Common shampoo ingredient stunts developing neurons of rats

An antimicrobial agent found in many shampoos and hand lotions and widely used in industrial settings inhibits the
development of particular neuron structures that are essential for transmitting signals between cells, according to a
University of Pittsburgh study presented today at Cell Biology 2004, the 44th annual meeting of the American Society for Cell
Biology. The meeting is being held Dec. 4 - 8 at the Washington Convention Center.


Prolonged exposure to low levels of methylisothiazolinone (MIT) restricted growth of axons and dendrites of immature rat
nerve cells in culture, apparently by disengaging the machinery of a key enzyme that is activated in response to cell-to-cell
contact, and may have potentially damaging consequences to a developing nervous system, the researchers report.


"While more research is needed to determine what effect MIT would have in rodent models, both at the cellular level and to a
developing nervous system, our results thus far suggest there is potential that everyday exposure to the chemical could also
be harmful to humans. I would be particularly concerned about occupational exposure in pregnant women and the possibility of
risk to the fetus," said senior author Elias Aizenman, Ph.D., professor of neurobiology at the University of Pittsburgh
School of Medicine.


Dr. Aizenman became interested in MIT as an offshoot to his primary area of research on the mechanisms of neuronal cell
death. The first he heard of the chemical was when its name came up in a literature search for compounds with specific
chemical properties that he thought would incite a particular cell death pathway he recently had identified. As it turned
out, MIT activated a different, novel pathway, but Dr. Aizenman remained intrigued, in large part because of the considerable
lack of scientific data about a compound that he came to realize was listed on numerous consumer product labels and was very
widely used in industry.


As an antimicrobial agent, or biocide, MIT and related compounds kill harmful bacteria that like to grow near moisture or
water. As such, they often are found in personal care products, as well as in water-cooling systems and at factories that
require water for manufacturing. Since learning about MIT, Dr. Aizenman has not found any published neurotoxicity reports, or
concrete data in any public documents filed with the Environmental Protection Agency.


The first set of studies he and his team published in 2002 in The Journal of Neuroscience involved acute exposure to mature
rat neurons. They reported that 10-minute exposure at a high concentration - roughly 100 times the dose used in their current
study - was lethal to these cells.


To understand what effect chronic exposure would have on immature, developing neurons, the researchers kept cells in a media
solution containing low concentrations of MIT for 18 hours. In a standard culture, an immature neuron will in such time
develop an axon, the extension from the cell body used for sending signals to other cells, and several dendrites, elaborate
projections that receive incoming information. But after exposure to MIT, the cells had few, if any, axons and dendrites,
with the inhibition of their growth being dose-dependent, reported Kai He, Ph.D., a postdoctoral fellow working with Dr.
Aizenman who presented the data at Cell Biology 2004.















Additional studies revealed that MIT significantly hindered tyrosine phosphorylation, a process that initiates molecular
events during cell-to-cell contact, and that a particular protein enzyme was its target. This enzyme, focal adhesion kinase
(FAK), is known to be important for outgrowth of axons and dendrites as well as necessary for cell signaling. But to kick
into action, FAK must undergo tyrosine phosphorylation, whereby collections of molecules called phosphate groups are added to
FAK's sequence of amino acids. Like all proteins, FAK is comprised of a unique sequence of some 20 amino acids, including
tyrosine, so any change, such as through phosphorylation, essentially changes its function. To pinpoint the exact site along
FAK's sequence where tyrosine phosphorylation was being inhibited, the researchers had to determine which of its tyrosines
were targeted by MIT. Digging further, they found that one tyrosine in particular (amino acid 576 in the protein sequence)
was more substantially affected by MIT, an indication that this particular amino acid residue is important for axon and
dendrite growth and development.


"Since we know FAK to be highly expressed in the nerve tissue during brain development, this study suggests that
phosphorylation of tyrosine 576 may be critical for axon and dendrite outgrowth," Dr. He reported.


The authors, who also included Carl F. Lagenaur, Ph.D., associate professor of neurobiology at the University of Pittsburgh
School of Medicine, plan additional research to further understand the molecular mechanisms underlying MIT's neurotoxic
effect on cells, as well as studies involving whole animals. They are hopeful that their work will stimulate additional
research by other groups as well as bring heightened awareness about the potential risks from human exposure.


"This chemical is being used more and more extensively, yet there have been no neurotoxicity studies in humans to indicate
what kind and at what level exposure is safe. I realize it's a big leap to suggest there may be a parallel between
environmental exposure and the noticeably higher rates of diagnosed childhood developmental disabilities, but I would caution
that based on our data, there very well could be neurodevelopmental consequences from MIT. Clearly, more study is needed,
with both scientists and government regulators equally engaged," added Dr. Aizenman.


NOTE TO EDITORS: This abstract, Lack of Phosphorylation of Tyrosine 576 of Focal Adhesion Kinase Correlates to Neurite
Outgrowth Deficiency Following Methylisothiazolinone Treatment in Cultured Cortical Neurons (Poster Presentation 300), is
being presented at 1:30 p.m., Sunday, Dec. 5 in Session 123 Focal Adhesions. To arrange interviews with the authors, please
call Lisa Rossi at 412-916-3315 (cell). The press contact for the American Society for Cell Biology (ASCB) is John
Fleischman, who can be reached at jfleischmanacsb or at 202-249-4019 between Dec. 4 - 8. The ASCB Newsroom is located in
Room 101 of the Washington Convention Center.


Contact: Lisa Rossi

RossiLupmc

(Cell - 412-916-3315)

Phone: 412-647-3555

Fax: 412-624-3184


Craig Dunhoff

DunhoffCCupmc

Phone: 412-647-3555

Fax: 412-624-3184

University of Pittsburgh Medical Center