Shigella strains are human pathogens. The O antigen gene cluster of Shigella dysenteriae O7 was sequenced and analyzed. It contains genes for synthesis of nucleotide sugars including UDP-2-acetamido-2-deoxy-D-galacturonamide, UDP-2-acetamido-2-deoxy-D-galacturonic acid and dTDP-4-amino-4,6-dideoxy-D-glucose. Also found in the gene cluster are genes encoding O unit flippase, O antigen polymerase and sugar transferases. The Escherichia coli O121 O antigen, which is present in an important Shiga toxin-producing strain, has the same structure as that of S. dysenteriae O7, and we found that the gene clusters also had the same genes and organization. Four genes specific to S. dysenteriae O7 and E. coli O121 were identified by PCR screening against representatives of 186 E. coli (including Shigella) O serotypes. E. coli O121 and S. dysenteriae O7 isolates can be distinguished by PCR of the H antigen fliC gene.

Programmed death ligand 1 (PD-L1) which is upregulated in various epithelial tumors, plays a central role in the evasion of the immune system. In addition to monoclonal antibodies that blocking PD1/PD-L1 axis, finding small molecule compounds that can suppress PD-L1 expression might be another substitutable strategy for PD1/PD-L1 based therapy. Here, we found that dihydropyridine calcium channel blockers dose-dependently reduced the expression of PD-L1, both in the cytoplasm and cell surface. IFNγ induced PD-L1 transcription was consistently suppressed by Lercanidipine in 24 h, whereas, the half-life of PD-L1 protein was not significantly affected. IFNγ trigged significant STAT1 phosphorylation, which was eliminated by Lercanidipine. Similarly, STAT1 phosphorylation could also be abolished by extracellular calcium chelating agent EGTA and intracellular calcium chelator BAPTA-AM. Furthermore, Lercanidipine enhanced killing ability of T cells by down-regulating PD-L1. Taken together, our studies suggest that calcium signal is a crucial factor that mediates the transcription of PD-L1 and regulation of calcium can be used as a potential strategy for PD-L1 inhibition.

Recent studies have pointed to the role of Krüpple-like factor 12 (KLF12) in cancer-associated processes, including cancer proliferation, apoptosis, and metastasis. However, the role of KLF12 in tumor immunity remains obscure. Here, we found that KLF12 expression was significantly higher in non-small cell lung cancer (NSCLC) cells with higher programmed death-ligand 1 (PD-L1) expression. Additionally, a positive correlation between KLF12 and PD-L1 was observed in clinical patient tumor tissues. By chromatin immunoprecipitation (ChIP) analysis, KLF12 was identified to bind to the CACCC motif of the PD-L1 promoter. Overexpression of KLF12 promoted PD-L1 transcription, whereas silencing of KLF12 inhibited PD-L1 transcription. Furthermore, signal transducer and activator of transcription 1 (STAT1)- and STAT3-triggered PD-L1 transcription was abolished in the absence of KLF12, and KLF12 knockdown weakened the binding of STAT1 and STAT3 to the PD-L1 promoter. Mechanistically, KLF12 physically interacted with P300, a histone acetyltransferase. In addition, KLF12 silencing reduced P300 binding to the PD-L1 promoter, which subsequently caused decreased acetylation of histone H3. PD-L1 transcription driven by KLF12 overexpression was eliminated by EP300 silencing. In immunocompetent mice, KLF12 knockout inhibited tumor growth and promoted infiltration of CD8+ T cells. However, this phenomenon was not observed in immunodeficient mice. Overall, this study reveals KLF12-mediated transcriptional regulation of PD-L1 in NSCLC; targeting KLF12 may be a potential therapeutic strategy for NSCLC.

Background. Portal cavernoma (PC) is the most critical condition with risk or variceal hemorrhage in pediatric patients. We retrospectively investigated the patients with PC focusing on the predictors for recurrent variceal bleeding. Methods. Between July 2003 and June 2013, we retrospectively enrolled all consecutive patients admitted to our department with a diagnosis of PC without abdominal malignancy or liver cirrhosis. The primary endpoint of this observational study was recurrent variceal bleeding. Independent predictors of recurrent variceal bleeding were identified using the logistic regression model. Results. A total of 157 patients were enrolled in the study. During the follow-up period, 24 patients exhibited onset of recurrent variceal bleeding. Acute variceal bleeding was subjected to conservative symptomatic treatment and emergency endoscopic sclerotherapy. Surgical procedure selection was based on the severity of vascular dilation and collateral circulation. Multivariate logistic regression analysis demonstrated that the presence of ascites, collateral circulation, and portal venous pressure were independent prognostic factors of recurrent variceal bleeding for patients with portal cavernoma. Conclusions. The presence of ascites, collateral circulation, and portal venous pressure evaluation are important and could predict the postsurgical recurrent variceal bleeding in patients with portal cavernoma.

To survive in its sand fly vector, the trypanosomatid protozoan parasite Leishmania first attaches to the midgut to avoid excretion, but eventually it must detach for transmission by the next bite. In Leishmania major strain Friedlin, this is controlled by modifications of the stage-specific adhesin lipophosphoglycan (LPG). During differentiation to infective metacyclics, d-arabinopyranose (d-Arap) caps the LPG side-chain galactose residues, blocking interaction with the midgut lectin PpGalec, thereby leading to parasite detachment and transmission. Previously, we characterized two closely related L. major genes (FKP40 and AFKP80) encoding bifunctional proteins with kinase/pyrophosphorylase activities required for salvage and conversion of l-fucose and/or d-Arap into the nucleotide-sugar substrates required by glycosyltransferases. Whereas only AFKP80 yielded GDP-d-Arap from exogenous d-Arap, both proteins were able to salvage l-fucose to GDP-fucose. We now show that Δafkp80- null mutants ablated d-Arap modifications of LPG as predicted, whereas Δfkp40- null mutants resembled wild type (WT). Fucoconjugates had not been reported previously in L. major, but unexpectedly, we were unable to generate fkp40-/afkp80- double mutants, unless one of the A/FKPs was expressed ectopically. To test whether GDP-fucose itself was essential for Leishmania viability, we employed "genetic metabolite complementation." First, the trypanosome de novo pathway enzymes GDP-mannose dehydratase (GMD) and GDP-fucose synthetase (GMER) were expressed ectopically; from these cells, the Δfkp40-/Δafkp80- double mutant was now readily obtained. As expected, the Δfkp40-/Δafkp80-/+TbGMD-GMER line lacked the capacity to generate GDP-Arap, while synthesizing abundant GDP-fucose. These results establish a requirement for GDP-fucose for L. major viability and predict the existence of an essential fucoconjugate(s).

Given the limitations of the existing antibody-based therapies, including immune-related adverse events, poor response rates, and intravenous route of dosing, small molecules inhibitors targeting PD-L1 are highly desirable. By cell-based screening, we found that tyrosine kinase inhibitor Bafetinib dramatically suppresses PD-L1 protein expression in a dose-dependent manner. In parallel, cell membrane PD-L1 is also reduced by Bafetinib. We confirm that Bafetinib doesn't affect the protein half-life of PD-L1 but significantly inhibits the transcription of PD-L1. Among the transcription factors that regulate PD-L1 expression, c-Myc is downregulated by Bafetinib. Bafetinib caused PD-L1 inhibition is abolished when c-Myc is knocked-down. Further, we identified that Bafetinib reduced c-Myc expression because of transcription inhibition. By using the CT26 tumor model, we further confirm that Bafetinib suppressed PD-L1 expression in vivo. In conclusion, our study shows that Bafetinib inhibits the transcription of PD-L1 through transcription factor c-Myc, suggesting that Bafetinib might be a small molecule drug targeting PD-L1.

Fecal microbiota transplantation (FMT) has been used successfully to treat a variety of gastroenterological diseases. The alterations of microbiota in mouse models of necrotizing enterocolitis (NEC) as well as in patients suggested the possibility of treating NEC with FMT. Here we show that FMT caused an improvement in the histopathology and symptoms of NEC in WT mice, but not Grx1-/- mice. FMT eliminated O2•- production and promoted NO production in experimental NEC mice though the modulation of S-glutathionylation of eNOS (eNOS-SSG). FMT decreased the extent of TLR4-mediated proinflammatory signaling though TLR9 in the intestinal mucosa tissue. FMT also suppressed intestinal apoptosis and bacterial translocation across the intestinal barrier, which was accompanied by decreased inflammatory cytokine levels, altered bacterial microbiota, and regulated lymphocyte proportions. FMT is effective in a mouse model of NEC through the modulation of oxidative stress and reduced colon inflammation.

The aim of the present study was to clarify the correlations between SPARC expression in gastric cancer-associated fibroblasts (GCAFs) and the prognosis of patients with gastric cancer and to elucidate the role of GCAF-derived SPARC in stemness transformation and 5-fluorouracil resistance in gastric cancer.

Dexmedetomidine (Dex) has been proven to exert protective effects on multiple organs in response to ischaemia-reperfusion injury, but the specific mechanism by which this occurs has not been fully elucidated. The purpose of this study was to investigate whether Dex attenuates spinal cord ischaemia-reperfusion injury (SCIRI) by inhibiting endoplasmic reticulum stress (ERS). Our team established a model of SCIRI and utilized the endoplasmic reticulum agonist thapsigargin. Dex (25 g/kg) was intraperitoneally injected 30 minutes before spinal cord ischaemia. After 45 minutes of ischaemia, the spinal cord was reperfused for 24 hours. To evaluate the neuroprotective effect of Dex on SCIRI, neurological function scores were assessed in rats and apoptosis of spinal cord cells was determined by TUNEL staining. To determine whether the endoplasmic reticulum apoptosis pathway CNPY2-PERK was involved in the neuroprotective mechanism of Dex, the expression levels of related proteins (CNPY2, GRP78, PERK, CHOP, caspase-12, caspase-9 and caspase-3) were detected by western blot analysis and RT-PCR. We observed that Dex significantly increased the neurological function scores after SCIRI and decreased apoptosis of spinal cord cells. The expression of ERS-related apoptosis proteins was significantly increased by SCIRI but was significantly decreased in response to Dex administration. Taken together, the results of this study indicate that Dex may attenuate SCIRI by inhibiting the CNPY2-ERS apoptotic pathway.

Gastric cancer (GC), known for high morbidity and mortality, is poorly prognosed with traditional chemotherapy and biological agents. Current studies have found that over-activation of AKT is a common molecular characteristic in GC. Although the development of this targeted inhibitor has entered clinical phases, limited success is reported because of its compensatory signaling pathways. Here, we found that GC cell lines with high phosphorylation of AKT show different sensitivity to AKT inhibitors (AKTis), but a reduction of p-GSK3β related sensitivity of AKTis in GC cells. Besides, we revealed that Ceritinib exerted a strongly synergistic antitumor effect with AKT inhibitors both in vitro and in vivo. Obviously, Ceritinib improved the sensitivity of Capivasertib (AZD5363, AKTs) and Afuresertib (GSK2110183, AKTis) in gastric cancer cells, as illustrated by a significant reduction in the GC cell proliferation and enhanced apoptosis. The drug combination showed tumor regression in BALB/c (nu/nu) mouse MKN45 (Gastric cancer), tumor model. Also, the combination strategy indicated significantly low p-AKT levels due to AKTis compensation and reduced the levels of p-GSK3β in both GC cell lines and GC patient-derived cells. These findings may provide a novel combination strategy for gastric cancer treatment.

Prostaglandin E2 (PGE2) is implicated in intestinal inflammation and intestinal blood flow regulation with a paradoxical effect on the pathogenesis of necrotizing enterocolitis (NEC), which is not yet well understood. In the current study, we found that PGE2, EP4, and COX-2 varied at different distances from the most damaged area in the terminal ileum obtained from human infants with NEC. PGE2 administration alleviated the phenotype of experimental NEC and the intestinal microvascular features in experimental NEC, but this phenomenon was inhibited by eNOS depletion, suggesting that PGE2 promoted intestinal microcirculatory perfusion through eNOS. Furthermore, PGE2 administration increased the VEGF content in MIMECs under TNFα stress and promoted MIMEC proliferation. This response to PGE2 was involved in eNOS phosphorylation and nitric oxide (NO) production and was blocked by the EP4 antagonist in vitro, suggesting that targeting the PGE2-EP4-eNOS axis might be a potential clinical and therapeutic strategy for NEC treatment. The study is reported in accordance with ARRIVE guidelines ( https://arriveguidelines.org ).

Understanding the regulatory mechanisms of PD-L1 expression in tumors provides key clues for improving immune checkpoint blockade efficacy or developing novel oncoimmunotherapy. Here, we showed that the FDA-approved sodium-glucose cotransporter-2 (SGLT2) inhibitor canagliflozin dramatically suppressed PD-L1 expression and enhanced T cell-mediated cytotoxicity. Mechanistic study revealed that SGLT2 colocalized with PD-L1 at the plasma membrane and recycling endosomes and thereby prevented PD-L1 from proteasome-mediated degradation. Canagliflozin disturbed the physical interaction between SGLT2 and PD-L1 and subsequently allowed the recognition of PD-L1 by Cullin3SPOP E3 ligase, which triggered the ubiquitination and proteasome-mediated degradation of PD-L1. In mouse models and humanized immune-transformation models, either canagliflozin treatment or SGLT2 silencing significantly reduced PD-L1 expression and limited tumor progression - to a level equal to the PD-1 mAb - which was correlated with an increase in the activity of antitumor cytotoxic T cells. Notably, prolonged progression-free survival and overall survival curves were observed in the group of PD-1 mAb-treated patients with non-small cell lung cancer with high expression of SGLT2. Therefore, our study identifies a regulator of cell surface PD-L1, provides a ready-to-use small-molecule drug for PD-L1 degradation, and highlights a potential therapeutic target to overcome immune evasion by tumor cells.

Fucose is a common component of eukaryotic cell-surface glycoconjugates, generally added by Golgi-resident fucosyltransferases. Whereas fucosylated glycoconjugates are rare in kinetoplastids, the biosynthesis of the nucleotide sugar GDP-Fuc has been shown to be essential in Trypanosoma brucei. Here we show that the single identifiable T. brucei fucosyltransferase (TbFUT1) is a GDP-Fuc: β-D-galactose α-1,2-fucosyltransferase with an apparent preference for a Galβ1,3GlcNAcβ1-O-R acceptor motif. Conditional null mutants of TbFUT1 demonstrated that it is essential for both the mammalian-infective bloodstream form and the insect vector-dwelling procyclic form. Unexpectedly, TbFUT1 was localized in the mitochondrion of T. brucei and found to be required for mitochondrial function in bloodstream form trypanosomes. Finally, the TbFUT1 gene was able to complement a Leishmania major mutant lacking the homologous fucosyltransferase gene (Guo et al., 2021). Together these results suggest that kinetoplastids possess an unusual, conserved and essential mitochondrial fucosyltransferase activity that may have therapeutic potential across trypanosomatids.

Trypanosoma cruzi is the protozoan parasite causing Chagas disease, a Neglected Tropical Disease that affects 8 million people and causes 12,000 deaths per year, primarily because of cardiac pathology. Effective vaccination for T. cruzi remains an elusive goal. The use of a live vaccine vector, especially one that mimics the pathogen target, may be superior to the use of recombinant protein or DNA vaccine formulations.