Increased Th17 activation and gut microbiota diversity are associated with pembrolizumab-triggered tuberculosis.

INTRODUCTION: A hypersensitivity response akin to immune reconstitution inflammatory syndrome (IRIS) has been proposed as a mechanism responsible for anti-PD-1 therapy-induced tuberculosis. IRIS is associated with enhanced activation of IL-17A-expressing CD4 + T cells (Th17). Gut microbiota is thought to be linked to pulmonary inflammation through the gut-lung axis. MATERIALS AND METHODS: We used ImmuCellAI to investigate the T cell population in lung cancer and tuberculosis samples. Then, we applied flow cytometry to monitor the expression levels of the Th17 cell activation marker CD38 in the peripheral blood of a patient experiencing adverse events, including tuberculosis, in response to pembrolizumab. The gut microbiome was examined by 16S rRNA sequencing to examine the alterations caused by pembrolizumab. RESULTS: The percentage of Th17 cells was increased in both lung cancer and tuberculosis. FACS analysis showed that pembrolizumab induced substantial CD38 expression in Th17 cells. The patient's fecal samples showed that the diversity of the gut microbiota was significantly increased in response to the pembrolizumab cycle. One enriched genus was Prevotella, which has previously been linked to lung inflammation and Th17 immune activation. DISCUSSION: The observed Th17 activation in our patient was consistent with a role of Th17-mediated IRIS in pembrolizumab-triggered tuberculosis. Pembrolizumab might trigger airway inflammation with a Th17 phenotype through microbiota interactions in the gut-lung axis.

Extraction of nucleobases, nucleosides and nucleotides by employing a magnetized graphene oxide functionalized with hydrophilic phytic acid and titanium(IV) ions.

A magnetite@graphene oxide nanocomposite was first coated with polyethylenimine and then modified with phytic acid and titanium(IV) ions. The high loading with Ti(IV) and the good hydrophilicity of PEI and PA result in a material that can be applied to the efficient extraction of highly polar nucleobases, nucleosides and nucleotides. The physicochemical properties of the composite were investigated by scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, water contact angle measurements, thermogravimetric analysis, and vibrating sample magnetometry. A series of parameters that affect extraction and elution under the conditions of immobilized metal affinity chromatography (IMAC) and hydrophilic interaction liquid chromatography (HILIC) were examined. The analytes were eluted from the nanocomposites using 10 mM trisodium phosphate as the elution solution in the IMAC mode, and 50% methanol-water as elution solution in the HILIC mode. Figures of merit include (a) an intra-day precision of 0.1-1.0% in the IMAC mode; (b) an intra-day precision of 0.4%-0.8% in the HILIC mode; (c) detection limits between 1.8-2.8 ng mL-1 in the IMAC mode; and (d) detection limits of 4.0-10.5 ng mL-1 in the HILIC mode. The method was applied to the extraction of the nucleotides cytidine-5'-monophosphate (CMP), uridine-5'-monophosphate (UMP), guanosine-5'-monophosphate (GMP), and adenosine-5'-monophosphate (AMP), and the nucleobases and nucleosides hypoxanthine, adenosine, cytosine, inosine and cytidine from Cordyceps sinensis, Lentinus edodes and plasma samples. Graphical abstract Schematic presentation of the workflow for the extraction of nucleobases, nucleosides and nucleotides using phytic acid-Ti(IV) functionalized magnetite@graphene oxide nanocomposites under two distinct modes.

Evaluation of interactions between RAW264.7 macrophages and small molecules by capillary electrophoresis.

In this study, the affinity interactions between RAW 264.7 macrophages and three small molecules including naringin, oleuropein and paeoniflorin were evaluated by affinity capillary electrophoresis (ACE), partial filling affinity capillary electrophoresis (PFACE) and frontal analysis capillary electrophoresis (FACE), respectively. The result indicated that ACE (varying concentrations of cell suspension were filled in the capillary as receptor) may not be suitable for the evaluation of interactions between cell and small molecules due to the high viscosity of cell suspension; PFACE can qualitatively evaluate the interaction, but the difference in viscosity between RAW264.7 suspension and buffer effects on the liner relationship between filling length and injection time, which makes the calculation of binding constant difficult. Furthermore, based on the PFACE results, naringin showed stronger interaction with macrophages than the other two molecules; taking advantage of the aggregation phenomenon of cell induced by electric field, FACE was successfully used to determine the stoichiometry (n = 5×109 ) and binding constant (Kb = 1×104 L/mol) of the interaction between RAW264.7 and naringin.

Evaluation of affinity interaction between small molecules and platelets by open tubular affinity capillary electrochromatography.

In this paper, an open tubular affinity capillary electrochromatography (OT-ACEC) was developed by physical adsorption of rabbit platelets on the inner surface of capillary. The interactions between small molecules include adenosine diphosphate (ADP) (positive control), protocatechuic acid (negative control) and seven natural products (salvianolic acid B, salvianic acid A sodium, hydroxysafflor yellow A, ferulic acid, chlorogenic acid, sinapic acid, caffeic acid) and platelets were evaluated by their retention factors and binding constants obtained based on peak-shift assay. Then, the activities of anti-platelet aggregation induced by thrombin (THR), ADP and arachidonic acid (AA) for those small molecules (except ADP) were evaluated by turbidimetric method. The results indicate that: (i) ADP, a platelet aggregation inducer, had strong interaction with platelet, while protocatechuic acid that had no inhibition on platelet aggregation behaved no specific interaction; (ii) there was a positive correlation between the anti-platelet aggregation activities of small molecules and their interactions with platelet, generally those compounds with higher binding constants with platelet exhibited higher activities. Therefore, the OT-ACEC method developed in the present study can be a potential method to evaluate affinity interactions between small molecules and platelets, so as to predict the biological activities such as anti-platelet aggregation for the small molecules.