Lamina Propria Research Articles

ORF virus causes tumor-promoting inflammation in sheep and goats.

ORF virus (ORFV) causes contagious ecthyma ("ORF"), a disease of sheep and goats characterized by lesions ranging from vesicles and pustules to atypical papilloma-like and angiomatous lesions in the skin and mucosae. The authors investigated the molecular factors leading to the ORF-associated atypical tumor-like changes. Fifteen lambs, 15 kids, and an adult ram clinically affected by natural ORFV infection were enrolled in the study and examined by several methods. ORFV was detected by viral culture or real-time polymerase chain reaction (RT-PCR) in the lesioned tissues and in the blood of the clinically affected sheep and goats. Surprisingly, ORFV was also detected in the blood of healthy goats from an affected herd. Microscopically, they found a pseudo-papillomatous proliferation of the epithelium, while the dermis and lamina propria were expanded by a proliferating neovascular component that highly expressed the viral vascular endothelial growth factor (vVEGF) and its host receptor vascular endothelial growth factor receptor 2 (VEGFR2). Immunohistochemistry, immunofluorescence, and in situ hybridization for mRNA showed that epidermal growth factor receptor (EGFR) was expressed in the fibrovascular component, in the infiltrating CD163+ macrophages, and in the basal stratum of the epidermis. Confocal immunofluorescence microscopy demonstrated that CD163+ macrophages were associated with VEGF and VEGFR2. Finally, they found by quantitative RT-PCR the overexpression of the interleukin-6 and VEGFR2 genes in the lesioned tissues. These findings suggest that ORFV activates an inflammatory reaction characterized by CD163+ macrophages expressing EGFR and VEGFR2, which might play an oncogenic role through synergistic action with vVEGF signaling.

Blimp-1 and c-Maf regulate immune gene networks to protect against distinct pathways of pathobiont-induced colitis.

Intestinal immune responses to microbes are controlled by the cytokine IL-10 to avoid immune pathology. Here, we use single-cell RNA sequencing of colon lamina propria leukocytes (LPLs) along with RNA-seq and ATAC-seq of purified CD4+ T cells to show that the transcription factors Blimp-1 (encoded by Prdm1) and c-Maf co-dominantly regulate Il10 while negatively regulating proinflammatory cytokines in effector T cells. Double-deficient Prdm1fl/flMaffl/flCd4Cre mice infected with Helicobacter hepaticus developed severe colitis with an increase in TH1/NK/ILC1 effector genes in LPLs, while Prdm1fl/flCd4Cre and Maffl/flCd4Cre mice exhibited moderate pathology and a less-marked type 1 effector response. LPLs from infected Maffl/flCd4Cre mice had increased type 17 responses with increased Il17a and Il22 expression and an increase in granulocytes and myeloid cell numbers, resulting in increased T cell-myeloid-neutrophil interactions. Genes over-expressed in human inflammatory bowel disease showed differential expression in LPLs from infected mice in the absence of Prdm1 or Maf, revealing potential mechanisms of human disease.

Establishment of an intragastric surgical model using C57BL/6 mice to study the vaccine efficacy of OMV-based immunogens against Helicobacter pylori.

Chronic gastritis is one of the major symptoms of gastro-duodenal disorders typically induced by Helicobacter pylori (H. pylori). To date, no suitable model is available to study pathophysiology and therapeutic measures accurately. Here, we have presented a successful surgical infection model of H. pylori-induced gastritis in C57BL/6 mice that resembles features similar to human infection. The proposed model does not require any preparatory treatment other than surgical intervention. C57BL/6 mice were injected with wild-type SS1 (Sydney strain 1, reference strain) directly into the stomach. Seven days post infection, infected animals showed alterations in cytokine responses along with inflammatory cell infiltration in the lamina propria, depicting a prominent inflammatory response due to infection. To understand the immunogenicity and protective efficacy, the mice were immunized with outer membrane vesicles (OMVs) isolated from an indigenous strain with putative virulence factors of H. pylori [A61C (1), cag+/vacA s1m1]. In contrast to the non-immunized cohort, the OMV-immunized cohort showed a gradual increase in serum immunoglobulin(s) levels on the 35th day after the first immunization. This conferred protective immunity against subsequent challenge with the reference strain (SS1). Direct inoculation of H. pylori into the stomach influenced infection in a short time and, more importantly, in a dose-dependent manner, indicating the usefulness of the developed model for pathophysiology, therapeutic and prophylactic studies.

Identification of neutrophils and eosinophils in upper airway mucosa with immunofluorescence multiplex image cytometry.

Characterization of inflammation in chronic rhinosinusitis with (CRSwNP) and without nasal polyps (CRSsNP) is an ongoing research process. To overcome limitations of current cytologic techniques, we investigated whether immunofluorescence multiplex image cytometry could quantify intact neutrophils, eosinophils, and other immune cells in solid upper airway mucosa. We used a four-channel immunofluorescence-microscopy technique for the simultaneous detection of the leukocyte marker CD45, the neutrophil marker myeloperoxidase, two eosinophil markers, i.e., major basic protein and eosinophil peroxidase, and DAPI (4',6-diamidin-2-phenylindole), in formalin-fixed paraffin-embedded upper airway tissue samples of patients with CRSwNP and CRSsNP, as well as of patients free of CRS with inferior turbinate hypertrophy (controls). Image acquisition and analysis were performed with TissueFAXS and StrataQuest (TissueGnostics, Vienna, Austria), respectively. Positive and negative immunostaining were differentiated with a specific fluorescence signal/background signal ratio. Isotype controls were used as negative controls. In six controls, nine patients with CRSsNP, and 11 patients with CRSwNP, the median area scanned and median cell count per patient were 14.2mm2 and 34,356, respectively. In CRSwNP, the number of eosinophils was three times higher (23%) than that of neutrophils (7%). Three times more immune cells were encountered in CRSwNP (33%) compared to CRSsNP (11%). In controls, inflammation was balanced between the epithelial layer and lamina propria, in contrast to CRS (three times more pronounced inflammation in the lamina propria). The quantification of intact neutrophils, eosinophils, and other immune cells in solid tissue with undisrupted architecture seems feasible with immunofluorescence multiplex image cytometry.

A review on Sarvasara Mukharoga in Ayurveda w.s.r. Oral Submucous Fibrosis

Introduction: An inflammatory response juxta epithelial, fibroelastic changes in the lamina propria, epithelial atrophy, trismus, and recurrent glossitis attacks are the hallmarks of the subtle chronic disease known as oral submucous fibrosis (OSMF). It is one of the most common premalignant disorders in India is. OSMF is gaining popularity these days. A study on OSMF and its management according to Ayurveda values is more in the scientific field because Ayurveda’s main focus is on maintaining and promoting health care. In this conceptual paper, OSMF will be highlighted, evaluated, elaborated on, and discussed. Aim and Objectives: The aim of the study is to gain a better understanding of OSMF and its relationship to Sarvasara Mukharoga in Ayurveda. Materials and Methods: The contents of this article are based on modern medicine textbooks as well as Ayurvedic Samhitas. Clinical research published in both indexed and non-indexed journals was also consulted to obtain relevant content. Results: According to the Lakshanas described by Brihatrayi, OSMF can be equated symptomatically with the Sarvasara Mukharoga. In the event of OSMF, the management of Sarvasara Mukharoga should be applied. Conclusion: On the basis of signs and symptoms, OSMF can be correlated with Sarvasara Mukharoga. Early treatment of OSMF using Ayurvedic medications and the cessation of habits helps in the easy cure of the disease.

Differential expression of neuropeptide F in the digestive organs of female freshwater prawn, Macrobrachium rosenbergii, during the ovarian cycle.

Neuropeptide F is a key hormone that controls feeding in invertebrates, including decapod crustaceans. We investigated the differential expression of Macrobrachium rosenbergii neuropeptide F (MrNPF) in the digestive organs of female prawns, M. rosenbergii, during the ovarian cycle. By using RT-qPCR, the expression of MrNPF mRNA in the esophagus (ESO), cardia (CD), and pylorus (PY) of the foregut (FG) gradually increased from stage II and peaked at stage III. In the midgut (MG), hindgut (HG), and hepatopancreas (HP), MrNPF mRNA increased from stage I, reaching a maximal level at stage II, and declined by about half at stages III and IV (P < 0.05). In the ESO, CD, and PY, strong MrNPF-immunoreactivities were seen in the epithelium, muscle, and lamina propria. Intense MrNPF-ir was found in the MG cells and the muscular layer. In the HG, MrNPF-ir was detected in the epithelium of the villi and gland regions, while MrNPF-ir was also more intense in the F-, R-, and B-cells in the HP. However, we found little colocalization between the MrNPF and PGP9.5/ChAT in digestive tissues, implying that most of the positive cells might not be neurons but could be digestive tract-associated endocrine cells that produce and secrete MrNPF to control digestive organ functions in feeding and utilizing feed. Taken together, our first findings indicated that MrNPF was differentially expressed in digestive organs in correlation with the ovarian cycle, suggesting an important link between MrNPF, the physiology of various digestive organs in feeding, and possibly ovarian maturation in female M. rosenbergii.

Histological changes of the esophageal mucosa in patients with achalasia

Background. Morphological changes in the esophageal mucosa (EM) depend on the type of esophageal achalasia (ЕA). In types I and II of ЕA, morphological changes are subtle, but in types III and IV, dystrophic changes are clearly visible. ЕA can transform from one type to another. This mainly concerns the pathological transition from inflammation to fibrosis. The purpose is to investigate the histological and morphometric differences of the EM depending on the type of ЕA. Materials and methods. We studied 51 biopsy specimens of the EM in patients with ЕA. The distribution of biopsies for histo-morphometric characterisation of changes in the EM in ЕA was used according to the radiological classification of the Japanese Society of the Esophagus (2017). To study the general histological structure of the EM, biopsy sections of 5–7 μm thickness were cut using a rotary microtome RM60-ECA, stained with hematoxylin-­eosin according to the standard method, and staining for fibrosis was performed according to the Mallory-Slinchenko method. For morphometric study, the sections were photographed using a light microscope XSZ-21 (Ukraine) and measured with the Image J.45S software (USA). Results. In histological examination, the development of structural changes in the EM on the background of ЕA in 24.1 % of cases of type III was accompanied by migration of eosinophils into the lamina propria, and in 100 % — by epithelial atrophy. Dense lymphoplasmacytic infiltration was noted in all patients with types III and IV of ЕA, as well as in 93.3 % of patients with type II, whereas in type I of ЕA, this sign was diagnosed in 20 % of cases (p = 0.005 and p = 0.0001 compared with types II and III of ЕA). Ballooning degeneration was detected in 20 % of patients with EA type II, in 57.1 % with type III and in 100 % of patients with type IV. Fibrosis in the EM was observed only in ЕA type IV. Thickening of the basal layer of the squamous epithelium of the EM was noted in all patients with type IV ЕA and 86.2 % with type III ЕA, while in types I and II, it was observed in only 20 and 13.3 % of cases, respectively (p = 0.007 and p &lt; 0.001 compared with type III ЕA). Expansion of the intercellular space was detected in 100 % of patients with types II, III and IV, and in only 40 % with type I (p = 0.009 and p = 0.0017 compared with types II and III ЕA). According to the morphometric study, the squamous epithelial height was largest in type IV ЕA — 343.40 μm versus 325.45 μm in type I ЕA (p = 0.028). The height of the basal layer was the lo­west in type II ЕA (about 40 µm) and the largest in type IV ЕA (about 50 µm) (p = 0.074). An increase in the intercellular space by 41.6 % was noted with the progression of ЕA (p = 0.001). The height of the papillae is the lowest in type II ЕA — 125.05 μm and the largest in type IV ЕA — 134.70 μm (p = 0.006). Conclusions. All types of AE are accompanied by esophagitis, histological examination of which revealed an increase in the height of the basal layer, papillae height, expansion of the intercellular space, balloon degeneration, fibrous changes in the lamina propria, lymphoplasmacytic and eosinophilic infiltration of the EM that progress depending on the type of ЕA. Morphometric studies of the EM in terms of increasing the height of the epithelium (p = 0.028), the height of the papillae (p = 0.006), the height of the basal layer (p = 0.074), and the expansion of the intercellular space (p = 0.001) allow to detect the type of ЕA.

Intestinal Stromal Cells in the Turmoil of Inflammation and Defective Connective Tissue Remodeling in Inflammatory Bowel Disease.

In steady state, intestinal subepithelial myofibroblasts form a thin layer below the basement membrane. Unlike the rest of the stromal cells in the lamina propria, they express tensile proteins, guide epithelial regeneration, and sense luminal microbiota. Upon inflammation in inflammatory bowel disease (IBD), they express activation markers, accept trophic signaling by infiltrating neutrophils and macrophages, and are activated by cytokines from helper T cells to produce a narrow spectrum of cytokines and a wider spectrum of chemokines, attract cells of innate and adaptive immunity, orchestrate inflammatory responses, and qualitatively and quantitatively modify the extracellular matrix. Thus, beyond being structural tissue components, they assume active roles in the pathogenesis of complicated IBD. Discrimination between myofibroblasts and fibroblasts may be an oversimplification in light of single-cell sequencing data unveiling the complexity of multiple phenotypes of stromal cells with distinct roles and plasticity. Spatial transcriptomics revealed distinct phenotypes by histologic localization and, more intriguingly, the assembly of mucosal neighborhoods that support spatially distinct functions. Current IBD treatments target inflammation but fail in fibrostenotic or fistulizing disease. Baseline and recent findings on stromal cells, molecules, and pathways involved in disrupted extracellular matrix homeostasis are reviewed to provide relevant pharmacologic targets.

Immune microniches shape intestinal Treg function.

The intestinal immune system is highly adapted to maintaining tolerance to the commensal microbiota and self-antigens while defending against invading pathogens1,2. Recognizing how the diverse network of local cells establish homeostasis and maintains it in the complex immune environment of the gut is critical to understanding how tolerance can be re-established following dysfunction, such as in inflammatory disorders. Although cell and molecular interactions that control T regulatory (Treg) cell development and function have been identified3,4, less is known about the cellular neighbourhoods and spatial compartmentalization that shapes microorganism-reactive Treg cell function. Here we used in vivo live imaging, photo-activation-guided single-cell RNA sequencing5-7 and spatial transcriptomics to follow the natural history of T cells that are reactive towards Helicobacter hepaticus through space and time in the settings of tolerance and inflammation. Although antigen stimulation can occur anywhere in the tissue, the lamina propria-but not embedded lymphoid aggregates-is the key microniche that supports effector Treg (eTreg) cell function. eTreg cells are stable once their niche is established; however, unleashing inflammation breaks down compartmentalization, leading to dominance of CD103+SIRPα+ dendritic cells in the lamina propria. We identify and validate the putative tolerogenic interaction between CD206+ macrophages and eTreg cells in the lamina propria and identify receptor-ligand pairs that are likely to govern the interaction. Our results reveal a spatial mechanism of tolerance in the lamina propria and demonstrate how knowledge of local interactions may contribute to the next generation of tolerance-inducing therapies.

Comparative ultrastructure of the olfactory system in the East African root rat (Tachyoryctes splendens) and the naked mole rat (Heterocephalus glaber).

The ultrastructure of the olfactory system of most fossorial rodents remains largely unexplored. This study sought to investigate the functional structure of the olfactory mucosa and olfactory bulb of two species of fossorial rodents that have distinct behaviour and ecology, the East African root rat (RR) and the naked mole rat (NMR). Transmission electron microscopy and scanning electron microscopy were employed. The basic ultrastructural design of the olfactory system of the two species was largely comparable. In both species, the olfactory mucosa comprised an olfactory epithelium and an underlying lamina propria. The olfactory epithelium revealed olfactory knobs, cilia and microvilli apically and sustentancular cells, olfactory receptor neurons and basal cells in the upper, middle and basal zones, respectively. The lamina propria was constituted by Bowman's glands, olfactory nerve bundles and vasculature supported by loose connective tissue. Within the olfactory bulb, intracellular and extracellular structures including cell organelles, axons and dendrites were elucidated. Notable species differences were observed in the basal zone of the olfactory epithelium and on the luminal surface of the olfactory mucosa. The basal zone of the olfactory epithelium of the RR consisted of a single layer of flattened electron-dense horizontal basal cells while the NMR had juxtaposed electron-dense and electron-lucent heterogenous cells, an occurrence seen as being indicative of quiescent and highly proliferative states of the olfactory epithelia in the two species, respectively. The olfactory epithelial surface of the NMR comprised an elaborate cilia network that intertwined extensively forming loop-like structures whereas in the RR, the surface was rugged and consisted of finger-like processes and irregular masses. With gross and histological studies showing significant differences in the olfactory structures of the two species, these findings are a further manifestation that the olfactory system of the RR and the NMR have evolved differently to reflect their varied olfactory functional needs.

NLRC5 expression profile in the oviduct of laying hens and its changes following estradiol treatment in induced molting hens

The innate immune defense system is initiated by recognizing components of microorganisms by different families of receptors such as TLRs and the intracellular NLR family. Among the NLR family, NLRC5 recognizes pathogenic components of microorganisms. The current study aimed to determine the expression profile of NLRC5 in the mucosal tissue of the oviduct of laying hens and to examine the effects of oviduct regression and estradiol benzoate (EB) on its expression. In this study, we used two groups of laying and molting hens. The molting group was treated by receiving a single dose of sesame oil or estradiol benzoate through intramuscular injection. To examine the histological differences in the oviduct between laying, molting, and estradiol-injected groups, Tissue samples from all segments of the oviduct of all groups were fixed in formalin and stained with hematoxylin and eosin. To observe the profile of NLRC5 gene and the effects of estradiol treatment, RNA was obtained from the surface epithelium and lamina propria of the whole segments of the oviduct of normal laying hens, and mucosal tissue of magnum, isthmus, uterus, and the vagina of molting groups. Both mucosal surface epithelium and lamina propria expressed NLRC5. The expression of NLRC5 was higher in the molting than in the laying group, and it was lower in the EB group. These results suggested that NLRC5 may have a role in recognizing pathogens invading the oviduct of laying and molting hens, and its expression is changed in association with oviduct growth. Bangladesh J. Zool. 51(3): 345-359, 2023                                            

444 Deciphering the role of IL-4 in post-colitis repair

OBJECTIVES/GOALS: Incomplete mucosal healingand dysbiosis prevent long-term remission after colitis. IL4 may restore colon homeostasis through its action on immune cells and the microbiome. We will demonstrate this mechanism using genetically modified mice and molecular tools. This may result in target therapies that prolong remission in patients with IBD. METHODS/STUDY POPULATION: Mice were treated with 3% dextran sulfate sodium (DSS) in drinking water for 5 days to induce colitis. Mice were monitored daily for changes in body weight, and to monitor colitis severity. At each endpoint, mice were sacrificed and colon length was measured. For disease severity assessment, mouse colons were prepared in paraffin sections by the 'swiss-rolling' method. For flow cytometry, lamina propria mononuclear cell isolation was performed and cellular populations were stained with fluorophore-conjugated antibodies. IL4-eGFP-expressing (4get) mice were used to analyze the cellular expression of IL4 after colitis. Cell-specific IL4 deletion mice were generated using the cre-lox system. RESULTS/ANTICIPATED RESULTS: IL4-deficient mice had worse colitis compared with wild-type controls. Flow cytometry of lamina propria cells from 4get mice showed that most IL4-producing cells after colitis are eosinophils (CD11b+SiglecF+). Flow cytometry of C57bl6 mice showed an influx of IL4Ra+ monocytes (CD11b+Ly6C+) and macrophages (CD11b+F480+). IL4-stimulated bone marrow-derived macrophages demonstrated an increase in HB-EGF mRNA transcription. Myeloid-specific IL4R deleted mice had no difference in colitis severity compared with controls. Neutrophil-specific IL4R-deleted mice had increased colitis severity and mortality. Co-housing of littermate mice rescued recovery after DSS in IL4 deficient mice. DISCUSSION/SIGNIFICANCE: IL4 appears to play a role in restoring homeostasis after colitis. The mechanism depends on eosinophil-derived IL4, and action through neutrophils. However, the reparative function of IL4 can be shared with deficient mice through the microbiome. I will study the cellular and microbial mechanism by which IL4 restores homeostasis after colitis.

Characterization of Th17 tissue-resident memory cells in non-inflamed intestinal tissue of Crohn's disease patients

Crohn's disease (CD) is a chronic inflammatory disorder affecting the bowel wall. Tissue-resident memory T (Trm) cells are implicated in CD, yet their characteristics remain unclear. We aimed to investigate the transcriptional profiles and functional characteristics of Trm cells in the small bowel of CD and their interactions with immune cells. Seven patients with CD and four with ulcerative colitis as controls were included. Single-cell RNA sequencing and paired T cell receptor sequencing assessed T cell subsets and transcriptional signatures in lamina propria (LP) and submucosa/muscularis propria-enriched fractions (SM/MP) from small bowel tissue samples. We detected 58,123 T cells grouped into 16 populations, including the CD4+ Trm cells with a Th17 signature and CD8+ Trm clusters. In CD, CD4+ Trm cells with a Th17 signature, termed Th17 Trm, showed significantly increased proportions within both the LP and SM/MP areas. The Th17 Trm cluster demonstrated heightened expression of tissue-residency marker genes (ITGAE, ITGA1, and CXCR6) along with elevated levels of IL17A, IL22, CCR6, and CCL20. The clonal expansion of Th17 Trm cells in CD was accompanied by enhanced transmural dynamic potential, as indicated by significantly higher migration scores. CD-prominent Th17 Trm cells displayed an increased interferon gamma (IFNγ)-related signature possibly linked with STAT1 activation, inducing chemokines (i.e., CXCL10, CXCL8, and CXCL9) in myeloid cells. Our findings underscored the elevated Th17 Trm cells throughout the small bowel in CD, contributing to disease pathogenesis through IFNγ induction and subsequent chemokine production in myeloid cells.

Tacrolimus-Associated Thrombotic Microangiopathy Presenting with Ischemic Colitis After Kidney Transplantation: A Case Report

Calcineurin inhibitors (CNIs) are a significant component of the immunosuppressive regimen after kidney transplantation. By inhibiting cytokine gene transcription, CNIs suppress T cell and T cell-dependent B cell activation. Tacrolimus is preferred in most patients undergoing kidney transplantation. Thrombotic microangiopathy (TMA) is a severe but rare complication of CNIs therapy. TMA defines a specific pathologic lesion of arterioles and capillaries that leads to microvascular thrombosis. A 45-year-old male underwent kidney transplantation five months ago due to autosomal dominant polycystic kidney disease (ADPKD). His triple-maintenance immunosuppressive therapy includes tacrolimus. Abdominal pain and bloody diarrhea occurred in the fifth month of posttransplant. The edematous and erythematous mucosa of the ascending colon was detected on the colonoscopy. The foci of microthrombi inside the vessel lumen in the lamina propria were shown biopsy. The thrombosis or occlusion was excluded with computerized tomography (CT) angiography in abdominal vessels. The fragmented red blood cells and moderate thrombocytopenia were detected on the peripheral blood smear. Eventually, TMA diagnosis was established through laboratory and histological findings. Tacrolimus was suspected as the trigger of the pathological process and promptly switched to the everolimus. Afterward, laboratory abnormalities and clinical symptoms were improved. In this case, we intend to emphasize drug-associated TMA and atypical presentations, such as colonic microvasculature involvement.

Excess of circulating apotransferrin enhances dietary iron absorption in mice

AbstractIntravenous injection of excess apotransferrin enhances dietary iron absorption in mice and triggers accumulation of plasma non–transferrin-bound iron. Injected fluorescent-labeled transferrin colocalizes with lamina propria macrophages, consistent with the recently proposed iron absorption checkpoint involving macrophage-mediated transferrin degradation.

The developmental architecture of the crop of quail embryos (Coturnix coturnix): Morphometric, light and scanning electron microscopy.

The present study focused on the histogenesis of the crop in quail embryos. The developmental sequence was recorded from day 10 of incubation to the hatching day (17th day) by gross anatomy, morphometric analysis and by using light and scanning electron microscope (SEM). Grossly, the crop was distinguished as an enlarged pouch of the cervical oesophagus on the 12th day of incubation. Histologically, the development of the crop began on the 10th day of incubation. The muscularis mucosae was observed as a small, thin layer of myoblast cells on the 10th day, then it appeared as longitudinally arranged smooth muscle fibres with progression of the developmental age till hatching day. The submucosal layer was also observed on the 10th day, while the lamina propria, muscular layer (inner circular and outer longitudinal layers of smooth muscle fibre) and glandular primordia were detected on the 12th day. The epithelial cells of crop revealed affinity for PAS stain and AB-PAS on the 10th day of incubation till the 17th day, while the affinity for AB stain appeared on the 14th day of incubation. Additionally, the glandular cells showed the positive for AB, PAS and AB-PAS on the 12th day of incubation. SEM sections showed variable sizes of the openings of the oesophageal glands. In conclusion, the present study provided a detailed overview of the histogenesis of the crop in quail embryos.

Long-Term Triclocarban Exposure Induced Enterotoxicity by Triggering Intestinal AhR-Mediated Inflammation and Disrupting Microbial Community in Mice.

Exposure to triclocarban (TCC), a commonly used antibacterial agent, has been shown to induce significant intestine injuries and colonic inflammation in mice. However, the detailed mechanisms by which TCC exposure triggered enterotoxicity remain largely unclear. Herein, intestinal toxicity effects of long-term and chronic TCC exposure were investigated using a combination of histopathological assessments, metagenomics, targeted metabolomics, and biological assays. Mechanically, TCC exposure caused induction of intestinal aryl hydrocarbon receptor (AhR) and its transcriptional target cytochrome P4501A1 (Cyp1a1) leading to dysfunction of the gut barrier and disruption of the gut microbial community. A large number of lipopolysaccharides (LPS) are released from the gut lumen into blood circulation owing to the markedly increased permeability and gut leakage. Consequently, toll-like receptor-4 (TLR4) and NF-κB signaling pathways were activated by high levels of LPS. Simultaneously, classic macrophage phenotypes were switched by TCC, shown with marked upregulation of macrophage M1 and downregulation of macrophage M2 that was accompanied by striking upregulation of proinflammatory factors such as Il-1β, Il-6, Il-17, and Tnf-α in the intestinal lamina propria. These findings provide new evidence for the TCC-induced enterotoxicity.

All-Trans-Retinoic Acid-Adjuvanted mRNA Vaccine Induces Mucosal Anti-Tumor Immune Responses for Treating Colorectal Cancer.

Messenger RNA (mRNA) cancer vaccines are a new class of immunotherapies that can activate the immune system to recognize and destroy cancer cells. However, their effectiveness in treating colorectal cancer located on the mucosal surface of the gut is limited due to the insufficient activation of mucosal immune response and inadequate infiltration of cytotoxic T cells into tumors. To address this issue, a new mRNA cancer vaccine is developed that can stimulate mucosal immune responses in the gut by co-delivering all-trans-retinoic acid (ATRA) and mRNA using lipid nanoparticle (LNP). The incorporation of ATRA has not only improved the mRNA transfection efficiency of LNP but also induced high expression of gut-homing receptors on vaccine-activated T cells. Additionally, the use of LNP improves the aqueous solubility of ATRA, eliminating the need for toxic solvents to administer ATRA. Upon intramuscular injections, ATRA-adjuvanted mRNA-LNP significantly increase the infiltration of antigen-specific, cytotoxic T cells in the lamina propria of the intestine, mesenteric lymph nodes, and orthotopic colorectal tumors, resulting in significantly improved tumor inhibition and prolonged animal survival compared to conventional mRNA-LNP without ATRA. Overall, this study provides a promising approach for improving the therapeutic efficacy of mRNA cancer vaccines against colorectal cancer.

Number of intraepithelial lymphocytes and presence of a subepithelial band in normal colonic mucosa differs according to stainings and evaluation method

Chronic watery diarrhea is a frequent symptom. In approximately 10% of the patients, a diagnosis of microscopic colitis (MC) is established. The diagnosis relies on specific, but sometimes subtle, histopathological findings. As the histology of normal intestinal mucosa vary, discriminating subtle features of MC from normal tissue can be challenging and therefore auxiliary stainings are increasingly used. The aim of this study was to determine the variance in number of intraepithelial lymphocytes (IELs) and presence of a subepithelial band in normal ileum and colonic mucosa, according to different stains and digital assessment. Sixty-one patients without diarrhea referred to screening colonoscopy due to a positive feacal blood test and presenting with endoscopically normal mucosa were included. Basic histological features, number of IELs, and thickness of a subepithelial band was manually evaluated and a deep learning-based algorithm was developed to digitally determine the number of IELs in each of the two compartments; surface epithelium and cryptal epithelium, and the density of lymphocytes in the lamina propria compartment. The number of IELs was significantly higher on CD3-stained slides compared with slides stained with hematoxylin-and-eosin (HE) (p<0.001), and even higher numbers were reached using digital analysis. No significant difference between right and left colon in IELs or density of CD3-positive lymphocytes in lamina propria was found. No subepithelial band was present in HE-stained slides while a thin band was visualized on special stains. Conclusively, in this cohort of prospectively collected ileum and colonic biopsies from asymptomatic patients, the range of IELs and detection of a subepithelial collagenous band varied depending on the stain and method used for assessment. As assessment of biopsies from patients with diarrhea constitute a considerable workload in the pathology departments digital image analysis is highly desired. Knowledge provided by the present study highlight important differences that should be considered before introducing this method in the clinic.

Verrucous gastritis-like lesion in intramucosal Helicobacter pylori-uninfected signet ring cell carcinoma with poorly differentiated adenocarcinoma.

In Japan, accessible Helicobacter pylori (Hp) eradication therapy is associated with an increase in the prevalence of gastric cancers (GCs) in Hp uninfected stomachs. Signet ring cell carcinoma (SRCC) is the most common of these GCs. Intramucosal SRCC with poorly differentiated adenocarcinoma (PDA) occurring in Hp uninfected gastric mucosa is rare; furthermore, many Hp uninfected pure SRCCs exhibit discoloration and flat or slightly depressed lesions, and morphological elevation is relatively rare. We report a case of intramucosal SRCC with PDA with an elevated, verrucous gastritis-like lesion in a 57-year-old male patient. In the present case, the PDA area showed dense tumor cell growth and coexisting desmoplastic and fibrotic reactions. Histopathology and immunohistochemical staining identified extensive fibromuscular obliteration with smooth muscle bundles extending from the muscularis mucosa into the lamina propria. The patient underwent curative endoscopic submucosal dissection. The reporting and analysis of such rare cases may lead to a better understanding of the characteristics of advanced Hp uninfected GCs.