Wearable Drug Delivery Device Helps Frogs Regenerate Their Hindlimbs

A team of scientists from Tufts University and the University of Florida has found that delivering progesterone to an amputation injury site can induce the regeneration of limbs in otherwise non-regenerative adults of the African clawed frog (Xenopus laevis). The team created a wearable bioreactor attached to the wound site to deliver the progesterone locally for a 24-hr period and observed that it had a lasting beneficial effect on tissue regrowth, allowing the frogs to partially regenerate their hindlimbs.

An African clawed frog (Xenopus laevis) swimming in a tank. Image credit: Celia Herrera-Rincon / Tufts University.

Many animals are capable of regeneration — in fact, planarian worms and sea cucumbers can spawn entire individuals from fragments when cut into pieces.

Partial regeneration is observed in other species — lizards regrow tails, some crabs regrow claws, and deer regrow antlers each year.

African clawed frogs can regenerate limbs when in their tadpole and froglet stages, but gradually lose that capability as they develop into adults. Until now, it was not known whether adult frogs were capable of significant regeneration response.

“At best, adult frogs normally grow back only a featureless, thin, cartilaginous spike,” said study senior author Professor Michael Levin, a researcher at the Allen Discovery Center at Tufts University.

“Our procedure induced a regenerative response they normally never have, which resulted in bigger, more structured appendages. The bioreactor device triggered very complex downstream outcomes that bioengineers cannot yet micromanage directly.”

Professor Levin and co-authors 3D printed the bioreactor out of silicon and filled it with hydrogel.

They laced the hydrogel with hydrating silk proteins that promote healing and regeneration, then added progesterone, which is best known for its role in preparing the uterus for pregnancy, but it has also been shown to promote nerve, blood vessel, and bone tissue repair.

Adult African clawed frogs (Xenopus laevis) are capable of induced increased regenerative response. Image credit: Herrera-Rincon et al, doi: 10.1016/j.celrep.2018.10.010.

“We looked at progesterone because it showed promise for promoting nerve repair and regeneration. It also modulates the immune response to promote healing, and triggers the re-growth of blood vessels and bone,” said study first author Dr. Celia Herrera-Rincon, a postdoctoral researcher at Tufts University.

“Progesterone can also regulate the bioelectric state of cells, caused by cells passing ions across their outer membranes, which is known to drive regeneration and body pattern formation.”

The researchers split the frogs into three groups: experimental, control, and sham. For the experimental and sham group, they sutured the device on the frogs immediately after limb amputation.

In the experimental group, the bioreactor released progesterone onto the amputation site. In all cases, they removed the devices after 24 hours.

When they looked at the experimental group frogs at different time points over 9.5 months, they noticed that the bioreactor seemed to trigger a degree of limb regeneration not observed in the other groups.

Instead of a typical spike-like structure, the bioreactor treatment resulted in a paddle-like formation closer to a fully formed limb than unaided regeneration could create.

“The bioreactor device created a supportive environment for the wound where the tissue could grow as it did during embryogenesis,” Professor Levin said.

“A very brief application of bioreactor and its payload triggered months of tissue growth and patterning.”

The study authors then took a closer look at the regenerated structures using molecular and histology analyses.

They saw that, unlike in the control and sham groups, the regenerating limbs of the bioreactor-treated frogs were thicker with more developed bones, innervation, and vascularization.

Analyzing video footage of the frogs in their tanks, they also noticed that the frogs could swim more like unamputated frogs.

The research is published in the journal Cell Reports.

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Celia Herrera-Rincon et al. 2018. Brief Local Application of Progesterone via a Wearable Bioreactor Induces Long-Term Regenerative Response in Adult Xenopus Hindlimb. Cell Reports 25 (6): 1593-1609; doi: 10.1016/j.celrep.2018.10.010