The influence of ion composition, pH, and peptide concentration on the conformation and activity of the 37-residue human antibacterial peptide LL-37 has been studied. At micromolar concentration in water, LL-37 exhibits a circular dichroism spectrum consistent with a disordered structure. The addition of 15 mM HCO 3 ؊ , SO 4 2؊ , or CF 3 CO 2 ؊ causes the peptide to adopt a helical structure, with approximately equal efficiency, while 160 mM Cl ؊ is less efficient. A cooperative transition from disordered to helical structure is observed as the peptide concentration is increased, consistent with formation of an oligomer. The extent of ␣-helicity correlates with the antibacterial activity of LL-37 against both Gram-positive and Gram-negative bacteria. Two homologous peptides, FF-33 and SK-29, containing 4 and 8 residue deletions at the N terminus, respectively, require higher concentrations of anions for helix formation and are less active than LL-37 against Escherichia coli D21. Below pH 5, the helical content of LL-37 gradually decreases, and at pH 2 it is entirely disordered. In contrast, the helical structure is retained at pH over 13. The minimal inhibitory concentration of LL-37 against E. coli is 5 M, and at 13-25 M the peptide is cytotoxic against several eukaryotic cells. In solutions containing the ion compositions of plasma, intracellular fluid, or interstitial fluid, LL-37 is helical, and hence it could pose a danger to human cells upon release. However, in the presence of human serum, the antibacterial and the cytotoxic activities of LL-37 are inhibited.

The epithelia constitute a major barrier to the environment and provide the first line of defense against invading microbes. Antimicrobial peptides are emerging as participants in the defense system of epithelial barriers in general. Originally we isolated the human antimicrobial peptide LL-37 from granulocytes. The gene (CAMP or cathelicidin antimicrobial peptide) coding for this peptide belongs to the cathelicidin family, whose members contain a conserved pro-part of the cathelin type. The human genome seems to have only one gene of this family, whereas some mammalian species have several cathelicidin genes. In the present work we demonstrate up-regulation of this human cathelicidin gene in inflammatory skin disorders, whereas in normal skin no induction was found. By in situ hybridization and immunohistochemistry the transcript and the peptide were located in keratinocytes throughout the epidermis of the inflammatory regions. In addition, the peptide was detected in partially pure fractions derived from psoriatic scales by immunoblotting. These fractions also exhibited antibacterial activity. We propose a protective role for LL-37, when the integrity of the skin barrier is damaged, participating in the first line of defense, and preventing local infection and systemic invasion of microbes.Epithelia provide a barrier between the body and the environment. In addition, the epithelial cells have an active immunological role with antigen processing and presentation and production of cytokines and defense effector molecules such as microbicidal peptides. Thus, the epithelia mediate an active protection against invading microbes (1).Several broad spectrum microbicidal peptides have been identified in mammalian mucosal epithelium; bovine tracheal mucosa produces a ␤-defensin, TAP (tracheal antimicrobial peptide) (2), paneth cells of the gastrointestinal mucosa of human and mouse synthesize defensins (3, 4), and another ␤-defensin, LAP (lingual antimicrobial peptide) is expressed by bovine tongue epithelial cells (5). Thus, peptide antibiotics appear at the surface epithelium where they are likely to act as key components in the first line of defense and in the wound healing process (5). So far, all mammalian antimicrobial peptides identified at the mucosal interface belong to the defensin family. Defensins are cysteine-rich peptides folded in ␤-pleated sheets with a broad activity against bacteria, enveloped viruses, fungi, and parasites (6).We have isolated a clone for a novel human antibacterial peptide and named the putative peptide FALL-39 (7). Recently the mature active peptide LL-37 (two amino acids shorter at the N terminus than the putative peptide) was isolated from granulocytes and characterized (amino acid sequence is shown in Fig. 2B) (8). The preproprotein of LL-37 has also been named human CAP18 by another group (9). In contrast to the defensins, LL-37 is a cysteine-free peptide that can adopt an amphipathic ␣-helical conformation. The preproprotein belongs to the cathelicidin protein family. The common...

The urinary tract functions in close proximity to the outside environment, yet must remain free of microbial colonization to avoid disease. The mechanisms for establishing an antimicrobial barrier in this area are not completely understood. Here, we describe the production and function of the cathelicidin antimicrobial peptides LL-37, its precursor hCAP-18 and its ortholog CRAMP in epithelial cells of human and mouse urinary tract, respectively. Bacterial contact with epithelial cells resulted in rapid production and secretion of the respective peptides, and in humans LL-37/hCAP-18 was released into urine. Epithelium-derived cathelicidin substantially contributed to the protection of the urinary tract against infection, as shown using CRAMP-deficient and neutrophil-depleted mice. In addition, clinical E. coli strains that were more resistant to LL-37 caused more severe urinary tract infections than did susceptible strains. Thus, cathelicidin seems to be a key factor in mucosal immunity of the urinary tract.

Cathelicidins are a family of peptides thought to provide an innate defensive barrier against a variety of potential microbial pathogens. The human and mouse cathelicidins (LL-37 and CRAMP, respectively) are expressed at select epithelial interfaces where they have been proposed to kill a number of gram-negative and gram-positive bacteria. To determine if these peptides play a part in the protection of skin against wound infections, the anti-microbial activity of LL-37 and CRAMP was determined against the common wound pathogen group A Streptococcus, and their expression was examined after cutaneous injury. We observed a large increase in the expression of cathelicidins in human and murine skin after sterile incision, or in mouse following infection by group A Streptococcus. The appearance of cathelicidins in skin was due to both synthesis within epidermal keratinocytes and deposition from granulocyctes that migrate to the site of injury. Synthesis and deposition in the wound was accompanied by processing from the inactive prostorage form to the mature C-terminal peptide. Analysis of anti-microbial activity of this C-terminal peptide against group A Streptococcus revealed that both LL-37 and CRAMP potently inhibited bacterial growth. Action against group A Streptococcus occurred in conditions that typically abolish the activity of anti-microbial peptides against other organisms. Thus, cathelicidins are well suited to provide defense against infections due to group A Streptococcus, and represent an important element of cutaneous innate immunity.

Type 1 diabetes develops over many years and is characterized ultimately by the destruction of insulin-producing pancreatic beta cells by autoreactive T cells. Nonetheless, the role of innate cells in the initiation of this disease remains poorly understood. Here, we show that in young female nonobese diabetic mice, physiological beta cell death induces the recruitment and activation of B-1a cells, neutrophils and plasmacytoid dendritic cells (pDCs) to the pancreas. Activated B-1a cells secrete IgGs specific for double-stranded DNA. IgGs activate neutrophils to release DNA-binding cathelicidin-related antimicrobial peptide (CRAMP), which binds self DNA. Then, self DNA, DNA-specific IgG and CRAMP peptide activate pDCs through the Toll-like receptor 9-myeloid differentiation factor 88 pathway, leading to interferon-α production in pancreatic islets. We further demonstrate through the use of depleting treatments that B-1a cells, neutrophils and IFN-α-producing pDCs are required for the initiation of the diabetogenic T cell response and type 1 diabetes development. These findings reveal that an innate immune cell crosstalk takes place in the pancreas of young NOD mice and leads to the initiation of T1D.