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In a landmark study published in Nature Communications, scientists at Children’s Hospital Los Angeles demonstrate an in vitro kidney model that could change the course of research for diseases like CKD.

The kidney contains specialized structures called glomeruli. Within each glomerulus is a filtration barrier made up of two thin layers of highly specialized cells and a membrane that acts as a selective filter. As blood moves through each glomerulus, toxins and small molecules can pass through, while proteins and other important components are kept in the bloodstream. “This filtration process breaks down in patients with kidney disorders,” explains Laura Perin, PhD, who is co-senior author on the study along with Stefano Da Sacco, PhD. “But because we haven’t had a good in vitro model, we still don’t know the mechanisms of injury to the glomerulus in CKD.”

Dr. Perin and Dr. Da Sacco conduct research in the GOFARR Laboratory for Organ Regenerative Research and Cell Therapeutics in Urology along with co-director Roger De Filippo, MD, at CHLA’s Saban Research Institute. The lead author on the study was CHLA postdoctoral research fellow Astgik Petrosyan. Together, the team studies the structure of the glomerulus to better understand how and why their ability to filter blood breaks down.

“A big challenge in the kidney research field has been trying to replicate the glomerulus in vitro,” says Dr. Da Sacco. “In particular, the glomerular filtration barrier is very difficult to recreate in a lab using standard techniques.” Because of this, most published studies have used an artificial membrane between the two cell layers. While fluid can be exchanged, the cells cannot communicate across this membrane in the same way they do biologically. “This results in a model that doesn’t really filter properly,” he explains.

The critical component missing from current experiments is a filter that is selective and allows proper cell-to-cell communication. Dr. Da Sacco and Dr. Perin set out to grow healthy kidney cells in a way that allowed for the natural glomerular barrier to form, just as it does in the body. Using specialized, compartmented containers called OrganoplatesTM, the investigators did exactly that.

The result?

A model glomerulus that functions nearly identically to that found in real kidneys. They are calling this model, which is derived entirely from healthy, human kidney tissue, a glomerulus on a chip.

On one side of the cells, investigators add fluid and, on the other side, they collect what the ‘glomerulus’ filters, which is called the filtrate. In their experiment, the scientists added blood serum from healthy individuals. Without the use of a manufactured filter, the team’s in vitro glomerulus behaved as human kidneys are expected to act: proteins remained in the serum while smaller molecules passed into the filtrate. “The barrier that our cells naturally formed is selective, just as it would be in a fully-functioning kidney,” says Dr. Da Sacco. “It is remarkable.”

This model represents a substantial leap forward from the current standard of in vitro kidney research. “Our system behaves like a biologically, physiologically correct glomerulus,” says Dr. Perin. “This opens up the door for us to understand what we still don’t know – the molecular mechanisms of injury in CKD and, more importantly, how to prevent damage.”

While this seemed a distant goal in the past, Dr. Da Sacco and Dr. Perin are already recreating and studying the disease state in their model. When the investigators added serum from patients with CKD, they found that the glomerulus exhibited the same type of damage observed clinically: proteins began to leak through the compromised filter. Protein levels measured in the experimental filtrate matched patient clinical filtrate samples with a correlation of approximately 90%.

This breakthrough paves the way for numerous clinical applications. In the burgeoning era of personalized medicine, a preparation such as this can be used to examine molecular mechanisms of kidney damage in individual patients. Disease progression can also be monitored over time using serial blood sampling. Also, the model could be used for screening new drugs prior to human clinical testing.

Dr. Perin and Dr. Da Sacco, who are also Assistant Professors in the Keck School of Medicine at USC, are co-senior authors on the publication. Other authors are Paolo Cravedi of Icahn School of Medicine at Mount Sinai, NY; Valentina Villani of CHLA; Andrea Angeletti of the University of Bologna, Italy; Joaquin Manrique of Complejo Hospitalario de Navarra, Pamplona, Spain; Alessandra Renieri of Azienda Ospedaliera Universitaria Senese, Siena, Italy; and Roger De Filippo of CHLA.

The study was funded by the GOFARR Foundation and the Schenkman Family, grants from the Alport Syndrome Foundation, a TSRI Research Career Development Award, and a Wright Foundation Pilot Award. OrganoplatesTM were purchased from MIMETAS, who the authors wish to thank for providing invaluable training, technical support, and assistance.

Healthy human kidney cells (visible in red and green in this fluorescent image) grow in layers to form a filtration barrier in vitro. Image courtesy of Dr. Perin and Dr. Da Sacco.

Many infants born premature or with other complications often forego their first weeks or months at home for a stay in the neonatal intensive care unit. The NICCU is designed to deliver critical medical care to babies in need but can be traumatic for infants and their families, alike. In the Early Childhood Mental Health Program at Children’s Hospital Los Angeles, clinical psychologists Marian Williams, PhD, Patricia Lakatos, PhD, and a team of infant-family mental health specialists work towards greater mental health awareness in the NICCU.

Infants may not be the first age group called to mind in discussions of mental health. Yet, for babies in critical medical condition, Dr. Lakatos says an “infant mental health-informed perspective” could reduce stress and improve bonding with parents. This means not only focusing on the physical needs of the child but also the emotional and mental needs, not an easy task for newborn infants who cannot make their voices heard.

In an article published in Journal of Clinical Psychology in Medical Settings, Dr. Lakatos, Dr. Williams, and co-authors Tamara Matic, MD, and Melissa Carson, MD, advocate for a third component of the NICCU family – the relationship between baby and parents. “A lot of mental health work in NICCUs currently focus on either the mental health of parents or on the baby’s development,” says Dr. Williams, who is also the Director of the Stein Tikun Olam Infant-Family Mental Health Initiative at CHLA. “We also want to focus on the relationship between babies and their parents.”

Many parents of children in intensive care units experience symptoms of post-traumatic stress, which can threaten bonding with a newborn baby. In order to support the developing relationship between parents and their new baby, the CHLA infant mental health team turned to a model of intervention that has demonstrated success in families who have undergone trauma. Child-Parent Psychotherapy – or CPP – addresses the parent-child relationship directly, nurturing and advocating for it in its own right.

With funding from the Stein Tikun Olam Infant-Family Health Initiative, Drs. Williams and Lakatos, and the team were able to adapt CPP to the NICCU setting at Children’s Hospital Los Angeles. Their publication describes how the established, evidence-based CPP model can be used to nurture developing infant-parent relationships in the NICCU. While it has been implemented in other settings, CPP is not commonly integrated into NICCU patient care.

CPP is a flexible model that has multiple levels of intervention, depending upon individual family needs. Sessions with trained CPP providers can vary in number or duration, with the aim of restoring a developmental trajectory for parent and child. CPP providers advocate for mental health needs of parents and babies, working alongside their medical and social work colleagues. “When babies are in the hospital, we need to think about them, their parents, and their relationships,” says Dr. Lakatos.

Peterson went on to say that, at birth, the architecture of the brain is very sparse compared to what it will soon become. Soon after birth, he said, the brain cells begin to sprout and grow dendrites – the branching, bushy structures that receive information from other brain cells. Cells begin to connect to one another, forming dense, complex circuitry. “Unbelievably quickly,” Peterson continued, “in the first months of life, the spokes begin to fill in.” The architecture of the brain is being built at lightning speed in this first year of life. It is this growing complexity, he said, that allows new information to be passed from one part of the brain to another. This dense web of interconnected brain cells sets the very foundation of all emerging capacities in early childhood – things like walking, talking, thinking, and relating socially to others.

Sometimes, the brain is compared to a computer. This analogy speaks to the brain’s incredible computational capacity, though it does not acknowledge some of its more mysterious aspects– things like personality or a person’s ability to love. Peterson expanded upon the intrigue of the brain’s capabilities, explaining that the ability to learn and physically reshape itself sets it apart from its machine counterpart. “A one-year-old is just beginning to walk and say a few words. In the next year or so, that child will be capable of hundreds of words. No computer can teach itself to do that. This ability is all due to the developing architecture of the human brain.”

This ability to learn can be found in our DNA long before a brain even begins to develop. Written into the very code from which all our proteins, cells, and tissues are born is the ability of the brain to be molded by our surroundings, a stunning feat of adaptability. Unlike a computer, the human brain grows and learns in a way that is largely dependent upon experience, which actually shapes the physical architecture of the brain.  Peterson explained that the dense, bushy web of interconnected cells in a two-year-old’s brain is actually richer in connections than in an adult’s brain – 50% more dense. The environment, he said, guides the brain in which connections will survive and become permanent, while pruning the irrelevant circuits. “It’s a dance between the developing organism, with all its hard-wired programming, and the environment – each interacts with, and affects, the other.” What we are left with is imagery of a blossoming array of neural connections that begins to explode within the first two years of life, and is then masterfully whittled away as we are shaped by our experience. The mature brain is perhaps less a computer than it is a sculpted work of art made by our genes and the world that surrounds us.

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“We have learned a lot about how to prevent the large vessel strokes,” says Dr. Wood. He is referring to strokes that have immediate, catastrophic effects such as blindness or paralysis. “But patients with SCD still suffer from silent strokes.” Though they are called silent, these strokes can have debilitating effects on the brain’s executive function, which is the ability to execute complex tasks needed for things like maintaining a job or doing well in school. On the surface, the reason for these strokes seems clear: damaged blood cells carry less oxygen, which causes oxygen deprivation in the brain. Yet, Dr. Wood measured total oxygen delivery to the brain and found no decrease in SCD patients. This is because the body compensates for decreased blood oxygen by increasing total blood flow to the brain.

Dr. Wood was left with the question of why patients with SCD continued to suffer strokes. “I began to think, ‘maybe there’s a distribution problem,’” Dr. Wood says. “Total blood flow is fine, but is it going where it needs to go?” Using an advanced imaging technique called arterial spin labeling to measure blood flow, Dr. Wood’s team found a disparity among brain areas. While total oxygen delivery to the brain was unchanged, oxygen delivery to the white matter was reduced by more than a third – 35% – in patients with SCD. Critically, this part of the brain – the white matter –  is where the majority of silent strokes occur in these patients.

Grey matter is composed of neurons, the brain cells that store information. White matter is the network of highways in the brain that neurons use to transmit this information. Dr. Wood’s finding demonstrates that the body differentiates between grey and white matter, clearly prioritizing neurons. This makes sense because keeping neurons alive is critical for survival. Strokes in grey matter are immediately debilitating, while strokes in white matter appear silent because they simply cause informational processing to slow. Though they may not cause paralysis or major motor deficits, effects of white matter silent strokes can greatly impede important aspects of a person’s day-to-day life. Dr. Wood’s study gives us a much better understanding of why these strokes are occurring, knowledge that is necessary before medicine can effectively treat this aspect of the disease.

Results of this study were published recently in the American Journal of Hematology by lead author Yaqiong Chai, a senior PhD candidate. The study was funded by the National Heart Lung and Blood Institute of the National Institutes of Health.

Other authors on the study include Adam M Bush, PhD; Julie Coloigner, PhD; Aart J Nederveen, PhD; Benita Tamrazi, MD; Chau Vu, MS; Soyoung Choi; Thomas D Coates, MD; and Natasha Lepore, PhD.

Dr. Harrison works in the laboratory of Ching-Ling (Ellen) Lien, PhD, where the overall research goal is to uncover the mechanisms used by zebrafish to recover from injury. The answers gained from this research could help develop new treatments for babies with congenital heart defects or individuals who have suffered heart attack or other cardiac injury.

In addition to arteries and veins, many animals – including humans – have a network of lymphatic vessels. This system clears fluid and debris from tissue. Their malfunction may cause fluid build-up or limited clearing of damaged cells after heart injury in humans. “We don’t see that occurring in the zebrafish,” says Dr. Harrison, “so we began to investigate the cardiac lymphatic system in our laboratory.”

Lymphatic vessels are clear, unlike their larger and more colorful blood vessel counterparts. In addition to being hard to see literally, they have also flown under the radar in cardiac regeneration research. In fact, it was not known whether zebrafish even have cardiac lymphatic vessels. Dr. Harrison and Dr. Lien determined that zebrafish do have these vessels and that they could be a critical factor in heart repair. These findings suggest that the cardiac lymphatic system could be an important research focus in the field of cardiac repair.

In zebrafish with heart damage, the team made a striking observation – the cardiac lymphatic vessels grew and expanded to cover the wound area. When the investigators blocked this process, the fish were unable to repair their heart damage. “These experiments showed that the lymphatic vessels in the zebrafish heart are doing something critical,” says Dr. Harrison, “there’s definitely an important role there.”

Characterizing this role will be the subject of ongoing research. Understanding how the lymphatic vessels are aiding in repair could point scientists toward a treatment target. “We are looking very closely at these vessels, says Dr. Lien. “We think this is perhaps a new frontier in heart regeneration research.”

New information systems at CHLA and other institutions nationwide are focusing on collecting data into a central warehouse of sorts, a big data repository, from which it can be analyzed for research and clinical purposes. Instead of identifying problems and then collecting data based on the nature of the questions at hand, researchers can now identify relevant data, generate questions and then mine the data to find the answer.

For example, scientists at CHLA recently developed a pandemic disaster triage methodology based on over 150,000 critically ill children and conducted a computer simulated emergency in children that demonstrated 25%  improved survival compared to current triage models.

However, there are precautions and concerns to address when it comes to utilizing big data methods in medicine. Data security, particularly when involving patient information, is a must according to Wetzel, as well as avoiding disruption of the clinical workflow.