Publications by authors named "Fernández-González Ana"

14 Publications

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The Role of Exosomes Derived From Mesenchymal Stromal Cells in Dermatology.

Front Cell Dev Biol 2021 7;9:647012. Epub 2021 Apr 7.

Cell Production and Tissue Engineering Unit, Virgen de las Nieves University Hospital, Granada, Spain.

The skin is the largest organ of the human body and its main functions include providing protection from external harmful agents, regulating body temperature, and homeostatic maintenance. Skin injuries can damage this important barrier and its functions so research focuses on approaches to accelerate wound healing and treat inflammatory skin diseases. Due to their regenerative and immunomodulatory properties, mesenchymal stromal cells (MSCs) have been reported to play a significant role in skin repair and regeneration. However, it seems that the secretome of these cells and exosomes in particular may be responsible for their functions in skin regeneration and the immunomodulation field. The present review aims to gather the available information about the role of MSC-derived exosomes for both and models of different skin conditions and to highlight the need for further research in order to overcome any limitations for clinical translation.
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http://dx.doi.org/10.3389/fcell.2021.647012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8058372PMC
April 2021

Cytotoxicity and Epidermal Barrier Function Evaluation of Common Antiseptics for Clinical Use in an Artificial Autologous Skin Model.

J Clin Med 2021 Feb 8;10(4). Epub 2021 Feb 8.

Cell Production and Tissue Engineering Unit, Virgen de las Nieves University Hospital, 18014 Granada, Spain.

Bioengineered artificial skin substitutes (BASS) are the main treatment used in addition to autografts when skin injuries involve a large body surface area. Antiseptic/antibiotic treatment is necessary to prevent infections in the BASS implant area. This study aims to evaluate the effect of antiseptics and antibiotics on cell viability, structural integrity, and epidermal barrier function in BASS based on hyaluronic acid during a 28 day follow-up period. Keratinocytes (KTs) and dermal fibroblasts (DFs) were isolated from skin samples and used to establish BASS. The following antibiotic/antiseptic treatment was applied every 48 h: colistin (1%), chlorhexidine digluconate (1%), sodium chloride (0.02%), and polyhexanide (0.1%). Cell viability (LIVE/DEAD assay), structural integrity (histological evaluation), and epidermal barrier function (trans-epidermal water loss, (TEWL), Tewameter) were also evaluated. Cell viability percentage of BASS treated with chlorhexidine digluconate was significantly lower ( ≤ 0.001) than the other antiseptics at day 28. Compared to other treatments, chlorhexidine digluconate and polyhexanide significantly affected the epithelium. No significant differences were found regarding epidermal barrier. These results may be useful for treatment protocols after implantation of BASS in patients and evaluating them in clinical practice. BASS represent a suitable model to test in vitro the impact of different treatments of other skin wounds.
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http://dx.doi.org/10.3390/jcm10040642DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7914612PMC
February 2021

Skin Barrier Function in Psoriasis and Atopic Dermatitis: Transepidermal Water Loss and Temperature as Useful Tools to Assess Disease Severity.

J Clin Med 2021 Jan 19;10(2). Epub 2021 Jan 19.

Dermatology Department, Hospital Universitario Virgen de las Nieves, Avenida de Madrid, 15, 18012 Granada, Spain.

Multiple diagnostic tools are used to evaluate psoriasis and atopic dermatitis (AD) severity, but most of them are based on subjective components. Transepidermal water loss (TEWL) and temperature are skin barrier function parameters that can be objectively measured and could help clinicians to evaluate disease severity accurately. Thus, the aims of this study are: (1) to compare skin barrier function between healthy skin, psoriatic skin and AD skin; and (2) to assess if skin barrier function parameters could predict disease severity. A cross-sectional study was designed, and epidermal barrier function parameters were measured. The study included 314 participants: 157 healthy individuals, 92 psoriatic patients, and 65 atopic dermatitis patients. TEWL was significantly higher, while stratum corneum hydration (SCH) (8.71 vs. 38.43 vs. 44.39 Arbitrary Units (AU)) was lower at psoriatic plaques than at uninvolved psoriatic skin and healthy controls. Patients with both TEWL > 13.85 g·mh and temperature > 30.85 °C presented a moderate/severe psoriasis (psoriasis area severity index (PASI) ≥ 7), with a specificity of 76.3%. TEWL (28.68 vs. 13.15 vs. 11.60 g·m h) and temperature were significantly higher, while SCH (25.20 vs. 40.95 vs. 50.73 AU) was lower at AD eczematous lesions than uninvolved AD skin and healthy controls. Patients with a temperature > 31.75 °C presented a moderate/severe AD (SCORing Atopic Dermatitis (SCORAD) ≥ 37) with a sensitivity of 81.8%. In conclusion, temperature and TEWL values may help clinicians to determine disease severity and select patients who need intensive treatment.
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http://dx.doi.org/10.3390/jcm10020359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7833436PMC
January 2021

Epidermal barrier changes in patients with psoriasis: The role of phototherapy.

Photodermatol Photoimmunol Photomed 2020 Dec 30. Epub 2020 Dec 30.

Dermatology Department, Hospital Universitario Virgen de las Nieves, Granada, Spain.

Background: Some skin diseases may modify epidermal barrier function. Psoriasis is a chronic multi-systemic inflammatory disease that affects the epidermal barrier. Phototherapy is an option for treating psoriasis, but little is known about how epidermal barrier function is modified by phototherapy in psoriatic patients.

Objectives: (a) To compare skin homeostasis between involved and uninvolved skin in psoriatic patients with healthy controls (b) To evaluate changes in the epidermal barrier function in psoriatic patients treated with phototherapy.

Methods: Sixty patients with plaque-type psoriasis and sixty gender and age-matched healthy controls were enrolled. Temperature, transepidermal water loss (TEWL), stratum corneum hydration (SCH), pH, elasticity, erythema and melanin index were measured using non-invasive tools in the healthy control and involved and uninvolved psoriatic skin before and after phototherapy.

Results: Healthy controls had lower TEWL and erythema index and higher SCH than psoriatic patients, both at uninvolved psoriatic skin and psoriasis plaques. TEWL was higher at psoriasis plaques than at uninvolved skin (19.20 vs 11.57 g/h/m ; P < .001). Following phototherapy, a decreasing trend was observed for TEWL, of 1.03 (SD 0.75) and 0.97 (SD 0.81) g/h/m for uninvolved and involved skin respectively. SCH was significantly lower at psoriatic plaques than at uninvolved skin (7.32 vs 36.62Arbitrary Units [AU]; P < .001). SCH increased by 1.15AU (SD 0.26) on psoriatic plaques after the phototherapy session (P < .001).

Conclusion: Psoriatic plaques showed epidermal barrier dysfunction compared to uninvolved skin and healthy controls. Phototherapy may improve epidermal barrier function in psoriatic patients. SCH increased after a phototherapy session on the psoriatic plaques.
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http://dx.doi.org/10.1111/phpp.12650DOI Listing
December 2020

From Grafts to Human Bioengineered Vascularized Skin Substitutes.

Int J Mol Sci 2020 Nov 2;21(21). Epub 2020 Nov 2.

Cell Production and Tissue Engineering Unit. Virgen de las Nieves University Hospital, 18014 Granada, Spain.

The skin plays an important role in the maintenance of the human's body physiological homeostasis. It acts as a coverage that protects against infective microorganism or biomechanical impacts. Skin is also implied in thermal regulation and fluid balance. However, skin can suffer several damages that impede normal wound-healing responses and lead to chronic wounds. Since the use of autografts, allografts, and xenografts present source limitations and intense rejection associated problems, bioengineered artificial skin substitutes (BASS) have emerged as a promising solution to address these problems. Despite this, currently available skin substitutes have many drawbacks, and an ideal skin substitute has not been developed yet. The advances that have been produced on tissue engineering techniques have enabled improving and developing new arising skin substitutes. The aim of this review is to outline these advances, including commercially available skin substitutes, to finally focus on future tissue engineering perspectives leading to the creation of autologous prevascularized skin equivalents with a hypodermal-like layer to achieve an exemplary skin substitute that fulfills all the biological characteristics of native skin and contributes to wound healing.
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http://dx.doi.org/10.3390/ijms21218197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7662999PMC
November 2020

Bioengineered Skin Intended as In Vitro Model for Pharmacosmetics, Skin Disease Study and Environmental Skin Impact Analysis.

Biomedicines 2020 Oct 31;8(11). Epub 2020 Oct 31.

Cell Production and Tissue Engineering Unit, Virgen de las Nieves University Hospital, 18014 Granada, Spain.

This review aims to be an update of Bioengineered Artificial Skin Substitutes (BASS) applications. At the first moment, they were created as an attempt to replace native skin grafts transplantation. Nowadays, these in vitro models have been increasing and widening their application areas, becoming important tools for research. This study is focus on the ability to design in vitro BASS which have been demonstrated to be appropriate to develop new products in the cosmetic and pharmacology industry. Allowing to go deeper into the skin disease research, and to analyze the effects provoked by environmental stressful agents. The importance of BASS to replace animal experimentation is also highlighted. Furthermore, the BASS validation parameters approved by the OECD (Organisation for Economic Co-operation and Development) are also analyzed. This report presents an overview of the skin models applicable to skin research along with their design methods. Finally, the potential and limitations of the currently available BASS to supply the demands for disease modeling and pharmaceutical screening are discussed.
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http://dx.doi.org/10.3390/biomedicines8110464DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7694072PMC
October 2020

Adherence and long-term outcomes of growth hormone therapy with easypod™ in pediatric subjects: Spanish ECOS study.

Endocr Connect 2019 Sep;8(9):1240-1249

Merck S.L.U., Madrid, Spain.

Background: Non-adherence to r-hGH treatments occurs in a variable percentage of subjects. One problem found when evaluating adherence is the great variability in methods of detection and definitions utilized in studies. This study assessed the level of adherence in subjects receiving r-hGH with the easypod™ electronic device.

Methods: National, multicenter, prospective and observational study involving 238 subjects (144 with GH deficiency (GHD), and 86 with small for gestational age (SGA), 8 with Turner Syndrome), who received r-hGH with easypod™ for at least 3 months before inclusion. The follow-up period was 4 years.

Results: Overall adherence was 94.5%; 97.5% after 6 months, 95.3% after 1 year, 93.7% after 2, 94.4% after 3 and 95.5% after 4 years of treatment. No differences in adherence were observed between prepubertal and pubertal groups and GHD and SGA groups. Change in height after 1 and 2 years, change in height SDS after 1 and 2 years, HV after 1 year, HV SDS after at 1 and 4 years, change in BMI after 1 year and change in BMI SDS at 1 and 2 years showed significant correlation with adherence. No significant differences in adherence according to IGF-I levels were found in follow-up visits or between groups.

Conclusions: The easypod™ electronic device, apart from being a precise and objective measure of adherence to r-hGH treatment, allows high compliance rates to be achieved over long periods of time. Adherence significantly impacts growth outcomes associated with r-hGH treatment.
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http://dx.doi.org/10.1530/EC-19-0325DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6733364PMC
September 2019

Control Materials and Digital PCR Methods for Evaluation of Circulating Cell-Free DNA Extractions from Plasma.

Methods Mol Biol 2018 ;1768:45-65

Molecular and Cell Biology Team, LGC, Teddington, UK.

Cell-free DNA is an accessible source of genetic material found naturally in plasma that could be used in many diagnostic applications. Translation of cfDNA analysis methods from research laboratories into the clinic would benefit from controls for monitoring the efficiency of patient sample purification and for quality control of the whole workflow from extraction through to analysis. Here we describe two types of control materials that can be "spiked" into plasma samples to monitor and evaluate different aspects of the workflow. The first control material is an internal control that enables evaluation of extraction efficiency, fragment size bias, and sample inhibition. The second control material serves as a parallel quality control material for measurement of specific genetic targets such as tumor mutations.
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http://dx.doi.org/10.1007/978-1-4939-7778-9_4DOI Listing
February 2019

Development of a highly sensitive liquid biopsy platform to detect clinically-relevant cancer mutations at low allele fractions in cell-free DNA.

PLoS One 2018 16;13(3):e0194630. Epub 2018 Mar 16.

Inivata Ltd, Granta Park, Cambridge, United Kingdom.

Introduction: Detection and monitoring of circulating tumor DNA (ctDNA) is rapidly becoming a diagnostic, prognostic and predictive tool in cancer patient care. A growing number of gene targets have been identified as diagnostic or actionable, requiring the development of reliable technology that provides analysis of multiple genes in parallel. We have developed the InVision™ liquid biopsy platform which utilizes enhanced TAm-Seq™ (eTAm-Seq™) technology, an amplicon-based next generation sequencing method for the identification of clinically-relevant somatic alterations at low frequency in ctDNA across a panel of 35 cancer-related genes.

Materials And Methods: We present analytical validation of the eTAm-Seq technology across two laboratories to determine the reproducibility of mutation identification. We assess the quantitative performance of eTAm-Seq technology for analysis of single nucleotide variants in clinically-relevant genes as compared to digital PCR (dPCR), using both established DNA standards and novel full-process control material.

Results: The assay detected mutant alleles down to 0.02% AF, with high per-base specificity of 99.9997%. Across two laboratories, analysis of samples with optimal amount of DNA detected 94% mutations at 0.25%-0.33% allele fraction (AF), with 90% of mutations detected for samples with lower amounts of input DNA.

Conclusions: These studies demonstrate that eTAm-Seq technology is a robust and reproducible technology for the identification and quantification of somatic mutations in circulating tumor DNA, and support its use in clinical applications for precision medicine.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0194630PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5856404PMC
July 2018

Comparative Study of Shrinkage in Human Skin, Artificial Human Skin, and Mouse Skin.

Am J Dermatopathol 2018 Apr;40(4):240-246

Dermatology Unit, Granada University Hospital Complex, Granada, Spain.

Introduction: The shrinkage of surgical specimens (SS) is known in human skin (HS) but has not been studied in an artificial skin (AS) or mouse skin (MS).

Objectives: To quantify the degree of shrinkage of SS and establish its timing in HS and an in vitro and animal model to explore the possible causes of this phenomenon.

Methodology: We collected 100 SS of HS, 50 SS of AS synthesized with fibrin-agarose biomaterials and 21 SS of MS. The width and length of specimens were measured before the surgical excision (pre-SE), at 5 minutes postsurgery (ex vivo), and after 24 hours of fixation in formalin (postfixation). Histological staining was performed to analyze the differences between HS, AS, and MS that may explain the differences in shrinkage.

Results: Between pre-SE and postfixation, the width and length shrank by 16.1% and 17.1% in HS, 14.5% and 8.5% in AS, and 26.5% and 23.1% in MS (P < 0.01), respectively. Shrinkage largely occurred between pre-SE and ex vivo. Cells and interstitial fibers were scant in AS and abundant in MS.

Conclusions: Almost all of the shrinkage occurred during the first 5 minutes postsurgery. According to the AS model findings, 53.6% of SS shrinkage would be explained by the action of dermal fibers and other cellular components of the dermis.
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http://dx.doi.org/10.1097/DAD.0000000000000951DOI Listing
April 2018

International Interlaboratory Digital PCR Study Demonstrating High Reproducibility for the Measurement of a Rare Sequence Variant.

Anal Chem 2017 02 18;89(3):1724-1733. Epub 2017 Jan 18.

Dana Farber Cancer Institute , Belfer Center for Applied Cancer Science and Department of Medical Oncology, Boston, Massachusetts 02115, United States.

This study tested the claim that digital PCR (dPCR) can offer highly reproducible quantitative measurements in disparate laboratories. Twenty-one laboratories measured four blinded samples containing different quantities of a KRAS fragment encoding G12D, an important genetic marker for guiding therapy of certain cancers. This marker is challenging to quantify reproducibly using quantitative PCR (qPCR) or next generation sequencing (NGS) due to the presence of competing wild type sequences and the need for calibration. Using dPCR, 18 laboratories were able to quantify the G12D marker within 12% of each other in all samples. Three laboratories appeared to measure consistently outlying results; however, proper application of a follow-up analysis recommendation rectified their data. Our findings show that dPCR has demonstrable reproducibility across a large number of laboratories without calibration. This could enable the reproducible application of molecular stratification to guide therapy and, potentially, for molecular diagnostics.
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http://dx.doi.org/10.1021/acs.analchem.6b03980DOI Listing
February 2017

Worldwide dissemination of acquired carbapenem-hydrolysing class D β-lactamases in Acinetobacter spp. other than Acinetobacter baumannii.

Int J Antimicrob Agents 2014 Apr 12;43(4):375-7. Epub 2014 Feb 12.

Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Goldenfelsstr. 19-21, 50935 Cologne, Germany. Electronic address:

The aim of this study was to identify acquired OXA-type carbapenemases in Acinetobacter spp. other than Acinetobacter baumannii. From a total of 453 carbapenem-susceptible and -resistant Acinetobacter isolates collected worldwide, 23 were positive for blaOXA genes by multiplex PCR. These isolates were identified as Acinetobacter pittii (n=18), Acinetobacter nosocomialis (n=2), Acinetobacter junii (n=1) and Acinetobacter genomic species 14TU/13BJ (n=2). The blaOXA genes and associated insertion sequence (IS) elements were sequenced by primer walking. In 11 of these isolates, sequencing of the PCR products revealed that they were false-positive for blaOXA. The remaining 12 isolates, originating from Europe, Asia, South America, North America and South Africa, harboured OXA-23 (n=4), OXA-58 (n=5), OXA-40-like (n=1) and OXA-143-like (n=1); one A. pittii isolate harboured both OXA-23 and OXA-58. IS elements were associated with blaOXA in 10 isolates. OXA multiplex PCR showed a high degree of false-positive results (47.8%), indicating that detection of blaOXA in non-baumanniiAcinetobacter spp. should be confirmed using additional methods.
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http://dx.doi.org/10.1016/j.ijantimicag.2014.01.012DOI Listing
April 2014

Detection of intrinsic blaOXA-51-like by multiplex PCR on its own is not reliable for the identification of Acinetobacter baumannii.

Int J Med Microbiol 2013 Mar 1;303(2):88-9. Epub 2013 Feb 1.

Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Cologne, Germany.

Three clinical A. baumannii isolates Ab-508, Ab-511, and Ab-653 were recovered from South Africa, South Korea, and Turkey, respectively. Multiplex PCR to detect OXA-type carbapenemases showed atypical blaOXA-51-like amplification products. The aim of this study was to investigate the background of changes in blaOXA-51-like PCR products. Isolates were confirmed as A. baumannii using gyrB multiplex and rpoB sequencing and were epidemiologically unrelated by rep-PCR-based DiversiLab. Sequencing of blaOXA-51-like revealed an insertion of ISAba15 in blaOXA-66 (isolate Ab-511) and an insertion of the novel ISAba19 in blaOXA-78 (isolates Ab-508 and Ab-653). Detection of the intrinsic blaOXA-51-like by OXA-multiplex PCR should not be considered a fully reliable method for identification of A. baumannii when used without an additional independent method. Other species identification methods such as gyrB multiplex PCR and rpoB sequencing should be used to reliably identify A. baumannii.
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http://dx.doi.org/10.1016/j.ijmm.2012.12.007DOI Listing
March 2013

A dual requirement for Iroquois genes during Xenopus kidney development.

Development 2008 Oct 20;135(19):3197-207. Epub 2008 Aug 20.

Centro Andaluz de Biología del Desarrollo, CSIC/UPO, Sevilla, Spain.

The Iroquois (Irx) genes encode evolutionary conserved homeoproteins. We report that Xenopus genes Irx1 and Irx3 are expressed and required during different stages of Xenopus pronephros development. They are initially expressed during mid-neurulation in domains extending over most of the prospective pronephric territory. Expression onset takes place after kidney anlage specification, but before pronephric organogenesis occurs. Later, during nephron segmentation, expression becomes restricted to the intermediate tubule region of the proximal-distal axis. Loss- and gain-of-function analyses, performed with specific morpholinos and inducible wild-type and dominant-negative constructs, reveal a dual requirement for Irx1 and Irx3 during pronephros development. During neurula stages, these genes maintain the specification of the pronephric territory and define its size. This seems to occur, at least in part, through positive regulation of Bmp signalling. Subsequently, Irx genes are required for proper formation of the intermediate tubule. Finally, we find that retinoic acid signalling activates both Irx1 and Irx3 genes in the pronephros.
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http://dx.doi.org/10.1242/dev.023697DOI Listing
October 2008