Comparative study on the epidemiological characteristics and hazards of respiratory syncytial virus and influenza virus infections among elderly people.

OBJECTIVE: To investigate the epidemiological characteristics and infections of respiratory syncytial virus (RSV) and influenza viruses in hospitalized elderly patients with respiratory tract infections in Suzhou City, China, and to compare the differences in clinical characteristics and economic burden associated with these two infections. METHODS: In this prospective study, pathogenetic testing and clinical data for hospitalized patients aged 60 years and older with respiratory tract infections were collected in five hospitals through stratified cluster sampling from December 2023 to May 2024. Comparative study on epidemic characteristics, clinical features and costs of cases who infected RSV alone and influenza alone were conducted. RESULTS: Among 1,894 cases included, the RSV positivity rate was 5.91% during the 2023-2024 winter-spring season, while the influenza positivity rate was 9.61%. RSV-B was the predominant subtype of RSV, and influenza A (primarily H3N2) was the dominant strain among the influenza-positive cases. Compared with cases infected influenza virus alone, those infected RSV alone had lower occurrence frequency of fever (18.8% vs. 35.7%, P = 0.004), higher occurrence frequency of complications of lower respiratory tract infections (70.8% vs. 54.8%, P = 0.011), higher direct medical costs ($996.2 vs. $841.1, P = 0.017) and total costs ($1019.7 vs. $888.1, P = 0.036). RSV single infection is more common in female cases (P = 0.007) and diabetic cases (P = 0.007) than influenza virus single infection. CONCLUSIONS: During the winter and spring months, RSV is the second most common pathogen after influenza virus among older adults hospitalized for respiratory infections in Suzhou, China. Patients infected RSV are more likely to develop complications with lower respiratory tract infections and have higher medical costs than the influenza. RSV infection in the elderly should be emphasized, especially in female patients and diabetic patients.

Spatial multi-omics reveal intratumoral humoral immunity niches associated with tertiary lymphoid structures in pancreatic cancer immunotherapy pathologic responders.

Pancreatic adenocarcinoma (PDAC) is a rapidly progressing cancer that responds poorly to immunotherapies. Intratumoral tertiary lymphoid structures (TLS) have been associated with rare long-term PDAC survivors, but the role of TLS in PDAC and their spatial relationships within the context of the broader tumor microenvironment remain unknown. We generated a spatial multi-omics atlas encompassing 26 PDAC tumors from patients treated with combination immunotherapies. Using machine learning-enabled H&E image classification models and unsupervised gene expression matrix factorization methods for spatial transcriptomics, we characterized cellular states within TLS niches spanning across distinct morphologies and immunotherapies. Unsupervised learning generated a TLS-specific spatial gene expression signature that significantly associates with improved survival in PDAC patients. These analyses demonstrate TLS-associated intratumoral B cell maturation in pathological responders, confirmed with spatial proteomics and BCR profiling. Our study also identifies spatial features of pathologic immune responses, revealing TLS maturation colocalizing with IgG/IgA distribution and extracellular matrix remodeling. HIGHLIGHTS: Integrated multi-modal spatial profiling of human PDAC tumors from neoadjuvant immunotherapy clinical trials reveal diverse spatial niches enriched in TLS.TLS maturity is influenced by tumor location and the cellular neighborhoods in which TLS immune cells are recruited.Unsupervised machine learning of genome-wide signatures on spatial transcriptomics data characterizes the TLS-enriched TME and associates TLS transcriptomes with survival outcomes in PDAC.Interactions of spatially variable gene expression patterns showed TLS maturation is coupled with immunoglobulin distribution and ECM remodeling in pathologic responders.Intratumoral plasma cell and immunoglobin gene expression spatial dynamics demonstrate trafficking of TLS-driven humoral immunity in the PDAC TME. Significance: We report a spatial multi-omics atlas of PDAC tumors from a series of immunotherapy neoadjuvant clinical trials. Intratumorally, pathologic responders exhibit mature TLS that propagate plasma cells into malignant niches. Our findings offer insights on the role of TLS-associated humoral immunity and stromal remodeling during immunotherapy treatment.

Multifunctional coating with hydrophobicity, antibacterial and flame-retardant properties on cotton fabrics by layer-by-layer self-assembly curing of phytic acid and a tyrosine-derived hyperbranched benzoxazine.

The inherent hydrophilicity and biocompatibility of cotton fabrics facilitated bacterial proliferation and safety concerns, limiting their applications. To address these issues, tyrosine-derived polyetherimide, bis(3-aminopropyl)-terminated poly(dimethylsiloxane), and paraformaldehyde were used to synthesize hyperbranched benzoxazine THB-BOZs-PDMS with potent antibacterial and antibiofilm activity. The protonated amino groups of benzoxazine facilitated electrostatic interactions with negatively charged bacteria, and hydrophobic interactions disrupted the cell membrane, leading to bacteria death. Notably, phytic acid interacts with benzoxazines through intermolecular forces, with its phosphoric acid groups facilitating the curing of benzoxazines, thereby imparting flame-retardant properties to the material. Consequently, a multifunctional coating was developed via LBL self-assembly and in-situ curing of benzoxazines and phytic acid on the fabric surfaces. The successful deposition of the coating was confirmed through compositional analysis and morphological characterization. After 4 cycles of LBL modification, the fabrics TBP + PA-CF-4 displayed outstanding antibacterial efficacy, bacterial anti-adhesion properties, and heat resistance. Furthermore, TBP + PA-CF-4 exhibited notable washing and mechanical durability, attributed to the stability conferred by in-situ cured of layers. Compared with other reported modified fabrics, TBP + PA-CF-4 displayed more comprehensive overall performances. These multifunctional fabrics provided a sustainable approach for advancing personal protective materials and public decoration, particularly suited for use in high-humidity environments or military settings.

Mapping the breast tumor microenvironment: proximity analysis reveals spatial relationships between macrophage subtypes and metastasis-initiating cancer cells.

Metastasis is responsible for the majority of cancer-related fatalities. We previously identified specific cancer cell populations responsible for metastatic events which are cytokeratin-14 (CK14) and E-cadherin positive in luminal tumors, and E-cadherin and vimentin positive in triple-negative tumors. Since cancer cells evolve within a complex ecosystem comprised of immune cells and stromal cells, we sought to decipher the spatial interactions of these aggressive cancer cell populations within the tumor microenvironment (TME). We used imaging mass cytometry to detect 36 proteins in tumor microarrays containing paired primary and metastatic lesions from luminal or triple-negative breast cancers (TNBC), resulting in a dataset of 1,477,337 annotated cells. Focusing on metastasis-initiating cell populations, we observed close proximity to specific fibroblast and macrophage subtypes, a relationship maintained between primary and metastatic tumors. Notably, high CK14 in luminal cancer cells and high vimentin in TNBC cells correlated with close proximity to specific macrophage subtypes (CD163intCD206intPDL1intHLA-DR+ or PDL1highARG1high). Our in-depth spatial analysis demonstrates that metastasis-initiating cancer cells consistently colocalizes with distinct cell populations within the TME, suggesting a role for these cell-cell interactions in promoting metastasis.

Phosphodiesterase-5 inhibition collaborates with vaccine-based immunotherapy to reprogram myeloid cells in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is highly lethal and resistant to immunotherapy. Although immune recognition can be enhanced with immunomodulatory agents including checkpoint inhibitors and vaccines, few patients experience clinical efficacy because the tumor immune microenvironment (TiME) is dominated by immunosuppressive myeloid cells that impose T cell inhibition. Inhibition of phosphodiesterase-5 (PDE5) was reported to downregulate metabolic regulators arginase and inducible NOS in immunosuppressive myeloid cells and enhance immunity against immune-sensitive tumors, including head and neck cancers. We show for the first time to our knowledge that combining a PDE5 inhibitor, tadalafil, with a mesothelin-specific vaccine, anti-programmed cell death protein 1, and anti-cytotoxic T lymphocyte-associated protein 4 yields antitumor efficacy even against immune-resistant PDAC. To determine immunologic advantages conferred by tadalafil, we profiled the TiME using mass cytometry and single-cell RNA-sequencing analysis with Domino to infer intercellular signaling. Our analyses demonstrated that tadalafil reprograms myeloid cells to be less immunosuppressive. Moreover, tadalafil synergized with the vaccine, enhancing T cell activation including mesothelin-specific T cells. Tadalafil treatment was also associated with myeloid/T cell signaling axes important for antitumor responses (e.g., Cxcr3, Il12). Our study shows that PDE5 inhibition combined with vaccine-based immunotherapy promotes pro-inflammatory states of myeloid cells, activation of T cells, and enhanced myeloid/T cell crosstalk to yield antitumor efficacy against immune-resistant PDAC.

3D histology reveals that immune response to pancreatic precancers is heterogeneous and depends on global pancreas structure.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer for which few effective therapies exist. Immunotherapies specifically are ineffective in pancreatic cancer, in part due to its unique stromal and immune microenvironment. Pancreatic intraepithelial neoplasia, or PanIN, is the main precursor lesion to PDAC. Recently it was discovered that PanINs are remarkably abundant in the grossly normal pancreas, suggesting that the vast majority will never progress to cancer. Here, through construction of 48 samples of cm3-sized human pancreas tissue, we profiled the immune microenvironment of 1,476 PanINs in 3D and at single-cell resolution to better understand the early evolution of the pancreatic tumor microenvironment and to determine how inflammation may play a role in cancer progression. We found that bulk pancreatic inflammation strongly correlates to PanIN cell fraction. We found that the immune response around PanINs is highly heterogeneous, with distinct immune hotspots and cold spots that appear and disappear in a span of tens of microns. Immune hotspots generally mark locations of higher grade of dysplasia or locations near acinar atrophy. The immune composition at these hotspots is dominated by naïve, cytotoxic, and regulatory T cells, cancer associated fibroblasts, and tumor associated macrophages, with little similarity to the immune composition around less-inflamed PanINs. By mapping FOXP3+ cells in 3D, we found that regulatory T cells are present at higher density in larger PanIN lesions compared to smaller PanINs, suggesting that the early initiation of PanINs may not exhibit an immunosuppressive response. This analysis demonstrates that while PanINs are common in the pancreases of most individuals, inflammation may play a pivotal role, both at the bulk and the microscopic scale, in demarcating regions of significance in cancer progression.

Unveiling the Impact of Smoking on Allergic Rhinitis: Disease Severity and Efficacy of Subcutaneous Immunotherapy.

OBJECTIVE: To evaluate the impact of smoking statuses on disease severity and subcutaneous immunotherapy (SCIT) efficacy in allergic rhinitis (AR). STUDY DESIGN: Open observational cohort study. SETTING: Tertiary referral center. METHODS: Five hundred and five AR patients undergoing dust mite allergen SCIT were categorized into never smokers, former smokers, and current smokers. AR severity was assessed using widely employed questionnaires. The changes in questionnaire scores pre- and post-SCIT were evaluated for SCIT efficacy. The differences in disease severity and SCIT efficacy were compared for different smoking statuses among AR patients. RESULTS: Compared to never smokers, former and current smokers exhibited higher proportion of male, alcohol, and asthma (P < .05). Current smokers had a greater prevalence of allergic conjunctivitis than former smokers (P < .05). Before SCIT, AR severity was similar across 3 groups, even after adjusting for confounders (P > .05). Current smokers reported lower SCIT efficacy in the first year (P < .05). By the third year, 3 groups showed comparable long-term efficacy (P > .05). However, current smokers experienced a significant decrease in benefits 2 years post-SCIT (P < .05) and lower improvement rates at the end of the 3-years SCIT period and 2 years following SCIT (P < .05). CONCLUSION: AR patients across different smoking statuses demonstrated similar baseline disease severity and long-time SCIT efficacy. Active smoking was associated with increased asthma risk, delayed early SCIT efficacy perception, reduced improvement over 3 years, and diminished benefits 2 years after SCIT. Prompt smoking cessation is crucial to mitigate these effects.

YY1 Contributes to the Inflammatory Responses of Mycobacterium tuberculosis-Infected Macrophages Through Transcription Activation-Mediated Upregulation TLR4.

Tuberculosis (TB) is a chronic respiratory infectious disease and is induced by Mycobacterium tuberculosis (M.tb) infection. Macrophages serve as the cellular home in immunoreaction against M.tb infection, which is tightly regulated through Toll-like receptor 4 (TLR4) expression. Therefore, this study is designed to explore the role and mechanism of TLR4 in mycobacterial injury in human macrophages (THP-1 cells) after M.tb infection. Cell proliferation and apoptosis were assessed using MTT, EdU, and flow cytometry assays. ELISA kits were utilized to assess the levels of Interleukin-6 (IL-6), IL-1ß, and tumor necrosis factor α (TNF-α). The binding between Yin-Yang-1 (YY1) and TLR4 promoter was predicted by JASPAR and verified using Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays. M.tb infection might repress THP-1 cell proliferation, and induce cell apoptosis and inflammatory response in a multiplicity of infection (MOI)-dependent manner. Moreover, M.tb infection increased the expression of TLR4 in HTP-1 cells in an MOI-dependent way, and its downregulation might overturn M.tb infection-mediated HTP-1 cell damage and inflammatory response. At the molecular level, YY1 was a transcription factor of TLR4 and promoted TLR4 transcription via binding to its promoter region. Besides, YY1 might activate the NF-kB signaling pathway via regulating TLR4. Meanwhile, TLR4 inhibitor BAY11-7082 might overturn the repression effect of TLR4 on M.tb-infected HTP-1 cell damage. YY1-activated TLR4 might aggravate mycobacterial injury in human macrophages after M.tb infection by the NF-kB pathway, providing a promising therapeutic target for TB treatment.

Natural Changbai mineral water reduces obesity risk through regulating metabolism and gut microbiome in a hyperuricemia male mouse model.

Access to clean and safe drinking water is essential. This study aimed to evaluate the effect of a kind of small molecular natural mineral water, C-cell mineral water on hyperuricemia male mice metabolism condition. A 13-week drinking water intervention study was conducted in Uox-knockout mice (KO). The hepatic metabolite profiling and related genes expression were detected by UPLC-TOF-MS and transcriptomic, and the gut microbiota of KO mice was determined by metagenomics sequencing. Results showed that the body weight of mice fed with C-cell water was remarkably lower than that of control mice on D 77 and D 91. Hepatic metabolite profiling revealed a shift in the pathway of glycine, serine and threonine metabolism, pantothenate and CoA biosynthesis, and biosynthesis of cofactors in KO mice fed with C-cell mineral water. Increased energy metabolism levels were related to increased hepatic expression of genes responsible for coenzyme metabolism and lipid metabolism. Gut microbiota was characterized by increasing activity of beneficial bacteria Blautia, and reducing activity of pathobiont bacteria Parasutterella. These genera have been reported to be associated with obesity. Small molecular mineral-rich natural water ingestion regulates metabolism and gut microbiota, protecting against obesity induced by hyperuricemia through mediating a microbiota-liver axis.

A stable liver-specific urate oxidase gene knockout hyperuricemia mouse model finds activated hepatic de novo purine biosynthesis and urate nephropathy.

Urate oxidase (Uox)-deficient mice could be an optimal animal model to study hyperuricemia and associated disorders. We develop a liver-specific conditional knockout Uox-deficient (UoxCKO) mouse using the Cre/loxP gene targeting system. These UoxCKO mice spontaneously developed hyperuricemia with accumulated serum urate metabolites. Blocking urate degradation, the UoxCKO mice showed significant de novo purine biosynthesis (DNPB) in the liver along with amidophosphoribosyltransferase (Ppat). Pegloticase and allopurinol reversed the elevated serum urate (SU) levels in UoxCKO mice and suppressed the Ppat up-regulation. Although urate nephropathy occurred in 30-week-old UoxCKO mice, 90 % of Uox-deficient mice had a normal lifespan without pronounced urate transport abnormality. Thus, UoxCKO mice are a stable model of human hyperuricemia. Activated DNPB in the UoxCKO mice provides new insights into hyperuricemia, suggesting increased SU influences purine synthesis.

Does skin prick test response intensity predict symptom severity and efficacy of subcutaneous immunotherapy in allergic rhinitis?

OBJECTIVES: To investigate the effect of response intensity of allergen skin prick test (SPT) on symptom severity and long-term efficacy of dust mite subcutaneous immunotherapy (SCIT) in allergic rhinitis (AR). METHODS: AR Patients diagnosed with dust mite allergy and completed 3 years of SCIT were collected and classified into three groups: grade 2 (SPT of + +), grade 3 (SPT of + + +) and grade 4 (SPT of + + + +). Comparisons between groups were performed to examine the associations of SPT categories and symptom severity and the long-term efficacy of SCIT in AR. RESULTS: 181 AR patients were included. There was no significant difference in the baseline TNSS, SMS, RQLQ and VAS, and particularly to symptom severity grading among three SPT grade groups (P > 0.05). The moderate-severe AR was more likely to be smoking and accompany with asthma and had higher prevalence of sensitization to cockroach, mixed grass and tree pollen than mild AR (P < 0.05). Prevalence of sensitization to cockroach, mixed grass, ragweed and animal dander was increased in AR patients with asthma and allergic conjunctivitis (P < 0.05). Furthermore, after 3 years of SCIT, no statistical differences in TNSS, SMS, RQLQ, VAS and long-term efficacy were observed among the three SPT grade groups (P > 0.05). Similarly, long-term outcomes of patients with different SPT grades did not differ among different clinical characteristics and different efficacy determination criteria (P > 0.05). CONCLUSIONS: The SPT response intensity cannot be used as an objective evaluation index for symptom severity and the long-term efficacy of SCIT in AR patients.

COVID-19 Vaccination and Gout Flare Risk in Patients With Infrequent or Frequent Flares: A Prospective Cohort Study.

OBJECTIVE: To assess post-COVID-19 vaccination gout flare risk with differing baseline flare burden. METHODS: We prospectively studied gout patients with infrequent or frequent flares, defined as ≤1 flare/year or ≥2 flares/year, respectively. COVID-19 vaccine-naive patients managed with urate-lowering therapy between February and June 2021 were included and voluntarily decided on vaccination. Participants were followed for 12 weeks after enrollment or first vaccine dose. Gout flares and risk factors were compared between groups. RESULTS: Of 530 participants, 308 (58.1%) had infrequent flares and 222 (41.9%) had frequent flares at baseline, with 248 (142 infrequent and 106 frequent) receiving two-dose COVID-19 vaccination. Vaccination increased cumulative flare incidence at 12 weeks in the infrequent but not the frequent flare group (26.1% vs 10.8%, P = 0.001, compared with 60.4% vs 65.5%, P = 0.428). Flare incidence in the final 4 weeks of observation decreased significantly only in the vaccinated infrequent flare group (4.3% vs 12.0%, P = 0.017). Multivariable analyses showed that vaccination (odds ratio [OR] 2.82, 95% confidence interval [95% CI] 1.50-5.30, P = 0.001), flare in the preceding year (OR 1.95, 95% CI 1.03-3.71, P = 0.04), and body mass index (OR 1.09, 95% CI 1.01-1.19, P = 0.03) were independently associated with increased flare risk in the infrequent flare group. Baseline serum urate (mg/dl) was an independent risk factor in the frequent flare group (OR 1.23, 95% CI 1.05-1.45, P = 0.012). CONCLUSION: COVID-19 vaccination was associated with increased early gout flares only in patients with previously infrequent flares.

Superiority of Low-Dose Benzbromarone Add-On to Low-Dose Febuxostat Compared With Febuxostat Monotherapy in Gout With Combined-Type Hyperuricemia.

OBJECTIVE: There is an unmet need for simpler urate-lowering therapy (ULT) regimens that achieve the serum urate target and improve the overall quality of gout care. We report a comparative effectiveness trial of febuxostat monotherapy versus benzbromarone add-on to low-dose febuxostat in gout specifically with combined renal urate underexcretion and overload. METHODS: A prospective randomized trial was conducted on patients with combined-type hyperuricemia and estimated glomerular filtration rate >60 mL/min/1.73 m2 1:1 randomly assigned to febuxostat and benzbromarone combination therapy (initially febuxostat at 20 mg/day, with benzbromarone at 25 mg/day added onto 20 mg/day of febuxostat if not at target) or febuxostat monotherapy (initially 20 mg/day, escalating to 40 mg/day if not at target). The primary end point at 12 weeks was the proportion achieving a serum urate (SU) level <360 µmol/L. Other outcomes included altered liver and kidney function, new-onset urolithiasis, and gout flares. RESULTS: There were 250 participants randomized; 219 completed 12-week treatment. More patients in the febuxostat and benzbromarone combination group achieved the SU target compared to patients in the febuxostat monotherapy group (75.5% vs 47.7%; odds ratio 3.37 [95% confidence interval 1.90-5.98]). Safety profiles were comparable between the two groups. CONCLUSION: Simply adding on low-dose benzbromarone (25 mg/day) to low-dose (20 mg/day) febuxostat showed superior urate lowering compared to febuxostat monotherapy in gout with a combined-type hyperuricemia. For selected patients, expedited achievement of the SU target in more than 75% of patients using one titration step and low xanthine oxidase inhibitor and uricosuric doses is a potential alternative to standard ULT regimens.

Fabrication of Schiff-base crosslinked films modified dialdehyde starch with excellent UV-blocking and antibacterial properties for fruit preservation.

Starch-based films have received considerable attention, owing to their commendable biocompatible and biodegradable properties; however, their poor ultraviolet (UV)-blocking and antibacterial performances limit their application in fruit preservation. Herein, bio-based bifunctional benzoxazine (Bi-BOZ) compounds with different carbon chain lengths were synthesized, and the influence of chain lengths on the antibacterial effect was explored. Benzoxazine with 1,12-dodecanediamine as the amine source (BOZ-DDA) exhibited excellent antibacterial and antibiofilm activities, with minimum inhibitory concentrations of 21.7 ± 2.2 and 23.3 ± 2.6 µg/mL against Escherichia coli and Staphylococcus aureus, respectively, mainly because the electrostatic attraction and hydrophobic effect of BOZ-DDA, effectively disrupted the bacterial integrity. DS/DDA films with hydrophobic, antibacterial, and UV-resistant abilities were prepared by the Schiff-base reaction between BOZ-DDA and dialdehyde starch (DS). The interactions between the films increased with BOZ-DDA content, enhanced mechanical and barrier properties. DS/DDA films exhibited acid-responsive antibacterial activity attributed to the acid hydrolysis of Schiff bases, released of BOZ-DDA from the films, and the protonation of BOZ-DDA. DS/DDA films exhibited commendable antibacterial and anti-ultraviolet characteristics compared to commercially available films, allowing them to prevent the degradation of mangoes and grapes. As sustainable antibacterial materials, the multifunctional DS/DDA films manifest promising prospects in fruit preservation packaging applications.

Long-Term Efficacy and Safety of Subcutaneous Immunotherapy in Monosensitized and Polysensitized Children With Allergic Rhinitis.

OBJECTIVE: To evaluate the efficacy and safety of dust mite subcutaneous immunotherapy (SCIT) in monosensitized and polysensitized children with allergic rhinitis (AR). STUDY DESIGN: Prospective cohort study. SETTING: Tertiary referral center. METHODS: One hundred thirty children were enrolled and categorized into 2 groups: monosensitized to only dust mites and polysensitized to at least 1 additional allergen beyond dust mites. All patients received SCIT targeting dust mites for 3 years, followed by a 5-year monitoring period. The Total Nasal Symptom Score (TNSS), Symptom and Medication Score (SMS), Visual Analogue Scale (VAS), and Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) were assessed before SCIT (T0); at 1 (T1) and 2 (T2) years of SCIT; immediately after SCIT (T3); and 2 years post-SCIT (T5). Safety was assessed based on adverse events (AEs). RESULTS: Fifty-one monosensitized and 50 polysensitized children completed the study. At T3, 47 monosensitized and 46 polysensitized children were effectively treated, with no significant between-group difference in efficacy (P > .05). The TNSS, SMS, VAS scores, and RQLQ score were significantly lower at T1, T2, T3, and T5 than at T0 in both groups (P < .05). The differences in the TNSS, SMS, VAS score, and RQLQ score between the 2 groups were nonsignificant at T0, T1, T2, and T3 (P > .05), but significant at T5 (P < .05). No serious AEs were reported. CONCLUSION: Monosensitized and polysensitized children exhibited similar beneficial efficacy and safety after 3 years of dust mite SCIT. Monosensitized children derived more benefits 2 years after discontinuation.

Immune landscape of tertiary lymphoid structures in hepatocellular carcinoma (HCC) treated with neoadjuvant immune checkpoint blockade.

Neoadjuvant immunotherapy is thought to produce long-term remissions through induction of antitumor immune responses before removal of the primary tumor. Tertiary lymphoid structures (TLS), germinal center-like structures that can arise within tumors, may contribute to the establishment of immunological memory in this setting, but understanding of their role remains limited. Here, we investigated the contribution of TLS to antitumor immunity in hepatocellular carcinoma (HCC) treated with neoadjuvant immunotherapy. We found that neoadjuvant immunotherapy induced the formation of TLS, which were associated with superior pathologic response, improved relapse free survival, and expansion of the intratumoral T and B cell repertoire. While TLS in viable tumor displayed a highly active mature morphology, in areas of tumor regression we identified an involuted TLS morphology, which was characterized by dispersion of the B cell follicle and persistence of a T cell zone enriched for ongoing antigen presentation and T cell-mature dendritic cell interactions. Involuted TLS showed increased expression of T cell memory markers and expansion of CD8+ cytotoxic and tissue resident memory clonotypes. Collectively, these data reveal the circumstances of TLS dissolution and suggest a functional role for late-stage TLS as sites of T cell memory formation after elimination of viable tumor.

Cadmium exposure dysregulates purine metabolism and homeostasis across the gut-liver axis in a mouse model.

Cadmium (Cd) exposure has been associated with the development of enterohepatic circulation disorders and hyperuricemia, but the possible contribution of chronic low-dose Cd exposure to disease progression is still need to be explored. A mouse model of wild-type mice (WT) and Uox-knockout mice (Uox-KO) to find out the toxic effects of chronic low-dose Cd exposure on liver purine metabolism by liquid chromatography-mass spectrometry (LC-MS) platform and associated intestinal flora. High throughput omics analysis including metabolomics and transcriptomics showed that Cd exposure can cause disruption of purine metabolism and energy metabolism. Cd changes several metabolites associated with purine metabolism (xanthine, hypoxanthine, adenosine, uridine, inosine) and related genes, which are associated with elevated urate levels. Microbiome analysis showed that Cd exposure altered the disturbance of homeostasis in the gut. Uox-KO mice were more susceptible to Cd than WT mice. Our findings extend the understanding of potential toxicological interactions between liver and gut microbiota and shed light on the progression of metabolic diseases caused by Cd exposure.

Comparison of the different monosodium urate crystals in the preparation process and pro-inflammation.

OBJECTIVES: The deposition of monosodium urate (MSU) crystals within synovial joints and tissues is the initiating factor for gout arthritis. Thus, MSU crystals are a vital tool for studying gout's molecular mechanism in animal and cellular models. This study mainly compared the excellence and worseness of MSU crystals prepared by different processes and the degree of inflammation induced by MSU crystals. METHODS: MSU crystals were prepared using neutralization, alkali titration, and acid titration methods. The crystals' shape, length, quality, and uniformity were observed by polarized light microscopy and calculated by the software Image J. The foot pad and air pouch models were used to assess the different degrees of inflammation induced by the MSU crystals prepared by the three different methods at different time points. Paw swelling was evaluated by caliper. In air pouch lavage fluid, inflammatory cell recruitment was measured by hemocytometer, and the level of IL-1ß, TNF-α, and IL-18 by ELISA. Inflammatory cell infiltration was assayed by immunohistochemistry of air pouch synovial slices. RESULTS: For the preparation of MSU crystals with the same uric acid, the quantity acquired by the alkalization method was highest, followed by neutralization, with the acid titration method being the lowest. The crystals prepared by neutralization were the longest. The swelling index of the foot pad induced by MSU crystals prepared by acid titration was significantly lower than that of the other methods at 24 h. The inflammatory cell recruitment and level of IL-1ß, TNF-α, and IL-18 in air pouch lavage fluid were lowest in animals with crystals prepared by acid titration. IL-1ß secretion induced by MSU crystals prepared by acid titration was significantly lower than that of the other two groups, but there was no significant difference in IL-18 secretion between the three groups in THP-1 macrophages and BMDMs. CONCLUSIONS: All three methods can successfully prepare MSU crystals, but the levels of inflammation induced by the crystals prepared by the three methods were not identical. The degree of inflammation induced by MSU crystals prepared by neutralization and alkalization is greater than by acid titration, but the quantity of MSU crystals obtained by the alkalization method is higher and less time-consuming. Apparently, the window of inflammation triggered by acid titration preparation is shorter compared to other forms of crystal preparation. Overall, MSU crystals prepared by the alkaline method should be recommended for studying the molecular mechanisms of gout in animal and cellular models.

Neoadjuvant Pembrolizumab in Localized Microsatellite Instability High/Deficient Mismatch Repair Solid Tumors.

PURPOSE: Pembrolizumab significantly improves clinical outcomes in advanced/metastatic microsatellite instability high (MSI-H)/deficient mismatch repair (dMMR) solid tumors but is not well studied in the neoadjuvant space. METHODS: This is a phase II open-label, single-center trial of localized unresectable or high-risk resectable MSI-H/dMMR tumors. Treatment is pembrolizumab 200 mg once every 3 weeks for 6 months followed by surgical resection with an option to continue therapy for 1 year followed by observation. To continue on study, patients are required to have radiographic or clinical benefit. The coprimary end points are safety and pathologic complete response. Key secondary end points are response rate and organ-sparing at one year for patients who declined surgery. Exploratory analyses include interrogation of the tumor immune microenvironment using imaging mass cytometry. RESULTS: A total of 35 patients were enrolled, including 27 patients with colorectal cancer and eight patients with noncolorectal cancer. Among 33 evaluable patients, best overall response rate was 82%. Among 17 (49%) patients who underwent surgery, the pathologic complete response rate was 65%. Ten patients elected to receive one year of pembrolizumab followed by surveillance without surgical resection (median follow-up of 23 weeks [range, 0-54 weeks]). An additional eight did not undergo surgical resection and received less than 1 year of pembrolizumab. During the study course of the trial and subsequent follow-up, progression events were seen in six patients (four of whom underwent salvage surgery). There were no new safety signals. Spatial immune profiling with imaging mass cytometry noted a significantly closer proximity between granulocytic cells and cytotoxic T cells in patients with progressive events compared with those without progression. CONCLUSION: Neoadjuvant pembrolizumab in dMMR/MSI-H cancers is safe and resulted in high rates of pathologic, radiographic, and endoscopic response, which has implications for organ-sparing strategies.

CyTOF protocol for immune monitoring of solid tumors from mouse models.

Techniques for robust immune profiling of mouse tumor and blood are key to understanding immunological responses in mouse models of cancer. Here, we describe mass cytometry (cytometry by time-of-flight) procedures to facilitate high-parameter profiling of low-volume survival blood samples and end-of-study tumor samples. We employ live-cell barcoding systems to mark all cells from each tumor and blood to improve cost-effectiveness and minimize batch effects. For complete details on the use and execution of this protocol, please refer to Charmsaz et al. (2021).1.