Dean Billheimer

PMID: 19622371;PMCID: PMC2927990;Abstract:

The northern leopard frog (Rana pipiens or Lithobates pipiens) is historically found in most of the provinces of Canada and the northern and southwest states of the United States. In the last 50 years, populations have suffered significant losses, especially in the western regions of the species range. Using a peptidomics approach, we show that the pattern of expressed antimicrobial skin peptides of frogs from three geographically separated populations are distinct, and we report the presence of four peptides (brevinin-1Pg, brevinin-1Pl, ranatuerin-2Pb, and ranatuerin-2Pc) that have not previously been found in skin secretions. The differences in expressed peptides reflect differences in the distribution of alleles for the newly described Brevinin1.1 locus in the three populations. When enriched peptide mixtures were tested for their ability to inhibit growth of the pathogenic amphibian chytrid (Batrachochytrium dendrobatidis), peptides from Minnesota or Vermont frogs were more effective that peptides from Michigan frogs. Four of the purified peptides were tested for their ability to inhibit growth of two bacterial pathogens (Aeromonas hydrophila and Staphylococcus epidermidis) and B. dendrobatidis. Three of the four were effective inhibitors of B. dendrobatidis and S. epidermidis, but none inhibited A. hydrophila. We interpret these differences in expression and activity of antimicrobial peptides as evidence to suggest that each population may have been selected to express a suite of peptides that reflects current and past encounters with skin microbes. © 2009 Elsevier Ltd. All rights reserved.

The objective was to use the United States Renal Data System (USRDS) to quantify the relationship between immunosuppressant therapy (IST) adherence and risk of graft failure among adult renal transplant recipients (RTRs). A secondary objective was to examine the relationship among select patient characteristics and IST adherence. The study sample included adult RTRs who: received primary transplant between January 1, 1999 and December 31, 2005; experienced graft survival for at least 12 months post-transplant and had at least 12 months of data in the USRDS; utilized Medicare coverage for IST; and were prescribed cyclosporine or tacrolimus. IST adherence was measured by medication possession ratio (MPR). Pearson chi-square tests were used to examine associations between patient characteristics and MPR quartiles. Cox proportional hazards regression was used to assess relationships among time to graft failure, MPR, and patient characteristics. Thirty-one thousand nine hundred and thirteen RTRs met inclusion criteria. Older age, female gender, white race, deceased donors, and tacrolimus were associated with greater adherence (p < 0.001). Cox proportional hazard modeling indicated greater adherence, white race, and having a living donor were significantly associated with longer graft survival (p < 0.05). Future prospective studies should further examine the clinical significance of IST nonadherence as it relates to graft failure.

Abstract:

Symbiotic microorganisms influence health and disease and may contribute to the innate immune defenses of amphibians. The mountain yellow-legged frog, Rana muscosa, is currently undergoing unprecedented population declines. One cause of recent declines is the pathogenic chytrid fungus, Batrachochytrium dendrobatidis (Bd). Skin swabs for detection of Bd, skin peptide secretions, and symbiotic skin bacteria were collected from 70 adult R. muscosa from two populations designated "Sixty Lake" and "Conness" in 2004-2005. The Conness population has persisted with the presence of Bd for at least 6 years whereas the Sixty Lake population is newly infected and declining. Of the frogs sampled at Conness, 67.5% were infected; whereas 96.7% of the Sixty Lake frogs were infected. Sixty Lake frogs were also more intensely infected than frogs at Conness. We isolated symbiotic bacteria that may contribute to immune defense. A significantly greater proportion of individuals with at least one anti-Bd bacterial species present were found at Conness (85%) than at Sixty Lake (62%). We observed no apparent differences in total skin peptides recovered; however, peptide mixtures from frogs at Sixty Lake showed better growth inhibitory activity against Bd than peptides from frogs at Conness. By MALDI-TOF MS analysis, there were no differences between the two populations in the previously described antimicrobial peptides (ranatuerin-2Ma, ranatuerin-2Mb, and temporin-1M). Antimicrobial skin peptides are only one factor in the resistance of R. muscosa to Bd infection. We suggest that symbiotic bacteria with the ability to persist in the presence of mucosal peptides may inhibit infection and colonization of the skin by Bd and increase the effectiveness of innate defense mechanisms in the skin. © 2007 Elsevier Ltd. All rights reserved.

Abstract:

Alpha-2 macroglobulin (A2M) functions as a universal protease inhibitor in serum and is capable of binding various cytokines and growth factors. In this study, we investigated if immunoaffinity enrichment and proteomic analysis of A2M protein complexes from human serum could improve detection of biologically relevant and novel candidate protein biomarkers in prostate cancer. Serum samples from six patients with androgen-independent, metastatic prostate cancer and six control patients without malignancy were analyzed by immunoaffinity enrichment of A2M protein complexes and MS identification of associated proteins. Known A2M substrates were reproducibly identified from patient serum in both cohorts, as well as proteins previously undetected in human serum. One example is heat shock protein 90 alpha (HSP90α), which was identified only in the serum of cancer patients in this study. Using an ELISA, the presence of HSP90α in human serum was validated on expanded test cohorts and found to exist in higher median serum concentrations in prostate cancer (n = 18) relative to control (n = 13) patients (median concentrations 50.7 versus 27.6 ng/mL, respectively, p = 0.001). Our results demonstrate the technical feasibility of this approach and support the analysis of A2M protein complexes for proteomic-based serum biomarker discovery. © 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Proteomic approaches identifying biomarkers have been applied to asthma to only a very limited extent.