Leaders in Pharmaceutical Business Intelligence (LPBI) Group

Observing the spleen colonies in mice and proving the existence of stem cells – Till and McCulloch

Larry H. Bernstein, MD, FCAP, Curator

Leaders in Pharmaceutical Innovation

Series E. 2; 7.2

 

Till & McCulloch are Doctors James Till and Ernest McCulloch who, while studying the effect of radiation on the bone marrow of mice at the Ontario Cancer Institute, in Toronto, demonstrated the existence of multipotent stem cells in 1961.

Now recognized as the Fathers of Stem Cell Science, Till & McCulloch exemplified the importance of multidisciplinary collaboration in scientific research and have received many awards for their collaborative and ground-breaking research.

They first published their findings of the discovery of stem cells in the journal Radiation Research.[1][2] In later work, joined by graduate student Andy Becker, they cemented their stem cell theory and published the results in the journal Nature in 1963.[3]

After their pioneering discovery, Till & McCulloch continued to help this new field develop; not only by continuing to expand their research activities, but also by mentoring other young scientists. Together, Till & McCulloch spawned successive generations of scientists who continue to deepen the understanding of how the different types of stem cells work and their application to different diseases and medical conditions—many have also become globally recognized leaders in their field.

Dr. Till’s focus shifted increasingly towards the evaluation of cancer therapies and quality of life issues in the 1980s. He has held a wide range of positions in organizations ranging from the Stem Cell Network to Project Open Source to the Canadian Breast Cancer Foundation, and many others.

Dr. McCulloch continued to expand the depth of work in his field with a heavy emphasis on cellular and molecular mechanisms affecting the growth of malignant blast stem cells from the blood of patients with Acute Myeloblastic Leukemia. Unfortunately, Dr. McCulloch died on January 20, 2011, shortly before the 50th anniversary of the publication of the 1961 paper in Radiation Research.

Lifetime Achievement: Drs. James Till and Ernest McCulloch

http://oicr.on.ca/news/portal-news/lifetime-achievement-drs-james-till-and-ernest-mcculloch

In the early 1960s, two Canadian scientists started a series of experiments involving injection of bone marrow cells into irradiated mice.

Dr. James E. Till, a native of Saskatchewan who completed his PhD in biophysics at Yale, and Dr. Ernest McCulloch, a Toronto-born doctor who completed his research training in England, were working together on research related to leukemia at the Ontario Cancer Institute. Their immediate aim was to investigate a controversial new finding by Colorado scientist Theodore Puck, which seemed to show that normal cells are just as susceptible to radiation as cancer cells. At the time, scientists believed radiation “melted” away cancer cells while leaving normal tissue intact. While there was no doubt that radiation is an effective way to kill cancer cells, Puck’s research suggested scientists must be wrong about the way it acts on cells.

Till and McCulloch’s study proved Puck’s finding was correct. But this wasn’t all that their research proved.

In the mouse experiments, they observed nodules in the animals’ spleens when the bone marrow cells were injected. These nodules appeared in proportion to the number of cells injected, leading the two young scientists to speculate that the nodules – which they termed “spleen colonies” – were arising from a single marrow cell. If this were true, the experiment would be a breakthrough, since scientists had not yet proved that it was possible for cells to act in this fashion.

Till and McCulloch conducted further experiments that proved the cells they were observing were indeed stem cells. The rest, as they say, is history.

Still a groundbreaking field

Stem cell research is often discussed in the media as a new, groundbreaking field, but the idea that certain special cells might be responsible for creating many other types of cell goes back quite a bit further than Till and McCulloch’s experiments in the 1960s. The problem of where cells come from is fundamental to biology; for centuries, or perhaps longer, scientists have searched for the origin of the building blocks of life.

Since early in the 1900s, scientists had suspected that there must be some sort of stem cell in the blood forming system. But stem cells proved extraordinarily tricky to observe.

By observing the spleen colonies in mice and proving the existence of stem cells, Till and McCulloch sparked worldwide interest. Once they had established proof that spleen colonies originate from stem cells, there was solid reason to believe that other cells originate from them too – something that has been confirmed through further research.

Developments in technology, biology and research ethics have recently propelled stem cell research to the forefront of public debates on science. Scientists now know that embryonic stem cells can differentiate into all of the specialized embryonic tissues, while adult organisms’ stem cells and progenitor cells can act as a repair system for the body, replenishing specialized cells and maintaining the normal turnover of regenerative organs, such as blood, skin or intestinal tissues.
In the United States, and to a lesser extent in other countries, controversy has erupted as scientists have proposed to explore using human embryonic stem cells – which, by definition, have to be harvested from human embryos – as treatments for disease.

While they tend to garner fewer headlines, there are also many projects exploring the use of adult stem cells in medicine to regenerate parts of the body affected by disease or injury. Research in this area has become very promising since 2006, when Shinya Yamanaka, a researcher at Kyoto University in Japan, showed that adult somatic cells can be “reprogrammed” to act like embryonic stem cells – opening the possibility of using pluripotent stem cells in medicine without harvesting cells from human embryos. The reprogrammed cells, called induced pluripotent stem cells, are an area of intense research activity. In the few years since Yamanaka’s discovery, researchers have already refined and improved techniques for creating induced pluripotent stem cells.

Remarkable careers

In the decades after their discovery, Till and McCulloch continued their research on stem cells, publishing several groundbreaking papers and eventually developing the framework through which stem cells are currently understood. They later moved on to other projects, with McCulloch focusing on cellular and molecular mechanisms affecting the growth of malignant blast stem cells obtained from the blood of patients with acute myeloblastic leukemia, and Till branching out into a number of other health-related fields including evaluation of cancer therapies, quality of life issues and the ethics of Internet research.

Till and McCulloch have received many honours for their research, including the Albert Lasker Award for Basic Medical Research and the Gairdner International Award, Canada’s major award for biomedical research. Both are University Professors Emeritus at the University of Toronto, Officers of the Order of Canada and members of the Order of Ontario and the Canadian Medical Hall of Fame. Till’s research on the impact of the Internet and advocacy for open access to research publications continues to this day. McCulloch is now retired.

Although Till and McCulloch are no longer working in the stem cell field, there are plenty of Ontario scientists who are. The University of Toronto and Ontario Cancer Institute have retained their early lead, developing programs to harness stem cell research for a wide range of applications in medicine. The province rose to international prominence again in the 1990s when Dr. John Dick, a scientist at the Ontario Cancer Institute, proved the existence of cancer stem cells – a subpopulation of cancer cells that are responsible for the growth and spread of cancer.

In the years since, Dick has established a major hub of cancer stem cell research in Ontario. In 2007 the Ontario Institute for Cancer Research appointed Dick as Director of a new Cancer Stem Cell Program to develop and implement a strategy to further understand cancer stem cells and use the concept as the basis for developing new treatments. The program has already recruited rising stars in the cancer stem cell field and has begun working on its ambitious research plan.

“The truly remarkable thing about Drs. Till and McCulloch is that the stem cell discovery was just one part of two very outstanding careers. They also worked tirelessly behind the scenes as builders, teachers and mentors in the decades when Ontario solidified its presence in cancer research,” says Dr. Bob Phillips, Deputy Director of OICR and a former colleague of Till and McCulloch’s at the Ontario Cancer Institute.

“And the remarkable thing about the discovery itself is that we’re just starting to realize the potential of stem cells for medicine. In the 1960s, scientists recognized that Drs. Till and McCulloch’s discovery was important, but I don’t think anyone could have imagined that more than 45 years down the road their work would still be laying the basis for new ideas, new strategies, even new research institutes built around the concept of stem cells.”

Ernest McCulloch: Cell Biology – Conducted a series of experiments that would eventually result in the first proof of the existence of stem cells, a discovery that would revolutionize our understanding of human biology and disease.

“I learned enough about myself to settle on a career in medicine: I did not like discipline – therefore I wanted to work for myself – to be my own boss.”

On an ordinary Sunday more than half a century ago, so ordinary a day that its exact date would later be forgotten, a young faculty member at the Ontario Cancer Institute in Toronto went to work to perform a routine check on his experimental animals. Many years later, he only remembered that it was a cold day, perhaps in the autumn. Navigating his way through quiet streets, Dr. Ernest McCulloch arrived at the Institute and entered the building. After donning his lab coat, McCulloch went to the animal quarters and checked his experimental mice. McCulloch followed a routine process for obtaining samples of their blood-forming tissues, a process which he had done many times before. His goal, working with his research partner James Till, was to determine if, by irradiating mouse bone marrow cells before transplanting them into irradiated mice, changes might later be found in the kinds of cells responsible for blood formation. It was a routine collection of samples on an ordinary day, noteworthy only because it was a Sunday.

After the samples were processed McCulloch, ever the sharp-eyed observer, noticed the unexpected presence of several small rounded bumps on the spleens of mice that had received bone marrow cells, and he decided to count them. He found that the number of nodules on each spleen was directly related to the number of bone marrow cells the mouse had received.

Suddenly things got very exciting for this unlikely duo of researchers. McCulloch was short, a medical doctor, raised in affluent downtown Toronto, with a penchant for classical literature, cinema and poetry. Till, on the other hand, was tall and athletic, a straight-shooting biophysicist who grew up on the Canadian Prairies and loved the sport of curling.

Although it had long been postulated that a single type of cell—a so-called stem cell— could give rise to multiple different cell types, no definitive evidence proved that they existed. The potential of such a “stem cell”, if discovered, would be dramatic, because its ability to regenerate different human body tissues could be used to treat all sorts of diseases. Following this cold, ordinary yet ultimately incredibly exciting day, McCulloch and Till went on to perform a series of seminal experiments in the 1960s that proved, for the first time, the existence of stem cells detected by their “spleen colony formation” assays.

The initial discovery of a direct relationship between the number of colonies and the number of transplanted cells suggested that single rare cells were able to initiate these colonies, but the suggestion required further validation. They knew that they were onto something very interesting, because they found that the colonies contained a variety of precursors of mature blood cell types—red cells, white cells and platelets—the normal cellular components of blood. These foundational observations were published in the specialty journal “Radiation Research” in 1961 under the un-dramatic title “A Direct Measurement of Radiation Sensitivity of Normal Bone Marrow Cells”. The paper did not use the words ”stem cell”, because Till and McCulloch, being rigorous scientists, required stronger evidence before making such a bold interpretation of their findings. Hence, their paper went unnoticed by the general biology community.

Their next paper, published in Nature in 1963, changed this and really brought Till and McCulloch to the forefront of hematological biology —the study of blood. Till’s PhD student Andy Becker found a way to trace the source of the cells in the spleen colonies to demonstrate that they originated from individual cells (not clusters of cells) in the bone marrow and could generate three types of progenitors required to make blood. The paper, titled “Cytological Demonstration of the Clonal Nature of Spleen Colonies Derived from Transplanted Mouse Marrow Cells”, still did not use the word “stem cell” as this was not the nature of these exacting scientists, who demanded that any degree of doubt be extinguished before making such claims.

McCulloch and Till went on to publish a number of subsequent papers, which have now been cited thousands of times, unequivocally demonstrating the presence of special cells within the bone marrow. They, with colleague Louis Siminovitch, offered the first biological definition of stem cells, which included two key characteristics: 1) self renewal – to be a stem cell, a cell must be able to give rise to new copies of itself; 2) differentiation – stem cells are able to divide and generate more mature cells that, following subsequent divisions, are eventually able to generate the highly specialized and functional cells essential for complex multi-cellular organisms work. An example of this can be seen in the hematopoietic (e.g. blood forming) stem cells they described, with a single undifferentiated stem cell being able to eventually form all the different types of cells that comprise our blood.

After these breakthroughs in the 1960s, the pair continued to work together in the field of experimental hematology for the next two decades.  Although they continued to make more discoveries, it was those first findings that caused a huge impact on biology today by demonstrating the presence of stem cells. The field of stem cell biology has expanded dramatically and is now on the verge of a potential revolution in how we understand health and treat disease.

Born in an affluent neighborhood of Toronto, on Warren Road south of St. Clair Avenue, Ernest “Bun” McCulloch was raised well, with a private school education at Upper Canada College and summers at the cottage in the country. Given the nickname “Bun” by his grandmother, the name stuck with him for his entire life. McCulloch was educated as a medical doctor at the University of Toronto, graduating with an MD in 1948, then going on to the Lister Institute in London, England, where he had his first experience with scientific research.

“Bun” returned to Canada in 1949 where he interned at the Toronto General Hospital, specializing in internal medicine. His medical career began at the Sunnybrook Hospital in Toronto where he became an assistant resident and a research fellow in pathology at the Banting Institute. In 1954, McCulloch joined the University of Toronto as a teacher in the Department of Medicine. His next move, taking on the Head of Hematology in the Biology Division at the Ontario Cancer Institute in 1957, would result in his most famous work. He became part of a team of new promising young cancer researchers in the newly founded Department of Medical Biophysics, McCulloch quickly partnered up with James Till to study the effects of radiation on mouse bone marrow cells. The pair conducted a series of experiments that would eventually result in the first proof of the existence of stem cells, a discovery that would revolutionize our understanding of human biology and disease.

Ernest McCulloch was a man of incredible personality and charm. He was extremely well read and enjoyed discussing a wide variety of poetry, classical literature and theatre with his colleagues. He is known for his long-lasting impact on the Canadian medical research community. A list of the notable scientists mentored by Till and McCulloch is a who’s who of Canadian medical scientists, including (but not limited to): former president of the Canadian Institute for Health Research, Alan Bernstein; the discoverer of the T-cell receptor, Tak Mak, and a world leader in the field of hematopoietic stem cell biology, Connie Eaves.

McCulloch and Till’s work resulted in almost every top honor in science, except for the Noble Prize. Widely expected to be a joint winner of this top prize in science with Jim Till, sadly McCulloch passed away in 2011 preventing him from receiving this distinction. Till and McCulloch’s legacy in Canadian biomedical research cannot be understated, with their foundational work in establishing the presence of stem cells within bone marrow and prolific scientific mentorship. With two recent Nobel prizes, 2007 and 2012, going to stem cell researchers who worked on embryonic stem cells and induced pluripotent stem cells, respectively, it is still expected by many scientists that Till’s seminal experiments on adult stem cells will garner him the Nobel prize in the future.

by Ben Paylor