Construction of [89Zr]Zr-Labeled HuL13 for ImmunoPET Imaging of LAG-3 Checkpoint Expression on Tumor-Infiltrating T Cells.

Lymphocyte activation gene 3 (LAG-3) has attracted much attention as a potentially valuable immune checkpoint. Individual identification of LAG-3 expression at screening and during treatment could improve the successful implementation of anti-LAG-3 therapies. HuL13 is a human IgG1 monoclonal antibody that binds to the LAG-3 receptor in T cells. Here, we used [89Zr]Zr-labeled HuL13 to delineate LAG-3+ T-cell infiltration into tumors via positron emission tomography (PET) imaging. A549/LAG-3 cells, which stably express LAG-3, were generated by infection with lentivirus. The uptake of [89Zr]Zr-DFO-HuL13 in A549/LAG-3 cells was greater than that in the negative control (A549/NC) cells at each time point. The equilibrium dissociation constant (Kd) of [89Zr]Zr-DFO-HuL13 for the LAG-3 receptor was 8.22 nM. PET imaging revealed significant uptake in the tumor areas of A549/LAG-3 tumor-bearing mice from 24 h after injection (SUVmax = 2.43 ± 0.06 at 24 h). As a proof of concept, PET imaging of the [89Zr]Zr-DFO-HuL13 tracer was further investigated in an MC38 tumor-bearing humanized LAG-3 mouse model. PET imaging revealed that the [89Zr]Zr-DFO-HuL13 tracer specifically targets human LAG-3 expressed on tumor-infiltrating lymphocytes (TILs). In addition to the tumors, the spleen was also noticeably visible. Tumor uptake of the [89Zr]Zr-DFO-HuL13 tracer was lower than its uptake in the spleen, but high uptake in the spleen could be reduced by coinjection of unlabeled antibodies. Coinjection of unlabeled antibodies increases tracer activity in the blood pool, thereby improving tumor uptake. Dosimetry evaluation of the healthy mouse models revealed that the highest absorbed radiation dose was in the spleen, followed by the liver and heart wall. In summary, these studies demonstrate the feasibility of using the [89Zr]Zr-DFO-HuL13 tracer for the detection of LAG-3 expression on TILs. Further clinical evaluation of the [89Zr]Zr-DFO-HuL13 tracer may be of significant help in the stratification and management of patients suitable for anti-LAG-3 therapy.

Effect of endogenous sodium and potassium ions in plants on the quality of alfalfa silage and bacterial community stability during fermentation.

This study investigated the impact of endogenous sodium and potassium ions in plants on the quality of alfalfa silage, as well as the stability of bacterial communities during fermentation. Silage was produced from the fermented alfalfa, and the chemical composition, fermentation characteristics, and microbiome were analyzed to understand their interplay and impact on silage fermentation quality. The alfalfa was cultivated under salt stress with the following: (a) soil content of <1‰ (CK); (b) 1‰-2‰ (LP); (c) 2‰-3‰ (MP); (d) 3‰-4‰ (HP). The results revealed that the pH of silage was negatively correlated with the lactic acid content. With the increase of lactic acid (LA) content increased (26.3-51.0 g/kg DM), the pH value decreased (4.9-5.3). With the increase of salt stress, the content of Na+ in silage increased (2.2-5.4 g/kg DM). The presence of endogenous Na+ and K+ ions in plants significantly affected the quality of alfalfa silage and the dynamics of bacterial communities during fermentation. Increased salt stress led to changes in microbial composition, with Lactococcus and Pantoea showing a gradual increase in abundance, especially under high salt stress. Low pH inhibited the growth of certain bacterial genera, such as Pantoea and Pediococcus. The abundance of Escherichia-Shigella and Comamonas negatively correlated with crude protein (CP) content, while Enterococcus and Lactococcus exhibited a positive correlation. Furthermore, the accumulation of endogenous Na+ in alfalfa under salt stress suppressed bacterial proliferation, thereby reducing protein degradation during fermentation. The pH of the silage was high, and the LA content was also high. Silages from alfalfa under higher salt stress had higher Na+ content. The alpha diversity of bacterial communities in alfalfa silages showed distinct patterns. Desirable genera like Lactococcus and Lactobacillus predominated in silages produced from alfalfa under salt stress, resulting in better fermentation quality.

[Effects of Eucalyptus grandis leaf litter decomposition on the growth and photosynthetic characteristics of Cichorium intybus].

A pot experiment was conducted to study the effects of Eucalyptus grandis leaf litter during its early stage decomposition on the growth and the photosynthesis of Cichorium intybus. Each pot contained 12 kg soil mixed with different amounts of E. grandis leaf litter (30 g x pot(-1), A1; 60 g x pot(-1), A2; 90 g x pot(-1), A3; and 0 g x pot(-1), CK), and sowed with C. intybus. The growth indicators and the photosynthetic characteristics of C. intybus were measured after the third leaf of C. intybus seedlings fully expanded in treatment A3. At the early stage of leaf litter decomposition, the C. intybus biomass accumulation, leaf area growth, and synthesis of photosynthetic pigments were inhibited significantly, and the inhibition effect was getting stronger with the increasing amount of the leaf litter addition. The intercellular CO2 concentration of C. intybus was increased by litter addition, while the net photosynthetic rate, stomatal conductance, and transpiration rate were significantly lower than those of the control. With the increase of leaf litter addition, all the parameters of C. intybus light response and CO2 response except CO2 compensation point showed an obvious downward trend, and there existed significant differences between the treatments of litter additions and the control. It was suggested that during the decomposition of E. grandis leaf litter, its allelopathic substances released gradually and acted on receptor plants, inhibited the synthesis of photosynthetic pigments and the photosynthesis of the receptors, decreased the receptors environmental adaptation ability, and accordingly, inhibited the growth of C. intybus.