Download Taurine depletion caused by knocking out the taurine transporter

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Journal of Molecular and Cellular Cardiology 44 (2008) 927 – 937
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Original article
Taurine depletion caused by knocking out the taurine transporter gene leads
to cardiomyopathy with cardiac atrophy
Takashi Ito a , Yasushi Kimura a , Yoriko Uozumi a , Mika Takai a , Satoko Muraoka a ,
Takahisa Matsuda a , Kei Ueki b , Minoru Yoshiyama c , Masahito Ikawa d , Masaru Okabe d ,
Stephen W. Schaffer e , Yasushi Fujio a , Junichi Azuma a,⁎
a
Department of Clinical Pharmacology and Pharmacogenomics, Graduate School of Pharmaceutical Sciences, Osaka University, Japan
b
Japan Clinical Laboratories, Inc., Bioassay Division, Japan
c
Department of Internal Medicine and Cardiology, Osaka City University Medical School, Japan
d
Research Institute for Microbial Diseases, Osaka University, Japan
e
Department of Pharmacology, University of South Alabama, College of Medicine, USA
Received 26 November 2007; received in revised form 9 February 2008; accepted 1 March 2008
Available online 18 March 2008
Abstract
The sulfur-containing β-amino acid, taurine, is the most abundant free amino acid in cardiac and skeletal muscle. Although its physiological
function has not been established, it is thought to play an important role in ion movement, calcium handling, osmoregulation and cytoprotection.
To begin examining the physiological function of taurine, we generated taurine transporter− (TauT−) knockout mice (TauTKO), which exhibited
a deficiency in myocardial and skeletal muscle taurine content compared with their wild-type littermates. The TauTKO heart underwent
ventricular remodeling, characterized by reductions in ventricular wall thickness and cardiac atrophy accompanied with the smaller cardiomyocytes. Associated with the structural changes in the heart was a reduction in cardiac output and increased expression of heart cardiac failure
(fetal) marker genes, such as ANP, BNP and β-MHC. Moreover, ultrastructural damage to the myofilaments and mitochondria was observed.
Further, the skeletal muscle of the TauTKO mice also exhibited decreased cell volume, structural defects and a reduction of exercise endurance
capacity. Importantly, the expression of Hsp70, ATA2 and S100A4, which are upregulated by osmotic stress, was elevated in both heart and
skeletal muscle of the TauTKO mice. Taurine depletion causes cardiomyocyte atrophy, mitochondrial and myofiber damage and cardiac
dysfunction, effects likely related to the actions of taurine. Our data suggest that multiple actions of taurine, including osmoregulation, regulation
of mitochondrial protein expression and inhibition of apoptosis, collectively ensure proper maintenance of cardiac and skeletal muscular structure
and function.
© 2008 Elsevier Inc. All rights reserved.
Keywords: Taurine; Taurine transporter; Heart; Cardiomyopathy; Atrophy; Transgenic mice; Mitochondrial defect; Cell volume; Osmoregulation
1. Introduction
The sulfur-containing amino acid, taurine (2-ethanesulfonic
acid), is the most abundant free amino acid in mammalian tissue,
reaching concentrations as high as 5–20 μmol/g wet wt [1,2].
⁎ Corresponding author. Department of Clinical Pharmacology and Pharmacogenomics, Graduate School of Pharmaceutical Sciences, Osaka University. 1-6
Yamadaoka, Suita, Osaka, 565-0871, Japan. Tel.: +81 6 6879 8258; fax: +81 6
6879 8259.
E-mail address: [email protected] (J. Azuma).
0022-2828/$ - see front matter © 2008 Elsevier Inc. All rights reserved.
doi:10.1016/j.yjmcc.2008.03.001
Since the capacity to synthesize taurine in most tissues including
the heart and skeletal muscle is limited, maintenance of the large
intracellular taurine pool depends upon uptake of the amino acid
from the blood. This transport process requires the accumulation
of taurine against a substantial concentration gradient, as the
concentration of taurine is 100 fold less in the plasma (20–
100 μM) than in the tissues (5–20 μmol/g wet wt.) [1,2]. Taurine
uptake is mediated by an ubiquitous Na+ and Cl− dependent
transporter, which uses the Na+ gradient across the cell membrane
to drive taurine accumulation [3–5]. The expression of the taurine
transporter (TauT; SLC6a6) is regulated by osmotic stress and
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T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
transcription factors, such as NFAT5, MEF-2 and p53 [4,6,7],
which control intracellular taurine content.
Accumulating evidences appear that taurine plays cytoprotective roles in the hearts. Oral supplementation of taurine is effective
to animal model and human patients with congestive heart failure
and cardiomyopathy [8–11]. Although the essential role of
taurine in heart has not been clarified, taurine exerts several
actions that could potentially benefit the diseased heart. First, it
modulates ion transport and regulates intracellular calcium levels
[12–14]. Maintenance of Ca2+ homeostasis is of paramount
importance in the heart because either reductions in [Ca2+]i or
impaired Ca2+ sensitivity of the myofibrils can lead to the
development of heart failure. Second, it possesses antioxidant
[11–13] and anti-apoptotic activity [15,16], which would be
expected to limit ventricular remodeling. Finally, taurine is a key
osmoregulator in the heart [13], an action that should limit
damaging osmotic imbalances that develop in conditions, such as
ischemia. Interestingly, taurine content is altered in various
pathological states [10,17]. Based on these findings, it has been
suggested that severe reductions in the size of the intracellular
taurine pool, as occurs in ischemia, may contribute to cell
shrinkage and the development of pathological lesions. On the
other hand, increases in taurine levels in conditions, such as
failing and hypertrophic heart. Taken together, it is hypothesized
that taurine would play a critical role in compensatory adaptation
against the pathophysiological loads associated with the development of myocardial hypertrophy and/or heart failure.
In a small number of species (fox and cat), taurine levels can be
dramatically diminished merely by reductions in dietary taurine
content [18,19]. After prolonged exposure to the taurine depletion
condition, the nutritionally deprived animals develop cardiomyopathy as well as retinopathy or immune deficiency [18,20,21]. In
contrast to fox and cat, the size of the intracellular taurine pool of
most animal species remains fairly constant even with significant
reductions in dietary taurine content. This occurs because a
decline in plasma taurine levels is accompanied by enhanced
cellular retention of taurine [22]. Despite resistance to depletion,
tissue taurine levels can be decreased by treatment of these
animals with a taurine transport inhibitor, such β-alanine or
guanidinoethane sulfonate, which interferes with taurine uptake
by the tissues [10,23,24]. However, treatment of these animals
with a taurine transport inhibitor does not generally cause sufficient taurine deficiency to promote the development of severe,
overt pathology. This has been attributed to the limited capacity of
the taurine transport inhibitors to cause severe taurine deficiency.
However, resistant species, such as rodents, also exhibit a greater
capacity to synthesize taurine than cats or fox. Therefore, a more
effective means of producing taurine deficiency in the rodent is
the formation of TauT null animals. The present study shows that
depletion of taurine in the TauT null mouse is associated with the
development of a cardiomyopathy.
2. Methods
2.1. Generation of TauT-null Mice
All animal experiments were performed in accordance with
protocols approved by the Animal Care and Use Committee of
Graduate School of Pharmaceutical Sciences, Osaka University. A
clone of the murine TauT gene including exon 2 to exon 5 was
isolated from the 129SVJ murine genomic library in bacterial
artificial clone, as identified by PCR with specific primers for the
TauT gene (Table 1). The targeting vector was designed to replace
the Stu I–Xba I fragment including the end of exon 2 to exon 4
with a gene conferring neomycin resistance (Fig. 1A). The Xba I
fragment (9.6 kb) containing exon 5 was inserted into the Xba I site
between hsv-thymidine kinase and the neomycin-resistant genes
of pPTN plasmid [25]. A 1.5-kb Stu I fragment containing intron 1
was inserted at the blunt-ended Xho I site, which is upstream of the
neomycin resistance gene. The targeting vector was transfected
Table 1
PCR primers used in RT-PCR
Gene
TauT
Primer sequence
exon2
exon4
exon5
Hsp70
(NM_010478)
Hsp40
(NM_018808)
Slc38a2
(NM_175121)
s100a4
(NM_011311)
s100a9
(NM_009114)
tnfrsf12a
(NM_013749)
GAPDH
(NM_008084)
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Forward:
Reverse:
Amplicon size
5′-AAAACGAAGAGATGGCCACGA-3′
5′-AGCAGAGGTACGGGAAACGC-3′
5′-TCATTGTGTCCCTCCTGAACGT-3′
5′-GTGAGGTGAAGTTGGCAGTGCT-3′
5′-AGCGCAATGTGCTCAGCC-3′
5′-CCTTCCAGATGCAGAAAAAACAG-3′
5′-AGAACGCGCTCGAATCCTATG-3′
5′-TCCTGGCACTTGTCCAGCAC-3′
5′-TTTTCGACCGCTATGGAGAGG-3′
5′-AACTCAGCAAACATGGCATGG-3′
5′-TGCAGGCCACGCTATTTCA-3′
5′-GGCTGGCGGCTCTTTAGCTCT-3′
5′-AGGACACCCTGACACCCTGAA-3′
5′-CCTGGTTTGTGTCCAGGTCCT-3′
5′-TCAAGCTGAACAAGACAGAGCTCA-3′
5′-TCCCTGTTGCTGTCCAAGTTG-3′
5′-GCCGCCGGAGAGAAAAGTT-3′
5′-TCCTCACTGGATCAGTGCCAC-3′
5′-GCCGGTGCTGAGTATGTCGT-3′
5′-CCCTTTTGGCTCCACCCTT-3′
225 bp
196 bp
123 bp
113 bp
126 bp
121 bp
134 bp
121 bp
82 bp
87 bp
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
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GAC CAC TTC TCC CTC-3′) and neoR (5′-AAG CGC ATG
CTC CAG ACT G-3′) (Data not shown). The mice which backcrossed at least 4 times into C57BL/6 line were used for analysis.
2.2. Reverse transcript-(RT-) PCR analysis
Total RNA was isolated from the cardiac and skeletal muscles
of TauTKO and wild-type mice by using Qiazol (Qiagen) according to manufacture's protocol. Total RNA (1 μg) was subjected to the reverse transcription with Rever Tra Ace (Toyobo),
using oligo(dT)12–18 primer (Invitrogen) at 42 °C for 60 min,
followed by PCR. For TauT gene analyses, PCR amplification
was carried out by using the primers specific for either exon 2 or
exon 4 or exon 5 of taut gene shown in Table 1, and then PCR
products was size-fractionated by 2% agarose gel electrophoresis
and detected with ethidium bromide.
Quantitative RT-PCR analyses were performed by using
ABI7700 (Applied Biosystems) with SYBR green and TaqGold DNA polymerase (Applied Biosystems), as described
previously [26]. The PCR primers used were shown in Table 1.
GAPDH was used as an internal control.
2.3. Measurement of taurine transporter activity in cultured
cardiomyocytes
Fig. 1. Outline of strategy for targeted disruption of TauT gene. A; Restriction
maps of the wild-type mice, targeting vector and predictional mutant allele. Opened
boxes indicate exons (2–5). The genomic fragments used 5′-probe and 3′-probe are
shown in bold gray bars. neo: PGK-neomycin resistance gene, tk: PGK-thymidine
kinase gene. B; Southern blot analysis of the mutant mice. SmaI and BglII-degested
genome samples were used for the analysis. Southern blot with the 5′-probes
detected the predicted restriction fragments shown in A. C; Total RNA from hearts
of wild-type mice (+/+), TauT+/− and TauT−/− mice was subjected to reverse
transcript-PCR. PCR was performed at 36cycles by using the specific primers for
exons 2, 4 and 5. D; Taurine uptake activity of cardiomyocytes from neonatal mice
(TauT+/+, +/− and −/−) was measured in vitro by using 1,2-[3H]-taurine. Data
were normalized by cellular protein content. Data are mean ± S.E., n = 5 (TauT−/−),
5 (+/−) and 3 (+/+). ⁎; p b 0.001. E; HPLC analyses for the measurement of cardiac
taurine content. Representative charts are shown. An arrow indicates the peak for
taurine. Quantified results are shown in Table 2.
into the D3 embryonic stem cells (ES cells) by electroporation, and
selected with G418 and gancyclovir. The targeting events were
screened by PCR and confirmed by Southern blot analysis using
digestion with restriction enzymes and both 3′- and 5′-end probes
(Data not shown). The recombinant cells were karyotyped to
ensure that 2 N chromosomes were present in the majority of the
metaphase spreads. Chimeric mice, which were derived from
correctly targeted ES cells, were mated to C57BL/6 mice to obtain
TauT+/− mice, and backcrossed into C57BL/6 mice. The mice
were maintained under controlled environmental conditions (22 ±
1 °C; 12:12-h light–dark cycle, lighting on at 08:00 h; food and
water ad libitum). To identify the genotypes, the mouse genomic
DNA (5 µg) was digested with restriction enzymes and was
subjected to Southern blot with 5′-specific probes (Fig. 1B). The
genotypes of the mice were also determined by PCR, with the
genomic DNA isolated from mouse tails used as a template and
specific primers, as follows; taut/intron1F (5′-AGG TTG GAA
CTG GTC TTA AGT CAC TG-3′), taut/exon2R (5′-TTG CTG
Taurine transport activity was measured as previously
described [6]. Cardiac myocytes were isolated from the hearts
of neonatal wild-, hetero- or homozyogote TauTKO mice, and
were cultured in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum for 1 day before use. After washed,
cells were incubated with uptake buffer containing 5 μM [1,
2-3H] taurine (0.1 μCi/mL) for 10 min, washed twice with cold
phosphate buffered saline (PBS) and lysed with 0.1 M NaOH.
Radioactivity of the lysate was measured by liquid scintillation
spectrometry, and normalized with the cellular protein content.
2.4. Measurement of tissue taurine concentration
Tissue taurine concentration was measured, as reported
previously [27]. In brief, tissues from mice were homogenized
with 9 volumes of ethanol, and homogenates were centrifuged at
5000 ×g for 10 min. After supernatants were dried, samples were
dissolved with drying solution (two parts water, two parts ethanol,
and one part triethylamine). After drying again, samples were
dissolved with derivatizing solution (seven parts ethanol and one
part each phenylisothiocyanate, triethylamine, and water),
vortexed and incubated at room temperature for 20 min. After
drying again, the phenylisothiocyanate derivatives were dissolved
in 10 mM phosphate buffer, pH 6.4, filtered and separated by a
high-pressure liquid chromatography system (Wakosil-PTC
column; Wako), as described in the manufacturer's protocol.
2.5. Histological analysis
For Hematoxylin and Eosin staining and Masson's trichrome
staining tissues from mice were fixed in 4% paraformaldehyde/
phosphate-buffered saline (PBS), washed, and embedded in
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T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
paraffin. Sections were stained by Hematoxylin and Eosin
staining method and Masson's trichrome staining method.
Myocyte cross-sectional area was measured from images
captured from Hematoxylin and Eosin-stained sections. Suitable
cross sections were defined as having nearly circular myocyte
sections. The outline of over 100 myocytes was traced in at least
3 sections. The Scion Image software (Scion Corporation) was
used to determine myocyte cross-sectional area.
For electron microscopic analyses, tissues were fixed in 2%
paraformaldehyde and 2% glutaraldehyde/0.1 M HEPES buffer,
washed, postfixed with 2% osmium tetroxide solution, dehydrated in ethanol series, and embedded in epoxy resin. Sections
were viewed by transmission electron microscope, H-7100
(Hitachi, Japan) at a magnification of × 3,000–10,000.
Staining the tissue section for succinate dehydrogenase
activity was performed as described previously [28,29]. In brief,
the hearts were frozen, and 5-µm sections were prepared with a
cryostat, Leica CA 1850 (Leica). The sections were incubated
in 0.1 M phosphate buffer (pH 7.4) containing 0.1 M sodium
succinate and 0.1% nitro blue tetrazolium at 37 °C for 5 min. The
staining intensity (blue color) of each section was quantified by
using Scion Image software (Scion Corporation).
2.6. Doppler echocardiography
Echocardiographic measurements were performed as
described previously [30,31]. The mice were lightly anesthetized
with an injection of ketamine hydrochloride (25–50 mg/kg, i.p.)
and xylazine (5–10 mg/kg, i.p.). Echocardiograms were performed with an echocardiography system equipped with a 7.5-MHz
phased-array transducer (SONOS 5500; Philips Medical System,
Best, The Netherlands). To measure LV end-diastolic dimension
(LVDd) and LVend-systolic dimension (LVDs), two-dimensional
short-axis views of the left ventricle were obtained through
the anterior and posterior LV walls at the papillary muscle level. Fractional shortening (%FS) and ejection fraction (%EF)
calculated by the cubed method according to previous report [32].
2.7. Northern blot analysis
Total RNA isolated from each heart was assessed by Northern
blotting as previously described [33]. cDNA probes for BNP and
GAPDH were labeled with the Megaprime DNA Labeling
System (Amersham Bioscience, USA) according to the protocol.
The following oligonucleotides (Invitrogen, USA) were labeled
with γ-32P ATP by using T4 polynucleotide kinase (TOYOBO,
Japan) according to the protocol before being used as probes: βMHC: 5′-GAG GGC TTC ACG GGC ACC CTT AGA GCT
GGG TAG CAC AAG ATC TAC TCC TCA TTC AGG CC-3′,
ANP: 5′-CCG GAA GCT GCA GCC TAG TCC ACT CTG GGC
TCC ATT CCT GTC AAT CCT ACC CCC GAA GCA GCT
GAA-3′.
[34]. The mice which were attached on the tail with a load (6% of
body weight) placed in the swimming pool filled with fresh water
at 30± 1 °C (depth: N 20 cm). The swimming time until the point at
which mice could not return to the surface of the water more than
5 s after sinking was measured.
2.9. Microarray analysis
Total RNA from hearts or skeletal muscle of 3 independent
control and TauTKO mice were pooled. CDNA was synthesized
from 2 μg of pooled total RNA and annealed to oligo-(dT)24
primer. Reverse transcription was performed with Superscript II
reverse transcriptase. Second-strand cDNA synthesis was performed with DNA polymerase I with the appropriate reagents.
Synthesis of biotin-labeled cRNA was performed by in vitro
transcription. The cRNA was fragmented and hybridized to the
GeneChip Mouse 430 ver.2 array Set (Affymetrix) in the
appropriate hybridization solution. After washing and staining,
probe arrays were scanned with the GeneChip Scanner 3000
controlled by GeneChip operating software (Microarray Suite
5.0 Expression Analysis Program, Affymetrix). Genes which
expression calls are “present”, expression levels are over 1000
in both control and TauTKO samples and changes are more than
1.8-fold compared with controls were regarded as differently
expressed.
2.10. Statistic analysis
Each value was expressed as the mean ± standard error (SE).
Statistical significance was determined by Student's t-test.
Differences were considered statistically significant when the
calculated p value was less than 0.05.
3. Results
3.1. Generation of TauT-null mice
A targeting construct was generated to replace exons 2–4 of
the TauT gene with a cassette containing a neomycin-resistance
gene (see Methods and Fig. 1A). Germline transmission of the
mutant allele was confirmed by Southern blotting (Fig. 1B) and
PCR. Disruption of the TauT gene in heart resulted in an truncated
transcript that included exon 5 but not exon 2–4; in other tissues
reverse transcript-PCR was used to evaluate knockout of the TauT
(Fig. 1C). As expected, cellular taurine uptake activity was
severely diminished in cultured cardiomyocytes obtained from
Table 2
Taurine concentration in tissues of control and TauTKO mice
Wild
Heart
Skeletal muscle
Eye
Brain
6.7 ± 2.7
(4)
ND
(4)
16.9 ± 5.1
(4)
0.70 ± 0.65⁎
(4)
5.3 ± 1.0
(4)
ND
(4)
1.6 ± 0.1
(4)
0.24 ± 0.02⁎
(4)
2.8. Exercise endurance test
KO
Exercise endurance capacity was determined by weight-loaded
swimming test as described previously with some modifications
μmol/g tissue wet weight (n). Data are mean ± SE. ⁎: p b 0.01 v.s. wild. ND; not
detectable.
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
931
Table 3
Body weight, tissue weight and food intake
Gene type
wild
hetero
KO
BW
HW
TAW
TBL
HW/TBL
Food intake
water intake
g (n)
mg (n)
mg (n)
cm (n)
mg/cm (n)
g/day (n)
ml/day (n)
28.23 ± 0.40
(5)
27.32 ± 0.66
(15)
23.41 ± 0.85⁎⁎,##
(12)
145.0 ± 1.3
(5)
136.2 ± 4.5
(15)
110.8 ± 3.8⁎⁎,##
(12)
56.7 ± 0.5
(4)
57.0 ± 0.1
(8)
46.2 ± 2.6⁎,##
(11)
1.81 ± 0.03
(4)
1.77 ± 0.07
(4)
1.78 ± 0.05
(4)
0.080 ± 0.014
(4)
0.080 ± 0.011
(4)
0.061 ± 0.004⁎,#
(4)
3.70 ± 0.28
(4)
4.82 ± 0.28
(4)
–
4.02 ± 0.31
(4)
–
6.06 ± 0.61
(4)
Measurement of body weight (BW), heart weight (HW), tibial anterior muscle weight (TAW), tibial bone length (TBL), HW/TBL ratio and the amount of food and
water intake of male 3- to 4-month-old control and TauTKO mice. Data are mean ± SE. ⁎:p b 0.05, ⁎⁎: p b 0.01 vs wild-, #:p b 0.05,##:p b 0.01 vs hetero-littermates.
TauTKO mice (Fig. 1D), indicating the selectivity of the mutant
allele for TauT function. Consistent with the loss of TauT activity,
tissue taurine levels decreased (Fig. 1E, Table 2).
Mating between male and female TauT heterozygote mice
yielded taut+/+ (wild-type), taut+/− and taut−/− at a 1:2:1 Mendelian
ratio. TauTKO mice exhibited a lower body weight than their
control littermates, concomitant with the decrease in heart and
skeletal muscle weight (Table 3). In those experiments, the
littermate controls used were taut+/+ and taut+/− mice; no phenotypic differences were observed between these genotypes. Food
and water intake were identical in the TauTKO and control mice
(Table 3).
3.2. TauTKO mice exhibit decreased myocyte size of both the heart
To examine whether knockout of the TauT affected cardiac
morphology, hearts were stained with Hematoxylin and Eosin. It
was found that the ventricular wall was thinner in the TauTKO
hearts compared to their control littermates (Figs. 2A–C). Based
on the cross-sectional area of the left ventricle, the size of the
cardiomyocyte was markedly decreased in the TauTKO heart
(Figs. 2D–F).
3.3. Taurine depletion caused cardiac dysfunction
Based on echocardiographic analysis of the TauTKO and
control mice, there was no significant difference in left ventricular
end-diastolic dimensions, while end-systolic dimensions were
modestly increased in TauTKO compared with control mice
(p b 0.1) (Fig. 3A). Importantly, both fractional shortening and
ejection fraction were diminished in the older (N 9-months old)
TauTKO mice, indicating that cardiac function was eventually
compromised in the TauTKO mice (Fig. 3A). Histological examination using Masson's trichrome staining showed no fibrosis
in the aged TauTKO hearts (data not shown). However, the
mRNA levels of the ANP, BNP and β-MHC genes, which are
pathological markers of heart failure, were significantly increased
in the TauTKO mice (Figs. 3B, C). Importantly, the levels of
Fig. 2. Histological characterization of TauTKO heart and skeletal muscle. A,B; Representative heart transverse sections of 3-month-old control (A) and TauTKO (B)
mice. Hematoxylin–Eosin staining was used. C; Quantification of wall dimension of left (LV), right ventricles (RV) and septum of hearts from wt and KO mice (n = 4).
Data are mean ± SE. ⁎: p b 0.01. D,E; Representative micrographs of left ventricular heart sections from 3-month-old control (D) and TauTKO (E) mice stained with
Hematoxylin–Eosin. Scale bars = 50 μm. F; Quantification of cross-sectional area of left ventricular heart from control and KO mice. A total of N100 individual
myocytes from 4 different mice. Data are mean ± SE. ⁎: p b 0.01.
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T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
Fig. 3. Analyses of in vivo cardiac function and gene expression in TauTKO mice. A; Echocardiographic analyses of TauTKO mice. left ventricular diastolic and
systolic dimension (LVIDd and LVIDs) and percent of fractional shortening (FS) and ejection fraction (EF) in N9-month-old (aged) control and mutant littermates were
determined by echocardiography. Data are mean ± SE, n = 5 (wild) and 7 (TauTKO). ⁎: p b 0.05. B; Northern blot analysis for cardiac failure markers in hearts of
control and TauTKO mice. The total ventricular RNA from 3–5-month-old (adult) or N9-month-old (aged) control and TauTKO mice were subjected to Northern
blotting with various probes, as indicated. C; Quantification of the intensity of the bands from 4 independent experiments by densitometry. Relative intensities
(normalized to GAPDH) are shown. Data are mean ± SE. ⁎: p b 0.01.
these genes were more severely upregulated in the aged TauTKO
mice than in the 3-month-old mice. These data demonstrate that
knockout of the TauT leads to an age-dependent dilated
cardiomyopathy characterized by thinning of the ventricular wall.
3.4. Breakdown of myofiber and mitochondrial ultrastructure
in TauTKO heart
In contrast to the orderly myofibrillar architecture present in
wild-type cardiac muscle, electron microscopy revealed pronounced myofibrillar fragmentation, mitochondrial swelling,
disruption of the outer mitochondrial membrane and cellular
vacuolization in aged, TauTKO hearts (Figs. 4A, B).
3.5. TauT-knockout resulted in the downregulation of succinate
dehydrogenase
To further investigate the mitochondrial phenotype, the hearts
of TauTKO mice were stained for SDH activity. Consistent with
a defect in mitochondrial morphology, both 3 month old and
aged TauTKO hearts showed decreased SDH activity compared
with their wild-type littermates (Fig. 5).
3.6. TauTKO muscles exhibited decreased cell volume, structural
defects and lowered exercise endurance capacity
Since TauTKO mice also lost muscle mass, the skeletal muscle
was analyzed histologically. As expected for atrophic muscle, the
myofibrillar cross-sectional area of the tibial anterior muscle of
the mutant mice was remarkably reduced compared to that of their
littermate controls (Figs. 6A–C). Based on electron microscopic
analyses, the wild-type showed normal myofibrillar ultrastructure
while skeletal muscle from TauTKO mice displayed myofiber
disruption, which was more severe than the lesions seen in the
TauTKO heart (Figs. 6 D,E).
Swimming endurance time of the TauTKO mice was also
reduced, suggesting that taurine deficiency leads to an impairment in skeletal muscle function (Fig. 6F).
3.7. Induction of osmotic stress-sensing genes in the TauTKO
heart and skeletal muscle
Microarray analyses of pooled total RNA from hearts or skeletal
muscle of 3 independent control and TauTKO mice revealed that
several genes were induced in the TauTKO compared to their
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
933
Fig. 4. Ultrastructural defects in TauTKO hearts. Transmission electron microscopic analysis of sections of left ventricle from N9-month-old control (A) and TauTKO (B).
Representative electron micrographs are shown. Magnification; ×3000 and ×10,000 (as indicated). Scale bars = 3.3 μm (×3000) and 1 μm (×10,000). Similar results were
obtained from different examinations. m: mitochondrion, arrows: mitochondria breakdown, arrowheads; myofilament breakdown.
control littermates (Table 4). Fig. 7 reveals that similar data were
obtained using qRT-PCR. These included genes encoding stressinducible chaperones (hspa1a/b; Hsp70 and dnajb1; Hsp40), the
amino acid transporters (slc38a2; ATA2), S100 calcium binding
proteins (S100A4 and S100A9) and the TNFα receptor superfamily
(tnfrsf12a, TWEAK Receptor). It is noteworthy that Hsp70, ATA2
and S100A4, as well as TauT, are regulated by hypertonic stress
[35–37]. Taken together, it is obvious that signaling pathways
responsible for osmoregulation are activated in the heart and
skeletal muscle of the TauTKO mice.
which are accompanied with an increase in myocyte volume. In
the case of dilated cardiomyopathy, while ventricular wall is
thinned, myocyte is commonly enlarged because of the increase
4. Discussion
The present study demonstrates that the downregulation of
the taut gene leads to cardiac dysfunction, ventricular remodeling, upregulation of cardiac failure marker genes, loss of body
weight and a decrease in exercise capacity. At the cellular level,
it was shown that myocytes from the heart and skeletal muscle
of TauTKO mice lost cell volume, develop mitochondrial
defects and undergo myofibrillar disruption. These findings are
consistent with the actions of taurine as an osmoregulator,
mitochondrial modulator and a cytoprotective agent.
In the present study, taurine deficiency leads thinned ventricular wall and cardiac atrophy with decreased myocyte volume.
It is well known that cardiac workload influences myocyte
morphology and function. Volume and pressure overload induce eccentric and concentric cardiac hypertrophy, respectively,
Fig. 5. Enzymatic histological staining for mitochondrial enzyme A,B;
Succinate dehydrogenase (SDH) activity was determined in sections of left
ventricles from N9-month-old control (A) and TauTKO mice (B). Representative micrographs are shown. Intensity of blue color indicates SDH activity of
each section. Magnification; x200. C; The intensity of SDH activity (blue) were
quantified by from 4 independent experiments by densitometry. Relative
intensity is shown. Data are mean ± SE. ⁎: p b 0.01.
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T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
Fig. 6. Histological characterization of TauTKO skeletal muscle. A,B; Representative micrographs of tibial anterior muscle sections of 3-month-old control (A) and
TauTKO (B) mice stained with Hematoxylin–Eosin. Scale bars = 50 μm. C; Quantification of cross-sectional area of tibial anterior muscle sections from control and
KO mice. A total of N100 individual myocytes from 4 different mice. Data are mean ± SE. ⁎: p b 0.01. D,E; Transmission electron microscopic analysis of sections of
tibial anterior muscle from control (D) and TauTKO (E). Representative electron micrographs are shown. Magnification; ×3000. Scale bars = 3.3 μm. Similar results
were obtained from different examinations. F; Total exercise capacity of 3-month-old control and TauTKO mice. Mice were subjected to a swimming endurance test.
Data shows swimming endurance time to sinking to the bottom from 5 different mice. Data are mean ± SE. ⁎: p b 0.01.
of wall stress, in contrast to TauTKO model. Importantly, sideto-side slippage of myocytes contributes to ventricular dilatation, such as the ventricles after myocardial infarction [38]. This
idea indicates that slippage of myocytes would occur in heart of
TauTKO mice as adaptive response to myocyte atrophy, which
results in maintaining ventricular volume. On the other hand,
cardiac unloading, which is induced by heart transplantation
and ventricular assist devises, results in cardiac atrophy and a
decrease in myocyte volume [39,40]. Although histological and
functional features of cardiac atrophy has not been well understood, mechanical unloading does not influence cardiac output
despite of myocyte atrophy in unloading model of dogs in vivo
[41]. Similarly, titin N2B region knockout mice shows reduced
heart size but normal cardiac output[42]. Notably, in the present
study, echocardiographic analyses revealed that diastolic and
systolic ventricular dimensions are not changed and modestly
increased in TauTKO mice, respectively, while both ventricular
dimensions are reduced concomitant with the decreased
myocyte volume in the unloaded dogs and the N2B-knockout
mice [41,42]. Thus, to our knowledge, we consider that
TauTKO mice show a novel cardiac phenotype.
While taurine mediates a number of biological actions in
mammalian cells, one of the most important actions is osmoregulation [3,12,13]. When a cell is subjected to hyperosmotic
stress, organic osmolytes, such as taurine, accumulate, an effect
that minimizes the movement of water and the resulting decrease
in cell volume [43]. By contrast, hypo-osmotic stress, which
increases cell volume, leads to a loss of taurine and a decrease in
intracellular osmolality. Although the regulatory volume change
is usually transient, in the case of the cardiomyocyte of the
Table 4
Upregulated genes in both heart and skeletal muscle of TauTKO mice identified by microarray analysis
Genbank ID
Symbol
Gene description
Fold change (Heart)
Fold change (Quadri)
M12573
AW763765
AW413169
NM_009114
AV306676
AK002290
D00208
AV067695
NM_013749
AK016563
NM_022018
Hspa1b
Hspa1a
Slc38a2
S100a9
Homo sapiens heat shock 70 kDa protein 1B
Homo sapiens heat shock 70 kDa protein 1A
Solute carrier family 38, member 2 (amino acid transporter A2)
S100 calcium binding protein A9 (calgranulin B)
EST
DnaJ (Hsp40) homolog, subfamily B, member 1
S100 calcium binding protein A4 (metastasin)
EST
Tumor necrosis factor receptor superfamily, member 12a
EST
Niban protein
14.86
6.57
4.03
2.85
2.53
2.09
2.02
2.00
1.89
1.88
1.84
15.23
10.34
2.38
3.83
2.25
3.27
2.18
2.36
2.80
2.15
2.58
Dnajb1
S100a4
Tnfrsf12a
Niban
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
Fig. 7. Osmosensitive genes and transcriptional factor were induced in the
TauTKO heart and skeletal muscle. Quantitative RT-PCR analyses for indicated
genes in heart (A) and skeletal muscle (quadriceps; B). The total RNA from 3–5month-old control and TauTKO mice were subjected to qRT-PCR. Data were
normalized with the expression level of GAPDH mRNA. Data are mean ± SE,
n = 4–7. ⁎; p b 0.05, ⁎⁎; p b 0.01 v.s. control littermates.
TauTKO mouse there is a chronic decrease in cell volume.
Interestingly, cardiomyocyte volume is also chronically decreased
in cells rendered taurine deficient by treatment with the taurine
transport inhibitor, β-alanine [44]. Similar observation has been
reported in mice lacking transcriptional factor NFAT5 [45] which
plays a central role for adaptation against osmotic stress and the
induction of genes responsible for the accumulation of osmolytes,
including taurine. Loss of NFAT5 leads to renal atrophy accompanied with the downregulation of transporters specific for
osmolytes [45]. Importantly, NFAT5-deficient mice displayed
lower body weights. Taken together, chronic dysfunction of
osmoregulatory system impairs the cell volume regulation, even
in the ordinary (isotonic) condition, and causes the reduction of
cell volume.
The present study also raises the possibility that mitochondrial defects might accelerate the onset of heart failure. This idea
is consistent with recent reports suggesting that mitochondrial
defects are a major cause of cardiac dysfunction [46,47]. Several
actions of taurine point to the mitochondria as a logical cause of
the cardiomyopathy. First, taurine is a modulator of mitochondrial apoptosis [15,16]. The appearance of matrix swelling and
rupture of the outer mitochondrial membrane imply initiation of
the mitochondrial permeability transition in the taurine deficient
heart. It has been established that the mitochondrial permeability
935
transition pore is activated by Ca2+ and that taurine is recognized
as an important modulator of [Ca2+]i. However, taurine does not
appear to directly modulate the mitochondrial permeability
transition, as it also exhibits no direct effect on Ca2+-induced
activation of the mitochondrial permeability transition in
isolated mitochondria [48]. Thus, taurine apparently exerts an
indirect effect on the mitochondrial permeability transition pore,
either through the modulation of [Ca2+]i or activation of a
survival pathway that regulates the pore. Second, taurine is a
constituent of mitochondrial t-RNA [49,50]. Therefore, taurine
deficiency might affect the expression of mitochondrially encoded proteins, whose function is to ensure proper maintenance
of the electron transport chain. It has been documented that
slowing of electron transport can lead to excessive oxidative
stress, as electrons are diverted from the electron transport chain
to form superoxide. Third, mitochondrial function depends upon
the maintenance of an appropriate osmotic balance between the
mitochondrial matrix and the cytosol. As an osmolyte, taurine
deficiency might alter this balance, leading to excessive mitochondrial swelling, which could weaken the outer mitochondrial
membrane.
It has been previously reported that nutritional depletion of
taurine leads to the development of a dilated cardiomyopathy in
cat and fox [18,19]. However, in apparent contrast to the
nutritional studies, Warskulat et al. failed to detect abnormal
function and morphology in hearts of TauTKO mice [51]. Their
findings also contrast with the present observation that TauTKO
mice exhibit cardiac dysfunction and undergo ventricular remodeling, effects associated with an elevation in the mRNA levels of
ANP, BNP and β-MHC, which are fetal genes commonly
upregulated in cardiomyopathy. Although the basis for the
differences between our results and those of Warskulat et al. [51]
remain unclear, it is possible that the differences are caused by
additional mutations, difference of genetic background, the aging
process and/or diet. We used mice which backcrossed at least 4
times into C57BL/6 line to minimize genetic differences other
than the absence of the TauT gene, while Warskulat et al. [51]
reported to use mixed genetic background (C57BL/6 (Bl6) × 129/
SvJ (129)). Thus, it is possible that this discrepancy could relate to
differences in the genetic backgrounds of the Bl6 and 129 mice,
which clearly have different cardiovascular phenotypes [52] and
different susceptibilities against pathological stressors [53]. Since
we did not measure cardiac output in the mixed genetic mice, we
are presently unsure if the differences in cardiac function between
the two TauTKO models primarily reside with the strains
themselves. The difference between the two models could be
explained if the Bl6 mice had intrinsically lower cardiac output or
if they were more susceptible to the effects of taurine deficiency.
In summary, the present study provides evidence that taurine
is necessary for myocyte volume regulation and organelle
stability in heart and skeletal muscle. Further, our data indicate
that chronic taurine deficiency potentiates age-related, cardiac
dysfunction and skeletal muscle atrophy, implying that taurine
deficiency somehow affects the aging process. Clearly, further
studies are required to clarify the mechanism by which taurine
deficiency triggers severe myocardial and skeletal muscle
defects.
936
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
5. Funding sources
This study was supported in part by a Grants-in-Aid from the
Ministry of Health, Labour and Welfare and from the Ministry
of Education, Science, Sports and Culture of Japan. This study
was also partly granted by Taisho Pharmaceutical Ltd. and The
Nakatomi Foundation.
Acknowledgments
We thank Ms. Yasuko Murao for her excellent secretary
work, Mr. Eizi Oiki (Osaka University) for his technical support
for electron microscopic analyses, and Dr. Nishiya and Dr.
Nakata (Osaka City University) for their technical support for
echocardiographic analyses.
References
[1] Chapman RA, Suleiman MS, Earm YE. Taurine and the heart. Cardiovasc
Res 1993;27:358–63.
[2] Chesney RW. Taurine: its biological role and clinical implications. Adv
Pediatr 1985;32:1–42.
[3] Uchida S, Nakanishi T, Kwon HM, Preston AS, Handler JS. Taurine
behaves as an osmolyte in Madin–Darby canine kidney cells. Protection by
polarized, regulated transport of taurine. J Clin Invest 1991;88:656–62.
[4] Uozumi Y, Ito T, Hoshino Y, Mohri T, Maeda M, Takahashi K, et al.
Myogenic differentiation induces taurine transporter in association with
taurine-mediated cytoprotection in skeletal muscles. Biochem J
2006;394:699–706.
[5] Takahashi K, Azuma M, Yamada T, Ohyabu Y, Schaffer SW, Azuma J.
Taurine transporter in primary cultured neonatal rat heart cells: a comparison between cardiac myocytes and nonmyocytes. Biochem Pharmacol
2003;65:1181–7.
[6] Ito T, Fujio Y, Hirata M, Takatani T, Matsuda T, Muraoka S, et al.
Expression of taurine transporter is regulated through the TonE (tonicityresponsive element)/TonEBP (TonE-binding protein) pathway and contributes to cytoprotection in HepG2 cells. Biochem J 2004;382:177–82.
[7] Han X, Patters AB, Chesney RW. Transcriptional repression of taurine
transporter gene (TauT) by p53 in renal cells. J Biol Chem 2002;277: 39266–73.
[8] Azuma J, Hasegawa H, Sawamura A, Awata N, Ogura K, Harada H, et al.
Therapy of congestive heart failure with orally administered taurine. Clin
Ther 1983;5:398–408.
[9] Azuma J, Sawamura A, Awata N, Ohta H, Hamaguchi T, Harada H, et al.
Therapeutic effect of taurine in congestive heart failure: a double-blind
crossover trial. Clin Cardiol 1985;8:276–82.
[10] Takihara K, Azuma J, Awata N, Ohta H, Hamaguchi T, Sawamura A, et al.
Beneficial effect of taurine in rabbits with chronic congestive heart failure.
Am Heart J 1986;112:1278–84.
[11] Oudit GY, Trivieri MG, Khaper N, Husain T, Wilson GJ, Liu P, et al. Taurine
supplementation reduces oxidative stress and improves cardiovascular
function in an iron-overload murine model. Circulation 2004;109: 1877–85.
[12] Huxtable RJ. Physiological actions of taurine. Physiol Rev 1992;72:
101–63.
[13] Schaffer S, Takahashi K, Azuma J. Role of osmoregulation in the actions
of taurine. Amino Acids 2000;19:527–46.
[14] Satoh H, Sperelakis N. Review of some actions of taurine on ion channels
of cardiac muscle cells and others. Gen Pharmacol 1998;30:451–63.
[15] Takatani T, Takahashi K, Uozumi Y, Matsuda T, Ito T, Schaffer SW, et al.
Taurine prevents the ischemia-induced apoptosis in cultured neonatal rat
cardiomyocytes through Akt/caspase-9 pathway. Biochem Biophys Res
Commun 2004;316:484–9.
[16] Takatani T, Takahashi K, Uozumi Y, Shikata E, Yamamoto Y, Ito T, et al.
Taurine inhibits apoptosis by preventing formation of the Apaf-1/caspase-9
apoptosome. Am J Physiol Cell Physiol 2004;287:C949–53.
[17] Huxtable R, Bressler R. Taurine concentrations in congestive heart failure.
Science 1974;184:1187–8.
[18] Pion PD, Kittleson MD, Rogers QR, Morris JG. Myocardial failure in cats
associated with low plasma taurine: a reversible cardiomyopathy. Science
1987;237:764–8.
[19] Moise NS, Pacioretty LM, Kallfelz FA, Stipanuk MH, King JM, Gilmour
Jr RF. Dietary taurine deficiency and dilated cardiomyopathy in the fox.
Am Heart J 1991;121:541–7.
[20] Schuller-Levis GB, Park E. Taurine: new implications for an old amino
acid. FEMS Microbiol Lett 2003;226:195–202.
[21] Wen GY, Sturman JA, Wisniewski HM, Lidsky AA, Cornwell AC, Hayes
KC. Tapetum disorganization in taurine-depleted cats. Invest Ophthalmol
Vis Sci 1979;18:1200–6.
[22] Hamaguchi T, Azuma J, Schaffer S. Interaction of taurine with methionine:
inhibition of myocardial phospholipid methyltransferase. J Cardiovasc
Pharmacol 1991;18:224–30.
[23] Lake N. Loss of cardiac myofibrils: mechanism of contractile deficits
induced by taurine deficiency. Am J Physiol 1993;264:H1323–6.
[24] Schaffer SW, Solodushko V, Kakhniashvili D. Beneficial effect of taurine
depletion on osmotic sodium and calcium loading during chemical
hypoxia. Am J Physiol Cell Physiol 2002;282:C1113–20.
[25] Tybulewicz VL, Crawford CE, Jackson PK, Bronson RT, Mulligan RC.
Neonatal lethality and lymphopenia in mice with a homozygous disruption
of the c-abl proto-oncogene. Cell 1991;65:1153–63.
[26] Ito T, Asakura K, Tougou K, Fukuda T, Kubota R, Nonen S, et al.
Regulation of cytochrome P450 2E1 under hypertonic environment
through TonEBP in human hepatocytes. Mol Pharmacol 2007;72:173–81.
[27] Lippincott SE, Friedman AL, Siegel FL, Pityer RM, Chesney RW. HPLC
analysis of the phenylisothiocyanate (PITC) derivatives of taurine from
physiologic samples. J Am Coll Nutr 1988;7:491–7.
[28] Larsson NG, Wang J, Wilhelmsson H, Oldfors A, Rustin P, Lewandoski M,
et al. Mitochondrial transcription factor A is necessary for mtDNA
maintenance and embryogenesis in mice. Nat Genet 1998;18:231–6.
[29] Nebigil CG, Etienne N, Messaddeq N, Maroteaux L. Serotonin is a novel
survival factor of cardiomyocytes: mitochondria as a target of 5-HT2B
receptor signaling. FASEB J 2003;17:1373–5.
[30] Nishiya D, Omura T, Shimada K, Matsumoto R, Kusuyama T, Enomoto S,
et al. Effects of erythropoietin on cardiac remodeling after myocardial
infarction. J Pharmacol Sci 2006;101:31–9.
[31] Yoshiyama M, Takeuchi K, Omura T, Kim S, Yamagishi H, Toda I, et al.
Effects of candesartan and cilazapril on rats with myocardial infarction
assessed by echocardiography. Hypertension 1999;33:961–8.
[32] Bao X, Lu CM, Liu F, Gu Y, Dalton ND, Zhu BQ, et al. Epinephrine is
required for normal cardiovascular responses to stress in the phenylethanolamine N-methyltransferase knockout mouse. Circulation 2007;116:
1024–31.
[33] Ito T, Fujio Y, Takahashi K, Azuma J. Degradation of NFAT5, a transcriptional
regulator of osmotic stress-related genes, is a critical event for doxorubicininduced cytotoxicity in cardiac myocytes. J Biol Chem 2007;282:1152–60.
[34] Tanaka M, Nakamura F, Mizokawa S, Matsumura A, Nozaki S, Watanabe
Y. Establishment and assessment of a rat model of fatigue. Neurosci Lett
2003;352:159–62.
[35] Woo SK, Lee SD, Na KY, Park WK, Kwon HM. TonEBP/NFAT5
stimulates transcription of HSP70 in response to hypertonicity. Mol Cell
Biol 2002;22:5753–60.
[36] Trama J, Go WY, Ho SN. The osmoprotective function of the NFAT5
transcription factor in T cell development and activation. J Immunol
2002;169:5477–88.
[37] Rivard CJ, Brown LM, Almeida NE, Maunsbach AB, PihakaskiMaunsbach K, Andres-Hernando A, et al. Expression of the calciumbinding protein S100A4 is markedly up-regulated by osmotic stress and is
involved in the renal osmoadaptive response. J Biol Chem
2007;282:6644–52.
[38] Olivetti G, Capasso JM, Sonnenblick EH, Anversa P. Side-to-side slippage
of myocytes participates in ventricular wall remodeling acutely after
myocardial infarction in rats. Circ Res 1990;67:23–34.
[39] Campbell SE, Korecky B, Rakusan K. Remodeling of myocyte dimensions
in hypertrophic and atrophic rat hearts. Circ Res 1991;68:984–96.
T. Ito et al. / Journal of Molecular and Cellular Cardiology 44 (2008) 927–937
[40] Welsh DC, Dipla K, McNulty PH, Mu A, Ojamaa KM, Klein I, et al.
Preserved contractile function despite atrophic remodeling in
unloaded rat hearts. Am J Physiol Heart Circ Physiol 2001;281:
H1131–6.
[41] Lisy O, Redfield MM, Jovanovic S, Jougasaki M, Jovanovic A, Leskinen
H, et al. Mechanical unloading versus neurohumoral stimulation on myocardial structure and endocrine function In vivo. Circulation 2000;102:
338–43.
[42] Radke MH, Peng J, Wu Y, McNabb M, Nelson OL, Granzier H, et al.
Targeted deletion of titin N2B region leads to diastolic dysfunction and
cardiac atrophy. Proc Natl Acad Sci U S A 2007;104:3444–9.
[43] Burg MB, Kwon ED, Kultz D. Regulation of gene expression by
hypertonicity. Annu Rev Physiol 1997;59:437–55.
[44] Schaffer SW, Ballard-Croft C, Azuma J, Takahashi K, Kakhniashvili DG,
Jenkins TE. Shape and size changes induced by taurine depletion in
neonatal cardiomyocytes. Amino Acids 1998;15:135–42.
[45] Lopez-Rodriguez C, Antos CL, Shelton JM, Richardson JA, Lin F,
Novobrantseva TI, et al. Loss of NFAT5 results in renal atrophy and lack of
tonicity-responsive gene expression. Proc Natl Acad Sci U S A 2004;101:
2392–7.
[46] Marin-Garcia J, Goldenthal MJ, Moe GW. Mitochondrial pathology in
cardiac failure. Cardiovasc Res 2001;49:17–26.
937
[47] Russell LK, Finck BN, Kelly DP. Mouse models of mitochondrial
dysfunction and heart failure. J Mol Cell Cardiol 2005;38:81–91.
[48] Palmi M, Youmbi GT, Sgaragli G, Meini A, Benocci A, Fusi F, et al. The
mitochondrial permeability transition and taurine. Adv Exp Med Biol
2000;483:87–96.
[49] Suzuki T, Wada T, Saigo K, Watanabe K. Taurine as a constituent of
mitochondrial tRNAs: new insights into the functions of taurine and
human mitochondrial diseases. EMBO J 2002;21:6581–9.
[50] Suzuki T, Wada T, Saigo K, Watanabe K. Novel taurine-containing uridine
derivatives and mitochondrial human diseases. Nucleic Acids Res Suppl
2001:257–8.
[51] Warskulat U, Flogel U, Jacoby C, Hartwig HG, Thewissen M, Merx MW,
et al. Taurine transporter knockout depletes muscle taurine levels and
results in severe skeletal muscle impairment but leaves cardiac function
uncompromised. FASEB J 2004;18:577–9.
[52] Deschepper CF, Olson JL, Otis M, Gallo-Payet N. Characterization of
blood pressure and morphological traits in cardiovascular-related organs in
13 different inbred mouse strains. J Appl Physiol 2004;97:369–76.
[53] Barrick CJ, Rojas M, Schoonhoven R, Smyth SS, Threadgill DW. Cardiac
response to pressure overload in 129S1/SvImJ and C57BL/6J mice: temporaland background-dependent development of concentric left ventricular
hypertrophy. Am J Physiol Heart Circ Physiol 2007;292:H2119–30.