Sections of Retinas Regenerated and Visual Function Increased With Stem Cells from Skin

Sections of Retinas Regenerated and Visual Function Increased With Stem Cells from Skin

  1:02:00 PM    Science Lover

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Scientists from Schepens Eye Research Institute are the first to regenerate large areas of damaged retinas and improve visual function using IPS cells (induced pluripotent stem cells) derived from skin. The results of their study, which is published in PLoS ONE this month, hold great promise for future treatments and cures for diseases such as age-related macular degeneration, retinitis pigmentosa, diabetic retinopathy and other retinal diseases that affect millions worldwide.

Histological staining of a teratoma containing Rho-/- eye
at 21 days post-injection of a heterogeneous population of
SSEA1-containing D33 differentiated cells. (Credit: Tucker
et al.,DOI: 10.1371/journal.pone.0018992)

"We are very excited about these results," says Dr. Budd A. Tucker, the study's first author. "While other researchers have been successful in converting skin cells into induced pluripotent stem cells (iPSCs) and subsequently into retinal neurons, we believe that this is the first time that this degree of retinal reconstruction and restoration of visual function has been detected," he adds. Tucker, who is currently an Assistant Professor of Ophthalmology at the University of Iowa, Carver College of Medicine, completed the study at Schepens Eye Research Institute in collaboration with Dr. Michael J. Young, the principle investigator of the study, who heads the Institute's regenerative medicine center.

Today, diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD) are the leading causes of incurable blindness in the western world. In these diseases, retinal cells, also known as photoreceptors, begin to die and with them the eye's ability to capture light and transmit this information to the brain. Once destroyed, retinal cells, like other cells of the central nervous system have limited capacity for endogenous regeneration.

"Stem cell regeneration of this precious tissue is our best hope for treating and someday curing these disorders," says Young, who has been at the forefront of vision stem cell research for more than a decade.

While Tucker, Young and other scientists were beginning to tap the potential of embryonic and adult stem cells early in the decade, the discovery that skin cells could be transformed into "pluripotent" cells, nearly identical to embryonic cells, stirred excitement in the vision research community. Since 2006 when researchers in Japan first used a set of four "transcription factors" to signal skin cells to become iPSCs, vision scientists have been exploring ways to use this new technology. Like embryonic stem cells, iPSCs have ¬the ability to become any other cell in the body, but are not fraught with the ethical, emotional and political issues associated with the use of tissue from human embryos.

Tucker and Young harvested skin cells from the tails of red fluorescent mice. They used red mice, because the red tissue would be easy to track when transplanted in the eyes of non-fluorescent diseased mice.

By forcing these cells to express the four Yamanaka transcription factors (named for their discoverer) the group generated red fluorescent IPSCs, and, with additional chemical coaxing, precursors of retinal cells. Precursor cells are immature photoreceptors that only mature in their natural habitat -- the eye.

Within 33 days the cells were ready to be transplanted and were introduced into the eyes of a mouse model of retina degenerative disease. Due to a genetic mutation, the retinas of these recipient mice quickly degenerate, the photoreceptor cells die and at the time of transplant electrical activity, as detected by ERG (electroretinography), is absent.

Within four to six weeks, the researchers observed that the transplanted "red" cells had taken up residence in the appropriate retinal area (photoreceptor layer) of the eye and had begun to integrate and assemble into healthily looking retinal tissue.

The team then retested the mice with ERG and found a significant increase in electrical activity in the newly reconstructed retinal tissue. In fact, the amount of electrical activity was approximately half of what would be expected in a normal retina. They also conducted a dark adaption test to see if connections were being made between the new photoreceptor cells and the rest of the retina. In brief, the group found that by stimulating the newly integrated photoreceptor cells with light they could detect a signal in the downstream neurons, which was absent in the other untreated eye.

Based on the results of their study, Tucker and Young believe that harvesting skin cells for use in retinal regeneration is and will continue to be a promising resource for the future.

The two scientists say their next step will be to take this technology into large animal models of retinal degenerative disease and eventually toward human clinical trials.

Other scientists involved in the PLoS ONE study include In-Hyun Park, Sara D. Qi, Henry J. Klassen, Caihui Jiang, Jing Yao, Stephen Redenti, and George Q. Daley.

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Why the Eye Is Better Than a Camera at Capturing Contrast and Faint Detail Simultaneously

Why the Eye Is Better Than a Camera at Capturing Contrast and Faint Detail Simultaneously

  8:52:00 AM    Science Lover

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The human eye long ago solved a problem common to both digital and film cameras: how to get good contrast in an image while also capturing faint detail.

Cones normally release the neurotransmitter glutamate 
in the dark, while light decreases glutamate release. This 
graph of neurotransmitter release shows what 
happens when cone cells are exposed to a dark spot in a 
light background (top) under various scenarios, including 
no feedback (green trace) and only negative feedback 
from horizontal cells (red trace). Negative feedback to 
many cones enhances edges, but would decrease 
detail in dark areas were it not for newly discovered 
positive feedback that is localized to only a few cone
cells (blue trace). (Credit: Richard Kramer lab, UC Berkeley)

Nearly 50 years ago, physiologists described the retina's tricks for improving contrast and sharpening edges, but new experiments by University of California, Berkeley, neurobiologists show how the eye achieves this without sacrificing shadow detail.

"One of the big success stories, and the first example of information processing by the nervous system, was the discovery that the nerve cells in the eye inhibit their neighbors, which allows the eye to accentuate edges," said Richard Kramer, UC Berkeley professor of molecular and cell biology. "This is great if you only care about edges. But we also want to know about the insides of objects, especially in dim light."

Kramer and former graduate student Skyler L. Jackman, now a post-doctoral fellow at Harvard University, discovered that while light-sensitive nerve cells in the retina inhibit dozens of their close neighbors, they also boost the response of the nearest one or two nerve cells.

That extra boost preserves the information in individual light detecting cells -- the rods and cones -- thereby retaining faint detail while accentuating edges, Kramer said. The rods and cones thus get both positive and negative feedback from their neighbors.

"By locally offsetting negative feedback, positive feedback boosts the photoreceptor signal while preserving contrast enhancement," he said.

Jackman, Kramer and their colleagues at the University of Nebraska Medical Center in Omaha report their findings May 3 in the journal PLoS Biology. Kramer also will report the findings at the 2011 annual meeting of the Association for Research in Vision and Ophthalmology in Ft. Lauderdale, Fla.

From horseshoe crabs to humans

The fact that retinal cells inhibit their neighbors, an activity known as "lateral inhibition," was first observed in horseshoe crabs by physiologist H. Keffer Hartline. That discovery earned him a share of the 1967 Nobel Prize in Physiology or Medicine. This form of negative feedback was later shown to take place in the vertebrate eye, including the human eye, and has since been found in many sensory systems as a way, for example, to sharpen the discrimination of pitch or touch.

Lateral inhibition fails, however, to account for the eye's ability to detect faint detail near edges, including the fact that we can see small, faint spots that ought to be invisible if their detection is inhibited by encircling retinal cells.

Kramer noted that the details of lateral inhibition are still a mystery half a century after Hartline's discovery. Neurobiologists still debate whether the negative feedback involves an electrical signal, a chemical neurotransmitter, or protons that change the acidity around the cell.

"The field is at an impasse," Kramer said. "And we were surprised to find this fundamental new phenomenon, despite the fact that the anatomy of the retina has been known for more than 40 years."

The retina in vertebrates is lined with a sheet of photoreceptor cells: the cones for day vision and the rods for night vision. The lens of the eye focuses images onto this sheet, and like the pixels in a digital camera, each photoreceptor generates an electrical response proportional to the intensity of the light falling on it. The signal releases a chemical neurotransmitter (glutamate) that affects neurons downstream, ultimately reaching the brain.

Unlike the pixels of a digital camera, however, photoreceptors affect the photoreceptors around them through so-called horizontal cells, which underlie and touch as many as 100 individual photoreceptors. The horizontal cells integrate signals from all these photoreceptors and provide broad inhibitory feedback. This feedback is thought to underlie lateral inhibition, a process that sharpens our perception of contrast and color, Kramer said.

The new study shows that the horizontal cells also send positive feedback to the photoreceptors that have detected light, and perhaps to one or two neighboring photoreceptors.

"Positive feedback is local, whereas negative feedback extends laterally, enhancing contrast between center and surround," Kramer said.

Electrical vs. chemical signals

The two types of feedback work by different mechanisms, the researchers found. The horizontal cells undergo an electrical change when they receive neurotransmitter signals from the photoreceptors, and this voltage change quickly propagates throughout the cell, affecting dozens of nearby photoreceptors to lower their release of the glutamate neurotransmitter.

The positive feedback, however, involves chemical signaling. When a horizontal cell receives glutamate from a photoreceptor, calcium ions flow into the horizontal cell. These ions trigger the horizontal cell to "talk back" to the photoreceptor, Kramer said. Because calcium doesn't spread very far within the horizontal cell, the positive feedback signal stays local, affecting only one or two nearby photoreceptors.

The discovery of a new and unsuspected feedback mechanism in a very well-studied organ is probably related to how the eye is studied, Kramer said. Electrodes are typically stuck into the retina to both change the voltage in cells and record changes in voltage. Because the new signal is chemical, not electrical, it would have been easily missed.

Jackman and Kramer found the same positive feedback in the cones of a zebrafish, lizard, salamander, anole (whose retina contains only cones) and rabbit, proving that "this is not just some weird thing that happens in lizards; it seems to be true across all vertebrates and presumably humans," Kramer said.

The research was supported by the National Institutes of Health and the organization Research to Prevent Blindness.

Coauthors with Kramer and Jackman are Norbert Babai and Wallace B. Thoreson of the Department of Ophthalmology at the University of Nebraska Medical Center and James J. Chambers of the Department of Chemistry at the University of Massachusetts, Amherst.

Source:Science Daily

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Blind Mice Can 'See' Thanks to Special Retinal Cells

Blind Mice Can 'See' Thanks to Special Retinal Cells

  9:36:00 AM    Science Lover

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It would make the perfect question for the popular television show "Are You Smarter than a 5th Grader:" What parts of the eye allow us to see?

Mice without rods and cones function can still see -- 
and not just light, but also patterns and images -- 
thanks to a third kind of photosensitive cell in the 
retina, according to new research. 
(Credit: iStockphoto/Steven Maltby)

The conventional wisdom: rods and cones. The human retina contains about 120 million rods, which detect light and darkness, shape and movement, and about 7 million cones, which in addition detect color. Without them, or so we are taught, our eyesight simply would not exist.

But that might not be true, according to a study -- published July 15 in the journal Neuron -- that provides new hope to people who have severe vision impairments or who are blind.

A team led by biologist Samer Hattar of The Johns Hopkins University's Krieger School of Arts and Sciences found that mice that didn't have any rods and cones function could still see -- and not just light, but also patterns and images -- courtesy of special photosensitive cells in the rodents' retinas. Until now, it was presumed that those cells, called intrinsically photosensitive Retinal Ganglion Cells, (or ipRGCs), didn't play a role in image formation, but instead served other functions, such as dictating when the animals went to sleep or woke up. (All mammals, including humans, have ipRGCs, as well as rods and cones.)

"Up until now, it was assumed that rods and cones were the only cells capable of detecting light to allow us to form images," said Hattar, who as an assistant professor in the Department of Biology, studies mammals' sleep-wake cycles, also called "circadian rhythms." "But our study shows that even mice which were blind could form low-acuity yet measurable images, using ipRGCs. The exciting thing is that, in theory at least, this means that a blind person could be trained to use his or her ipRGCs to perform simple tasks that require low visual acuity."

"Visual acuity" refers to the sharpness or clarity of a person's (or animal's) vision. Someone with so-called "20/20 vision" can see clearly at a distance of 20 feet what the "average" human being can see at that distance. In contrast, a person with "20/100" vision would have to stand 20 feet away from, for instance, an eye chart that the average person could read from 100 feet away. People with very low visual acuity (worse than "20/100" with corrective lenses) are considered "legally blind."

In addition to providing hope for people with serious vision problems, Hattar's findings hint that, in the past, mammals may have used their ipRGCs for sight/image formation, but during the course of evolution, that function was somehow taken over by rods and cones.

The study also concludes that, far from being homogenous, ipRGCs come in five different subtypes, with the possibility that each may have different light-detecting physiological functions.

To conduct the study, the team used a special system to genetically label cells and then "trace" them to the rodents' brains before subjecting the mice to a number of vision tests. In one, mice followed the movements of a rotating drum, a test that assessed the animals' ability to track moving objects. In another, the rodents were placed within a "Y"-shaped maze and challenged to escape by selecting the lever that would let them out. That lever was associated with a certain visual pattern. The mice that were blind -- they lacked rods, cones and ipRGCs -- couldn't find that lever. But those with only ipRGCs could.

"These studies are extremely exciting to me, because they show that even a simple light-detecting system like ipRGCs has incredible diversity and may support low-acuity vision, allowing us to peer into evolution to understand how simple vision may have originally evolved before the introduction of the fancy photoreceptors rods and cones," Hattar said.

Hattar's team worked on this study in collaboration with groups led by David Berson of Brown University and Glen Prusky of Weill Cornell Medical College. It was supported by grants from the National Institutes of Health, the David and Lucile Packard Foundation and the Alfred P. Sloan Foundation.

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