Pictured above: Michigan State University researchers Ripla Arora, PhD, (foreground) and Kaylie Chiles review images from a 3D model they helped develop. The model can reveal structural lung changes due to genetic mutations. Photo credit: Dr. Jonathan Hardy.

The 3D images from a recently published study are striking: Compared to healthy lungs, the lungs of adult mice with mutated TBX4 genes appear overwhelmed with soft muscle tissue. TBX4 normally controls the embryonic development of the umbilical vessels, lungs and lower limbs (a closely related gene, TBX5, controls the heart and upper limbs).

When a TBX4 mutation occurs spontaneously or is passed down from a parent, this development can go awry, leading to TBX4 syndrome, which can cause lower limb skeletal disorders, underdeveloped or diseased lungs, and pulmonary arterial hypertension.

The development of a 3D model was critical to understanding this change, according to Ripla Arora, associate professor in the Michigan State University College of Human Medicine’s obstetrics, gynecology and reproductive biology department.

The model reveals complex structural changes that aren’t visible on standard two-dimensional tissue samples.

Excess smooth muscle tissue

Normally, smooth muscle tissue controls countless involuntary muscular movements such as breathing, uterine contractions and blood flow. The development of PH seems largely related to how the mutation causes excessive development of smooth muscle tissue.

When her research team genetically altered mice to have TBX4 mutations and looked at samples in 3D, “smooth muscle tissue was present in a lot of places it should not be,” Arora said.

“It’s not just more smooth muscle around the blood vessels, which is a feature of pediatric PH. It’s also more muscle wrapping the bronchial tree, the interstitial spaces of the lung and even around the lungs.”

Possible treatment pathway

Importantly, the researchers didn’t see the same problem in 3D models of fetal mice with altered TBX4 genes. Meaning, the overgrowth of smooth muscle tissue is progressive – opening up a possible treatment window.

“Once the babies are born and you identify you have a problem, can you stop too much muscle formation once you diagnose it?” Arora said. “That is the therapeutic problem that can now be solved, now that we know there is a window of time to solve it.”

Arora sits on the scientific advisory committee for the nonprofit TBX4Life. The organization was founded in 2020 by Anton Morkin, whose toddler developed PH at 16 months old and was later found to have TBX4 syndrome.

A once-mysterious disorder

Prior to widespread genetic sequencing, parents in years past were often left in the dark about their child’s seemingly disparate developmental problems, such as small or missing kneecaps and PH.

But researchers now think the mutation is the second most common heritable cause of pediatric PH.

In its most severe form, TBX4 causes death in just the first few days of life. This fatal version also is currently being investigated, Arora said.

PHA grant arrived just in time

To develop the 3D model, Arora worked with fellow MSU researchers and partners at Stanford University School of Medicine. Along with funding from PHA, the teams received support from MSU, Stanford, the American Thoracic Society and NIH.

PHA’s grant arrived at a critical time, Arora said. Worried she wouldn’t be able to build out the model due to budget cuts at the federal level, she “breathed a sigh of relief” when the grant came through.

“We were in a sea of challenges last year. Thanks to PHA, I was able to push the project forward,” she said. “This is a very important research question to solve. And I think this model can help us solve that.”