Publications by authors named "Michael J Clare-Salzler"

58 Publications

Optimizing COVID-19 testing capabilities and clinical management using pathology informatics.

JAMIA Open 2020 Dec 5;3(4):523-529. Epub 2020 Dec 5.

Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, Florida, USA.

Coronavirus disease 2019, first reported in China in late 2019, has quickly spread across the world. The outbreak was declared a pandemic by the World Health Organization on March 11, 2020. Here, we describe our initial efforts at the University of Florida Health for processing of large numbers of tests, streamlining data collection, and reporting data for optimizing testing capabilities and superior clinical management. Specifically, we discuss clinical and pathology informatics workflows and informatics instruments which we designed to meet the unique challenges of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing. We hope these results benefit institutions preparing to implement SARS-CoV-2 testing.
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http://dx.doi.org/10.1093/jamiaopen/ooaa055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7717304PMC
December 2020

Influence of Vitamin D on Islet Autoimmunity and Beta-Cell Function in Type 1 Diabetes.

Nutrients 2019 Sep 11;11(9). Epub 2019 Sep 11.

Department of Systems Medicine, University of Rome "Tor Vergata", Via Montpellier 1, 00133 Rome, Italy.

Type 1 diabetes (T1D) is a chronic autoimmune disease leading to immune-mediated destruction of pancreatic beta cells, resulting in the need for insulin therapy. The incidence of T1D is increasing worldwide, thus prompting researchers to investigate novel immunomodulatory strategies to halt autoimmunity and modify disease progression. T1D is considered as a multifactorial disease, in which genetic predisposition and environmental factors interact to promote the triggering of autoimmune responses against beta cells. Over the last decades, it has become clear that vitamin D exerts anti-inflammatory and immunomodulatory effects, apart from its well-established role in the regulation of calcium homeostasis and bone metabolism. Importantly, the global incidence of vitamin D deficiency is also dramatically increasing and epidemiologic evidence suggests an involvement of vitamin D deficiency in T1D pathogenesis. Polymorphisms in genes critical for vitamin D metabolism have also been shown to modulate the risk of T1D. Moreover, several studies have investigated the role of vitamin D (in different doses and formulations) as a potential adjuvant immunomodulatory therapy in patients with new-onset and established T1D. This review aims to present the current knowledge on the immunomodulatory effects of vitamin D and summarize the clinical interventional studies investigating its use for prevention or treatment of T1D.
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http://dx.doi.org/10.3390/nu11092185DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6769474PMC
September 2019

Dual-Sized Microparticle System for Generating Suppressive Dendritic Cells Prevents and Reverses Type 1 Diabetes in the Nonobese Diabetic Mouse Model.

ACS Biomater Sci Eng 2019 May 26;5(5):2631-2646. Epub 2019 Mar 26.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, 1275 Center Drive, Gainesville, Florida 32611, United States.

Antigen specificity is a primary goal in developing curative therapies for autoimmune disease. Dendritic cells (DCs), as the most effective antigen presenting cells in the body, represent a key target to mediate restoration of antigen-specific immune regulation. Here, we describe an injectable, dual-sized microparticle (MP) approach that employs phagocytosable ∼1 μm and nonphagocytosable ∼30 μm MPs to deliver tolerance-promoting factors both intracellularly and extracellularly, as well as the type 1 diabetes autoantigen, insulin, to DCs for reprogramming of immune responses and remediation of autoimmunity. This poly(lactic--glycolic acid) (PLGA) MP system prevented diabetes onset in 60% of nonobese diabetic (NOD) mice when administered subcutaneously in 8 week old mice. Prevention of disease was dependent upon antigen inclusion and required encapsulation of factors in MPs. Moreover, administration of this "suppressive-vaccine" boosted pancreatic lymph node and splenic regulatory T cells (Tregs), upregulated PD-1 on CD4 and CD8 T cells, and reversed hyperglycemia for up to 100 days in recent-onset NOD mice. Our results demonstrate that a MP-based platform can reeducate the immune system in an antigen-specific manner, augment immunomodulation compared to soluble administration of drugs, and provide a promising alternative to systemic immunosuppression for autoimmunity.
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http://dx.doi.org/10.1021/acsbiomaterials.9b00332DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6518351PMC
May 2019

CYP2D6-guided opioid therapy improves pain control in CYP2D6 intermediate and poor metabolizers: a pragmatic clinical trial.

Genet Med 2019 08 23;21(8):1842-1850. Epub 2019 Jan 23.

Department of Pharmacotherapy and Translational Research, College of Pharmacy, University of Florida, Gainesville, FL, USA.

Purpose: CYP2D6 bioactivates codeine and tramadol, with intermediate and poor metabolizers (IMs and PMs) expected to have impaired analgesia. This pragmatic proof-of-concept trial tested the effects of CYP2D6-guided opioid prescribing on pain control.

Methods: Participants with chronic pain (94% on an opioid) from seven clinics were enrolled into CYP2D6-guided (n = 235) or usual care (n = 135) arms using a cluster design. CYP2D6 phenotypes were assigned based on genotype and CYP2D6 inhibitor use, with recommendations for opioid prescribing made in the CYP2D6-guided arm. Pain was assessed at baseline and 3 months using PROMIS measures.

Results: On stepwise multiple linear regression, the primary outcome of composite pain intensity (composite of current pain and worst and average pain in the past week) among IM/PMs initially prescribed tramadol/codeine (n = 45) had greater improvement in the CYP2D6-guided versus usual care arm (-1.01 ± 1.59 vs. -0.40 ± 1.20; adj P = 0.016); 24% of CYP2D6-guided versus 0% of usual care participants reported ≥30% (clinically meaningful) reduction in the composite outcome. In contrast, among normal metabolizers prescribed tramadol or codeine at baseline, there was no difference in the change in composite pain intensity at 3 months between CYP2D6-guided (-0.61 ± 1.39) and usual care (-0.54 ± 1.69) groups (adj P = 0.540).

Conclusion: These data support the potential benefits of CYP2D6-guided pain management.
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http://dx.doi.org/10.1038/s41436-018-0431-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6650382PMC
August 2019

Immature Dendritic Cell Therapy Confers Durable Immune Modulation in an Antigen-Dependent and Antigen-Independent Manner in Nonobese Diabetic Mice.

J Immunol Res 2018 14;2018:5463879. Epub 2018 Feb 14.

Department of Pathology, Immunology and Laboratory Medicine, Center for Immunology and Transplantation, University of Florida, Gainesville, FL 32610, USA.

Dendritic cell (DC) immunotherapy has been effective for prevention of type 1 diabetes (T1D) in NOD mice but fails to protect if initiated after active autoimmunity. As autoreactivity expands inter- and intramolecularly during disease progression, we investigated whether DCs unpulsed or pulsed with cell antigenic dominant determinants (DD), subdominant determinants (SD), and ignored determinants (ID) could prevent T1D in mice with advanced insulitis. We found that diabetes was significantly delayed by DC therapy. Of interest, DCs pulsed with SD or ID appeared to provide better protection. T lymphocytes from DC-treated mice acquired spontaneous proliferating capability during culture, which could be largely eliminated by IL-2 neutralizing antibodies. This trend maintained even 29 weeks after discontinuing DC therapy and appeared antigen-independent. Furthermore, CD4+Foxp3+ T regulatory cells (Tregs) from DC-treated mice proliferated more actively compared to the controls, and Tregs from DC-treated mice showed significantly enhanced immunosuppressive activities in contrast to those from the controls. Our study demonstrates that DC therapy leads to long-lasting immunomodulatory effects in an antigen-dependent and antigen-independent manner and provides evidence for peptide-based intervention during a clinically relevant window to guide DC-based immunotherapy for autoimmune diabetes.
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http://dx.doi.org/10.1155/2018/5463879DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5832131PMC
September 2018

Multisite Investigation of Outcomes With Implementation of CYP2C19 Genotype-Guided Antiplatelet Therapy After Percutaneous Coronary Intervention.

JACC Cardiovasc Interv 2018 01 1;11(2):181-191. Epub 2017 Nov 1.

Department of Medicine, University of Maryland, Baltimore, Maryland.

Objectives: This multicenter pragmatic investigation assessed outcomes following clinical implementation of CYP2C19 genotype-guided antiplatelet therapy after percutaneous coronary intervention (PCI).

Background: CYP2C19 loss-of-function alleles impair clopidogrel effectiveness after PCI.

Methods: After clinical genotyping, each institution recommended alternative antiplatelet therapy (prasugrel, ticagrelor) in PCI patients with a loss-of-function allele. Major adverse cardiovascular events (defined as myocardial infarction, stroke, or death) within 12 months of PCI were compared between patients with a loss-of-function allele prescribed clopidogrel versus alternative therapy. Risk was also compared between patients without a loss-of-function allele and loss-of-function allele carriers prescribed alternative therapy. Cox regression was performed, adjusting for group differences with inverse probability of treatment weights.

Results: Among 1,815 patients, 572 (31.5%) had a loss-of-function allele. The risk for major adverse cardiovascular events was significantly higher in patients with a loss-of-function allele prescribed clopidogrel versus alternative therapy (23.4 vs. 8.7 per 100 patient-years; adjusted hazard ratio: 2.26; 95% confidence interval: 1.18 to 4.32; p = 0.013). Similar results were observed among 1,210 patients with acute coronary syndromes at the time of PCI (adjusted hazard ratio: 2.87; 95% confidence interval: 1.35 to 6.09; p = 0.013). There was no difference in major adverse cardiovascular events between patients without a loss-of-function allele and loss-of-function allele carriers prescribed alternative therapy (adjusted hazard ratio: 1.14; 95% confidence interval: 0.69 to 1.88; p = 0.60).

Conclusions: These data from real-world observations demonstrate a higher risk for cardiovascular events in patients with a CYP2C19 loss-of-function allele if clopidogrel versus alternative therapy is prescribed. A future randomized study of genotype-guided antiplatelet therapy may be of value.
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http://dx.doi.org/10.1016/j.jcin.2017.07.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5775044PMC
January 2018

Type 1 Diabetes and Type 1 Interferonopathies: Localization of a Type 1 Common Thread of Virus Infection in the Pancreas.

EBioMedicine 2017 Aug 21;22:10-17. Epub 2017 Jun 21.

Department of Microbiology and Immunology, Infection, Inflammation, and Immunity (I3) Research Group, Life Sciences Institute, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada. Electronic address:

Type 1 diabetes (T1D) has been associated with both genetic and environmental factors. Increasing incidence of T1D worldwide is prompting researchers to adopt different approaches to explain the biology of T1D, beyond the presence and activity of autoreactive lymphocytes. In this review, we propose inflammatory pathways as triggers for T1D. Within the scope of those inflammatory pathways and in understanding the pathogenesis of disease, we suggest that viruses, in particular Coxsackieviruses, act by causing a type 1 interferonopathy within the pancreas and the microenvironment of the islet. As such, this connection and common thread represents an exciting platform for the development of new diagnostic, treatment and/or prevention options.
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http://dx.doi.org/10.1016/j.ebiom.2017.06.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5552106PMC
August 2017

Institutional profile: University of Florida Health Personalized Medicine Program.

Pharmacogenomics 2017 Apr 27;18(5):421-426. Epub 2017 Mar 27.

Department of Pharmacotherapy & Translational Research, College of Pharmacy, University of Florida, Gainesville, FL 32610, USA.

The University of Florida (UF) Health Personalized Medicine Program launched in 2012 with CYP2C19 genotyping for clopidogrel response at UF Health Shands Hospital. We have since expanded CYP2C19 genotyping to UF Health Jacksonville and established the infrastructure at UF Health to support clinical implementation for five additional gene-drug pairs: TPMT-thiopurines, IFNL3 (IL28B)-PEG IFN-α-based regimens, CYP2D6-opioids, CYP2D6/CYP2C19-antidepressants and CYP2C19-proton pump inhibitors. We are contributing to the evidence based on outcomes with genotype-guided therapy through pragmatic studies of our clinical implementations. In addition, we have developed a broad array of educational programs for providers, trainees and students that incorporate personal genotype evaluation to enhance participant learning.
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http://dx.doi.org/10.2217/pgs-2017-0028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5558508PMC
April 2017

Macrophage integrins modulate response to ultra-high molecular weight polyethylene particles and direct particle-induced osteolysis.

Biomaterials 2017 01 27;115:128-140. Epub 2016 Oct 27.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611, USA. Electronic address:

Aseptic loosening due to peri-prosthetic osteolysis is one of the primary causes for failure of artificial joint replacements. Implant-derived wear particles, often ultra-high molecular weight polyethylene (UHMWPE) microparticles, initiate an inflammatory cascade upon phagocytosis by macrophages, which leads to osteoclast recruitment and activation, ultimately resulting in osteolysis. Investigation into integrin receptors, involved in cellular interactions with biomaterial-adsorbed adhesive proteins, is of interest to understand and modulate inflammatory processes. In this work, we investigate the role of macrophage integrins Mac-1 and RGD-binding integrins in response to UHMWPE wear particles. Using integrin knockout mice as well as integrin blocking techniques, reduction in macrophage phagocytosis and inflammatory cytokine secretion is demonstrated when these receptors are either absent or blocked. Along this line, various opsonizing proteins are shown to differentially modulate microparticle uptake and macrophage secretion of inflammatory cytokines. Furthermore, using a calvarial osteolysis model it is demonstrated that both Mac-1 integrin and RGD-binding integrins modulate the particle induced osteolysis response to UHMWPE microparticles, with a 40% decrease in the area of osteolysis by the absence or blocking of these integrins, in vivo. Altogether, these findings indicate Mac-1 and RGD-binding integrins are involved in macrophage-directed inflammatory responses to UHMWPE and may serve as therapeutic targets to mitigate wear particle induced peri-prosthetic osteolysis for improved performance of implanted joints.
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http://dx.doi.org/10.1016/j.biomaterials.2016.10.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5431751PMC
January 2017

Omega-3 fatty acids are associated with a lower prevalence of autoantibodies in shared epitope-positive subjects at risk for rheumatoid arthritis.

Ann Rheum Dis 2017 Jan 17;76(1):147-152. Epub 2016 May 17.

Department of Epidemiology, Colorado School of Public Health, Aurora, Colorado, USA.

Objectives: Previously, we found that omega-3 fatty acids (n-3 FAs) were inversely associated with anti-cyclic citrullinated peptide (anti-CCP) positivity in participants at risk for future rheumatoid arthritis (RA). We investigated whether n-3 FAs were also associated with rheumatoid factor (RF) positivity and whether these associations were modified by shared epitope (SE) positivity.

Methods: The Studies of the Etiology of RA (SERA) cohort includes RA-free participants who are at increased risk for RA. We conducted a nested case-control study (n=136) to determine the association between RF and anti-CCP2 positivity and n-3 FA percentage in erythrocyte membranes (n-3 FA% in red blood cells (RBCs)). Additionally, in the baseline visit of the SERA cohort (n=2166), we evaluated the association between reported n-3 FA supplement use and prevalence of RF and anti-CCP2. We assessed SE positivity as an effect modifier.

Results: In the case-control study, increasing n-3 FA% in RBCs was inversely associated with RF positivity in SE-positive participants (OR 0.27, 95% CI 0.10 to 0.79), but not SE-negative participants. Similar associations were seen with anti-CCP positivity in SE-positive participants (OR 0.42, 95% CI 0.20 to 0.89), but not SE-negative participants. In the SERA cohort at baseline, n-3 FA supplement use was associated with a lower prevalence of RF positivity in SE-positive participants (OR 0.32, 95% CI 0.12 to 0.82), but not SE-negative participants; similar but non-significant trends were observed with anti-CCP2.

Conclusions: The potential protective effect of n-3 FAs on RA-related autoimmunity may be most pronounced in those who exhibit HLA class II genetic susceptibility to RA.
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http://dx.doi.org/10.1136/annrheumdis-2016-209154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5371398PMC
January 2017

Discovery, Total Synthesis and Key Structural Elements for the Immunosuppressive Activity of Cocosolide, a Symmetrical Glycosylated Macrolide Dimer from Marine Cyanobacteria.

Chemistry 2016 06 3;22(24):8158-66. Epub 2016 May 3.

Department of Medicinal Chemistry and Center for Natural Products, Drug Discovery and Development (CNPD3), University of Florida, Gainesville, Florida, 32610, USA.

A new dimeric macrolide xylopyranoside, cocosolide (1), was isolated from the marine cyanobacterium preliminarily identified as Symploca sp. from Guam. The structure was determined by a combination of NMR spectroscopy, HRMS, X-ray diffraction studies and Mosher's analysis of the base hydrolysis product. Its carbon skeleton closely resembles that of clavosolides A-D isolated from the sponge Myriastra clavosa, for which no bioactivity is known. We performed the first total synthesis of cocosolide (1) along with its [α,α]-anomer (26) and macrocyclic core (28), thus leading to the confirmation of the structure of natural 1. The convergent synthesis featured Wadsworth-Emmons cyclopropanation, Sakurai annulation, Yamaguchi macrocyclization/dimerization reaction, α-selective glycosidation and β-selective glycosidation. Compounds 1 and 26 potently inhibited IL-2 production in both T-cell receptor dependent and independent manners. Full activity requires the presence of the sugar moiety as well as the intact dimeric structure. Cocosolide also suppressed the proliferation of anti-CD3-stimulated T-cells in a dose-dependent manner.
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http://dx.doi.org/10.1002/chem.201600674DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5224530PMC
June 2016

A cell-based microarray to investigate combinatorial effects of microparticle-encapsulated adjuvants on dendritic cell activation.

J Mater Chem B 2016 Mar 30;4(9):1672-1685. Epub 2015 Sep 30.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, 130 BME/PO Box 116131, Gainesville, Florida, 32611-6131, USA.

Experimental vaccine adjuvants are being designed to target specific toll-like receptors (TLRs) alone or in combination, expressed by antigen presenting cells, notably dendritic cells (DCs). There is a need for high-content screening (HCS) platforms to explore how DC activation is affected by adjuvant combinations. Presented is a cell-based microarray approach, "immunoarray", exposing DCs to a large number of adjuvant combinations. Microparticles encapsulating TLR ligands are printed onto arrays in a range of doses for each ligand, in all possible dose combinations. Dendritic cells are then co-localized with physisorbed microparticles on the immunoarray, adherent to isolated islands surrounded by a non-fouling background, and DC activation is quantified. Delivery of individual TLR ligands was capable of eliciting high levels of specific DC activation markers. For example, either TLR9 ligand, CpG, or TLR3 ligand, poly I:C, was capable of inducing among the highest 10% expression levels of CD86. In contrast, MHC-II expression in response to TLR4 agonist MPLA was among the highest, whereas either MPLA or poly I:C, was capable of producing among the highest levels of CCR7 expression, as well as inflammatory cytokine IL-12. However, in order to produce robust responses across all activation markers, adjuvant combinations were required, and combinations were more represented among the high responders. The immunoarray also enables investigation of interactions between adjuvants, and each TLR ligand suggested antagonism to other ligands, for various markers. Altogether, this work demonstrates feasibility of the immunoarray platform to screen microparticle-encapsulated adjuvant combinations for the development of improved and personalized vaccines.
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http://dx.doi.org/10.1039/C5TB01754HDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4790840PMC
March 2016

Csf2 and Ptgs2 Epigenetic Dysregulation in Diabetes-prone Bicongenic B6.NODC11bxC1tb Mice.

Genet Epigenet 2015 11;7:5-17. Epub 2015 Oct 11.

Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, FL, USA. ; Sanford-Burnham Medical Research Institute, Diabetes and Obesity Center, Lake Nona-Orlando, FL, USA. ; Florida Hospital Cancer Institute, Orlando, FL, USA.

In Type 1 diabetic (T1D) human monocytes, STAT5 aberrantly binds to epigenetic regulatory sites of two proinflammatory genes, CSF2 (encoding granulocyte-macrophage colony-stimulating factor) and PTGS2 (encoding prostaglandin synthase 2/cyclooxygenase 2). Bicongenic B6.NOD C11bxC1tb mice re-create this phenotype of T1D monocytes with only two nonobese diabetic (NOD) Idd subloci (130.8 Mb-149.7 Mb, of Idd5 on Chr 1 and 32.08-53.85 Mb of Idd4.3 on Chr11) on C57BL/6 genetic background. These two Idd loci interact through STAT5 binding at upstream regulatory regions affecting Csf2 (Chr 11) and Ptgs2 (Chr 1) expression. B6.NODC11bxC1tb mice exhibited hyperglycemia and immune destruction of pancreatic islets between 8 and 30 weeks of age, with 12%-22% penetrance. Thus, B6.NODC11bxC1tb mice embody NOD epigenetic dysregulation of gene expression in myeloid cells, and this defect appears to be sufficient to impart genetic susceptibility to diabetes in an otherwise genetically nonautoimmune mouse.
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http://dx.doi.org/10.4137/GEG.S29696DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4603573PMC
October 2015

Effects of type 1 diabetes-associated IFIH1 polymorphisms on MDA5 function and expression.

Curr Diab Rep 2015 Nov;15(11):96

Department of Pathology, Immunology and Laboratory Medicine, University of Florida, 2033 Mowry Rd., P.O. Box 103633, Gainesville, FL, 32611, USA.

Recent evidence has highlighted the role of the innate immune system in type 1 diabetes (T1D) pathogenesis. Specifically, aberrant activation of the interferon response prior to seroconversion of T1D-associated autoantibodies supports a role for the interferon response as a precipitating event toward activation of autoimmunity. Melanoma differentiation-associated protein 5 (MDA5), encoded by IFIH1, mediates the innate immune system's interferon response to certain viral species that form double-stranded RNA (dsRNA), the MDA5 ligand, during their life cycle. Extensive research has associated single nucleotide polymorphisms (SNPs) within the coding region of IFIH1 with T1D. This review discusses the different risk and protective IFIH1 alleles in the context of recent structural and functional analysis that relate to MDA5 regulation of interferon responses. These studies have provided a functional hypothesis for IFIH1 T1D-associated SNPs' effects on MDA5-mediated interferon responses as well as supporting the genome-wide association (GWA) studies that first associated IFIH1 with T1D.
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http://dx.doi.org/10.1007/s11892-015-0656-8DOI Listing
November 2015

Lower omega-3 fatty acids are associated with the presence of anti-cyclic citrullinated peptide autoantibodies in a population at risk for future rheumatoid arthritis: a nested case-control study.

Rheumatology (Oxford) 2016 Feb 13;55(2):367-76. Epub 2015 Sep 13.

Colorado School of Public Health, Department of Epidemiology, Aurora,

Objective: The aim of this study was to investigate omega-3 fatty acid (FA) supplement use and omega-3 FAs in erythrocyte membranes [omega-3 FA % in erythrocyte membranes (RBC)] and their association with anti-CCP autoantibodies in a population without RA, but who are at genetic risk for RA.

Methods: The multicentre Studies of the Etiology of RA (SERA) cohort includes RA-free subjects who are first-degree relatives of RA probands or are enriched with the HLA-DR4 allele. In a nested case-control study, 30 SERA cases were identified who were anti-CCP2 antibody positive. We further identified 47 autoantibody negative controls, frequency matched to cases on age at study visit, sex, race and study site. Anti-CCP2 status, self-reported omega-3 FA supplement use and omega-3 FA % in RBCs were obtained from a single visit.

Results: Anti-CCP2 positive cases were less likely than controls to report omega-3 FA supplement use (odds ratio: 0.14; 95% CI 0.03, 0.68). In addition, the likelihood of anti-CCP2 positivity was inversely associated with total omega-3 FA % in RBCs (odds ratio: 0.47; 95% CI 0.24, 0.92, for a s.d. increase).

Conclusion: The inverse association between anti-CCP2 positivity and self-reported omega-3 FA supplement use and omega-3 FA % in RBCs suggests that omega-3 FAs may protect against the development of RA-related autoimmunity in pre-clinical RA.
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http://dx.doi.org/10.1093/rheumatology/kev266DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5009416PMC
February 2016

A combination dual-sized microparticle system modulates dendritic cells and prevents type 1 diabetes in prediabetic NOD mice.

Clin Immunol 2015 Sep 2;160(1):90-102. Epub 2015 Apr 2.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611, USA; Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, FL 32611, USA. Electronic address:

We developed a novel poly(lactic-co-glycolic acid)-based, microparticle (MP) system providing concurrent delivery of multiple encapsulated immuno-suppressive factors and antigen, for in vivo conditioning of dendritic cells (DCs) toward a tolerance promoting pathway. Subcutaneous administration prevents onset of type 1 diabetes (T1D) in NOD mice. Two MP sizes were made: phagocytosable MPs were fabricated encapsulating vitamin D3 or insulin B(9-23) peptide, while unphagocytosable MPs were fabricated encapsulating TGF-β1 or GM-CSF. The combination of Vit D3/TGF-β1 MPs confers an immature and LPS activation-resistant phenotype to DCs, and MP-delivered antigen is efficiently and functionally presented. Notably, two subcutaneous injections into 4week old NOD mice using the combination of MPs encapsulating Vit D3, Ins B, TGF-β1 and GM-CSF protected 40% of mice from T1D development, significant in comparison to the control. This work represents one of the first applications of a biomaterial-based, MP vaccine system to successfully prevent autoimmune diabetes.
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http://dx.doi.org/10.1016/j.clim.2015.03.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4554803PMC
September 2015

Increased IFN-α-producing plasmacytoid dendritic cells (pDCs) in human Th1-mediated type 1 diabetes: pDCs augment Th1 responses through IFN-α production.

J Immunol 2014 Aug 27;193(3):1024-34. Epub 2014 Jun 27.

Department of Pathology, Immunology, and Laboratory Medicine, University of Florida, Gainesville, FL 32610;

Increasing evidence suggests that type 1 IFN (IFN-αβ) is associated with pathogenesis of Th1-mediated type 1 diabetes (T1D). A major source of IFN-αβ is plasmacytoid dendritic cells (pDCs). In this study, we analyzed peripheral blood pDC numbers and functions in at-risk, new-onset, and established T1D patients and controls. We found that subjects at risk for T1D and new-onset and established T1D subjects possessed significantly increased pDCs but similar number of myeloid DCs when compared with controls. pDC numbers were not affected by age in T1D subjects but declined with increasing age in control subjects. It was demonstrated that IFN-α production by PBMCs stimulated with influenza viruses was significantly higher in T1D subjects than in controls, and IFN-α production was correlated with pDC numbers in PBMCs. Of interest, only T1D-associated Coxsackievirus serotype B4 but not B3 induced majority of T1D PBMCs to produce IFN-α, which was confirmed to be secreted by pDCs. Finally, in vitro studies demonstrated IFN-α produced by pDCs augmented Th1 responses, with significantly greater IFN-γ-producing CD4(+) T cells from T1D subjects. These findings indicate that increased pDCs and their IFN-αβ production may be associated with this Th1-mediated autoimmune disease, especially under certain viral infections linked to T1D pathogenesis.
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http://dx.doi.org/10.4049/jimmunol.1303230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4500116PMC
August 2014

C-Abl inhibitor imatinib enhances insulin production by β cells: c-Abl negatively regulates insulin production via interfering with the expression of NKx2.2 and GLUT-2.

PLoS One 2014 16;9(5):e97694. Epub 2014 May 16.

Department of Pathology, Immunology and Laboratory Medicine, Diabetes Center of Excellence, University of Florida, Gainesville, Florida, United States of America.

Chronic myelogenous leukemia patients treated with tyrosine kinase inhibitor, Imatinib, were shown to have increased serum levels of C-peptide. Imatinib specifically inhibits the tyrosine kinase, c-Abl. However, the mechanism of how Imatinib treatment can lead to increased insulin level is unclear. Specifically, there is little investigation into whether Imatinib directly affects β cells to promote insulin production. In this study, we showed that Imatinib significantly induced insulin expression in both glucose-stimulated and resting β cells. In line with this finding, c-Abl knockdown by siRNA and overexpression of c-Abl markedly enhanced and inhibited insulin expression in β cells, respectively. Unexpectedly, high concentrations of glucose significantly induced c-Abl expression, suggesting c-Abl may play a role in balancing insulin production during glucose stimulation. Further studies demonstrated that c-Abl inhibition did not affect the major insulin gene transcription factor, pancreatic and duodenal homeobox-1 (PDX-1) expression. Of interest, inhibition of c-Abl enhanced NKx2.2 and overexpression of c-Abl in β cells markedly down-regulated NKx2.2, which is a positive regulator for insulin gene expression. Additionally, we found that c-Abl inhibition significantly enhanced the expression of glucose transporter GLUT2 on β cells. Our study demonstrates a previously unrecognized mechanism that controls insulin expression through c-Abl-regulated NKx2.2 and GLUT2. Therapeutic targeting β cell c-Abl could be employed in the treatment of diabetes or β cell tumor, insulinoma.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0097694PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4023982PMC
August 2015

Combinatorial delivery of immunosuppressive factors to dendritic cells using dual-sized microspheres.

J Mater Chem B 2014 May;2(17):2562-2574

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611.

Microparticulate systems are beginning to show promise for delivery of modulatory agents for immunotherapeutic applications which modulate dendritic cell (DC) functions. Co-administration of multiple factors is an emerging theme in immune modulation which may prove beneficial in this setting. Herein, we demonstrate that localized, controlled delivery of multiple factors can be accomplished through poly (lactic-co-glycolic acid) (PLGA) microparticle systems fabricated in two size classes of phagocytosable and unphagocytosable microparticles (MPs). The immunosuppressive ability of combinatorial multi-factor dual MP systems was evaluated by investigating effects on DC maturation, DC resistance to LPS-mediated maturation and proliferation of allogeneic T cells in a mixed lymphocyte reaction. Phagocytosable MPs (~2 μm) were fabricated encapsulating either rapamycin (RAPA) or all-trans retinoic acid (RA), and unphagocytosable MPs (~30 μm) were fabricated encapsulating either transforming growth factor beta-1 (TGF-β1) or interleukin-10 (IL-10). Combinations of these MP classes reduced expression of stimulatory/costimulatory molecules (MHC-II, CD80 and CD86) in comparison to iDC and soluble controls, but not necessarily to single factor MPs. Dual MP-treated DCs resisted LPS-mediated activation, in a manner driven by the single factor phagocytosable MPs used. Dendritic cells treated with dual MP systems suppressed allogeneic T cell proliferation, generally demonstrating greater suppression by combination MPs than single factor formulations, particularly for the RA/IL-10 MPs. This work demonstrates feasibility of simultaneous targeted delivery of immunomodulatory factors to cell surface receptors and intracellular locations, and indicates that a combinatorial approach can boost immunoregulatory responses for therapeutic application in autoimmunity and transplantation.
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http://dx.doi.org/10.1039/C3TB21460EDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4000038PMC
May 2014

Anti-CD3 antibody treatment induces hypoglycemia and super tolerance to glucose challenge in mice through enhancing glucose consumption by activated lymphocytes.

J Immunol Res 2014 11;2014:326708. Epub 2014 Feb 11.

Department of Pathology, Immunology and Laboratory Medicine, University of Florida College of Medicine, Gainesville, FL 32610, USA.

Anti-CD3 antibody has been employed for various immune-mediated disorders. However, whether anti-CD3 administration leads to rapid metabolic alternation has not been well investigated. In the current study, we studied how anti-CD3 treatment affected blood glucose levels in mice. We found that anti-CD3 treatment induced immediate reduction of blood glucose after administration. Furthermore, a single dose of anti-CD3 treatment corrected hyperglycemia in all nonobese diabetic mice with recently diagnosed diabetes. This glucose-lowering effect was not attributable to major T cell produced cytokines. Of interest, when tested in a normal strain of mice (C57BL/6), the serum levels of C-peptide in anti-CD3 treated animals were significantly lower than control mice. Paradoxically, anti-CD3 treated animals were highly tolerant to exogenous glucose challenge. Additionally, we found that anti-CD3 treatment significantly induced activation of T and B cells in vitro and in vivo. Further studies demonstrated that anti-CD3 treatment lowered the glucose levels in T cell culture media and increased the intracellular transportation of 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2 deoxyglucose (2-NBDG) particularly in activated T and B cells. In addition, injection of anti-CD3 antibodies induced enhanced levels of Glut1 expression in spleen cells. This study suggests that anti-CD3 therapy-induced hypoglycemia likely results from increased glucose transportation and consumption by the activated lymphocytes.
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http://dx.doi.org/10.1155/2014/326708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3987876PMC
January 2015

Characterization of Bone Marrow-Derived Dendritic Cells Developed in Serum-Free Media and their Ability to Prevent Type 1 Diabetes in Nonobese Diabetic Mice.

J Blood Disord Transfus 2014 Apr 22;5(4). Epub 2014 Mar 22.

Department of Pathology, Immunology and Laboratory Medicine, University of Florida, USA.

Dendritic cells (DC) have been investigated as a cell-based therapy for Type 1 Diabetes (T1D). BM-DC expanded with GM-CSF and IL-4 is typically cultured with fetal bovine serum (FBS). The effect of FBS on NOD BM-DC has not been extensively studied. In the present study we compare BM-DC generated in serum-free culture media (X-VIVO20; FBS-) with BM-DC generated in media containing 10% FBS (RPMI1640/10%FBS; FBS+). We show that FBS- BM-DC display a phenotype and cytokine-producing profile distinct from FBS+ BMDC. Additionally, compared to FBS+ BM-DC, we show evidence of an altered T cell response induced by FBS- BM-DC. Finally, we demonstrate that only FBS- BM-DC prevent the onset of T1D and induce increased levels of CD4Foxp3 regulatory T cells as well as a long-lasting β cell-specific T cell response. This study indicates that serum-free media generates a more tolerogenic BM-DC capable of preventing T1D in the NOD mice.
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http://dx.doi.org/10.4172/2155-9864.1000206DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4712959PMC
April 2014

Clinical pharmacogenetics implementation: approaches, successes, and challenges.

Am J Med Genet C Semin Med Genet 2014 Mar 10;166C(1):56-67. Epub 2014 Mar 10.

Current challenges exist to widespread clinical implementation of genomic medicine and pharmacogenetics. The University of Florida (UF) Health Personalized Medicine Program (PMP) is a pharmacist-led, multidisciplinary initiative created in 2011 within the UF Clinical Translational Science Institute. Initial efforts focused on pharmacogenetics, with long-term goals to include expansion to disease-risk prediction and disease stratification. Herein we describe the processes for development of the program, the challenges that were encountered and the clinical acceptance by clinicians of the genomic medicine implementation. The initial clinical implementation of the UF PMP began in June 2012 and targeted clopidogrel use and the CYP2C19 genotype in patients undergoing left heart catheterization and percutaneous-coronary intervention (PCI). After 1 year, 1,097 patients undergoing left heart catheterization were genotyped preemptively, and 291 of those underwent subsequent PCI. Genotype results were reported to the medical record for 100% of genotyped patients. Eighty patients who underwent PCI had an actionable genotype, with drug therapy changes implemented in 56 individuals. Average turnaround time from blood draw to genotype result entry in the medical record was 3.5 business days. Seven different third party payors, including Medicare, reimbursed for the test during the first month of billing, with an 85% reimbursement rate for outpatient claims that were submitted in the first month. These data highlight multiple levels of success in clinical implementation of genomic medicine.
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http://dx.doi.org/10.1002/ajmg.c.31390DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4076109PMC
March 2014

Integrin-directed modulation of macrophage responses to biomaterials.

Biomaterials 2014 Apr 24;35(11):3504-15. Epub 2014 Jan 24.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611, USA. Electronic address:

Macrophages are the primary mediator of chronic inflammatory responses to implanted biomaterials, in cases when the material is either in particulate or bulk form. Chronic inflammation limits the performance and functional life of numerous implanted medical devices, and modulating macrophage interactions with biomaterials to mitigate this response would be beneficial. The integrin family of cell surface receptors mediates cell adhesion through binding to adhesive proteins nonspecifically adsorbed onto biomaterial surfaces. In this work, the roles of integrin Mac-1 (αMβ2) and RGD-binding integrins were investigated using model systems for both particulate and bulk biomaterials. Specifically, the macrophage functions of phagocytosis and inflammatory cytokine secretion in response to a model particulate material, polystyrene microparticles were investigated. Opsonizing proteins modulated microparticle uptake, and integrin Mac-1 and RGD-binding integrins were found to control microparticle uptake in an opsonin-dependent manner. The presence of adsorbed endotoxin did not affect microparticle uptake levels, but was required for the production of inflammatory cytokines in response to microparticles. Furthermore, it was demonstrated that integrin Mac-1 and RGD-binding integrins influence the in vivo foreign body response to a bulk biomaterial, subcutaneously implanted polyethylene terephthalate. A thinner foreign body capsule was formed when integrin Mac-1 was absent (~30% thinner) or when RGD-binding integrins were blocked by controlled release of a blocking peptide (~45% thinner). These findings indicate integrin Mac-1 and RGD-binding integrins are involved and may serve as therapeutic targets to mitigate macrophage inflammatory responses to both particulate and bulk biomaterials.
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http://dx.doi.org/10.1016/j.biomaterials.2014.01.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970928PMC
April 2014

It's time to bring dendritic cell therapy to type 1 diabetes.

Diabetes 2014 Jan;63(1):20-30

Department of Medicine, Columbia Center for Translational Immunology and Naomi Berrie Diabetes Center, Columbia University Medical Center, New York, NY.

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http://dx.doi.org/10.2337/db13-0886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3968436PMC
January 2014

Infection and persistence of erythrovirus B19 in benign and cancerous thyroid tissues.

J Med Virol 2014 Sep 21;86(9):1614-20. Epub 2013 Nov 21.

Departments of Psychiatry and Pediatrics, University of Florida College of Medicine, Gainesville, Florida.

Human erythrovirus B19 (EVB19) is a small, pathogenic DNA virus that has been associated with a wide range of illnesses. The primary site of replication is in bone marrow-derived erythroid progenitor cells, but EVB19 DNA has been detected in a wide range of organs. Recently, studies have linked EVB19 to thyroid cancers and other thyroid diseases. Previous studies from multiple laboratories have detected EVB19 capsid proteins in Graves' disease, Hashimoto's thyroiditis, and thyroid cancer tissues. Data on viral gene expression and mechanism of infection in the thyroid are lacking. To investigate EVB19 infection and persistence in the thyroid, previously archived adult and pediatric tissue sections were examined for EVB19 DNA, RNA, and capsid proteins, as well as EVB19 receptor P-antigen and co-receptor α5β1 integrin. EVB19 DNA and protein were detected in a majority of tissues examined (87% and 68%, respectively). Detection was similar in adult and pediatric samples. Quantification of viral genomes revealed no significant difference in the amount of viral DNA in benign, cancerous, or metastatic thyroid tissues. EVB19 capsid RNA was detected in 67% of the tissues examined, confirming at least low-level viral gene expression. Immunohistochemical staining for P-antigen and α5β1 detected the receptor and co-receptor most frequently on normal thyroid epithelial cells. EVB19 capsid staining could be detected in tumors lacking viral receptors. These results suggest that normal thyroid epithelial cells are the initial target for EVB19 infection in the thyroid and allow for continued persistence in both normal and cancerous thyroid tissues.
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http://dx.doi.org/10.1002/jmv.23852DOI Listing
September 2014

Persistent STAT5 phosphorylation and epigenetic dysregulation of GM-CSF and PGS2/COX2 expression in Type 1 diabetic human monocytes.

PLoS One 2013 18;8(10):e76919. Epub 2013 Oct 18.

Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, Florida, United States of America.

STAT5 proteins are adaptor proteins for histone acetylation enzymes. Histone acetylation at promoter and enhancer chromosomal regions opens the chromatin and allows access of transcription enzymes to specific genes in rapid response cell signals, such as in inflammation. Histone acetylation-mediated gene regulation is involved in expression of 2 key inflammatory response genes: CSF2, encoding granulocyte-macrophage colony stimulating factor (GM-CSF), and PTGS2, encoding prostaglandin synthase 2/cyclooxygenase 2 (PGS2/COX2). Prolonged CSF2 expression, high GM-CSF production, and GM-CSF activation of PTGS2 gene expression all are seen in type 1 diabetes (T1D) monocytes. Persistent phosphorylation activation of monocyte STAT5 (STAT5Ptyr) is also found in individuals with or at-risk for T1D. To examine whether elevated T1D monocyte STAT5Ptyr may be associated with aberrant inflammatory gene expression in T1D, blood monocytes from non-autoimmune controls and T1D patients were analyzed by flow cytometry for STAT5Ptyr activation, and by chromatin immuno-precipitation (ChIP) analyses for STAT5Ptyr's ability to bind at CSF2 and PTGS2 regulatory sites in association with histone acetylation. In unstimulated monocytes, STAT5Ptyr was elevated in 59.65% of T1D, but only 2.44% of control subjects (p<0.0001). Increased STAT5Ptyr correlated with T1D disease duration (p = 0.0030, r(2) = 0.0784). Unstimulated (p = 0.140) and GM-CSF-stimulated (p = 0.0485) T1D monocytes, had greater STAT5Ptyr binding to epigenetic regulatory sites upstream of CSF2 than control monocytes. Increased STAT5Ptyr binding in T1D monocytes was concurrent with binding at these sites of STAT6Ptyr (p = 0.0283), CBP/P300 histone acetylase, acetylated histones H3, SMRT/NCoR histone deacetylase (p = 0.0040), and RNA Polymerase II (p = 0.0040). Our study indicates that in T1D monocytes, STAT5Ptyr activation is significantly higher and that STAT5Ptyr is found bound to CSF2 promoter and PTGS2 enhancer regions coincident with histone acetylation and RNA polymerase II. These findings suggest that the persistent activation of STAT5 by GM-CSF may be involved in altering the epigenetic regulation of these inflammatory response genes in T1D monocytes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0076919PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3799903PMC
August 2014

Institutional profile: University of Florida and Shands Hospital Personalized Medicine Program: clinical implementation of pharmacogenetics.

Pharmacogenomics 2013 May;14(7):723-6

Department of Pharmacotherapy & Translational Research & Center for Pharmacogenomics, University of Florida, PO Box 100486, Gainesville, FL 32610-0486, USA.

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http://dx.doi.org/10.2217/pgs.13.59DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3684075PMC
May 2013

Anti-thymocyte globulin (ATG) differentially depletes naïve and memory T cells and permits memory-type regulatory T cells in nonobese diabetic mice.

BMC Immunol 2012 Dec 14;13:70. Epub 2012 Dec 14.

Department of Hematology, Xuanwu Hospital, Capital Medical University, Beijing, P.R. China.

Background: ATG has been employed to deplete T cells in several immune-mediated conditions. However, whether ATG administration affects naïve and memory T cell differently is largely unknown. THE CONTEXT AND PURPOSE OF THE STUDY: In this study, we assessed how murine ATG therapy affected T cell subsets in NOD mice, based on their regulatory and naïve or memory phenotype, as well as its influence on antigen-specific immune responses.

Results: Peripheral blood CD4+ and CD8+ T cells post-ATG therapy declined to their lowest levels at day 3, while CD4+ T cells returned to normal levels more rapidly than CD8+ T cells. ATG therapy failed to eliminate antigen-primed T cells. CD4+ T cell responses post-ATG therapy skewed to T helper type 2 (Th2) and possibly IL-10-producing T regulatory type 1 (Tr1) cells. Intriguingly, Foxp3+ regulatory T cells (Tregs) were less sensitive to ATG depletion and remained at higher levels following in vivo recovery compared to controls. Of note, the frequency of Foxp3+ Tregs with memory T cell phenotype was significantly increased in ATG-treated animals.

Conclusion: ATG therapy may modulate antigen-specific immune responses through inducing memory-like regulatory T cells as well as other protective T cells such as Th2 and IL-10-producing Tr1 cells.
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http://dx.doi.org/10.1186/1471-2172-13-70DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3547787PMC
December 2012

Total synthesis of grassystatin A, a probe for cathepsin E function.

Bioorg Med Chem 2012 Aug 7;20(15):4774-80. Epub 2012 Jun 7.

School of Pharmacy, Fudan University, Shanghai 201203, China.

The linear depsipeptide grassystatin A, a valuable probe for the study of cathepsin E function, has been synthesized by a [4+6] strategy. It exhibited specific inhibitory activity against cathepsin E with an IC(50) value of 0.8 nM. Our studies indicated that inhibition of cathepsin E did not have an impact on ovalbumin antigen processing and peptide presentation, unique from studies of other aspartic protease inhibitors.
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http://dx.doi.org/10.1016/j.bmc.2012.05.077DOI Listing
August 2012
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