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Wednesday 12 November 2003

amyloidosis, amyloid disorders


Definition: Amyloidosis is a group of diseases in which an abnormal protein, known as amyloid fibrils, make deposits in the extracellular space.

Amyloidosis is not a single disease but a term for diseases that share a common feature: the extracellular deposition of pathologic insoluble fibrillar proteins in organs and tissues.

Digital Cases

- HPC:125 : Salivary amyloidosis.
- JRC:6103 : Pulmonary amyloidosis.
- JRC:18909 : Thyroid medullary carcinoma with amyloid stroma.
- JRC:18927 : Thyroid amyloidosis.


- Renal amyloidosis

- thyroid medullary carcinoma (TMC)

- Hepatic amyloidosis


- renal amyloidosis
- pulmonary amyloidosis
- hepatic amyloidosis
- cutaneous amyloidosis
- digestive amyloidosis


- amyloid deposits


In the mid-19th century, Virchow adopted the botanical term "amyloid," meaning starch or cellulose, to describe abnormal extracellular material seen in the liver at autopsy. Subsequently, amyloid was found to stain with Congo red, appearing red microscopically in normal light but apple green when viewed in polarized light. Almost a century after Virchow’s observations, the fibrillar nature of amyloid was described with the use of electron microscopy and the characteristic beta-pleated–sheet configuration, now believed to be responsible for the typical staining properties, was identified.

The amyloidoses constitute a large group of diseases in which misfolding of extracellular protein has a prominent role. This dynamic process, which occurs in parallel with or as an alternative to physiologic folding, generates insoluble, toxic protein aggregates that are deposited in tissues in bundles of beta-sheet fibrillar protein. A beta-sheet consists of strands of polypeptides in zigzag formation.

The two most common forms of systemic amyloidosis are light-chain (AL) amyloidosis, with an incidence of approximately 1 case per 100,000 person-years in Western countries and reactive amyloidosis due to chronic inflammatory diseases (e.g., rheumatoid arthritis and chronic infections).

Hereditary amyloidosis is an ever-expanding group of disorders that pose difficult diagnostic problems.

The deposition of amyloid in brain tissue underlies Alzheimer disease which affects more than 12 million people worldwide. The approximately 1 million patients who are receiving dialysis worldwide are at risk for symptomatic amyloidosis.

According to the localization ot the deposits, two forms must be distingished: localized amyloidosis and diffuse amyloidosis.


The final pathway in the development of amyloidosis is the production of amyloid fibrils in the extracellular matrix. The process by which precursor proteins produce fibrils appears to be multifactorial and to differ among the various types of amyloid.

In AL amyloidosis, the demonstration that substitutions of particular amino acids at specific positions in the light-chain variable region occur at significantly higher frequencies than in nonamyloid immunoglobulins has led to the suggestion that these replacements destabilize light chains, increasing the likelihood of fibrillogenesis.

A similar situation may exist in ATTR amyloidosis. Normal transthyretin is a tetrameric protein with four identical subunits.

Inherently unstable variant monomers, produced by the substitution of amino acids, may allow the protein to precipitate when provoked by physical or chemical stimuli, such as the local surface pH, electric field, and hydration forces on cellular surfaces. Such stimuli may be responsible for the deposition of amyloid in both AL and ATTR amyloidosis and may explain the organ specificity of amyloid deposits.

An intriguing role for aging has been postulated in the formation of amyloid fibrils, since patients with variant transthyretin do not have clinically apparent disease until midlife, despite the lifelong presence of abnormal transthyretin.

Once the symptoms start, however, disease progression is usually rapid, suggesting an age-related trigger. Further evidence of such a trigger is that senile cardiac amyloidosis, caused by the deposition of fibrils derived from normal transthyretin, is exclusively a disease of elderly people.

A role for the precipitation of amyloid fibrils by "seeding" has been demonstrated in vivo, suggesting that amyloid begets amyloid. This again may partly explain why extensive amyloid deposition is sometimes concentrated in certain organs in an individual patient and does not progress in other parts of the body.

Studies of fibrillogenesis may help explain the aggressiveness of the disease with some amyloidogenic precursor proteins and the slow progression with others.

The three most common forms of amyloidosis — namely AL, ATTR, and amyloid protein A (AA) — differ entirely in their pathogenesis. Although there are overlapping characteristics, certain clinical features may suggest one form of the disease or another.

Amyloid deposits

These amyloid deposits are identified on the basis of their apple-green birefringence under a polarized light microscope after staining with Congo red and the presence of rigid, nonbranching fibrils 7.5 to 10 nm in diameter, on electron microscopy.

For example, lysozyme, transthyretin, apolipoprotein A-I and immunoglobulin light chain chains converge into a cross-beta super-secondary structure that has been well characterized by x-ray diffraction, with prototypical interstrand and intersheet distances of 4.7 and 10 to 13 Å, respectively.

The original conformation of the precursor protein can no longer be distinguished at this stage. Contiguous beta-sheet polypeptide chains constitute a protofilament. Several (four to six) protofilaments are wound around one another to form an amyloid fibril, with a distinct diameter of 7.5 to 10 nm visible on transmission electron microscopy (x100,000).

This ultrastructure of the fibril allows the regular intercalation of Congo red dye, conferring a diagnostic optical property to amyloid such as apple-green birefringence under polarized light microscopy.

To date, at least 21 different proteins have been recognized as causative agents of amyloid diseases. Despite having heterogeneous structures and functions, all these proteins can generate morphologically indistinguishable amyloid fibrils.

The generic fibrillar form of proteins can be regarded as a primordial structure dominated by hydrogen bonding between the amide and the carbonyl groups of the main chain, rather than by specific interactions of the side chains, which dictate the structure of functional globular proteins.

The essence of amyloidosis lies in the capacity of these proteins to acquire more than one conformation, a feature that has earned them the sobriquet of chameleon proteins.

The conversion of the structure of the native protein into a predominantly antiparallel beta-sheet secondary structure (in which the N- and C-terminals are oriented in opposite directions) is a pathologic process closely related to physiologic protein folding.

Extracellular protein misfolding

The folding of a newly synthesized polypeptide occurs in a rapid sequence of conformational modifications in the cytoplasm. According to the "folding energy landscape theory," the process follows a funnel-like pathway in which the conformational intermediates progressively merge into a final species.

In addition, at a minimum of energy similar to that reached by the native protein, the polypeptide can acquire an alternative and relatively stable "misfolded state," which is prone to aggregation. Once the folding process has been completed and the native protein secreted, many proteins are in dynamic equilibrium with a partially folded conformation, and in this state, they retrace the final part of the folding pathway, ultimately forming either a native or misfolded protein.

From a random coil conformation, the unfolded polypeptide enters a funnel-like pathway in which the conformational intermediates become progressively more organized as they merge, resulting in the most stable native state. In this state, there is a minimum of free energy, which results from the balance between the level of enthalpy, the internal energy that in folded protein is mainly determined by the kind and number of intramolecular bonds, and the level of conformational entropy, the level of randomness of the polypeptide in solution.

The protein chains undergo synthesis in the endoplasmic reticulum (ER), fold, pass through the cellular quality-control mechanism, and are secreted. In the extracellular environment, the mutants may change from a fully folded to a partially folded state and then retrace the final part of the folding pathway. Normal proteins are functionally active and are normally metabolized. The partially folded polypeptides can generate misfolded molecules, which have a high propensity to self-aggregate. Environmental conditions, chemical modifications, and common constituents favor the pathologic pathway. Oligomers, or protofibrils, may mediate cellular toxicity through a mechanism that activates apoptosis in cells of the target tissues. The green arrows indicate therapeutic targets.

In amyloid disease (amyloidosis), potentially pathogenic misfolded proteins can form in different ways. The protein may have an intrinsic propensity to assume a pathologic conformation, which becomes evident with aging (e.g., normal transthyretin in patients with senile systemic amyloidosis) or at persistently high concentrations in serum (e.g., beta2-microglobulin in patients undergoing long-term hemodialysis).

Another mechanism is the replacement of a single amino acid in the protein, as occurs in hereditary amyloidosis.

A third mechanism is proteolytic remodeling of the protein precursor, as in the case of -amyloid precursor protein (APP) in Alzheimer’s disease.

These mechanisms can act independently or in association with one another. In addition to the intrinsic amyloidogenic potential of the pathogenic protein, other factors may act synergistically in amyloid deposition. For example, the protein precursor must reach a critical local concentration to trigger fibril formation, a process enhanced by local environmental factors and by interactions with extracellular matrixes.


Although the molecular mechanisms involved are diverse and characteristic of each disease, almost all result in beta-linkages formed by hydrogen bonding between peptide loops and sheets.

Some larger, highly ordered proteins, such as the cystatins, can form intermolecular linkages by domain swapping, as with the loop-sheet linkages of the serpins. The result is the formation of polymers in which the individual molecules substantially retain their ordered structure.

In other proteins, such as beta2-microglobulin, lysozyme and transthyretin, linkage occurs by the remarkable realignment of peptide segments to give the sequential layering of beta-structures known as amyloid.

The characteristic features of amyloid are its staining with the dye Congo red and its birefringence to polarized light.

The determinant feature of amyloid is the formation of a specific pattern on X-ray diffraction, which has been interpreted as being due to the formation of layered arrays of extended beta-sheets.

Amyloid is most frequently observed as large tissue or pericellular deposits; in several systemic amyloidoses this end product interferes with organ function (for example, in the heart, lung or liver) and so is directly responsible for the disease pathology.

The dementias, however, arise specifically from the cumulative loss of neurons, and the pathology is likely to occur directly at the cellular level.

Protein instability

The property shared by these amyloidogenic variants and confirmed in studies of cystatin C (CST3), immunoglobulin light chains (IGL and IGK), and gelsolin (GSN) is a native conformation that is thermodynamically less stable than that of the normal counterpart.

A reduction in the stability of transthyretin (TTR) should make it easier for the tetramer to dissociate into monomers, whereas lysozyme (LYZ)mutations destabilize the tertiary structure and thus give rise to partially folded conformers (alternative spatial arrangements of the same polypeptide).

Monomers of transthyretin (TTR( and partially folded conformers of lysozyme (LYZ) have a strong propensity to self-aggregate and assemble into fibrils.

The role of protein stability in the formation of fibrils in vivo has been clarified by studies of natural, nonamyloidogenic variants of both transthyretin (TTR) and lysozyme (LYZ).

Destabilization is necessary but probably not sufficient to confer an amyloidogenic propensity on a protein; other structural features are required for the formation of fibrils. Recently, the role of charged residues in modulating the aggregation process, acting as structural gatekeepers by means of repulsive forces, has been highlighted.

In certain proteins, such as gelsolin (GSN), the partial unfolding caused by the mutation renders the protein susceptible to the attack of proteases, thus provoking the release of highly amyloidogenic polypeptides.

Amyloidogenic and normal counterparts are synthesized and secreted as native proteins, but the system of intracellular quality control appears to be incapable of recognizing and removing dangerous mutants.

Outside the cell, the amyloidogenic variants ultimately reach a state of equilibrium between fully folded and partially folded forms, but there is a much greater fluctuation in the concentrations of the two forms than would be expected.

All factors that perturb the three-dimensional structure — such as a low pH, oxidation, increased temperature, limited proteolysis, metal ions, and osmolytes — can shift the equilibrium toward the partially folded amyloidogenic state.

For example, urea at the concentrations present in the inner renal medulla enhances the formation of fibrils by reducing the time required for a nucleus to form, which in turn initiates rapid growth of the fibril.

In addition, local microenvironmental conditions affect the ultrastructural organization of protein deposits. For example, pH influences the processing of immunoglobulin light chains, causing them to form either fibrillar amyloid aggregates or amorphous aggregates characteristic of light-chain–deposition disease.

Common components of amyloid deposits, such as glycosaminoglycans and serum amyloid P (SAP) component, may exert identical effects by hastening the integration of a soluble polypeptide into a more stable fibril.


In certain amyloidoses, only a limited portion of the amyloid protein precursor forms the fibril.

The prototypical example is Alzheimer disease, in which the fibrils consist of proteolytic fragments of 39 to 43 residues derived from the 753-residue APP.

In lysozyme amyloidosis (LYZ), the full-length protein is detectable in natural fibrils.

There is a wide gradation of proteolytic remodeling of the protein precursor in all types of amyloidosis, but the remodeling of light chains in AL amyloidosis can be considered the archetype of the heterogeneity of this process.

Proteolysis is generally ascribed to extracellular, or pericellular, enzymes, such as those that cleave serum amyloid A.

However, in the cases involving gelsolin (GSN) or the amyloid ABri protein, the proteases act in the Golgi apparatus.

The structural flexibility of the target protein allows limited proteolysis and therefore the release of polypeptides that cannot display conformational plasticity in the constrained structure of the original protein.

A Conformational Disease

Amyloidosis properly belongs to the category of conformational diseases because pathologic protein aggregation is largely due to reduced folding stability and a strong propensity to acquire more than one conformation.

The risk of protein aggregation, which poses toxic threats to the cell, is minimized by protein sequences that confer the properties of high stability and fast folding kinetics, both of which minimize the concentration of easily aggregating, partially folded proteins.

Certain proteins, however, seem to require a high degree of structural disorder in their native states to fulfill their function.

For instance, the structural plasticity of the emerging class of "natively unfolded proteins" favors their interaction with ligands.

These proteins represent an intriguing gamble of molecular evolution in which the subtle border between risky self-aggregation and sophisticated function is easily crossed.

Amyloidogenic lipoproteins are examples. On the basis of its amino acid composition, lipid-free apolipoprotein A-I should behave as a natively unfolded protein; this state guarantees the plasticity of the protein, which partially unfolds when lipids are released and refolds when lipids are taken up.

These properties are particularly evident in the N-terminal domain of apolipoprotein A-I, the major constituent of apolipoprotein A-I amyloid fibrils.

Other lipoproteins, such as apolipoprotein A-II, apolipoprotein E, and serum amyloid A, also form amyloid or are implicated in amyloidogenesis and thus constitute a unique group of proteins.

They have structural similarities that confer the conformational plasticity necessary for their function and at the same time favor the formation of amyloid.

A Protein-Mediated Transmissible Disease?

Certain aspects of animal models of the amyloidosis caused by serum amyloid A and apolipoprotein A-II have introduced the possibility that amyloidosis is transmissible.

In mice, amyloid protein A amyloidosis (AA) is caused by an inflammatory reaction that results in overproduction of the acute-phase protein serum amyloid A.

Injection or oral administration of amyloid-enhancing factor, a crude homogenate of natural amyloid fibrils, accelerates the deposition of amyloid during the inflammatory process.

These findings are consistent with the capacity of fibrillar seeds to catalyze conformational changes in the soluble protein. The capacity of preformed fibrils to trigger fibrillogenesis has been demonstrated in vitro for amyloid (A) peptides, lysozyme, and beta2-microglobulin.

With the exception of prion diseases, there is no evidence that amyloidosis is transmissible in humans. However, the formation of amyloid can be accelerated by the presence of fibril nuclei in tissues.

A pertinent example is the patient with transthyretin variants who has cardiac involvement and receives a liver transplant. The transplanted liver minimizes the production of amyloidogenic transthyretin but does not halt the progression of amyloid deposition in the heart. Wild-type instead of mutant transthyretin continues to accumulate in the heart.

This finding is reminiscent of the capacity of the pathologic prion protein (PrPsc) to convert its normal counterpart (PrPc) into a pathologic conformation. The main difference is that the dominant negative effect, in the case of transthyretin, is due entirely to fibrils within the patient and is not transmitted from one person to another, as in prion disease.

Common Constituents of Amyloid

All amyloid deposits contain SAP (APCS), a glycoprotein that belongs to the pentraxin family (PTXs) and binds amyloid independently of the protein of origin. It has a specific binding motif for the common conformation of amyloid fibrils. This property makes radiolabeled SAP a diagnostic tool for the imaging of amyloid deposits. SAP (APCS) is highly protected against proteolysis and thus makes amyloid fibrils resistant to degradation.

Proteoglycans are also common in amyloid deposits and contribute extensively to the carbohydrate composition of natural amyloid. Heparan sulfate proteoglycans, in particular, have kinetics of deposition in tissue similar to that of fibrillar proteins and localize with constitutive elements of the extracellular matrix, such as perlecan, laminin, entactin, and collagen IV.

These molecules can constitute a scaffold, facilitating the initial phases of fibril nucleation, and could have a targeting role in the localization of amyloid deposits in tissue. For example, apolipoprotein E is reportedly a common constituent of amyloid deposits, and epidemiologic studies have shown an increased risk of Alzheimer disease among white persons carrying the 4 allele of apolipoprotein E. However, the role of apolipoprotein E in systemic amyloidoses is less clear.

Tissue Specificity of Amyloid Deposition

The remarkable diversity in the organ distribution of amyloid deposits remains one of the most important unsolved problems in amyloid research.

Specific proteins aggregate predominantly in defined target organs: beta2-microglobulin in joints, the fibrinogen A chain in the kidney, and the transthyretin Met30 variant in peripheral nerves. In light-chain amyloidosis, the deposits can involve virtually any organ.

One quarter of patients present with clinical involvement of a single organ, and the organ affected establishes the prognosis. Localized deposition of proteins that are normally deposited systemically can also occur, such as in localized AL amyloidosis, an intriguing condition characterized by localized growth of monoclonal plasma cells and the restriction of amyloid deposits to sites adjacent to the synthesis of the precursor.

The site of deposition may depend on the concurrence of several factors favoring the formation of fibrils, such as a high local protein concentration, a low pH, the occurrence of proteolytic processing, and the presence of fibril seeds.

Specific interactions with tissue glycosaminoglycans or cell-surface receptors such as the receptor for advanced glycation end-products (RAGE) may be important.

In AL amyloidosis, recognition of particular tissue constituents (i.e., collagen) by amyloidogenic light chains may determine the specificity of tissue deposition. A specific kidney tropism of the light chains derived from the 6a germ-line gene has been demonstrated; the tropism may occur because of the interaction of these proteins with mesangial cells.

Mechanism of Tissue Damage

There is lively debate about the mechanism by which aggregation causes tissue damage and organ dysfunction. The deposition of large amounts of fibrillar material can subvert the tissue architecture and consequently cause organ dysfunction.

Amyloid fibrils may also cause organ dysfunction by interacting with local receptors, such as RAGE.

In Alzheimer disease, an inflammatory response in the cerebral cortex elicited by the progressive accumulation of A contributes to the pathogenesis of the disease.

In A and transthyretin amyloidosis, soluble oligomeric intermediates of fibril assembly are cytotoxic in vitro and in vivo.

Soluble fibril precursors are likely to be the quaternary structures that mediate cellular toxicity through a mechanism that causes oxidative stress and activates the apoptotic pathway.

According to this hypothesis, mature amyloid fibrillary deposits are inactive proteinaceous reservoirs that are in equilibrium with smaller, putatively toxic assemblies (ordered aggregates).

Several clinical clues suggest that in AL amyloidosis as well, soluble oligomers are cytotoxic and contribute to organ dysfunction.

For example, peripheral neuropathy and renal and cardiac function improve dramatically after chemotherapy has halted the production of amyloidogenic light chains but before the expected resolution of amyloid deposits. The ongoing elucidation of the mechanism of tissue damage by other amyloid proteins may ultimately redirect therapeutic efforts.


- AA amyloidosis
- AL amyloidosis


- systemic amyloidosis
- localized amyloidosis

See also

- amyloidogenic proteins
- conformational diseases
- conformational dementias
- intermolecular linkages
- protein aggregation


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- Sungur, C. I., van der Hilst, J. C.H., Simon, A., Drenth, J. P.H., Merlini, G., Bellotti, V. (2003). Molecular Mechanisms of Amyloidosis. NEJM 349: 1872-1873.

- Merlini G, Bellotti V. Molecular mechanisms of amyloidosis.
N Engl J Med. 2003 Aug 7;349(6):583-96. PMID: 12904524

- Sigurdsson EM, Wisniewski T, Frangione B. Infectivity of amyloid diseases. Trends Mol Med. 2002 Sep;8(9):411-3. PMID: 12223307