Lathering the sunscreen for a trip to the lake might be good to avoid skin cancer and sunburn, but it might not be so good for the lake’s inhabitants.

An ISU research team is the first to look at the long-term effects of nano-titanium dioxide, a common ingredient in household items, on fish.

In the study, fathead minnows and their embryos — standards for EPA toxicology tests — as well as zebrafish were placed in tanks and exposed to different concentrations of nano-TiO2 for an extended period of time. After two days, the fish were examined and the researchers found that the nano-TiO2 caused an increase in Neutrophil Extracellular Trap release and an increase in cell death. 

"The higher the concentration of nanoparticles, the higher the oxcidity burst response, meaning the cells have been primed to respond to regular stimulus more strongly, or more potently," said Dusan Palic, assistant professor of biomedical sciences. "Which sometimes in the immune system isn't a good thing because it can cause more damage than good."

Neutrophils are white blood cells that kill bacteria, viruses and fungi. During their normal life cycle, they catch foreign cells and, in essence, eat them. However, when near the end of their life cycle, the neutrophils may respond to an infection with a NET release. The exposure to TiO2 affects the NET release in fish.

"The neutrophils when nearing the end of their life, if stimulated correctly, release their DNA in the form of a net, like Spiderman trying to catch somebody, and it is used to catch bacteria," Palic said. "It is also a sign of their increased tendency to die. The pre-exposure to TiO2 increases that NET release response in neutrophils."

When near the end of their life cycle, the neutrophils may respond to an infection with a NET release. The exposure to TiO2 affects the NET release in fish. In other words, exposure to TiO2 causes the fish's immune system to attack foreign cells, like bacteria and viruses, far more strongly than they normally would. The neutrophils release their DNA in the form of a net in an attempt to kill the foreign cells. This kills the defenders, leaving the fish open to other diseases that may be fatal.

However, individual cells that die by NETosis are not always the only ones to die. Upon a neutrophil's death, there is respiratory burst which may form superoxides, like hydrogen peroxide and dioxide, which act as a signal for other neutrophils to react the same way — similar to the way a bee sting will send a signal to other bees to attack.

The respiratory burst response in other cells does eventually decrease and eventually cease. Additionally, the effects from the fish injected with TiO2 caused fewer neutrophils to be produced in addition to causing an increase of NET release.

The TiO2 is not the direct killer of the fish. Fish exposed to TiO2 have weakened immune systems, but if there are no bacteria, fungi or viruses, the fish will remain unharmed.

Similarly, should the fish be exposed to bacteria, viruses, or fungi but not TiO2, they would generally be resistant. It is only when the two are present together that a threat is posed to the fish.

"Let's say you have a small town in Iowa that uses a number of products that have titanium dioxide in them," said Boris Jovanovic, graduate assistant in biomedical sciences. "Let's say they're building up a new subdivision and there's a lot of powder or dust coming from the material, and the workers go home and shower with a lot of stuff that has titanium dioxide like shampoos and soaps.

"Their little sewage treatment plant can't process all the increase in titanium dioxide, and some of that overflows into a nearby stream. But the stream has a field on the other side that has a facility that produces hogs and pigs," he said. "The seepage of antibiotic-resistant bacteria from the hogs and pigs enters that stream as well. If there wasn't titanium and impairment of the immune system caused by that titanium, the fish would be more resilient to the bacteria and they wouldn't be endangered by that in any way."

"I think this study raises a concern regarding that the concentrations can cause this effect were close to the environmental estimates," Jovanovic said. "Given the fact that the product of nanoparticles is increasing rapidly every year, probably in the near future we will double or triple the amount of titanium dioxide entering the environment from human sources, and we may start seeing changes in the populations of fish responses in five, 10 or 15 years."

As of now, there is no actual study of the concentrations of nanoparticles in real environments in the United States, Jovanovic said. The study did expose fish to the estimated levels of TiO2 in the environment. With the results indicating that TiO2 is severely weakening the immune systems of fish, and considering the United States currently produces 20 percent of the world's nanoparticles and is expected to increase its production, it is something that needs to be studied.

"We're not saying it's going to kill every fish out there in the next five years," Palic said. "It's just one of the pieces of the big picture that whoever started to look into health effects or potential health effects is finding reasons for concern."

TiO2 is not inherently dangerous to humans unless it gets into the lungs, or if a person is exposed to it in large amounts. Additionally, contaminated fish would not contain enough TiO2 to pose a health threat to humans.

The study was conducted by Jovanovic, Eric W. Rowe and Palic from the ISU College of Veterinary Medicine’s Department of Biomedical Science; Yanjie Zhang and Aaron R. Clapp from Vet Med's Department of Chemical and Biological Engineering; and Lora Anastasova from Trakia University’s Department of Veterinary Medicine.

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