Peritumoral stroma and systemic inflammatory response in cervical cancer.

OBJECTIVE: To compare cervical stroma in advanced cervical cancer with the control group; to compare, in the pre-treatment period, hemogram parameters in patients with advanced cervical cancer with the same parameters as the control group; and to verify if there is an association of stromal markers with prognostic factors in cervical cancer. MATERIALS AND METHODS: We prospectively evaluated 16 patients diagnosed with advanced invasive cervical cancer. A control group of 22 patients was used (uterine leiomyoma). Immunohistochemistry was performed to verify the stromal immunostaining of alpha-smooth muscle actin (SMA) and fibroblast activation protein alpha (FAP). Immunostainings and hemogram parameters were compared using Fisher's exact and Mann-Whitney Test, respectively. RESULTS: Strong FAP immunostaining was more frequent in patients with cervical cancer when compared with patients with leiomyoma (P = 0.0002). Regarding SMA, strong immunostaining was also found more in the group of cancer patients compared to the control group (P < 0.00001). The neutrophil-lymphocyte ratio (NLR) values were higher in the cancer patient group compared to the control group (P = 0.0019). There was no association of the parameters studied with prognostic factors. CONCLUSIONS: Strong FAP and SMA immunostaining was found more in patients with cervical cancer when compared to the control group. NLR values were also higher in cervical cancer.

Tumor-suppressive miR-4732-3p is sorted into fucosylated exosome by hnRNPK to avoid the inhibition of lung cancer progression.

BACKGROUND: Aberrant fucosylation observed in cancer cells contributes to an augmented release of fucosylated exosomes into the bloodstream, where miRNAs including miR-4732-3p hold promise as potential tumor biomarkers in our pilot study. However, the mechanisms underlying the sorting of miR-4732-3p into fucosylated exosomes during lung cancer progression remain poorly understood. METHODS: A fucose-captured strategy based on lentil lectin-magnetic beads was utilized to isolate fucosylated exosomes and evaluate the efficiency for capturing tumor-derived exosomes using nanoparticle tracking analysis (NTA). Fluorescence in situ hybridization (FISH) and qRT-PCR were performed to determine the levels of miR-4732-3p in non-small cell lung cancer (NSCLC) tissue samples. A co-culture system was established to assess the release of miRNA via exosomes from NSCLC cells. RNA immunoprecipitation (RIP) and miRNA pull-down were applied to validate the interaction between miR-4732-3p and heterogeneous nuclear ribonucleoprotein K (hnRNPK) protein. Cell functional assays, cell derived xenograft, dual-luciferase reporter experiments, and western blot were applied to examine the effects of miR-4732-3p on MFSD12 and its downstream signaling pathways, and the impact of hnRNPK in NSCLC. RESULTS: We enriched exosomes derived from NSCLC cells using the fucose-captured strategy and detected a significant upregulation of miR-4732-3p in fucosylated exosomes present in the serum, while its expression declined in NSCLC tissues. miR-4732-3p functioned as a tumor suppressor in NSCLC by targeting 3'UTR of MFSD12, thereby inhibiting AKT/p21 signaling pathway to induce cell cycle arrest in G2/M phase. NSCLC cells preferentially released miR-4732-3p via exosomes instead of retaining them intracellularly, which was facilitated by the interaction of miR-4732-3p with hnRNPK protein for selective sorting into fucosylated exosomes. Moreover, knockdown of hnRNPK suppressed NSCLC cell proliferation, with the elevated levels of miR-4732-3p in NSCLC tissues but the decreased expression in serum fucosylated exosomes. CONCLUSIONS: NSCLC cells escape suppressive effects of miR-4732-3p through hnRNPK-mediated sorting of them into fucosylated exosomes, thus supporting cell malignant properties and promoting NSCLC progression. Our study provides a promising biomarker for NSCLC and opens a novel avenue for NSCLC therapy by targeting hnRNPK to prevent the "exosome escape" of tumor-suppressive miR-4732-3p from NSCLC cells.

Domain-Targeted Membrane Partitioning of Specific Proteins with DNA Nanodevices.

The cell membrane exhibits a remarkable complexity of lipids and proteins that dynamically segregate into distinct domains to coordinate various cellular functions. The ability to manipulate the partitioning of specific membrane proteins without involving genetic modification is essential for decoding various cellular processes but highly challenging. In this work, by conjugating cholesterols or tocopherols at the three bottom vertices of the DNA tetrahedron, we develop two sets of nanodevices for the selective targeting of lipid-order (Lo) and lipid-disorder (Ld) domains on the live cell membrane. By incorporation of protein-recognition ligands, such as aptamers or antibodies, through toehold-mediated strand displacement, these DNA nanodevices enable dynamic translocation of target proteins between these two domains. We first used PTK7 as a protein model and demonstrated, for the first time, that the accumulation of PTK7 to the Lo domains could promote tumor cell migration, while sequestering it in the Ld domains would inhibit the movement of the cells. Next, based on their modular nature, these DNA nanodevices were extended to regulate the process of T cell activation through manipulating the translocation of CD45 between the Lo and the Ld domains. Thus, our work is expected to provide deep insight into the study of membrane structure and molecular interactions within diverse cell signaling processes.

Rigorous Comparison of Extracellular Vesicle Processing to Enhance Downstream Analysis for Glioblastoma Characterization.

Extracellular vesicles (EVs) are highly sought after as a source of biomarkers for disease detection and monitoring. Tumor EV isolation, processing, and evaluation from biofluids is convoluted by EV heterogeneity and biological contaminants and is limited by technical processing efficacy. This study rigorously compares common bulk EV isolation workflows (size exclusion chromatography, SEC; membrane affinity, MA) alongside downstream RNA extraction protocols to investigate molecular analyte recovery. EV integrity and recovery is evaluated using a variety of technologies to quantify total intact EVs, total and surface proteins, and RNA purity and recovery. A comprehensive evaluation of each analyte is performed, with a specific emphasis on maintaining user (n = 2), biological (n = 3), and technical replicates (n≥3) under in vitro conditions. Subsequent study of tumor EV spike-in into healthy donor plasma samples is performed to further validate biofluid-derived EV purity and isolation for clinical application. Results show that EV surface integrity is considerably preserved in eluates from SEC-derived EVs, but RNA recovery and purity, as well as bulk protein isolation, is significantly improved in MA-isolated EVs. This study concludes that EV isolation and RNA extraction pipelines govern recovered analyte integrity, necessitating careful selection of processing modality to enhance recovery of the analyte of interest.

Employing the Anchor DSPE-PEG as a Redox Probe for Ratiometric Electrochemical Detection of Surface Proteins on Extracellular Vesicles with Aptamers.

Quantitative analysis of surface proteins on extracellular vesicles (EVs) has been considered to be a crucial approach for reflecting the status of diseases. Due to the diverse composition of surface proteins on EVs and the interference from nonvesicular proteins, accurately detecting the expression of surface proteins on EVs remains a challenging task. While membrane affinity molecules have been widely employed as EVs capture probes to address this issue, their inherent biochemical properties have not been effectively harnessed. In this paper, we found that the electrochemical redox activity of the DSPE-PEG molecule was diminished upon its insertion into the membrane of EVs. This observation establishes the DSPE-PEG molecule modified on the Au electrode surface as a capture and a redox probe for the electrochemical detection of EVs. By utilizing methylene blue-labeled aptamers, the targeted surface proteins of EVs can be detected by recording the ratio of the oxidation peak current of methylene blue and DSPE-PEG. Without complicated signal amplification, the detection limit for EVs is calculated to be 8.11 × 102 particles/mL. Using this platform, we directly analyzed the expression of CD63 and HER2 proteins on the surface of EVs in human clinical plasma samples, demonstrating its significant potential in distinguishing breast cancer patients from healthy individuals.

Size-exclusion chromatography-based extracellular vesicle size subtyping and multiplex membrane protein profiling for differentiating gastrointestinal cancer prognosis.

Extracellular vesicles (EVs), as a type of subcellular structure, have been extensively researched for their potential for developing advanced diagnostic technologies for various diseases. However, the biomolecular and biophysical heterogeneity of EVs has restricted their application in clinical settings. In this article, we developed a size-exclusion chromatography-based technique for simultaneous EV size subtyping and protein profiling. By eluting fluorescent aptamer-treated patient plasma through a size-exclusion column, the mixture can be classified into 50 nm aptamer-bound EVs, 100 nm aptamer-bound EVs and free-floating aptamers, which could further enable multiplex EV membrane protein profiling by analyzing the fluorescence intensities of EV-bound aptamers. Using this technique, we successfully identified EV size subtypes for differentiating gastrointestinal cancer prognosis states. Overall, we developed a rapid, user-friendly and low-cost EV size subtyping and protein profiling technique, which holds great potential for identifying crucial EV size subtypes for disease diagnosis in the clinic.

Double-Labeling Method for Visualization and Quantification of Membrane-Associated Proteins in Lactococcus lactis.

Lactococcus lactis displaying recombinant proteins on its surface can be used as a potential drug delivery vector in prophylactic medication and therapeutic treatments for many diseases. These applications enable live-cell mucosal and oral administration, providing painless, needle-free solutions and triggering robust immune response at the site of pathogen entry. Immunization requires quantitative control of antigens and, ideally, a complete understanding of the bacterial processing mechanism applied to the target proteins. In this study, we propose a double-labeling method based on a conjugated dye specific for a recombinantly introduced polyhistidine tag (to visualize surface-exposed proteins) and a membrane-permeable dye specific for a tetra-cysteine tag (to visualize cytoplasmic proteins), combined with a method to block the labeling of surface-exposed tetra-cysteine tags, to clearly obtain location-specific signals of the two dyes. This allows simultaneous detection and quantification of targeted proteins on the cell surface and in the cytoplasm. Using this method, we were able to detect full-length peptide chains for the model proteins HtrA and BmpA in L. lactis, which are associated with the cell membrane by two different attachment modes, and thus confirm that membrane-associated proteins in L. lactis are secreted using the Sec-dependent post-translational pathway. We were able to quantitatively follow cytoplasmic protein production and accumulation and subsequent export and surface attachment, which provides a convenient tool for monitoring these processes for cell surface display applications.

Label-free quantitative proteomic analysis of milk fat globule membrane proteins in porcine colostrum and mature milk.

Milk fat globule membrane (MFGM) proteins are nutritional components with various biological functions. This study aimed to analyze and compare MFGM proteins in porcine colostrum (PC) and porcine mature milk (PM), via label-free quantitative proteomics. In total, 3917 and 3966 MFGM proteins were identified in PC and PM milk, respectively. A total of 3807 common MFGM proteins were found in both groups, including 303 significant differentially expressed MFGM proteins. Gene Ontology (GO) analysis revealed that the differentially expressed MFGM proteins were mainly related to the cellular process, cell, and binding. The dominant pathway of the differentially expressed MFGM proteins was related to the phagosome according to Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. These results reveal crucial insights into the functional diversity of MFGM proteins in porcine milk during lactation and provide theoretical guidance for the development of MFGM proteins in the future.

Cell surface glycoproteomics: deciphering glycoproteins through a unique analytical capture approach.

Cell surface proteins (CSPs) are often involved in various biological processes such as cell-cell interactions, immune responses, and molecular transport. The abnormal expression of CSP usually indicates the occurrence and development of human diseases. Most CSPs are glycosylated and have been explored as potential drug targets and disease biomarkers; however, efficient isolation of CSPs from intracellular proteins is difficult due to their low abundance and strong hydrophobicity. The comprehensive characterization of surface glycoproteins remains a great challenge and is often underrepresented in proteomics. In recent years, unprecedented progress has been made in the mass spectrometry analysis of surface proteins, and CSP capture methods and mass spectrometry have been greatly developed. In this article, we aim to give a comprehensive overview of innovative analytical methods that can enrich CSPs, including centrifugation-based separation, phase partitioning, adhesion-based capture of surface proteins, antibody or lectin affinity, and biotin-based chemical labeling. Surface glycoproteins are captured by chemical oxidation of glycans or click chemistry for carbohydrate metabolic labeling. These techniques offer a wide range of applications for studying the function of cell surface receptors and identifying markers for diagnostic and therapeutic development.

Hybrid Supported Lipid Bilayers for Bioinspired Bioelectronics with Enhanced Stability.

A promising new class of biosensors leverages the sensing mechanisms of living cells by incorporating native transmembrane proteins into biomimetic membranes. Conducting polymers (CPs) can further improve the detection of electrochemical signals from these biological recognition elements through their low electrical impedance. Supported lipid bilayers (SLBs) on CPs mimic the structure and biology of the cell membrane to enable such sensing, but their extrapolation to new target analytes and healthcare applications has been difficult due to their poor stability and limited membrane properties. Blending native phospholipids with synthetic block copolymers to create a hybrid SLB (HSLB) may address these challenges by allowing for the tuning of chemical and physical properties during membrane design. We establish the first example of HSLBs on a CP device and show that polymer incorporation enhances bilayer resilience and thus offers important benefits toward bio-hybrid bioelectronics for sensing applications. Importantly, HSLBs outperform traditional phospholipid bilayers in stability by exhibiting strong electrical sealing after exposure to physiologically relevant enzymes that cause phospholipid hydrolysis and membrane degradation. We investigate the impact of HSLB composition on membranes and device performance and demonstrate the ability to finely adjust the lateral diffusivity of HSLBs with modest changes in block copolymer content through a large compositional range. The inclusion of the block copolymer into the bilayer does not disrupt electrical sealing on CP electrodes, an essential metric for electrochemical sensors, or the insertion of a representative transmembrane protein. This work interfacing tunable and stable HSLBs with CPs paves the way for future bioinspired sensors that combine the exciting developments from both bioelectronics and synthetic biology.

Emergence of mass spectrometry detergents for membrane proteomics.

Detergents enable the investigation of membrane proteins by mass spectrometry. Detergent designers aim to improve underlying methodologies and are confronted with the challenge to design detergents with optimal solution and gas-phase properties. Herein, we review literature related to the optimization of detergent chemistry and handling and identify an emerging research direction: the optimization of mass spectrometry detergents for individual applications in mass spectrometry-based membrane proteomics. We provide an overview about qualitative design aspects including their relevance for the optimization of detergents in bottom-up proteomics, top-down proteomics, native mass spectrometry, and Nativeomics. In addition to established design aspects, such as charge, concentration, degradability, detergent removal, and detergent exchange, it becomes apparent that detergent heterogeneity is a promising key driver for innovation. We anticipate that rationalizing the role of detergent structures in membrane proteomics will serve as an enabling step for the analysis of challenging biological systems.

IgA-dominant glomerulonephritis with DNAJB9-negative fibrillar polytypic immunoglobulin deposits in the subepithelium.

Fibrillary glomerulonephritis (FGN), a rare disease is pathologically characterized by glomerular fibril accumulation ranging from 12 to 24 nm in diameter with negative Congo red staining. Recently, the identification of DnaJ homolog subfamily B member 9 (DNAJB9) as a highly sensitive and specific marker for FGN has revolutionized diagnosis of this disease. However, few recent studies have reported DNAJB9-negative glomerulonephritis with fibrillar deposits. As such, it remains unclear whether DNAJB9-negative cases can be considered equivalent to FGN. Here, we report the case of a 70-year-old woman who developed renal impairment and nephrotic-range proteinuria. Renal biopsy and pathological examination revealed focal glomerulonephritis with fibrocellular crescents. Immunofluorescence microscopy showed IgA-dominant deposition of polytypic IgG in the glomerulus. Electron microscopy revealed hump-like subepithelial electron dense deposits with fibrils of 15-25 nm in diameter. These findings were consistent with FGN; thus, Congo red and direct fast scarlet (DFS) staining, and immunohistochemistry for DNAJB9 were performed. In addition to negative Congo red/DFS/DNAJB9 staining, laser microdissection (LMD) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) resulted negative for DNAJB9, which is a highly sensitive and specific marker for FGN. The patient's renal function further declined, prompting administration of rituximab weekly for 2 weeks, similar to the treatment for FGN. This is a unique case of IgA-dominant glomerulonephritis with DNAJB9-negative fibrillar polytypic immunoglobulin deposits in the subepithelium, unlike previous DNAJB9-negative cases. Thus, DNAJB9-negative cases diagnosed based on accurate electron microscopic evaluation must be gathered, and LMD and LC-MS/MS must be used to analyze the organized fibrillar deposits to reveal the disease entity.

Characterization of the Multiple Domains of Pex30 Involved in Subcellular Localization of the Protein and Regulation of Peroxisome Number.

Pex30 is a peroxisomal protein whose role in peroxisome biogenesis via the endoplasmic reticulum has been established. It is a 58 KDa multi-domain protein that facilitates contact site formation between various organelles. The present study aimed to investigate the role of various domains of the protein in its sub-cellular localization and regulation of peroxisome number. For this, we created six truncations of the protein (1-87, 1-250, 1-352, 88-523, 251-523 and 353-523) and tagged GFP at the C-terminus. Biochemical methods and fluorescence microscopy were used to characterize the effect of truncation on expression and localization of the protein. Quantitative analysis was performed to determine the effect of truncation on peroxisome number in these cells. Expression of the truncated variants in cells lacking PEX30 did not cause any effect on cell growth. Interestingly, variable expression and localization of the truncated variants in both peroxisome-inducing and non-inducing medium was observed. Truncated variants depicted different distribution patterns such as punctate, reticulate and cytosolic fluorescence. Interestingly, lack of the complete dysferlin domain or C-Dysf resulted in increased peroxisome number similar to as reported for cells lacking Pex30. No contribution of this domain in the reticulate distribution of the proteins was also observed. Our results show an interesting role for the various domains of Pex30 in localization and regulation of peroxisome number.

Clinical implications of Golgi protein 73 and granulocyte colony-stimulating factor and their related factors in children with bronchopneumonia.

OBJECTIVE: To investigate the clinical implications of Golgi glycoprotein 73 (GP73) and granulocyte colony-stimulating factor (G-CSF) in children with bronchopneumonia (BP). METHODS: Seventy-two children with BP (observation group) and 81 healthy children (control group) consecutively brought to the present study's hospital between June 2019 and October 2020 were enrolled. GP73 and G-CSF levels were determined to analyze their diagnostic value for pediatric BP. High-sensitivity C-reactive protein (hs-CRP) was also measured. The clinical implications of GP73 and G-CSF in pediatric BP complicated with respiratory failure and their connections with the inflammatory response were discussed. RESULTS: GP73 and G-CSF levels were remarkably higher in the observation group (p < 0.05). The sensitivity and specificity of combined detection (GP73+G-CSF) in predicting pediatric BP were 72.22% and 86.42%, respectively (p < 0.001). GP73 and G-CSF, which are closely related to X-ray classification and complications in the observation group, decreased after treatment and were positively correlated with hs-CRP (p < 0.05), especially in children complicated with respiratory failure. Regression analysis identified the independence of the course of the disease, hs-CRP, X-ray classification, GP73, and G-CSF as influencing factors of respiratory failure in children with BP (p < 0.05). CONCLUSION: GP73 and G-CSF, with elevated levels in children with BP, are strongly linked to disease progression and are independent influencing factors of respiratory failure, which may be the key to diagnosing and treating pediatric BP in the future.

Proximity labeling methods for proteomic analysis of membrane proteins.

Membrane proteins constitute the filter that controls the cellular traffic of nutrients, ions and other essential molecules, as well as the transmission of signals across the membrane. These proteins interact with other proteins in the cytosol, cytoskeleton or the extracellular side of the membrane, giving rise to complex interactomes that are distributed throughout the various lipid microdomains of the membrane plane. In this manner, complex networks of protein-protein and protein-lipid interactions are formed which regulate the most diverse biological functions, and disturbance of these networks can lead to disease. Therefore, characterization of these interactomes is a priority for current biomedical sciences. Traditionally, such studies have largely depended on solubilization/dissociation of the essential components of multiprotein complexes with detergents of various strength. However, this technique may result in the loss of certain components of such complexes, especially those whose binding is weak or transient. Moreover, protein solubilization can lead to the formation of non-native spurious interactions. As an alternative, proximity labelling (PL) techniques have been developed in recent years that can identify interactors of the protein of interest in a native cellular environment, prior to solubilization. In this article, we review the recent advances in PL and explore the new possibilities they offer for the characterization of membrane interactomes. SIGNIFICANCE: Membranes establish a series of complex protein-protein and protein-lipid interactions that are essential for cell physiology. For decades, they have been one of the central objects of study in Cell and Molecular Biology. However, knowledge of the structure of membrane proteins and their respective interactomes lags far behind that of soluble proteins, mainly due to technical difficulties in their handling and characterization caused by their insolubility. Recent research has developed various techniques to study these proteins in their native cellular environment. In this review article we address the application to membrane proteins of the so-called 'proximity labeling methods', which allows neighborhood relationships to be established between proteins in intact cells. The scarcity of alternatives for study of the components of membrane complexes make these methods especially attractive for analyzing this type of membrane associated supramolecular structures.

p63-positive prostate carcinoma-A very rare presentation.

Prostate cancer being the world's leading cause of cancer and also the second most common cancer in men is posing challenges in its diagnosis. Immunohistochemistry with markers like high molecular weight cytokeratin, p63 aid in the diagnosis. The absence of p63 and high molecular weight cytokeratin and presence of p504s in the biopsies indicate malignant lesions. Yet, there is a loophole to this too. A rare case of p63-positive prostatic adenocarcinoma in an 87-year-old patient, with immunohistochemistry results showing overexpression of p63 in the nuclei of the malignant glands. This tumor shows high molecular weight cytokeratin negativity, and p504s positivity. Prognosis of this variant of the tumor is mostly favorable. Prompt treatment will halt the progression of this tumor and prevent paraplegia. Radical prostatectomy could be avoided by treatment modalities like androgen blockade and brachytherapy, as morbidity is very high with radical prostatectomy surgery.

Identification of Novel Prognostic Signatures for Clear Cell Renal Cell Carcinoma Based on ceRNA Network Construction and Immune Infiltration Analysis.

Objective: Clear cell renal cell carcinoma (ccRCC) carries significant morbidity and mortality globally and is often resistant to conventional radiotherapy and chemotherapy. Immune checkpoint blockade (ICB) has received attention in ccRCC patients as a promising anticancer treatment. Furthermore, competitive endogenous RNA (ceRNA) networks are crucial for the occurrence and progression of various tumors. This study was aimed at identifying reliable prognostic signatures and exploring potential mechanisms between ceRNA regulation and immune cell infiltration in ccRCC patients. Methods and Results: Gene expression profiling and clinical information of ccRCC samples were obtained from The Cancer Genome Atlas (TCGA) database. Through comprehensive bioinformatic analyses, differentially expressed mRNAs (DEmRNAs; n = 131), lncRNAs (DElncRNAs; n = 12), and miRNAs (DEmiRNAs; n = 25) were identified to establish ceRNA networks. The CIBERSORT algorithm was applied to calculate the proportion of 22 types of tumor-infiltrating immune cells (TIICs) in ccRCC tissues. Subsequently, univariate Cox, Lasso, and multivariate Cox regression analyses were employed to construct ceRNA-related and TIIC-related prognostic signatures. In addition, we explored the relationship between the crucial genes and TIICs via coexpression analysis, which revealed that the interactions between MALAT1, miR-1271-5p, KIAA1324, and follicular helper T cells might be closely correlated with the progression of ccRCC. Ultimately, we preliminarily validated that the potential MALAT1/miR-1271-5p/KIAA1324 axis was consistent with the ceRNA theory by qRT-PCR in the ccRCC cell lines. Conclusion: On the basis of the ceRNA networks and TIICs, we constructed two prognostic signatures with excellent predictive value and explored possible molecular regulatory mechanisms, which might contribute to the improvement of prognosis and individualized treatment for ccRCC patients.

Protein expression of transmembrane protease serine 4 in the gastrointestinal tract and in healthy, cancer, and SARS­CoV­2 infected lungs.

In addition to the angiotensin­converting enzyme 2 (ACE2), a number of host cell entry mediators have been identified for severe acute respiratory syndrome coronavirus­2 (SARS­CoV­2), including transmembrane protease serine 4 (TMPRSS4). The authors have recently demonstrated the upregulation of TMPRSS4 in 11 different cancers, as well as its specific expression within the central nervous system using in silico tools. The present study aimed to expand the initial observations and, using immunohistochemistry, TMPRSS4 protein expression in the gastrointestinal (GI) tract and lungs was further mapped. Immunohistochemistry was performed on tissue arrays and lung tissues of patients with non­small cell lung cancer with concurrent coronavirus disease 2019 (COVID­19) infection using TMPRSS4 antibody. The results revealed that TMPRSS4 was abundantly expressed in the oesophagus, stomach, small intestine, jejunum, ileum, colon, liver and pancreas. Moreover, the extensive TMPRSS4 protein expression in the lungs of a deceased patient with COVID­19 with chronic obstructive pulmonary disease and bronchial carcinoma, as well in the adjacent normal tissue, was demonstrated for the first time, at least to the best of our knowledge. On the whole, the immunohistochemistry data of the present study suggest that TMPRSS4 may be implicated in the broader (pulmonary and extra­pulmonary) COVID­19 symptomatology; thus, it may be responsible for the tropism of this coronavirus both in the GI tract and lungs.

A case of juvenile-onset fibrillary glomerulonephritis diagnosed by mass spectrometry and immunohistochemistry of DNAJB9.

Fibrillary glomerulonephritis (FGN) is a rare glomerular disease. FGN is characterized by the deposition of randomly arranged, nonbranching microfibrils in the mesangium and glomerular basement membrane. The discovery of DNAJ homolog subfamily B member 9 (DNAJB9) in 2017 was a breakthrough, and DNAJB9 has been proven to be extremely useful for the definitive diagnosis of FGN. While FGN often occurs in middle-aged individuals, this case was diagnosed at a relatively young age of 17. We performed renal biopsy, and light microscopic study revealed mesangial proliferation with expansion and subepithelial deposits. Electron microscopic study showed glomerular deposition of randomly oriented nonbranching fibrils with a mean of 20 nm. However, direct first scarlet stain for amyloidosis was weakly positive. Therefore, we confirmed the diagnosis of FGN and eliminated the presence of amyloidosis with mass spectrometry. This is the first case in Japan in which the complication of amyloidosis was ruled out with mass spectrometry and FGN was diagnosed using immunostaining and mass spectrometry of DNAJB9. We began treatment with cyclosporine A. One and a half years after the start of the treatment, kidney function continues to be normal.

Matrix remodeling-associated protein 8 is a marker of a subset of cancer-associated fibroblasts in pancreatic cancer.

Cancer-associated fibroblasts (CAFs), a compartment of the tumor microenvironment, were previously thought to be a uniform cell population that promotes cancer progression. However, recent studies have shown that CAFs are heterogeneous and that there are at least two types of CAFs, that is, cancer-promoting and -restraining CAFs. We previously identified Meflin as a candidate marker of cancer-restraining CAFs (rCAFs) in pancreatic ductal adenocarcinoma (PDAC). The precise nature of rCAFs, however, has remained elusive owing to a lack of understanding of their comprehensive gene signatures. Here, we screened genes whose expression correlated with Meflin in single-cell transcriptomic analyses of human cancers. Among the identified genes, we identified matrix remodeling-associated protein 8 (MXRA8), which encodes a type I transmembrane protein with unknown molecular function. Analysis of MXRA8 expression in human PDAC samples showed that MXRA8 was differentially co-expressed with other CAF markers. Moreover, in patients with PDAC or syngeneic tumors developed in MXRA8-knockout mice, MXRA8 expression did not affect the roles of CAFs in cancer progression, and the biological importance of MXRA8+ CAFs is still unclear. Overall, we identified MXRA8 as a new CAF marker; further studies are needed to determine the relevance of this marker.