Pathophysiology of lupus nephritis

1. Systemic lupus erythematosus
Dr. Mohit Mathur
14/10/14

2. Introduction
 The term “lupus erythematosus” was
introduced by 19th-century physicians to
describe skin lesions, it took almost 100
years to realize that the disease is systemic
and spares no organ and that it is caused by
an aberrant autoimmune response.
 In 1941, Klemperer, Pollack and Baehr first
described systemic lupus erythematosus (SLE)
as one of the CTD

3. Definition
 Systemic lupus erythematosus (SLE) is an
autoimmune disease in which organs and
cells undergo damage mediated by tissue-
binding autoantibodies and immune
complexes

4.  The prevalence ranges from 20 to 150 cases
per 100,000 population, with the highest
prevalence reported in Brazil.
 Prevalance appears to be increasing as the
disease is recognized more readily and
survival is increasing.
 The onset of disease peaks between 15 and
45 years of age, and more than 85% of
patients are younger than 55 years of
age.
 The 10-year survival rate is about 70%.

5.  Females outnumber males by about 10 to 1
 Males with SLE have the same incidence of
renal disease as females.
 SLE is more likely to be associated with
severe nephritis in children and in males and
is less likely in elderly individuals

6.  Females outnumber males by about 10 to 1
 Males with SLE have the same incidence of
renal disease as females.
 SLE is more likely to be associated with
severe nephritis in children and in males and
is less likely in elderly individuals

7. Pathogenesis and Etiology
There are multiple susceptibility factors,
which result in abnormal immune responses,
which vary among different patients.
These factors include:
 Genetic factors
 Environmental factors

9. Abnormal immune responses
 Those responses may include:
(1) Activation of innate immunity
(dendritic cells,monocyte/macrophages) by
CpG DNA, DNA in immune complexes, viral RNA,
and RNA in RNA/protein self-antigens;
(2) Abnormal activation pathways in adaptive
immunity cells (T and B lymphocytes);
(3) Ineffective regulatory CD4+ and CD8+ T cells;
(4) Reduced clearance of immune complexes and
of apoptotic cells.

10. Pathogenesis
 Self-antigens (nucleosomal DNA/protein;
RNA/protein in Sm, Ro, and La;
phospholipids) are available for recognition
by the immune system in surface blebs of
apoptotic cells;

11. …pathogenesis
 Immune cell activation is accompanied by increased
secretion of multiple cytokines and inflammatory mediators
like:
a) Type 1 and 2 interferons (IFNs),
b) Tumor necrosis factor
c) Interleukin (IL)-17 and IL-10.
d) B cell–maturation/survival cytokines B lymphocyte stimulator
(BLyS/BAFF)
Note: These cytokines produce fever, malaise, myalgia, weight loss etc
(Similar to viral infections)
 Decreased production of other cytokines also contributes
to SLE:
Lupus T and natural killer (NK) cells fail to produce enough IL-2
and transforming growth factor to induce and sustain regulatory
CD4+ and CD8+ T cells.

12. …pathogenesis
 The result of these abnormalities is sustained
production of autoantibodies and immune
complexes;

13. …pathogenesis
 Due to the defects listed above antigens,
autoantibodies, and immune complexes persist for
prolonged periods of time, allowing inflammation
and disease to develop.
 Pathogenic antibodies/immunecomplex bind target
tissues, with activation of complement, leading to
release of cytokines, chemokines, vasoactive
peptides, oxidants, and destructive enzymes.
 This is accompanied by influx into target tissues of T
cells, monocyte/macrophages, and dendritic cells,
activation of resident macrophages and dendritic
cells.

15.  In the setting of chronic inflammation,
accumulation of growth factors and products
of chronic oxidation contribute to irreversible
tissue damage, including fibrosis/sclerosis, in
glomeruli, arteries, brain, lungs, and other
tissues.

17. Clinical features
 The clinical heterogeneity of the disease led
to the establishment of 11 criteria with 4
needed for the formal diagnosis of systemic
lupus erythematosus (SLE).

18. Clinical features

19. As early as the 13th century, medical writings by Rogerius (c. 1230) and Paracelsus (c.
1500) proposed the Latin term lupus, meaning wolf, to describe the erythematous
ulcerative lesions affecting the skin of the cheeks

20. Clinical features

23. Diagnosis
Autoantibody testing
 In 1948 Malcom Hargrave, Helen Richmond and the medical
resident Robert Morton noted the presence of previously
unknown cells in the bone marrow of a patient with SLE.
 They called these LE cells and described them as mature
polymorphonuclear leukocytes which had phagocytosed the
liberated nuclear material of another leukocyte
 This extremely important discovery laid the foundation of
research for ANA.

24. Presently the ANA have been categorized in
to 2 main groups:
 Autoantibodies to DNA and histones
These include antibodies against
single/doublestranded DNA (dsDNA) and histones
 Autoantibodies to extractable nuclear antigens
(ENA).
i. These nuclear antigens were named ENA as they
are extracted from the nuclei with saline.
ii. Autoantibody to Smith antigen (Sm),
ribonucleoproteins (RNP), SSA/Ro, or SSB/La, Scl-
70, Jo-1 and PM1.

25. ANA – the two broad subtypes

27. Techniques for ANA detection
 Indirect immunofluorescence antinuclear
antibody test (IF-ANA)
 Enzyme immunoassay (EIA)/enzyme linked
immunosorbent assay (ELISA)

28. IF-ANA: The standard ANA testing
technique
 It is inexpensive and easy to perform, with
high sensitivity and specificity
 The test detects the presence of ANA in the
blood of the patient which adhere to HEp-2
cell substrate, forming distinct fluorescence
patterns.

31. ANA titer
 It is directly proportional to antibody
concentration and expressed with a
quantitative scale of values.
 Its evaluation is crucial as low titer is less
significant, and may be seen even in healthy
individuals.
 A titer of 1:160 is taken as significant.

32. ELISA
 ELISA is both highly specific and sensitive and
substantially decreases the time involved when
screening large numbers of patient samples.
 The test is simple to perform, can be automated and
does not require highly trained operators who can
recognize microscopic patterns.
 The EIA/ELISA is therefore becoming the most
widely used method not only for routine screening
but also for detection of specific ANA.

34. Complement Levels
 Levels of total hemolytic complement (CH50) and
complement components are usually decreased
during active SLE and especially active LN.
 Levels of C4 and C3 often decline before a
clinical flare of SLE.
 Serial monitoring of complement levels, with a
decline in levels predicting a flare, is considered
more useful clinically than an isolated depressedC3
or C4 value.
 Normalization of depressed serum complement
levels is often associated with improved outcomes.

35. Pathogenesis of Lupus
Nephritis

36. DISEASE PATHOMECHANISMS AT
THE KIDNEY LEVEL
 IgG, IgM, IgA deposition and complement activation
via classical pathway.
(a) Glomerulus:
Mesangial compartment (glomerular inflammation,
class I and II);
Sub-endothelial compartment (vascular
inflammation, class III and IV);
Subepithelial compartment (podocyte injury,class V).
(b) Tub.-Int: Peritubular vascular injury.

37. Disease patho-mechanism at the kidney
level
 Fc, Toll-like, and complement receptor
activation.
 Increased, local expression of cytokines,
chemokines and adhesion molecules.
(a) Glomerulus: Activation of glomerular
parenchymal cells and infiltrating leukocytes.
(b) Tub.-Int: Activation of peritubular
endothelial cells and tubular epithelial cells.

38. Cont…
 Recruitment of leukocytes with pro-
inflammatory effector functions.
(a) Glomerulus: Amplification of inflammation
via release of cytokines and cytotoxic factors.
(b) Tub.-Int: Similar to glomerulus, tertiary
lymphoid organ formation, specifically
local immunoglobulin production.

39.  Programmed death of renal parenchymal
cells – reparative hyperproliferation.
(a) Glomerulus: Mesangio-proliferative
glomerulonephritis (GN), endocapillary GN,
podocyte loss, parietal cell hyperproliferation
and cellular crescent formation.
(b) Tub.-Int: Tubular atrophy, loss of
peritubular vasculature causing hypoxia.

40.  Insufficient regeneration and scarring
(a) Glomerulus: Focal segmental
glomerulosclerosis, fibrocellular crescents,
global glomerulosclerosis, i.e. nephron loss
(class VI).
(b) Tub.-Int: Tubular atrophy and nephron
loss, interstitial fibrosis.

41. When to biopsy?
 Renal biopsy is generally indicated in a
patient with any combination of:
a) Acute increase in serum creatinine,
b) Proteinuria >500 mg/24 h,
c) Hematuria in presence of any level of
proteinuria.
d) Active sediment/cellular casts
e) To detect class switch/transformation in
an established case of LN

42. Role of renal biopsy
 Glomerulonephritis (GN) is the most common form
of renal disease in patients with SLE, but is
frequently accompanied by tubulointerstitial and/or
vascular lesions (such as thrombosis secondary to
antiphospholipid syndrome).
 A variety of other nephropathies have also been
described like
a) Renal amyloidosis,
b) Focal segmental glomerulosclerosis,
c) Minimal-change disease,
d) IgA and IgM nephropathy,
e) Necrotizing glomerulitis,
f) Sarcoidal and NSAID-induced tubulointerstitial
nephritis

43. Clinico-pathological correlation
 Several studies have focused on the
discrepancy between clinical presentation
and pathologic findings at renal biopsy in
patients with SLE.
 Silent LN has been reported not only in class
II but also in class IV.
 Even patients with low-level proteinuria
(<1g/24h) have demonstrated significant
renal involvement with proliferative LN
(classes III or IV)

44. Glomerular pathology
Terminology
 The major histologic abnormalities of the
glomerulus in lupus nephritis include immune
deposits, glomerular proliferation, influx of
leukocytes, glomerular necrosis, and
scarring.

45. Wire loop
 Wire loops, a classic sign of active lupus
nephritis, are segmental areas of refractile,
eosinophilic, thickening of the glomerular
capillary seen by light microscopy in
haematoxylin and eosin stained sections
 They correspond to massive subendothelial
electron-dense deposits on electron
microscopy, that when large enough to
completely involve the peripheral
circumference of the glomerular capillary wall

46. Wire loop lesions

47. Hyaline thrombi
 Hyaline thrombi are large, acellular,
eosinophilic, intracapillary deposits that
occlude the glomerular capillary lumens
 The term is actually a misnomer because
they do not represent true fibrin thrombi but
are instead massive intracapillary immune
deposits

48. Hyaline thrombi

49. Hypercellularity
 Proliferation of glomerular endothelial, mesangial,
and epithelial cells and infiltration of leukocytes is
the most frequent histological finding in lupus
nephritis.
 Mesangial hypercellularity and matrix expansion are
the first observable responses to mesangial
deposits.
 The endocapillary hypercellularity results from the
proliferation of glomerular endothelial and mesangial
cells, as well as by leukocyte infiltration that
occludes the glomerular capillary

51. Crescents
 Cellular crescents are a feature of active lupus
nephritis.
 Cellular crescents commonly overlie necrosis of the
glomerular tuft, and are formed by proliferating
parietal epithelial cells with infiltrating mononuclear
cells (monocytes or macrophages).
 The greater the proportion of glomerular
involvement (i.e > 50%), the worse the prognosis.
 With evolution of the glomerular injury, there is
progressive scarring of cellular crescents, forming
fibrocellular and fibrous crescents.

53. Necrosis
 Glomerular necrosis is a destructive inflammatory
lesion that heals with scarring and is frequently
associated with crescent formation.
 The histological diagnosis of necrosis is established
by the presence of fibrin, ruptures (breaks) of the
glomerular basement membrane, and neutrophilic
infiltrates with karyorrhexis.
 Karyorrhexis refers to apoptosis of infiltrating
neutrophils producing pyknotic and fragmented
nuclear debris(“nuclear dust”)

55. Classification of Lupus Nephritis
 The original World Health Organization
(WHO) classification of LN introduced in 1974
has evolved into the 2003 International
Society of Nephrology (ISN)/Renal Pathology
Society (RPS) classification

57.  The changes introduced in the new ISN/RPS
classification include:
a) The “normal” category of LN was
eliminated from the previous WHO
classifications, and mesangial LN was thus
divided into two categories to maintain the
relevant number designation of other classes.
b) ISN/RPS classification has more
detailed definitions of specific categories, and
this has resulted in improved reproducibility.

58. ISN/RPS Class I
 Minimal mesangial lupus nephritis
 Class I is characterized by normal glomeruli
by light microscopy.
 Purely mesangial immune deposits identified
by immunofluorescence.
 This is the mildest form of lupus nephritis,
 Class I patients have minimal clinical renal
disease

59. ISN/RPS Class II
 The glomeruli in Class II may
have any degree of mesangial
proliferation by light
microscopy, associated with
mesangial immune deposits by
immunofluorescence.
 Mesangial hypercellularity is
defined as greater than three
cells in mesangial regions
distant from the vascular pole
in 3-μm–thick sections.
 Any active or inactive lesion or
significant subendothelial
deposits (wire loops) disqualify
the biopsy from this class.
 The clinical renal
manifestations of Class II
patients are mild and the
prognosis is excellent.

60. ISN/RPS Class III
 ISN/RPS class III, focal LN, is defined as focal
(< 50% Gloms) - segmental or global endocapillary
or extracapillary GN..
 Endocapillary and extracapillary proliferation.
 Class III biopsies may have active (proliferative),
inactive (sclerosing), or active and inactive
lesions subclassified as A, C, or A/C,
respectively.
 Active lesions may display cellular crescents,
fibrinoid necrosis, nuclear pyknosis or karyorrhexis,
wireloop lesions, hyaline bodies and rupture of the
glomerular basement membrane (GBM).

61.  Hematoxylin bodies, swollen basophilic
nuclear material acted upon by antinuclear
antibodies, are occasionally found within the
necrotizing lesions (Specific Lesion)
 In class III biopsies, glomeruli adjacent to
those with severe histologic changes may
show only mesangial abnormalities by LM.
 In class III, diffuse mesangial and focal and
segmental subendothelial immune deposits
are typically identified by IF and EM.

63. ISN/RPS class IV
 ISN/RPS class IV or Diffuse proliferative
LN, has qualitatively similar glomerular
endocapillary proliferation as class III, but
the proliferation involves more than 50%
of the glomeruli
 ISN/RPS class IV is subdivided into:
i) Diffuse segmental proliferation (class IV-S)
More than 50% of affected glomeruli
have segmental lesions
ii) Diffuse global proliferation (class IV-G)
More than 50% of affected glomeruli have global
lesions.

64.  The most controversial of these changes in the
ISN/RPS classification system is the subdivision of
class IV into diffuse global and diffuse segmental
LN.
 The Lupus Nephritis Collaborative Study Group
showed the Class IV-S biopsies had more extensive
fibrinoid necrosis and less prominent immune
deposits, and despite similar treatment, Class IV-S
had a worse prognosis than Class IV-G.
 However, several studies have shown that suggest
lupus nephritis Class IV-G has a similar or worse
outcome than lupus nephritis Class IV-S

65. ISN/RPS class IV
 All of the active features described for class
III may be encountered in ISN/RPS class IV
LN.
 There is more extensive peripheral
capillary wall subendothelial immune
deposition and extracapillary proliferation in
the form of crescents is common.
 Occasionaly MPGN pattern or Necrotising
vasculitis may also be seen

66.  Class IV patients usually have evidence of
active systemic disease and have the most
severe and active clinical renal presentation.
 Proteinuria is universal, haematuria occurs to
variable degrees in 80% - 90% of patients,
and renal insufficiency is detected in more
than 50% of patients.
 These patients have the worst prognosis
despite optimal treatment

67. ISN/RPS class V
 Defined by regular subepithelial immune deposits producing a
membranous pattern
 The coexistence of mesangial immune deposits and mesangial
hypercellularity helps to distinguish membranous LN from
primary membranous glomerulopathy.
 There is typically thickening of the glomerular capillary walls and
“spike” formation between the subepithelial deposits.
 When the membranous alterations are accompanied by focal or
diffuse endocapillary proliferative lesions and subendothelial
immune complex deposition, they are classified as class V +
III and V + IV, respectively.
 Because sparse subepithelial deposits may also be encountered
in other classes (III or IV) of LN, a diagnosis of pure lupus
membranous LN should be reserved only for cases in which the
membranous pattern predominates.

69. ISN/RPS class VI
 ISN/RPS class VI, advanced sclerosing LN or
end-stage LN, is reserved for biopsies with
more than 90% of the glomeruli sclerotic.
 There are no active lesions, and it may
be difficult to even establish the diagnosis
of LN without the identification of residual
glomerular immune deposits by IF and EM.

72. Activity and chronicity Index
 The biopsy is graded on a scale of 0 to 3+ for each
of six histologic features.
 The severe lesions of crescents, and fibrinoid
necrosis are assigned double weight.
 The sum of the individual components yields a total
histologic activity index score of from 0 to 24.
 Likewise, a chronicity index of 0 to 12 is
derived.
 High activity index (>12) and especially a high
chronicity index (>4) have a poor 10-year renal
survival.
 However, in several large studies, neither the
activity index nor the chronicity index correlated
well with long-term prognosis.

74. Immunofluoroscence
 In LN, immune deposits can be found in the
glomeruli, tubules, interstitium, and blood vessels.
 IgG is almost universal, with co-deposits of IgM
and IgA in most specimens.
 Both C3 and C1q are commonly identified.
 The presence of all three immunoglobulins, IgG,
IgA, and IgM,along with the two complement
components, C1q and C3, is known as “full house”
staining and is highly suggestive of LN.
 Staining for fibrin-fibrinogen is common in the
distribution of crescents and segmental
necrotizing lesions.

75. Electron Microscopy
 The distribution of glomerular, tubulointerstitial, and vascular
deposits seen by EM correlates closely with that observed by IF.
 Deposits are typically electron dense and granular.
 Some exhibit focal organization with a “fingerprint” substructure
composed of curvilinear parallel arrays measuring 10 to 15 nm
in diameter.
 Tubuloreticular inclusions (TRIs), intracellular branching
tubular structures measuring 24 nm in diameter located within
dilated cisternae of the endoplasmic reticulum of glomerular and
vascular endothelial cells, are commonly observed in SLE
biopsies.
 TRIs are inducible upon exposure to INF-α(so-called “INF
footprints”) and are also present in biopsies of HIV-infected
patients and those with some other viral infections

76. Lupus nephritis class IV. Electron micrograph
showing a large subendothelial electron-dense
deposit as well as a few small subepithelial
deposits (arrow) (×1200)

77. Prognostic Factors Not Addressed in the ISN/RPS
classification – Tubulointerstitial and vascular
involvement
 The ISN/RPS classification is based entirely on an
assessment of the glomerular alterations;
 Tubulointerstitial and vascular lesions were not
factored into the previous WHO classifications nor
the 2003 ISN/RPS classification.
 However, the 2003 ISN/RPS classification
stipulates that the pathology report should include
grading of the degree of tubular atrophy and
interstitial fibrosis, interstitial inflammation and
vascular disease.

78.  Severity of tubulointerstitial lesions correlates with glomerular
proliferative lesions in LN.
 Active tubulointerstitial lesions include edema and inflammatory
infiltrates, T lymphocytes (CD4 and CD8), monocytes, plasma cells and
destructive tubulitis.
 Tubulointerstitial immune deposits of Ig’s or complement may be
present along the basement membranes of tubules and interstitial
capillaries.
 Interstitial fibrosis or tubular atrophy seen in chronic phases of LN and
are significant independent risk factors for renal outcome.

87. Biomarkers in detecting lupus activity
 Conventional markers of active renal disease are serum levels of
anti-dsDNA, anti-C1q antibody and complement levels have
variable sensitivity between 50 and 75% for active renal disease.
 Urinary biomarker candidates have included adhesion molecules,
cytokines, chemokines, and their receptors, including VCAM-1,
P-selectin, IL-6, IP-10, RANTES, MCP-1, CXCL16, CX3CL1,
TWEAK, and TNFR1
 Urinary proteomic approaches have been used to identify
proteins like transferrin, ceruloplasmin, hepcidin 20, hepcidin 25,
α1-acid-glycoprotein, lipocalintype prostaglandinD-synthetase,
and neutrophil gelatinase associated lipocalin.
 The utility of above biomarkers need independent confirmation in
large scale prospective studies.

88. Conclusions
 Lupus nephritis can present as nephritic
and/or nephrotic syndrome with various
combinations of edema, constitutional
symptoms, proteinuria, hematuria, impaired
renal function, abnormal lipid profile and
hypertension.
 Kidney biopsies remain the gold standard for
establishing the
diagnosis/prognosis/treatment of LN.

89. Thank you