Illuminating the Invisible: Bringing the Smallest Protein Clusters into Focus

by Ian Scheffler

The bright white spots represent tiny clusters of proteins detected by CluMPS. (Photo by: Thomas Mumford)

Penn Engineers have pioneered a new way to visualize the smallest protein clusters, skirting the physical limitations of light-powered microscopes and opening new avenues for detecting the proteins implicated in diseases like Alzheimer’s and testing new treatments.

In a paper in Cell Systems, Lukasz Bugaj, Assistant Professor in Bioengineering, describes the creation of CluMPS, or Clusters Magnified by Phase Separation, a molecular tool that activates by forming conspicuous blobs in the presence of target protein clusters as small as just a few nanometers. In essence, CluMPS functions like an on/off switch that responds to the presence of clusters of the protein it is programmed to detect.

Normally, says Bugaj, detecting such clusters requires laborious techniques. “With CluMPS, you don’t need anything beyond the standard lab microscope.” The tool fuses with the target protein to form condensates orders of magnitude larger than the protein clusters themselves that resemble the colorful blobs in a lava lamp. “We think the simplicity of the approach is one of its main benefits,” says Bugaj. “You don’t need specialized skills or equipment to quickly see whether there are small clusters in your cells.”

Read the full story in Penn Engineering Today.

Penn Partners in Multi-University Research Center Supporting Healthy Pregnancies

by Andrew Smith

How does the placenta keep harmful substances away from developing babies while still providing proper nutrition?

(Photo: Getty Images)

The exact mechanisms remain unknown, which is why the University of Pennsylvania, Rutgers University, Tulane University, the University of North Carolina at Chapel Hill and the University of Rochester have joined together to launch a research center dedicated to solving this mystery and ensuring healthy pregnancies.

A $5 million grant from the National Institutes of Health (NIH) will help fund the Integrated Transporter Elucidation Center (InTEC), which will operate from the Rutgers Biomedical Health Sciences campus in Piscataway under the leadership of Lauren Aleksunes, a professor of pharmacology and toxicology at Rutgers’ Ernest Mario School of Pharmacy and resident scientist in the Environmental and Occupational Health Sciences Institute (EOHSI).

“Since my time working as a community pharmacist, I have found the lack of high-quality information about the safety of everyday products on the health of a pregnancy frustrating,” says Aleksunes.  “People need to know whether the chemicals in their diet, personal care products and medications can impact their babies. Our goal at InTEC is to better understand how these chemicals travel in and out of the placenta and if they can reach the baby and influence their development.”

Aleksunes will study how transporter proteins carrying nutrients, dietary supplements, medications and toxic chemicals work during pregnancies. Some of the work will test how individual placenta cells respond to various stimuli in the laboratory. Others on the team will examine how environmental factors influence placental transporters during healthy and unhealthy or complicated pregnancies. 

Key to this work will be Dan Huh, Associate Professor in Bioengineering in Penn Engineering, who will lead a team with an innovative approach to modeling the transfer of molecules across the human placenta. 

As a pioneer of organ-on-a-chip technology, the Huh group will use a novel microengineered system in which maternal tissue engineered from a layer of primary human trophoblasts is grown adjacent to a three-dimensional network of perfusable fetal blood vessels to mimic the human placental barrier. This microphysiological system will be employed as an in vitro platform to simulate and quantitatively analyze the exchange of various substances between maternal and fetal circulation without the need for laboratory animals or placenta explants.

Read the full story in Penn Engineering Today.

Two Penn Bioengineers Receive NIH Director Award

by Nathi Magubane

Jina Ko (left) and Kevin Johnson (right), both from the School of Engineering and the Perelman School of Medicine with appointments in Bioengineering, have received the National Institute of Health Director’s Award to support their “highly innovative and broadly impactful” research projects through the High-Risk, High-Reward program.

The National Institutes of Health (NIH) has awarded grants to three researchers from the University of Pennsylvania through the NIH Common Fund’s High-Risk, High-Reward Research program. The research of Kevin B. Johnson, Jina Ko, and Sheila Shanmugan will be supported through the program, which funds “highly innovative and broadly impactful” biomedical or behavioral research by exceptionally creative scientists.

The High-Risk, High-Reward Research program catalyzes scientific discovery by supporting highly innovative research proposals that, due to their inherent risk, may struggle in the traditional peer-review process despite their transformative potential. Program applicants are encouraged to think “outside the box” and pursue trail-blazing ideas in any area of research relevant to the NIH’s mission to advance knowledge and enhance health.

Two Penn Bioengineering faculty, Johnson and Ko, are among 85 recipients for 2023.

Johnson, the David L. Cohen University Professor of Pediatrics, is a Penn Integrates Knowledge University Professor who holds appointments in the Department of Computer and Information Science in the School of Engineering and Applied Science and the Department of Biostatistics, Epidemiology, and Informatics in the Perelman School of Medicine. He also holds secondary appointments in Bioengineering, Pediatrics, and in the Annenberg School for Communication. He is widely known for his work with e-prescribing and computer-based documentation and, more recently, work communicating science to lay audiences, which includes a documentary about health-information exchange. Johnson has authored more than 150 publications and was elected to the American College of Medical Informatics, Academic Pediatric Society, National Academy of Medicine, International Association of Health Science Informatics, and American Institute for Medical and Biological Engineering.

Ko is an assistant professor in the Department of Pathology and Laboratory Medicine in the Perelman School of Medicine and Department of Bioengineering in the School of Engineering and Applied Science. She focuses on developing single molecule detection from single extracellular vesicles and multiplexed molecular profiling to better diagnose diseases and monitor treatment efficacy. Ko earned her Ph.D. in bioengineering at Penn in 2018, during which time she developed machine learning-based microchip diagnostics that can detect blood-based biomarkers to diagnose pancreatic cancer and traumatic brain injury. For her postdoctoral training, she worked at the Massachusetts General Hospital and the Wyss Institute at Harvard University as a Schmidt Science Fellow and a NIH K99/R00 award recipient. Ko developed new methods to profile single cells and single extracellular vesicles with high throughput and multiplexing.

Read the full announcement in Penn Today.

Understanding the Physics of Kidney Development

Abstract image of tubules repelling each other and shifting around.
The model of tubule packing developed by the Hughes Lab shows the tubules repelling each other and shifting around.

A recent study by Penn Bioengineering researchers sheds new light on the role of physics in kidney development. The kidney uses structures called nephrons and tubules to filter blood and pass urine to the bladder. Nephron number is set at birth and can vary over an order of magnitude (anywhere from 100,000 to over a million nephrons in an individual kidney). While the reasons for this variability remain unclear, low numbers of nephrons predispose patients to hypertension and chronic kidney disease. 

Now, research published in Developmental Cell led by Alex J. Hughes, Assistant Professor in the Department of Bioengineering, demonstrates a new physics-driven approach to better visualize and understand how a healthy kidney develops to avoid organizational defects that would impair its function. While previous efforts have typically approached this problem using molecular genetics and mouse models, the Hughes Lab’s physics-based approach could link particular types of defects to this genetic information and possibly highlight new treatments to prevent or fix congenital defects.

During embryonic development, kidney tubules grow and the tips divide to make a branched tree with clusters of nephron stem cells surrounding each branch tip. In order to build more nephrons, the tree needs to grow more branches. To keep the branches from overlapping, the kidney’s surface grows more crowded as the number of branches increase. “At this point, it’s like adding more people to a crowded elevator,” says Louis Prahl, first author of the paper and Postdoctoral Fellow in the Hughes Lab. “The branches need to keep rearranging to accommodate more until organ growth stops.”

To understand this process, Hughes, Prahl and their team investigated branch organization in mouse kidneys as well as using computer models and a 3D printed model of tubules. Their results show that tubules have to actively restructure – essentially divide at narrower angles – to accommodate more tubules. Computer simulations also identified ‘defective’ packing, in which the simulation parameters caused tubules to either overlap or be forced beneath the kidney surface. The team’s experimentation and analysis of published studies of genetic mouse models of kidney disease confirmed that these defects do occur.

This study represents a unique synthesis of different fields to understand congenital kidney disease. Mathematicians have studied geometric packing problems for decades in other contexts, but the structural features of the kidney present new applications for these models. Previous models of kidney branching have approached these problems from the perspective of individual branches or using purely geometric models that don’t account for tissue mechanics. By contrast, The Hughes Lab’s computer model demonstrates the physics of how tubule families interact with each other, allowing them to identify ‘phases’ of kidney organization that either relate to normal kidney development or organizational defects. Their 3D printed model of tubules shows that these effects can occur even when one sets the biology aside.

Hughes has been widely recognized for his research in the understanding of kidney development. This new publication is the first fruit of his 2021 CAREER Award from the National Science Foundation (NSF) and he was recently named a 2023 Rising Star by the Cellular and Molecular Bioengineering (CMBE) Special Interest Group. In 2020 he became the first Penn Engineering faculty member to receive the Maximizing Investigators’ Research Award (MIRA) from the National Institutes of Health (NIH) for his forward-thinking work in the creation of new tools for tissue engineering.

Pediatric nephrologists have long worked to understand the cause of these childhood kidney defects. These efforts are often confounded by a lack of evidence for a single causative mutation. The Hughes Lab’s approach presents a new and different application of the packing problem and could help answer some of these unsolved questions and open doors to prevention of these diseases. Following this study, Hughes and his lab members will continue to explore the physics of kidney tubule packing, looking for interesting connections between packing organization, mechanical stresses between neighboring tubule tips, and nephron formation while attempting to copy these principles to build stem cell derived tissues to replace damaged or diseased kidney tissue. Mechanical forces play an important role in developmental biology and there is much scope for Hughes, Prahl and their colleagues to learn about these properties in relation to the kidney.

Read The developing murine kidney actively negotiates geometric packing conflicts to avoid defects” in Developmental Cell.

Other authors include Bioengineering Ph.D. students and Hughes Lab members John Viola and Jiageng Liu.

This work was supported by NSF CAREER 2047271, NIH MIRA R35GM133380, Predoctoral Training Program in Developmental Biology T32HD083185, and NIH F32 fellowship DK126385.

Bushra Raj Receives NIH Grant Through High-risk, High-reward Research Program

Bushra Raj, Ph.D.

Eight researchers from the Perelman School of Medicine have received research grants designed to invest in high-risk, high-reward projects.

Bushra Raj, Assistant Professor of Cell and Developmental Biology in the Perelman School of Medicine and member of the Penn Bioengineering Graduate Group, was one of three Penn winners of the NIH Director’s New Innovator Award for independent projects developed by early-career investigators. More additional Penn scientists who received NIH Director’s Transformative Research Award for a project focusing on cancer research.

Raj’s project focuses on “testing a novel technology that uses CRISPR/Cas gene-editing tools to genomically record inputs from two signaling pathways in the developing zebrafish brain.”

Established in 2009, the Transformative Research Award promotes cross-cutting, interdisciplinary science and is open to individuals and teams of investigators who propose research that could potentially create or challenge existing paradigms.

Read the full list of grant recipients in Penn Medicine News.

Defining Neural “Representation”

by Marilyn Perkins

Neuroscientists frequently say that neural activity ‘represents’ certain phenomena, PIK Professor Konrad Kording and postdoc Ben Baker led a study that took a philosophical approach to tease out what the term means.

Monitors Show EEG Reading and Graphical Brain Model. In the Background Laboratory Man Wearing Brainwave Scanning Headset Sits in a Chair with Closed Eyes. In the Modern Brain Study Research Laboratory
Neuroscientists use the word “represent” to encompass multifaceted relationships between brain activity, behavior, and the environment.

One of neuroscience’s greatest challenges is to bridge the gaps between the external environment, the brain’s internal electrical activity, and the abstract workings of behavior and cognition. Many neuroscientists rely on the word “representation” to connect these phenomena: A burst of neural activity in the visual cortex may represent the face of a friend or neurons in the brain’s memory centers may represent a childhood memory.

But with the many complex relationships between mind, brain, and environment, it’s not always clear what neuroscientists mean when they say neural activity “represents” something. Lack of clarity around this concept can lead to miscommunication, flawed conclusions, and unnecessary disagreements.

To tackle this issue, an interdisciplinary paper takes a philosophical approach to delineating the many aspects of the word “representation” in neuroscience. The work, published in Trends in Cognitive Sciences, comes from the lab of Konrad Kording, a Penn Integrates Knowledge University Professor and senior author on the study whose research lies at the intersection of neuroscience and machine learning.

“The term ‘representation’ is probably one of the most common words in all of neuroscience,” says Kording, who has appointments in the Perelman School of Medicine and School of Engineering and Applied Science. “But it might mean something very different from one professor to another.”

Read the full story in Penn Today.

Konrad Kording is a Penn Integrates Knowledge University Professor with joint appointments in the Department of Neuroscience the Perelman School of Medicine and in the Department of Bioengineering in the School of Engineering and Applied Science.

Ben Baker is a postdoctoral researcher in the Kording lab and a Provost Postdoctoral Fellow. Baker received his Ph.D. in philosophy from Penn.

Also coauthor on the paper is Benjamin Lansdell, a data scientist in the Department of Developmental Neurobiology at St. Jude Children’s Hospital and former postdoctoral researcher in the Kording lab.

Funding for this study came from the National Institutes of Health (awards 1-R01-EB028162-01 and R01EY021579) and the University of Pennsylvania Office of the Vice Provost for Research.

Konrad Kording’s CENTER is Part of a New NIH Education Initiative on Scientific Rigor

by Melissa Pappas

Konrad Kording (Photo by Eric Sucar)

In 2005, John Ioannidis published a bombshell paper titled “Why Most Published Research Findings Are False.” In it, Ioannidis argued that a lack of scientific rigor in biomedical research — such as poor study design, small sample sizes and improper assessment of the significance of data— meant that a large percentage of experiments would not return the same results if they were conducted again.

Since then, researchers’ awareness of this “replication crisis” has grown, especially in fields that directly impact the health and wellbeing of people, where lapses in rigor can have life-or-death consequences. Despite this attention and motivation, however, little progress has been made in addressing the roots of the problem. Formal training in rigorous research practices remains rare; while mentors advise their students on how to properly construct and conduct experiments to produce the most reliable evidence, few educational resources exist to support them.

To address this discrepancy, the National Institute of Neurological Disorders and Stroke (NINDS), part of the National Institutes of Health (NIH), has launched the Initiative to Improve Education in the Principles of Rigorous Research.

Konrad Kording, a Penn Integrates Knowledge Professor with appointments in the Departments of Bioengineering and Computer and Information Science in Penn Engineering and the Department of Neuroscience in Penn’s Perelman School of Medicine, has been awarded one of the initiative’s first five grants.

“The replication crisis is real,” says Kording. “I’ve tried to replicate the research of others and failed. I’ve reanalyzed my own data and found major mistakes that needed to be corrected. I was never properly taught how to do rigorous science, and I want to improve that for the next generation.”

Read the full story in Penn Engineering Today.

With NIH Pioneer Award, Jennifer E. Phillips-Cremins Will Study Genome Folding’s Role in Long-term Memory

by Evan Lerner

Jennifer E. Phillips-Cremins (upper left) and members of her lab.

Each year, the National Institutes of Health (NIH) recognizes exceptionally creative scientists through its High-Risk, High-Reward Research Program. The four awards granted by this program are designed to support researchers whose “out of the box” and “trailblazing” ideas have the potential for broad impact.

Jennifer E. Phillips-Cremins, Associate Professor and Dean’s Faculty Fellow in Penn Engineering’s Department of Bioengineering and the Perelman School of Medicine’s Department of Genetics, is one such researcher. As a recipient of an NIH Director’s Pioneer Award, she will receive $3.5 million over five years to support her work on the role that the physical folding of chromatin plays in the encoding of neural circuit and synapse properties contributing to long-term memory.

Phillips-Cremins’ award is one of 106 grants made through the High-Risk, High-Reward program this year, though she is only one of 10 to receive the Pioneer Award, which is the program’s largest funding opportunity.

“The science put forward by this cohort is exceptionally novel and creative and is sure to push at the boundaries of what is known,” said NIH Director Francis S. Collins.

Phillips-Cremins’ research is in the general field of epigenetics, the molecular and structural modifications that allow the genome — an identical copy of which is found in each cell — to express genes differently at different times and in different parts of the body. Within this field, her lab focuses on higher-order folding patterns of the DNA sequence, which bring distant sets of genes and regulatory elements into close proximity with one another as they are compressed inside the cell’s nucleus.

Previous work from the Cremins lab has investigated severe genome misfolding patterns common across a class of genetic neurological disorders, including fragile X syndrome, Huntington’s disease, ALS and Friedreich’s ataxia.

With the support of the Pioneer Award, she and the members of her lab will extend that research to a more fundamental question of neuroscience: how memory is encoded over decades, despite the rapid turnover of the relevant proteins and RNA sequences within the brain’s synapses.

“Our long-term goals are to understand how, when and why pathologic genome misfolding leads to synaptic dysfunction by way of disrupted gene expression,” said Phillips-Cremins, “as well as to engineer the genome’s structure-function relationship to reverse pathologic synaptic defects in debilitating neurological diseases.”

Originally posted in Penn Engineering Today.

BE Seminar: “Dynamics of 3D Cell Migration and Organ Formation” (Kenneth Yamada)

Our next Penn Bioengineering seminar will be held on zoom next Thursday.

Kenneth Yamada, MD, PhD

Speaker: Kenneth Yamada, M.D., Ph.D.
NIH Distinguished Investigator
Cell Biology Section
National Institute of Dental and Craniofacial Research, National Institutes of Health (NIH)

Date: Thursday, September 9, 2021
Time: 3:30-4:30 PM EDT
Zoom – check email for link or contact ksas@seas.upenn.edu
Location: Moore Room 216, 200 S. 33rd Street

Abstract: Real-time microscopy of the dynamics of cells and tissues in 3D environments is opening new windows to understanding the biophysical mechanisms of complex biological processes. Direct visualization is allowing us to explore fundamental questions in more depth that include: How do cells migrate in 3D? How do cancer cells invade? How is the extracellular matrix assembled? How are organs formed? Visualizing how cells move and organize into tissues is not only providing descriptive insights, but is also leading to the identification of novel, unexpected physical and mechanical mechanisms relevant to tissue engineering. Cells can use varying combinations of cell adhesion to adjacent cells and to the surrounding extracellular matrix with localized cellular contractility to migrate, invade, and produce the complex tissue architecture needed for organ formation.

Kenneth Yamada Bio: Kenneth Yamada has been an NIH Distinguished Investigator since 2011. He received MD and PhD degrees from Stanford. He was a Section Chief at the National Cancer Institute for 10 years and has been a Section Chief at NIDCR since 1990. He is an elected Fellow of the AAAS and American Society for Cell Biology. His research focuses on discovering novel mechanisms and regulators of cell interactions with the extracellular matrix and their roles in embryonic development and cancer. His research group focuses on the mechanisms by which three-dimensional (3D) extracellular matrix mediates key biological events, including cell migration, tissue morphogenesis, and cancer cell invasion. His research places particular emphasis on characterizing the dynamic movements of cells and their extracellular matrix as tissues are remodeled in 3D in real time. The biological systems they study include human primary cells migrating in 3D, human tumor cells and tissues, and mouse organ development. He places particularly high priority on developing future independent research leaders.

Penn Bioengineering’s Tsourkas Lab and Penn Start-up AlphaThera Awarded $667,000 SBIR Phase II Grant to Improve COVID-19 Detection Assays

To combat the COVID-19 pandemic caused by the SARS-CoV2 virus, Dr. Andrew Tsourkas’s Targeted Imaging Therapeutics and Nanomedicine (Titan) Lab in Penn Bioengineering, in collaboration with the Penn-based startup, AlphaThera, was recently awarded a $667,000 SBIR Phase II Grant Extension to support its efforts in commercializing COVID-19 detection technology. The grant supports work to address the growing need for anti-viral antibody testing. Specifically, the Tsourkas Lab and AlphaThera hope to leverage their expertise with antibody conjugation technologies to reduce the steps and complexity of existing detection assays to enable greater production and higher sensitivity tests. AlphaThera was founded in 2016 by Andrew Tsourkas, PhD, Professor of Bioengineering and James Hui, MD, PhD, a graduate of the Perelman School of Medicine and Penn Bioengineering’s doctoral program.

During this pandemic it is crucial to characterize disease prevalence among populations, understand immunity, test vaccine efficacy and monitor disease resurgence. Projections have indicated that millions of daily tests will be needed to effectively control the virus spread. One important testing method is the serological assay: These tests detect the presence of SARS-CoV2 antibodies in a person’s blood produced by the body’s immune system responding to infection. Serological tests not only diagnose active infections, but also establish prior infection in an individual, which can greatly aid in forecasting disease spread and contact tracing. To perform the serological assays for antibody detection, well-established immunoassay methods are used such as ELISA.

A variety of issues have slowed the distribution of these serological assays for antibody testing. The surge in demand for testing has caused shortages in materials and reagents that are crucial for the assays. Furthermore, complexity in some of the assay formats can slow both production and affect the sensitivity of test results. Recognizing these problems, AlphaThera is leveraging its novel conjugation technology to greatly improve upon traditional assay formats.

With AlphaThera’s conjugation technology, the orientation of antibodies can be precisely controlled so that they are aligned and uniformly immobilized on assay detection plates. This is crucial as traditional serological assays often bind antibodies to plates in a non-uniform manner, which increases variability of results and reduces sensitivity. See Fig 1 below. With AlphaThera’s uniform antibody immobilization, assay specificity could increase by as much as 1000- fold for detection of a patient’s SaRS-CoV2 antibodies.

Fig 1: Uniform vs Non-Uniform Immobilized Antibodies on Surface: Top is AlphaThera improvement, showing how antibodies would be uniformly immobilized and oriented on a plate for detection. Bottom is how many traditional serological assays immobilize antibodies, resulting in variability of results and lower specificity.

Furthermore, AlphaThera is addressing the shortage of assay reagents, specifically secondary antibody reagents, by removing certain steps from traditional serological assays. Rather than relying on secondary antibodies for detection of the patient antibodies, AlphaThera’s technology can label the patient SaRS-CoV2 primary antibodies directly in serum with a detection reagent. This eliminates several processing steps, reducing the time of the assay by as much as 50%, as well as the costs.

The Tsourkas Lab and AlphaThera have initiated their COVID-19 project, expanding into the Pennovation Center and onboarding new lab staff. Other antibody labeling products have also become available and are currently being prepared for commercialization. Check out the AlphaThera website to learn more about their technology at https://www.alphathera.com.

NIH SBIR Phase II Grant Extension— 5-R44-EB023750-03 (PI: Yu)  — 10/07/2020 – 10/07/2021