The Importance Of Human Pluripotent Stem Cell

Pluripotent stem cells are the stem cells that can only differentiate into a limited range of differentiated cells. (2) They have the ability to give rise to all somatic cells from ectoderm, mesoderm and endoderm, as well as gametes. Naturally it can be found in embryos as Embryonic stem cells (ES cells). Induced pluripotent stem cells (iPS cells or iPSC) are the pluripotent stem cells that are generated directly from adult cells, first discovered by Shinya Yamanaka in 2006 by using a set of reprogramming factors (Oct4, Sox2, Klf4, and c-Myc or LIN28 and Nanog) (3) to reprogram mature cells back to a pluripotent state (4).

The research of human embryonic stem cells (hESC) is talked about a lot in the field of medical research, not just by the scientists, but also by politicians, religious groups, etc. The discovery of stem cells is known as a medical sensation, with its research having the potential to cure many diseases. But there are still ethical issues standing in the way of this research, and due to this, different parts of the world have different rules and regulations regarding the research of stem cells and their use in cellular therapy, and it is even illegal in a number of countries. Before I dive straight into the advantages and disadvantages of their use in cellular therapy, I should talk about the basics of hESC and how we can make them. Embryonic stem cells are pluripotent, i.e. they can differentiate into any type of cell in our body, and they can replicate indefinitely. We can create an embryonic stem cell line in more than one method, and most of them involve creating a fertilized egg (zygote), which eventually forms into a blastocyst. The inner cell mass of the blastocyst can be transferred onto a petri-dish containing a culture medium, where it is encouraged to divide. The diagram below illustrates this process.

Embryonic stem cells are found in human blastocysts (Marcovitz 17). A blastocyst is a very young embryo (just a few days old) that contains around 200 undifferentiated stem cells (Marcovitz 17). German Zoologist Valentin Hacker coined the term “stem cell” after he discovered them in a blastocyst of a crustacean (Marcovitz 18). Embryonic stem cells were collected for the first time in 1988 by Dr. James Thomson of University of Wisconsin and by Dr. John Gearheart of Johns Hopkins (Panno 76). These stem cells are unspecialized; they do not perform a specific function like cells such as muscle and nerve do (“Stem Cells”). They are also pluripotent, meaning they have the ability to divide and become specialized cells (“Stem Cells”). This is why stem cells hold so

Coat a T75-cm2 culture flasks with 0.2% gelatin to provide better adhesion surface for mEFs.

Over time, the use of human stem cells has proven to be monumental in research, particularly in the field of disease modeling and drug development for treatments of diseases. However, there are many barriers that come with using human stem cells, specifically embryonic stem cells (ESCs), mainly due to the fact that they must come from either

Human embryonic stem cells (hESCs) are pluripotent and are obtained from the inner mass of a 4-5 day old human blastocyst that consists of approximately 100 cells (“Stem cell research,” 2009).

Picture 3. This picture shows the stock cultures for both colony 1 and colony 2. The left test tube shows the growth from colony 1 on the TSA slant. The right test tube shows the growth from colony 2 on the TSA slant.

The creation of induced pluripotent stem cells by direct reprogramming has allowed for the circumvention of using embryonic stem cells while still leaving the cells with the ability to maintain pluripotency. Instead of ES cells which were originally derived from the epiblast of mouse embryos, IPS cells were generated from mouse embryonic fibroblasts. This eliminated both any ethical concerns for whether those cells were a living being or not and the need to destroy embryos at the blastocyst stage. An advantage of IPS cells is that they are derived from human somatic cells which makes them easy to acquire due to the possibility of using skin or blood cells. They can also be grown and differentiated individually for each person that the sample of somatic cells is taken from which eliminates the possibility of having any immune reaction and rejection to the differentiated cells during transplantation. These characteristics of IPS cells are important because they are what enables us to safely and accurately transform these affected cells from patients cells into neurons and confidently study them.

diH20 acted as a negative control in the experiment, all cells, both in intrafollicular zone and in interfollicular zone stained purple. Cells stained brown in the negative control indicate non-specific staining.

Stem cells are unspecialized cells, which have the potential to develop into several different cell types—muscle cells, brain cells, skin cells, or red blood cells—during early development. Stem cells continuously divide in order to replace other cells. As stem cells divide, they have the potential to develop into a specialized cell or remain a stem cell. Induced pluripotent stem cells (iPSCs) play an essential role in tissue regeneration, and can circumvent immune system concerns. There are a variety of stem cells, including embryonic stem cells, adult stem cells, and induced pluripotent stem cells. However, the main focus of this paper will be the usage of induced pluripotent stem cells.

Before plating the strains on agar plates, dilutions of the three strains of cells were prepared with LB broth.

provide new specialized cells to take the place of those that die or that are lost. Current studies are researching how these different stem cells may be used to prevent or cure diseases and injuries such as Parkinson’s disease, type 1 diabetes, heart disease, spinal cord injury, Duchene’s muscular dystrophy, Alzheimer’s disease, strokes, burns, osteoarthritis, rheumatoid arthritis, vision, and hearing loss (American Medical Association, 2013). However, there is specific embryonic stem cell that has potential to contribute to the medical world. It is known as the Pluripotent stem cell. In Ian Murnaghan’s article (2011), he refers to pluripotent as an isolated embryo stem cell that can produce almost all of the cells in the body, but it can no longer do this once the embryo is done developing. The Pluripotent stem cells provide the potential ability to renew sources of healthy cells and tissues to treat a variety of diseases. This topic is especially important for people who are suffering from disease that are capable of being treated through stem cell use. For these people, the treatment has the potential to change their outlook on life to a positive one. Scientists are striving to find ways to have stem cells repair damaged tissue in the human body, and this will lead to further medical advancements for researchers.

Human embryonic stem cells (ESCs) are pluripotent cells isolated from blastocysts, and are highly useful in studying human development (Itzkovitz-Eldor et al., 2000 p. 88). Although the National Institute of Health states that “it is not known if iPSCs and embryonic stem cells differ in clinically significant ways”, iPSCs are already being used to achieve the same results as ESCs in some applications without the use of embryos, removing the ethical concern associated with ESCs (National Institutes of Health, 2009). ESCs are capable of differentiating into all cell types, and can be used as a source of differentiated cells. In the report by Itskovitz-Eldor et al., they discuss the induced differentiation of ESCs in suspension into embryoid bodies, including the three embryonic germ layers. The authors state that “the ability to induce formation of human embryoid bodies that contain cells of neuronal, hematopoietic and cardiac origins will be useful in studying early human embryonic development” (Itzkovitz-Eldor et al., 2000 p. 88).

In order to obtain well-isolated discrete colonies, the quadrant streak and spread technique was used. This allowed dilution of the original microbial material over the entire surface of the plate. As the original sample was diluted by streaking and spreading it, over successive quadrants the number of organism decreases. Usually by the third or fourth quadrant only a few organism were transferred on the by the inoculating loop and theses produce a few isolated

Sylgard plates were prepared using 100 mm diameter cell culture plates filled with 15 ml sylgard (Dow Chemical Corp., Midland, MI, USA; type 184 silicon elastomer) and allowed to cure for 3 weeks at room temperature. Prior to use, plates were decontaminated with UV light (wavelength 253.7 nm) for 60 min and rinsed with 70% EtOH and DPBS.