A three amino acid peptide, Gly-Pro-Arg, protects and rescues cell death induced by amyloid β-peptide☆
Introduction
Alzheimer's disease (AD) is a progressive neurodegenerative disorder with dementia as a major clinical symptom. The brain of AD patients is characterized by deposition of amyloid β-peptide (Aβ), a 39- to 43-amino acid peptide (Yankner, 1996). Although Aβ has been implicated as a cause of neuronal degeneration (Mattson, 1997), the precise mechanism of its neurotoxic action remains unclear. Post-mortem AD brains have shown a large number of neurons with fragmented DNA Dragunow et al., 1995, Lassmann et al., 1995, Masliah et al., 1998. Although not all neurons showed other morphological features of apoptosis, for example, cell surface blebbing, cell shrinkage, and nuclear condensation Lassmann et al., 1995, Lucassen et al., 1987, Stadelmann et al., 1998, neurons did show altered expression of apoptosis-related proteins, such as caspase-3, caspase-8, caspase-9 Lu et al., 2000, Rohn et al., 2001, Par-4, bak, bad, bax, bcl-2, p53, CPP32, and fas Guo et al., 1998, Kitamura et al., 1997, Kitamura et al., 1998, Nishimura et al., 1995, Shinohara et al., 1999, Stadelmann et al., 1999, Su et al., 1996, Su et al., 1997. What causes neuronal apoptosis in the brain of AD patients has not been determined; however, Aβ induces apoptosis in cultured neurons Gschwind and Huber, 1995, Li et al., 1996 and inhibition of p53, a key regulatory molecule of apoptosis, was shown to prevent Aβ-induced neuronal death (Culmsee et al., 2001). These in vivo and in vitro observations suggest that Aβ induces apoptosis in the brain of AD patients.
Studies of the mechanisms whereby Aβ induces neuronal degeneration have led to the identification of compounds capable of protecting neurons. They include peptide aggregation-blocking agents Permanne et al., 2002, Soto et al., 1998, neurotrophic factors (Guo and Mattson, 2000), antioxidants Behl et al., 1992, Goodman and Mattson, 1994, Kumar et al., 1994, drugs that affect calcium signals (Weiss et al., 1994), and estrogens (Goodman and Mattson, 1994). Several proteins and peptides were also reported to prevent Aβ-induced neuronal degeneration. They include tachykinin neuropeptides (substance P) (Yankner et al., 1990), vasoactive intestinal peptide (VIP) (Gozes et al., 1999), and activity-dependent neuroprotective protein (ADNP; i.e., VIP fragment of either 8 or 14 amino acids) Brenneman and Gozes, 1996, Zemylak et al., 2000, and IGF-1 (Doré et al., 1997). Interestingly, peptide Gly-Pro-Glu (GPE), a product of IGF-I, was shown to protect against NMDA-induced toxicity (Saura et al., 1999), hypoxic–ischemic injury (Sizonenko et al., 2001), and 6-OHDA-induced nigral lesions (Guan et al., 2000). In this study, we examined the neuroprotective action of small peptides, GPE, Gly-Glu (GE), Gly-Pro-Asp (GPD), and Gly-Pro-Arg (GPR) against Aβ toxicity. GPR, but not other peptides, effectively prevented neuronal degeneration by preventing Aβ-induced activation of caspase-3 and increased expression of p53, suggesting that GPR prevents Aβ-mediated apoptosis.
Section snippets
Cell culture and Aβ peptide
Hippocampal neurons were obtained from 17E Sprague–Dawley rats. Cell suspension with a density of 1 to 3 × 105 cells/ml in neurobasal media with B27 supplement (Life Technologies) was plated in 24-well tissue culture plates coated with polyethylenimine (PEI). Cultures were incubated at 37°C in 5% CO2-humidified atmosphere. All pups from each pregnant rat are used as an experimental unit (n). Aβ25–35 and Aβ1–40 (Bachem) were dissolved in deionized water at a concentration of 6 mg/ml and stored
GPR prevents Aβ-mediated increase in LDH
Neuronal death, cultured rat hippocampal neurons were exposed to Aβ25–35 with or without peptides. Aβ-induced increase in LDH levels was about 70% greater than that in the control culture (Fig. 1). Three peptides, GE (50 μM), GPD (50 μM), and GPE (50 and 100 μM) did not prevent Aβ-mediated LDH increase (data not shown). However, GPR (10 and 100 μM) was effective in preventing Aβ-mediated increase in LDH levels [Fig. 1; F(2,36) = 4.5, P = 0.02]. When GPR was added together with Aβ or even added
Discussion
It was shown that 75% of hippocampal neurons die after 6-day incubation with Aβ25–35 (25–50 μM) (Lockhart et al., 1994). The present study demonstrated that Aβ-induced neurotoxicity can be protected by GPR. Most importantly, LDH and MTT assays suggest that GPR effectively rescues neurons that have been pre-exposed to aggregated Aβ for up to 2 days. Similar findings have been reported for IGF-I (Doré et al., 1997). Furthermore, GPE, a peptide structurally identical to the N-terminal tripeptide
Acknowledgements
The authors thank Cathy Martens for her technical support.
References (56)
- et al.
Hypoxia-induced apoptosis: effect of hypoxic severity and role of p53 in neuronal cell death
Brain Res.
(1998) - et al.
Vitamin E protects nerve cells from amyloid toxicity
Biochem. Biophys. Res. Commun.
(1992) - et al.
Capase-3 activation and DNA fragmentation in primary hippocampal neurons following glutamate excitotoxicity
Mol. Brain Res.
(2001) - et al.
Correlates of p53- and Fas (CD95)-mediated apoptosis in Alzheimer's disease
J. Neurol. Sci.
(1997) - et al.
Staurosporin and K-252 compounds protect hippocampal neurons against amyloid β-peptide toxicity and oxidative injury
Brain Res.
(1994) - et al.
N-terminal tripeptide of IGF-1 (GPE) prevents the loss of TH positive neurons after 6-OHDA induced nigral lesion in rats
Brain Res.
(2000) - et al.
Neuronal apoptosis induced by β-amyloid is mediated by caspase-8
Neurobiol. Dis.
(1999) - et al.
Apoptosis and expression of p53 response proteins and cyclin D1 after cortical impact in rat brain
Brain Res.
(1999) - et al.
Changes of p53 in the brains of patients with Alzheimer's disease
Biochem. Biophys. Res. Commun.
(1997) - et al.
Alteration of proteins regulating apoptosis, Bcl-2, Bcl-x, Bax, Bak, Bad, ICH-1 and CPP32, in Alzheimer's disease
Brain Res.
(1998)
Beta-amyloid induces apoptosis in human-derived neurotypic SH-SY5Y cells
Brain Res.
β-Amyloid-induced activation of caspase-3 in primary culture of rat neurons
Mech. Ageing Dev.
Fas antigen expression in brains of patients with Alzheimer-type dementia
Brain Res.
Activation of caspase-8 in the Alzheimer's disease brain
Neurobiol. Dis.
Neuroprotective effects of the N-terminal tripeptide of IGF-1, glycine-proline-glutamate, in the immature rat brain after hypoxic–ischemic injury
Brain Res.
Activation of caspase-3 in single neurons and autophagic granules of granulovascular degeneration in Alzheimer's disease
Am. J. Physiol., Lung Cell. Mol. Physiol.
Toxic effect of a β-amyloid peptide (β22–35) on the hippocampal neuron and its prevention
Neurosci. Lett.
AMPA/kainate receptor-mediated damage to NADPH-diaphorase containing neurons is Ca2+ dependant
Neurosci. Lett.
Sublethal concentrations of prion peptide PrP106-126 or the amyloid beta peptide of Alzheimer's disease activates expression of proapoptotic markers in primary cortical neurons
Neurobiol. Dis.
Mechanisms of neuronal degeneration in Alzheimer's disease
Neuron
Multiple caspases are involved in beta-amyloid-induced neuronal apoptosis
J. Neurosci. Res.
Caspase-cleaved amyloid precursor protein in Alzheimer's disease
Brain Pathol.
A femtomolar-acting neuroprotective peptide
J. Clin. Invest.
Induction of cytochrome c-mediated apoptosis by amyloid β 25–35 requires functional mitochondria
Brain Res.
Bax-dependent caspase-3 activation is a key determinant in p53-induced apoptosis in neurons
J. Neurosci.
A synthetic inhibitor of p53 protects neurons against death induced by ischemic and excitotoxic insults, and amyloid β-peptide
J. Neurochem.
Insulin-like growth factor I protects and rescues hippocampal neurons against β-amyloid- and human amylin-induced toxicity
Proc. Natl. Acad. Sci. U. S. A.
In situ evidence for DNA fragmentation in Huntington's disease striatum, and Alzheimer's disease temporal lobes
NeuroReport
Cited by (25)
Organometallic Chemistry of Drugs Based on Iron
2022, Comprehensive Organometallic Chemistry IV: Volume 1-15Effects of food-derived bioactive peptides on cognitive deficits and memory decline in neurodegenerative diseases: A review
2021, Trends in Food Science and TechnologyDevelopment of glycine-α-methyl-proline-containing tripeptides with neuroprotective properties
2016, European Journal of Medicinal ChemistryCitation Excerpt :The role of acidic amino acids, such as Glu in P3 position of GPE, was extensively studied; biological studies showed that the replacement of Glu with Arg resulted effective in protecting neurons from Aβ toxicity. In fact, such GPR peptide was noticeable for its aptitude of powerfully preventing Aβ-induced apoptosis in rat hippocampal neurons by interfering with caspase-3 and p53 expression [17]. Although, GPR is a promising compound for the management of AD, its use is hindered by the peptidic nature that makes it exposed to the proteases digestion.
Ferrocene tripeptide Gly-Pro-Arg conjugates: Synthesis and inhibitory effects on Alzheimer's Aβ<inf>1-42</inf> fibrillogenesis and Aβ-induced cytotoxicity in vitro
2013, Bioorganic and Medicinal ChemistryCitation Excerpt :As seen from Figure 1, when Aβ1–42 was incubated in the presence of GPR, the fluorescence intensity increases initially with incubation time, reaching its maximum at 4 days. However, a reduction in fluorescence intensity shows that GPR is able to inhibit the fibril formation of Aβ1–42, in agreement with the neuroprotective behavior of GPR in vivo.16 GPR–Fca exhibits similar ThT fluorescence pattern as that of GPR.
Selective and sensitive determination of peptides using 3,4-dihydroxyphenylacetic acid as a fluorogenic reagent
2012, Analytica Chimica ActaCitation Excerpt :For instance, opioid peptides such as enkephalins, dynorphins, and endorphins contain Tyr at their N-termini [7], and growth factors such as insulin-like growth factor-I (IGF-I) and liver cell growth factor, GHK, have Gly at their N-termini [8,9]. In addition, N-terminal Gly-containing small oligopeptides such as GPE and GPR have physiological roles, including neuro-protective activity [10–12]. Furthermore, various N-terminal Gly-containing small oligopeptides are derived from the enzymatic digestion of collagen with collagenase; these oligopeptides might be associated with pathophysiological disorders [13].
- ☆
This study was supported by a grant from the Alzheimer's Association (IIRG-00-2189).