scholarly journals Reduced serum methods for contact-based coculture of human dermal fibroblasts and epidermal keratinocytes

BioTechniques ◽  
2020 ◽  
Vol 69 (5) ◽  
pp. 347-355
Author(s):  
Snehal Kadam ◽  
Madhusoodhanan Vandana ◽  
Karishma S Kaushik
Keyword(s):  
Wound Closure ◽  
Fetal Bovine Serum ◽  
Cell Types ◽  
High Serum ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Human Dermal Fibroblasts ◽  
Fetal Bovine ◽  
High Concentrations ◽  
Serum Free Conditions

Direct contact-based coculture of human dermal fibroblasts and epidermal keratinocytes has been a long-standing and challenging issue owing to different serum and growth factor requirements of the two cell types. Existing protocols employ high serum concentrations (up to 10% fetal bovine serum), complex feeder systems and a range of supplemental factors. These approaches are technically demanding and labor intensive, and pose scientific and ethical limitations associated with the high concentrations of animal serum. On the other hand, serum-free conditions often fail to support the proliferation of one or both cell types when they are cultured together. We have developed two reduced serum approaches (1–2% serum) that support the contact-based coculture of human dermal fibroblasts and immortalized keratinocytes and enable the study of cell migration and wound closure.

scholarly journals To Serum or Not to Serum: Reduced-serum based methods for contact-based co-culture of human dermal fibroblasts (HDFa) and epidermal keratinocytes (HaCaT) for wound bed studies

2020 ◽  
Author(s):  
Snehal Kadam ◽  
Madhusoodhanan Vandana ◽  
Karishma Kaushik
Keyword(s):  
Direct Contact ◽  
Cell Types ◽  
High Serum ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Human Dermal Fibroblasts ◽  
Serum Concentrations ◽  
High Concentrations ◽  
Serum Free Conditions ◽  
Structure And Functions

Abstract Background: Contact-based co-culture of human dermal fibroblasts and epidermal keratinocytes is important to study wound bed structure and functions. Co-culture of these two cell types in direct contact with each other has been a long-standing and challenging issue, owing to specific and different serum and growth factor requirements of the two cell types. Existing protocols employ high-serum concentrations (up to 10% FBS), complex cell feeder systems, and a range of supplemental factors. These approaches are not only technically demanding, labor- and material-intensive but also pose scientific and ethical limitations associated with high concentrations of animal serum. On the other hand, serum-free conditions often fail to support the attachment and proliferation of one or both cell types when cultured together. Results: Given this, we have developed two reduced-serum based approaches (1-2% serum), using commonly-available media components, to support the contact-based co-culture of HDFa and HaCaT cells. Using these approaches, HDFa and HaCaT were co-cultured by layering keratinocytes over confluent fibroblasts or by co-seeding the two cell types simultaneously. Under these conditions, both cell types showed robust attachment and proliferation, and characteristic cellular morphology in co-culture. Further, these co-cultured platforms enabled the study of important wound bed functions such as cell migration and wound closure.Conclusions: We believe that these methods can be leveraged for a range of wound and skin studies, and tissue bioengineering applications, potentially reducing concerns with high-serum formulations.

scholarly journals To Serum or Not to Serum: Reduced-serum Based Approaches for Contact-based Co-culture of Fibroblasts and Keratinocytes for Wound Bed Studies

2020 ◽  
Author(s):  
Snehal Kadam ◽  
Madhusoodhanan Vandana ◽  
Karishma Kaushik
Keyword(s):  
Wound Closure ◽  
Culture Media ◽  
Cell Types ◽  
High Serum ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Serum Concentrations ◽  
Human Epidermal Keratinocytes ◽  
Cell Culture Media ◽  
Structure And Functions

Contact-based co-culture of fibroblasts and keratinocytes is important to study the structure and functions of the wound bed. Co-culture of these two cell types in direct contact with each other has been challenging, requiring high serum concentrations (up to 10%), feeder systems and a range of supplemental factors. These approaches are not only technically demanding, but also present scientific, cost and ethical limitations associated with high-serum concentrations. We have developed two reduced-serum approaches (1-2%) to support contact-based co-culture of human dermal fibroblasts (HDFa) and human epidermal keratinocytes (HaCaT). The two approaches include (1) Specialized cell culture media for each cell type mixed in a 1:1 ratio (KGM+FGM), and (2) Minimal media supplemented with cell-specific growth factors (MEM+GF). Co-culture could be successfully achieved by co-seeding (two cell types were introduced simultaneously), or in a layered fashion (keratinocytes seeded on top of confluent fibroblasts). With wound scratch assays, the co-cultured platforms could demonstrate cell proliferation, migration and wound closure. The reduced-serum conditions developed are simple, easy to formulate and adopt, and based on commonly-available media components. These contact-based co-culture approaches can be leveraged for wound and skin studies, and tissue bioengineering applications, potentially reducing concerns with high-serum formulations.

scholarly journals To Serum or Not to Serum: Reduced-serum Based Approaches for Contact-based Co-culture of Fibroblasts and Keratinocytes for Wound Bed Studies

2020 ◽  
Author(s):  
Snehal Kadam ◽  
Madhusoodhanan Vandana ◽  
Karishma Kaushik
Keyword(s):  
Wound Closure ◽  
Culture Media ◽  
Cell Types ◽  
High Serum ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Serum Concentrations ◽  
Human Epidermal Keratinocytes ◽  
Cell Culture Media ◽  
Structure And Functions

Contact-based co-culture of fibroblasts and keratinocytes is important to study the structure and functions of the wound bed. Co-culture of these two cell types in direct contact with each other has been challenging, requiring high serum concentrations (up to 10%), feeder systems and a range of supplemental factors. These approaches are not only technically demanding, but also present scientific, cost and ethical limitations associated with high-serum concentrations. We have developed two reduced-serum approaches (1-2%) to support contact-based co-culture of human dermal fibroblasts (HDFa) and human epidermal keratinocytes (HaCaT). The two approaches include (1) Specialized cell culture media for each cell type mixed in a 1:1 ratio (KGM+FGM), and (2) Minimal media supplemented with cell-specific growth factors (MEM+GF). Co-culture could be successfully achieved by co-seeding (two cell types were introduced simultaneously), or in a layered fashion (keratinocytes seeded on top of confluent fibroblasts). With wound scratch assays, the co-cultured platforms could demonstrate cell proliferation, migration and wound closure. The reduced-serum conditions developed are simple, easy to formulate and adopt, and based on commonly-available media components. These contact-based co-culture approaches can be leveraged for wound and skin studies, and tissue bioengineering applications, potentially reducing concerns with high-serum formulations.

scholarly journals Extracellular-Vesicle-Based Coatings Enhance Bioactivity of Titanium Implants—SurfEV

Nanomaterials ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1445
Author(s):  
Taisa Nogueira Pansani ◽  
Thanh Huyen Phan ◽  
Qingyu Lei ◽  
Alexey Kondyurin ◽  
Bill Kalionis ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Titanium Surface ◽  
Wound Closure ◽  
Cell Types ◽  
Extracellular Vesicle ◽  
Tissue Growth ◽  
Titanium Implants ◽  
The Body ◽  
High Concentrations ◽  
And Migration

Extracellular vesicles (EVs) are nanoparticles released by cells that contain a multitude of biomolecules, which act synergistically to signal multiple cell types. EVs are ideal candidates for promoting tissue growth and regeneration. The tissue regenerative potential of EVs raises the tantalizing possibility that immobilizing EVs on implant surfaces could potentially generate highly bioactive and cell-instructive surfaces that would enhance implant integration into the body. Such surfaces could address a critical limitation of current implants, which do not promote bone tissue formation or bond bone. Here, we developed bioactive titanium surface coatings (SurfEV) using two types of EVs: secreted by decidual mesenchymal stem cells (DEVs) and isolated from fermented papaya fluid (PEVs). For each EV type, we determined the size, morphology, and molecular composition. High concentrations of DEVs enhanced cell proliferation, wound closure, and migration distance of osteoblasts. In contrast, the cell proliferation and wound closure decreased with increasing concentration of PEVs. DEVs enhanced Ca/P deposition on the titanium surface, which suggests improvement in bone bonding ability of the implant (i.e., osteointegration). EVs also increased production of Ca and P by osteoblasts and promoted the deposition of mineral phase, which suggests EVs play key roles in cell mineralization. We also found that DEVs stimulated the secretion of secondary EVs observed by the presence of protruding structures on the cell membrane. We concluded that, by functionalizing implant surfaces with specialized EVs, we will be able to enhance implant osteointegration by improving hydroxyapatite formation directly at the surface and potentially circumvent aseptic loosening of implants.

Fetuin: its enigmatic property of growth promotion

1992 ◽  
Vol 263 (3) ◽  
pp. C551-C562 ◽  
Author(s):  
Z. Nie
Keyword(s):  
Fetal Bovine Serum ◽  
Growth Promotion ◽  
Cell Types ◽  
Specific Cell ◽  
Biological Functions ◽  
Cellular Attachment ◽  
Fetal Bovine ◽  
Growth Promoting ◽  
Free Media ◽  
Inhibit Trypsin Activity

A variety of cell types in culture respond to fetuin, a glycoprotein from fetal bovine serum, which is often an important supplement to many serum-free media. Bovine fetuin preparation has been shown to inhibit trypsin activity and promote cellular attachment, growth, and differentiation in many different culture systems. In addition, fetuin associates with various growth factors or growth-promoting substances. However, whether the growth-promoting activity of fetuin preparation is due to fetuin per se or to its minor contaminant(s) has been a long-standing puzzle. The present review surveys the literature concerning this enigmatic property of fetuin and summarizes three possibilities: 1) fetuin itself is active, although the majority of studies do not support this; 2) various contaminants of fetuin preparations, including potentially unidentified ones, are responsible for the activity, a possibility supported by numerous reports; and 3) one of the fetuin subspecies, one of its contaminants, or a combination of both of these is responsible for growth of a specific cell type. In addition, the basic physicochemical properties and other biological functions of fetuin have also been presented.

Effect of fetal bovine serum on the growth and survival of insect cell cultures

1970 ◽  
Vol 48 (3) ◽  
pp. 427-432 ◽  
Author(s):  
S. S. Sohi ◽  
Cheryl Smith
Keyword(s):  
Adverse Effect ◽  
Cell Lines ◽  
Insect Cell ◽  
Bovine Serum ◽  
Fetal Bovine Serum ◽  
Maximum Growth ◽  
Growth And Survival ◽  
Fetal Bovine ◽  
High Concentrations ◽  
Insect Cell Lines

The effect of different concentrations of fetal bovine serum (FBS) on the growth and survival of three insect cell lines—Aedes aegypti, Antheraea eucalypti, and Bombyx mori—was studied in a series of 10 experiments. Cell viability was low and no growth occurred in any of the cell lines when FBS was omitted from the medium. Maximum growth of A. aegypti cells was obtained with 10% FBS. There was no further increase in this growth when FBS was increased to 20 and 30%, and neither did the increased concentrations have any appreciable adverse effect on the growth or survival of these cells. Maximum growth of A. eucalypti and B. mori cells was obtained in 5% FBS; the growth of these cells was significantly less in 20 and 30% FBS. Viability of A. eucalypti cells was quite low in 20 and 30% FBS. There was, however, no adverse effect on the viability of B. mori cells at these high concentrations.

scholarly journals The Imbalance of MMP-2/TIMP-2 and MMP-9/TIMP-1 Contributes to Collagen Deposition Disorder in Diabetic Non-Injured Skin

2021 ◽  
Vol 12 ◽  
Author(s):  
Peng Zhou ◽  
Chao Yang ◽  
Shan Zhang ◽  
Zun-Xiang Ke ◽  
Dian-Xi Chen ◽  
...  
Keyword(s):  
High Glucose ◽  
Cell Types ◽  
Collagen Type I ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Type I ◽  
Collagen Deposition ◽  
Skin Cell ◽  
Treatment Of Diabetes ◽  
Injured Skin

The importance of the early diagnosis and treatment of diabetes and its cutaneous complications has become increasingly recognized. When diabetic non-injured skin was stained with Masson’s trichrome, its dermal collagen was found to be disordered, its density was variable, and it was dispersed or arranged in vague fascicles. The collagen type I sequencing results of RNA sequencing-based transcriptome analysis of three primary human skin cell types—dermal fibroblasts, dermal microvascular endothelial cells, and epidermal keratinocytes—under high glucose were analyzed. The results showed that both COL1A1 and COL1A2 mRNA expressions were reduced in human dermal fibroblasts (HDFs). The ratio of matrix metalloproteinase (MMP)-2/tissue inhibitors of metalloproteinase (TIMP)-2 and MMP-9/TIMP-1 in HDFs increased when treated with high glucose. By inhibiting MMP-2 and MMP-9 with SB-3CT, collagen deposition disorder of the skin in streptozotocin-induced diabetes mice was alleviated. The imbalance of MMP2/TIMP2 and MMP9/TIMP1 contributes to the non-injured skin disorder of collagen deposition in diabetes, suggesting a possibility for early treatment of diabetes skin complications.

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