Photo by Glen Carrie at Unsplash
Each of the following research stories are highlights selected from previous editions of “Regenerative Medicine News Under the Microscope.” Vote for the one that inspired you most this year!
- Vision science: Making organoids cry
This story is nothing to blink at.
While many of us don’t think much about our tear glands on any given day, they are critical organs that keep our eyes well-lubricated. Why is this important? Well, when tear glands malfunction, dry eye syndrome might result, leaving patients with irritated eyes, blurred vision and, in severe cases, corneal damage.
At the moment, mild cases of dry eye syndrome can be treated with eye drops, but serious cases may require surgery. Surprisingly, the limited nature of these options is due to the fact that we don’t know much about lacrimal glands, or tear glands.
Bannier-Hélaouët et al. developed both human and mouse lacrimal gland organoids; showed that the gene Pax6 is a must for differentiation of this cell type; offered up a single cell atlas of both human lacrimal gland organoids and dissected tissue (scRNAseq buffs: they also conducted an RNA velocity analysis); and, proved that their organoids can cry, just like the real organs. To see what this looks like, look no further than the gif at the top of this blog post! The organoids are the spheroid structures shown. Our tear glands respond to a mix of neurotransmitters released by our bodies when we’re about to cry. The researchers replicated this effect by simply applying the neurotransmitters to their organoids, causing the structures to fill up with tear fluid, thus expanding! Why do they expand? The organoids themselves don’t have structurally sound ducts to outlet the fluid so, instead, the tears are trapped inside.
To top it all off, the researchers transplanted their dish-grown human tear glands into mice, and showed that they seem to self-organize into the host and engraft successfully. They even see the formation of a structure resembling a duct, and the accumulation of tear proteins therein. Though the transplant evidence was preliminary, more studies will reveal how safe and how truly functional such an approach might be. For further reading, check out this article by Nature’s Heidi Ledford, which features additional details concerning how salivary gland transplants may set the stage for tear glands in the future, and just what exactly crocodile tears have to do with this story!
- The terrestrial benefits of stem cell research – in space
Have you ever wondered what happens to stem cells in microgravity?
An investigation by NASA called MVP Cell-03 was designed to answer this question. Heart precursor cells are cultured on the International Space Station to determine how cell survival and cell proliferation is affected by microgravity. Interestingly, prior studies had shown that culturing these cells in simulated microgravity actually increased their production efficiency.
As you can imagine, sending cells up to space without killing them is a difficult task. How did scientists solve this problem? The same way we protect cells when shipping them between cities and countries: cryopreservation. Not only was this protocol not found to affect the cells adversely, but it also protected the cells from the extreme gravity during launch.
How does this help us earthling researchers? The team also developed a culture medium that supports cells without necessitating carbon dioxide. Carbon dioxide adds mass to any payload – more mass, more money spent.
The above strategies were successfully applied in March 2020 to generate beating heart cells – in space – which journeyed back to Earth after 22 days in orbit. NASA reported the results this year so that the findings might help other teams, both on the ground and in the sky.
- The “super liver”
You might not expect me to start an item about a “super liver” by telling you about the pancreas, but that’s what’s about to happen.
Our pancreas, which is just behind the stomach, produces critical enzymes that aid in digestion, plus hormones for sugar processing, aka insulin. A condition called pancreatitis will strike some of us, resulting in pathological inflammation of the organ. It can present and resolve quickly, or it can become a chronic condition, causing cellular damage, reduced pancreatic function, and severe pain. Some cases can even be life-threatening. In extreme cases the pancreas may be completely removed, relieving the patient of pain; however, this will render the patient diabetic and in need of insulin from external sources. If a patient doesn’t want to go down this route, they may end up on heavy narcotics to manage the pain.
Enter a relatively new treatment: total pancreatectomy and islet autotransplantation. This involves the complete removal of the pancreas followed by extraction of the insulin-producing cells from the inflamed organ, and injection of these cells into the liver. There, they will take root and thrive. This regenerative medicine solution both relieves the painful condition and satisfies the patient’s need for insulin using their own cells.
This year, CTV reported on one of the first two patients to undergo this procedure at Toronto General Hospital. Seeing as the incidence of pancreatitis seems to be increasing worldwide for unknown reasons, this treatment option is an invaluable asset to our city. By the way, the islet cells were isolated, purified and concentrated at CCRM’s Centre for Cell and Vector Production, a Good Manufacturing Practices-compliant facility. The cells were then returned to the hospital to be implanted into the patient’s liver.
- A significant field-first: In vitrogametogenesis
Prior to this year, the only way to produce female gametes outside of the body was by incubating a stem-cell-derived primordial egg cell with harvested tissue from an animal’s reproductive organ – the ovary. That has changed, thanks to this work, published in Science.
Yes, that’s right, it’s another “-oid.”
The “ovarioid,” derived from stem cells, provides the niche necessary for oocytes to grow and mature into eggs in vitro. The authors fertilized some of them to prove they were functional, and healthy newborn mice were born.
The significance of this work is that a few decades in the future, this technology might help women who have, for whatever reason, lost reproductive function. They could have eggs grown in a lab with their own genetic material. It could also help couples in the LGBTQIA2S+ community produce children who share their genetics.
Of course, whenever a new technology is developed with the power to create or edit a life, ethical concerns are raised. More on that here.
- SARS-CoV-2 and the central nervous system: Another new organoid model
It seems we’re quickly approaching a point where we’ll have a full fleet of in vitro systems we can use to examine the effects of COVID-19 on our physiology. I’ve talked about many of these in previous blogs.
The one I’d like to highlight here is the neural-perivascular assembloid, which was developed to determine how exactly the virus is able to cross the blood-brain barrier (BBB) to trigger ischemic strokes, seizures, hemorrhaging, and much more. All in all, central nervous system (CNS) symptoms present in up to 85% of intensive care unit patients.
Human neurons themselves are not susceptible to infection by SARS-CoV-2, so another cell type needed to be identified; the “Trojan horse,” identified in Nature Medicine this year, turned out to be pericytes.
Assembloids are essentially cortical organoids with pericytes introduced into the system, making for a more sophisticated model. These cells wrap around the blood vessel and are vulnerable to infection due to expression of the ACE2 receptor. They’re positioned such that they form an interface between the blood vessel and astrocytes, which have been found to be susceptible as well. Pericytes serve to form a connection between neighboring cells, plus coordinate and regulate signaling critical for CNS functions including BBB permeability and neuroinflammation.
Another possibility is that infection of these cells may lead to inflammation of the blood vessels, which in turn may result in clotting, stroke, or hemorrhaging.
Next steps include attempts to develop assembloids which include blood vessels to produce a more accurate model.
- Liver regeneration: Cracking a two-thousand-year-old mystery
The liver’s regenerative properties have been known to us since ancient times (see: The ancient Greek myth of Prometheus). It’s the only organ that can be cut down to one third of its size and still grow back. It’s a remarkable capability that has been the subject of investigation for years, mostly because it could hold the key to human regeneration more broadly.
This year, we finally have some answers on the matter, and it isn’t a totally unexpected result: The key seems to have been in the epigenetic code all along.
Researchers at NYU Abu Dhabi found that while the liver was in a state of quiescence, an epigenetic mark that represses the expression of certain proliferation-related genes was present (H3K27me3, a tri-methylation of histone H3 on lysine residue 27). However, when the liver is regenerating, this repressive epigenetic mark is removed from these particular gene promoters, facilitating expression.
The authors of the study suggest that in order to manipulate this system, more work will be required, but this is a strong step in the right direction.
- When science stares back: 2021’s latest organoids
I’m sure we all remember August’s Pick of the Month: brain organoids with impressive, light-sensitive optic vesicles, termed “optic vesicle-containing brain organoids” (OVB-organoids).
Researchers who study the eye can often be overheard saying that a person’s eyes are the only part of the brain you can see without… well, intervening. This is because your retina and optic nerve develop as outgrowths of the brain. In fact, the retina is technically part of your central nervous system.
Framed by this context, OVB-organoids make plenty of sense. By modifying the culture conditions usually employed to grow brain organoids, researchers created an in vitro environment conducive to the continued development and specification of the organoid, allowing for the generation of bilaterally symmetric optic vesicles over the course of 60 days. These extra structures contained functional circuitry and were composed of cells expressing genes related to the developing lens, corneal epithelium, routing of axons at the optic chiasm, the retina, retinal ganglion cells, and more.
One of my favourite parts: The researchers test for light sensitivity using electroretinography, a technique that allows us to record and quantify retinal signaling. When they changed the light intensity, the cells would respond accordingly in a dose-dependent manner.
Future studies might address the outlined limitations of this work, as follows: first, the viability of OVB-organoids past 60 days is apparently questionable, meaning that this model can’t necessarily be used to study mature retinal cells. Also, not all induced pluripotent stem cell donor cells were equally consistent in yielding these organoids following their protocol, so this may be another avenue requiring investigation and optimization.
At this stage, the organoids may be useful in generating retinal pigmented epithelium for transplants by reprogramming patient cells, and may help model the retinopathies associated with early neurodevelopmental disorders.
If, like me, you’re familiar with retinal organoids and are wondering what advantage this system might offer in comparison, here’s the brief version: The optic vesicle-like tissues generated by retinal organoids must be excised and further cultured for a period of several weeks, and thus do not form an in vivo-like structure. You lose the influence of surrounding tissues like the forebrain on the developmental process. Retinal organoids are still remarkable model systems, but each in vitro culture has its pros and cons.
- Tissue regeneration may not be a stretch: Blocking mechanotransduction enhances wound healing and regeneration
The achievement of scar-less tissue regeneration has been referred to as the holy grail of biomedical research – it’s that critical. In human tissue, healing is achieved through fibrosis and scar formation, not regeneration. Paradoxically, this can adversely affect the organ – take a heart attack, for instance. The subsequent formation of scar tissue on the heart can lead to congestive heart failure and arrhythmias, both potentially deadly conditions. Other examples include liver cirrhosis and fibrosis of the lungs. Taken together, fibrosis costs us billions of health-care dollars each year and can cause lifelong disabilities, furthering economic impact.
As large mammals, our tissues have evolved to accommodate significant mechanical forces associated with our size. If we look at human skin, it withstands significant mechanical stress as we move, and has adapted by employing hypertrophic healing responses that result in the formation of scar tissue that’s essentially dysfunctional, characterized by a thicker dermis, increased stiffness, an absence of features like hair follicles and intradermal adipose tissue. Pigs are similar to us in this sense, so Chen et al. studied porcine wound healing responses in this important Nature Communications paper.
After inhibiting focal adhesion kinase (FAK), a molecular force transducer, the authors were able to promote regenerative healing and, by that, I mean that they observed a restoration in biomechanical properties of the skin, re-growth of hair follicles, and normal collagen architecture. They also aimed to confirm their findings in human tissue by studying 3D cultured human fibroblasts at the transcriptomic level, finding that FAK can pull fibroblasts away from fibrotic transcriptional states and activate regenerative phenotypes.
Interestingly, pharmacological FAK inhibition has been shown to be safe in clinical trials for cancer treatment, so there’s real promise here for the future of regenerative medicine.
- A new hope for intervertebral disc disease
Between the bones of your spine lie shock-absorbing, fibro-cartilaginous discs that keep the spine flexible and are designed to resist strong forces on different axis of motion. Degenerative disc disease (DDD), though less a disease and more a condition, occurs when the spinal discs begin to break down. Unfortunately, it is one of the leading causes of disability worldwide. Though these discs show wear with age in everyone, some of us will accumulate more damage than others, causing pain ranging from mild to disabling. Progressive wear can also lead to complications like a herniated or bulging disc.
Treatment options for DDD include nonsurgical and surgical interventions, including cell-based therapies that have shown promise. Of special interest here is the injection of mesenchymal stem cell-derived chondrocytes, which are responsible for cartilage formation. However, there have been some limitations to this treatment, including hypertrophy of the differentiated chondrocytes, resulting in undesirable outcomes including apoptosis (cell death), abnormal formation, and extracellular matrix calcification. Such complications may lead to graft failure and, ultimately, unsuccessful tissue regeneration.
Bello et al. aimed to advance this promising therapy by developing a way to circumvent negative outcomes. They incorporated gelatin microparticles that were co-loaded with matrilin 3 and transforming growth factor beta 3 into mesenchymal cell spheroids (3D cell clusters or aggregates). This promoted differentiation to chondrocytes while simultaneously preventing hypertrophy. This strategy also induced the release of cytokines that promoted regeneration in vitro. The intervention worked in vivo as well, promoting regeneration of the disc tissue in rodent models.
- Quantitative 3D-imaging platform for the cranial microvascular environment: Visualization at single-cell resolution
The dynamic spatial interaction between blood vessels and osteoprogenitors during critical processes, including the growth of craniofacial bone, its healing processes, and remodeling, posed a serious knowledge gap prior to this year. An understanding of these physiological phenomena will contribute to the development of treatments for vascular abnormalities in craniofacial bone, which have been linked to syndromes including cleft palate, craniosynostosis (aberrantly early closure of skull sutures that, in its most severe forms, can lead to physical deformities, visual/sleeping impairments, headaches and developmental delays), mandibular hypoplasia (underdevelopment of the lower jaw), and more. In future, advanced regenerative medicine solutions for both congenital conditions and injuries will greatly benefit from, and will be informed by, research into the craniofacial regenerative niche and its healing processes.
Historically, older imaging technologies have limited the amount of information we’ve been able to collect in this field, but researchers have capitalized on a combination of newer technologies to overcome these limitations. Using whole-mount immunostaining, optical tissue clearing, light-sheet microscopy, and 3D image analysis, Rindone et al. built maps of vessel subtypes and skeletal progenitors in frontoparietal bones at single-cell resolution. They then used this technology to show how the spatial distribution of these elements vary during growth, remodeling and healing. In addition, the authors emphasize that this technology can be extended to study other cell types not explored here, such as neurons and immune cells, highlighting that this is truly the beginning of a series of investigations that wouldn’t have been possible in previous years.
- Paralyzed mice walk again thanks to reparative “dancing” molecules
The video is stunning. A mouse shuffles towards a researcher, sluggishly putting one hindpaw ahead of the other, in eager pursuit of an object. Looking at the little guy, it’s hard to imagine that this mouse had a spinal cord injury that eliminated any and all movement in its hind limbs, and that this injury had been largely healed following treatment with a sophisticated new injectable therapy. In the original video on YouTube, you’ll also see an untreated paraplegic mouse that can only pull itself forward using its forepaws, highlighting just how significant the movements in the clip above truly are.
When I think of “regenerative medicine,” restoring movement to paralyzed individuals comes to mind as one of our ultimate goals in the field. This year in Science, Álvarez et al. do just this in rodent models of severe spinal cord injury.
What sets their molecularly complex treatment apart? They engineered short, modified peptides that promote axon regeneration, cell proliferation and blood vessel formation – but most importantly, the peptides are designed to remain on the move. This makes contact with receptors in the tissue more likely, because the receptors are moving around in the membrane as well. The treatment is a liquid when injected into the spinal cord, but when it comes into contact with the body, a hydrogel is formed that mimics the spinal cord’s extracellular matrix. That being said, the molecules themselves are contained within a nanofiber network, and yet they’re able to move around within it. This novel approach, described by the authors as a “supramolecular peptide fibril scaffold,” was the key to their success.
Mice were able to achieve functional recovery four weeks after a single injection; the reason is that the treatment stimulated: 1) the regeneration of severed axons in the spine; 2) re-formation of the myelin sheath around neurons; 3) growth of blood vessels in the injury site; 4) better motor neuron survival; and, 5) reduced scar tissue formation.
The icing on the cake? The treatment is biodegradable, breaking down into nutrients for the patient’s cells within 12 weeks and then disappearing.
In early 2022, the authors will be approaching the Food and Drug Administration to request permission for human trials.
Click here to cast your vote! The poll will close on December 31st, 2021.
Lyla El-Fayomi
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