Publications by authors named "Emily Dando"

7 Publications

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Correction to: ISSLS prize in basic science 2021: a novel inducible system to regulate transgene expression of TIMP1.

Eur Spine J 2021 Mar 7. Epub 2021 Mar 7.

Ferguson Laboratory for Orthopaedic and Spine Research, Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA.

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http://dx.doi.org/10.1007/s00586-021-06783-7DOI Listing
March 2021

ISSLS prize in basic science 2021: a novel inducible system to regulate transgene expression of TIMP1.

Eur Spine J 2021 Feb 1. Epub 2021 Feb 1.

Ferguson Laboratory for Orthopaedic and Spine Research, Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA.

Purpose: Inflammatory and oxidative stress upregulates matrix metalloproteinase (MMP) activity, leading to intervertebral disc degeneration (IDD). Gene therapy using human tissue inhibitor of metalloproteinase 1 (hTIMP1) has effectively treated IDD in animal models. However, persistent unregulated transgene expression may have negative side effects. We developed a recombinant adeno-associated viral (AAV) gene vector, AAV-NFκB-hTIMP1, that only expresses the hTIMP1 transgene under conditions of stress.

Methods: Rabbit disc cells were transfected or transduced with AAV-CMV-hTIMP1, which constitutively expresses hTIMP1, or AAV-NFκB-hTIMP1. Disc cells were selectively treated with IL-1β. NFκB activation was verified by nuclear translocation. hTIMP1 mRNA and protein expression were measured by RT-PCR and ELISA, respectively. MMP activity was measured by following cleavage of a fluorogenic substrate.

Results: IL-1β stimulation activated NFκB demonstrating that IL-1β was a surrogate for inflammatory stress. Stimulating AAV-NFκB-hTIMP1 cells with IL-1β increased hTIMP1 expression compared to unstimulated cells. AAV-CMV-hTIMP1 cells demonstrated high levels of hTIMP1 expression regardless of IL-1β stimulation. hTIMP1 expression was comparable between IL-1β stimulated AAV-NFκB-hTIMP1 cells and AAV-CMV-hTIMP1 cells. MMP activity was decreased in AAV-NFκB-hTIMP1 cells compared to baseline levels or cells exposed to IL-1β.

Conclusion: AAV-NFκB-hTIMP1 is a novel inducible transgene delivery system. NFκB regulatory elements ensure that hTIMP1 expression occurs only with inflammation, which is central to IDD development. Unlike previous inducible systems, the AAV-NFκB-hTIMP1 construct is dependent on endogenous factors, which minimizes potential side effects caused by constitutive transgene overexpression. It also prevents the unnecessary production of transgene products in cells that do not require therapy.
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http://dx.doi.org/10.1007/s00586-021-06728-0DOI Listing
February 2021

The asymmetric function of Dph1-Dph2 heterodimer in diphthamide biosynthesis.

J Biol Inorg Chem 2019 09 28;24(6):777-782. Epub 2019 Aug 28.

Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, 14853, USA.

Diphthamide, the target of diphtheria toxin, is a post-translationally modified histidine residue found in archaeal and eukaryotic translation elongation factor 2 (EF2). In the first step of diphthamide biosynthesis, a [4Fe-4S] cluster-containing radical SAM enzyme, Dph1-Dph2 heterodimer in eukaryotes or Dph2 homodimer in archaea, cleaves S-adenosylmethionine and transfers the 3-amino-3-carboxypropyl group to EF2. It was demonstrated previously that for the archaeal Dph2 homodimer, only one [4Fe-4S] cluster is necessary for the in vitro activity. Here, we demonstrate that for the eukaryotic Dph1-Dph2 heterodimer, the [4Fe-4S] cluster-binding cysteine residues in each subunit are required for diphthamide biosynthesis to occur in vivo. Furthermore, our in vitro reconstitution experiments with Dph1-Dph2 mutants suggested that the Dph1 cluster serves a catalytic role, while the Dph2 cluster facilitates the reduction of the Dph1 cluster by the physiological reducing system Dph3/Cbr1/NADH. Our results reveal the asymmetric functional roles of the Dph1-Dph2 heterodimer and may help to understand how the Fe-S clusters in radical SAM enzymes are reduced in biology.
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http://dx.doi.org/10.1007/s00775-019-01702-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6893874PMC
September 2019

Intralesional 5-Fluorouracil for the Nonsurgical Management of Low-Risk, Invasive Squamous Cell Carcinoma.

Dermatol Surg 2020 01;46(1):126-130

Department of Dermatology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania.

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http://dx.doi.org/10.1097/DSS.0000000000001740DOI Listing
January 2020

The rise of transcutaneous drug delivery for the management of alopecia: a review of existing literature and an eye towards the future.

J Cosmet Laser Ther 2019 Aug 9;21(5):247-254. Epub 2018 Oct 9.

a Department of Dermatology , University of Minnesota , Minneapolis , Minnesota , United States.

: Fractional lasers and microneedling devices are increasingly used with topical drugs to treat various conditions, including alopecia, as they grant access to dermal structures such as hair follicles and cutaneous vasculature. : To perform a comprehensive review on transcutaneous drug delivery for the management of alopecia. : PubMed, Embase, and Ovid Medline databases were searched using terms including: alopecia, microneedling, lasers, androgenetic alopecia (AGA), alopecia areata (AA), drug delivery. Articles were examined for inclusion criteria: diagnosis of alopecia regardless of type, use of fractional laser or microneedling devices, and subsequent administration of topical medication. : 8 studies, 6 prospective clinical trials and 2 case series, examining either AA or AGA were identified. For AA, five studies examined microneedling together with topical triamcinolone in three of these, while two studies used photodynamic therapy. Regarding AGA, two studies used topical minoxidil plus microneedling, and one examined topical finasteride with fractional erbium glass laser. Improvement was seen in 6 of the 8 studies. : Transcutaneous drug delivery via fractional laser and microneedling is a promising modality with preliminary evidence for increased hair regrowth over topical therapy alone. Further studies are needed to elucidate treatment parameters and appropriate device selection for drug delivery.
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http://dx.doi.org/10.1080/14764172.2018.1525743DOI Listing
August 2019

A Cost-Utility Analysis of 5 Strategies for the Management of Acute Otitis Media in Children.

J Pediatr 2017 10 28;189:54-60.e3. Epub 2017 Jun 28.

Division of General Internal Medicine, University of Pittsburgh, Pittsburgh, PA.

Objective: To assess whether antimicrobial therapy in young children with acute otitis media reduces time to resolution of symptoms, overall symptom burden, and persistence of otoscopic evidence of infection. We used a cost-utility model to evaluate whether immediate antimicrobial treatment seems to be worthwhile, and if so, which antimicrobial agent is most cost effective.

Study Design: We compared the cost per quality-adjusted life-day of 5 treatment regimens in children younger than 2 years of age with acute otitis media: immediate amoxicillin/clavulanate, immediate amoxicillin, immediate cefdinir, watchful waiting, and delayed prescription (DP) for antibiotic.

Results: The 5 treatment regimens, listed in order from least effective to most effective were DP, watchful waiting, immediate cefdinir, immediate amoxicillin, and immediate amoxicillin/clavulanate. Listed in order from least costly to most costly, the regimens were DP, immediate amoxicillin, watchful waiting, immediate amoxicillin/clavulanate, and immediate cefdinir. The incremental cost-utility ratio of immediate amoxicillin compared with DP was $101.07 per quality-adjusted life-day gained. The incremental cost-utility ratio of immediate amoxicillin/clavulanate compared with amoxicillin was $2331.28 per quality-adjusted life-day gained.

Conclusions: In children younger than 2 years of age with acute otitis media and no recent antibiotic exposure, immediate amoxicillin seems to be the most cost-effective initial treatment.
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http://dx.doi.org/10.1016/j.jpeds.2017.05.047DOI Listing
October 2017

Dph3 is an electron donor for Dph1-Dph2 in the first step of eukaryotic diphthamide biosynthesis.

J Am Chem Soc 2014 Feb 22;136(5):1754-7. Epub 2014 Jan 22.

Department of Chemistry and Chemical Biology, Cornell University , Ithaca, New York 14853, United States.

Diphthamide, the target of diphtheria toxin, is a unique posttranslational modification on translation elongation factor 2 (EF2) in archaea and eukaryotes. The biosynthesis of diphthamide was proposed to involve three steps. The first step is the transfer of the 3-amino-3-carboxypropyl group from S-adenosyl-l-methionine (SAM) to the histidine residue of EF2, forming a C-C bond. Previous genetic studies showed this step requires four proteins in eukaryotes, Dph1-Dph4. However, the exact molecular functions for the four proteins are unknown. Previous study showed that Pyrococcus horikoshii Dph2 (PhDph2), a novel iron-sulfur cluster-containing enzyme, forms a homodimer and is sufficient for the first step of diphthamide biosynthesis in vitro. Here we demonstrate by in vitro reconstitution that yeast Dph1 and Dph2 form a complex (Dph1-Dph2) that is equivalent to the homodimer of PhDph2 and is sufficient to catalyze the first step in vitro in the presence of dithionite as the reductant. We further demonstrate that yeast Dph3 (also known as KTI11), a CSL-type zinc finger protein, can bind iron and in the reduced state can serve as an electron donor to reduce the Fe-S cluster in Dph1-Dph2. Our study thus firmly establishes the functions for three of the proteins involved in eukaryotic diphthamide biosynthesis. For most radical SAM enzymes in bacteria, flavodoxins and flavodoxin reductases are believed to serve as electron donors for the Fe-S clusters. The finding that Dph3 is an electron donor for the Fe-S clusters in Dph1-Dph2 is thus interesting and opens up new avenues of research on electron transfer to Fe-S proteins in eukaryotic cells.
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http://dx.doi.org/10.1021/ja4118957DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3985478PMC
February 2014