Overlapping peptides (p1-p14) are underlined in the chemerin sequence.

<p>The N-terminal signal peptide is indicated by italics. Peptide 4 is shown in bold.</p

The chemerin-derived peptide 4 (Val⁶⁶-Pro⁸⁵) strongly inhibits growth of <i>E. coli</i>.

<p>Chemically synthesized chemerin peptides (p1-p14) were tested against <i>E. coli</i> HB101 using the microtitre broth dilution assay (A) or against <i>E. coli</i> ATCC 25922 using radial diffusion assay (RDA) in physiological 0.15 M NaCl (B) or low salt concentration (C). Bacteria were incubated with the peptides at 100 µM. The results are expressed as the mean ± SD of three independent experiments.</p

Synthetic chemerin peptides¹.

<p>Peptide 4 is shown in bold. The net charge at pH 6 (netchg), mean hydrophobicity (H), relative mean hydrophobic moment assuming a β structure and α-helix, (rHMβ) and (rHMα) respectively, and rHMβ/rHMα ratio are indicated for each peptide. Data for the antibacterial peptide protegrin-1 (pg-1) and magainin-2 (Mag-2) known to adopt β structure and α-helical conformation, respectively when bound to the lipid membrane <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058709#pone.0058709-Gesell1" target="_blank">[36]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058709#pone.0058709-Mani1" target="_blank">[37]</a>, are shown for comparison. The rHM values and rHMβ/rHMα ratio for preferred peptide conformation are shown in bold.</p>1<p>chemerin peptides-patent pending.</p

Chemerin peptide 4 exhibits pH- and salt-dependent lytic activity against <i>E.coli</i>.

<p>Chemically synthesized peptide 4 was tested against <i>E. coli</i> JM83 using cytoplasmic beta-galactosidase release assay. Bacteria were incubated with 10 µM peptide 4 for 0.5 h. Maximum (100%) lysis was set by the beta-galactosidase activity present in supernatants from bacteria treated with 1% Triton ×100. The results are expressed as the mean ± SD of three independent experiments.</p

Chemerin peptide 4 exhibits anti-microbial activity against variety of microbial species.

<p>The indicated microorganisms (<i>E. coli</i> ATCC 25922, <i>S. aureus</i> ATCC 29213, <i>P. aeruginosa</i> ATCC 27853 and <i>C. albicans</i> ATCC 90028) were tested for antimicrobial activity of chemerin peptide 4 or LL-37 (both at 100 µM), using RDA assay. The results are expressed as the mean ± SD of three independent experiments. * p<0.005 (Student's <i>t</i> test).</p

Keratinocyte-derived chemerin displays anti-bacterial activity.

<p>Paraffin sections of normal, shoulder skin biopsies (A) or chest keratinocytes grown in 3D culture for 1 week (B) were stained for chemerin or control rabbit Abs (red), with Hoechst counterstain to detect cell nuclei (blue). The slides were examined by fluoresce microscopy. Dotted lines in A indicate location of epidermis. Scale bar = 10 µm. Data are representative of three different donors. The antimicrobial activity of conditioned media from 3D cultures of keratinocytes (conditioned media) was tested against <i>E. coli</i> using the microtitre broth dilution assay (C). Where indicated, the conditioned media were first treated with sepharose-conjugated anti-chemerin Ab (anti-chemerin IgG), sepharose-conjugated control IgG (control IgG), or anti-chemerin Ab followed by recombinant chemerinS157 (chemS157) at 20 ng/ml. The results are expressed as the mean ± SD of four independent experiments. Statistically significant differences are indicated by asterisks (p≤0.01, Student's <i>t</i> test).</p

Chemerin levels in lysates isolated from epidermis of healthy donors.

<p>Chemerin levels in lysates isolated from epidermis of healthy donors.</p

Closure of cuteneous wounds in the HO-1^+/+ (WT), HO-1^+/− (HT), or HO-1^−/− (KO) C57BLxFVB mice.

<p>A – 3-month old animals. B – 6-month old animals. Each point represents individual animal (N = 4–5), lines connect the median values. Crossed points represent animals subjected to euthanasia. * P<0.05, ** P<0.01 in comparison to WT. C – representative pictures showing the wounds in 6-month old animals immediately after wounding and on day 10^th. Scale bar = 5 mm.</p

A – Closure of cutaneous wounds in the HO-1^+/+ wild type and HO-1^Tg mice.

<p>Each bar represents mean+SD. N = 10 animals per group. * P<0.05, ** P<0.01, *** P<0.001 in comparison to HO-1^+/+ mice. B – Representative pictures demonstrating CD31 staining of endothelial cells in the wounded skin (3 days after wounding) in the 3-month old mice of different genotypes. Scale bar = 100 µm. C – Number of vessels in wounded skin (3 days after wounding, CD31 staining) in the 3-month old mice of different genotypes. Each bar represents analysis of samples from 5–8 animals. Data are presented as mean+SD. * P<0.05 in comparison to HO-1^+/+ animals.</p

Effect of HO-1 transgene delivery on wounds.

<p>A – Effect of HO-1 transgene delivery on wound closure in the db/db diabetic mice. Adenoviral vectors (2.3×10⁷ IU in 100 µL of PBS) were injected subcutaneously near the wound immediately after injury. Control animal were injected with the same amount of AdGFP carriers. Each bar represents mean+SD; N = 5–8 animals per group. * P<0.05 in comparison to control, AdGFP treated mice. B – representative pictures showing blood vessels in the wounded skin of db/db mice injected with AdHO-1 or AdGFP vectors. CD31 staining of the skin cross-section. Scale bar = 100 µm. C – Number of vessels in wounded skin in the db/db mice injected with AdHO-1 or AdGFP, on the 3^rd and 14^th days after wounding. Analysis of specimens stained for CD31 to visualize endothelial cells. Each bar represents mean+SD values for 5–8 animals. * P<0.05 in comparison to control, AdGFP injected animals.</p