Umbilical Cord Stem Cells Normalize Blood Glucose Levels in Diabetic Mice


Diabetes mellitus results from an insufficiency of insulin (Type 1 diabetes) or an inability to properly respond to insulin (Type 2 diabetes). Type 1 diabetes is caused by an attack by the patient’s own immune system on their pancreatic beta cells, which synthesize and secrete insulin. It is a disease characterized by inflammation in the pancreas. This suggests that abatement of inflammation in the pancreas might provide relief and delay the onset of diabetes.

Mesenchymal stem cells isolated from umbilical cord connective tissue, which is also known as Wharton’s jelly (WJ-MSCs), have the ability to reverse inflammatory destruction and might provide a way to delay or even reverse the onset of Type 1 diabetes.

To test this possibility, Jianxia Hu, Yangang Wang, and their colleagues took 60 non-obese diabetic mice and divided them into four groups: a normal control group, a normal diabetic group, a WJ-MSCs prevention group that was treated with WJ-MSCs before the onset of diabetes, and a WJ-MSCs treatment group that was treated with WJ-MSCs after the onset of diabetes.

After their respective treatments, the onset time of diabetes, levels of fasting plasma glucose (FPG), fed blood glucose levels and C-peptide (an indication of the amount of insulin synthesized), regulation of cytokines, and islet cells were examined and evaluated.

After WJ-MSCs infusion, fasting and fed blood glucose levels in WJ-MSCs treatment group decreased to normal levels in 6-8 days and were maintained for 6 weeks. The levels of fasting C-peptide of the WJ-MSC-treated mice was higher compared to diabetic control mice. In the WJ-MSCs prevention group, WJ-MSCs protected mice from the onset of diabetes for 8-weeks, and the fasting C-peptide in this group was higher compared to the other two diabetic groups.

Other comparisons between the WJ-MSC-treated group and the diabetic control group, showed that levels of regulatory T-cells (that down-regulate autoinflammation), were high and levels of pro-inflammatory molecules such as IL-2, IFN-γ, and TNF-α. The degree of inflammation in the pancreas was also examined, and pancreatic inflammation was depressed, especially in the WJ-MSCs prevention group.

These experiments show that infusions of WJ-MSCs can down-regulate autoimmunity and facilitate the recovery of islet β-cells whether given before or after onset of Type 1 Diabetes Mellitus. THis suggests that WJ-MSCs might be an effective treatment for Type 1 Diabetes Mellitus.

See March 2014 edition of the journal Endocrine.

Artificial Skin Created Using Umbilical Cord Stem Cells


Major burn patients usually must wait weeks for artificial skin to be grown in the laboratory to replace their damaged skin, buy a Spanish laboratory has developed new protocols and techniques that accelerate the growth of artificial skin from umbilical cord stem cells. Such laboratory-grown skin can be frozen and stored in tissue banks and used when needed.

Growing skin in the laboratory requires the acquisition of keratinocytes, those cells that compose the skin and the mucosal covering inside our mouths.  Keratinocytes can be cultured in the laboratory, but they have a long cell cycle, which means that they take a really long time to divide.  Consequently, cell cultures of keratinocytes tend to take a very long time to grow.

Keratinocytes in culture
Keratinocytes in culture

As they grow, the keratinocytes respond to connective tissue underneath them to receive the cues that tell them how to connect with each other and form either skin or oral mucosa.  In patients with severe burns, however, the underlying connective tissue is also often damaged.  Therefore, finding a way to not only accelerate the growth of cultured keratinocytes, but also to provide the underlying structure that directs the cells to form a proper epithelium is essential.

Remember that severe burn patients are living on borrowed time.  Without a proper skin covering, water loss is severe and dehydration is a genuine threat.  Also, infection is another looming threat.  Therefore, the treatment of a burn patient is a race against time.

Because umbilical cord stem cells grow quickly and effectively in culture, they might be able to differentiate into keratinocytes and form the structures associated with oral mucosa and skin.

University of Granada researchers used a new type of epithelial covering to grow their artificial skin in addition to a biomaterial made of fibrin (the stiff, cable-like protein that forms clots) and agarose to provide the underlying connective tissue. In case you might need a refresher, an epithelium refers to a layer of cells that have distinct connects with each other and form a discrete layer. Epithelia can form single or multiple layers and can be composed of long, skinny cells, short, flat cells, or boxy cells.  An epithelium is a membrane-like tissue composed of one or more layers of cells separated by very little intervening substances.  Epithelia cover most internal and external surfaces of the body and its organs.

Previous work from this same research group showed that stem cells from Wharton’s jelly (connective tissue within the umbilical cord), could be converted into epithelial cells. This current study confirms and extends this previous work and applies it to growing skin, and oral mucosa.

“Creating this new type of skin suing stem cells, which can be stored in tissue banks, mains that it can be used instantly when injuries are caused, and which would bring the application of artificial skin forward many weeks,” said Antonio Campos, professor of histology and one of the authors of this study.

By growing the Wharton’s jelly stem cells on their engineered matrix in a three-dimensional culture system, Campos and his colleagues saw that the stem cells stratified (formed layers), and expressed a bunch of genes that are peculiar to skin and other types of epithelia that cover surfaces (e.g., cytokeratins 1, 4, 8, and 13; plakoglobin, filaggrin, and involucrin).  When examined with an electron microscope, the cells had truly formed the kinds of tight connections and junctions that are so common to skin epithelia.

Electron microscopy analysis of controls and three-dimensional bioactive models of H-hOM and H-hS. SEM images (top) corresponding to N-hOM and N-hS controls showed a tight superficial layer of flat polygonal cells with desquamation signs in which cells were covering the entire surface, whereas samples kept in vitro for 2 weeks showed immature differentiation patterns, and samples implanted in vivo for 40 days tended to resemble the structure of the native control tissues, with flattened cells and evident signs of desquamation. Scale bars = 50 μm. TEM samples (bottom) were analyzed after 40 days of in vivo implantation and demonstrated that in vivo-implanted tissues were mature and well-differentiated, with numerous intercellular junctions, abundant cell organelles, and a collagen-rich stroma. Scale bars = 1 μm. Abbreviations: H-hOM, heterotypical human oral mucosa; H-hS, heterotypical human skin; N-hOM, native human oral mucosa; N-hS, native human skin; SEM, scanning electron microscopy; TEM, transmission electron microscopy.
Electron microscopy analysis of controls and three-dimensional bioactive models of H-hOM and H-hS. SEM images (top) corresponding to N-hOM and N-hS controls showed a tight superficial layer of flat polygonal cells with desquamation signs in which cells were covering the entire surface, whereas samples kept in vitro for 2 weeks showed immature differentiation patterns, and samples implanted in vivo for 40 days tended to resemble the structure of the native control tissues, with flattened cells and evident signs of desquamation. Scale bars = 50 μm. TEM samples (bottom) were analyzed after 40 days of in vivo implantation and demonstrated that in vivo-implanted tissues were mature and well-differentiated, with numerous intercellular junctions, abundant cell organelles, and a collagen-rich stroma. Scale bars = 1 μm. Abbreviations: H-hOM, heterotypical human oral mucosa; H-hS, heterotypical human skin; N-hOM, native human oral mucosa; N-hS, native human skin; SEM, scanning electron microscopy; TEM, transmission electron microscopy.

The authors conclude the article with this statement: “All these findings support the idea that HWJSCs could be useful for the development of human skin and oral mucosa tissues for clinical use in patients with large skin and oral mucosa injuries.”  Think of it folks – new skin for burn patients, quickly, safely and ethically.

Now back to reality – this is exciting, but it is a a pre-clinical study.  Larger animals studies must show the efficacy and safety of this protocol before human trials can be considered, but you must admit that it looks exciting; and without killing any embryos.

See I. Garzón, et al., Stem Cells Trans MedAugust 2013 vol. 2 no. 8625-632.

Mesenchymal Stem Cells from Umbilical Cord Promote Repair After Heart Attacks in Minipigs


The umbilical cord contains a major umbilical vein and an umbilical artery, but these blood vessels are embedded in a gel-like matrix called “Wharton’s jelly.” Wharton’s jelly is home to a population of mesenchymal stem cells that have peculiar properties.

You might first say, “what on earth is a mesenchymal stem cell?” Fair enough. Mesenchymal stem cells were first discovered in bone marrow. In bone marrow, mesenchymal stem cells (MSCs) do not make blood cells; that;’s the job of the hematopoietic stem cells (HSCs). MSCs in bone marrow serve an important support role for HSCs in bone marrow. Traditionally, MSCs have the capacity to differentiate into fat cells, bone cells, and cartilage cells. However, further has shown that MSCs can also form a variety of other cell types as well if manipulated in the laboratory. MSCs also express are characteristic cadre of cell surface proteins (CD10, CD13, CD29, CD44, CD90, and CD105 for those who are interested).

MSCs, however, are found in more places that just bone marrow. As it turns out, MSCs have been found in fat, muscle, liver, tendons, synovial membrane (the membranes that surround joints, skin, and so on. Some scientists think that every organ in the body may harbor a MSC population. Furthermore, these MSC populations differ in the genes they express, their capability to differentiate into different cell types, and their cell surface proteins (see this article on this website for a rather exhaustive foray into this topic).

Now that you are more savvy about MSCs, Wharton’s jelly contains a MSC population, but this population seems to have a younger profile than MSCs from other parts of the body.  They are more plastic and more invisible to the immune system than other types of MSCs.  For that reason,  they might be good candidates for treating a sick heart after a heart attack.  A recent paper by Wei Zhang and others from the TEDA International Cardiovascular Hospital and the Tianjin Medical Cardiovascular Clinical College examined the ability of MSCs from the Wharton’s jelly of human umbilical cords to heal the hearts of minipigs after a heart attack.  Oh, before I forget – this paper was published in the journal Coronary Artery Disease.

Twenty-three minipigs were subjected to open-heart surgery and given heart attacks.  Then the pigs were divided into three groups, a control group, a group that received injections of saline into their hearts, and a third group that received injections of 40 million human Wharton’s jelly derived MSCs into the region of the infarct.  The animals were sewn up and given antibiotics to prevent infection.

Six weeks after surgery, each animal was examined by means of Technetium-sestamibi myocardial perfusion imaging, and electrocardiography. For those who do not know what Technetium-sestamibi myocardial perfusion imaging is for, it works like this.  Cardiolite is the trade name of a large, fat-soluble molecule that flows through the heart in a fashion proportion to the blood flow through the heart muscle.  Single photon emission computed tomography or SPECT is used to detect the Cardiolite.    Areas of the heart without blood flow are the regions damaged during the heart attack.  Therefore, this technique is extremely useful to determine the area of damage in the heart.

Cardiolite
Cardiolite

After the animals were examined, they were put down and their hearts were extracted, sectioned, and stained for areas or cell death, and the areas where the injected stem cells resided.  All injected stem cells were labeled before injection so that they were easily detectable.

The results were clear.  The heart injected with MSCs from umbilical cord did not show any decrease in ejection fraction, whereas the other two groups showed an average reduction in injection fraction of around 10%.  In fact the stem cell-injected hearts showed an average 1 % increase in ejection fraction.  The blood flow in the hearts was even more different.  blood flow is measured as a ratio of dead heart tissue to total heart tissue.  The control of saline-injected hearts had an average ratio of about 4%, whereas the stem cell-injected hearts had a slightly negative percentage.  This is a significant difference.   Echocardiography confirmed that the wall thickness of the stem cell-injected hearts was significantly thicker than the walls of the control or the saline-injected hearts; some 14 times thicker!!

When the dissected hearts were examined, the MSC-injected hearts had lots of stem cells still in them.  The cells not only survived, but, according to Zhang and his colleagues, differentiated into heart muscle cells.  Their rationale for this conclusion is three-fold – the cells had the same shape and form or native heart muscle cells, they expressed heart specific Troponin T and vWF proteins, and electrically coupled with other heart muscle cells by expressing connexin.  Connexin is a protein that traverses the membranes of two closely apposed cells and forms small pores between two cells that allows the exchange of SMALL molecules such as ions, ATP, and things like that.  These connexin constructed pores are called “gap junctions” and they are the reason heart muscle cells work as a single unit, since any electrochemical change in one cell immediately spreads to all other nearby, connected cells.

gap junctions

As much as I would like to believe Zhang and his colleagues, I remain skeptical that these cells differentiated into heart muscle cells.  I say this because MSCs can be differentiated in culture to form cells that look and act like heart muscle cells.  These cells will even express some heart-specific genes.  However, they lack the calcium handling machinery of true heart muscle cells and do not function as true heart muscle.  To convince that these Wharton jelly MSCs truly are heart muscle cells, they will need to show that they contain heart specific calcium handling proteins (see Shake JG, Gruber PJ, Baumgartner WA et al. Ann Thorac Surg 2002;73:1919–1925; Davani S, Marandin A, Mersin N et al.  Circulation 2003;108(suppl 1):II253–258; Hou M, Yang KM, Zhang H et al. Int J Cardiol 2007;115:220 –228).  If they can show this, then I will believe them.

However, there are two findings of this paper that are not in doubt.  The number of blood vessels in the hearts of the MSC-treated animals far exceeded the number found in the control or the saline-treated hearts (3-4 times the number of blood vessels).  Therefore, the Wharton’s jelly MSCs induced lots and lots of blood vessels.  Many of these blood vessels contained labeled cells, which shows that the MSCs differentiated into endothelial and smooth muscle cells, Also, the Wharton’s jelly MSCs clearly induced resident cardiac stem cell (CSC) populations in the hearts of the minipigs, since several cells that expressed CSC surface molecules were found in the heart muscle tissue.  Previous work by Hatzistergos and others showed that MSCs induce the endogenous CSC population and this is one of the ways that MSCs help heal ailing hearts (Circulation Research 2010 107:913-22).

Zhang’s paper is interesting and it shows that Wharton’s jelly MSCs are safe and efficacious for treating the heart after a heart attack.  Also, none of the minipigs in this experiment were treated with drugs to suppress the immune system.  No immune response against the cells was reported.  Therefore, the invisibility of these cells to the immune system seems to last, at least in this experiment.