Anti-Microbial Peptides and Their Speculative Role in Periodontitis

Humans often require a multilayered defense mechanism to function smoothly and combat micro-organisms and pathogens. The fundamental guarding complex for almost all human organisms comprises of innate immunity [1]. It is this defense mechanism that helps discern a wide variety of agents, known as pathogens, and distinguish them from the organism’s own healthy tissue [1]. The oral cavity is a manifold interaction where sundry organisms, both commensal and destructive types, intercommunicate and escalate in the environment [2]. The noteworthy quality of the oral cavity lies in the presence of specialized interaction between tooth (hard tissue) presenting itself from the underlying gingival epithelium (soft tissue).


Introduction
Humans often require a multilayered defense mechanism to function smoothly and combat micro-organisms and pathogens. The fundamental guarding complex for almost all human organisms comprises of innate immunity [1]. It is this defense mechanism that helps discern a wide variety of agents, known as pathogens, and distinguish them from the organism's own healthy tissue [1]. The oral cavity is a manifold interaction where sundry organisms, both commensal and destructive types, intercommunicate and escalate in the environment [2]. The noteworthy quality of the oral cavity lies in the presence of specialized interaction between tooth (hard tissue) presenting itself from the underlying gingival epithelium (soft tissue).
Ideally in the oral cavity, the tooth structure often encompasses a layer of pathogenic biofilm termed "Dental plaque," that compromises the surrounding epithelium via its incessant exposure to microorganisms [3]. This is when the comeback of the epithelium to these insults determines the overall condition of the gingival sulcus. It is evident that the oral epithelium works in several ways for the conservation of the underlying tissues. As a physical barricade, it can counter unbroken microbial oppositions from dental plaque by the production of chemokines, cytokines, and antimicrobial peptides (AMPs), which enhance inflammation and immune response in oral epithelial tissues [4]. These epithelial antimicrobial peptides are considered to be paramount for the innate immunity of the host [5].
AMPs are also prime contributors that enable the stabilization between health and disease in this complex ecosystem [6]. Exhibiting a wide spectrum of antimicrobial activity against grampositive and gram-negative bacteria, yeasts and certain viruses, they possess the ability to prevent various oral periodontal diseases including bacteria, fungal and viral infections [7]. The reaction involving multiple AMPs to a single pathogen of infection prevents the consequences of antimicrobial resistance. Several families of antimicrobial peptides have been studied in the oral cavity which includes α-defensins, β-defensins, calprotectin, adrenomedullin, histatins, and cathelicidin [8]. The aim of this review paper is to view the crucial role of these molecules against periodontal diseases and its function in host immunity. The article also sheds light on the mechanism of action and the types of identified AMPs. signal molecules like interleukin, chemokine and cytokines that attract neutrophils at the site and caution the host response [7]. Conducive with the amount of microbial exposure, they also produce natural AMPs and proteins. By acting as an integral part of the hosts natural innate environment, the oral epithelium, polymorph nuclear leukocytes (neutrophils) and saliva, all concurrently and solitarily bestow to this response [9,10]. These responses involve several salivary antimicrobial peptides, the β-defensins manifested in the epithelium, the α-defensins expressed in neutrophils, and the cathelicidin, LL-37, in both epithelium and neutrophils [11][12][13].

Defensins
The first AMPs identified in the oral epithelium were defensin, Lingual Antimicrobial Peptide (LAP) [14]. The defensin families are prominent AMPs that reside in the host environment, which include the alpha-defensins (human neutrophil peptide; HNP) and human beta-defensins (hBD) [15]. Defensins are stored in secretory granules, with expanding levels observed during periodontitis ( Figure 1). Their mechanism of action involves destroying phagocytosed bacteria upon fusion with the secretory granules from the phagocytic vacuoles they reside in [16]. Being a part of the host's natural environment, expressed in gingiva, tongue, salivary glands and mucosa, they are present in conditions like oral inflammation, carcinomas, etc. Defensins have also shown to inhibit LPS-stimulated inflammatory responses in host cells, which is a primary causal mechanism in the pathogenesis of periodontal disease [16]. Defensins are allocated into subfamilies of α-and β-defensins. Human beta-defensins (hBD) are elementally revealed in epithelial cells, while alpha-defensin are predominantly expressed in neutrophils [17]. Segregated in terms of their cysteine motifs, dually both of them share a homogenous secondary structure, and are opulent in cationic residues [18,19]. In the α-defensin subfamily, four of the six α-defensins, human neutrophil peptide -1, -2, -3, and -4, are fabricated and gathered in neutrophil granules while the other two α-defensins, HD-5 and -6, are synthesized and stored in the granules of paneth cells, specialized epithelial cells at the crypts of Lieberkuhn of the small intestine [17,20,21]. In the β-defensin subfamily only hBD -1, -2 and -3 are substantially expressed in the oral cavity [16,22]. hBD-3 along with the respective three mentioned above are markedly expressed in epithelial cells that encrust some tissues and organs, fundamentally skin [23]. In non-inflamed gingival tissues and gingival crevicular fluid, hBD-1 and hBD-2 have been expressed, with maximum levels at gingival margin [22]. Elevated levels of HNP1-3 have been noted in cases of periodontal infections, indicating a strong correlation See Table 1 for summary. Around 99% of hNP 1-3 is found in the saliva.
hNP-4 levels are lower hNP-5 and hNP-6 are not found in the oral cavity and are expressed in enteric system.
hNP-1 keeps a check on epithelial cell adhesion and spread.
hNP levels are seen to be higher with oral diseases like oral lichen plans, leukoplakia, SCC.
hBD-3: It is expressed in undifferentiated epithelial cells, which is within the basal layer.
hBD-1 is constitutively expressed hBD-2 and hBD-3 are unregulated in inflamed skin and other epithelial cells, by bacterial and pro-inflammatory stimuli. Low levels of unstimulated hBD-2 have been observed.
Principle AMPs holding major role in initial innate comeback against bacterial agents in tissues. Part of protective barrier in periodontal situations.

Anti-fungal
Anti-viral Additionally, they play a role in wound healing by rejuvenating the damaged epithelium.
They function as both proinflammatory and antiinflammatory agents in pathogenesis of diseases.

Chemical appeal of dendritic cells and t cells, macrophages
hBD-3 acts as a mediator to signal the underlying cells in the connective tissue.

Up-regulated levels hoods have been observed in inflammatory situations and infections.
Having higher levels at gingival margins close to plaque, hBD-1 and hBD-2 are expressed in noninflamed tissues.

Cathelicidin (LL-37)
Used markedly as one of the predominant identifying tools in inflammatory periodontal disorders, this class of AMPs incorporates a mature peptide and cathelin domain. It is a multifunctional peptide, comprising of 37 amino acids [24]. Active against gram positive and gram negative bacterias, it directly binds to the LPS of bacterial cells, and is native to the oral cavity in several sites, including the buccal and tongue mucosa, gingival crevicular fluid [24]. By activating antigen-presenting cells, it presents as a hemoattractant for immune cells, including monocytes, T cells, etc. There is a specific correlation amongst multiplication in LL-37 levels and periodontal inflammation [25][26][27][28]. It is also known to cause an elevation in mucosal thickness in the gingiva. Furthermore, studies have also demonstrated its tissue specific effects on cancer cells [27,28]. Their mode of action involves intracellular killing of the phagosomes after phagocytosis of the bacteria, where the AMP in the neutrophil is severed into a fully developed peptide (Table 2). Table 2: Summary of the role of Cathelicidin.

Gingival crevicular fluid in gingival sulcus
Primary antibacterial-Gram positive and gram negative bacterias Diagnostic tool in Inflammatory periodontal disorders Increased levels observed in chronic periodontitis, inflammatory conditions. Tissue specific in terms of cancer cells

Histatins
These AMPs are exclusively resided in the salivary glands, i.e. parotid and submandibular salivary duct cells. Histamine 1, 3 and 5 are found to be predominant of the total histatin proteins (85%) in the saliva [29][30][31][32]. They have a major role in fungicidal activity, having a noteworthy role in oral candidiasis restraint, especially histatin-5, which is the most significantly active against candida species as well as bactericidal activities against Porphyromonas gingivalis and Streptococcus mutans, which play a key role in the etiopathogenesis of periodontal disease and dental caries ,respectively [30,32]. Significant linkage occurs between xerostomia and oral candidiasis. It is also noteworthy that the antifungal action is shared between alpha-defensin and histatin, which require ATP transport found in active mitochondria.

Secretory Leuko Protease Inhibitor (SLPI)
Another AMP identified from the parotid salivary gland is SLPI. Manifested in neutrophils, epithelial cells and salivary glands, this protein suggests anti-inflammatory action at the site of infection [33]. Supplementary to having fungicidal action, it has shown to inhibit contamination of human monocytes and is also conveyed in oral tumor tissues as well as inhibition of the HIV virus [34,35]. SLPI is associated with wound repair and its expression is elevated following wound healing [35,36]. The elevated quantity of SLPI following periodontal treatment is an evidence of inflammation resolution activities [36].

Summary
The capacity of these inborn antibiotics is only at its inception of its recognition, as amplified natural announcement or as novel relieving agents [8,37]. This class of proteins is a captivating target for periodontal diseases [37]. In times of periodontal disease, AMPs amalgamate with other inflammatory proteins and maintain inflammatory molecules and pathways. It can be concluded that salivary AMPs have a prospective capacity to be recognized as initial markers of periodontal infections [38]. Thus, a dual approach is mandatory to comprehend the task of hosts' immunity in response to, against periodontitis [37]. This involves more comprehensive statistics about HNP and hBD, LL-37, and other oral antimicrobial peptides and proteins, along with their mode of action and clinical significance [38][39][40][41][42][43][44][45][46][47][48][49][50]. These peptides are peculiar in keeping the level of bacteria in control, having distinctive as well as dual roles in maintaining oral health. Not only can they act as diagnostic or prognostic biomarkers, but also the enhancement of new peptide agents can be a signaling method for future investigation and testing (Table 3).
Deficiency in LL-37 in neutrophils, plasma, and saliva Alpha-defensins deficiency ( by 70% ) Normal Lactoferrin levels Papillon-Lefe`vre syndrome [48,49] Haim-Munk syndrome [50] Palm plantar keratoderma and severe periodontitis Allelic mutations of the cathepsin C(CTSC) gene Moreover, Antimicrobial peptides are potentially important as novel therapeutic agents against periodontal disease diagnosis and potential treatment and probably show most immediate promise for development as topical adjuvant agents in conjunction with conventional periodontal therapy in the treatment of periodontal disease. This makes them promising for oral diseases, as topical application of antimicrobial agents is easy and appropriate, and their use would not contribute to resistance to antibiotics normally used in treatment of more life-threatening bacterial infections. All these findings will have direct implications for new understanding of oral innate immune responses and the development of potential new and innovative therapeutic interventions for periodontal disease.