**2. Origin, structure and function of the Paneth cell**

Paneth cells are found in the small intestine of humans and other vertebrates, including horses, sheep and rodents. Paneth cells are one of the four main derivatives of the intestinal stem cell [10, 11]. The other cells derived from the intestinal stem cells are enterocytes, goblet cells and neuroendocrine cells. These cells are found towards the luminal surface of the epithelium of the small intestine, whilst Paneth cells are located at the base of the intestinal crypt. They mingle with the crypt-based, Leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5+) intestinal stem cells in a 1:1 ratio [4, 12, 13]. Refer to **Figure 1** for more detail on the position of Paneth cells. Paneth cells are distinguishable from the Lgr5+ stem cells and other cells in the epithelium of the small intestine, which are found at the base of the intestinal crypt because they are secretory and have eosinophilic apical cytoplasmic granules and an extensive network of endoplasmic retinaculi [1, 12, 14, 15].

Paneth cells appear in utero from the 13th week of gestation [6, 16]. The number of Paneth cells progressively increase with a much more rapid expansion after the 29th week of pregnancy. A sufficient number of matured and functional Paneth cells is only attained at term [16]. Although Paneth cells are found in the entire colon of human embryos, they disappear soon after birth and are only found in the caecum and the ascending colon [17]. However, the location of Paneth cells in the epithelium of the small intestine is initially haphazard and more towards the luminal surface of the epithelium. Unlike the other derivatives of the intestinal stem cell, the Paneth cells later migrate downwards and end up at the base of the intestinal crypts. The stable number of Paneth cells in the crypt of a particular individual gets established in early adulthood. The overall number of Paneth cells in the intestinal crypt of the small

*Paneth Cells: The Gatekeepers of the Gut DOI: http://dx.doi.org/10.5772/intechopen.104402*

### **Figure 1.** *A schematic showing the structure and position of Paneth cells in the villus.*

intestine is influenced by factors such as the gestational age at birth, the mode of delivery, breastfeeding [6], weaning diet and period, dietary preference and disease states [17, 18]. The timing of colonization and the type of organisms involved play a role in both the development and number of Paneth cells [9, 19].

Once they are fully established, the Paneth cells are found at the base of the intestinal crypt, and in a healthy person, the number and distribution of Paneth cells remain relatively constant for up to 20 years [20]. Each intestinal crypt contains five to fifteen Paneth cells. The terminal ileum, which is the small intestine area with the highest load of microorganisms, contains the most Paneth cells. The overall number of Paneth cells in the small intestine may increase following a viral infection or a course of nutritional supplementation. Paneth cells may appear in ectopic sites due to metaplasia in certain diseased states, such as chronic inflammation [7, 21]. Similarly, the number of Paneth cells may be reduced due to malnutrition, the infestation of the GIT by parasites, chronic HIV disease, radiation enteritis, smoking, high fat diet, and aging [1–3].

There are three tiers of intestinal stem cells. The 1st tier of intestinal stem cells are found at the base of the crypt and are the so-called crypt base columnar cells (CBC cells) or Lgr5+ cells. The 1st tier stem cells are found in the most protected environment in the region where the most mature and furnished Paneth cells are. They are paired with Paneth cells on a 1:1 basis for intimate contact and direct communication. These Lgr5+ stem cells are the most vulnerable to radiation injury and require the most protection [22, 23]. It is the reason why they are closely associated with the Paneth

cells. They rely on the Paneth cells for both protection and nourishment. The Lgr5+ intestinal stem cells also have the least capacity to repair any damage to their DNA. The second tier of the intestinal stem cells is found high up along the crypt-villous axis at around cell position 4. The 3rd tier of cells with stem cell capability is higher up and closer to the crypt-villous transition zone and are also called intermediate cells. The 2nd and 3rd tiers are normally quiescent and only become activated on-demand, such as following an injury to the intestinal epithelium.

Other cells found in the epithelium of the small intestine are enterocytes and goblet cells. The enterocytes are the most populous derivatives of the intestinal stem cells and are found on the luminal surface of the small intestine. The small intestine has microvilli to increase the surface area to absorb nutrients. The function of the enterocytes is not limited to digestion and absorption of nutrients, as they also participate in the innate immunity of the small intestine. Combined with junctional proteins, enterocytes provide an uninterrupted physical barrier [10, 16, 22]. The enterocytes also secrete cytokines and chemokines, which recruit immune response elements. The lifespan of enterocytes is around five days, and they are being replaced continuously. Apoptotic enterocytes remain structurally intact until they are replaced to prevent the creation of defects on the surface of the epithelium and thus increased permeability. The enterocytes can de-differentiate into the stem cells following the destruction of the intestinal stem cells and Paneth cells. The enterocytes and goblet cells can de-differentiate into stem cells if they have not yet undergone terminal division. However, the plasticity of Paneth cells is far greater as they can be de-differentiating and acquire stemness even after they have been terminally divided [22]. The responsibility of de-differentiating to stem cells is first reserved for the Paneth cells afferent task. The intermediate cells, i.e. intermediaries of the other derivatives, only get involved and regain stemness if the Paneth cells have been irreversibly damaged and cannot play the role [23, 24].

The goblet cells are the second most populous cell type in the epithelium of the small intestine. They are interspersed among the enterocytes on the luminal surface of the epithelium. The lifespan of goblet cells is around five days and thus similar to that of enterocytes. Goblet cells are also found in the colon, where they play a role similar to that performed by Paneth cells in the small intestine [14, 16, 17]. Goblet cells secrete various types of mucins. The mucins produced by the goblet cells are the main constituent of the mucus, which coats the mucosal surface of the epithelium of the small intestine and colon. Even though the layer of mucus which lines the luminal surface of the small intestine is thin and attenuated, it can assist in maintaining a high concentration of antimicrobial peptides and protein in the area adjacent to the surface epithelial cells—the other function of the goblet cells secretion of trophic factors such as the trefoil factor. The goblet cells can de-differentiate and acquire stemness if the stem cells and Paneth cells have been damaged [10, 17, 22].

Paneth cells are tall columnar cells that are pyramidal in shape because of a broader base, have supra-nuclear Golgi apparatus, and zinc-rich apical orientated cytoplasmic granules [6]. The granules contain more than 50 constituents, including antimicrobial peptides like human alpha defensin 5 and 6 (HD5 and HD6), lysozyme, secretory phospholipase A2, osteopontin, and associated pancreatitis peptide, trypsinogen, IgA, TNF-alpha and alpha 1-antitrypsin and catecholamines [6]. The predominant constituent which is contained in the granules of Paneth cell is HD5, which make up about 90% of the components. The HD5 is the main antimicrobial peptide that is secreted by the Paneth cells of the small intestine, which is responsible for the control of luminal microbiota [6]. Degranulation of Paneth cells is induced following stimulation, and the granules are replenished expeditiously, usually within 24 hours.

### *Paneth Cells: The Gatekeepers of the Gut DOI: http://dx.doi.org/10.5772/intechopen.104402*

Paneth cells have autocrine, paracrine and endocrine functions. Similarly, Paneth cells respond to autocrine, paracrine or endocrine signals. Paneth cells also play a key role in innate and adaptive immunity by providing a direct line of communication between the two subsystems. They protect and regulate the functioning of the Lgr5+ intestinal stem cell and its derivatives, ensuring that exfoliating surface epithelial cells are regularly and timeously replaced [23]. Secretions from Paneth cells in the proximal parts of the small intestine influence the growth and function of distally situated small intestine stem cells and their derivatives [9]. The area at the base of the intestinal crypt (stem cell niche) is among the most active region in the body. The proliferation and differentiation of the Lgr5+ intestinal stem cells in the niche area is tightly regulated by secretions of Paneth cells and the cells situated in the connective tissue of the lamina propria around the base of the intestinal crypt [23]. The niche factors predominantly released by the Paneth cells influence the timing and type of division the intestinal stem cells should undergo while balancing the maintenance of the stemness and production of their progenies. The intestinal stem cell division may be symmetrical or asymmetrical [6, 10, 17].

A symmetrical division of the intestinal stem cells leads to the proliferation and production of daughter stem cells to expand the pool, whereas differentiation is prioritized during asymmetric division. The derivatives of the intestinal stem cell are generated following an asymmetric division. Both symmetrical and asymmetrical divisions are initiated and regulated by the niche factors which are produced by the Paneth cells and cells in the adjacent mesenchyme of the lamina propria in the peri-crypt space region. These niche factors, which include Wnt and Notch act as signals for the intestinal stem cells [23]. Even though they are in intimate physical contact, Paneth cells also have ligands for engagement with factors produced by the intestinal stem cells for cross-talk [23]. As indicated previously, When conditions are extremely hostile, and the stem cells have been destroyed, paneth cells can de-differentiate and acquire stem-ness.

Paneth cells have pattern recognition receptors that include nucleotide-binding oligomerization domain-like (NOD) and toll-like (TLR) receptors which they use to continuously sample the microbiome's composition in the lumen of the gut to prevent dysbiosis and or invasion by pathogenic organisms [24]. Paneth cells in the terminal ileum where Peyer's patches jointly sample the luminal contents and directly communicate with the M-cells. Paneth cells communicate remotely with the active immune cells and mesenchymal tissue in the lamina propria of the gut epithelium. Secretion from the Paneth cells is continuous with augmentation following a stimulus. The net effect of the Paneth cells' secretions depends on both their composition and volume. The secretions from the Paneth cells help to regulate and fine-tune the gut microbiome [9, 13, 24, 25]. The antimicrobial peptides secreted by the Paneth cells selectively kill pathogenic organisms while sparing the commensals. Ultimately, several microbial niches are created along the entire length of the small intestines where the microorganisms are naturalized and live in a symbiotic relationship with the host. Some of the organisms in the established niches assist during the digestion of food. Additionally, the composition of the flora freely floating in the lumen of the small intestine is different from the area close to the surface epithelial cells. The region closest to the epithelial cells' surface has the highest concentration of antimicrobial peptides and, therefore, the most repulsive to non-commensal organisms [26, 27].

Once established, the commensal organisms in the gut assist with digestion and absorption of essential nutrients in either the small intestine or colon, preventing the overgrowth of potentially pathogenic microorganisms and immune regulation [24]. Some of the ingested compounds in the food would not be digestible were it

not for certain species of resident microorganisms. The overall number of nutrients available in a particular site is a secretion of antimicrobial molecules that creates areas of zonal dominance by some species of microorganisms. The task of the Paneth cells is to accept the dominant organisms in various niches and for the sustenance of the symbiotic relationship. A deviation from the established normality is detected by the Paneth cells through their pathogen recognition receptors leading to degranulation and release of antimicrobial peptides. The change in the microbiome may occur following the use of broadspectrum antibiotics, change in diet, change in the anatomy of the GIT, alteration of gut transient time or a state of suppressed systemic immunity [25].

The other role of the Paneth cells is to nourish the intestinal stem cells [11, 12, 22]. Paneth cells derive their energy from the glycolytic pathways, whereas the intestinal stem cells' Adenosine triphosphate (ATP) is derived from aerobic metabolism in their mitochondria. The lactic acid which is produced by a Paneth cell is shunted into adjacent intestinal stem cells for their metabolism. Paneth cells can sense the fedstate of the body and, after that, influence the intestinal stem cell activity accordingly. If the epithelium is damaged, they become more active. Paneth cells are found in the normal human small intestine from the duodenum to the terminal ileum [26, 27]. They are most abundant in the region of the terminal ileum. The hostility of the environment, which increases as one move distally along the small intestine due to increment in the number and species of microbes, is a plausible explanation of the need for more Paneth cells at the region of the terminal ileum [26]. Paneth cells are not found in a healthy stomach and colon, except for a few in the caecum and the ascending colon [26]. Paneth cells may develop following metaplasia associated with chronic gastritis and inflammatory bowel disease in the stomach and colon. The effects of paneth cell secretions are enumerable and continue to be added. The result of the secreted factor varies depending on the type, volume and concentration of secretions. These effects include antimicrobial activity, inflammation and regulation of intestinal stem cells'. The antimicrobial peptides secreted by Paneth cells help sterilize the intestinal crypt environment, the so-called "stem cell zone; and thus protect intestinal stem cells [7, 13, 22, 28–30].

The antimicrobial peptides which are produced by the Paneth cells are of three types: Type 1 is cationic, Type 2 is amphipathic, and Type 3 is composed of hydrophobic peptides [31, 32]. The micro-biocidal effects of Type 1 and Type 2 are based on induction of damage to the surface of the cell membrane and creation of large pores as it penetrates deeper into the hydrophobic cell membrane and its bi-layer, respectively. Type 3 peptides cause micellization of the cell membrane of pathogens. The antimicrobial peptides produced by various cells in humans include cathelicidins and alpha and beta-defensins. Beta-defensins are produced in almost every cell in the body, including neutrophils. The Paneth cell is the only source of cathelicidins, HD5 and HD6 in humans. The synthesis of cathelicidins ceases when the foetus reaches term and is then replaced with HD5 and HD6 [6, 24]. The antimicrobial peptides from human Paneth cells are secreted in a pro-active form and become activated in the lumen of the small intestine [32]. The Paneth cell secretions are secreted together with water and chloride ions and assisted by peristalsis to bathe the crypt environment to make it conducive to functioning the tiers of intestinal stem cells [6, 22, 33]. The solvent load and anionic composition in the secretion assist in the after-release potentiation of the antimicrobial peptides [31]. In addition, the bile salts in the small intestine influence the killing activity of paneth cell-derived antimicrobial peptides. The human alpha-defensins are active against bacteria, fungi, parasites, and viruses [31, 32].

### *Paneth Cells: The Gatekeepers of the Gut DOI: http://dx.doi.org/10.5772/intechopen.104402*

The HD5 is microbicidal, and the HD6 binds the antigens to prevent invasion until it is eliminated. The action of HD6 is similar to that of the IgA antibodies. The binding of antigens by HD6 buys time for the other elements of innate immunity of the gut to arrive [31, 32].

Matured Paneth cells are found at the base of the intestinal crypt of the small intestine, where the most vulnerable but essential intestinal stem cells are found. The loss of senescent Paneth cells like absorptive enterocytes, goblet cells and neuroendocrine cells is programmed [6]. The senescent Paneth cells die and are removed through phagocytosis following apoptosis. The other mechanism involved in the death of Paneth cells is autophagy, which ensures that some essential constituents found in the cytoplasm are recycled. Apoptotic enterocytes located at the tip of villi remain structurally intact until they are replaced by a carpet of new cells arriving from the stem cell zone. A perfect balance between the rate of proliferation and loss through exfoliation is sustained to ensure that defects are not created, and the epithelium of the small intestine becomes permeable to microorganisms and their products [11, 16].

A healthy life without a normally functioning small intestine is not possible. The influence of the Paneth cells on the GIT starts soon after birth. Henceforth, the Paneth cell influences everything that happens in the gut, whether physical or biochemical barrier, absorption of nutrients, and linkage with the body's overall immune system. Paneth cells play a role in the development and maturation of the innate immunity of the gut and subsequently of the entire body. The immune system is a dominant player in systemic immunity, including adaptive immunity. Paneth cells maintain the integrity of the gut by controlling the microbiome, regulating proliferation and differentiation of the intestinal stem cells, influencing the quality of mucin in the mucous, and keeping the crypt environment relatively sterile for the protection of the intestinal stem cells. Paneth cells also release growth signals that influence the growth and function of the enterocytes, goblet cells, and neuroendocrine cells [7, 9, 11]. Chemokines and cytokines produced by Paneth cells can also recruit and influence components of adaptive immunity in the adjacent lamina propria [34]. Among the cytokines which are produced by the Paneth cells is TNF-alpha. There is also cross-talk between the Paneth cells and elements of adaptive immunity [34, 35].

A plethora of acute and chronic conditions are driven from the gut. These conditions may be initiated by a changing diet, starvation, trauma or sepsis. The normal development of the crypts and villi of the small intestine and control of the microbiome is dependent on the Paneth cells. Paneth cells continuously sample the luminal contents for the gut microbiota composition to prevent dysbiosis. Dysbiosis leads to increased gut permeability and translocation of bacteria and endotoxin [3]. Usually, when the body experiences significant physiological stress, the gut mucosa is strategically sacrificed; a typical example is the shunting of blood from the GIT in various shock states. If prolonged, what is meant to be a short-term survival strategy leads to dysbiosis, across the intestinal epithelium. Translocation of bacteria and endotoxins is the driver of systemic inflammation. The gut is the most trusted and potent site for eliminating invading pathogens [35, 36]. Shunting of invading pathogens to the GIT also applies to viruses, including HIV. Should the gut immune system fail to eliminate the pathogen, as it happens following HIV infection, the gut ultimately becomes a long-term reservoir and haven of mutated strains of the virus, which is currently impossible to eradicate despite the availability of potent antiviral drugs [36, 37].
