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Muscularis macrophages, as the most abundant immune cells in the intestinal muscularis externa, exhibit tissue protective phenotype in the steady state. Owing to tremendous advances in technology, we now know the fact that muscularis macrophages are a heterogeneous population of cells which could be divided into different functional subsets depending on their anatomic niches. There is emerging evidence showing that these subsets, through molecular interactions with their neighbours, take part in a wide range of physiological and pathophysiological processes in the gut. In this review, we summarize recent progress (particularly over the past 4 years) on distribution, morphology, origin and functions of muscularis macrophages and, where possible, the characteristics of specific subsets in response to the microenvironment they occupy, with particular emphasis on their role in muscular inflammation. Furthermore, we also integrate their role in inflammation-related gastrointestinal disorders, such as post-operative ileus and diabetic gastroparesis, in order to propose future therapeutic strategies. [ABSTRACT FROM AUTHOR]

Background: Development of the human fetal stomach starts at the 4th week of intrauterine life. The knowledge of histogenesis of the musculature of the stomach is important for understanding, diagnosis, and treatment of gastric adenomyoma, congenital anomalies like congenital hypertrophic pyloric stenosis, and abnormalities in the development of the enteric nervous system. The aim of this study was to find out the histogenesis of the submucosa (SM), muscularis mucosa (MM), and muscularis externa (ME) in the human fetal stomach. Methodology: Sixty-three freshly aborted fetuses which were free from any gross abnormality were collected from the obstetrics and gynecology department and studied for 2 years. The fetuses were dissected, the stomach was removed, and the tissue was processed for a histological study. Staining was done by hematoxylin and eosin, Masson's trichrome, and Van Gieson's stain. Slides were studied under Zeiss branded light microscope using ×10 and ×40 objectives. Results: MM was discontinuous initially and became well defined by 21 weeks in the fundus and body and by the 23rd week in the pyloric region. The density of collagen fibers increased gradually in SM and was seen in bundles from the 23rd week onward. The thickness of SM was more than ME till 22 weeks, and after 27 weeks, it was less than that of ME. In the fundus and body of the stomach, till 18 weeks, ME consisted of two layers, with the inner oblique layer being thicker than the circular layer. The circular layer thickened gradually, and the longitudinal layer appeared by the 25th week. In the pyloric region, initially, there was only a circular layer. The oblique and longitudinal muscle layers appeared at 17–18 weeks and 25th week, respectively. The thickness of the circular layer was found to be more than that of the body from the 21st week onward and became much thicker at around the 27th week. The histological feature of MM, ME, and SM of the stomach resembled an adult pattern by the 28th week of gestational age. Conclusion: The findings of the present study about the time of appearance and further development of MM and ME and the thickness and formation of SM of the human fetal stomach are quite comprehensive. These findings are expected to aid in the knowledge of histogenesis of the stomach in the human fetuses and help in the diagnosis and treatment of various congenital anomalies and clinical conditions involving gastric connective tissue and musculature.

The primitive gut consists of two parts: an anterior and a posterior part joined by the midgut, which is open anteriorly and which communicates with the secondary yolk sac. The stomach is first visible in the fourth week, as a small fusiform dilatation located below the transverse septum (the future diaphragm). The stomach rotates during weeks 7 and 8. It rotates around the craniocaudal axis 90 degrees, from left to right, so that the initially left side of the stomach becomes anterior and the initially right side of the stomach becomes posterior. The stomach consists of four layers, which from the inside to the outside are: the tunica mucosa, the tunica submucosa, the tunica muscularis, and the tunica serosa. The stomach mucosa consists of the surface epithelium, below which are the lamina propria and muscularis mucosae. The submucosa consists of connective tissue. The muscular tunic or the muscularis externa is formed by three layers of smooth muscle fibers. The serosa is the outermost layer, being formed by a layer of squamous epithelium. [ABSTRACT FROM AUTHOR]

Background: Intestinal resident macrophages play a crucial role in homeostasis and have been implicated in numerous gastrointestinal diseases. While historically believed to be largely of hematopoietic origin, recent advances in fate‐mapping technology have unveiled the existence of long‐lived, self‐maintaining populations located in specific niches throughout the gut wall. Furthermore, the advent of single‐cell technology has enabled an unprecedented characterization of the functional specialization of tissue‐resident macrophages throughout the gastrointestinal tract. Purpose: The purpose of this review was to provide a panorama on intestinal resident macrophages, with particular focus to the recent advances in the field. Here, we discuss the functions and phenotype of intestinal resident macrophages and, where possible, the functional specialization of these cells in response to the niche they occupy. Furthermore, we will discuss their role in gastrointestinal diseases. [ABSTRACT FROM AUTHOR]

The intestinal macrophage pool represents the largest population of macrophages present within the body. Nevertheless, flow cytometry analysis of intestinal macrophages remains challenging due to historical lack of consensus on surface markers, variations in sample preparation, and a certain capriciousness of the isolation procedure itself. Furthermore, recent studies have uncovered a hitherto unknown heterogeneity of intestinal macrophages, accompanied by a vast increase of subset-identifying surface markers. Here, the isolation procedure for intestinal tissue for flow cytometry analysis is laid out, with particular attention toward the procedures for isolated intestinal layers, and a trouble-shooting section with strategies to avoid common pitfalls and mistakes., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Postoperative ileus (POI) is an iatrogenic complication of abdominal surgery, mediated by a severe inflammation of the muscularis externa (ME). We demonstrated that orally applicated CPSI-2364 prevents POI in rodents by blockade of p38 MAPK pathway and abrogation of NO production in macrophages. In the present experimental swine study we compared the effect of orally and intravenously administered CPSI-2364 on POI and examined CPSI-2364 effect on anastomotic healing. Methods: CPSI-2364 was administered preoperatively via oral or intravenous route. POI was induced by intestinal manipulation of the small bowel. ME specimens were examined by quantitative PCR for CCL2 chemokine gene expression and myeloperoxidase activity. Functional analyzes included measurement of ileal smooth-muscle ex vivo contractility, in vivo intestinal and colonic transit. Furthermore, anastomotic healing of a rectorectostomy after CPSI-2364 treatment was assessed by perianastomotic hydroxyproline concentration, a histochemically evaluated healing score and anastomotic bursting pressure (ABP). Results: CPSI-2364 abolished inflammation of the ME and improved postoperative smooth muscle contractility and intestinal transit independently of its application route. Hydroxyproline concentration and ABP measurement revealed no wound healing disturbances after oral or intravenous CPSI-2364 treatment whereas histological scoring demonstrated delayed anastomotic healing after intravenous treatment. Conclusion: CPSI-2364 effectively prevents POI in swine independently of its application route. Impairment of anastomotic healing could be observed after intravenous but not oral preoperative CPSI-2364 treatment. Subsumed, an oral preoperative administration of CPSI-2364 appears to be a safe and efficient strategy for prophylaxis of POI.

A full description of the terminal architecture of sympathetic axons innervating the gastrointestinal (GI) tract has not been available. To label sympathetic fibers projecting to the gut muscle wall, dextran biotin was injected into the celiac and superior mesenteric ganglia (CSMG) of rats. Nine days postinjection, animals were euthanized and stomachs and small intestines were processed as whole mounts (submucosa and mucosa removed) to examine CSMG efferent terminals. Myenteric neurons were counterstained with Cuprolinic Blue; catecholaminergic axons were stained immunohistochemically for tyrosine hydroxylase. Essentially all dextran-labeled axons (135 of 136 sampled) were tyrosine hydroxylase-positive. Complete postganglionic arbors (n = 154) in the muscle wall were digitized and analyzed morphometrically. Individual sympathetic axons formed complex arbors of varicose neurites within myenteric ganglia/primary plexus and, concomitantly, long rectilinear arrays of neurites within circular muscle/secondary plexus or longitudinal muscle/tertiary plexus. Very few CSMG neurons projected exclusively (i.e., ∼100% of an arbor's varicose branches) to myenteric plexus (∼2%) or smooth muscle (∼14%). With less stringent inclusion criteria (i.e., ≥85% of an axon's varicose branches), larger minorities of neurons projected predominantly to either myenteric plexus (∼13%) or smooth muscle (∼27%). The majority (i.e., ∼60%) of all individual CSMG postganglionics formed mixed, heterotypic arbors that coinnervated extensively (>15% of their varicose branches per target) both myenteric ganglia and smooth muscle. The fact that ∼87% of all sympathetics projected either extensively or even predominantly to smooth muscle, while simultaneously contacting myenteric plexus, is consistent with the view that these neurons control GI muscle directly, if not exclusively. J. Comp. Neurol. 524:2577-2603, 2016. [ABSTRACT FROM AUTHOR]

Background: Postoperative ileus (POI) is an iatrogenic complication of abdominal surgery, mediated by a severe inflammation of the muscularis externa (ME). We demonstrated that orally applicated CPSI-2364 prevents POI in rodents by blockade of p38 MAPK pathway and abrogation of NO production in macrophages. In the present experimental swine study we compared the effect of orally and intravenously administered CPSI-2364 on POI and examined CPSI-2364 effect on anastomotic healing. Methods: CPSI-2364 was administered preoperatively via oral or intravenous route. POI was induced by intestinal manipulation of the small bowel. ME specimens were examined by quantitative PCR for CCL2 chemokine gene expression and myeloperoxidase activity. Functional analyzes included measurement of ileal smooth-muscle ex vivo contractility, in vivo intestinal and colonic transit. Furthermore, anastomotic healing of a rectorectostomy after CPSI-2364 treatment was assessed by perianastomotic hydroxyproline concentration, a histochemically evaluated healing score and anastomotic bursting pressure (ABP). Results: CPSI-2364 abolished inflammation of the ME and improved postoperative smooth muscle contractility and intestinal transit independently of its application route. Hydroxyproline concentration and ABP measurement revealed no wound healing disturbances after oral or intravenous CPSI-2364 treatment whereas histological scoring demonstrated delayed anastomotic healing after intravenous treatment. Conclusion: CPSI-2364 effectively prevents POI in swine independently of its application route. Impairment of anastomotic healing could be observed after intravenous but not oral preoperative CPSI-2364 treatment. Subsumed, an oral preoperative administration of CPSI-2364 appears to be a safe and efficient strategy for prophylaxis of POI. © 2013 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Purpose: Postoperative ileus (POI) is an iatrogenic complication of abdominal surgery, mediated by a severe inflammation of the muscularis externa (ME). Previously, we demonstrated that intravenous application of the tetravalent guanylhydrazone semapimod (CNI-1493) prevents POI, but the underlying mode of action could not definitively be confirmed. Herein, we investigated the effect of a novel orally active salt of semapimod (CPSI-2364) on POI in rodents and distinguished between its inhibitory peripheral and stimulatory central nervous effects on anti-inflammatory vagus nerve signaling. Methods: Distribution of radiolabeled orally administered CPSI-2364 was analyzed by whole body autoradiography and liquid scintillation counting. POI was induced by intestinal manipulation with or without preoperative vagotomy. CPSI-2364 was administered preoperatively via gavage in a dose- and time-dependent manner. ME specimens were assessed for p38-MAP kinase activity by immunoblotting, neutrophil extravasation, and nitric oxide production. Furthermore, in vivo gastrointestinal (GIT) and colonic transit were measured. Results: Autoradiography demonstrated a near-exclusive detection of CPSI-2364 within the gastrointestinal wall and contents. Preoperative CPSI-2364 application significantly reduced postoperative neutrophil counts, nitric oxide release, GIT deceleration, and delay of colonic transit time, while intraoperatively administered CPSI-2364 failed to improve POI. CPSI-2364 also prevents postoperative neutrophil increase and GIT deceleration in vagotomized mice. Conclusions: Orally administered CPSI-2364 shows a near-exclusive dispersal in the gastrointestinal tract and effectively reduces POI independently of central vagus nerve stimulation. Its efficacy after single oral dosage affirms CPSI-2364 treatment as a promising strategy for prophylaxis of POI. [ABSTRACT FROM AUTHOR]

The gross and microscopical dimensions of small intestines from 12-week old streptozotocin-diabetic rats receiving no therapeutic intervention were compared with those from animals receiving insulin alone or in conjunction with the aldose reductase inhibitor, ponalrestat. Four regions along each intestine were analysed stereologically. Insulin had significant beneficial effects on body weight as well as on intestinal length, width, surface area and volume. In contrast, ponalrestat did not improve body weight deficits and was associated with crypt hypertrophy and a reduced villous surface/crypt volume ratio. There were interaction effects between insulin and ponalrestat for intestinal length and primary mucosal surface area. All groups displayed significant regional differences in surface area of primary mucosa and volume of muscularis externa. Only untreated diabetic rats failed to reveal regional variation in the surface area and volume of villi. Ratios of villous surface area/crypt volume varied from region-to-region in insulin-treated diabetic rats but not in other groups. The study fails to reveal any beneficial effect of aldose reductase inhibition on the changes in intestinal morphology seen in experimental diabetes. [ABSTRACT FROM AUTHOR]

Electron-microscopic studies have revealed a heterogeneous distribution of gap junctions in the muscularis externa of mammalian intestines. This heterogeneity is observed at four different levels: among species; between small and large intestines; between longitudinal and circular muscle layers; and between subdivisions of the circular muscle layer. We correlated results obtained with two immunomethods, using an antibody to the known gap-junctional protein (connexin43) with ultrastructural findings, and further evaluated the respective sensitivity of these two approaches. For comparative reasons we also included the vascular smooth muscle of coronary arteries into our study. Two versions of the immunotechnique (peroxidase-antiperoxidase and fluorescence methods) were applied to frozen sections of murine, canine, and human small and large intestines, as well as to pig coronary artery. In the small intestine of all three species a very strong reactivity marked the outer main division of the circular muscle layer, while the longitudinal muscle layer as well as the inner thin division of the circular muscle layer were negative. In murine and human colon both muscle layers were negative, while in canine colon the border layer between the circular muscle and the submucosa reacted strongly, and scattered activity was found in the portion of the circular muscle layer (one tenth of its thickness) closest to the submucosa. The remainder of the circular muscle layer and the entire longitudinal muscle layer were negative in the canine colon. In the coronary artery we could not confirm the positive, specific labeling reported by other investigators (l.c.). In conclusion, we found close correlations at all four above-mentioned levels in the distribution of gap junctions in the gut musculature, as determined by binding of anticonnexin43 in comparison to conventional ultrastructural studies. Since no significant immunostaining was found in (i) the outer border of the circular muscle layer of the canine colon and (ii) the border layer between the submucosa and the circular muscle layer of human colon, where rare gap junctions have been identified at the ultrastructural level, we conclude that the electron-microscopic analysis is the more sensitive of the two methods. [ABSTRACT FROM AUTHOR]

Interactions between the peripheral nervous system and resident macrophages (MMs) modulate intestinal homeostatic functions. Activation of β2-adrenergic receptors on MMs has been shown to reduce bacterial challenges. These MMs are also crucial for the development of bowel inflammation in postoperative ileus (POI), an iatrogenic, noninfectious inflammation-based motility disorder. However, the role of the sympathetic nervous system (SNS) in the immune modulation of these MMs during POI or other noninfectious diseases is largely unknown. By employing 6-OHDA-induced denervation, we investigated the changes in the muscularis externa by RNA-seq, quantitative PCR, and flow cytometry. Further, we performed transcriptional phenotyping of sorted CX3CR1+ MMs and ex vivo LPS/M-CSF stimulation on these MMs. By combining denervation with a mouse POI model, we explored distinct changes on CX3CR1+ MMs as well as in the muscularis externa and their functional outcome during POI. Our results identify SNS as an important mediator in noninfectious postoperative inflammation. Upon denervation, MMs anti-inflammatory genes were reduced, and the muscularis externa profile is shaped toward a proinflammatory status. Further, denervation reduced MMs anti-inflammatory genes also in the early phase of POI. Finally, reduced leukocyte infiltration into the muscularis led to a quicker recovery of bowel motility in the late phase of POI. [ABSTRACT FROM AUTHOR]

Competing Interests: Competing interests: None declared.

Background & Aims: Postoperative ileus (POI) is a common consequence of abdominal surgery that increases the risk of postoperative complications and morbidity. We investigated the cellular mechanisms and immune responses involved in the pathogenesis of POI., Methods: We studied a mouse model of POI in which intestinal manipulation leads to inflammation of the muscularis externa and disrupts motility. We used C57BL/6 (control) mice as well as mice deficient in Toll-like receptors (TLRs) and cytokine signaling components (TLR-2(-/-), TLR-4(-/-), TLR-2/4(-/-), MyD88(-/-), MyD88/TLR adaptor molecule 1(-/-), interleukin-1 receptor [IL-1R1](-/-), and interleukin (IL)-18(-/-) mice). Bone marrow transplantation experiments were performed to determine which cytokine receptors and cell types are involved in the pathogenesis of POI., Results: Development of POI did not require TLRs 2, 4, or 9 or MyD88/TLR adaptor molecule 2 but did require MyD88, indicating a role for IL-1R1. IL-1R1(-/-) mice did not develop POI; however, mice deficient in IL-18, which also signals via MyD88, developed POI. Mice given injections of an IL-1 receptor antagonist (anakinra) or antibodies to deplete IL-1α and IL-1β before intestinal manipulation were protected from POI. Induction of POI activated the inflammasome in muscularis externa tissues of C57BL6 mice, and IL-1α and IL-1β were released in ex vivo organ bath cultures. In bone marrow transplantation experiments, the development of POI required activation of IL-1 receptor in nonhematopoietic cells. IL-1R1 was expressed by enteric glial cells in the myenteric plexus layer, and cultured primary enteric glia cells expressed IL-6 and the chemokine monocyte chemotactic protein 1 in response to IL-1β stimulation. Immunohistochemical analysis of human small bowel tissue samples confirmed expression of IL-1R1 in the ganglia of the myenteric plexus., Conclusions: IL-1 signaling, via IL-1R1 and MyD88, is required for development of POI after intestinal manipulation in mice. Agents that interfere with the IL-1 signaling pathway are likely to be effective in the treatment of POI., (Copyright © 2014 AGA Institute. Published by Elsevier Inc. All rights reserved.)