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Alzheimer disease: An imbalance of proteolytic regulation?
r
1
Medical Hypotheses
Alzheimer Disease: An Imbalance of Proteolytic Regulation? M. A. SMITH and G. PERRY Division of Neuropathology, Institute of Pathology, Case Western Reserve University, 2085 Adelbert Road, Cleveland, OH 44106-4901, USA
Abstract - Alzheimer disease is typified by the accumulation of protein and neuronal death. We propose that neuronal death creates a proteolytic imbalance that generates the pathological lesions. Our hypothesis explains the morphology and topographic distribution of neuritic plaques and neurofibrillary tangles.
Proteolytic homeostasis is crucial for normal physiological function. A delicate balance governs proteolytic removal of denatured and excess or unwanted proteins and prevents autolysis of functional proteins. Proteolytic regulation relies on a multitude of cofactors creating a harmony of proenzyme (zymogen) activation and suppression by inhibitor molecules. We speculate that an imbalance in proteolytic regulation may be responsible for generating the pathological features of Alzheimer disease. Alzheimer disease is characterized by three separate, but presumably connected events. First, neuronal atrophy and death occurs in a regionally specific manner and leads to the cognitive impairment seen clinically (1). Second, intraneuronal and, following cell death, extracellular fibrillary protein aggregates termed neurofibrillary tangles (NFT). NFT are filamentous aggregates of the cytoskeletal proteins z, neurofilament protein and tropomyosin (2). Third, senile plaques (SP) are extracellular focal deposits of amyloid-P (AB) protein surrounded by a rim of dystrophic neurites. AB is a 39-43 residue polypeptide generated by proteolytic cleavage of the P-protein precursor (BPP) (3).
Efforts by a number of laboratories have concentrated on elucidating specific AP amyloidogenicgenerating pathways and individual proteases (4, 5). However, quantitation of AP in sera and CSF shows no difference between AD and control. Nevertheless, since SP are focal deposits of AP it seems possible that localized elevations in AP production could be responsible for the formation of SP. Indeed, the notion that a slow insidious build up of AP generates SP must be seriously questioned in light of the very rapid accumulation of AP in trauma patients(6). One mechanism in which AP levels could be focally increased is through a perturbation of PPP processing, i.e. an imbalance in proteolytic regulation. An underlying role for proteases in AD is indicated by an array of protease-related molecules found in association with NFT and SP. For example, ubiquitin is a protein that covalently tags denatured or unwanted proteins marking them for subsequent degradation(7). Also, a number of proteases, including cathepsins B, D and G (8), thromin (9) and attrypsin (IO); and protease inhibitors, including atantichymotrypsin (I l), at-antitrypsin (12), and antithrombin III (13); and several components of the
Date received 19 August 1993 Date accepted 23 September 1993
277
278
MEDICALHYPOTHESES
1:1 Pmteasdinhibttor
1:1 Pmteadlnhibitor
Complex
QJmMX
t
Neuronai Degeneration
~~j
1 ,..:.:,:,..: ,......._ :...,xg$,#@.&
I
f
I
knyioid Precursor Protein (BPP)
Protease inhibitor
Tau & Ubiquitin Accumulation
Fig. The seminal features of Alzheimer aberrant proteolytic regulation.
Aggregation& Degeneration of Neumnai Processes
disease (neuronal loss, neuroftbrillary tangles and
complement system (14) are found in association with Alzheimer pathology. Interestingly, secreted PPP, or protease nexin-II, can form inhibitory complexes with serine (15,16) and metailoproteinase (17) and the PPP Kunitz domain is a potent inhibitor of a number of serine proteases (18-20). Therefore, we suggest that PPP, protease nexin-II, along with other inhibitors and proteases are intimately involved in the regulation of proteolytic function in the central nervous system, and that a disturbance of this system could contribute towards the pathogenesis of AD. The senile plaque, a core of radially extending AP fibrils surrounded by dystrophic neurites (21) might arise through diffusion of ‘amyloidogenic factors’, including proteolytically-related molecules. Each component released following cell death’ has a specific diffusion distance. The diffusion constant will vary from region to region within the brain and is dependent upon size (Stoke’s radius), charge (isoelectric point, glycosylation/phosphorylation state), specific extracellular matrix interactions, concentration and the stability of the released components. Leakage of ceil contents from neurons including proteases, their inhibitors and substrates, would induce a number of biochemical and cellular events in a
senile plaques) could all be
generated by
radial diffusion pattern. Proteolysis would be confined to a focalized spherical area of damage. Proteases might also initiate focal increases in AP production. The excess Al3 protein could precipitate, perhaps by exceeding the critical concentration2 of polymerization induced by a specific nidus. Different regions within the brain could result in distinct responses, such that the hippocampus, amygdala and temporal and frontal cortices might be particularly vulnerable to this phenomenon, whereas as other areas within the brain would be less affected, i.e. paralleling the pathological distribution seen in AD (22). Inappropriate radial proteolysis would continue until either concentrations were diminished or ‘neutralized’ by inhibitor molecules, i.e. parallelling an inflammatory response where inhibitor molecule production at the periphery of the necrotic region limits autolytic damage. Inhibitors interact with proteases in a 1:l stochiometry, therefore, at a defined radius the spread of proteolysis is halted. Morphological data on senile plaque size distribution are consistent with diffusable factors, i.e. spherical with an exponential size distribution (23). Excess inhibitor molecules released peripheral to the ‘neutralization point’ would also create an imbalance in proteoiytic regulation with equiva-
IProteins released from degenerating or atrophied cells might be quantitatively far more important in this process. Such actively transcribing cells are able to secrete greater quantities of material into the extracellular millet& whereas cell death only results in spillage of cytoplasmic contents. *Released lysosomes would also focally decrease the pH which might further decrease the solubility of released AD (24, 25).
279
ALZHEIMER DISEASE: AN IMBALANCE OF PROEOLYTIC REGULATION?
lent detrimental cellular consequences. Protease inhibitors (presumably including some forms of PPP) can inititate the aggregation and accumulation of neuronal processes with concomitant z and ubiquitin increases (26). Therefore, protease inhibitors could affect intracellular events and induce the peripheral degeneration seen around SP. Further, neuritic degeneration would explain the topographic distribution of pathology since neuritic degeneration precipitates atrophy of the neuronal cell body. Proteolytic imbalance would then, like Alzheimer pathology, proceed along corticocortical connections (27). The schematic represented in the Figure accommodates many of the key elements associated with Alzheimer disease, including regionally specific cell death, AP deposition and NFI formation. Following initial cell damage a self perpetuating cycle of degeneration might arise. Indeed, normal neuronal senescence might explain the presence of senile plaques and NFT in almost every aged brain. The quantitative difference in pathology between normal aging and Alzheimer disease might be a reflection of the numbers of dying and senescent neurons.
IO. Smith M A, Kalaria
II.
12.
13.
14.
15.
16.
17.
18.
References 19. I.
2. 3.
4.
5.
6.
7.
8.
9.
Terry R D, Peck A, DeTeresa R, Schechter R, Horoupian D S. Some morphometric aspects of the brain in senile dementia of the Alzheimer type. Ann Neurol 1981; 10: 184-192. Perry G. Alzheimer disease. In: Smith B, Adelman G. eds. Neuroscience year. Boston: Birkatiser, 1992: 5-8. Tanzi R E, Gusella J F, Watkins P C et al. Amyloid beta protein gene: cDNA, mRNA distribution, and genetic linkage near the Alzheimer locus. Science 1987; 235: 880-884. Golde T E, Estus S, Younkin L H, Selkoe D J, Younkin S G. Processing of the amyloid protein precursor to potentially amyloidogenic derivatives. Science 1992; 255: 728-730. Haass C, Hung A Y, Schlossmacher M G, Teplow D B. Selkoe D J. beta-Amyloid peptide and a 3-kDa fragment are derived by distinct cellular mechanisms. J Biol Chem 1993; 268: 3021-3024. Roberts G W, Gentleman S M, Lynch A, Graham D I. beta A4 amyloid protein deposition in brain after head trauma. Lancet 1991; 338: 1422-1423. Perry G. Friedman R, Shaw G, Chau V. Ubiquitin is detected in neurofibrillary tangles and senile plaque neurites of Alzheimer disease brains. Proc Nat1 Acad Sci USA 1987; 84: 3033-3036. Cataldo A M, Nixon R A. Enzymatically active lysosomal proteases are associated with amyloid deposits in Alzheimer brain. Proc Nat1 Acad Sci USA 1990; 87: 3861-3865. Akiyama H, lkeda K, Kondo H, McGeer P L. Thrombin accumulation in brains of patients with Alzheimer’s disease. Neurosci Lett 1992: 146: 152-154.
20.
21. 22.
23.
24.
2s.
26.
27.
R N, Perry G. at-Trypsin immunoreactivity in Alzheimer disease. Biochem Biophys Res Commun 1993; 193: 579-584. Abraham C R, Selkoe D J, Potter H C. lmmunochemical identification of the serine protease inhibitor at-antichymotrypsin in the brain amyloid deposits of Alzheimer’s disease. Cell 1988; 52: 487-501. Gollin P A, Kahuia R N, Eikelenhoom P, Rozemuller A, Perry G. at-antitrypsin and at-antichymotrypsin are in the lesions of Alzheimer disease. Neuroreport 1992; 3: 201-203. Kalaria R N. Golde T, Kroon S N, Perry G. Serine protease inhibitor antithrombin Ill and its mRNA in the pathogenesis of Alzheimer’s disease. Am J Pathol 1993; (in press). Walker D G, McGeer P L. Complement gene expression in human brain: a comparison between normal and Alzheimer disease cases. Brain Res Mol Brain Res 1992; l-2: 109-116. Baker J B, Low D A, Simmer R L, Cunningham D D. Protease-nexin: a cellular component that links thrombin and plasminogen activator and mediates their binding to cells. Cell 1980; 21: 37-45. Knauer D J, Cunningham D D. Epidermal growth factor carrier protein binds to cells via a complex with released carried protein nexin. Proc Nat1 Acad Sci USA 1982; 79: 3210-3214. Miyazaki K, Hasegawa M, Funahashi K, Umeda M. A metalloproteinase inhibitor domain in Alzheimer amyloid protein precursor. Nature 1993; 362: 839-841. Kitaguchi N, Takahashi Y_ Tokushima Y, Shiojiri S, lto H. Novel precursor of Alzheimer’s disease amyloid protein shows protease inhibitory activity. Nature 1988; 33 I: 530532. Kitaguchi N, Takahashi Y, Oishi K et al. Enzyme specificity of proteinase inhibitor region in amyloid precursor protein of Alzheimer’s disease: different properties compared with protease nexin I. Biochim Biophys Acta 1990; 1038: JO5-113. Sinha S, Dovey H F, Seubert P et al. The protease inhibitory properties of Alzheimer’s beta-amyloid precursor protein. J Biol Chem 1990; 265: 8983-8985. Kidd M. Alzheimer’s disease - an electron microscopical study. Brain 1964; 87: 307-320. Ball M 1. Neuronal loss, neurotibrillary tangles and granulovacuolar degeneration in the hippocampus with ageing and dementia. Acta Neuropathol 1977; 37: 111-I 18. Kawai M, Kalaria R N, Harik S I, Perry G. The relationship of amyloid plaques to cerebral capillaries in Alzheimer’s disease. Am J Path01 1990; 137: 1435-1446. Pike C J, Walencewicz A J, Glabe C G, Cotman C W. In vitro aging of beta-amyloid protein causes peptide aggregation and neurotoxicity. Brain Res 1991; 563: 311-314. Barrow C J, Zagorski M G. Solution structures of beta peptide and its constituent fragments: relation to amyloid deposition. Science 1991; 253: 179-182. Ivy G 0, Kitani K, lhara Y. Anomalous accumulation of tau and uhiquitin immunoreativities in rat brain caused by protease inhibition and by normal aging: a clue to PHF pathogenesis? Brain Res 1989; 498: 360-365. Pearson R C, Esiri M M, Hioms R W, Wilcock G K. Powell T P. Anatomical correlates of the distribution of the pathological changes in the neocortex in Alzheimer disease. Proc Nat1 Acad Sci USA 1985; 82: 45314534.
1
Medical Hypotheses
Alzheimer Disease: An Imbalance of Proteolytic Regulation? M. A. SMITH and G. PERRY Division of Neuropathology, Institute of Pathology, Case Western Reserve University, 2085 Adelbert Road, Cleveland, OH 44106-4901, USA
Abstract - Alzheimer disease is typified by the accumulation of protein and neuronal death. We propose that neuronal death creates a proteolytic imbalance that generates the pathological lesions. Our hypothesis explains the morphology and topographic distribution of neuritic plaques and neurofibrillary tangles.
Proteolytic homeostasis is crucial for normal physiological function. A delicate balance governs proteolytic removal of denatured and excess or unwanted proteins and prevents autolysis of functional proteins. Proteolytic regulation relies on a multitude of cofactors creating a harmony of proenzyme (zymogen) activation and suppression by inhibitor molecules. We speculate that an imbalance in proteolytic regulation may be responsible for generating the pathological features of Alzheimer disease. Alzheimer disease is characterized by three separate, but presumably connected events. First, neuronal atrophy and death occurs in a regionally specific manner and leads to the cognitive impairment seen clinically (1). Second, intraneuronal and, following cell death, extracellular fibrillary protein aggregates termed neurofibrillary tangles (NFT). NFT are filamentous aggregates of the cytoskeletal proteins z, neurofilament protein and tropomyosin (2). Third, senile plaques (SP) are extracellular focal deposits of amyloid-P (AB) protein surrounded by a rim of dystrophic neurites. AB is a 39-43 residue polypeptide generated by proteolytic cleavage of the P-protein precursor (BPP) (3).
Efforts by a number of laboratories have concentrated on elucidating specific AP amyloidogenicgenerating pathways and individual proteases (4, 5). However, quantitation of AP in sera and CSF shows no difference between AD and control. Nevertheless, since SP are focal deposits of AP it seems possible that localized elevations in AP production could be responsible for the formation of SP. Indeed, the notion that a slow insidious build up of AP generates SP must be seriously questioned in light of the very rapid accumulation of AP in trauma patients(6). One mechanism in which AP levels could be focally increased is through a perturbation of PPP processing, i.e. an imbalance in proteolytic regulation. An underlying role for proteases in AD is indicated by an array of protease-related molecules found in association with NFT and SP. For example, ubiquitin is a protein that covalently tags denatured or unwanted proteins marking them for subsequent degradation(7). Also, a number of proteases, including cathepsins B, D and G (8), thromin (9) and attrypsin (IO); and protease inhibitors, including atantichymotrypsin (I l), at-antitrypsin (12), and antithrombin III (13); and several components of the
Date received 19 August 1993 Date accepted 23 September 1993
277
278
MEDICALHYPOTHESES
1:1 Pmteasdinhibttor
1:1 Pmteadlnhibitor
Complex
QJmMX
t
Neuronai Degeneration
~~j
1 ,..:.:,:,..: ,......._ :...,xg$,#@.&
I
f
I
knyioid Precursor Protein (BPP)
Protease inhibitor
Tau & Ubiquitin Accumulation
Fig. The seminal features of Alzheimer aberrant proteolytic regulation.
Aggregation& Degeneration of Neumnai Processes
disease (neuronal loss, neuroftbrillary tangles and
complement system (14) are found in association with Alzheimer pathology. Interestingly, secreted PPP, or protease nexin-II, can form inhibitory complexes with serine (15,16) and metailoproteinase (17) and the PPP Kunitz domain is a potent inhibitor of a number of serine proteases (18-20). Therefore, we suggest that PPP, protease nexin-II, along with other inhibitors and proteases are intimately involved in the regulation of proteolytic function in the central nervous system, and that a disturbance of this system could contribute towards the pathogenesis of AD. The senile plaque, a core of radially extending AP fibrils surrounded by dystrophic neurites (21) might arise through diffusion of ‘amyloidogenic factors’, including proteolytically-related molecules. Each component released following cell death’ has a specific diffusion distance. The diffusion constant will vary from region to region within the brain and is dependent upon size (Stoke’s radius), charge (isoelectric point, glycosylation/phosphorylation state), specific extracellular matrix interactions, concentration and the stability of the released components. Leakage of ceil contents from neurons including proteases, their inhibitors and substrates, would induce a number of biochemical and cellular events in a
senile plaques) could all be
generated by
radial diffusion pattern. Proteolysis would be confined to a focalized spherical area of damage. Proteases might also initiate focal increases in AP production. The excess Al3 protein could precipitate, perhaps by exceeding the critical concentration2 of polymerization induced by a specific nidus. Different regions within the brain could result in distinct responses, such that the hippocampus, amygdala and temporal and frontal cortices might be particularly vulnerable to this phenomenon, whereas as other areas within the brain would be less affected, i.e. paralleling the pathological distribution seen in AD (22). Inappropriate radial proteolysis would continue until either concentrations were diminished or ‘neutralized’ by inhibitor molecules, i.e. parallelling an inflammatory response where inhibitor molecule production at the periphery of the necrotic region limits autolytic damage. Inhibitors interact with proteases in a 1:l stochiometry, therefore, at a defined radius the spread of proteolysis is halted. Morphological data on senile plaque size distribution are consistent with diffusable factors, i.e. spherical with an exponential size distribution (23). Excess inhibitor molecules released peripheral to the ‘neutralization point’ would also create an imbalance in proteoiytic regulation with equiva-
IProteins released from degenerating or atrophied cells might be quantitatively far more important in this process. Such actively transcribing cells are able to secrete greater quantities of material into the extracellular millet& whereas cell death only results in spillage of cytoplasmic contents. *Released lysosomes would also focally decrease the pH which might further decrease the solubility of released AD (24, 25).
279
ALZHEIMER DISEASE: AN IMBALANCE OF PROEOLYTIC REGULATION?
lent detrimental cellular consequences. Protease inhibitors (presumably including some forms of PPP) can inititate the aggregation and accumulation of neuronal processes with concomitant z and ubiquitin increases (26). Therefore, protease inhibitors could affect intracellular events and induce the peripheral degeneration seen around SP. Further, neuritic degeneration would explain the topographic distribution of pathology since neuritic degeneration precipitates atrophy of the neuronal cell body. Proteolytic imbalance would then, like Alzheimer pathology, proceed along corticocortical connections (27). The schematic represented in the Figure accommodates many of the key elements associated with Alzheimer disease, including regionally specific cell death, AP deposition and NFI formation. Following initial cell damage a self perpetuating cycle of degeneration might arise. Indeed, normal neuronal senescence might explain the presence of senile plaques and NFT in almost every aged brain. The quantitative difference in pathology between normal aging and Alzheimer disease might be a reflection of the numbers of dying and senescent neurons.
IO. Smith M A, Kalaria
II.
12.
13.
14.
15.
16.
17.
18.
References 19. I.
2. 3.
4.
5.
6.
7.
8.
9.
Terry R D, Peck A, DeTeresa R, Schechter R, Horoupian D S. Some morphometric aspects of the brain in senile dementia of the Alzheimer type. Ann Neurol 1981; 10: 184-192. Perry G. Alzheimer disease. In: Smith B, Adelman G. eds. Neuroscience year. Boston: Birkatiser, 1992: 5-8. Tanzi R E, Gusella J F, Watkins P C et al. Amyloid beta protein gene: cDNA, mRNA distribution, and genetic linkage near the Alzheimer locus. Science 1987; 235: 880-884. Golde T E, Estus S, Younkin L H, Selkoe D J, Younkin S G. Processing of the amyloid protein precursor to potentially amyloidogenic derivatives. Science 1992; 255: 728-730. Haass C, Hung A Y, Schlossmacher M G, Teplow D B. Selkoe D J. beta-Amyloid peptide and a 3-kDa fragment are derived by distinct cellular mechanisms. J Biol Chem 1993; 268: 3021-3024. Roberts G W, Gentleman S M, Lynch A, Graham D I. beta A4 amyloid protein deposition in brain after head trauma. Lancet 1991; 338: 1422-1423. Perry G. Friedman R, Shaw G, Chau V. Ubiquitin is detected in neurofibrillary tangles and senile plaque neurites of Alzheimer disease brains. Proc Nat1 Acad Sci USA 1987; 84: 3033-3036. Cataldo A M, Nixon R A. Enzymatically active lysosomal proteases are associated with amyloid deposits in Alzheimer brain. Proc Nat1 Acad Sci USA 1990; 87: 3861-3865. Akiyama H, lkeda K, Kondo H, McGeer P L. Thrombin accumulation in brains of patients with Alzheimer’s disease. Neurosci Lett 1992: 146: 152-154.
20.
21. 22.
23.
24.
2s.
26.
27.
R N, Perry G. at-Trypsin immunoreactivity in Alzheimer disease. Biochem Biophys Res Commun 1993; 193: 579-584. Abraham C R, Selkoe D J, Potter H C. lmmunochemical identification of the serine protease inhibitor at-antichymotrypsin in the brain amyloid deposits of Alzheimer’s disease. Cell 1988; 52: 487-501. Gollin P A, Kahuia R N, Eikelenhoom P, Rozemuller A, Perry G. at-antitrypsin and at-antichymotrypsin are in the lesions of Alzheimer disease. Neuroreport 1992; 3: 201-203. Kalaria R N. Golde T, Kroon S N, Perry G. Serine protease inhibitor antithrombin Ill and its mRNA in the pathogenesis of Alzheimer’s disease. Am J Pathol 1993; (in press). Walker D G, McGeer P L. Complement gene expression in human brain: a comparison between normal and Alzheimer disease cases. Brain Res Mol Brain Res 1992; l-2: 109-116. Baker J B, Low D A, Simmer R L, Cunningham D D. Protease-nexin: a cellular component that links thrombin and plasminogen activator and mediates their binding to cells. Cell 1980; 21: 37-45. Knauer D J, Cunningham D D. Epidermal growth factor carrier protein binds to cells via a complex with released carried protein nexin. Proc Nat1 Acad Sci USA 1982; 79: 3210-3214. Miyazaki K, Hasegawa M, Funahashi K, Umeda M. A metalloproteinase inhibitor domain in Alzheimer amyloid protein precursor. Nature 1993; 362: 839-841. Kitaguchi N, Takahashi Y_ Tokushima Y, Shiojiri S, lto H. Novel precursor of Alzheimer’s disease amyloid protein shows protease inhibitory activity. Nature 1988; 33 I: 530532. Kitaguchi N, Takahashi Y, Oishi K et al. Enzyme specificity of proteinase inhibitor region in amyloid precursor protein of Alzheimer’s disease: different properties compared with protease nexin I. Biochim Biophys Acta 1990; 1038: JO5-113. Sinha S, Dovey H F, Seubert P et al. The protease inhibitory properties of Alzheimer’s beta-amyloid precursor protein. J Biol Chem 1990; 265: 8983-8985. Kidd M. Alzheimer’s disease - an electron microscopical study. Brain 1964; 87: 307-320. Ball M 1. Neuronal loss, neurotibrillary tangles and granulovacuolar degeneration in the hippocampus with ageing and dementia. Acta Neuropathol 1977; 37: 111-I 18. Kawai M, Kalaria R N, Harik S I, Perry G. The relationship of amyloid plaques to cerebral capillaries in Alzheimer’s disease. Am J Path01 1990; 137: 1435-1446. Pike C J, Walencewicz A J, Glabe C G, Cotman C W. In vitro aging of beta-amyloid protein causes peptide aggregation and neurotoxicity. Brain Res 1991; 563: 311-314. Barrow C J, Zagorski M G. Solution structures of beta peptide and its constituent fragments: relation to amyloid deposition. Science 1991; 253: 179-182. Ivy G 0, Kitani K, lhara Y. Anomalous accumulation of tau and uhiquitin immunoreativities in rat brain caused by protease inhibition and by normal aging: a clue to PHF pathogenesis? Brain Res 1989; 498: 360-365. Pearson R C, Esiri M M, Hioms R W, Wilcock G K. Powell T P. Anatomical correlates of the distribution of the pathological changes in the neocortex in Alzheimer disease. Proc Nat1 Acad Sci USA 1985; 82: 45314534.