Accelerated involution of germinal center in palatine tonsils in IgA nephropathy.

BACKGROUND: Altered immunological responses in the palatine tonsils may be involved in the pathogenesis of IgA nephropathy (IgAN). The germinal center serves as the site for antigen-specific humoral immune responses in the palatine tonsils. Germinal center involution is frequently observed in the palatine tonsils of IgAN (IgAN tonsils). However, the pathogenic significance of these characteristic changes remains unclear. This study aimed to investigate the morphological changes in secondary lymphoid follicles in IgAN tonsils and to evaluate the correlation between the morphometric results and the clinicopathological severity of IgAN. METHODS: The tonsils of age-matched patients with recurrent tonsillitis (RT tonsils) were used as controls. The correlation between the degree of lymphoid follicular involution and histopathological severities in clinical or kidney biopsy was evaluated. RESULTS: In total, 87 patients with IgAN were included (48% male, median age 35 years, median estimated glomerular filtration rate: 74 mL/min/1.73 m2). Compared to RT tonsils, IgAN tonsils showed smaller median sizes of lymphoid follicles and germinal centers (P < 0.001). The relative areas of lymphoid follicles (%LFA) and germinal centers (%GCA) in the total tonsillar tissue were smaller in the IgAN tonsils than in the RT tonsils (P < 0.001). In contrast, the median proportion of mantle zones in the total tonsillar tissue was comparable between the groups. A lower %LFA was associated with a longer period from the onset of urinary abnormalities to biopsy diagnosis and higher urinary protein excretion (P = 0.01). %LFA showed significant negative correlations with frequencies of glomeruli with both global and segmental sclerosis. CONCLUSIONS: The present study confirmed accelerated germinal center involution in the tonsils of patients with IgAN. This characteristic change in the IgAN tonsil correlates with heavy proteinuria and advanced chronic histopathological changes in the kidneys, thereby suggesting the involvement of repeated tonsillar immunoreactions during IgAN progression.

Histological Correlation between Tonsillar and Glomerular Lesions in Patients with IgA Nephropathy Justifying Tonsillectomy: A Retrospective Cohort Study.

Tonsillectomy with steroid pulse therapy (SPT) has been established as an effective treatment for immunoglobulin A nephropathy (IgAN) in Japan. However, the underlying mechanisms supporting tonsillectomy remain unclear. This study assessed palatine tonsils from 77 patients with IgAN, including 14 and 63 who received SPT before and after tonsillectomy, respectively. Tonsils from 21 patients with chronic tonsillitis were analyzed as controls. Specific tonsillar lesions were confirmed in patients with IgAN, correlating with active or chronic renal glomerular lesions and SPT. T-nodule and involution of lymphoepithelial symbiosis scores in tonsils correlated with the incidence of active crescents and segmental sclerosis in the glomeruli, respectively. The study revealed an essential role of the tonsil-glomerular axis in early active and late chronic phases. Moreover, the SPT-preceding group demonstrated no changes in the T-nodule score, which correlated with active crescent formation, but exhibited a considerable shrinkage of lymphatic follicles that produced aberrant IgA1. The study underscores the involvement of innate and cellular immunity in IgAN and advocates for tonsillectomy as a necessary treatment alongside SPT for IgAN, based on a stepwise process.

Tissue-specific sex differences in pediatric and adult immune cell composition and function.

Sex-based differences in immune cell composition and function can contribute to distinct adaptive immune responses. Prior work has quantified these differences in peripheral blood, but little is known about sex differences within human lymphoid tissues. Here, we characterized the composition and phenotypes of adaptive immune cells from male and female ex vivo tonsils and evaluated their responses to influenza antigens using an immune organoid approach. In a pediatric cohort, female tonsils had more memory B cells compared to male tonsils direct ex vivo and after stimulation with live-attenuated but not inactivated vaccine, produced higher influenza-specific antibody responses. Sex biases were also observed in adult tonsils but were different from those measured in children. Analysis of peripheral blood immune cells from in vivo vaccinated adults also showed higher frequencies of tissue homing CD4 T cells in female participants. Together, our data demonstrate that distinct memory B and T cell profiles are present in male vs. female lymphoid tissues and peripheral blood respectively and suggest that these differences may in part explain sex biases in response to vaccines and viruses.

Predictive values of Mallampati score, tonsillar size, and BMI z-score in the presence and severity of obstructive sleep apnea in pediatric population.

Background/aim: Obstructive sleep apnea (OSA) is a common sleep-related breathing disorder in children. Determination of risk factors for the development of OSA is essential for early diagnosis and treatment of the disease and decreases the risk of negative consequences. This study aimed to investigate the predictive values of Mallampati score, tonsillar size, and BMI z-score in the presence and severity of OSA in children. Materials and methods: This prospective cross-sectional study included 114 children with OSA symptoms. All children were assessed by BMI z-score, Mallampati score, and tonsillar size and underwent overnight polysomnography. They were consecutively selected and assigned to 4 groups as follows: Group 1 included normal-weight with a low Mallampati score; Group 2 involved normal-weight with a high Mallampati score; Group 3 included obese with a low Mallampati score; and Group 4 involved obese with a high Mallampati score. Results: Of the 114 included children, 58 were female and 56 were male, with a mean age of 13.1 ± 2.9 years. OSA frequency and apnea-hypopnea index were significantly higher in group 4 compared with other groups (p = 0.003 and p < 0.0001, respectively), whereas average and minimum spO2 were significantly lower (for both, p = 0.001). Mallampati score and BMI z-score were found to be significant for predicting OSA (odds ratio = 4.147, 95% CI: 1.440-11.944; p = 0.008 and odds ratio = 1.760, 95% CI: 1.039-2.980; p = 0.035, respectively). Among OSA patients, the Mallampati score, tonsillar size, and BMI z-score were found to be significant for predicting OSA severity (odds ratio = 4.520, 95% CI: 1.332-15.335, p = 0.015, odds ratio = 9.177, 95% CI: 2.513-33.514, p = 0.001, and odds ratio = 2.820, 95% CI: 1.444-5.508; p = 0.002, respectively). Conclusion: The coexistence of the Mallampati score and BMI z-score significantly increases the presence of OSA in children. Mallampati score, tonsillar size, and BMI z-score are promising parameters for predicting OSA severity.

Ultrasensitive and Multiplexed Protein Imaging with Clickable and Cleavable Fluorophores.

Single-cell spatial proteomic analysis holds great promise to advance our understanding of the composition, organization, interaction, and function of the various cell types in complex biological systems. However, the current multiplexed protein imaging technologies suffer from low detection sensitivity, limited multiplexing capacity, or are technically demanding. To tackle these issues, here, we report the development of a highly sensitive and multiplexed in situ protein profiling method using off-the-shelf antibodies. In this approach, the protein targets are stained with horseradish peroxidase (HRP) conjugated antibodies and cleavable fluorophores via click chemistry. Through repeated cycles of target staining, fluorescence imaging, and fluorophore cleavage, many proteins can be profiled in single cells in situ. Applying this approach, we successfully quantified 28 different proteins in human formalin-fixed paraffin-embedded (FFPE) tonsil tissue, which represents the highest multiplexing capacity among the tyramide signal amplification (TSA) methods. Based on their unique protein expression patterns and their microenvironment, ∼820,000 cells in the tissue are classified into distinct cell clusters. We also explored the cell-cell interactions between these varied cell clusters and observed that different subregions of the tissue are composed of cells from specific clusters.

Oral ulcer treatment using human tonsil-derived mesenchymal stem cells encapsulated in trimethyl chitosan hydrogel: an animal model study.

BACKGROUND: Oral ulcers are a common side effect of chemotherapy and affect patients' quality of life. While stem cell transplantation is a potential treatment for oral ulcers, its efficacy is limited as the stem cells tend to remain in the affected area for a short time. This study aims to develop a treatment for oral ulcers by using trimethyl chitosan (TMC) hydrogel with human tonsil-derived stem cells (hTMSCs) to increase the therapeutic effect of stem cells and investigate their effectiveness. METHODS: Animals were divided into four experimental groups: Control, TMC hydrogel, hTMSCs, and hTMSCs loaded in TMC hydrogel (Hydrogel + hTMSCs) (each n = 8). Oral ulcers were chemically induced by anesthetizing the rats followed by injection of dilute acetic acid in the right buccal mucosa. After confirming the presence of oral ulcers in the animals, a single subcutaneous injection of 100 µL of each treatment was applied to the ulcer area. Histological analyses were performed to measure inflammatory cells, oral mucosal thickness, and fibrosis levels. The expression level of inflammatory cytokines was also measured using RT-PCR to gauge therapeutic the effect. RESULTS: The ulcer size was significantly reduced in the TMC hydrogel + hTMSCs group compared to the control group. The stem cells in the tissue were only observed until Day 3 in the hTMSCs treated group, while the injected stem cells in the TMC Hydrogel + hTMSCs group were still present until day 7. Cytokine analysis related to the inflammatory response in the tissue confirmed that the TMC Hydrogel + hTMSCs treated group demonstrated superior wound healing compared to other experimental groups. CONCLUSION: This study has shown that the adhesion and viability of current stem cell therapies can be resolved by utilizing a hydrogel prepared with TMC and combining it with hTMSCs. The combined treatment can promote rapid healing of oral cavity wounds by enhancing anti-inflammatory effects and expediting wound healing. Therefore, hTMSC loaded in TMC hydrogel was the most effective wound-healing approach among all four treatment groups prolonging stem cell survival. However, further research is necessary to minimize the initial inflammatory response of biomaterials and assess the safety and long-term effects for potential clinical applications.

Effect of palatine tonsil tumor resection on postoperative velopharyngeal insufficiency in transoral surgery.

BACKGROUND: Velopharyngeal insufficiency (VPI) is a known complication of transoral surgery (TOS) for oropharyngeal HPV-mediated squamous cell carcinoma. Controversy exists regarding adequate resection margins for balancing functional and oncologic outcomes. METHODS: This retrospective study was exempted by the IRB. Patients who underwent TOS from January 2017 to October 2022 were included. Patient characteristics, treatment details, and oncologic and functional outcomes were evaluated. RESULTS: Fifty-five patients were included. Mean and median follow-up was 34 months. 98% of patients were AJCC stage I/II. Recurrence-free survival was 96% with no local recurrences. Univariate analysis demonstrated an association between VPI and pT stage (p = 0.035), medial pterygoid resection (p = 0.049), and palatal attachment sacrifice (p < 0.001). Multivariate analysis showed sacrifice of the palatal attachments remained a significant risk for VPI (p = 0.009). CONCLUSION: Loss of soft palate pharyngeal attachments is an independent risk factor for VPI. When oncologically appropriate, the palatal attachments to the pharynx may be preserved.

Paired qualitative and quantitative analysis of bacterial microcolonies in the tonsils of patients with tonsillar hyperplasia.

The discovery of bacterial microcolonies in tonsillar tissue of patients with tonsillar hyperplasia has raised the question of their role in provoking the local immune response. Tonsils collected from patients undergoing tonsillectomy were stained for three clinically relevant bacterial taxa and lymphocytes. The bacterial composition and abundance of microcolonies was investigated using a combination of laser-microdissection, amplicon sequencing and Droplet Digital polymerase chain reaction. Microcolonies were detected in most samples (32/35) with a high prevalence of Haemophilus influenzae (78% of samples). B and T cell lymphocytes were significantly higher in the epithelium adjacent to microcolonies compared to epithelium distal to microcolonies. Furthermore, significant positive and negative correlations were identified between bacterial taxa and lymphocytes. Genus Streptococcus, which includes Group A Streptococcus (traditionally described as the main pathogen of tonsillar hyperplasia), was found in low abundance in this study. These results suggest other potential pathogens may be involved in stimulating the local immune response leading to tonsillar hyperplasia.

An Automated and Fast Sample Preparation Workflow for Laser Microdissection Guided Ultrasensitive Proteomics.

Spatial tissue proteomics integrating whole-slide imaging, laser microdissection, and ultrasensitive mass spectrometry is a powerful approach to link cellular phenotypes to functional proteome states in (patho)physiology. To be applicable to large patient cohorts and low sample input amounts, including single-cell applications, loss-minimized and streamlined end-to-end workflows are key. We here introduce an automated sample preparation protocol for laser microdissected samples utilizing the cellenONE robotic system, which has the capacity to process 192 samples in 3 h. Following laser microdissection collection directly into the proteoCHIP LF 48 or EVO 96 chip, our optimized protocol facilitates lysis, formalin de-crosslinking, and tryptic digest of low-input archival tissue samples. The seamless integration with the Evosep ONE LC system by centrifugation allows 'on-the-fly' sample clean-up, particularly pertinent for laser microdissection workflows. We validate our method in human tonsil archival tissue, where we profile proteomes of spatially-defined B-cell, T-cell, and epithelial microregions of 4000 µm2 to a depth of ∼2000 proteins and with high cell type specificity. We finally provide detailed equipment templates and experimental guidelines for broad accessibility.

Pain management after tonsil surgery in children and adults-A national survey related to pain outcome measures from the Swedish Quality Register for tonsil surgery.

OBJECTIVE: The primary aim of this study was to describe the current practice regarding pain management in relation to tonsil surgery among Ear Nose and Throat (ENT) clinics in Sweden. The secondary aim was to determine the impact of the provider's regime of rescue analgesics on the pain related Patient Reported Outcome Measures (pain-PROMs) from the Swedish Quality Register for Tonsil Surgery (SQTS). MATERIALS & METHODS: A descriptive cross-sectional study originating from a validated web-based questionnaire. The survey enrolled one respondent from each ENT clinic (47/48 participated) nationally. Pain-PROMs from the SQTS, recorded from October 2019 to October 2022, were included (8163 tonsil surgeries). RESULTS: Paracetamol was used by all enrolled ENT clinics as preemptive analgesia. The addition of COX inhibitors was used in 40% of the clinics. Betamethasone was usually administered, to prevent pain and nausea (92%). All clinics gave postdischarge instructions on multimodal analgesia with COX inhibitors and paracetamol. Rescue analgesics were prescribed after tonsillectomy for 77% of adults, 62% of older children, 43% of young children and less often after tonsillotomy. The most frequently prescribed rescue analgesic was clonidine in children (55%) and oxycodone in adults (72%). A high proportion of patients reported contact with health care services due to postoperative pain (pain-PROMs/ SQTS). Tonsillectomy procedures were associated with the highest rates of contacts (children/adolescents 13-15%; adults 26%), while tonsillotomy were associated with lower rates, (5-7% of children/adolescents). There was no significant difference in the frequency of health care contacts due to pain regarding whether clinics routinely prescribed rescue analgesics or not after tonsillectomy. CONCLUSION: The Swedish analgesic regimen after tonsil surgery is good overall. Nevertheless, there is a need for increased awareness and knowledge to achieve optimal patient recovery. Pain-PROM data demonstrate the call for improvement in pain management after tonsil surgery.

Clinical manifestations, endoscopic findings and outcomes of tonsillar fossa foreign bodies in dogs: seven cases (2020-2023).

OBJECTIVES: Clinical presentation and outcomes of dogs with tonsillar foreign bodies are not described in the literature. The objective of this case series is to describe the presence, clinical manifestations, endoscopic findings and outcomes of foreign bodies in the palatine tonsillar fossa of dogs. MATERIALS AND METHODS: Medical records of dogs in which tonsillar fossa foreign bodies were removed by endoscopy between 2020 and 2023 at a referral centre were reviewed. Dogs were included if complete medical records were available. RESULTS: Seven dogs were included. Among all clinical signs reported, acute onset of throat clearing occurred in all dogs. Palatine tonsil enlargement and protrusion from the tonsillar fossa were observed in four of seven dogs. Foreign bodies were removed under endoscopic guidance in all dogs; a vegetal foreign body was found in all but one dog (6/7). Complete resolution of clinical signs was reported in the five dogs for which follow-up information was available. CLINICAL SIGNIFICANCE: The tonsillar fossa is a rare but possible location for foreign bodies in dogs and should be explored even in the absence of macroscopic lesions of the tonsils, since some dogs could have a normal palatine tonsil appearance. Acute throat clearing should raise the suspicion of tonsillar fossa foreign bodies. The prognosis appears good after removal.

Low usefulness of reporting tonsillar PCR Ct-values in pharyngeal infections with Fusobacterium necrophorum.

Tonsillar Fusobacterium necrophorum PCR Ct-values were higher in participants with asymptomatic tonsillar carriage than patients with pharyngeal infections. However, Ct-values were not associated with severity of disease or predictive of development of complications and hence lacked clinical usefulness. The reporting of F. necrophorum Ct-values in clinical samples is not recommended.

The relationship between posttonsillectomy bleeding to hemogram and coagulation parameters.

PURPOSE: Tonsillectomy is a common surgery performed for indications such as chronic tonsilitis, tonsil hypertrophy and obsructive sleep apnea. Although posttonsillectomy bleeding (PTB) is rare and can be controlled with simple interventions in many patients, it is one of the most feared complications of tonsillectomy surgery. In our study, we investigated the effects of changes in hemogram and coagulation values and seasonal effects on PTB. METHODS: Pediatric and adult patients who underwent tonsillectomy with cold knife method between August 2020 and August 2023 in our clinic were retrospectively reviewed. Demographic data, hemogram and coagulation values of the patients in the control and study groups were recorded and the differences between the two groups in terms of these parameters were evaluated. RESULTS: Our study included 991 patients aged 1-51 years. The rate of PTB was calculated as 2.82%. No patient with primary PTB was found. The duration of bleeding development was 7.03 days. Age, WBC and neutrophil values were statistically significantly higher in the study group. There were no significant differences between two groups in terms of gender, season and other hemogram and coagulation parameters. CONCLUSIONS: Age, high WBC and neutrophil levels were determined as possible risk factors for PTB. Seasonal and gender distribution, aPTT and INR values were similar in the two groups. In order to prevent and predict bleeding, detailed infection investigation should be performed and the risk of bleeding should be considered to increase with increasing age.

Characterization of pig tonsils as niches for the generation of Streptococcus suis diversity.

Streptococcus suis is a gram-positive bacterium that causes meningitis, septicemia, endocarditis, and other disorders in pigs and humans. We obtained 42 and 50 S. suis isolates from lesions of porcine endocarditis and palatine tonsils, respectively, of clinically healthy pigs in Japan; we then determined their sequence types (STs) by multilocus sequence typing (MLST), cps genotypes, serotypes, and presence of classical major virulence-associated marker genes (mrp, epf, and sly). The 42 isolates from endocarditis lesions were assigned to a limited number of STs and clonal complexes (CCs). On the other hand, the 50 isolates from tonsils were diverse in these traits and seemingly in the degree of virulence, suggesting that tonsils can accommodate a variety of S. suis isolates. The goeBURST full algorithm using tonsil isolates obtained in this study and those retrieved from the database showed that major CCs as well as many other clusters were composed of isolates originating from different countries, and some of the STs were very similar to each other despite the difference in country of origin. These findings indicate that S. suis with not only different but also similar mutations in the genome have survived in tonsils independently across different geographical locations. Therefore, unlike the lesions of endocarditis, the tonsils of pigs seemingly accommodate various S. suis lineages. The present study suggests that S. suis acquired its diversity by natural mutations during colonization and persistence in the tonsils of pigs.

An atlas of cells in the human tonsil.

Palatine tonsils are secondary lymphoid organs (SLOs) representing the first line of immunological defense against inhaled or ingested pathogens. We generated an atlas of the human tonsil composed of >556,000 cells profiled across five different data modalities, including single-cell transcriptome, epigenome, proteome, and immune repertoire sequencing, as well as spatial transcriptomics. This census identified 121 cell types and states, defined developmental trajectories, and enabled an understanding of the functional units of the tonsil. Exemplarily, we stratified myeloid slan-like subtypes, established a BCL6 enhancer as locally active in follicle-associated T and B cells, and identified SIX5 as putative transcriptional regulator of plasma cell maturation. Analyses of a validation cohort confirmed the presence, annotation, and markers of tonsillar cell types and provided evidence of age-related compositional shifts. We demonstrate the value of this resource by annotating cells from B cell-derived mantle cell lymphomas, linking transcriptional heterogeneity to normal B cell differentiation states of the human tonsil.

Causal association of gastroesophageal reflux disease with chronic sinusitis and chronic disease of the tonsils and adenoids.

BACKGROUND: Exploring bidirectional causal associations between gastroesophageal reflux disease (GERD) and chronic disease of the tonsils and adenoids and chronic sinusitis, respectively. METHODS: We first conducted a TSMR (two-sample mendelian randomization) study using the results of the inverse variance weighting method as the primary basis and bidirectional MR to rule out reverse causation. Subsequently, MVMR (multivariate MR) analysis was performed to identify phenotypes associated with SNPs and to explore the independent effect of GERD on two outcomes. Finally, we calculated MR-Egger intercepts to assess horizontal polytropy and Cochran's Q statistic to assess heterogeneity and ensure the robustness of the study. RESULTS: For each standard deviation increase in genetically predicted GERD rate, there was an increased risk of chronic disease of the tonsils and adenoids (OR 1.162, 95% CI 1.036-1.304, P: 1.06E-02) and of developing chronic sinusitis (OR 1.365, 95% CI 1.185-1.572, P: 1.52E-05), and there was no reverse causality. Causality for TSMR was obtained on the basis of IVW (inverse variance weighting) and appeared to be reliable in almost all sensitivity analyses, whereas body mass index may be a potential mediator of causality between GERD and chronic sinusitis. CONCLUSION: There is a causal association between GERD and chronic disease of the tonsils and adenoids and chronic sinusitis, respectively, and the occurrence of GERD increases the risk of developing chronic disease of the tonsils and adenoids and chronic sinusitis.

Activation-neutral gene editing of tonsillar CD4 T cells for functional studies in human ex vivo tonsil cultures.

The molecular and immunological properties of tissue-resident resting CD4 T cells are understudied due to the lack of suitable gene editing methods. Here, we describe the ex vivo culture and gene editing methodology ediTONSIL for CD4 T cells from human tonsils. Optimized CRISPR-Cas9 RNP nucleofection results in knockout efficacies of over 90% without requiring exogenous activation. Editing can be performed on multiple cell types in bulk cultures or on isolated CD4 T cells that can be labeled and reintroduced into their tissue environment. Importantly, CD4 T cells maintain their tissue-specific properties such as viability, activation state, or immunocompetence following reassembly into lymphoid aggregates. This highly efficient and versatile gene editing workflow for tonsillar CD4 T cells enables the dissection of molecular mechanisms in ex vivo cultures of human lymphoid tissue and can be adapted to other tonsil-resident cell types.

Tonsillar cytotoxic CD4 T cells are involved in the control of EBV primary infection in children.

CD4 T cells play a key role in Epstein Barr virus (EBV) infection, by modulating latent antigen expression, and exhibiting cytotoxic and regulatory properties. Our aim was to evaluate the presence of Granzyme B (GZMB) and Foxp3 CD4 T cells at different EBV infection status and latency profiles. We examined CD4, GZMB, Foxp3, IL10, TGF-ß, CD4-GZMB and CD4-Foxp3 expression at the tonsils of pediatric patients with different infective status and EBV latency profiles. CD4+, GZMB+, Foxp3+, CD4-GZMB+ and CD4-Foxp3+ cell counts were higher at the interfollicular region. Higher expression of CD4-GZMB was found in primary infected patients compared to healthy carriers. In patients that expressed latency III antigens, we demonstrated lower CD4+, CD4-GZMB+, CD4-Foxp3+ expression; a negative correlation between the immunoregulatory cytokine IL-10+ and GZMB+ as well as a positive correlation of IL-10+ and CD4+. In patients expressing the lytic protein BMRF1, a positive correlation of TGF-ß+ with CD4-GZMB+ and CD4-Foxp3+ was observed. Our findings indicate that CD4-GZMB+ cells are involved in the restriction of primary EBV infection in pediatric patients, which could partially explain the lack of symptoms, whereas both CD4-GZMB+ and CD4-Foxp3+ cells could be involved in the modulation of latency.