Through their lens: Genevieve de Kervor assess chromosomal irregularities in iPS cells

This summer we’re sponsoring high school interns in stem cell labs throughout California. We asked those students to contribute to our Instagram photos and YouTube videos about life in the lab, and write about their experiences.

Genevieve de Kervor worked in the lab of Eric Topol at Scripps Research Institute.

Genevieve de Kervor working in the lab. She submitted this photo through Instagram to CIRM’s #CIRMStemCellLab collection.

I am currently working with Scripps Translational Science Institute as a member of Dr. Eric Topol’s Lab. The primary goal of my portion of the project is to capture images of cells to determine if there are any gross chromosomal abnormalities in the induced pluripotent stem cells (iPSCs) that have been generated thus far. The secondary goal is to optimize existing protocols for the assay used to do the metaphase spreads. Chromosomal analysis is an important step in validating that the iPSCs generated thus far are indeed stem cells.

At the start of the internship, I was given the opportunity to read existing protocols and scientific papers in order to better understand the importance of both my portion of the project as well as the long term goals laid out.

Then, I learned to count chromosomes of existing iPSC slides on Photoshop. I organized cell line image files on our lab’s shared folder. I counted to make sure that each cell had its full genetic makeup of forty six chromosomes. Many of the metaphase spreads were hard to count as a result of excessive overlap in chromosomes. The challenge of counting the chromosomes reiterated the importance of improving the metaphase spread protocols in order to take decent photos on the microscope. Currently, only 15% of cells are captured in metaphase after completing the protocol which makes it difficult to image adequate numbers of cells. Increasing the numbers of cells in metaphase is an example of an optimization I am seeking to make.

Aside from my specific role in the large scale project, I was able to work with my mentor Lauren on one of the initial steps of the project: isolating cells to use for iPSC derivation. I also helped organize the reagents used in the iPSC project by cataloging lot numbers. Helping others in my lab work on a collective project has provided broader insight on many different aspects of a career in academic research.

I have been able to manage my time and project on my own. Lauren and I laid out the last five weeks of my project and I am in charge of being self-sufficient. I am learning to answer most of my questions on my own and to solve problems. So far, my internship has been an incredible experience. I have enjoyed being immersed in this completely new environment with such welcoming people. Thanks CIRM, I am extremely grateful for this opportunity!

Genevieve de Kervor

Genevieve submitted this video about her experience:

Finding should result in safer stem cell transplants from donors

Blood-forming stem cells donated by family members and good Samaritans save the lives of thousands of patients with blood cancers such as leukemia every year. But of the 20,000 such transplants each year around half result in a severe and frequently fatal complication known as Graft Versus Host Disease (GVHD).

Now, a multi-institutional team has found a molecule in the blood of patients who are most likely to develop the severe form of GVHD. Knowing about this molecule should allow doctors tailor more aggressive treatment to those patients most at risk and intervene earlier, perhaps before the complication manifests in the patients.

GVHD occurs when the donated stem cells produce immune cells that recognize the tissues of the patient as foreign and attacks them. This attack from the new donor immune system often focuses on linings the intestinal system and can be quite painful. It typically does not strike until about 30 days after the transplant, but the research team found evidence of the biomarker called ST2 as early as 14 days after transplant pointing to the possibility of early intervention.

The team included researchers from Indiana University, the University of Michigan, the Fred Hutchison Cancer Center and the Dana-Farber Cancer Institute. It was published in today’s New England Journal of Medicine, and a press release from IU was picked up by this news site. It quoted the team’s senior author from IU Sophie Paczesny:

“This blood test, which is currently available to clinicians, will make informed treatment possible as the clinicians will now be able to adjust therapy to the degree of risk rather than treating every patient the same way.”

For patients lucky enough to have an immunologically matched relative, the chances of GVHD are around 40 percent. But for those who have to rely on unrelated donors the risk can be as high as 60 to 80 percent. So, knowing who to treat early and aggressively could save considerable pain and suffering as well as lives.

The blood-forming stem cells used to treat leukemia come from either bone marrow or cord blood transplants. For a primer on the various types of stem cells visit our Stem Cell Basics.

Don Gibbons

Through their lens: Erik Owen learns that stem cell biology is more complex than textbook descriptions

This summer we’re sponsoring high school interns in stem cell labs throughout California. We asked those students to contribute to our Instagram photos and YouTube videos about life in the lab, and write about their experiences.

Erik Owen worked in the lab of Bruce Torbett at teh Scripps Research Institute.

Erik Owen learning lab techniques. He submitted this photo through Instagram to CIRM’s #CIRMStemCellLab collection.

Multipotent hematopoietic stem cells (HSCs) develop into all the different types of blood cells (that’s why they named stem cells stem cells, because specialized cells stem from them—how clever!). We figured that out a long time ago. We even know what kind of blood cells are out there. We just don’t know what stimulates HSCs to mature in a certain way. Probability probably plays a part in deciding which HSCs develop into what but it’s also influenced by the epigenetic structure of the cell’s DNA and transcription factors.

Okay, enough with the technical background. Let’s go into specifics. Pu.1 is a transcription factor that causes HSCs to differentiate. My focus is on quantifying certain amounts of Pu.1 to specific types of differentiation. That’s more the big picture than anything though. The point of my project is to test the Pu.1 negative/positive system for leaks. The system is leaky if cell markers for CD11b, part of an integrin that is expressed when a cell is Pu.1 positive, appear in a Pu.1 “null” cell.

Just from asking around the lab about other people’s projects, I’ve realized how extensive (almost) every single project is. Even some of the aspects of my work, a relatively small project, are extensive. For example, isolating stem cells. Wow, who would’ve known that there is about 1 long term hematopoietic stem cell for every other 100,000 cells inside the bone marrow? Yeah, isolating .003% of blood cells in the bone marrow (or in umbilical cord blood which thankfully has a higher HSC concentration) gives a pretty low yield of HSCs.

Obtaining cells is only the start. Culturing cells is a nearly daily process. Although not as time consuming as some lab procedures, it is very important. Cells have to be kept alive and growing (although in the case of stem cells, they have to be kept undifferentiated as well). You have to count cell lines in order to make sure the media in the flask isn’t being too crowded. If too crowded, you have to split the cells (in suspension) by taking out a certain amount of cells, putting it in a new flask, and adding fresh media. It’s not actually as bad as it sounds though. It can be pretty fun to watch your cells grow and fill up the media more and more every single day.

One thing that has really been stressed as I have learned about my project is that a hematopoietic stem cell doesn’t just spontaneously become a neutrophil or eosinophil or basophil or erythrocyte or basically, any fully specialized blood cell. There are many precursor cells that gradually gain more and more characteristics of a specific differentiated cell. For example, a HSC will become a common myeloid progenitor cell, then a granulocyte-macrophage progenitor, then a myeloblast, promyeloblast, myelocyte, metamyelocyte, and finally a neutrophil! That’s seven different progenitor steps before a fully developed cell is created. That’s a lot more complicated than it might seem when a textbook says, “Neutrophils develop from stem cells.”

Overall, even Wikipedia couldn’t fully explain what makes stem cells develop into specific specialized cells. Well, maybe in the not-so-distant future it will, but that’s going to take more research—and that reminds me, I have to go split some K-562 cells now. Time for some more research.

Erik Owen

Erik submitted this video about his experience:

Applying lessons learned from the HeLa experience: making consent informative

NIH Director Francis Collins with members of the Lacks family. Collins posted this photo to his Twitter account @NIHdirector yesterday after announcing the historic agreement.

Yesterday the NIH announced that their director Francis Collins had personally worked with the Lacks family to give consent for the publication of the HeLa cell genome sequence. This agreement includes a working group to control access to the cells and requires crediting the Lacks family in any publication.

The HeLa cells have been widely used in research since they were originally obtained from a tumor biopsy in 1951 from a patient named Henrietta Lacks. According to a story in Nature, that cell line now turns up in more than 75,000 publications. The cell line has contributed to countless advances in science, but it was obtained without permission from Henrietta Lacks or her family. The New York Times has more details about the cells and their history in a story today.

Most recently, the DNA sequence of the HeLa cell line was published without the knowledge of the family. The family was understandably upset by the lack of consultation and in response the research team removed the genome data from public access. Dr. Collins intervened and worked with the family and researchers to come to agreement about how genetic information could be released to researchers. Here’s what Collins said about the agreement in his blog:

It’s most unfortunate that Ms. Lacks did not receive the thanks she deserved from researchers during her lifetime. However, I’m glad that we now have a chance to thank the Lacks family for continuing to share her enduring legacy with the biomedical research community. Their generosity extends to the millions of people who have benefited, or will benefit in the future, from research using HeLa cells.

This story is the latest chapter in the story of the HeLa line. Rebecca Skloot’s book The Immortal Life of Henrietta Lacks recounts the history of the line and the family’s ordeal. Ironically, the Lacks family has been supportive of research. Skloot documents a very moving account of a visit by family members to Christoph Lengauer’s lab at Johns Hopkins. The family members were provided the opportunity to learn about the research being conducted with HeLa cells and they were able to observe the cells under the microscope.

CIRM has benefited from these efforts. We are currently supporting an initiative to collect tissue samples from thousands of people with a range of incurable diseases and create reprogrammed iPS cells from those tissues (here’s more about that initiative). These cells will be a resource for scientists worldwide working to understand and treat diseases. Part of this initiative includes a consent process to make sure people who donate fully understand how their cells will be used. (This process is formally called informed consent.)

The informed consent process includes a form that identifies the purposes of the research and describes the way cells will be used. We are also developing education materials that will help potential donors quickly and easily understand the basic aspects of research that will be conducted with those cells. The end result of this collaboration with our grantees will be a process that is truly informative to donors.

The informed consent process can’t entirely eliminate all future questions on the part of the donor, but it does ensure that donors have a chance to understand how their cells will be used and what information will be made public—something Henrietta Lacks and her family never had.

Geoff Lomax