Linking To And Excerpting From "Introduction to the Concept of Signal Toxicity"

Today, I link to and excerpt from Introduction to the Concept of Signal Toxicity. [PubMed Abstract] [Full-Text HTML] [Full-Text PDF]. J Toxicol Sci. 2016;41(Special):SP105-SP109. doi: 10.2131/jts.41.SP105.

The above article has been cited by nine articles in PubMed.

All that follows is from the above article, Introduction to the Concept of Signal Toxicity.

ABSTRACT — Silent Spring by Rachel Carson (1962) established a role for environmental chemicals in cancer and Our Stolen Future by Theo Colbone, Dianne Dumanoski and John Peterson Myers (1996) coined the concept of “Endocrine Disrupting Chemicals (EDCs)” with its mechanistic plausibility for all

the living organisms. For basic biologists, seeing a non-monotonic dose-response curve was a matter of course. In contrast, for the toxicologists at that time, the dose-response curves should be monotonic. It took some time for toxicologists to accept the plausibility that animals and humans are subject to the effects of EDCs act in a way that is explained by the new paradigm of receptor-mediated toxicity or in other words “signal toxicity.” In classical toxicology, a toxic substance reaches a cellular target and induc- es malfunction. The target molecules are proteins including enzymes, lipid membranes, DNA, and other components of the cell which are damaged by the toxic substances. On the other hand, in the case of sig-

nal toxicity, a chemical binds to a speciﬁc receptor – after that, the chemical itself is not important. The signal from the receptor initiates a cascade of molecular events that leads to various changes in the cells and organs. When the signal is abnormal for a cell or an organ in terms of quality, intensity and timing,

then the signal will induce adverse effects to the target. An extreme example of signal toxicity is the 1981 Nobel Prize in Physiology or Medicine work by Drs. Hubel and Wiesel. They blocked the signal of sharp images from the retina to the brain and found that the visual cortex needed this signal at the correct time for its proper development. In humans, such signal disruption is well known to induce “form-deprivation amblyopia” in infants. The concept of signal toxicity widens the range of systems vulnerable to EDCs and facilitates the understanding of their biological characteristics. For example, compared with intrinsic ligands, xenobiotic chemicals usually act as weak agonists and/or weak antagonists of receptor sys-

tems; the dose-response characteristics and the dose range will depend on the signaling system of concern. If the signal is used for organogenesis and functional maturation, there would be a critical period in the development during which the disturbance of such signals may cause irreversible changes. Since recepter-based signaling mechanisms are usually an ampliﬁcation systems, it is hard to set a threshold in its dose response, and the outcome of signal toxicity is often stochastic at low doses. This review attempts to explain the beneﬁts of incorporating the concept of signal toxicology for widening the range of toxicology for the better protection of human and environmental health in modern civilized life, where chemicals are designed to be less toxic in terms of traditional toxicity but not yet in “signal toxicity.”

Key words:Signal toxicity, Receptor, Non-monotonous dose response, Critical period, Irriversible effect, Omic tools

Signal toxicity

In classical toxicology, a toxic substance (chemical)

or its metabolite(s) reaches the target molecule and inter-

acts with it to induce malfunction. In order to understand

how the chemicals reach the target, Absorption, Distri-

bution, Metabolism and Excretion (ADME) were con-

sidered important. The target molecules are proteins,

including enzymes, lipid membranes, DNA, and other

components of the organs and cells. Depending on the tar-

get molecules, the nature of toxicity differs. If the target is

involved in energy production, then the effect appears as

acute toxicity. When the malfunction leads to cell degen-

eration or cell death, the outcome can be chronic.

On the other hand, in case of signal toxicity, a chem-

ical only has to bind to a receptor on the cell membrane

or inside the cell. After binding to the receptor, the chem-

ical itself does not need to move to the target site where

the key events of toxicity take place. The signal from the

receptor initiates a cascade of molecular events that leads

to various changes in the cells and organs. When the sig-

nal is abnormal for a cell or an organ in terms of intensi-

ty and timing, then the signal will induce adverse effects

to them (Fig. 1).

Nearly all receptor systems have intrinsic or natural

ligands in the body. This is the case for estrogen, andro-

gen and thyroid hormone receptors. Intrinsic ligands are

often the best binders; they show the highest afﬁ nity to

the receptor. On the other hand, many xenobiotic chem-

icals can bind to the receptors at lower afﬁ nity. In such

cases, those chemicals are assumed to act as weak ago-

nists and/or weak antagonists.

Signal transduction is normally an amplifying systemto pick up a very small change in concentration of the liand to the receptor. Therefore, essentially speaking, the dose-response characteristics of the signaling system are understood to have no threshold. The biological effect of signal disruption by a chemical that binds to a receptor can be either transient or long-lasting. Recent advance in biology provide explanations for signal-mediated long-lasting alterations in gene expression by alteration of DNA methylation and histone modiﬁcation or other epigenetic modiﬁcations.

Signal disruption can elicit different effects in mature,

versus developing organisms. This difference is an impor-

tant characteristic of signal toxicity. Although effects

on adults can be transient, signal toxicity in the embry-

os, fetuses, neonates and infants can be more long-

lasting or even permanent. During development and

maturation, various signals are used to coordinate org-

anogenesis and establishment of proper organ func-

tions. An extreme example of signal toxicity by sig-

nal disruption can be found in the 1981 Nobel Prize

in Physiology or Medicine work by Drs. Hubel and

Wiesel. They sutured one eyelid of a baby monkey, then

checked the vision in its adulthood; the vision of the

sutured eye was severely and irreversibly impaired. In

short, the visual cortex needed sharp images from the ret-

ina to develop properly; therefor the neuronal networks

required to comprehend the “vision” were not developed

and resulted in ipsilateral “cortical blindness .” In Hubel’s

Nobel Lecture, he says that “The design of these experi-

ments was undoubtedly inﬂuenced by the observation that

children with congenital cataract still have substantial and

often permanent visual deﬁcits after removal of the cat-

aract and proper refraction” (http://www.nobelprize.org/

nobel_prizes/medicine/laureates/1981/wiesel-lecture.pdf).

Among ophthalmologists, it is well known that an even

shorter period of disrupting sharp images from retina

results in visual impairment. This is called “form-depri-

vation amblyopia” or “occlusion amblyopia,” and can be

caused by more than 2 days of eyepatch use in a baby less

than 2 years of age. Likewise, various parts of the devel-

oping brain need proper signals at proper timing or criti-

cal periods for their normal development. Pronunciation

of a mother tongue and sense of absolute pitch are some

typical examples.

With respect to developmental toxicology, signals at

the molecular level are frequently used for brain devel-

opment. As an example, estrogen and its receptor(s) are

found in a particular series of neurons, and disruption of

the estrogenic signal by exogenous estrogens or antiestro-

gens during a critical period can leave permanent effects in brain structure and function. Acetylcholine is one of the

major neurotransmitters. During development, again, sig-

naling via neurotransmitters is used for proper formation

of the brain. Recepter agonists and antagonists at signal-

ing doses have been shown to leave irreversible function-

al changes on the developing brain when applied during

the critical period of the development of the target sys-

tems (Tanemura et al., 2009; Furukawa et al., 2016).

There are four major characteristic features of sig-

nal toxicity:

(1) The disruption of the signal does not necessari-

ly result in morphological damaging or killing of the tar-

get cell. Rather signal disruption just has to change the

function, for example altering gene expression and pro-

tein synthesis.

(2) The signaling mechanisms in the body, especially

those mediated by receptors, are usually an ampliﬁcation

system. Therefore, it is difﬁcult to set a threshold in its

dose-response characteristics. When the system is already

activated by the intrinsic ligands, the additional agonistic

or antagonistic effects elicited by signal disrupting chemi-

cals directly affects the system.

(3) The outcome of signal toxicity is often stochas-

tic in dose response. The symptoms of high-dimension-

al function, such as impairment in memory and cognitive

function, are the result of multiple stages in development

and maturation. The disturbance of signals in those stages

at lower doses would not induce weak changes in all sub-

jects; rather it would result in a low frequency of affect-

ed subjects among those exposed. Similar stochastic dose

responses can be seen in carcinogenesis studies or in radi-

ation studies (both of which are multi-stage events) where

the frequency or probability in numbers of cancerous ani-

mals increases with dose. If the gene expression proﬁles

are altered irreversibly by the signal toxicity, it is pre-

sumed that the underlying molecular mechanism(s) are

epigenetic alterations in the genome. There are examples

of stochastic effects monitored as early onset of persistent

estrus cyclicity in female offspring exposed to estrogen in

their prenatal periods (Shirota et al. 2012; Kanno et al. in

preparation).

(4) The dose range of the signal toxicity depends on the

sensitivity of the targeted receptor system. For example,

the natural ligands for the estrogen receptor system work

at low Pico molar range (~10-11M). Although bisphenol A

is 10,000 to 100,000 times less potent than 17-beta estra-

diol in transient transactivation assays (Ohta et al., 2012;

Grimaldi et al., 2015); its signaling dose is in the range of

20 to 200 micrograms/L. If orally administered bisphenol

A is absorbed 100% and distributed evenly throughout the

body ﬂuid, 20 to 200 micrograms/kg is the signaling dosefor bisphenol A. This amount of bisphenol A is much less

than the NOAEL (5 mg/kg) given by traditional toxicity

studies. On the other hand, the androgen receptor system

works at an about 1,000 times higher range of concentra-

tion than does the estrogen receptor system (Wilson et al.,

2002). Therefore, androgenic chemicals 10,000 times less

potent than intrinsic androgens are calculated to work at

the range of 20 to 200 milligrams/kg.

Endocrine Disrupting Chemicals (EDCs)

The “low-dose effect” (Melnick et al., 2002) is under-

stood as an effect of a chemical observed in a dose range

lower that the NOEL or NOAEL (no observed (adverse)

effect level) given by toxicological studies conducted in

compliance with established test guidelines such as the

OECD Test Guidelines. Toxicologists had been divid-

ed in opinion as to whether or not the low-dose effect is

real. The identiﬁcation of toxicity in classical studies is

more or less dependent on histopathological diagnosis as

the deterministic diagnosis. Such diagnosis is made based

on changes that are usually a reflection of cytotoxicity

at the levels of intracellular organelle. Such cytotoxici-

ty results in atrophy, degeneration, apoptosis or necrosis,

and some of those changes can be directed to cell pro-

liferation (hypertrophy, hyperplasia), and neoplasia. Inter-

stitial changes such as ﬁbrosis and other kinds of deposi-

tions are also triggered.

In contrast to the overt morphological changes found

in classical toxicity studies, the low-dose effects of EDCs,

in general, are of developmental and functional in nature.

The concept of signal toxicity can explain the effects of

EDCs across a broader range. The discussion of EDCs

began with observation of estrogenic (anti-estrogenic)

and androgenic (anti-androgenic) compounds targeting

the reproductive systems. Mechanistically speaking, such

chemicals act by binding, to some extent, to estrogen

receptors or androgen receptors and aberrant signals from

these receptors initiate the events that lead to adversity.

In adults, these signaling systems have established

their own homeostatic regulation. Therefore, disturbances

by xenobiotic chemicals are well cancelled. On the other

hand, embryos and neonates have premature homeostasis.

Moreover, the receptor signals are used for the develop-

ment and maturation of various organ systems. There-

fore, once disrupted by xenobiotic chemicals at the crit-

ical period of a developing system, irreversible adversity

appears either immediately or after a certain latent peri-

od until the affected system starts to function at its full

potency (e.g., the reproductive system). From the view-

point of signal toxicity, the developing neuro-immuno-

endocrine network is a well-established target of EDCs.

Relatively recent news on “obesogens” (Janesick and

Blumberg, 2011) indicates that “signal toxicity” can be

systemic beyond the framework of the neuro-immuno-en-

docrine network.

The key to understanding the outcomes of signal tox-

icity of a chemical of concern is to identify the receptor

system(s) that the chemical can act on together with when

and where the receptor system(s) operates in the organ-

ism under development and maturation. There are neu-

rons that express estrogen receptors in the developing

brain. These neurons are sensitive to estrogenic/antiestro-

genic chemicals at a signaling dose that is lower than the

cytotoxic dose. Pesticides directed against neurotransmit-

ter receptors can directly (e.g. neonicotinoids) or indirect-

ly (e.g. acetylcholinesterase inhibitors) trigger signal tox-

icity in the developing brain, because the neurotransmitter

signals are intensively used for the development and func-

tional maturation of the brain (Shelton et al., 2014;

Kimura-Kuroda et al., 2012; Cimino et al., 2017).

It is interesting to note that aryl hydrocarbon receptor

(AhR) gene knockout mice are resistant to dioxin-mediat-

ed teratogenesis and toxicity at large. This ﬁnding clearly

shows that “if there is no receptor, there is no signal tox-

icity.” Neuronal stem cells under development are known

to change their response to differentiation inducers. In the

mouse brain, neuronal stem cells of gestational day (GD)

11.5 can be differentiated only into neurons, whereas

those of GD 14.5 can be differentiated into neurons, astro-

cytes and oligodendrocytes (Namihira et al., 2008). This

process is called “maturation” of the stem cell and it is

plausible that the neuronal stem cell at GD 11.5 and 14.5

has different sensitivity to exogenous signals brought by

neuroactive chemicals. Indeed, transplacental administra-

tion of domoic acid at GD 11.5 and 14.5 resulted in dif-

ferent neurobehavioral anomalies at the age of 11 weeks

(Tanemura et al., 2009). This experiment clearly showed

that signal toxicity is dependent on the status of the target

cells and organs.

As the signaling mechanisms involved at particular

developmental stages are shared by neuronal, immuno-

logical and endocrine systems, the target organs or sys-

tems of signal toxicity can be found in all three systems.

DISCUSSION