Journal of APPLIED BIOMEDICINE
ISSN 1214-0287 (on-line)
ISSN 1214-021X (printed)
Volume 8 (2010), No 1, p 19-22
DOI 10.2478/v10136-009-0003-y
Alternative haematotoxicological testing
Josef Berger
Address: Josef Berger, Faculty of Health and Social Studies, University of South Bohemia, Emy Destinove, 370 05 Ceske Budejovice, Czech Republic
berger@jcu.cz
Received 16th October 2009.
Revised 22nd November 2009.
Published online 5th January 2010.
Full text article (pdf)
Abstract in xml format
Summary
Key words
Introduction
Non-animal assaya
Non-mammalian biomodels
Current trends
References
SUMMARY
Standard haematological procedures in preclinical subchronic and chronic toxicity studies are carried out on dogs, rats and other mammals. In
vitro clonic assays CFU-GM, BFU-E, CFU-E, CFU-Mgkr and pluripotent stem cells are used in screening candidate compounds to predict acute
cytopenias, but they are not able to reveal the risk of suppression which can develop after repeated administration. No in silico studies
have yet been published in haematotoxicology. More recent haematotoxicological in vivo models represent invertebrates: they can be used in
both ecotoxicology and in the screening of myelotoxicity.
KEY WORDS
myelotoxicity; alternative testing; in vitro; in silico; invetebrate
INTRODUCTION
Haematological studies were among the first to be
incorporated into routine toxicology. Classical
preclinical haematoxicological examinations, which
detect the adverse effects of various xenobiotics, are
carried out on laboratory animals in subchronic and
chronic organismal studies (Berger 1987). The use of
laboratory animal models is limited by restrictions
driven by social and economical concerns (cf. Knight
2008).
Alternatives to mammalian models can be in vitro
assays, in silico analyses or the use of invertebrates.
In vitro clonogenic assays have been established
for forty years; in toxicology for the last twenty years.
Computerised methods have also been evaluated for
several decades, but primarily for the prediction of the
desired effects of newly developed drugs. A goal in
predictive toxicology is the use of in vitro methods
and database development to make toxicological
predictions (Guengerich and MacDonald 2007).
Invertebrate models are necessary in many areas
of biomedical research but it seems that they go often
unheeded in ecotoxicology (Berger 2009a).
The aim of our contribution is shortly to discuss
the main problems current in haematoxicology.
NON-ANIMAL ASSAYS
Cytotoxic drugs can be also evaluated by in vitro
clonogenic assays to investigate changes in the
proliferation and differentiation of stem cells (Pessina
et al. 2005, for review). The prediction of in vivo
acute neutropenia using a CFU-GM (colony forming
unit - granulocyte-macrophage) assay seems to be the
most valuable (Masubuchi 2006). Pluripotent
haematopoietic stem cell, CFU-Mgkr
(megakaryocyte) and BFU-E or CFU-E (erythroid)
assays are also useful. They can predict acute
thrombocytopenia, anaemia and aplastic anaemia.
On the other hand, colony growth inhibition
granulocyte-macrophage colony-forming-units
(GM-CFC) may not be correlated with the degree of
myelotoxicity in the same patient (Sessa et al. 1999).
Comparing the differential effects of antitumour drugs
on mature haematopoietic stem cells such as
GM-CFC and undifferentiated long-term
culture-initiating cells, haematopoietic progenitors in
vitro may not represent a valid model for the pattern
of myelotoxicity observed in clinical studies carried
on human subjects (Ghielmini et al. 1999).
Haematological in vitro tests, moreover, seem not to
be able to reveal the risk of the suppression which can
develop after repeated administration of an evaluated
compound.
In silico analysis is asuming some importance in
toxicology (Knight 2008, Nigsch et al. 2009),
especially for predicting preclinical toxicological
endpoints, clinical adverse effects, and the
metabolism of pharmaceutical substances (Valerio
2009). A more recent summary of in silico systems
for toxicity predictions is available in the literature
(Muster et al. 2008). Although included in
haematology for the last ten years, this approach is
limited to the analysis of genes and gene expression
related to malignancy (Stegmaier 2009).
Toxicological database development also covers
haematological data but no in silico studies have yet
been published in haematotoxicology.
NON-MAMMALIAN BIOMODELS
Invertebrates are phylogenetically distant from
humans and their use for the prediction of the
side-effects of various compounds in human subjects
can therefore be disputed. On the other hand, we have
known for many years that the Ames test on
microorganisms is a very useful screening of
mutagenicity. Moreover, systematic reviews of
toxicological results demonstrate that mammalian
models have not been substantially consistent with
clinical outcomes in many studies (Knight 2007).
Invertebrates are already used in several areas of
biomedical research, for example, ageing research
(Flatt and Schmidt 2009), genetics and neuroscience
(Lu 2009, Spokony and Restifo 2009, Yang and Lu
2009), paediatrics and physiology (Zhou et al. 2009),
and oncology (Vaccari and Bilder 2009). There are
also several studies which used invertebrates in
toxicological research (e.g., Berger et al. 2003,
Cummings and Kavlock 2005, Park and Lee 2007,
Verhofstad et al. 2008, Choudhuri 2009).
Xenobiotics influence the total number of
circulating haemocytes; they usually decrease it and
change differentials (e.g., Berger et al. 2003, Auffret
et al. 2004, Yildiz and Benli 2004, Szczerbina et al.
2008, Oweson and Hernroth 2009). A very important
immune function of haemocytes as well as human
neutrophils and monocytes is phagocytosis (Bouchard
et al. 1999, Auffret et al. 2004, Cammarata et al.
2007, Russo et al. 2007, De Guise et al. 2008, Gagne et al. 2008, Oweson et al. 2008). Alterations in
phagocytosis are a consequence of changes in the
cytoskeleton (Cima et al. 2008, Menin et al. 2008).
Some xenobiotics induce programmed cell death,
which can be diagnosed also in haemocytes (Cima et
al. 2008, De Guise et al. 2008). A similar situation
concerns the detection of haemocyte necrosis (Sung
et al. 2003, De Guise et al. 2008).
Haemocytes divide in haemolymph. Their mitotic
index (Oweson 2008) has a similar diagnostic role as
the percentage of mitosis in the bone marrow of
mammals.
Haemocytes make it possible to detect genotoxic
injuries by the micronucleus test (Dopp et al. 1996, Carvalho
Pinto-Silva et al. 2005, Binelli et al. 2008).
CURRENT TRENDS
The main area where invertebrates are used for
haematotoxicology, seems to be ecotoxicology
because contaminants can adversely affect immune
functions in both human subjects and wildlife
(Galloway and Depledge 2001).
The morphology of invertebrate haemocytes is
similar to the mammalian leucocyte (Berger 2009a).
Haemocytes (or coelomocytes) may synthesize
immune proteins and agglutinins and phagocytise
foreign particles (Lavine and Strand 2002). Although
invertebrates have no erythrocytes, haemolytic
potential can be tested by detecting changes in
haemocyte morphology (Berger 2009b).
Invertebrates can be used as in vivo models for
haematotoxicological screening of newly developed
drugs or chemicals in occupational environments. To
reach a better prediction of adverse effects, further
haematology research of invertebrate species and
their haematokinetics seems to be desirable.
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