulin treatment, which was increased from (0.21±0.04) mm/ms to (0.30±0.07) mm/ms in the HG group, but there was no significant difference in CV between the HG+INS group and the HG group (P>0.05). The induction experiment showed that 10 µmol/L of sotalol could prolong the FPDc of hiPSC-CMs by (78.9±11.6)% in the HG+INS group, but no arrhythmia was induced in each well. CONCLUSIONS: High glucose can induce FPD/FPDc of hiPSC-CMs prolongation and increase the risk of arrhythmia induced by drugs. Insulin can reduce the FPD/FPDc prolongation and the risk of induced arrhythmia by high glucose.These results are important to understand the electrophysiological changes of the myocardium in diabetic patients and the impact of insulin therapy on its electrophysiology. Further study on the mechanism may provide new ideas and methods for the treatment of acquired and even inherited long QT syndrome.

Biochanin A as an α-hemolysin inhibitor for combating methicillin-resistant Staphylococcus aureus infection.

Methicillin-resistant Staphylococcus aureus (MRSA) is a multidrug-resistant pathogen that poses a significant risk to global health today. In S. aureus, α-hemolysin is an important virulence factor as it contributes to the capacity of the bacteria to infect the host. Here, we showed that biochanin A (bioA), an isoflavone present in red clover, cabbage and alfalfa, effectively inhibited hemolytic activity at a dose as low as 32 µg/mL. Further, western blot and RT-qPCR data showed that bioA reduced the production and expression of MRSA hemolysin in a dose-dependent manner. In addition, when different concentrations of bioA were added to a coculture system of A549 cells and S. aureus, it could significantly decrease cell injury. Importantly, the in vivo study showed that bioA could protect mice from pneumonia caused by a lethal dose of MRSA, as evidenced by improving their survival and reducing the number of bacterial colonies in lung tissues, the secretion of hemolysin into alveolar lavage fluid and the degree of pulmonary edema. In conclusion, biochanin A protected the host from MRSA infection by inhibiting the expression of the hemolysin of MRSA, which may provide experimental evidence for its development to a potential anti-MRSA drug.

The influence of porcine epidemic diarrhea virus on pig small intestine mucosal epithelial cell function.

Porcine epidemic diarrhea (PED) is a highly contagious, acute enteric tract infectious disease of pigs (Sus domesticus) caused by porcine epidemic diarrhea virus (PEDV). PED is characterized by watery diarrhea, dehydration, weight loss, vomiting and death. PEDV damages pig intestinal epithelial tissue, causing intestinal hyperemia and atrophy of intestinal villi, with formation of intestinal epithelial cell cytoplasmic vacuoles. Since pig small intestinal epithelial cells (IECs) are target cells of PEDV infection, IEC cells were utilized as a model for studying changes in cellular activities post-PEDV infection. Monitoring of Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities demonstrated that PEDV infection decreased these activities. In addition, IECs proliferation was shown to decrease after PEDV infection using an MTT assay. Moreover, IECs apoptosis detected by flow cytometry with propidium iodide (PI) staining was clearly shown to increase relative to the control group. Meanwhile, animal experiments indicated that PEDV virulence for IEC cells was greater than viral virulence for Vero cells, although this may be due to viral attenuation after numerous passages in the latter cell line. Collectively, these studies revealed viral pathogenic mechanisms in PEDV-infected IECs and offer a theoretical basis for PEDV prevention and control.

Cryptosporidium suis infection in post-weaned and adult pigs in Shaanxi province, northwestern China.

Cryptosporidium spp., ubiquitous enteric parasitic protozoa of vertebrates, recently emerged as an important cause of economic loss and zoonosis. The present study aimed to determine the distribution and species of Cryptosporidium in post-weaned and adult pigs in Shaanxi province, northwestern China. A total of 1,337 fresh fecal samples of post-weaned and adult pigs were collected by sterile disposable gloves from 8 areas of Shaanxi province. The samples were examined by Sheather's sugar flotation technique and microscopy at × 400 magnification for Cryptosporidium infection, and the species in positive samples was further identified by PCR amplification of the small subunit (SSU) rRNA gene. A total of 44 fecal samples were successfully amplified by the nested PCR of the partial SSU rRNA, with overall prevalence of 3.3%. The average prevalence of Cryptosporidium infection in each pig farms ranged from 0 to 14.4%. Species identification by sequencing of SSU rRNA gene revealed that 42 (3.1%) samples were Cryptosporidium suis and 2 (0.15%) were Cryptosporidium scrofarum. C. suis had the highest prevalence (7.5%) in growers and the lowest in breeding pigs (0.97%). C. suis was the predominant species in pre-weaned and adult pigs, while C. scrofarum infected pigs older than 3 months only. A season-related difference of C. suis was observed in this study, with the highest prevalence in autumn (5.5%) and the lowest (1.7%) in winter. The present study provided basic information for control of Cryptosporidium infection in pigs and assessment of zoonotic transmission of pigs in Shaanxi province, China.

Dicrocoelium chinensis and Dicrocoelium dendriticum (Trematoda: Digenea) are distinct lancet fluke species based on mitochondrial and nuclear ribosomal DNA sequences.

Lancet flukes parasitize the bile ducts and gall bladder of a range of mammals, including humans, causing dicrocoeliosis. In the present study, we sequenced and characterized the complete mitochondrial (mt) genomes as well as the first and second internal transcribed spacers (ITS-1 and ITS-2=ITS) of nuclear ribosomal DNA (rDNA) of two lancet flukes, Dicrocoelium chinensis and D. dendriticum. Sequence comparison of a conserved mt gene and nuclear rDNA sequences among multiple individual lancet flukes revealed substantial nucleotide differences between the species but limited sequence variation within each of them. Phylogenetic analysis of the concatenated amino acid and multiple mt rrnS sequences using Bayesian inference supported the separation of D. chinensis and D. dendriticum into two distinct species-specific clades. Results of the present study support the proposal that D. dendriticum and D. chinensis represent two distinct lancet flukes. While providing the first mt genomes from members of the superfamily Plagiorchioidea, the novel mt markers described herein will be useful for further studies of the diagnosis, epidemiology and systematics of the lancet flukes and other trematodes of human and animal health significance.

The protective role of cardiovascular health against kidney stones: A study based on LE8 score.

Objectives: The American Heart Association recently released an updated algorithm for evaluating cardiovascular health (CVH)-Life's Essential 8 (LE8) score. Our objective was to investigate the correlation between levels of CVH, as determined by the LE8 score, and the risk of kidney stones among a representative sample of adults in the United States. Methods: We included data from the National Health and Nutrition Examination Survey (NHANES) covering the years 2007-2016 for further analysis. The LE8 score, a comprehensive measurement ranging from 0 to 100, was used to evaluate overall CVH and classified into three categories: low (0-49), moderate (50-79), and high (80-100) CVH. Logistic regression was employed to assess the association between the LE8 score and kidney stones. Furthermore, sensitivity analysis was conducted to validate the findings, and the presence of a non-linear relationship was examined using restricted cubic spline (RCS) regression methods. Results: A total of 19,988 participants were included in this study (weighted mean age, 47.99 years; 95 % confidence interval [CI]: 47.46-48.53 years), with 10,319 being female (weighted percentage, 51.98 %; 95 % CI: 51.26-52.71 %) and 1923 identified as having kidney stones (weighted percentage, 9.95 %; 95 % CI: 9.41-10.53 %). In the fully-adjusted multivariable model, higher LE8 scores were associated with prevalence of self-reported kidney stones (odds ratio [OR] for a 10-unit increase in score, 0.86; 95 % CI: 0.82-0.91), presenting a linear dose-response relationship. Compared to the low CVH group, participants in the moderate and high CVH groups exhibited a lower prevalence of kidney stones (OR = 0.80; 95 % CI: 0.69-0.92; OR = 0.54; 95 % CI: 0.43-0.69, respectively). Similar trends were observed when assessing the association between health behavior scores and kidney stones. Moreover, the negative correlation between the LE8 score and the prevalence of kidney stones was markedly more pronounced in various stratified analyses. Conclusion: Our study suggests that a higher level of CVH, as assessed by the LE8 metrics, is independently associated with a lower prevalence of self-reported kidney stones in a linear relationship. Further research, particularly through longitudinal or intervention studies, is required to establish whether actively promoting optimal CVH levels can effectively reduce the incidence of kidney stones.

Loganin reduces diabetic kidney injury by inhibiting the activation of NLRP3 inflammasome-mediated pyroptosis.

Diabetic kidney disease (DKD) is an essential cause of end-stage renal disease. The ongoing inflammatory response in the proximal tubule promotes the progression of DKD. Timely and effective blockade of the inflammatory process to protect the kidney during DKD progression is a proven strategy. The purpose of this study was to investigate the protective effect of loganin on diabetic nephropathy in vivo and in vitro and whether this effect was related to the inhibition of pyroptosis. The results indicated that loganin reduced fasting blood glucose, blood urea nitrogen and serum creatinine concentrations, and alleviated renal pathological changes in DKD mice. In parallel, loganin downregulated the expression of pyroptosis related proteins in the renal tubules of DKD mice and decreased serum levels of interleukin-1beta (IL-1ß) and interleukin-18 (IL-18). Furthermore, in vitro experiments showed that loganin attenuated high glucose-induced HK-2 cell injury by reducing the expression of pyroptosis-related proteins, and cytokine levels were also decreased. These fundings were also confirmed in the polyphyllin VI (PPVI) -induced HK-2 cell pyroptosis model. Loganin reduces high glucose induced HK-2 cells pyroptosis by inhibiting reactive oxygen species (ROS) production and NOD-like receptor protein 3 (NLRP3) inflammasome activation. In conclusion, the inhibition of pyroptosis via inhibition of the NLRP3/Caspase-1/Gasdermin D (GSDMD) pathway might be an essential mechanism for loganin treatment of DKD.

Hypoxia switches TET1 from being tumor-suppressive to oncogenic.

The classical oxidizing enzymatic activity of Ten Eleven Translocation 1 (TET1) and its tumor suppressor role are well known. Here, we find that high TET1 expression is associated with poor patient survival in solid cancers often having hypoxia, which is inconsistent with its tumor suppressor role. Through a series of in vitro and in vivo studies, using thyroid cancer as a model, we demonstrate that TET1 plays a tumor suppressor function in normoxia and, surprisingly, an oncogenic function in hypoxia. Mechanistically, TET1 mediates HIF1α-p300 interaction by acting as a co-activator of HIF1α to promote CK2B transcription under hypoxia, which is independent of its enzymatic activity; CK2 activates the AKT/GSK3ß signaling pathway to promote oncogenesis. Activated AKT/GSK3ß signaling in turn maintains HIF1α at elevated levels by preventing its K48-linked ubiquitination and degradation, creating a feedback loop to enhance the oncogenicity of TET1 in hypoxia. Thus, this study uncovers a novel oncogenic mechanism in which TET1 promotes oncogenesis and cancer progression through a non-enzymatic interaction between TET1 and HIF1α in hypoxia, providing novel therapeutic targeting implications for cancer.

Ayanin, a natural flavonoid inhibitor of Caseinolytic protease, is a promising therapeutic agent to combat methicillin-resistant Staphylococcus aureus infections.

Antimicrobial resistance (AMR) is a global health threat. The dramatic increase of Methicillin-resistant Staphylococcus aureus (MRSA) infections emphasizes the need to find new anti-infective agents with a novel mode of action. The Caseinolytic protease (ClpP) is a central virulence factor in stress survival, virulence, and antibiotic resistance of MRSA. Here, we found ayanin, a flavonoid isolated from Callicarpa nudiflora, was an inhibitor of MRSA ClpP with an IC50 of 19.63 µM. Using quantitative real-time PCR, ayanin reduced the virulence of Staphylococcus aureus (S. aureus) by down-regulating the level of some important virulence factors, including agrA, RNAⅢ, hla, pvl, psmα and spa. The results of cellular thermal shift assay and thermal shift assay revealed a binding between ayanin and ClpP. Molecular docking showed that ASP-168, ASN-173 and ARG-171 were the potential binding sites for ClpP binding to ayanin. ClpP mutagenesis study further indicated that ARG-171 and ASN-173 were the main active sites of ClpP. The affinity constant (KD) value of ayanin with ClpP was 3.15 × 10-5 M measured by surface plasmon resonance. In addition, ayanin exhibited a significant therapeutic effect on pneumonia infection induced by S. aureus in mice in vivo, especially in combination with vancomycin. This is the first report of ayanin with in vivo and in vitro efficacy against S. aureus infection. In conclusion, ayanin is a promising therapeutic agent to combat MRSA infections by targeting ClpP.

Isovanillic acid protects mice against Staphylococcus aureus by targeting vWbp and Coa.

Aim: Our primary objective was to investigate the protective effects and mechanisms of isovanillic acid in mice infected with Staphylococcus aureus Newman. Methods: In vitro coagulation assays were used to validate vWbp and Coa as inhibitory targets of isovanillic acid. The binding mechanism of isovanillic acid to vWbp and Coa was investigated using molecular docking and point mutagenesis. Importantly, a lethal pneumonia mouse model was used to assess the effect of isovanillic acid on survival and pathological injury in mice. Results & Conclusion: Isovanillic acid reduced the virulence of S. aureus by directly binding to inhibit the clotting activity of vWbp and Coa, thereby reducing lung histopathological damage and improving the survival rate in mice with pneumonia.

Transjugular Approach to Closure of Patent Foramen Ovale Under the Guidance of Fluoroscopy and Transthoracic Echocardiography: A Case Report.

Background: We describe a rare case of patent foramen ovale (PFO) associated stroke in a patient with pulmonary embolism, inferior vena cava thrombosis and undergoing filter implantation who successfully underwent PFO closure using the right internal jugular venous approach. Case Summary: This is a rare case of a 42-year-old patient who presented with stroke and pulmonary embolism and was diagnosed with a PFO, inferior vena cava thrombosis and underwent filter implantation. The patient suffered from stroke and pulmonary embolism successively; that is, embolic events occurred in both the arterial and venous systems. Transesophageal echocardiography (TEE) showed a PFO with an atrial septal aneurysm (ASA), which we considered a "pathological" PFO. Due to the obstructive nature of the inferior vena cava approach, we successfully performed PFO closure via the right internal jugular venous approach under the guidance of X-ray and transthoracic echocardiography (TTE). Discussion: The right jugular venous approach provides a simple technical solution for patients who require PFO closure when femoral venous access is unavailable, which can be performed under X-ray and TTE guidance.

A Prospective, Single-Center, Phase I Clinical Trial to Evaluate the Value of Transesophageal Echocardiography in the Closure of Patent Foramen Ovale With a Novel Biodegradable Occluder.

Objective: Traditional metal alloy occluders for the closure of patent foramen ovale (PFO) may be associated with some potential complications, and may restrict the trans-septal access to the left atrium for future treatment of left-sided heart disease. Increasing attention has been paid to novel biodegradable occluders (NBOs) to achieve PFO closure. We aimed to evaluate the role of transesophageal echocardiography (TEE) in the diagnostic and anatomical evaluation of PFO, as well as in the Post-procedural assessment after transcatheter closure with a NBO. Methods: We conducted a prospective, single-center clinical study of 44 patients who were diagnosed with PFO by contrast transthoracic echocardiography (c-TTE) and TEE from June 2019 to June 2020. All patients underwent PFO occlusion with NBO under TTE guidance. Follow-up was performed at 2 days and 3 months after the procedure with TTE, and at 6 months and 1 year after the procedure with c-TTE, TTE, and TEE. Results: Interventional treatment was successfully performed in all patients. The left and right sides of the occluder device disc were significantly reduced at 3, 6, and 12 months compared to 2 days after the procedure (all P < 0.01), and decreased gradually. The thickness was significantly reduced at 12 months compared to the first three time points (all P < 0.01). Thrombus was found on the surface of the occluder device in three patients (6.4%) at 3 and 6 months after occlusion. At 6 months after procedure, there were 3 (6.8%) cases of extensive residual right-to-left shunt (RLS), 2 (4.5%) cases of moderate shunt, and 7 (15.9%) cases of small shunts. One year after procedure, 2 (4.5%) cases had a extensive residual shunt, 6 (13.6%) cases of small shunts were confirmed to originate from pulmonary veins by TEE, and the PFO-RLS occlusion rate reached 95.5%. Conclusion: This study demonstrates the feasibility, safety, and effectiveness of NBO for the closure of PFO in humans, with a high rate of complete shunt closure. Accurate TEE assessment of the PFO anatomy before closure with NBO is important to ensure that the procedure remains safe and effective. Furthermore, TEE plays a crucial role in the Post-procedure follow-up.