Induced Pluripotent Stem Cells from Diabetic Foot Ulcer Fibroblasts


Dr. Jonathan Garlick is professor of Oral Pathology at Tufts University and has achieved some notoriety among stem cell scientists by publishing a stem-cell rap on You Tube to teach people about the importance of stem cells.

Garlick and his colleagues have published a landmark paper in the journal Cellular Reprogramming in which cells from diabetic patients were reprogrammed into induced pluripotent stem cells (iPSCs).

Garlick and his colleagues have established, for the first time, that skin cells from diabetic foot ulcers can be reprogrammed iPSCs. These cells can provide an excellent model system for diabetic wounds and may also used, in the future, to treat chronic wounds.

ESC and iPSCs differentiation to fibroblast fate. ESC and iPSC were differentiated and monitored at various stages of differentiation. Representative images show the morphology of ESC and 2 iPSC lines after days 1, 4, 7, 10, 14, 21 and 28 of differentiation. Early morphologic changes showed differentiation beginning at the periphery of colonies (day 1). At later stages cells acquired fibroblast features of elongated, stellate cells (day 10 at days 21 and 28 of differentiation.
ESC and iPSCs differentiation to fibroblast fate. ESC and iPSC were differentiated and monitored at various stages of differentiation. Representative images show the morphology of ESC and 2 iPSC lines after days 1, 4, 7, 10, 14, 21 and 28 of differentiation. Early morphologic changes showed differentiation beginning at the periphery of colonies (day 1). At later stages cells acquired fibroblast features of elongated, stellate cells (day 10 at days 21 and 28 of differentiation.

Garlick’s team at Tufts University School of Dental Medicine and the Sackler School of Graduate Biomedical Sciences at Tufts, have also used their diabetic-derived iPSCs to show that a protein called fibronectin is linked to a breakdown in the wound-healing process in cells from diabetic foot ulcers.

One of the goals of Garlick’s research is to develop efficient protocols to make functional cell types from iPSCs and to use them to generate 3D tissues that demonstrate a broad range of biological functions. His goal is to use the 3D model system to develop human therapies to replace or regenerate damaged human cells and tissues and restore their normal function.

In this paper, Garlick and his colleagues showed that not only can fibroblasts from diabetic wounds form iPSCs, but they can also participate in 3D skin-like tissues. This model system is more than a disease-in-a-dish system but disease-in-a-tissue system.

Fabrication of three-dimensional tissue construction. (A) A collagen gel embedded with human dermal fibroblasts is layered onto a polycarbonate membrane. (B) After dermal fibroblasts contract and remodel the collagen matrix, keratinocytes are then seeded onto it to create a monolayer that will form the basal layer of the tissue. (C) Tissues are raised to an air-liquid interface to initiate tissue development that mimics in vivo skin.
Fabrication of three-dimensional tissue construction. (A) A collagen gel embedded with human dermal fibroblasts is layered onto a polycarbonate membrane. (B) After dermal fibroblasts contract and remodel the collagen matrix, keratinocytes are then seeded onto it to create a monolayer that will form the basal layer of the tissue. (C) Tissues are raised to an air-liquid interface to initiate tissue development that mimics in vivo skin. From this site.

“The results are encouraging. Unlike cells taken from healthy human skin, cells taken from wounds that don’t heal – like diabetic foot ulcers – are difficult to grow and do not restore normal tissue function,” said Garlick. “By pushing these diabetic wound cells back to this earliest, embryonic stage of development, we have ‘rebooted’ them to a new starting point to hopefully make them into specific cell types that can heal wounds in patients suffering from such wounds.”

Scientists in Garlick’s laboratory used these 3D tissues to test the properties of cells from diabetic foot ulcers and found that cells from the ulcers get are not able to advance beyond synthesizing an immature scaffold made up predominantly of a protein called fibronectin.  Fibronectin, unfortunately, seems to prevent proper closure of wounds.

Fibronectin Sigma

Fibronectin has been shown to be abnormal in other diabetic complications, such as kidney disease, but this is the first study that directly connects it to cells taken from diabetic foot ulcers.

Deriving more effective therapies for foot ulcers has been slow going because of a lack of realistic wound-healing models that mimic the extracellular matrices of human tissues. This scaffolding is critical for wound repair in skin, and other tissue as well.

The work in this paper builds on earlier experiments that showed that cells from diabetic ulcers have fundamental defects that can be simulated using laboratory-grown 3D tissue models. These 3D models will almost certainly be a good model system to test new therapeutics that could improve wound healing and prevent those limb amputations that result when treatments fail.

Garlick’s 3D model will allow him and other researchers to push these studies forward. Can they differentiate their cells into more mature cell types that can be studied in 3D models to see if they will improve healing of chronic wounds?

More than 29 million Americans have diabetes. Diabetic foot ulcers, often resistant to treatment, are a major complication. The National Diabetes Statistics Report of 2014 stated that about 73,000 non-traumatic lower-limb amputations in 2010 were performed in adults aged 20 years or older with diagnosed diabetes, and approximately 60 percent of all non-traumatic lower-limb amputations occur in people with diabetes.

This paper appeared in: Behzad Gerami-Naini, et al., Cellular Reprogramming. June 2016, doi:10.1089/cell.2015.0087.

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mburatov

Professor of Biochemistry at Spring Arbor University (SAU) in Spring Arbor, MI. Have been at SAU since 1999. Author of The Stem Cell Epistles. Before that I was a postdoctoral research fellow at the University of Pennsylvania in Philadelphia, PA (1997-1999), and Sussex University, Falmer, UK (1994-1997). I studied Cell and Developmental Biology at UC Irvine (PhD 1994), and Microbiology at UC Davis (MA 1986, BS 1984).