New technique maps out diversity and location of cells in tissue or tumor

Image Description: Alex Marson is part of a team of researchers who developed a new technique to map the specialized diversity and spatial location of individual cells within a tissue or tumor. Photo Credit: Anastasiia Sapon

All the cells in your body work together and each can have a different role. Their individual function not only depends on cell type, but can also depend on their specific location and surroundings.

A CIRM supported and collaborative study at the Gladstone Institutes, UC San Francisco (UCSF), and UC Berkeley has developed a more efficient method than ever before to simultaneously map the specialized diversity and spatial location of individual cells within a tissue or a tumor.

The technique is named XYZeq and involves segmenting a tissue into microscopic regions. Within each of these microscopic grids, each cell’s genetic information is analyzed in order to better understand how each particular cell functions relative to its spacial location.

For this study, the team obtained tissue from mice with liver and spleen tumors. A slice of tissue was then placed on a slide that divides the tissue into hundreds of “microwells” the size of a grain of salt. Each cell in the tissue gets tagged with a unique “molecular barcode” that represents the microwell it’s contained in, much like a zip code. The cells are then mixed up and assigned a second barcode to ensure that each cell within a given square can be individually identified, similar to a street address within a zip code. Finally, the genetic information in the form of RNA from each cell is analyzed. Once the results are obtained, both barcodes tell the researchers exactly where in the tissue it came from.

The team found that some cell types located near the liver tumor were not evenly spaced out. They also found immune cells and specific types of stem cells clustered in certain regions of the tumor. Additionally, certain stem cells had different levels of some RNA molecules depending on how far they resided from the tumor.

The researchers aren’t entirely sure what this pattern means, but they believe that it’s possible that signals generated by or near the tumor affect what nearby cells do.

In a press release, Alex Marson, M.D., Ph.D., a senior author of the study, elaborates on what the XYZeq technology could mean for disease modeling.

“I think we’re actually taking a step toward this being the way tissues are analyzed to diagnose, characterize, or study disease; this is the pathology of the future.”

The full results of the study were published in Science Advances.

Stem Cell All-Stars, All For You

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Dr. Larry Goldstein, UC San Diego

It’s not often you get a chance to hear some of the brightest minds around talk about their stem cell research and what it could mean for you, me and everyone else. That’s why we’re delighted to be bringing some of the sharpest tools in the stem cell shed together in one – virtual – place for our CIRM 2020 Grantee Meeting.

The event is Monday September 14th and Tuesday September 15th. It’s open to anyone who wants to attend and, of course, it’s all being held online so you can watch from the comfort of your own living room, or garden, or wherever you like. And, of course, it’s free.

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Dr. Daniela Bota, UC Irvine

The list of speakers is a Who’s Who of researchers that CIRM has funded and who also happen to be among the leaders in the field. Not surprising as California is a global center for regenerative medicine. And you will of course be able to post questions for them to answer.

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Dr. Deepak Srivastava, Gladstone Institutes

The key speakers include:

Larry Goldstein: the founder and director of the UCSD Stem Cell Program talking about Alzheimer’s research

Irv Weissman: Stanford University talking about anti-cancer therapies

Daniela Bota: UC Irvine talking about COVID-19 research

Deepak Srivastava: Gladsone Institutes, talking about heart stem cells

Other topics include the latest stem cell approaches to COVID-19, spinal cord injury, blindness, Parkinson’s disease, immune disorders, spina bifida and other pediatric disorders.

You can choose one topic or come both days for all the sessions. To see the agenda for each day click here. Just one side note, this is still a work in progress so some of the sessions have not been finalized yet.

And when you are ready to register go to our Eventbrite page. It’s simple, it’s fast and it will guarantee you’ll be able to be part of this event.

We look forward to seeing you there.

Perseverance: from theory to therapy. Our story over the last year – and a half

Some of the stars of our Annual Report

It’s been a long time coming. Eighteen months to be precise. Which is a peculiarly long time for an Annual Report. The world is certainly a very different place today than when we started, and yet our core mission hasn’t changed at all, except to spring into action to make our own contribution to fighting the coronavirus.

This latest CIRM Annual Reportcovers 2019 through June 30, 2020. Why? Well, as you probably know we are running out of money and could be funding our last new awards by the end of this year. So, we wanted to produce as complete a picture of our achievements as we could – keeping in mind that we might not be around to produce a report next year.

Dr. Catriona Jamieson, UC San Diego physician and researcher

It’s a pretty jam-packed report. It covers everything from the 14 new clinical trials we have funded this year, including three specifically focused on COVID-19. It looks at the extraordinary researchers that we fund and the progress they have made, and the billions of additional dollars our funding has helped leverage for California. But at the heart of it, and at the heart of everything we do, are the patients. They’re the reason we are here. They are the reason we do what we do.

Byron Jenkins, former Naval fighter pilot who battled back from his own fight with multiple myeloma

There are stories of people like Byron Jenkins who almost died from multiple myeloma but is now back leading a full, active life with his family thanks to a CIRM-funded therapy with Poseida. There is Jordan Janz, a young man who once depended on taking 56 pills a day to keep his rare disease, cystinosis, under control but is now hoping a stem cell therapy developed by Dr. Stephanie Cherqui and her team at UC San Diego will make that something of the past.

Jordan Janz and Dr. Stephanie Cherqui

These individuals are remarkable on so many levels, not the least because they were willing to be among the first people ever to try these therapies. They are pioneers in every sense of the word.

Sneha Santosh, former CIRM Bridges student and now a researcher with Novo Nordisk

There is a lot of information in the report, charting the work we have done over the last 18 months. But it’s also a celebration of everyone who made it possible, and our way of saying thank you to the people of California who gave us this incredible honor and opportunity to do this work.

We hope you enjoy it.

“Mini” human liver made of stem cells successfully transplanted in rats

Miniature liver made from human skin cells turned stem cells turned specialized liver cells Photo Credit: University of Pittsburgh School of Medicine

According to the American Liver Foundation website, almost 14,000 patients are on the waiting list for a liver transplant. But what if there was a way to generate a liver using your own cells so that you didn’t have to wait? Researchers at the University of Pittsburgh School of Medicine have gotten one step closer towards that goal.

Using human skin cells from volunteers, Dr. Alejandro Soto-Gutierrez and his team of researchers were able to create “mini” livers which were successfully transplanted into rats. In this proof of concept experiment, the “mini” livers survived inside the rats for four days. Additionally, they secreted bile acids and urea and produced proteins similar to a normal liver. Normally, liver maturation takes up to two years in a natural environment, but Dr. Soto-Gutierrez and his team were able to do this in under a month.

The researchers were able to do this by taking human skin cells and reprogramming them into induced pluripotent stem cells (iPSCs), a type of stem cell that has the ability to turn into virtually any other kind of cell. These newly formed iPSCs were then made into liver cells which were then seeded into a rat liver with all of its own cells removed. These newly formed “mini” livers were then transplanted into the rats.

In a press release, Dr. Soto-Gutierrez discusses what it was like observing the newly created “mini” livers.

“Seeing that little human organ there inside the animal – brown, looking like a liver – that was pretty cool. This thing that looks like a liver and functions like a liver came from somebody’s skin cells.”

Although these results were promising, there are still challenges that need to be addressed in future studies such as long-term survival and safety issues.

Even so, Dr. Soto-Gutierrez says his research could one-day benefit patients who are running out of options.

“The long-term goal is to create organs that can replace organ donation, but in the near future, I see this as a bridge to transplant. For instance, in acute liver failure, you might just need hepatic boost for a while instead of a whole new liver”.

The full results to this study were published in Cell Reports.

CIRM invests $1.3 million to study stem cells in metabolic liver disease

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Dr. Tracy Grikscheit. Image courtesy of Children’s Hospital LA.

Metabolic liver disease, is an emerging public health concern in Western countries, but has largely been overshadowed by health issues such as cancer and diabetes. Chronic liver disease (of which metabolic liver disease is a significant contributor) however, is a significant public health concern, evidenced by its contribution to nearly 2 million deaths per year worldwide.

The primary treatment option for metabolic liver disease is a liver transplant. In fact, of the liver transplants performed every year, 14% are due to damage associated with metabolic disorders. With any organ transplant, however, such a procedure comes with drawbacks, the most frustrating of which is the need for patients to wait for an organ donor.

As transplants are not a reasonable or feasible option for many people, alternative treatment options are necessary.  Enter Dr. Tracy Grikscheit, a doctor-scientist at the Children’s Hospital Los Angeles, who hopes to make liver transplant a thing of the past for the millions of people who live with metabolic liver disease.

Dr. Grikscheit was awarded a $1.3 million grant to study how stem cells can be used to treat liver disease caused by metabolic disorders. In a press release, Dr. Grikscheit details the importance and practicality of using stem cells to treat liver disease:

“Liver-based metabolic diseases are the perfect starting point to apply cellular therapy to liver disorders. The only current therapy — a liver transplant — is costly and in short supply. Plus, it requires suppressing the patient’s immune system, which has long-term consequences.”

The project, termed UPLiFT for Universal Pluripotent Stem Cell Therapy, aims to use pluripotent stem cells (cells that can turn into any cell in the body) to correct liver associated disorders like Crigler-Najjar Syndrome. A genetic mutation in liver cells of these patients makes them unable to covert bilirubin (a byproduct of red blood cell degradation) to its non-toxic form. Dr. Grikscheit hopes to bypass the need for a liver transplant by giving these patients pluripotent stem cells that can become liver cells without the genetic mutation, and are able to convert bilirubin to its non-toxic form. The use of pluripotent stem cells would also potentially eliminate the need for lifelong immunosuppressive therapy

Dr. Grikscheit will use the CIRM grant to test safety and efficacy of the stem cell treatment in pre-clinical trials to determine the optimal cell dosage that will be both safe and relieve disease symptoms, as well as assessing any off-target effects of the treatment. She has previously received a grant from CIRM to study stem cell therapy options for digestive neuromuscular condition, which you can read about here.

 

Making stem cell-derived liver cells to study fatty liver disease

Non-alcoholic fatty liver disease (NAFLD) affects approximately 30% of the population, with that number increasing to 75% in obese individuals. Shockingly, the number of cases is expected to increase 21% by the year 2030 in the United States alone.

liver_fattyliverNAFLD refers to a broad range of liver conditions, which are all characterized by abnormally high levels of fat deposits in the livers of people who do not drink excessive amounts of alcohol. While not always fatal, NAFLD can lead to liver cirrhosis, or extensive scaring of the liver tissue. Cirrhosis, in turn, can cause life-threatening conditions such as liver cancer or liver failure. Whether or not N

AFLD will lead to extensive liver damage is not well understood and the primary therapeutic option is weight loss with no FDA-approved drug options. The projected increase in NALD cases combined with the poor treatment options makes this disease a significant public health burden.

Studying NALD can be quite complicated because the liver is complex organ made up of multiple different cell types. Investigators at the University of Edinburgh have simplified some of this complexity by figuring out a way to generate liver cells in a dish.

In studies published in the Philosophical Transactions of the Royal Society B, these scientists used human embryonic stem cells to generate hepatocyte-like cells (HLCs), or cells that are highly similar to liver cells isolated from humans. When exposed to fatty acids, they saw that the HLCs exhibited hallmarks of NAFLD, such as fat accumulation in liver cells, and changes in gene expression that are indicative of NAFLD.

In a press release, Dr. David Hay, one of the two senior investigators of this study, states:

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Dr. David Hay

“Our ability to generate human hepatocytes from stem cells, using semi-automated procedures, allows us to study the mechanisms of human liver disease in a dish and at scale.”

 

This approach is particularly valuable because it would replace the need to use cancer cell lines for this type of work. While valuable for many reasons, research done in cancer cells lines can be difficult to draw therapeutic conclusions from, because cell lines have significant genetic alternations from normal cells. Generating liver cells from human stem cells provides an important tool for high throughput screening of medically relevant therapies for NALD.

 

Livers skip stem cells, build missing structures from scratch via direct cell identity conversion

Stem cells…eh, who needs them anyway?!

That’s what you might be thinking after today, at least for some forms of liver disease. That’s because a team of researchers from UCSF and Cincinnati Children’s Hospital Medical Center just published results in Nature showing liver cells can directly change identity, or transdifferentiate, in order to build, from scratch, structures missing due to disease.

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The liver contains a network of tubes called bile ducts that carry fat-digesting bile to the small intestine via the gallbladder.
Image: National Cancer Inst.

The extraordinary regenerative power of the liver in animals is well-documented. A human liver, for instance, can fully regrow from just 25% of its original mass. That’s thanks to the hepatocyte, the main type of liver cell, that has the ability to replenish pre-existing tissue lost due to disease or injury. What hasn’t been as clear cut, is whether the hepatocyte has the capacity to change identity and build functional liver structures from scratch that never developed in the first place due to genetic disorders.

To examine that possibility, the study – funded in part by CIRM – focused on an inherited liver disease called Alagille syndrome which is caused by abnormal bile ducts. Produced by the liver, bile helps digest fats in our diet. It travels from the liver via bile ducts – tree branch-like tube structures in the liver – to the gallbladder, where it’s stored before moving on to the small intestine. In Alagille syndrome, the bile ducts are fewer in number, narrower in size or altogether missing. As a result, the bile builds up in the liver causing scarring and severe damage. Nearly half of all those with Alagille syndrome, require a liver transplant, usually in childhood.

The research team mimicked the symptoms of Alagille syndrome in mice by genetically engineering the animals to lack cholangiocytes, the cells that form bile ducts. Sure enough, liver damage from bile buildup was observed in these mice at birth due to the missing bile duct structures, also called the biliary tree. However, 90% of the mice survived and eventually formed a functional biliary tree. The team went on to show, for the first time, that the hepatocytes had converted en masse into cholangiocytes and created the wholly new bile ducts.

liver cell switching

Mice that mimic Alagille syndrome are born without the branches of the biliary tree, an important “plumbing system” in the liver (A), but show a near-normal biliary system as adults (B). To build the missing branches, liver cells switch identity and form tubes, shown in green, that connect to the trunk of the biliary tree, shown in blue (C). Image: Cincinnati Children’s

The underlying molecular mechanisms of this process were further examined. The researchers showed that the lack of a particular gene activity pathway due to the absence of cholangiocytes during development causes a replacement pathway, stimulated by a protein called TGF-beta, to kick into gear. As a result, the hepatocytes convert into cholangiocytes and form bile ducts. To make a direct connection with the human form of the disease, the researchers found evidence that TGF-beta is active in the liver samples of some patients but not in the livers from healthy individuals.

With this Alagille syndrome mouse model in hand, the researchers want to identify which transcription factors – proteins that bind DNA and regulate gene activity – are involved in changing the liver cells into bile duct cells. Holger Willenbring, MD, PhD, a senior author and CIRM grantee, explained the rationale behind this approach in a press release:

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Holger Willenbring

“Using transcription factors to make bile ducts from hepatocytes has potential as a safe and effective therapy. With our finding that an entire biliary system can be ‘retrofitted’ in the mouse liver, I am encouraged that this eventually will work in patients.”

So rather than developing a stem cell-based therapy in the lab which is then transplanted into a patient, this approach would rely on stimulating the regenerative capacity of liver cells that are already inside the body. And if it eventually works in patients with Alagille syndrome, which only affects 1 in 30,000, it’s possible it could be applied to other liver diseases as well.

East Coast Company to Sell Research Products Derived from CIRM’s Stem Cell Bank

With patient-derived induced pluripotent stem cells (iPSCs) in hand, any lab scientist can follow recipes that convert these embryonic-like stem cells into specific cell types for studying human disease in a petri dish. iPSCs derived from a small skin sample from a Alzheimer’s patient, for instance, can be specialized into neurons – the kind of cell affected by the disease – to examine what goes wrong in an Alzheimer’s patient’s brain or screen drugs that may alleviate the problems.

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Neurons created from Alzheimer’s disease patient-derived iPSCs.
Image courtesy Elixirgen Scientific

But not every researcher has easy access to a bank of patient-derived iPSCs and it’s not trivial to coax iPSCs to become a particular cell type. The process is also a time sink and many scientists would rather spend that time doing what they’re good at: uncovering new insights into their disease of interest.

Since the discovery of iPSC technology over a decade ago, countless labs have worked out increasingly efficient variations on the original method. In fact, companies that deliver iPSC-derived products have emerged as an attractive option for the time-strapped stem cell researcher.

One of those companies is Elixirgen Scientific of Baltimore, Maryland. Pardon the pun but Elixirgen has turned the process of making various cell types from iPSCs into a science. Here’s how CEO Bumpei Noda described the company’s value to me:

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Bumpei Noda

“Our technology directly changes stem cells into the cells that make up most of your body, such as muscle cells or neural cells, in about one week. Considering that existing technology takes multiple weeks or even months to do the same thing, imagine how much more research can get done than before.”

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With Elixirgen’s technology, different “cocktails” of ingredients can quickly and efficiently turn iPSCs into many different human cell types. Image courtesy Elixirgen Scientific

Their technology is set to become an even greater resource for researchers based on their announcement yesterday that they’ve signed a licensing agreement to sell human disease cells that were generated from CIRM’s iPSC Repository.

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Stephen Lin

“The CIRM Repository holds the largest publicly accessible collection of human iPSCs in the world and is the result of years of coordinated efforts of many groups to create a leading resource for disease modeling and drug discovery using stem cells,” said Stephen Lin, a CIRM Senior Science Officer who oversees the cell bank.

 

The repository currently contains a collection of 1,600 cell lines derived from patients with diseases that are a source of active research, including autism, epilepsy, cerebral palsy, Alzheimer’s disease, heart disease, lung disease, hepatitis C, fatty liver disease, and more (visit our iPSC Repository web page for the complete list).

While this wide variety of patient cells lines certainly played a major role in Elixirgen’s efforts to sign the agreement, Noda also noted that the CIRM Repository “has rich clinical and demographic data and age-matched control cell lines” which is key information to have when interpreting the results of experiments and drug screening.

Lin also points out another advantage to the CIRM cells:

“It’s one of the few collections with a streamlined route to commercialization (i.e. pre-negotiated licenses) that make activities like Elixirgen’s possible. iPSC technology is still under patent and technically cannot be used for drug discovery without those legal safeguards. That’s important because if you do discover a drug using iPSCs without taking care of these licensing agreements, your discovery could be owned by that original intellectual property holder.”

At CIRM, we’re laser-focused on accelerating stem cell treatments to patients with unmet medical needs. That’s why we’re excited that Elixirgen Scientific has licensed access to the our iPSC repository. We’re confident their service will help researchers work more efficiently and, in turn, accelerate the pace of new discoveries.

Using stem cells to take an inside approach to fixing damaged livers

Often on the Stem Cellar we write about work that is in a clinical trial. But getting research to that stage takes years and years of dedicated work. Over the next few months we are going to profile some of the scientists we fund who are doing Discovery, or early stage research, to highlight the importance of this work in developing the treatments that could ultimately save lives.

 This first profile is by Pat Olson, Ph.D., CIRM’s Vice President of Discovery & Translation

liver

Most of us take our liver for granted.  We don’t think about the fact that our liver carries out more than 500 functions in our bodies such as modifying and removing toxins, contributing to digestion and energy production, and making substances that help our blood to clot.  Without a liver we probably wouldn’t live more than a few days.

Our liver typically functions well but certain toxins, viral infections, long-term excess alcohol consumption and metabolic diseases such as obesity and type 2 diabetes can have devastating effects on it.  Under these conditions, functional liver cells, called hepatocytes, die and are replaced with cells called myofibroblasts.  Myofibroblasts are cells that secrete excess collagen leading to fibrosis, a form of scarring, throughout the liver.  Eventually, a liver transplant is required but the number of donor livers available for transplant is small and the number of persons needing a functional liver is large.  Every year in the United States,  around 6,000 patients receive a new liver and more than 35,000 patients die of liver disease.

Searching for options

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Dr. Holger Willenbring

Dr. Holger Willenbring, a physician scientist at UCSF, is one of the CIRM-funded researchers pursuing a stem cell/regenerative medicine approach to discover a treatment for patients with severe liver disease.  There are significant challenges to treating liver disease including getting fully multi-functional hepatocytes and getting them to engraft and/or grow sufficiently to achieve adequate mass for necessary liver functions.

In previous CIRM–funded discovery research, Dr. Willenbring and his team showed that they could partially reprogram human fibroblasts (the most common cell found in connective tissue) and then turn them into immature hepatocytes.  (see our Spotlight on Liver Disease video from 2012 featuring Dr. Willenbring.) These immature hepatocytes, when transplanted into an immune-deficient mouse model of human liver failure, were shown to mature over time into hepatocytes that were comparable to normal human hepatocytes both in their gene expression and their function.

This was an important finding in that it suggested that the liver environment in a living animal (in vivo), rather than in a test tube (in vitro) in the laboratory, is important for full multi-functional maturation of hepatocytes.  The study also showed that these transplanted immature human hepatocytes could proliferate and improve the survival of this mouse model of chronic human liver disease.  But, even though this model was designed to emphasizes the growth of functional human hepatocytes, the number of cells generated was not great enough to suggest that transplantation could be avoided

A new approach

Dr. Willenbring and his team are now taking the novel approach of direct reprogramming inside the mouse.  With this approach, he seeks to avoid the challenge of low engraftment and proliferation of transplanted hepatocytes generated in the lab and transplanted. Instead, they aim to take advantage of the large number of myofibroblasts in the patient’s scarred liver by turning them directly into hepatocytes.

Recently, he and his team have shown proof-of principle that they can deliver genes to myofibroblasts and turn them into hepatocytes in a mouse. In addition these in vivo myofibroblasts-derived hepatocytes are multi-functional, and can multiply in number, and can even reverse fibrosis in a mouse with liver fibrosis.

From mice to men (women too)

Our latest round of funding for Dr. Willenbring has the goal of moving and extending these studies into human cells by improving the specificity and effectiveness of reprogramming of human myofibroblasts into hepatocytes inside the animal, rather than the lab.

He and his team will then conduct studies to test the therapeutic effectiveness and initial safety of this approach in preclinical models. The ultimate goal is to generate a potential therapy that could eventually provide hope for the 35,000 patients who die of liver disease each year in the US.

 

 

Stem cell study shows how smoking attacks the developing liver in unborn babies

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It’s no secret that smoking kills. According to the Centers for Disease Control and Prevention (CDC) smoking is responsible for around 480,000 deaths a year in the US, including more than 41,000 due to second hand smoke. Now a new study says that damage can begin in utero long before the child is born.

Previous studies had suggested that smoking could pose a serious risk to a fetus but those studies were done in petri dishes in the lab or using animals so the results were difficult to extrapolate to humans.

Researchers at the University of Edinburgh in Scotland got around that problem by using embryonic stem cells to explore how the chemicals in tobacco can affect the developing fetus. They used the embryonic stem cells to develop fetal liver tissue cells and then exposed those cells to a cocktail of chemicals known to be found in the developing fetus of mothers who smoke.

Dangerous cocktail

They found that this chemical cocktail proved far more potent, and damaged the liver far more, than individual chemicals. They also found it damaged the liver of males and females in different ways.  In males the chemicals caused scarring, in females it was more likely to negatively affect cell metabolism.

There are some 7,000 chemicals found in cigarette smoke including tar, carbon monoxide, hydrogen cyanide, ammonia, and radioactive compounds. Many of these are known to be harmful by themselves. This study highlights the even greater impact they have when combined.

Long term damage

The consequences of exposing a developing fetus to this toxic cocktail can be profound, including impaired growth, premature birth, hormonal imbalances, increased predisposition to metabolic syndrome, liver disease and even death.

The study is published in the Archives of Toxicology.

In a news release Dr. David Hay, one of the lead authors, said this result highlights yet again the dangers posed to the fetus by women smoking while pregnant or being exposed to secondhand smoke :

“Cigarette smoke is known to have damaging effects on the foetus, yet we lack appropriate tools to study this in a very detailed way. This new approach means that we now have sources of renewable tissue that will enable us to understand the cellular effect of cigarettes on the unborn foetus.”