scholarly journals Donor Age and Time in Culture Affect Dermal Fibroblast Contraction in a Hydrogel Skin Graft Model

2021 ◽  
Author(s):  
Amber Detwiler ◽  
Kathryn Polkoff ◽  
Lewis Gaffney ◽  
Donald Freytes ◽  
Jorge Piedrahita
Keyword(s):  
Cellular Proliferation ◽  
Dermal Fibroblast ◽  
Age Groups ◽  
Collagen Matrix ◽  
Scar Tissue ◽  
Dermal Fibroblasts ◽  
Future Research ◽  
Skin Grafts ◽  
Donor Age

Regenerating functional skin without the formation of scar tissue remains an important goal for Tissue Engineering. Current hydrogel-based grafts minimize contraction of full-thickness skin wounds and support skin regeneration using adult or neonatal foreskin dermal fibroblasts, which are often expanded in vitro and used after multiple passages. Based on the known effects of 2D tissue culture expansion on cellular proliferation and gene expression, we hypothesized that differences in donor age and time in culture may also influence the functionality of 3D skin constructs by affecting fibroblast-mediated graft contraction. To validate these predicted differences in fibroblast phenotype and resulting 3D graft model contraction, we isolated porcine dermal fibroblasts of varying donor age for use in a 2D proliferation assay and a 3D cell-populated collagen matrix contractility assay. In 2D cell culture, doubling time remained relatively consistent between all age groups from passage 1 to 6. In the contractility assays, fetal and neonatal groups contracted faster and generated more contractile force than the adult group at passage 1. However, after 5 passages in culture, there was no difference in contractility between groups. These results show how cellular responses differ based on donor age and time in culture, which could account for important differences in biomanufacturing of 3D hydrogel-based skin grafts. Future research and therapies using bioengineered skin grafts should consider how results may vary based on donor age and time in culture before seeding.

scholarly journals Cellular Proliferation of Equine Bone Marrow- and Adipose Tissue-Derived Mesenchymal Stem Cells Decline With Increasing Donor Age

2020 ◽  
Vol 7 ◽  
Author(s):  
Jasmin Bagge ◽  
James N. MacLeod ◽  
Lise C. Berg
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Cellular Proliferation ◽  
Age Groups ◽  
Mrna Levels ◽  
Donor Age ◽  
Autologous Cell

Background: Bone marrow (BM)- and adipose tissue (AT)-derived mesenchymal stem cells (MSCs) are used increasingly for autologous cell therapy in equine practice to treat musculoskeletal and other injuries. Current recommendations often call for 10–100 million MSCs per treatment, necessitating the expansion of primary cells in culture prior to therapeutic use. Of concern, human and rodent studies have shown a decline of both MSC recovery from sampled tissue and in vitro proliferative capacity with increasing donor age. This may be problematic for applications of autologous cell-based therapies in the important equine demographic of older patients.Objectives: To investigate the effect of donor age on the cellular proliferation of equine BM- and AT-MSCs.Study Design:In vitro study.Methods: BM- and AT-MSCs and dermal fibroblasts (biological control) were harvested from horses in five different age groups (n = 4, N = 60); newborn (0 days), yearling (15–17 months), adult (5–8 years), middle-aged (12–18 years), and geriatric (≥22 years). Proliferation of the cells was tested using an EdU incorporation assay and steady state mRNA levels measured for targeted proliferation, aging, and senescence biomarkers.Results: The cellular proliferation of equine BM- and AT-MSCs declined significantly in the geriatric cohort relative to the younger age groups. Proliferation levels in the two MSC types were equally affected by donor age. Analysis of steady state mRNA levels showed an up-regulation in tumor suppressors, apoptotic genes, and multiple growth factors in MSCs from old horses, and a down-regulation of some pro-cycling genes with a few differences between cell types.Main Limitations: Potential age-dependent differences in cell function parameters relevant to cell-therapy application were not investigated.Conclusions: The cellular proliferation of equine BM- and AT-MSCs declined at advanced donor ages. High levels of in vitro proliferation were observed in both MSC types from horses in the age groups below 18 years of age.

scholarly journals A Phytocomplex Consisting of Tropaeolum majus L. and Salvia officinalis L. Extracts Alleviates the Inflammatory Response of Dermal Fibroblasts to Bacterial Lipopolysaccharides

2020 ◽  
Vol 2020 ◽  
pp. 1-14 ◽  
Author(s):  
Tunde Jurca ◽  
Ioana Baldea ◽  
Gabriela Adriana Filip ◽  
Diana Olteanu ◽  
Simona Clichici ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Inflammatory Response ◽  
Dermal Fibroblast ◽  
Physicochemical Characterization ◽  
Antibacterial Activities ◽  
Dermal Fibroblasts ◽  
High Dose ◽  
Cytokine Levels ◽  
Bacterial Lipopolysaccharides

Background. The antimicrobial activity and effects of a phytocomplex consisting of Tropaeolum flos (T) and Salviae folium (S) extracts on the cytokine levels and transcription factors on dermal fibroblast BJ exposed to bacterial lipopolysaccharides were examined. Methods. In order to select the most optimal combination ratio of the two extracts for using in vitro, the physicochemical characterization of vegetal extract mixtures was performed and the antioxidant and antibacterial activities were evaluated on five different formulations of T : S, namely, 1 : 1, 1 : 2, 2 : 1, 3 : 1, and 1 : 3. The best combination of bioactive compounds with regard to antioxidant and antibacterial activities (T : S 1 : 2) was selected for in vitro evaluation of the anti-inflammatory effect. Human dermal fibroblast BJ cells were treated with two doses of the extract mixture and then exposed to bacterial lipopolysaccharides (LPS). The levels of the cytokines involved in inflammatory response, namely, interleukin- (IL-) 6, tumor necrosis factor- (TNF-) α, IL-31, and IL-33, were quantified by ELISA, and the expression of transcription factors, namely, signal transducer and activator of transcription (STAT) 3, nuclear factor kappa B (NFκB), and phosphorylated NFκB (pNFκB), were evaluated by western blot analysis. Results. The results have shown that the mixture of T : S 1 : 2 exhibited significant antibacterial effects on Staphylococcus aureus ATCC 25923. LPS exposure increased the cytokine levels in BJ cells and enhanced the NFκB expression. The pretreatment of BF cells exposed to LPS with the two doses of the extract mixture markedly inhibited the increase of IL-33 and TNF-α levels and amplified the NFκB expression and its activation, especially with the high dose. The low doses of the extract reduced NFκB expression but increased its activation. Conclusions. These experimental findings suggest that the mixture of T : S 1 : 2 can exert some protection against bacterial infections and inflammation induced by LPS in BJ cells being a good therapeutic option in related conditions associated with inflammation.

268 INFLUENCE OF DONOR AGE ON DEVELOPMENTAL COMPETENCE OF IN VITRO v. IN VIVO MATURED BOVINE OOCYTES OBTAINED BY REPEATED OVUM PICKUP FROM FOLLICLE-STIMULATING-HORMONE-STIMULATED AND NONSTIMULATED ANIMALS

2011 ◽  
Vol 23 (1) ◽  
pp. 232
Author(s):  
M. Matthiesen ◽  
H. D. Reichenbach ◽  
F. A. Habermann ◽  
M. Reichenbach ◽  
G. J. Arnold ◽  
...  
Keyword(s):  
Mixed Model ◽  
Ovarian Function ◽  
Age Groups ◽  
Developmental Competence ◽  
Reproductive Aging ◽  
Donor Age ◽  
Ovarian Aging ◽  
Bovine Oocytes

Recent findings on oogenesis, folliculogenesis, and ovarian aging in cows make the bovine system an attractive model for elucidating ovarian function and dysfunction as well as reproductive aging in women. The aim of the present study was to investigate the influence of donor age on the developmental competence of in vitro v. in vivo matured bovine cumulus–oocyte complexes (COC) obtained by ultrasound-guided repeated ovum pickup (OPU). Two groups (G1 and G2) of German Simmental heifers (14 months old at the beginning of the experiment, n = 5 and n = 7), first-lactation young cows (2–4 y old, n = 5 and n = 3), and old cows (10–15 y old, n = 5 and n = 3) were subjected to twice-weekly OPU without hormonal prestimulation 32 (G1) and 6 times (G2). Afterward, animals in G1 were punctured at 5-week intervals 9 times after FSH superstimulation to obtain in vivo matured COC at the metaphase II stage. Data were analysed using a mixed model (SAS). In the twice-weekly OPU for G1 and G2 combined, significantly (P < 0.05) more COC per animal and OPU session were obtained from the old cows (9.9 ± 1.0) compared with heifers and young cows (6.0 ± 0.8 and 7.0 ± 1.0, respectively). When G1 and G2 were regarded separately, lower numbers of COC (P < 0.01) were obtained in G1 than in G2 (2.7 ± 0.8, 4.4 ± 0.8, 7.0 ± 0.8 and 9.2 ± 1.5, 9.4 ± 2.3, 12.9 ± 2.3 for heifers, young cows, and old cows of G1 and G2, respectively). Cleavage rates (CR) on day 3 after IVF (day 0) were not affected by donor age and were not different between groups. Cultivation of COC from young cows in G1 led to higher blastocyst rates (BR) on day 7 (P < 0.05) compared with old cows and heifers. No differences in BR were observed between animals of G2. Significantly more COC (P < 0.01) were obtained in all age groups from FSH superstimulated donors (10.6 ± 0.8, 9.0 ± 0.9, and 11.7 ± 0.9 for heifers, young cows, and old cows, respectively). Cleavage rates and BR were significantly higher (P < 0.05) in all age groups after FSH superstimulation compared with those of nonstimulated donors. However, there were no differences in CR and BR between age groups (CR: 82.8 ± 7.0, 89.9 ± 7.0, 77.1 ± 6.2%; BR: 34.4 ± 7.2, 44.6 ± 7.2, 36.7 ± 7.2%). We conclude that although the numbers of COC obtained per animal and session were significantly different between G1 and G2, in vitro results were highly repeatable after OPU without hormonal prestimulation. Higher CR and BR were obtained after IVF of in vivo matured COC obtained from FSH superstimulated donors, regardless of animal age. This work was supported by the Deutsche Forschungsgemeinschaft (FOR 1041).

Evaluation of Gene Expression and In Vitro Enzyme Study for Antiaging Effect of Crocin and Lutein

2020 ◽  
Vol 10 (5) ◽  
pp. 595-604
Author(s):  
Archana A. Naik ◽  
Chhaya H. Gadgoli ◽  
Arvind B. Naik
Keyword(s):  
Gene Expression ◽  
Cell Line ◽  
Dermal Fibroblast ◽  
Dermal Fibroblasts ◽  
Major Constituent ◽  
Chromatographic Technique ◽  
Gene Expression Study ◽  
Expression Study ◽  
Enzyme Study

Background: Tubular calyx of flowers of Nyctanthes arbour-tristis contains an apocarotenoid crocin, a major constituent present in saffron stigma. The flowers of N. arbortristis are readily available, hence can be an economic substitute for saffron. Lutein from flowers of Tagetes patula, is another carotenoid which is a popular antioxidant. Objective: Oxidative stress is a major contributor to the process of aging. Carotenoids are powerful antioxidants. Hence, the study was carried out to evaluate anticollagenase activity and antielastase activity using gene expression study in Human dermal fibroblasts. Methods: Crocin was isolated from the tubular calyx of Nyctanthes arbortristis using flash chromatographic technique and lutein was isolated using column chromatography. Anticollagenase and antielastase activity of crocin and lutein were carried out using collagenase from Clostridium histolyticum as enzyme and porcine pancreatic elastase. Cytotoxicity of crocin and lutein was determined in Human Dermal Fibroblast cell line (HDF) through MTT assay. In gene expression study, the HDF Cell line was inoculated with Crocin (450 and 250 ppm) and lutein (100 and 50 ppm) separately for 24 hrs and the m-RNA expression levels of COL Type-1 and elastin were determined using RT-PCR. The results were compared with standards. Result: Crocin and lutein both showed inhibition of collagenase and elastase enzyme which are responsible for aging process. The cytotoxic concentration CTC 50 (ppm) for Crocin and lutein was found to be 790.2 ppm and 137.14 ppm. Gene expression study on crocin rich extract of Nyctanthes arbortristis showed upregulation of both collagen and elastin gene whereas lutein rich extract having concentration100 μg/ml showed up regulation by 0.02 fold and concentration 50 μg/ml showed down regulation. Conclusion: In vitro collagenase and elastase enzyme study and Gene expression study showed that these carotenoids are potential antiageing agents which can be substituted to synthetic cosmeceuticals as well as saffron.

scholarly journals Enhancement Of Dermal Fibroblast Isolation Method

2015 ◽  
Vol 16 (1) ◽  
pp. 65-69
Author(s):  
Milan Zaric ◽  
Ivana Nikolic ◽  
Ivanka Zelen ◽  
Marina Mitrovic ◽  
Zoran Milosavljevic
Keyword(s):  
Dermal Fibroblast ◽  
Petri Dish ◽  
Culture Conditions ◽  
Dermal Fibroblasts ◽  
Enzyme Digestion ◽  
Digestion Method ◽  
Primary Isolation ◽  
Donor Cells ◽  
Cell Culture Conditions

ABSTRACTCultivated fibroblasts have been widely used in a large number of in vitro studies. Although they readily proliferate under cell culture conditions, improvements in methods for their isolation are necessary. Here, we present our modified enzyme digestion method and compare its efficiency with commonly used techniques.Three foreskin samples from young, middle-aged and old donors were used. The classical explant, standard enzyme digestion method with collagenase and our improved enzyme digestion method were compared for efficiency of fibroblast isolation and the time needed to achieve 95% confluence in a 30-mm Petri dish.The explant method was the slowest to achieve fibroblast confluence, especially with the tissues from the older donors (up to 23 days). With the standard enzyme digestion method, the skin tissue was partially digested, but the fibroblasts reached confluence much faster (the younger donor cells needed approximately 7 days to reach confluence). Our modified “mixed” enzyme digestion method was the fastest (the fibroblasts from the younger donors needed up to 5 days to reach confluence).For studies requiring the primary isolation and cultivation of dermal fibroblasts, the best method to achieve this goal is the tissue digestion method with the multiple enzyme solution.

scholarly journals Loss of the Desmosomal Component Perp Impairs Wound HealingIn Vivo

2010 ◽  
Vol 2010 ◽  
pp. 1-11 ◽  
Author(s):  
Veronica G. Beaudry ◽  
Rebecca A. Ihrie ◽  
Suzanne B. R. Jacobs ◽  
Bichchau Nguyen ◽  
Navneeta Pathak ◽  
...  
Keyword(s):  
Wound Healing ◽  
Cell Adhesion ◽  
Wound Closure ◽  
Cellular Proliferation ◽  
Dermal Fibroblasts ◽  
Conditional Knockout ◽  
Keratinocyte Proliferation ◽  
Complex Biological Process ◽  
Cell Cell

Epithelial wound closure is a complex biological process that relies on the concerted action of activated keratinocytes and dermal fibroblasts to resurface and close the exposed wound. Modulation of cell-cell adhesion junctions is thought to facilitate cellular proliferation and migration of keratinocytes across the wound. In particular, desmosomes, adhesion complexes critical for maintaining epithelial integrity, are downregulated at the wound edge. It is unclear, however, how compromised desmosomal adhesion would affect wound reepithelialization, given the need for a delicate balance between downmodulating adhesive strength to permit changes in cellular morphology and maintaining adhesion to allow coordinated migration of keratinocyte sheets. Here, we explore the contribution of desmosomal adhesion to wound healing using mice deficient for the desmosomal component Perp. We find thatPerpconditional knockout mice display delayed wound healing relative to controls. Furthermore, we determine that while loss of Perp compromises cell-cell adhesion, it does not impair keratinocyte proliferation and actually enhances keratinocyte migration inin vitroassays. Thus, Perp's role in promoting cell adhesion is essential for wound closure. Together, these studies suggest a role for desmosomal adhesion in efficient wound healing.

Correlation of donor age and telomerase activity with in vitro cell growth and replicative potential for dermal fibroblasts and keratinocytes

2009 ◽  
Vol 18 (4) ◽  
pp. 109-116 ◽  
Author(s):  
M.H. Ng ◽  
B.S. Aminuddin ◽  
S. Hamizah ◽  
C. Lynette ◽  
A.L. Mazlyzam ◽  
...  
Keyword(s):  
Cell Growth ◽  
Telomerase Activity ◽  
Dermal Fibroblasts ◽  
Donor Age

Ultrastructural and metabolic effects of products of activated mononuclear leucocytes on cultured human dermal fibroblasts

1984 ◽  
Vol 42 ◽  
pp. 682-683
Author(s):  
I. Stachura ◽  
Matias Pardo ◽  
Jennifer Worrall ◽  
Theresa L. Whiteside
Keyword(s):  
Plasma Membranes ◽  
Dermal Fibroblast ◽  
Human Dermal Fibroblast ◽  
Dermal Fibroblasts ◽  
Metabolic Effects ◽  
Normal Human Skin ◽  
Normal Human ◽  
Mononuclear Leucocytes ◽  
Distinctive Phenotype

Products of antigen- or mitogen-activated mononuclear leucocytes (ML) are known to modulate fibroblast proliferation and collagen production in vitro. In tissue, ML accumulate at sites of inflammation and are probably involved in the process of fibrosis. We have established that supernatants (SN) of concanavalin A-activated ML increase synthesis of glycosaminoglycan (GAG) in human dermal fibroblast (DF) cultures). When explants of normal human skin were cultured in CMRL 1066 medium supplemented with 15% pooled human serum and containing MLSN the outgrowing DF acquired a distinctive phenotype. In comparison to control DF, the cells treated with MLSN exhibited a marked increase in the number of intracytoplasmic organelles especially dilated cisternae of RER filled with electrondense material, abundant lysosomes, prominent Golgi apparatus and bundles of microfilaments often extending beyond the cell boundaries. Cell surfaces were shaggy and floccular material accumulated in patches along the plasma membranes.

scholarly journals In Vitro Cytotoxic Protective Effect of Alginate-Encapsulated Capsaicin Might Improve Skin Side Effects Associated with the Topical Application of Capsaicin

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1455
Author(s):  
Ariana Hudita ◽  
Bianca Galateanu ◽  
Marieta Costache ◽  
Carolina Negrei ◽  
Rodica-Mariana Ion ◽  
...  
Keyword(s):  
Neuropathic Pain ◽  
Side Effects ◽  
Cell Viability ◽  
Dermal Fibroblast ◽  
Expression Profiles ◽  
Control Culture ◽  
Dermal Fibroblasts ◽  
Diabetic Patients ◽  
Cytotoxic Effects

Chronic neuropathic pain, particularly peripheral pain, is a cause of great concern for diabetic patients. Current treatments include numerous agents such as capsaicinoids, a known deterrent of neuropathic pain despite the inconvenience associated with local side effects. In this context, the current work aims to elucidate the potential mechanisms involved in cytotoxicity by capsaicin and proposes an efficient formulation of capsaicin in alginate microcapsules, which significantly reduces side effects from capsaicin topical administration. For this, human dermal fibroblast cells were treated with alginate-microencapsulated capsaicin extracts and screened for potential cytotoxic effects produced by the treatment. Cell viability and morphology were examined, as well as oxidative stress status and anti-inflammatory potential. Our results show that the alginate encapsulated formulation of capsaicin exerted lower cytotoxic effects on human dermal fibroblasts as measured by cell viability and reactive oxygen species (ROS) production. Furthermore, the expression profiles of inflammatory cytokines were significantly altered by the treatment as compared with the control culture.

scholarly journals In Vitro Differentiation Potential of Human Placenta Derived Cells into Skin Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Ruhma Mahmood ◽  
Mahmood S. Choudhery ◽  
Azra Mehmood ◽  
Shaheen N. Khan ◽  
Sheikh Riazuddin
Keyword(s):  
Stem Cells ◽  
Umbilical Cord ◽  
Human Placenta ◽  
Morphological Changes ◽  
Dermal Fibroblast ◽  
Placental Tissue ◽  
Dermal Fibroblasts ◽  
Differentiation Potential ◽  
Umbilical Cord Tissue

Skin autografting is the most viable and aesthetic technique for treatment of extensive burns; however, this practice has potential limitations. Harvesting cells from neonatal sources (such as placental tissue) is a simple, inexpensive, and noninvasive procedure. In the current study authors sought to evaluate in vitro potential of human placenta derived stem cells to develop into skin-like cells. After extensive washing, amniotic membrane and umbilical cord tissue were separated to harvest amniotic epithelial cells (AECs) and umbilical cord mesenchymal stem cells (UC-MSCs), respectively. Both types of cells were characterized for the expression of embryonic lineage markers and their growth characteristics were determined. AECs and UC-MSCs were induced to differentiate into keratinocytes-like and dermal fibroblasts-like cells, respectively. After induction, morphological changes were detected by microscopy. The differentiation potential was further assessed using immunostaining and RT-PCR analyses. AECs were positive for cytokeratins and E-Cadherin while UC-MSCs were positive for fibroblast specific makers. AECs differentiated into keratinocytes-like cells showed positive expression of keratinocyte specific cytokeratins, involucrin, and loricrin. UC-MSCs differentiated into dermal fibroblast-like cells indicated expression of collagen type 3, desmin, FGF-7, fibroblast activation protein alpha, procollagen-1, and vimentin. In conclusion, placenta is a potential source of cells to develop into skin-like cells.

Sign in / Sign up

Export Citation Format

Share Document