Implications of Paneth cell dysfunction on gastrointestinal health and disease : Current Opinion in Gastroenterology

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IMMUNOLOGY: Edited by Jocelyn A. Silvester

Implications of Paneth cell dysfunction on gastrointestinal health and disease

Lee, Vivian H.a,d; Gulati, Ajay S.b,c,d

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Current Opinion in Gastroenterology: November 2022 - Volume 38 - Issue 6 - p 535-540
doi: 10.1097/MOG.0000000000000887
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Abstract

INTRODUCTION

Paneth cells are professional secretory cells of the small intestine that reside at the base of the crypts of Lieberkühn. They were first described in the late 19th century by Gustav Schwalbe and Josef Paneth as columnar epithelial cells containing eosinophilic, apically clustered granules (Fig. 1) [1]. There are approximately 5–15 PCs per small intestinal crypt, which are derived from and ultimately interspersed between the intestinal stem cells (ISC) at the crypt base. In contrast to other small intestinal epithelial cell types that migrate upwards out of the crypt base toward the villi as they mature, Paneth cells migrate downwards toward the crypt base. They have a life span of approximately 30 days, which is much longer than other intestinal epithelial cells that turn over every 3–5 days. While primarily found in the small intestine and occasionally in the right colon of humans, Paneth cells have also been identified in esophageal, gastric, and colonic epithelium in the context of metaplasia. Paneth cell development occurs prenatally in humans [2] and postnatally in mice [3].

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FIGURE 1:
Microscopic appearance of Paneth cells. (a) Representative tissue section from mouse jejunum (H&E stain, 100× magnification). The Paneth cells are located at the base of the crypts, recognized by their highly distinct eosinophilic granules. (b) Electron micrograph of a mouse ileal crypt (2000× magnification). Paneth cells are characterized by their electron-dense, apically positioned cytoplasmic granules.

Broadly speaking, the primary function of Paneth cells is to support the epithelial barrier of the small intestine. This is accomplished in two key ways (Fig. 2). First, Paneth cells support the physical barrier of the epithelium by providing essential niche signals to their neighboring ISCs. These signals are mediated by factors such as epidermal grown factor, wingless/integrated ligands, and Notch ligand Dll4 [4], which are critical for epithelial cell renewal. Second, Paneth cells secrete antimicrobial peptides (AMPs) into the crypt lumen and unstirred mucus layer of the gut. This creates an antimicrobial barrier that protects the host from enteric pathogens, safeguards against bacterial translocation, and shapes the composition of the resident microbiota.

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FIGURE 2:
Paneth cell-mediated regulation of intestinal barrier function. (1) Paneth cells support proliferating stem cells by secreting epithelial growth factor (EGF), Wnt3, and Notch ligands. (2) Paneth cells also secrete antimicrobial peptides (AMPs) into the gut lumen that defend against enteric pathogens and regulate the composition of the enteric microbiota. Adapted from ‘Keystone Gut Microbiota Species Provide Colonization Resistance to Invading Bacteria’ (2020), by Biorender.com. Retrieved from https://app.biorender.com/biorender-templates.

AMPs are evolutionarily conserved, innate immune effector molecules found in Paneth cell granules that are secreted into the crypt lumen. Human Paneth cell-derived AMPs include two α-defensins known as human α-defensin (HD)-5 and HD-6. HD-5 has direct antimicrobial activity, and HD-5 transgenic mice are immune to infection by Salmonella typhimurium[5]. In contrast, HD-6 entraps both gram-positive and -negative bacteria in extracellular peptide nets [6]. Human Paneth cells also produce other AMPs including lysozyme, secretory phospholipase A2, and regenerating islet-derived protein IIIA [7]. In contrast, mice express a myriad of AMPs including α-defensin (cryptdins), cryptdin-like peptides, lysozyme (Lyz), phospholipases, C-type lectins (e.g. Reg3γ), and ribonucleases (e.g. Ang4) [8]. Collectively, these human and mouse AMPs have a broad spectrum of activity against bacterial, viral, and protozoal organisms. 

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Box 1:
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CROHN'S DISEASE

Crohn's disease is a subtype of inflammatory bowel disease characterized by skip lesions and transmural inflammation of the gastrointestinal tract. Approximately 50% of Crohn's disease patients have disease involvement of the terminal ileum and colon, 30% have isolated small bowel involvement, and 20% have isolated colonic disease [9]. Clinical manifestations include abdominal pain, diarrhea, and systemic symptoms such as fatigue, fever, and weight loss. Although the cause of Crohn's disease is multifactorial, several Crohn's disease-risk polymorphisms identified in genome-wide association studies have been linked to Paneth cell dysfunction. Examples of these include autophagy-related 16-like 1 (ATG16L1) [10], unfolded protein response transcription factor X-box binding protein-1 (XBP1) [11], leucine-rich repeat kinase 2 (LRRK2) [12], and nucleotide-binding oligomerization domain 2 (NOD2) [13].

Several of the Crohn's disease-risk alleles associated with Paneth cell dysfunction are involved in processes such as autophagy, the unfolded protein response, and the regulation of mitochondrial function. As Paneth cells are highly secretory cells, they may be subject to dysfunction when these intracellular housekeeping and stress responses go awry. For example, ATG16L1 encodes a protein critical for autophagy, which is an intracellular degradation pathway that allows for the clearance of pathogens and damaged organelles. ATG16L1 hypomorphic mice have disorganized and decreased numbers of Paneth cell granules, with impaired granule exocytosis [14]. A Paneth cell-specific deletion of mouse Xbp1, which encodes a transcription factor involved in the unfolded protein response, leads to induction of endoplasmic reticulum stress, compensatory autophagy, and spontaneous ileitis [15]. Interestingly, dual depletion of ATG16L1 and Xbp1 in mice results in a more severe Crohn's disease-like transmural ileitis [15].

Several human studies have also identified abnormal Paneth cells in the context of Crohn's disease. For example, the expression of α-defensins HD-5 and HD-6 is reduced in ileal Crohn's disease biopsy samples compared with colonic Crohn's disease, ulcerative colitis, pouchitis, or control biopsies [16]. The authors further identified a reduction in HD-5 protein levels and decreased total antimicrobial activity against Escherichia coli and Staphylococcus aureus in patients with ileal Crohn's disease compared with controls, demonstrating a biological outcome resulting from changes in HD-5 expression. Interestingly, mice with deficient α-defensin function also develop significant alterations in their enteric microbiota [17]. Collectively, these findings support the theory that Paneth cell dysfunction may lead to a dysbiotic microbiota, which in turn, could predispose to the development of Crohn's disease.

More recently, several studies have linked dysmorphic Paneth cells to disease outcome in Crohn's disease. For example, Crohn's disease patients with higher proportions of dysmorphic Paneth cells demonstrate a significantly shorter time to disease relapse after surgical resection, compared with patients with lower proportions of abnormal Paneth cells [13]. These dysmorphic Paneth cells are stable over time and not impacted by ongoing intestinal inflammation [18], making them an intriguing biomarker for disease prognostication. Importantly, dysmorphic Paneth cells are also associated with enteric dysbiosis. In a study of ileal mucosal samples isolated from pediatric ileal Crohn's disease patients, patients with a higher percentage of abnormal Paneth cells showed significantly reduced bacterial diversity compared with patients with a lower percentage of abnormal Paneth cells, reflecting a reduced abundance of anti-inflammatory microbes [19]. This again aligns with the purported pathogenesis of Crohn's disease, further supporting the possible importance of Paneth cells in certain patients with this disorder.

ENTERIC INFECTIONS

Several animal models demonstrate that impaired Paneth cell function increases host susceptibility to intestinal infection. Matrix metalloproteinase 7 (Mmp7) knockout mice are commonly used to model α-defensin deficiency. Mmp7 is a protease produced by Paneth cells that cleaves AMPs into their active forms. Mmp7 knockout mice have increased mortality to S. typhimurium infection [20]. This may be partly mediated by IL-22 signaling, as Paneth cell-specific knockdown of the interleukin 22 receptor (Il22ra1) results in decreased expression of Lyz and Mmp7 [21▪▪]. When infected with S. typhimurium, these mice demonstrate an increased dissemination of infection to the liver, suggesting that Paneth cells play a role in preventing systemic infection. On the other hand, transgenic mice that overexpress human HD-5 are resistant to oral S. typhimurium and demonstrate significantly reduced ileal S. typhimurium loads compared with wild-type mice [5]. Finally, knockout of Nod2 or Xbp1 in mice (both of which are Crohn's disease-risk alleles associated with Paneth cell dysfunction) increases susceptibility to Listeria monocytogenes infection [11,22].

Outcomes of aberrant Paneth cell function on viral and parasitic infections are less well understood. However, it is known that defensins can act at several steps of the viral life cycle of enveloped and nonenveloped viruses, including inhibition of viral binding and fusion to host receptor and postentry neutralization [23]. Regarding parasitic infections, a Paneth cell-specific deletion of the autophagy protein Atg5 leads to severe enteritis, hypomorphic Paneth cells, and increased acute mortality to Toxoplasma gondii infection [24].

GRAFT-VERSUS-HOST DISEASE

Graft-versus-host disease (GVHD), a potential complication of allogeneic hematopoietic stem cell transplantation, is a multisystem disorder that often affects the gastrointestinal system and liver, manifesting as abdominal pain and diarrhea. GVHD is histologically characterized by epithelial cell apoptosis, cryptitis, and crypt destruction. Prior studies utilizing mouse models of GVHD have demonstrated a decrease in Paneth cell number and α-defensin transcripts (Defa-1, 4, 5, 21, 22, rs1) compared with non-GVHD controls [25]. In human studies, lower numbers of duodenal Paneth cells correlate with more clinically severe GVHD and serve as better predictors of disease severity than classical pathologic grading [26]. Lower Paneth cell numbers were also associated with a decreased likelihood of response to treatment. These studies suggest a possible clinical utility for Paneth cell enumeration in patients with GVHD.

Mechanistically, decreased Paneth cell numbers promote enteric dysbiosis in GVHD, evidenced by decreased diversity of the gut microbiota [25]. This includes decreases in the commensal phyla Bacteroidetes and Firmicutes, and an expansion in E. coli. The former is negatively correlated, and the latter positively correlated with GVHD severity in mice [25]. Human studies mirror a similar loss in microbial diversity and demonstrate shifts in the phylum Firmicutes, specifically an increase in Lactobacillales (predominantly Lactobacillus johnsonii) and a decrease in Clostridiales [27]. As greater microbial perturbation following bone marrow transplant predicts the development of GVHD in this study, Paneth cells may play an indirect role in the pathogenesis of intestinal GVHD through modulation of the enteric microbiota. Interestingly, administration of recombinant human R-Spondin1, which stimulates differentiation of intestinal stem cells towards mature Paneth cells, prevents GVHD-mediated dysbiosis [28]. Similar results were observed in this study with recombinant α-defensin (cryptdin-4) administration. As such, provision and/or stimulation of Paneth cell-derived AMPs may represent novel therapeutic modalities to mitigate GVHD-mediated dysbiosis.

NEONATAL NECROTIZING ENTEROCOLITIS

Neonatal necrotizing enterocolitis (NEC) represents one of the most common gastrointestinal emergencies in preterm infants and is characterized by ischemic necrosis of the intestine. Initial signs and symptoms include abdominal distention, feeding intolerance, bilious emesis, and rectal bleeding. Pathognomonic imaging findings include pneumatosis intestinalis, pneumoperitoneum or sentinel bowel loops. Dysfunctional Paneth cells are thought to play a role in the multifactorial pathogenesis of NEC by contributing to inadequate intestinal barrier function. An analysis of small intestinal mucosa from 10 preterm and term newborn infants diagnosed with NEC showed a decrease in Paneth cells compared with controls [29]. In 9 of the 10 patients, no Paneth cells were detected; in the last patient, only rare Paneth cells were observed. Notably, the absence of Paneth cells was also demonstrated in nonnecrotic tissue. A subsequent human study that analyzed nonnecrotic ileal tissue also found a similar decrease in Paneth cells in infants with NEC compared with gestational age-matched controls [30]. These findings suggest that that loss of Paneth cells and/or diminished Paneth cell function may play a role in the pathogenesis of NEC.

Animal studies corroborate a potential role for Paneth cells in NEC. Ablation of Paneth cells with the heavy metal chelator dithizone, followed by enteric gavage of a human enteroinvasive E. coli (EIEC) strain, induces a NEC-like small intestinal injury and increases morbidity and mortality in newborn rats. Increased numbers of E. coli were found in jejunal and ileal luminal contents in dithizone-treated rats, suggesting diminished bacterial clearance in the setting of compromised Paneth cell function. Interestingly, mucosal-associated E. coli levels were elevated in the jejunum but not the ileum [31]. As such, there may be differential Paneth cell functions across the regions of the small intestine. Depletion of Paneth cells by treating genetically engineered mice (diptheria toxin receptor inserted into cryptdin-2 promoter) with diptheria toxin, followed by a single enteric gavage with Klebsiella pneumoniae, leads to decreased Paneth cell number, a NEC-like small intestinal injury and a sustained bloom of Enterobacteriacae in the cecum [32]. These studies suggest that diminished Paneth cell AMP function may provide conditions for engraftment by opportunistic pathogenic bacteria, predisposing the host to NEC.

ENVIRONMENTAL ENTERIC DYSFUNCTION

Environmental enteric dysfunction (EED) is a subclinical disorder characterized by systemic and small intestinal inflammation, typically in the absence of diarrhea. EED predominantly impacts children from low-income and middle-income countries and is hypothesized to be caused by recurrent fecal–oral contamination in the context of poor sanitation and hygiene. Histologic hallmarks of EED include blunting of small intestinal villi, crypt hyperplasia, and lymphocytic infiltration of the epithelium and lamina propria. Clinically, EED may present as malabsorption and growth stunting [33]. Paneth cells may be implicated in the pathogenesis of EED. A recent study of small intestinal biopsies from Zambian children with EED complicated by growth stunting were found to have depleted Paneth cells and goblet cells compared with control (non-EED) biopsies [34▪]. The resulting impairment in secretory function may compromise intestinal barrier function, further perpetuating the intestinal inflammation characteristic of EED. A separate study comparing duodenal biopsies from Zambian and Pakistani children with EED complicated by growth stunting identified a loss of Paneth cells and goblet cells in the Zambian cohort, but no change in the Pakistani cohort compared with controls [35]. Of note, the Zambian cohort was more severely malnourished and had higher C-reactive protein levels reflecting more severe EED than the Pakistani cohort. These findings suggest a possible geographic heterogeneity in EED, as well as an association of Paneth cell abnormalities with disease severity in this disorder.

CONCLUSION

Although Paneth cell alterations are associated with several gastrointestinal inflammatory conditions such as Crohn's disease, enteric infection, GVHD, NEC, and EED, whether these alterations are the result of inflammation, or the initial driver of inflammation remains unclear. Moreover, the precise mechanisms by which Paneth cell biology is regulated are poorly understood. For example, the influences of the gut microbiota, diet, and environmental influences on Paneth cell function require further investigation. Understanding how these external factors impact Paneth cell biology is necessary for the development of novel therapeutics that target Paneth cells to optimize intestinal mucosal barrier function and prevent chronic inflammatory responses. Finally, further understanding of how recombinant AMPs can be used to prevent and/or treat intestinal dysbiosis and infection is needed. To date, no clinical trials have utilized Paneth cell-specific AMPs in the context of gastrointestinal disease. Such therapies are appealing, as treatment with AMPs confers a theoretically low risk of bacterial resistance and may help mitigate enteric dysbiosis. In the future, enhanced understanding of Paneth cell biology has the potential to open a new class of safer, precision therapeutics for a wide range of gastrointestinal disorders.

Acknowledgements

None.

Financial support and sponsorship

This work was supported by grants to V.H.L. (NIH/NIDDK T32 DK007737, PI: Balfour Sartor) and A.S.G. (NIH/NIDDK R01 DK122042 and NIH/NIDDK P30 DK034987, PI: Robert Sandler).

Conflicts of interest

The authors have no conflicts of interest.

REFERENCES AND RECOMMENDED READING

Papers of particular interest, published within the annual period of review, have been highlighted as:

▪ of special interest

▪▪ of outstanding interest

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Keywords:

antimicrobial peptide; Crohn's disease; Paneth cell

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