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Background & Aims RING finger protein 43 (RNF43) is a tumor suppressor that frequently is mutated in gastric tumors. The link between RNF43 and modulation of Wingless-related integration site (WNT) signaling has not been shown clearly in the stomach. Because mutations in RNF43 are highly enriched in microsatellite-unstable gastric tumors, which show defects in DNA damage response (DDR), we investigated whether RNF43 is involved in DDR in the stomach. Methods DDR activation and cell viability upon γ-radiation was analyzed in gastric cells where expression of RNF43 was depleted. Response to chemotherapeutic agents 5-fluorouracil and cisplatin was analyzed in gastric cancer cell lines and xenograft tumors. In addition, involvement of RNF43 in DDR activation was analyzed upon Helicobacter pylori infection in wild-type and Rnf43ΔEx8 mice. Furthermore, a cohort of human gastric biopsy specimens was analyzed for RNF43 expression and mutation status as well as for activation of DDR. Results RNF43 depletion conferred resistance to γ-radiation and chemotherapy by dampening the activation of DDR, thereby preventing apoptosis in gastric cells. Upon Helicobacter pylori infection, RNF43 loss of function reduced activation of DDR and apoptosis. Furthermore, RNF43 expression correlated with DDR activation in human gastric biopsy specimens, and RNF43 mutations found in gastric tumors conferred resistance to DNA damage. When exploring the molecular mechanisms behind these findings, a direct interaction between RNF43 and phosphorylated H2A histone family member X (γH2AX) was observed. Conclusions We identified a novel function for RNF43 in the stomach as a regulator of DDR. Loss of RNF43 function in gastric cells confers resistance to DNA damage-inducing radiotherapy and chemotherapy, suggesting RNF43 as a possible biomarker for therapy selection., Graphical abstract

International journal of molecular sciences 23(23), 15126 (2022). doi:10.3390/ijms232315126 special issue: "Special Issue "Neutrophil Extracellular Traps (NETs) in Immunity and Diseases" / Special Issue Editor: Małgorzata Wachowska, Guest Editor", Published by Molecular Diversity Preservation International, Basel

Toll-like receptor TLR5 recognizes a conserved domain, termed D1, that is present in flagellins of several pathogenic bacteria but not in Helicobacter pylori. Highly virulent H. pylori strains possess a type IV secretion system (T4SS) for delivery of virulence factors into gastric epithelial cells. Here, we show that one of the H. pylori T4SS components, protein CagL, can act as a flagellin-independent TLR5 activator. CagL contains a D1-like motif that mediates adherence to TLR5+ epithelial cells, TLR5 activation, and downstream signaling in vitro. TLR5 expression is associated with H. pylori infection and gastric lesions in human biopsies. Using Tlr5-knockout and wild-type mice, we show that TLR5 is important for efficient control of H. pylori infection. Our results indicate that CagL, by activating TLR5, may modulate immune responses to H. pylori., Toll-like receptor TLR5 recognizes a domain, D1, that is present in flagellins of several pathogenic bacteria but not in Helicobacter pylori. Here, the authors show that TLR5 can be activated independently of flagellin by a component of the H. pylori type IV secretion system that contains a D1-like motif.

Background Coronavirus induced disease 2019 (COVID-19) can be complicated by severe organ damage leading to dysfunction of the lungs and other organs. The processes that trigger organ damage in COVID-19 are incompletely understood. Methods Samples were donated from hospitalized patients. Sera, plasma, and autopsy-derived tissue sections were examined employing flow cytometry, enzyme-linked immunosorbent assays, and immunohistochemistry. Patient findings Here, we show that severe COVID-19 is characterized by a highly pronounced formation of neutrophil extracellular traps (NETs) inside the micro-vessels. Intravascular aggregation of NETs leads to rapid occlusion of the affected vessels, disturbed microcirculation, and organ damage. In severe COVID-19, neutrophil granulocytes are strongly activated and adopt a so-called low-density phenotype, prone to spontaneously form NETs. In accordance, markers indicating NET turnover are consistently increased in COVID-19 and linked to disease severity. Histopathology of the lungs and other organs from COVID-19 patients showed congestions of numerous micro-vessels by aggregated NETs associated with endothelial damage. Interpretation These data suggest that organ dysfunction in severe COVID-19 is associated with excessive NET formation and vascular damage. Funding Deutsche Forschungsgemeinschaft (DFG), EU, Volkswagen-Stiftung

Diagnosing low grade intraepithelial neoplasia (LGIN) in patients with ulcerative colitis (UC) is difficult. Distinguishing between sporadic adenoma (SA) and UC associated LGIN is even more challenging but has clinical impact. We aimed to examine, if the morphological distinction between both entities is reliably possible, how it influences patient's outcome and the role of the endoscopist in this decision with respect to current endoscopy classification schemes.Seven pathologists retrospectively reevaluated 425 cases of LGIN in UC patients, diagnosed between 2009 and 2017 with preceding expert consensus and follow up in two separate readings, based on published morphological differentiation criteria. In the first evaluation, the observers were blinded to any clinical data. In the second evaluation, they knew patients' age as well as endoscopic features. They also rated their subjective diagnostic certainty.Diagnostic correctness improved significantly in the second assessment as did the pathologists' confidence in their diagnoses (p0.001 - p = 0.019). Knowledge of clinical and endoscopical data led to a higher percentage of SA (71.8% vs. 85.6%). UC associated LGIN showed significant earlier LGIN relapse as well as more high grade intraepithelial neoplasia and carcinoma during follow up (p0.001, p0.001, p = 0.005).Distinction between SA and UC associated LGIN is important as it has an impact on patients' follow up and treatment. Morphological distinction remains difficult with moderate interobserver variability. Adequate clinical information significantly improves pathologists' diagnoses as well as their confidence in their diagnoses.

Helicobacter pylori infection of the human stomach is associated with chronic gastritis, peptic ulcer disease or gastric carcinoma, and thus a high burden for the public health systems worldwide. Fortunately, only a small subfraction of up to 15-20% of infected individuals will develop serious complications. Unfortunately, it is not always known upfront, who will be affected by serious diesease outcome. For risk stratifications, it is therefore necessary to establish a common terminology and grading system, that can be applied worldwide to compare population data. The updated Sydney System for classification of gastritis with its semi-quantitative analogue scale is the system, that is currently used worldwide. Additionally, pathologists should always try to classify the etiology of the inflammatory infiltrates in the stomach to instruct the clinicians for choosing a proper treatment regime. Risk factors such as intestinal metaplasia, atrophy and scoring systems to classify these risk factors into a clinical context such as OLGA and OLGIM are discussed. Also, special forms of gastritis like lymphocytic gastritis, autoimmune gastritis and peptic ulcer disease are explained and discussed e.g. how to diagnose and how to treat. Extra-gastric sequelae of H. pylori infections inside and outside the stomach are shown in this chapter as well. Important host and bacterial risk factors such as pathogenicity islands are dicussed to draw a complete landscape around a H. pylori infection, that still can be diagnosed in patients. However, it needs to be noted that some countries have almost no H. pylori infection anymore, while others have still a very high frequency of infections with or without serious complications. The understanding and application of risk assessements may help to save money and quality of life. Extra-gastric H. pylori infections are rarely reported in the literature until today. The pathogenitiy is still under debate, but especially in the bile ducts and gallbladder, several pathological conditions may be also based on H. pylori infection, and will be also discussed.

Helicobacter pylori infection of the human stomach is associated with chronic gastritis, peptic ulcer disease or gastric carcinoma, and thus a high burden for the public health systems worldwide. Fortunately, only a small subfraction of up to 15–20% of infected individuals will develop serious complications. Unfortunately, it is not always known upfront, who will be affected by serious diesease outcome. For risk stratifications, it is therefore necessary to establish a common terminology and grading system, that can be applied worldwide to compare population data. The updated Sydney System for classification of gastritis with its semi-quantitative analogue scale is the system, that is currently used worldwide. Additionally, pathologists should always try to classify the etiology of the inflammatory infiltrates in the stomach to instruct the clinicians for choosing a proper treatment regime. Risk factors such as intestinal metaplasia, atrophy and scoring systems to classify these risk factors into a clinical context such as OLGA and OLGIM are discussed. Also, special forms of gastritis like lymphocytic gastritis, autoimmune gastritis and peptic ulcer disease are explained and discussed e.g. how to diagnose and how to treat. Extra-gastric sequelae of H. pylori infections inside and outside the stomach are shown in this chapter as well. Important host and bacterial risk factors such as pathogenicity islands are dicussed to draw a complete landscape around a H. pylori infection, that still can be diagnosed in patients. However, it needs to be noted that some countries have almost no H. pylori infection anymore, while others have still a very high frequency of infections with or without serious complications. The understanding and application of risk assessements may help to save money and quality of life. Extra-gastric H. pylori infections are rarely reported in the literature until today. The pathogenitiy is still under debate, but especially in the bile ducts and gallbladder, several pathological conditions may be also based on H. pylori infection, and will be also discussed.

We have designed and synthesized a series of 14 hybrid molecules out of the cholinesterase (ChE) inhibitor tacrine and a benzimidazole-based human cannabinoid receptor subtype 2 (hCB(2)R) agonist and investigated them in vitro and in vivo. The compounds are potent ChE inhibitors, and for the most promising hybrids, the mechanism of human acetylcholinesterase (hAChE) inhibition as well as their ability to interfere with AChE-induced aggregation of β-amyloid (Aβ), and Aβ self-aggregation was assessed. All hybrids were evaluated for affinity and selectivity for hCB(1)R and hCB(2)R To ensure that the hybrids retained their agonist character, the expression of cAMP-regulated genes was quantified, and potency and efficacy were determined. Additionally, the effects of the hybrids on microglia activation and neuroprotection on HT-22 cells were investigated. The most promising in vitro hybrids showed pronounced neuroprotection in an Alzheimer’s mouse model at low dosage (0.1 mg/kg, i.p.), lacking hepatotoxicity even at high dose (3 mg/kg, i.p.).

After Babylonian confusion over the histological classification of gastritis, the Sydney system brought standardization and reproducibility to the diagnostic field of gastric biopsies. Even some shortcomings do not reduce the importance of the Sydney system for classification of gastritis. Essentially gastritis is a purely histological diagnosis. Herein we describe further diagnostic criteria for the diagnosis of gastritis and show the time-dependent changes in frequencies of various types of gastritis over more than 25 years at the Institute of Pathology in Bayreuth. Pathologists should be encouraged to address the etiology of inflammatory infiltrates to enhance the clinical value of a histological diagnosis on gastric biopsies.

Objective:Tumours with neuroendocrine differentiation frequently express chromogranin A, synaptophysin and somatostatin receptors. The role of neuroendocrine differentiation in head and neck squamous cell carcinoma is not yet clear.Method:The presence of chromogranin A, synaptophysin and somatostatin receptors was studied immunohistochemically in 78 head and neck squamous cell carcinoma specimens.Results:Sparse chromogranin A expression was found in 41 per cent, associated with high chromogranin A messenger RNA expression and the presence of dense core granules. Low synaptophysin expression was found in 18 per cent. The highest staining scores were found for somatostatin receptor 5 (82 per cent), followed by somatostatin receptor 1 (69 per cent) and somatostatin receptor 2 (54 per cent), whereas somatostatin receptors 3 and 4 expression was low. Expression was not correlated with tumour stage or survival.Conclusion:Cells with neuroendocrine differentiation are sparsely scattered in some head and neck squamous cell carcinomas. Their pathophysiological role is elusive. In contrast, somatostatin receptor and particularly somatostatin receptor 5 expression is frequent in head and neck squamous cell carcinoma. Somatostatin receptor expression is not considered to indicate neuroendocrine differentiation in head and neck squamous cell carcinoma.

Fibroblasts (Fibs) contribution to neoplastic progression, tumor growth, angiogenesis, and metastasis has been recently reported by several research groups. In this study it was investigated if fibroblasts are the source of brain-derived neurotrophic factor (BDNF), which plays a crucial role in the progression of oral squamous cell carcinoma. In a novel in vitro system oral Fibs were cultured with SCC-25 lingual squamous cell carcinoma cells for 7 days. Factors related with this interaction were investigated by quantitative PCR and western blot. In the co-culture, fibroblasts were converted to carcinoma-associated fibroblasts (CAFs), which in return initiated epithelial–mesenchymal transition (EMT) of SCC-25 cells. The induced CAFs produced increased levels of BDNF, which interacted with the increased-expressed neurothrophin receptor B (TrkB) on EMT-converted SCC-25 cells. Possible regulatory factors of BDNF expression (tumor necrosis factor-α and interleukin-1-β) were detected both in CAFs and EMT-tumor cells. In CAFs: IL-1β-, in SCC-25 cells TNF-α-gene-expression was significantly increased in co-culture conditions. Activated fibroblasts (CAFs) and mesenchymal transitioned tumor cells might use the BDNF-TrkB axis and its regulation to harmonize their interaction in the process of tumor progression.

Hypothesis Growth hormones have beneficial effects on increasing height in adults with Turner syndrome (TS) and may also affect auditory function. Background Turner syndrome is the most common sex-linked chromosomal abnormality in female conceptions. Epidermal growth factor and its receptor (EGFR) affect differentiation, proliferation, and migration of epithelial cells and function as survival factors. The expression of EGFR is found in the developing and juvenile inner ear of experimental animals but is absent in adults. Methods To determine whether EGFR plays a role in TS, its expression was analyzed in the cochlea of healthy fetus and fetus with TS and in healthy adults. Results In healthy fetuses, EGFR protein expression was localized to the inner and outer hair cells and the Reissner membrane. The fetuses with TS on the 13th gestational week (GW) showed a similar pattern of immunoreactivity as the normal 16th and 20th GW cochlea. By the 23rd GW, EGFR immunoreactivity was not detectable in the TS hair cells or the Reissner membrane, and less intensive staining was found in the surrounding fibrocytes of the spiral ganglion. Conclusion This is the first demonstration of EGFR immunoreactivity in the human cochlea and illustrates how EGFR expression is altered during development in TS. These findings indicate the importance of growth hormone receptors for inner ear development in humans.

Summary Hereditary complete C4 deficiency (C4def) is a very rare condition that predisposes to immune complex disease and end-stage renal failure. Whether such patients should undergo renal transplantation is debated. The clinical outcome of five transplantations in three C4def patients is described. The first patient lost one allograft after 6 years because of chronic allograft rejection. Back on dialysis, he suffered from meningitis caused by Neisseria menigitidis and Aspergillus. One year after a second transplantation under alemtuzumab induction, he developed fulminant Kaposi’s sarcoma and died. His sister is now 6 years post-transplantation without complications. The third patient lost his first graft after 3 years because of chronic allograft nephropathy and recurrence of glomerulonephritis. He has now been living with a second graft for over 9 years. He suffered from pneumonia, a generalized varicella infection and Hemophilis parainfluenzae bronchitis. Patients with complete C4def are at increased risk for infection after kidney transplantation. Under certain precautions and with judicious use of immunosuppression, good long-term results are achievable.

In this study different malformations of the cochlea could be demonstrated. Nevertheless, we could not delineate a distinct malformation of the inner ear, that can be linked to a neural tube defect. Neural tube defects are a frequent and heterogeneous group of malformations, ranging from the survivable spina bifida to fatal anencephaly. In multiple animal models an involvement of the vestibulocochlear system has been demonstrated. In this article human fetal temporal bones of neural tube defects were analysed in a multimodular work-up. The morphologic study was performed with light microscopy, transmission electron microscopy and synchrotron radiation-based microcomputed tomography. Immunohistochemistry for different neuronal markers such as peripherin, beta-III-tubulin and vimentin helped to evaluate ontogenetic tissue development. Eight fetal temporal bones with neural tube defects and five control temporal bones were included into the morphologic study. The morphologic results of the neural tube defect temporal bones showed six regularly developed cochleas and two with only a single cochlear turn. Three of the neural tube defect temporal bones were further examined with immunohistochemical analysis. No differences in the staining pattern for peripherin, beta-III-tubulin and vimentin were detected.

BACKGROUND& AIMS: Lymphocytic esophagitis is ill defined and lymphocyte-rich inflammatory lesions of esophageal squamous epithelium are believed to show non-specific histological changes. This study was performed to investigate the histological and molecular criteria in lymphocytic esophagitis. METHODS: We collected 33 biopsies from 28 patients with lymphocytic esophagitis, which were diagnosed between 2010 and 2012 (17 men and 11 women, mean age = 60,8 and 61,1 years, respectively). Grading was performed by the number of lymphocytes (50/HPF cut off value) and according to Purdy et al. The inflammatory infiltrate was characterized by using the XTRACT robot (STRATIFYER, Cologne, Germany), quantitative real-time-PCR (FOXP3, CXCL9, CXCL13, IGHM, CD3, CD4 and CD8) and immunohistochemistry (CD4, CD8, TIA, GranzymB and Caspase 3). Spearman rank test was used to calculate the correlation between these parameters. RESULTS: Apoptoses could be detected in 83% of severe cases (15/18) and in 47% of mild cases (7/15) without differences between the two grading systems. High levels of cytotoxic T-cells were present in 80% of severe cases with apoptoses (12/15) and in 5 cases without visible apoptoses in H&E stain, mostly mild cases. The overall number of lymphocytes correlated with the number of apoptoses (p = 0,002) and the number of CD8 positive lymphocytes (p = 0,005). Cases with apoptoses showed high levels of CD8 positive lymphocytes. A positive correlation between spongiosis and levels of cytotoxic T-cells could be demonstrated (p = ,0,0001). Evaluation of clinical data revealed a correlation between high levels of cytotoxic lymphocytes and a wavey mucosal surface (p = 0,025), reddening of esophageal epithelium (p = 0,017) and a stipplelike exsudate (p = 0,015). Within the group with apoptoses a strong correlation between cytotoxic T-cells (= CD8) and regulatory T-cells (= FOXP3) (p = ,0,0001) was found. Expression of CXCL9 correlated with the expression of CD4 and CD8 (p = 0,05 and p = 0,0042, respectively). CONCLUSIONS: Our data suggest that the term "lymphocytic esophagitis" should only be used as an umbrella-term in several diseases and therefore we suggest "lymphocyte-rich esophagitis" to be used for further subdivisions. Two different subgroups of this lymphocyte-rich variant of chronic esophagitis can be distinguished: Lymphocyte-rich esophagitis with apoptosis (LEA) and lymphocytic esophagitis with no apoptosis (LENA). Apoptotic keratinocytes are usefull in grading LEA, as they can be found in large numbers predominantely in severe forms. The number and localization of apoptoses distinguishes esophageal Crohn's disease and lichen planus from LEA. Further studies should focus on the question, whether our findings are a hint to an autoimmunologic mechanism or not.