CARDIOVASCULAR JOURNAL OF AFRICA • Vol 21, No 1, January/February 2010
AFRICA
41
subcellular localisation and initiates different cell death-signal-
ling processes in the endoplasmic reticulum and mitochondria
in a rat dopaminergic neuronal cell line.
18
Bnip3 promoter was
identified to contain the binding sites for HIF-1
α
.
9,19
Elevated
levels of Bnip3 expression have been observed
in vivo
in an
animal model of chronic heart failure
20
and have been found to
be expressed in considerable amounts in the adult myocardium.
21
Moreover, Bnip3 expression has been reported to be upregulated
in neonatal rat ventricular myocytes subjected to hypoxia, result-
ing in mitochondrial dysfunction and subsequent cell death.
9,22
Bnip3 has been demonstrated to induce both necrotic and apop-
totic cell death.
22-24
However, investigations must be carried out to
determine whether HIF-1
α
or alternative cellular factors mecha-
nistically account for the induction of Bnip3 during hypoxic
injury of ventricular myocytes.
With western blot analysis, we observed an increase in the
expression level of Bnip3 in cells exposed to hypoxic conditions.
When YC-1 was added to cells exposed to hypoxic conditions,
there was a decrease in the expression level of HIF-1
α
and a
simultaneous decrease in the that of Bnip3. The concurrent
changes in the expression levels of two pro-apoptotic proteins of
the Bcl-2 family, Bax and Bad were unclear.
Due to a shortage of funds, we have not further tested the role
of Bnip3 expression in HIF-1
α
-mediated apoptosis in cultured
primary neonatal rat ventricular myocytes. The present results
could suggest a possible link between hypoxia, HIF-1
α
signalling
and the activation of genes encoding Bnip3, which are involved
in apoptosis in primary neonatal rat ventricular myocytes.
Conclusions
We found that exposure to hypoxic conditions for 24 hours
increased the levels of HIF-1
α
expression in a manner that
was dependent on the degree of hypoxia. The apoptotic rate in
ventricular myocytes was also increased in hypoxia-exposed cells
in a manner dependent on the degree of hypoxia. The expression
level of the pro-apoptotic protein Bnip3 was upregulated under
hypoxic conditions. When HIF-1
α
activity was suppressed by
treatment with YC-1, the hypoxia-induced apoptosis and Bnip3
expression were blocked.
We concluded that HIF-1
α
mediated apoptosis in primary
neonatal rat ventricular myocytes cultured under acute hypoxic
conditions, and that the pro-apoptotic protein Bnip3 may be one
of the key molecules involved in this effect of HIF-1
α
. Targeting
HIF-1
α
may represent a new strategy for reducing ventricular
myocyte apoptosis induced by hypoxia.
We thank Drs WuYan and Shan for valuable suggestions and technical assist-
ance. This work was supported by a grant from the National Natural Science
Foundation of China (No 30500212).
References
Wang GL, Semenza GL. Purification and characterization of hypoxia-
1.
inducible factor 1. J.
Biol Chem
1995;
270
: 1230–1237.
Wang GL, Jiang BH, Rue EA, Semenza GL. Hypoxia-inducible factor
2.
1 is a basic-helix–loop–helix-PAS heterodimer regulated by cellular O
2
tension.
Proc Natl Acad Sci USA
1995;
92
: 5510–5514.
Reyes H, Reisz-Porszasz S, Hankinson O. Identification of the Ah recep-
3.
tor nuclear translocator protein (Arnt) as a component of the DNA bind-
ing form of the Ah receptor.
Science
1992;
256
: 1193–1195.
Semenza GL. Surviving ischemia: adaptive responses mediated by
4.
hypoxia-inducible factor 1.
J Clin Invest
2000;
106
: 809–812.
Wenger RH, Stiehl DP, Camenisch G. Integration of oxygen signaling at
5.
the consensus HRE. Sci STKE 2005; re12.
Semenza GL. Development of novel therapeutic strategies that target
6.
HIF-1.
Expert Opin Ther Targets
2006;
10
: 267–280.
Bruick RK. Expression of the gene encoding the proapoptotic Nip3
7.
protein is induced by hypoxia.
Proc Natl Acad Sci USA
2000;
97
:
9082–9087.
Merighi S, Benini A, Mirandola P, Gessi S, Varani K, Leung E,
8.
et al
.
Hypoxia inhibits paclitaxel-induced apoptosis through adenosine-medi-
ated phosphorylation of Bad in glioblastoma cells.
Molec Pharmacol
2007;
72
: 162–172.
Chen JK, Hu LJ, Wang J, Lamborn KR, Kong EL, Deen DF. Hypoxia-
9.
induced BAX overexpression and radiation killing of hypoxic glioblas-
toma cells.
Radiat Res
2005;
163
: 644–653.
Piret JP, Mottet D, Raes M, Michiels C. Is HIF-1alpha a pro- or an anti-
10.
apoptotic protein?
Biochem Pharmacol
2002;
64
: 889–892.
Kim HL, Yeo EJ, Chun YS, Park JW. A domain responsible for
11.
HIF-1alpha degradation by YC-1, a novel anticancer agent.
Int J Oncol
2006;
29
: 255–260.
Görlach A, Diebold I, Schini-Kerth VB, Berchner-Pfannschmidt U,
12.
Roth U, Brandes RP,
et al
. Thrombin activates the hypoxia-inducible
factor-1 signaling pathway in vascular smooth muscle cells: Role of the
p22(phox)-containing NADPH oxidase.
Circ Res
2001;
89
: 47–54.
Richard DE, Berra E, Pouyssegur J. Nonhypoxic pathway mediates the
13.
induction of hypoxia-inducible factor 1alpha in vascular smooth muscle
cells.
J Biol Chem
2000;
275
: 26765–26771.
Fukuda R, Hirota K, Fan F, Jung YD, Ellis LM, Semenza GL. Insulin-
14.
like growth factor 1 induces hypoxia-inducible factor 1-mediated vascu-
lar endothelial growth factor expression, which is dependent on MAP
kinase and phosphatidylinositol 3-kinase signaling in colon cancer cells.
J Biol Chem
2002;
277
: 38205–38211.
Stiehl DP, Jelkmann W, Wenger RH, Hellwig-Burgel T. Normoxic induc-
15.
tion of the hypoxia-inducible factor 1alpha by insulin and interleukin-
1beta involves the phosphatidylinositol 3-kinase pathway.
FEBS Lett
2002;
512
: 157–162.
Chun YS, Yeo EJ, Choi E, Teng CM, Bae JM, Kim MS,
16.
et al
. Inhibitory
effect of YC-1 on the hypoxic induction of erythropoietin and vascular
endothelial growth factor in Hep3B cells.
Biochem Pharmacol
2001;
61
: 947–954.
Kamat CD, Thorpe JE, Shenoy SS, Antonio C, Green DE, Warnke LA,
17.
Ihnat MA. A long-term ‘memory’ of HIF induction in response to chron-
ic mild decreased oxygen after oxygen normalization.
BMC Cardiovasc
Disord
2007;
18
(7): 4.
Zhang L, Li L, Liu H, Borowitz JL, Isome GE. BNIP3 mediates cell
18.
death by different pathways following localization to endoplasmic
reticulum and mitochondrion
FASEB J
2009;
17
: online.
Sowter HM, Ratcliffe PJ, Watson P, Greenberg AH, Harris AL. HIF-1-
19.
dependent regulation of hypoxic induction of the cell death factors
BNIP3 and NIX in human tumors.
Cancer Res
2001;
61
: 6669–6673.
Regula KM, Ens K, Kirshenbaum LA. Inducible expression of BNIP3
20.
provokes mitochondrial defects and hypoxia-mediated cell death of
ventricular myocytes.
Circ Res
2002;
91
: 226–231.
Hamacher-Brady A, Brady NR, Logue SE, Sayen MR, Jinno M,
21.
Kirshenbaum LA,
et al
. Response to myocardial ischemia/reperfusion
injury involves Bnip3 and autophagy.
Cell Death Differen
2007;
14
:
146–157.
Kubasiak LA, Hernandez OM, Bishopric NH, Webster KA. Hypoxia and
22.
acidosis activate cardiac myocyte death through the Bcl-2 family protein
BNIP3.
Proc Natl Acad Sci USA
2002;
99
: 12825–12830.
Velde CV, Cizeau J, Dubik D, Alimonti J, Brown T, Israels S,
23.
et al
. BNIP3
and genetic control of necrosis-like cell death through the mitochondrial
permeability transition pore.
Molec Cell Biol
2000;
20
: 5454–5468.
Graham RM, Frazier DP, Thompson JW, Haliko S, Li H, Wasserlauf BJ,
24.
et al
. A unique pathway of cardiac myocyte death caused by hypoxia-
acidosis.
J Exp Biol
2004;
207
: 3189–3200.
