Amyloidosis Diagnosis and
Classification in Native Kidney
and Renal transplants
Prof. Dr. B. Handan Özdemir
Baskent University
Ankara-Turkey
Definition
Amyloidosis comprises a diverse
group of systemic and local diseases
Characterized by organ involvement
by the extracellular deposition of
fibrils composed of subunits of a
variety of normal serum proteins
Physical Nature
Amyloid fibril protein occurs in
tissue deposits as rigid, nonbranching fibrils 7-to 10 nm in dm
When analysed by X-ray
diffraction, the fibrils
exhibit a characteristic
cross Beta diffraction
pattern
Chemical Nature

Pentagonal molecule

95% Protein Fibril

5% Glycoprotein P component
Genetic factors may be involved in predisposing to
the development of fibrillogenesis and amyloidosis
•
Cofactors such as amyloid P component may have an
important role in the tissue deposition and resorption
Promoting fibrillogenesis
Act by
• Stabilization of the fibrils
• Binding to matrix proteins
• Affecting metabolism and proteolysis of formed fibrils
•
Husby G. Clin Immunol Immunopathol 1994:70:2
What factors allow some proteins to aggregate into
amyloid fibrils ?
Patients with AA amyloidosis have levels of
SAA protein no greater than those patients
with inflammatory diseases who do not have amyloid
Therefore some additional unknown stimulus is
required for amyloid fibrils to form and precipate
In AL amyloidosis biochemical characteristics
of the light chain is important in
determining amyloid formation
Cast nephropathy versus Amyloidosis
Certain light chains also may form high molecular
weight aggregates in vitro
Amyloid fibril protein nomenclature

The established amyloid fibril nomenclature is
based on the chemical nature of the fibril protein

Which is designated protein A and followed by a
suffix that is an abbreviated form of the parent
or precursor protein name

For example, when amyloid fibrils are derived from
immunoglobulin light chains, the amyloid fibril is AL
and the disease is AL amyloidosis
Amyloid, 2010; 17(3–4): 101–104
Amyloid fibril protein nomenclature

At least 25 different precursor proteins are known

They are associated with variety of
Inflammatory
Immune
Infectious
Hereditary conditions
TYPING OF RENAL AMYLOIDOSIS

Typing of amyloid deposits is important because of
the difference in their treatment strategies

Typing of the amyloid deposits can be performed
with various techniques

The most definitive method used is IF or IHC
staining of tissue using antibodies that are
directed against known amyloid proteins
Amyloid Typing and Pitfalls

IHC typing of amyloidosis poses several problems
and requires experience

Wide range of success in IHC amyloid typing,
ranging from 38% to 87% was reported

IHC diagnosis of AA type is relatively reliable,

But there is a problem in the differentiation of AL
and hereditary amyloidosis
Kebbel A, Röcken C. Am J Surg Pathol. 2006;30:673
Amyloid Typing and Pitfalls
•
Commercial antibodies are raised against the
constant regions of the Ig light chains
•
A subset of AL, in which amyloid fibrils
are derived from a truncated light chain
“containing only variable regions”
will be nonreactive with commercial antibodies
•
Therefore, negative light chain staining does not
rule out AL amyloidosis
Immunopathology in Renal Amyloidosis
The typical antibody panel should include
• Amyloid P component
• Kappa & lambda Ig light chains
• Amyloid A protein
• Transthyretin
• Fibrinogen
• Beta-2 microglobulin
•
•
This panel allowed definitive
typing of amyloid in 90% of kidney biopsies
Classification of Amyloidosis
Systemic Amyloidosis
Primary Amyloidosis
Secondary Amyloidosis
Localized amyloidosis
Senile cerebral
Senile cardiac
Type 2 diabetes
Primary Systemic Amyloidosis
Disease name
Type of amyloid
Precursor
M.Myeloma
AL
Light chains
Primary
AL
Light chains
Secondary Systemic Amyloidosis
Disease name
Chronic inflammatory
disease
Hemodialysis
associate
Type of amyloid
AA
Aβ2- micro globulin
Precursor
SAA
β2- micro globulin

The most common type of amyloidosis depends on
the population studied

In the USA and the Western World AL amyloidosis
is the most prevelant, followed by AA amyloidosis

In Turkey AA amyloidosis with an underlying
disease of FMF is the most common type
Ozdemir AI. Am J Gastroenterol 1969; 51:311

In developing countries AA amyloidosis is far more
common than AL amyloidosis (Tbc !)
AL Amyloidosis

Derived from the immunoglobulin light chain

Can occur alone or in association with

M. Myeloma

Malignant lymphomas

Macroglobulinemia
Kyle RA et al N Engl J Med. 2006;354:1362
AL Amyloidosis

Light chain deposition disease has a similar
pathogenesis with AL amyloidosis

Primary difference is that

Deposited light chain fragments do not form fibrils
and do not engender deposition of amyloid cofactors
AL Amyloidosis
•
Wide spectrum of organ system involvement
•
The kidneys are commonly affected
•
Proteinuria
•
Edema and hypoalbuminemia
•
Mild renal dysfunction is frequent
•
Rapidly progressing renal failure is rare
Waxy appearance of intradermal amyloid
deposition around the eye
Macroglossia showing
teeth indentations
Peri-orbital haemorrhage
(raccoon or panda eyes)
Gross lymphadenopathy
AA Amyloidosis

The AA amyloid proteins result from a
proteolytic cleavage SAA protein

SAA is an acute-phase reactant produced
by liver

Sustained high concentration of SAA
prerequisite for AA amyloidosis
Benditt EP et al. Ann N Y Acad Sci. 1982;389:183
AA Amyloidosis
Acquired and hereditary diseases can lead to AA
amyloidosis
Chronic inflammatory diseases
Rheumatoid arthritis
Rheumatoid arthritis
JCA
Other arthritides
Inflammatory bowel disease
Chronic sepsis
FMF
Periodic fevers
Crohn's
Bronchiectasis,
Castleman's
Other
Tuberculosis
Unknown
0
50
100
150
AA Amyloidosis
•
Kidney is the most frequently affected target organ
•
The primary clinical manifestation is proteinuria
•
The underlying disease usually is longstanding
•
Active inflammation typically is present when
amyloidosis becomes evident
Hereditary Amyloidosis

Caused by deposition of genetically variant proteins

Associated with mutations in the genes for

Transthyretin
Apolipoprotein AI,
Apolipoprotein AII
Lysozyme
Fibrinogen A.




Hereditary Amyloidosis

Typically they associated with polyneuropathy and
cardiac involvement but can affect kidneys

Renal deposits may be clinically silent

Isolated glomerular involvement with no amyloid in
the tubules, interstitium, or vessels has been
found to be characteristic of fibrinogen A

Renal failure develops rapidly
Blood. 1997;90:799–805
RENAL AMYLOIDOSIS

Clinically evident renal involvement occurs mainly in
AA and AL amyloidosis

The deposition of Abeta2 M occurs in patients on
prolonged dialysis

But diagnosis on the kidney biopsy is unexpected

Eight precursor fibrils are particularly important
for kidney
RENAL AMYLOIDOSIS

The incidence of amyloid in patients with nephrotic
syndrome or proteinuria was found in 2% to 12%
of native renal biopsies
N Engl J Med. 2003;349:583–596.
Arch Pathol Lab Med. 2007;131:917–922
Amyloidosis without therapy usually progresses to
endstage kidney disease
Deposits may also regress
Ozdemir BH. Transplant Proc. 2006;38:432–434.
Diagnosis of Amyloidosis
• Can be very difficult
• No blood test can diagnose or exclude amyloidosis
• Usually relies on clinical suspicion
• Possibility supported by
•
•
•
•
Underlying chronic inflammatory state – AA
Underlying plasma cell dyscrasia – AL
Family history - hereditary
Evidence of renal dysfunction
Detection of Renal Amyloid

In H&E stained sections, amyloid is recognized as
amorphous hyaline and eosinophilic material

Weakly PAS positive
Detection of Amyloid
•
Amyloid do not stain by silver staining
•
Occasionally may stain with silver stains and
show spicules (Jones silver)
Detection of Amyloid
•
Congo red is the gold standard
•
Slides must be examined under polarized light
•
Apple-green birefringent deposits is diagnostic
Detection of Amyloid

Caution is advised regarding ‘‘overinterpreting’’
collagen as amyloid

Because deposits of amyloid are frequently very
focal and irregularly distributed in tissue sections

Multiple and thicker (5–10 m) sections may need to
be examined
Picken MM. Curr Opinion Nephrol Hypertens. 2007;16:196
Detection of Amyloid






Other stains, or techniques
Fluorescence
Thioflavin S and T
Methyl violet
Sulphonated Alcian blue
They are less specific
Sen S. Arch Pathol Lab Med 2010: 134: 532
Curr Opinion Nephrol Hypertens. 2007;16:196
Electron Microscopy
•
Characterized by randomly disposed, rigid,
nonbranching, variably long, 7-to 10nm-dm fibrils
•
Ultrastructural immunogold labelling can depict the
precursor protein in amyloif fibrils
Electron Microscopy
•
Rare cases exhibit massive aggregates of amyloid
fibrils in subendothelium
•
They arranged in thightly packed, electron-dense
structures and can be easily confused with
•
MPGN
Cryoglobulinemic glomerulopathy
•
•
These cases are usually associated with monoclonal
kappa light cahins
Gross Pathology of Renal Amyloidosis
•
•
•
Enlarged kidneys
Pale, waxy appearing cut surfaces
Increase in the weight of kidney
Weight of the kidney did not correlate with
Renal function
The site of renal amyloid deposition
Inversely proportional to the amount of amyloid
Microscopy of Renal Amyloidosis
•
Amyloid can be found in any of the
renal compartment
•
The most common and the earliest
site of amyloid deposition in the
kidney is the glomeruli
•
Glomerular amyloid formations
begins in the mesangium
•
Extends into capillary walls
Microscopy of Renal Amyloidosis
•
Amyloid deposition in glomeruli may occur
•
Segmental
•
Diffuse mesangial
•
Nodular
•
Pure basement membrane patterns
A proposed histopathologic classification
Renal amyloidosis was divided into 6 classes
Similar to the classification of SLE
Sen S. Arch Pathol Lab Med 2010: 134: 532
Microscopy of Renal Amyloidosis
•
Early segmental deposits are small and confined to
mesangium without creating nodularity
•
It is very easy to miss this early form
Microscopy of Renal Amyloidosis
•
In the diffuse form
•
The mesangium is uniformly expanded by deposit
Microscopy of Renal Amyloidosis
In the nodular form
• Mesangium is asymmetrically expanded by amyloid
•
•
•
Distinguish from diabetic nephropathy
Other forms of nodular glomerulosclerosis
Microscopy of Renal Amyloidosis
•
Subepithelial amyloid deposition
Associated with spikes
Can be seen at the periphery of mesangial areas
Microscopy of Renal Amyloidosis
•
Rarely cresents can be seen
Highlighting the fact that capillary wall rupture
has occured
Microscopy of Renal Amyloidosis
•
Interstitial amyloid are seen in 50% cases
•
Generally begins in areas adjacent to blood vessels
•
Medullary amyloid deposits are more frequent
Microscopy of Renal Amyloidosis
•
•
AA amyloidosis
Certain mutants of transthyretin
•
May show amyloid deposition limited to the
interstitium and medulla
•
Such patients present with renal failure not
associated with proteinuria
Microscopy of Renal Amyloidosis
•
Renal vessels are often involved
•
Arteriolar deposits being most frequent
•
Followed by deposits in arteries, PTCs and veins
RENAL AMYLOIDOSIS
•
Therapies are not successful in patients diagnosed
at advanced stages of amyloidosis
•
Supportive therapy is essential
•
There are two choices for the therapy
•
Hemodialysis
•
Transplantation
Hemodialysis in Renal Amyloidosis
•
The survival of patients begining dialysis is poor
•
Mean survival is only 33 months
•
Worse than patients with other renal diseases
Hemodialysis in Renal Amyloidosis
•
The European experience showed a 76% survival at
2 years for all patients compared with 53% among
amyloidosis patients
•
Similarly in our center the 2-year survival was
50% among patients with amyloidosis
•
The mean survival of our patients on hemodialysis
was 33 months
Hemodialysis in Renal Amyloidosis
•
Common complications of amyloidosis are
•
Extrarenal progression of amyloidosis
•
Infections
•
Major causes of death
Hemodialysis in Renal Amyloidosis
•
It has been reported that 21 of the 31 deaths in
the group that underwent dialysis were due to
extrarenal progression of amyloidosis, namely to
cardiac amyloidosis
•
Similarly, we have observed that infection and
extrarenal progressive amyloid deposition was the
cause of death in 15 of our 30 patients who died
approximately 9 months after starting hemodialysis
Ozdemir BH et al. Transplant Int 2004: 17: 241–246
Other Choice of Therapy is Transplantation
Transplantation in Renal Amyloidosis
•
The outcomes of renal tx in patients with
amyloidosis is still controversial
•
Pasternack
3-year survival
Patients with amyloidosis 51%
Patients with glomerulonephritis 79%
•
•
•
Pasternack A et al. Transplantation 1986; 42:598
Transplantation in Renal Amyloidosis
•
Pras noted similar graft and patient survival rates
among patients with amyloidosis and GN
Pras M et al. Adv Nephrol Necker Hosp 1984; 13:261
•
Similarly our 5-year survival rates for amyloidosis
patients (78%) were equal to the survival rates of
patients with GN
Ozdemir BH et al. Transplant Proc 2006: 38, 432
Ozdemir BH et al. Transplant Int 2004: 17: 241–246
Recurrence of Amyloidosis

The rates of recurrent amyloidosis vary from
center to center
(13 to 50%)

This is mainly because only a small number of
patients have been studied in single-center series
The rates of recurrence is dependent
on the indication of graft biopsy
Since early amyloidosis could exist
without urinary abnormalities or
impairment of renal function
Recurrence of Amyloidosis

In our center we observed higher rates of amyloid
recurrence than have been reported previously

Recurrent amyloidosis was diagnosed by renal
allograft biopsy in 20 of our 30 cases

Of 20 grafts, 18 were from living and 2 were from
cadaveric donors (P<0.01)

Four of the five patients (80%) with HLA-identical
LRDs showed amyloid recurrence
Ozdemir BH et al. Transplant Int (2004) 17: 241–246
Recurrence of Amyloidosis

Previous reports document amyloid recurrence in
renal grafts at 6 months to 4 years after tx
Transplantation 1981; 32:6
Arch Intern Med 1979; 139:1135

In our patients, amyloidosis developed in the grafts
18 months to 10 years after tx

This is a very wide range, emphasising that amyloid
recurrence can develop at any time after tx
Ozdemir BH et al. Transplant Int (2004) 17: 241–246
Recurrence of Amyloidosis

Recipients with LRD showed shorter times to
recurrence (33.6±5.2 months) than for those with
cadaveric grafts (78±24 months; P<0.01)

The overall 3-, 5- and 10-year graft survival
rates for the recipients with amyloidosis
recurrence were 77%, 70%, and 36%, respectively

The corresponding patient survival rates were 93%,
78%, and 35%
Ozdemir BH et al. Transplant Int (2004) 17: 241–246
Why so many patients showed such a high
incidence of amyloidosis recurrence, even
though they were taking colchicine ?
Part of the reason for this may be poor
Drug compliance
Doubting the need for medication
Preference for self-care measures other than medication
Convenience, side effects and lack of demonstrated benefit
CONCLUSION
•
Amyloidosis patients maintained on chronic
dialysis have high mortality rates
•
Better survival was noted among renal
transplant patients even with a recurrence
•
These results encourage transplantation for
renal end-stage disease due to amyloidosis