Dermal white adipose tissue development and metabolism: The role of transcription factor Foxn1.

Intradermal adipocytes form dermal white adipose tissue (dWAT), a unique fat depot localized in the lower layer of the dermis. However, recognition of molecular factors regulating dWAT development, homeostasis, and bioactivity is limited. Using Foxn1-/- and Foxn1+/+ mice, we demonstrated that epidermally expressed Foxn1 regulates dWAT development and defines the adipogenic capacity of dermal fibroblasts. In intact and post-wounded skin, Foxn1 contributes to the initial stimulation of dWAT adipogenesis and participates in the modulation of lipid metabolism processes. Furthermore, Foxn1 activity strengthens adipogenic processes through Bmp2 and Igf2 signaling and regulates lipid metabolism in differentiated dermal fibroblasts. The results reveal the contribution of Foxn1 to dWAT metabolism, thus identifying possible targets for modulation and regulation of dWAT in physiological and pathological processes in the skin.

Hypoxia reveals a new function of Foxn1 in the keratinocyte antioxidant defense system.

Skin exposed to environmental threats, including injuries and oxidative stress, develops an efficient but not fully recognized system of repair and antioxidant protection. Here, using mass spectrometry analysis (LC-MS/MS), followed by in vitro and in vivo experiments, we provided evidence that Foxn1 in keratinocytes regulates elements of the electron transport chain and participates in the thioredoxin system (Txn2, Txnrd3, and Srxn1) induction, particularly in a hypoxic environment. We first showed that Foxn1 in keratinocytes upregulates glutathione thioredoxin reductase 3 (Txnrd3) protein expression, and high levels of Txnrd3 mRNA were detected in injured skin of Foxn1+/+ mice. We also showed that Foxn1 strongly downregulated the Ccn2 protein expression, participating in epidermal reconstruction after injury. An in vitro assay revealed that Foxn1 controls keratinocyte migration, stimulating it under normoxia and suppressing it under hypoxia. Keratinocytes overexpressing Foxn1 and exposed to hypoxia displayed a reduced ability to promote angiogenesis by downregulating Vegfa expression. In conclusion, this study showed a new mechanism in which Foxn1, along with hypoxia, participates in the activation of antioxidant defense and controls the functional properties of keratinocytes.

Comparative studies on the effect of pig adipose-derived stem cells (pASCs) preconditioned with hypoxia or normoxia on skin wound healing in mice.

Adipose-derived stem cells (ASCs) from human and animal fat have emerged as therapeutic alternatives for damaged tissues. Pre-conditioning of ASCs with hypoxia results in their functional enhancement, which might facilitate the process of healing. However, there is still a critical need for large-scale preclinical studies to reinforce the translation of these findings into clinical practice for humans and in veterinary medicine. Here, we adapted a full-thickness excisional skin wound mouse model to evaluate and compare the effect of pig adipose-derived stem cells (pASCs) cultured under normoxia (pASCs-Nor) or hypoxia (pASCs-Hyp) on the healing process. We show that pASCs-Hyp accelerated re-epithelialization, increased hyaluronic acid (HA) content, and decreased scar elevation index (SEI) during the late stage of healing (day 21). Transplantation of pASCs-Hyp also promoted expression of angiogenic marker VegfA and decreased levels of pro-scarring Tgfß1. Mice tolerated xenotransplantation of the pASCs with no impact on macrophage (CD68 -positive cell) content. However, wounds treated with pASCs-Hyp exhibited decreased elasticity at the early stage of healing and increased expression of Wnt signaling members including Wnt10a, Wnt11, and ß-catenin, which are associated with scar-forming wound repair. In conclusion, pASCs treatment may provide a critical step toward the evaluation of pASCs as therapeutically relevant cells in the context of wound healing.

Activation of Ethanol Transformation on Copper-Containing SBA-15 and MnSBA-15 Catalysts by the Presence of Oxygen in the Reaction Mixture.

The aim of this study was to get insight into the pathway of the acetaldehyde formation from ethanol (the rate-limiting step in the production of 1,3-butadiene) on Cu-SBA-15 and Cu-MnSBA-15 mesoporous molecular sieves. Physicochemical properties of the catalysts were investigated by XRD, N2 ads/des, Uv-vis, XPS, EPR, pyridine adsorption combined with FTIR, 2-propanol decomposition and 2,5-hexanedione cyclization and dehydration test reactions. Ethanol dehydrogenation to acetaldehyde (without and with oxygen) was studied in a flow system using the FTIR technique. In particular, the effect of Lewis acid and basic (Lewis and BrØnsted) sites, and the oxygen presence in the gas reaction mixture with ethanol on the activity and selectivity of copper catalysts, was assessed and discussed. Two different reaction pathways have been proposed depending on the reaction temperature and the presence or absence of oxygen in the flow of the reagents (via ethoxy intermediate way at 593 K, in ethanol flow, or ethoxide intermediate way at 473 K in the presence of ethanol and oxygen in the reaction mixture).

Transcatheter pulmonary denervation in patients with left heart failure with reduced ejection fraction and combined precapillary and postcapillary pulmonary hypertension: A prospective single center experience.

OBJECTIVES: The present study was a prospective, single-center, single-arm study to investigate the efficacy of transcatheter pulmonary artery denervation (TPADN) in patients with combined postcapillary and precapillary PH (Cpc-PH) associated with left heart failure with reduced ejection fraction (HF-rEF). BACKGROUND: Pulmonary hypertension (PH) in patients with left ventricular systolic dysfunction has a negative impact on outcome. METHODS: The combination of pulmonary artery systolic pressure (PAPs) ≥60 mmHg, transpulmonary pressure gradient (TPG) ≥12 mmHg, nonreversible mean PAP, and pulmonary vascular resistance (PVR) ≥3.5 Wood Units was considered as too high risk for heart transplantation (HTx). The clinical efficacy endpoint was an improvement in 6-min walking test and the hemodynamic endpoints were changes in PAPs, PVR, and TPG between baseline and 6 months. Circumferential radiofrequency applications were delivered around distal main, left and right pulmonary arteries. At each ablation point temperature was 45°C and energy 10 W. RESULTS: TPADN was performed in 10 patients. At 6-month in 5 patients we observed reduction in PAP, PVR, TPG, and DPG and then 1 had successful HTx, 2 are on HTx waiting list, 2 received LVADs, 2 patients did not improve, and 3 patients died. CONCLUSIONS: TPADN may be beneficial in selected patients with HF-rEF and Cpc-PH.

Effect of Pig-Adipose-Derived Stem Cells' Conditioned Media on Skin Wound-Healing Characteristics In Vitro.

The primary mechanism by which adipose-derived stem cells (ASCs) exert their reparative or regenerative potential relies predominantly on paracrine action. Secretory abilities of ASCs have been found to be amplified by hypoxia pre-conditioning. This study investigates the impact of hypoxia (1% O2) on the secretome composition of pig ASCs (pASCs) and explores the effect of pASCs' conditioned media (CM) on skin cell functions in vitro and the expression of markers attributed to wound healing. Exposure of pASCs to hypoxia increased levels of vascular endothelial growth factor (VEGF) in CM-Hyp compared to CM collected from the cells cultured in normoxia (CM-Nor). CM-Hyp promoted the migratory ability of pig keratinocytes (pKERs) and delayed migration of pig dermal fibroblasts (pDFs). Exposure of pKERs to either CM-Nor or CM-Hyp decreased the levels of pro-fibrotic indicators WNT10A and WNT11. Furthermore, CM-Hyp enhanced the expression of KRT14, the marker of the basal epidermis layer. In contrast, CM-Nor showed a stronger effect on pDFs manifested by increases in TGFB1, COL1A1, COL3A1, and FN1 mRNA expression. The formation of three-dimensional endothelial cell networks was improved in the presence of CM-Hyp. Overall, our results demonstrate that the paracrine activity of pASCs affects skin cells, and this property might be used to modulate wound healing.

Dermal White Adipose Tissue (dWAT) Is Regulated by Foxn1 and Hif-1α during the Early Phase of Skin Wound Healing.

Dermal white adipose tissue (dWAT) is involved in the maintenance of skin homeostasis. However, the studies concerning its molecular regulation are limited. In the present paper, we ask whether the introduction of two transcription factors, Foxn1 and Hif-1α, into the post-wounded skin of Foxn1-/- mice regulates dWAT during wound healing (days 3 and 6). We have chosen lentivirus vectors (LVs) as a tool to deliver Foxn1 and Hif-1α into the post-wounded skin. We documented that combinations of both transgenes reduces the number, size and diameter of dermal adipocytes at the wound bed area. The qRT-PCR analysis of pro-adipogenic genes, revealed that LV-Hif-1α alone, or combined with LV-Foxn1, increases the mRNA expression of Pparγ, Glut 4 and Fasn at post-wounding day 6. However, the most spectacular stimulatory effect of Foxn1 and/or Hif-1α was observed for Igf2, the growth factor participating in adipogenic signal transduction. Our data also shows that Foxn1/Hif-1α, at post-wounding day 3, reduces levels of CD68 and MIP-1γ mRNA expression and the percentage of CD68 positive cells in the wound site. In conclusion, the present data are the first to document that Foxn1 and Hif-1α cooperatively (1) regulate dWAT during the proliferative phase of skin wound healing through the Igf2 signaling pathway, and (2) reduce the macrophages content in the wound site.

Transcriptome and Proteome Analysis Revealed Key Pathways Regulating Final Stage of Oocyte Maturation of the Turkey (Meleagris gallopavo).

In birds, the zona pellucida (ZP) matrix that surrounds the ovulated oocyte-called the inner perivitelline layer-is involved in sperm-zona interaction and successful fertilization. To identify the important genes and proteins connected with the final step of egg development, next-generation sequencing and two-dimensional electrophoresis, combined with mass spectrometry, were used for the analysis of mature oocytes at the F1 developmental stage. A total of 8161 genes and 228 proteins were annotated. Six subfamilies of genes, with codes ZP, ZP1-4, ZPD, and ZPAX, were identified, with the dominant expression of ZPD. The main expression site for ZP1 was the liver; however, granulosa cells may also participate in local ZP1 secretion. A ubiquitination system was identified in mature oocytes, where ZP1 was found to be the main ubiquitinated protein. Analysis of transcripts classified in estrogen receptor (ESR) signaling indicated the presence of ESR1 and ESR2, as well as a set of estrogen-dependent genes involved in both genomic and nongenomic mechanisms for the regulation of gene expression by estrogen. Oxidative phosphorylation was found to be a possible source of adenosine triphosphate, and the nuclear factor erythroid 2-related factor 2 signaling pathway could be involved in the response against oxidative stress. Oocyte-granulosa cell communication by tight, adherens, and gap junctions seems to be essential for the final step of oocyte maturation.

From Supramolecular Hydrogels to Multifunctional Carriers for Biologically Active Substances.

Supramolecular hydrogels are 3D, elastic, water-swelled materials that are held together by reversible, non-covalent interactions, such as hydrogen bonds, hydrophobic, ionic, host-guest interactions, and metal-ligand coordination. These interactions determine the hydrogels' unique properties: mechanical strength; stretchability; injectability; ability to self-heal; shear-thinning; and sensitivity to stimuli, e.g., pH, temperature, the presence of ions, and other chemical substances. For this reason, supramolecular hydrogels have attracted considerable attention as carriers for active substance delivery systems. In this paper, we focused on the various types of non-covalent interactions. The hydrogen bonds, hydrophobic, ionic, coordination, and host-guest interactions between hydrogel components have been described. We also provided an overview of the recent studies on supramolecular hydrogel applications, such as cancer therapy, anti-inflammatory gels, antimicrobial activity, controlled gene drug delivery, and tissue engineering.

Examining the Impact of Squaric Acid as a Crosslinking Agent on the Properties of Chitosan-Based Films.

Hydrogels based on chitosan are very versatile materials which can be used for tissue engineering as well as in controlled drug delivery systems. One of the methods for obtaining a chitosan-based hydrogel is crosslinking by applying different components. The objective of the present study was to obtain a series of new crosslinked chitosan-based films by means of solvent casting method. Squaric acid-3,4-dihydroxy-3-cyclobutene-1,2-dione-was used as a safe crosslinking agent. The effect of the squaric acid on the structural, mechanical, thermal, and swelling properties of the formed films was determined. It was established that the addition of the squaric acid significantly improved Young's modulus, tensile strength, and thermal stability of the obtained materials. Moreover, it should be stressed that the samples consisting of chitosan and squaric acid were characterized by a higher swelling than pure chitosan. The detailed characterization proved that squaric acid could be used as a new effective crosslinking agent.

Switching and Swapping of Quantum Information: Entropy and Entanglement Level.

Information switching and swapping seem to be fundamental elements of quantum communication protocols. Another crucial issue is the presence of entanglement and its level in inspected quantum systems. In this article, a formal definition of the operation of the swapping local quantum information and its existence proof, together with some elementary properties analysed through the prism of the concept of the entropy, are presented. As an example of the local information swapping usage, we demonstrate a certain realisation of the quantum switch. Entanglement levels, during the work of the switch, are calculated with the Negativity measure and a separability criterion based on the von Neumann entropy, spectral decomposition and Schmidt decomposition. Results of numerical experiments, during which the entanglement levels are estimated for systems under consideration with and without distortions, are presented. The noise is generated by the Dzyaloshinskii-Moriya interaction and the intrinsic decoherence is modelled by the Milburn equation. This work contains a switch realisation in a circuit form-built out of elementary quantum gates, and a scheme of the circuit which estimates levels of entanglement during the switch's operating.

Synthesis, structure and biological evaluation of ruthenium(III) complexes of triazolopyrimidines with anticancer properties.

Six novel ruthenium(III) complexes of general formula [RuCl3(L)3] (1,3,5) and [RuCl3(H2O)(L)2] (2,4,6), where L stands for three different triazolopyrimidine-derived ligands, are reported. The compounds have been structurally characterized (IR, EPR, SCXRD), and their magnetic moments have been determined. The single-crystal X-ray diffraction study revealed a slightly distorted octahedral geometry of the Ru(III) complexes with mer configuration in 1 and 5, and fac configuration in 3. In 2 and 4, three chloride ions are in mer configuration and the two triazolopyrimidines are oriented trans mutually with the water molecule playing the role of the sixth ligand. All complexes have been thoroughly screened for their in vitro cytotoxicity against human breast cancer cell line MCF-7, human cervical cancer cell line HeLa, and L929 murine fibroblast cells, uncovering among others that the most lipophilic complexes 5 and 6, containing the bulky ligand dptp (5,7-diphenyl-1,2,4-triazolo[1,5-a]pyrimidine), display high cytotoxic activity against MCF-7, and HeLa cells. Moreover, it was also revealed that during the interaction of the complexes 1-6 with the cancer MCF-7 cell line, reactive oxygen species are released intracellularly, which could indicate that they are involved in cell apoptosis. Furthermore, extensive studies have been carried out to reveal the mechanism by which complexes 1-6 interact with DNA, albumin, and apotransferrin. The biological studies were complemented by detailed kinetic studies of the hydrolysis of the complexes in the pH range 5-8, to determine the stability of the complexes in solution. Six novel ruthenium(III) complexes with triazolopyrimidine derivatives demonstrated the potential for use as anticancer agents by maintaining the toxic effect on MCF-7 and HeLa cells.

Evidence for the presence of stem/progenitor cells in porcine endometrium.

A population of adult stem cells responsible for cyclic reconstructing and remodeling has been proposed to reside in the highly regenerative mammalian endometrium. Recently, stem/progenitor cells have been identified in the human and mouse endometrium, but less is known about these cells in livestock animals. Using Hoechst 33342 fluorescent dye staining and flow cytometry, we identified an emerging cell side population that may be responsible for the regeneration process of the porcine endometrium. The percentage of side-population cells on Day 19 of the estrous cycle was significantly higher than that on Days 2-4. Moreover, single cells were able to seed clones that could differentiate into three independent mesenchymal-cell lineages. We also demonstrated the expression of specific markers of self-renewal cells on these side-population cells and the presence of a population of cells among the stromal cells that possess markers for mesenchymal stem cells. These results indicate that the porcine endometrium contains a population of cells with the capacity for self-renewal and a high rate of proliferation, which depend on the phase of the estrous cycle. These cells could potentially be involved in the cyclic reconstruction of the porcine endometrium.

Inhibition of 5-lipoxygenase by U73122 is due to covalent binding to cysteine 416.

U73122 which was originally identified as a phospholipase C inhibitor represents a potent direct inhibitor of purified 5-lipoxygenase (5-LO) with an IC50 value of 30 nM. 5-LO catalyzes the conversion of arachidonic acid (AA) into leukotrienes which represent mediators involved in inflammatory and allergic reactions and in host defense reactions against microorganisms. Since the efficient inhibition of the human 5-LO enzyme depended on the thiol reactivity of the maleinimide group of U73122, we used this property to identify cysteine residues in the 5-LO protein that are important for 5-LO inhibition by U73122. We found by MALDI-MS that U73122 covalently binds to cysteine residues 99, 159, 248, 264, 416 and 449. Mutation of Cys416 to serine strongly reduces inhibition of 5-LO by U73122 and the additional mutation of three cysteines close to Cys416 further impairs 5-LO inhibition by the compound. Wash out experiments with U73122 and 5-LO indicated an irreversible binding of U73122. Together, our data suggest that the area around Cys416 which is close to the proposed AA entry channel to the active site is an interesting target for the development of new 5-LO inhibitors.

Psoriasis vulgaris and familial cancer risk- a population-based study.

BACKGROUND: Follow-up studies of psoriasis patients indicate an increased risk in the occurrence of malignancies at different sites of origin. Population stratification and/or complicated interpretation of evidence on the risk of cancer (due to the small number of patients included in most series) lead to inconsistent data. Herein we investigated the risk of occurrence of malignancies at different sites of origin in a series of 517 psoriasis patients and their 1st degree relatives. METHODS: We evaluated the tumour spectrum as well as the age of the patient at diagnosis of cancers in psoriasis families along with the observed and expected frequencies of malignancies. The distribution of 17 common mutations/polymorphisms in 10 known cancer susceptibility genes among psoriasis patients and 517 matched healthy controls were examined. No such study has been published to date. RESULTS: The statistical comparison of the observed and expected frequencies of cancers revealed a higher than expected occurrence of Hodgkin's lymphoma among males in psoriasis families when compared to the general population (OR=1.8, 95%CI 1.6-2.1, p=0.002). There was a non-significant tendency towards a younger age of onset and overrepresentation of laryngeal cancer and leukaemia in psoriasis families. We found no major differences in the distribution of cancer susceptibility mutations among our cases and the healthy controls. CONCLUSIONS: The results of our study suggest an increased risk of Hodgkin's lymphoma for male members of psoriasis families. Further studies are needed to confirm the findings and to evaluate whether or not the application of cancer surveillance protocols for Hodgkin's lymphoma, leukaemia and laryngeal cancer are justified in these families.

Structural Diversity, XAS and Magnetism of Copper(II)-Nickel(II) Heterometallic Complexes Based on the [Ni(NCS)6]4- Unit.

The new heterometallic compounds, [{Cu(pn)2}2Ni(NCS)6]n·2nH2O (1), [{CuII(trien)}2Ni(NCS)6CuI(NCS)]n (2) and [Cu(tren)(NCS)]4[Ni(NCS)6] (3) (pn = 1,2-diaminopropane, trien = triethylenetetramine and tren = tris(2-aminoethylo)amine), were obtained and characterized by X-ray analysis, IR spectra, XAS and magnetic measurements. Compounds 1, 2 and 3 show the structural diversity of 2D, 1D and 0D compounds, respectively. Depending on the polyamine used, different coordination polyhedron for Cu(II) was found, i.e., distorted octahedral (1), square pyramidal (2) and trigonal bipyramidal (3), whereas coordination polyhedron for nickel(II) was always octahedral. It provides an approach for tailoring magnetic properties by proper selection of auxiliary ligands determining the topology. In 1, thiocyanate ligands form bridges between the copper and nickel ions, creating 2D layers of sql topology with weak ferromagnetic interactions. Compound 2 is a mixed-valence copper coordination polymer and shows the rare ladder topology of 1D chains decorated with [CuII(tren)]2+ antennas as the side chains attached to nickel(II). The ladder rails are formed by alternately arranged Ni(II) and Cu(I) ions connected by N2 thiocyanate anions and rungs made by N3 thiocyanate. For the Cu(I) ions, the tetrahedral thiocyanate environment mixed N/S donor atoms was found, confirming significant coordination spheres rearrangement occurring at the copper precursor together with the reduction in some Cu(II) to Cu(I). Such topology enables significant simplification of the magnetic properties modeling by assuming magnetic coupling inside {NiIICuII2} trinuclear units separated by diamagnetic [Cu(NCS)(SCN)3]3- linkers. Compound 3 shows three discrete mononuclear units connected by N-H…N and N-H…S hydrogen bonds. Analysis of XAS proves that the average ligand character and the covalency of the unoccupied metal d-based orbitals for copper(II) and nickel(II) increase in the following order: 1 → 2 → 3. In 1 and 2, a weak ferromagnetic coupling between copper(II) and nickel(II) was found, but in 2, additional and stronger antiferromagnetic interaction between copper(II) ions prevailed. Compound 3, as an ionic pair, shows, as expected, a spin-only magnetic moment.

Specific expression of alternatively spliced genes in the turkey (Meleagris gallopavo) reproductive tract revealed their function in spermatogenesis and post-testicular sperm maturation.

The tissue-specific profile of alternatively spliced genes (ASGs) and their involvement in reproduction processes characteristic of turkey testis, epididymis, and ductus deferens were investigated for the first time in birds. Deep sequencing of male turkey reproductive tissue RNA samples (n = 6) was performed using Illumina RNA-Seq with 2 independent methods, rMATs and SUPPA2, for differential alternative splicing (DAS) event prediction. The expression of selected ASGs was validated using quantitative real-time reverse transcriptase-polymerase chain reaction. The testis was found to be the site of the highest number of posttranscriptional splicing events within the reproductive tract, and skipping exons were the most frequently occurring class of alternative splicing (AS) among the reproductive tract. Statistical analysis revealed 86, 229, and 6 DAS events in the testis/epididymis, testis/ductus deferens, and epididymis/ductus deferens comparison, respectively. Alternative splicing was found to be a mechanism of gene expression regulation within the turkey reproduction tract. In testis, modification was observed for spermatogenesis specific genes; the changes in 5' UTR could act as regulator of MEIG1 expression (a player during spermatocytes meiosis), and modification of 3' UTR led to diversification of CREM mRNA (modulator of gene expression related to the structuring of mature spermatozoa). Sperm tail formation can be regulated by changes in the 5' UTR of testicular SLC9A3R1 and gene silencing by producing dysfunctional variants of ODF2 in the testis and ATP1B3 in the epididymis. Predicted differentially ASGs in the turkey reproductive tract seem to be involved in the regulation of spermatogenesis, including acrosome formation and sperm tail formation and binding of sperm to the zona pellucida. Several ASGs were classified as cilia by actin and microtubule cytoskeleton organization. Such genes may play a role in the organization of sperm flagellum and post-testicular motility development. To our knowledge, this is the first functional investigation of alternatively spliced genes associated with tissue-specific processes in the turkey reproductive tract.

Cu-HKUST-1 and Hydroxyapatite-The Interface of Two Worlds toward the Design of Functional Materials Dedicated to Bone Tissue Regeneration.

A novel composite based on biocompatible hydroxyapatite (HA) nanoparticles and Cu-HKUST-1 (Cu-HKUST-1@HA) has been prepared following a layer-by-layer strategy. Cu-HKUST-1 was carefully selected from a group of four Cu-based metal-organic frameworks as the material with the most promising antimicrobial activity. The formation of a colloidal Cu-HKUST-1 layer on HA nanoparticles was confirmed by various techniques, e.g., infrared spectroscopy, powder X-ray diffraction, N2 sorption, transmission electron microscopy imaging, electron paramagnetic resonance, and X-ray absorption spectroscopy. Importantly, such a Cu-HKUST-1 layer significantly improved the nanomechanical properties of the composite, with Young's modulus equal to that of human cortical bone (13.76 GPa). At the same time, Cu-HKUST-1@HA has maintained the negative zeta potential (-16.3 mV in pH 7.4) and revealed biocompatibility toward human dermal fibroblasts up to a concentration of 1000 µg/mL, without inducing ex vivo hemolysis. Chemical stability studies of the composite over 21 days in a buffer-simulated physiological fluid allowed a detailed understanding of the transformations that the Cu-HKUST-1@HA undergoes over time. Finally, it has been confirmed that the Cu-HKUST-1 layer provides antibacterial properties to HA, and the synergism reached in this way makes it promising for bone tissue regeneration.

Gender Differences in Post-Operative Human Skin.

Although the impact of age, gender, and obesity on the skin wound healing process has been extensively studied, the data related to gender differences in aspects of skin scarring are limited. The present study performed on abdominal human intact and scar skin focused on determining gender differences in extracellular matrix (ECM) composition, dermal white adipose tissue (dWAT) accumulation, and Foxn1 expression as a part of the skin response to injury. Scar skin of men showed highly increased levels of COLLAGEN 1A1, COLLAGEN 6A3, and ELASTIN mRNA expression, the accumulation of thick collagen I-positive fibers, and the accumulation of α-SMA-positive cells in comparison to the scar skin of women. However, post-injured skin of women displayed an increase (in comparison to post-injured men's skin) in collagen III accumulation in the scar area. On the contrary, women's skin samples showed a tendency towards higher levels of adipogenic-related genes (PPARγ, FABP4, LEPTIN) than men, regardless of intact or scar skin. Intact skin of women showed six times higher levels of LEPTIN mRNA expression in comparison to men intact (p < 0.05), men post-injured (p < 0.05), or women post-injured scar (p < 0.05) skin. Higher levels of FOXN1 mRNA and protein were also detected in women than in men's skin. In conclusion, the present data confirm and extend (dWAT layer) the data related to the presence of differences between men and women in the skin, particularly in scar tissues, which may contribute to the more effective and gender-tailored improvement of skin care interventions.

Modification of the Human Amniotic Membrane Using Different Cross-Linking Agents as a Promising Tool for Regenerative Medicine.

Human amniotic membranes (hAMs) obtained during cesarean sections have proven to be clinically useful as an interesting biomaterial in a wide range of tissue engineering applications such as ocular surface reconstruction, burn treatments, chronic wounds, or bedsore ulcers. It presents antimicrobial properties, promotes epithelization, reduces inflammation and angiogenesis, contains growth factors, and constitutes the reservoir of stem cells. However, variability in hAM stiffness and its fast degradation offers an explanation for the poor clinical applications and reproducibility. In addition, the preparatory method of hAM for clinical use can affect its mechanical properties, and these differences can influence its application. As a directly applied biomaterial, the hAM should be available in a ready-to-use manner in clinical settings. In the present study, we performed an analysis to improve the mechanical properties of hAM by the addition of various reagents used as protein cross-linkers: EDC/NHS, PEG-dialdehyde, PEG-NHS, dialdehyde starch, and squaric acid. The effect of hAM modification using different cross-linking agents was determined via infrared spectroscopy, thermal analyses, mechanical properties analyses, enzymatic degradation, and cytotoxicity tests. The use of PEG-dialdehyde, PEG-NHS, dialdehyde starch, and squaric acid increases the mechanical strength and elongation at the breaking point of hAM, while the addition of EDC/NHS results in material stiffening and shrinkage. Also, the thermal stability and degradation resistance were evaluated, demonstrating higher values after cross-linking. Overall, these results suggest that modification of human amniotic membrane by various reagents used as protein cross-linkers may make it easier to use hAM in clinical applications, and the presented study is a step forward in the standardization of the hAM preparation method.