With the insertion of a single gene, blind mice recover their sight

It was surprisingly simple. Scientists at the University of California at Berkeley inserted a gene for a green light receptor into the eyes of blind mice, and a month later, they jumped through obstacles as easily as mice with no eye problems. vision. They were able to see the movement, the brightness changes over a range of a thousand times and the fine details on an iPad sufficient to distinguish the letters.

Researchers say that in just three years, gene therapy – delivered via an inactivated virus – could be tried in humans who have lost their eyesight because of retinal degeneration, ideally providing them with enough vision to move about. eventually allowing them to regain their capacity. read or watch videos.

"You would inject this virus into the eye of a person and a few months later she would see something," said Ehud Isacoff, professor of molecular and cellular biology at the University of Berkeley and director from the Helen Wills Neuroscience Research Institute. "With neurodegenerative diseases of the retina, we often try to stop or slow down the degeneration, but something that restores an image in a few months is an amazing thing to think about."

About 170 million people worldwide suffer from age-related macular degeneration, which strikes one in 10 people over the age of 55, while 1.7 million people in the world suffer from the most common form of hereditary blindness, retinitis pigmentosa, which usually blinds people over 15 years of age. of 40.

"I have friends who do not perceive the light and their lifestyle is heartbreaking," said John Flannery, professor of molecular and cellular biology at the University of Berkeley, a faculty member at the University of California. 39 School of Optometry. "They have to consider what the seers take for granted – for example, every time they go to a hotel, the room layout is a little different, and they need someone to guide them around." the room while building a 3D map Everyday objects, such as a coffee table, can pose a risk of falling The burden of disease is enormous for people with severe and disabling vision loss. could be the first candidates for this type of therapy. "

Currently, the options for these patients are limited to an electronic eye implant connected to a video camera based on a pair of glbades – a delicate, invasive and expensive configuration that produces on the retina an image equivalent to a few hundred times. pixels. Normal, clear vision involves millions of pixels.

The correction of the genetic defect responsible for retinal degeneration is not simple either, since there are more than 250 different genetic mutations responsible for retinitis pigmentosa alone. About 90% of these cells kill photoreceptor cells in the retina – penis-sensitive rods and cones for color perception in daylight. But retinal degeneration usually spares other layers of retinal cells, including bipolar and retinal ganglion cells, which can remain healthy even though they are insensitive to light for decades after people have become completely blind.

In their mouse trials, the UC Berkeley team was able to light-sensitive 90% of ganglion cells.

Isacoff, Flannery and colleagues at UC Berkeley will report on their success in an online article on March 15 in Nature Communications.

& # 39; You could have done this 20 years ago & # 39;

To reverse blindness in these mice, the researchers devised a virus targeting the retinal ganglion cells and loaded it with the corresponding gene for a photosensitive receptor, the green cone opsin (lenght). medium wave). Normally, this opsine is only expressed by conical photoreceptor cells and makes them sensitive to green-yellow light. When it was injected into the eye, the virus transported the gene into ganglion cells, which are normally insensitive to light, and made them sensitive to light and able to send light. signals to the brain interpreted as being on demand.

"As long as we can not test the mice, it's impossible to distinguish the behavior of optogenetically treated mice from normal mice without special equipment," Flannery said. "It remains to be seen what it means in a patient."

In mice, researchers were able to administer opsins to most retinal ganglion cells. To treat humans, they would need to inject a lot more virus particles because the human eye contains thousands of times more ganglion cells than the mouse eye. However, the UC Berkeley team has developed ways to improve viral delivery and hopes to insert the new light sensor into a similarly high percentage of ganglion cells, a quantity equivalent to the very high number of 'a camera.

Isacoff and Flannery have found the simple solution after more than a decade of testing more complex regimens, including the insertion into recombinant retinal cells of combinations of genetically engineered neurotransmitter receptors and light-sensitive chemical switches. These worked, but did not reach the sensitivity of normal vision. Opsins from microbes tested elsewhere also had lower sensitivity, requiring the use of light-amplified glbades.

To capture the high sensitivity of natural vision, Isacoff and Flannery turned to photoreceptor light receptor opsins. Using an adeno-badociated virus (AAV) that naturally infects ganglion cells, Flannery and Isacoff successfully delivered the retinal opsin gene into the ganglion cell genome. The previously blind mice acquired a vision that lasted all their lives.

"The fact that this system works is really very satisfying, partly because it's also very simple," Isacoff said. "Ironically, you could have done that 20 years ago."

Isacoff and Flannery are raising funds for gene therapy to be tested in humans three years from now. The FDA has approved similar AVA delivery systems for eye diseases in people with degenerative retinal disorders for whom there is no medical alternative.

It can not work

According to Flannery and Isacoff, most people working in the field of vision would wonder if opsines could act outside their photoreceptor cells specialized in cones and rods. The surface of a photoreceptor is decorated with opsines – rhodopsin rods and opsins red, green and blue cones – embedded in a complex molecular machine. A molecular relay – the G-protein coupled receptor signaling cascade – amplifies the signal so efficiently that we are able to detect individual photons of light. An enzyme system reloads the opsin once it has detected the photon and that it is "bleached". Feedback control adapts the system to very different background brightness levels. And a specialized ion channel generates a powerful voltage signal. Without transplanting all this system, it was reasonable to think that opsin would not work.

But Isacoff, a specialist in G protein-coupled receptors in the nervous system, knew that many of these parts exist in all cells. He suspected that an opsin would automatically connect to the retinal ganglion cell signaling system. Together, he and Flannery first tried rhodopsin, which is more sensitive to light than cones opsins.

To their great pleasure, when rhodopsin was introduced into the ganglion cells of mice whose rods and cones had completely degenerated, and which were therefore blind, the animals regained the ability to distinguish the dark from the light even the low ambient light. But rhodopsin proved too slow and failed in the recognition of images and objects.

They then tried the green cone opsin, which responded 10 times faster than rhodopsin. Remarkably, the mice were able to distinguish parallel lines from horizontal lines, lines closely spaced from widely spaced lines (standard human acuity task), moving lines relative to stationary lines. The restored vision was so sensitive that iPads could be used for visual displays instead of much brighter LEDs.

"This has powerfully brought the message home," said Isacoff. "After all, it would be wonderful if blind people regained the ability to read a standard computer screen, communicate via video, watch a movie."

These successes prompted Isacoff and Flannery to go further and discover if animals could navigate the world with a restored vision. Strikingly, here too, the green cone's opinion was a success. Mice that had been blind have regained their ability to perform one of their most natural behaviors: recognizing and exploring objects in three dimensions.

They then asked, "What would happen if a person with a restored vision went outside with a brighter light? Would she be blinded by the light?" Here, another striking feature of the system has emerged, said Isacoff: The opsin signaling path of the green cone is adapting. Animals that were previously blind have adjusted to the change in brightness and could perform the task just as well as sighted animals. This adaptation worked on a beach about a thousand times – the difference, essentially, between the average indoor and outdoor lighting.

"When everyone says that it will never work and you are crazy, it usually means that you are on something," said Flannery. Indeed, this is the first successful restoration of vision modeled using an LCD, the first to adapt to changes in ambient light and the first to restore the natural vision of objects.

The UC Berkeley team is currently testing variants on the theme that can restore color vision and increase acuity and adaptation.

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University of California, Berkeley