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Creating a database of equine science information and a system
of free access（2005-2007）
Purpose
Scientific knowledge about horses
is posted on the ERI website and also published in numerous printed
publications. In many cases, however, it is not easy to obtain
the desired scientific information accurately and in a short
time from these many sources.
Therefore, we have created
a database of equine science information and a system for obtaining
that information by accessing the ERI website, so that people
who are interested in scientific information on horses may obtain
that information easily.
Results
First, we selected and organized
the terms to be included in the database (DB), using “Uma no Igakusho”
(Textbook of Equine Veterinary Science) and “Uma Yougoshuu” (Dictionary of Equine Terminology),
among numerous other publications. We created a system for searching
about 500 of these terms, and posted this on the web in a dedicated
programming language (CDML) using search software (FileMaker
Pro Unlimited 6.0). However, we then discovered that the system
had no measures to prevent unauthorized access. To correct this,
we separated the server from the web server, using instead a
database server based on an international standard programming
language (XSLT) and the safest and most reputable web server
(Apache). Meanwhile, by installing the DB server inside an Internet
intruder prevention system (firewall), we eliminated the risk
of unauthorized access. We changed the program from the old HTML-based
website to a new, expandable web 2.0-based website. We changed
language encoding from Japanese standard (S-JIS) to international
standard (UTF-8), making it possible to display specialist terms
using kanji (Chinese characters) that could not previously
be displayed. Using this new system, we published a dictionary
of terms consisting of 1,776 terms (Equipedia). With this system,
the instability of the first system was eliminated, and it is
now possible to search for the desired terms safely and easily
from personal computers and mobile phones.
Research in connection with
the international project on the equine genome - Search for
the genome region Involved in tying-up syndrome - (2005-2007)
Purpose
It has been suggested that tying-up
syndrome may be a genetically governed disease. In this research,
we conducted the following studies based on tying-up syndrome
as a model, the purpose being to establish a method of clarifying
the gene loci of athletic performance or specific traits in Thoroughbreds.
1) First, we identified the region containing gene loci that
are involved in occurrences of tying-up syndrome. 2) Next, we
selected markers linked to the functions of genes in that region,
and investigated the possibility of gene diagnosis.
Results
1. Horses
that had been diagnosed as suffering from tying-up syndrome between
2000 and 2003 were selected from the medical information database
on racehorses (JARIS III). On analyzing the relationship between
those horses and their sires using the Gibbs Sampling for Threshold
Trait program, heritability was estimated at 0.42. From this,
a marked genetic involvement in tying-up syndrome was recognized.
Meanwhile, taking approximately 3,500 racehorses from which blood
had been sampled in 2002, the ten sires in which affected foals
were most numerous were selected, and linkage analysis on half-sibling
families was carried out based on the DNA type of their 478 foals
(72 affected, 406 unaffected). As a result, a strong linkage
was recognized between two microsatellites (UCDEQ411 and COR058)
on the 12th chromosome (ECA12) for one bloodline only. Finally,
144 horses affected by tying-up syndrome and 144 unaffected horses
were selected at random, and a case-control study was carried
out using about 1000 microsatellites. As a result, a significant
linkage disequilibrium was recognized between microsatellites
of the 20th chromosome (ECA20), 12th chromosome (ECA12), and
15th chromosome (ECA15). The microsatellites (TKY409 and COR058)
on ECA12, in particular, backed up the results of linkage analysis
on half-sibling families that had been conducted up to the previous
year.
2. After approximately 2,500
microsatellites had been newly developed and synteny analysis
with human orthologues had been conducted for about 200 of these,
they were then mapped on an existing linkage map. This is helping
to further improve the equine genome map being conducted as an
international cooperative project.
Investigation concerning the
international equine genome project (2000-2004)
The International Equine Genome
Project was started in 1995 and is being promoted through cooperation
between 25 research bodies around the world. In global terms,
the number of researchers working on equine genome research is
extremely limited. This needs to be addressed by an international
project in which research institutions in various countries share
the results and information obtained, with the ultimate aim of
putting this to use in research on equine genes. In Japan, however,
equine genome research is not included in nationally funded genome
research on livestock. Therefore, the ERI has chosen to tackle
this research independently, albeit with cooperation from the
government and the JRA.
The purpose of this study
is to contribute to the development of strong, healthy horses
in future by obtaining and using equine genes and technical information,
as well as fulfilling our role in the International Equine Genome
Project as a center laboratory for equine genome research in
Japan, through cooperation with national research bodies.
Results:
1. We created our own high-grade
(albeit small-scale) equine genome BAC library, and established
technology for creating BAC libraries. This makes it possible
to create a large-scale international BAC library that can be
permanently shared.
2. We developed approximately
2000 polymorphic microsatellite markers, analyzed approximately
1000 of these for half-sibling families, Newmarket full-sibling
families, and RH panels, and arranged these on their respective
maps, in cooperation with the University of California, the Animal
Health Trust, Texas A&M University, and others. As a result,
the precision of all maps was improved by more than double compared
to the previous level.
Studies on the clinical application
of equine chromogranin A (2002-2004)
If the activity of the sympathetic
nervous system and reaction to stress in horses could be measured
using a simple method, it would be possible to ascertain the
exercise adaptation and psychological state of racehorses, thus
providing useful information for daily horse management. So far,
there have been several reports including those of bio-active
substances such as catecholamines, known as indicators for activity
of the sympathetic nervous system. However, a practical method
of measurement that can be used in clinical sites has yet to
be established for any of these. In research so far, we have
focused attention on a neuro-peptide known as chromogranin A
(CgA), which is released into blood together with catecholamines
in reaction to stress. After conducting genetic analysis on this
substance and studying its specificity and method of measurement,
there appeared to be a high possibility that it could be used
as an indicator for a variety of stress stimuli. Moreover, it
has become clear that this CgA is also secreted in saliva in
reaction to stress, and is also very useful as a non-invasive
indicator. The purpose of this research is to establish standard
values for CgA in equine blood and saliva, as well as the range
of fluctuation under various stress conditions. Another aim is
to clarify changes when adapting and not adapting, differences
between individuals, etc., allowing us to draw up standard values
for actual clinical application. Last year, we measured the CgA
in blood and saliva of stud horses when mating, which generally
show a high state of excitement, with the aim of clarifying changes
in CgA in blood and saliva when reacting to stress such as exercise
and emotional disturbance. Furthermore, we are conducting basic
studies on changes in blood CgA values during regularly repeated
all-out exercise load experiments, using horses subjected to
exercise load on a treadmill, with the aim of evaluating the
adaptive response to exercise and condition of Thoroughbreds.
Results
1. The average CgA concentration
in plasma was 2.48±0.09 pmol/ml
with time-related variation (i.e. it was higher in the daytime,
when the sympathetic nervous system is more active, but lower
at night). On the other hand, CgA in saliva concentration showed
considerable variation at each sampling time, and no cyclical
variation was recognized.
2. We confirmed that, when the
parotid gland and submandibular gland are stimulated by Ach-NA
solution, mucous saliva containing high-concentration CgA is
secreted in large volumes, and, from the submandibular gland
in particular, CgA is secreted in a concentration-dependent manner.
 3.
Following stimulation by treadmill exercise, CgA concentration
in blood rose by a maximum 1.5 times compared to when at rest,
while CgA in saliva rose to 21 times compared to when at rest
5 minutes after the exercise. The rise in CgA concentration was
weakened by continued training, suggesting the influence of exercise
adaptation.
4. Increases in CgA concentrations
in blood and saliva were observed due to sexual excitement and
emotional stress when mating.
5. CgA concentrations in blood
and saliva rose due to novel environment stimulus. Furthermore,
when we divided horses into three groups in accordance with changes
in their heart rate due to novel stimuli, our analysis proved
that CgA concentration in saliva was higher in the group with
larger changes in heart rate. It was also shown that this value
has a correlation with adrenalin concentration in blood, and
that, in the group with larger changes in heart rate, it was
already high when at rest. From the above, it was demonstrated
that CgA in blood and saliva is useful as an indicator showing
the activation of the sympathetic nervous system-adrenal medulla
system. Since CgA in saliva has particularly high reactivity
to exercise and psychological stimuli, it should be useful as
a new stress indicator in combination with other biochemical,
electro-physiological, behavioral and psychological indicators.
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