CASE

PC :18 year old Malay boy referred for an incidental finding of proteinuria and microscopic hematuria. No other complaints System review was unremarkable PMH: Bronchial asthma( well controlled) Birth history: Full term baby. No complications. No issues during his childhood DH: Ventolin as needed SH: Currently studying at college. Denies smoking, illicit drugs or alcohol

CASE
FH : Uncle is kidney transplant recipient ( primary diagnosis unknown, ESRD occurred when he was in his late 20s

Examination
NORMAL NO OEDEMA

Investigations

US: Normal kidneys. RK 10.3cm LK 10.9 cm. No cyst or calculi.

Differential Diagnosis
Ig A nephropathy

FSGS
Alport syndrome

15 glomeruli at best level -One globally sclerosed

10x HE: One globally sclerosed glomerulus

10x HE: capsular reaction

40x HE: Mild mesangial proliferation

40x HE: One glomerulus with early prolapsing tip into the tubule

40x PAS: capsular reaction

Focal tubular atrophy

DIAGNOSIS

Blood vessels unremarkable Interstitium: mild lymphoplasmacytic infiltrates, no increase in fibrosis, no foam cells HPE: Early focal segmental glomerulosclerosis IF studies: IgM 1+ (capillary wall) IgG, IgA, C3 and C4 negative

WOULD YOU DO ELECTRON MICROSCOPY IN THIS PATIENT ?

Capillary loops

Thinning of the basement membrane

http://www.unckidneycenter.org/images/alport_gbm.gif

Capillary lumen

Split and laminated basement membrane

podocytes

Mesangium

Basement membrane

Renal Biopsy - Light Microscopy

Light microscopy findings are nonspecific. Can see focal and segmental glomerular hypercellularity of the mesangial and endothelial cells. Renal interstitial foam cells can be found and represent lipid-laden macrophages which can be seen in many renal diseases.

Renal Biopsy - EM
Earliest finding is thinning of GBM. Characteristic finding of longitudinal splitting of lamina densa of GBM. May not be seen in young AS patients. The proportion of GBM that shows splitting increases from 30% by age 10 to more than 90% by age 30.

Rumpelt, HJ. Hereditary nephropathy: Correlation of clinical data with GBM alterations. Clin Nephrol 1980; 13:203.

Basement Membrane
Electron microscopy reveals to us that the basement membrane actually consists three layers: the lamina lucida (electron-lucent), lamina densa (electron-dense), and lamina fibroreticularis (electron-lucent).

HISTORY of ALPORT SYNDROME

In 1927, 3 later generations of the same family was followed by Dr Cecil Alport and he recognized that deafness was a component The disorder tended to be more severe in males than females, that affected males died of uremia, while females lived to old age. Subsequently, many more families were described and the disease was named Alport Syndrome (AS) in 1961.

Pathophysiology
AS is a primary basement membrane disorder arising from mutations in genes encoding several members of the type IV collagen family. Basement membranes are assembled through an interweaving of type IV collagen Six genes, COL4A1, COL4A2, COL4A3, COL4A4, COL4A5 and COL4A6 encode the six chains of collagen IV, 1(IV) through 6(IV), respectively.

PATHOPHYSIOLOGY
The six collagen IV chains only form three sets of triple helical molecules called protomers: 1.1.2(IV), 3.4.5(IV) and 5.5.6(IV

Pathophysiology

Two NC1 trimers unite to form a hexamer. Four 7S domains form tetramers with other protomers The three protomers only form three sets of hexamers to form collagenous networks:

1.1.2(IV) - 1.1.2(IV) 3.4.5(IV) 3.4.5(IV) 1.1.2(IV) 5.5.6(IV)

GENETICS
X-linked accounts for 80% of the affected patients & due to mutation COL4A5 Autosomal Recessive 15% of patients and defects in COL4A3/4 Autosomal Dominant :5% with heterozygous mutations COL4A3/4

Women with X-linked : almost all carriers has hematuria. Variable course in women may be due to lyonization

Inheritance Patterns
Chain 1(IV) 2(IV) 3(IV) Genes COL4A1 COL4A2 Chromosom e 13 13 Ubiquitous Ubiquitous GBM, tubular basement membrane, Descemet membrane, Bruch membrane, anterior lens capsule, lungs, cochlea GBM, TBM, Descemet membrane, Bruch membrane, anterior lens capsule, lungs, cochlea Epidermal basement membrane (EBM), Bowmans capsule (BC), GBM, distal TBM, Descemet membrane, Bruch membrane, anterior lens capsule, lungs, cochlea BC, TBM, EBM Tissue Distribution Mutation Unknown Unknown

COL4A3

ARAS*/ADAS **

4(IV)

COL4A4

ARAS/ADAS

5(IV)

COL4A5

XLAS

6(IV)

COL4A6

Leiomyomatosi s

*Autosomal recessive Alport syndrome, ** Autosomal dominant AS X-linked AS ARAS with mutations spanning COL4A5 and COL4A6 genes

Chain distribution
Monoclonal antibody have been used to determine distribution of Alpha- 3/4/5 Located in Bowmans capsule, basement membrane of glomerulus, distal and collecting tubules and basement membrane of eye and cochlea. In normal patients, alpha-5(IV) chain is present in basement membrane underlying epidermis (alpha 5-5-6)

IMMUNOSTAINING
X-Linked Male: complete absence of immunostaining for alpha-3/4/5(IV) while heterozygous exhibit patchy loss of staining in GBM and tubular membranes SKIN: 20% of X-linked Alport have normal staining of renal basement Autosomal recessive: GBM no staining but Bowmans capsules and tubular membrane staining for alpha 5(IV) and skin (alpha 5,5,6)

Immunohistological analysis of the renal distribution of type IV collagen chains. The analysis was carried out in (AD) control, X-linked (E,F) male and (G,H) female Alport syndrome patients, and (IL) patients with autosomal recessive Alport syndrome, using antibodies to 1(IV) (A,E,I), 3(IV) (B,F,J), or 5(IV) (C,G,K,L) chains. Double labeling was made with anti-2(IV) in red, and anti-5(IV) in green (D,H). In control kidney, the 1(IV) chain is present in the mesangial matrix, Bowman's capsule, and the extraglomerular basement membranes (A). The 3(IV) and 5(IV) chains are distributed within the GBM (B and C, respectively). The Bowman's capsule is strongly 5(IV)-positive (C). In X-linked Alport syndrome, no 3(IV) expression was detected in a male patient (id for 45) (F), whereas the distribution is segmental in a female patient (G). In autosomal recessive Alport syndrome, no 3(IV) 5(IV) labeling is detected in the GBM (J) whereas 5(IV) is expressed in Bowman's capsule (K) and the basement membranes of the collecting ducts (L). In both types of Alport syndrome, 1(IV) is diffusely expressed in the GBM (E,I).

Schematic algorithm in case of Alport syndrome suspicion because of hematuria proteinuria.

Heidet L , and Gubler M JASN 2009;20:1210-1215

2009 by American Society of Nephrology

PROGNOSIS IN FEMALES
Characteristics of heterozygous girls and women belonging to the 195 families with proven COL4A5 mutation are compared with those of hemizygous boys and men. Hematuria was observed in 95% of carriers and consistently absent in the others. Proteinuria, hearing loss, and ocular defects developed in 75%,28%, and 15%, respectively. The probability of developing end-stage renal disease or deafness before the age of 40 yr was 12% and 10%, respectively, in female versus 90 an 80%, respectively, in boys and men. The risk of progression to end-stage renal disease appears to increase after the age of 60yr in women.
J Am Soc Nephrol 14: 26032610, 2003

INCIDENCE/PRESENTAT ION
Overall incidence is unknown but in USA, Alport accounts for 3% children with ESRD and 0.2% adult with ESRD Initial presentation: microscopic hematuria >>proteinuria >> ESRD Boys without hematuria by age 10 unlikely to have Alports.

Mutation
Patients with large deletions and nonsense mutations mutations have more severe disease. Risk of ESRD by 30 years 50% with missence mutations, 70% splice site mutations and 90% large deletion. Mutation located closer to the 5end of the gene associated with younger onset ESRD and increased risk of ocular changes and hearing loss closer to 3 end

Hearing
About 50% of male patients with XLAS show sensorineural deafness by age 25 years, and about 90% are deaf by age 40 years Rate of hearing loss is similar to progression to ESRD

Study of families with X-linked found hearing loss in 85% in boys and 18% in females. May reflect impaired adhesion of the Organ of Corti (which contain auditory sensory cells) to the basilar membrane of the inner ear.

Ocular Findings Anterior Lenticonus

Conical protrusion of the central portion of the lens into the anterior chamber.
It is most marked anteriorly because it is the region where the capsule is thinnest, the stresses of accommodation are greatest, and the lens is least supported. Occurs in approximately 15-20% of AS patients.

Treatment
ACE blockade: European Alport Registry followed a cohort of patient for 20 years and ACE inhibitor delayed dialysis in patients with proteinuria. To date, only small uncontrolled trials have demonstrated the effect of ACE inhibitors on reducing proteinuria in humans.

Preemptive therapy with ACE inhibitors in an 3(IV) knockout Alport mouse model prolonged lifespan until death from renal failure by more than 100%.
So a double-blinded RCT started recruiting pediatrics patient for early start of ace inhibitor (Safety and Efficacy of the ACE-Inhibitor
Ramipril in Alport Syndrome: The Double-Blind, Randomized, Placebo-Controlled, Multicenter Phase III EARLY PRO-TECT Alport Trial in Pediatric Patients)

Cohen, EP. In hereditary nephritis ACE inihibition decreases proteinuria and may slow the rate of progression. Am J Kidney Dis, 1996; 27:199. Gross, O et al. Preemptive ramipril therapy delays renal failure and reduces renal fibrosis in COL4A3knockout mice with Alport syndrome. KI 2003; 63: 438-446.

Early angiotensin-converting enzyme inhibition in Alport syndrome delays renal failure and improves life expectancy. Kidney International (2012) 81, 494501

Aggressive form of anti-glomerular basement membrane disease that targets the allograft in transplanted patients with XLAS Seen in approximately 3-5% of Alport patients post transplantation Usually occurs during the first year after the transplantation Devastating disease almost always destroys the allograft Retransplantation is rarely successful

Alport post-transplant anti-GBM nephritis (APTN)

Treatment Renal Transplant

For unclear reasons, certain patients are at very low risk for developing post-transplant anti-GBM nephritis, including patients with normal hearing, patients with late progression to ESRD, or females with XLAS. Unlike de novo anti-GBM nephritis, pulmonary hemorrhage is never observed because the patient's lung tissue does not contain the antigen. Treatment with plasmapheresis and cyclophosphamide is usually unsuccessful, and most patients lose the allograft.

Retransplantation in most patients results in recurrence of antiGBM nephritis despite the absence of detectable circulating antiGBM antibodies before transplantation.
Kashtan CE. Alport syndrome and thin glomerular basement membrane disease. J Am Soc Nephrol. 1998;9:1736.

Alport post-transplant anti-GBM nephritis (APTN)

Presenting symptoms

Morphology: mimics anti-GBM disease that develops in the setting of Goodpasture syndrome

hematuria rapid deterioration of renal function

LM: segmental glomerular necrosis and crescent formation with associated tubulointerstitial inflammation IF: liniar staining along the glomerular capillaries for IgG, kappa and lambda light chains, and linear or finely granular staining for C3 EM: absence of immune deposits and normal GBM ultrastructural organization

Renal Manifestations - ESRD

The risk of progression of renal failure is highest among males with XLAS and in both males and females with ARAS. ESRD develops in virtually all males with XLAS, usually between the ages of 16 and 35 years. Some evidence suggests that ESRD may occur even earlier in ARAS, whereas renal failure has a slower progression in ADAS. Approximately 90% of patients develop ESRD by age 40 years. The probability of ESRD in people younger than 30 years is significantly higher (90%) in patients with large rearrangements of the COL4A5 gene compared to those with minor mutations (50-70%).

Diagnosis that requires EM

Mark Hass. Touch Briefings.2007

No of biopsy Pearson 1994 Haas 1997 Wang 1998 Collan 2005 88 288 777 85

Essential for diag 25% 21% 18.5% 15.3%

Important 50% 24% 13.5% 58.8%

Not needed 25% 55% 68% 25.9%

Wagrowska 2007
Mubarak 2009 Darouich 2010

113
200 20

31%
43% 40% 23.5%

13.3%
51.5% 60% 41.7%

63%

Elhefnawy 2011 120

33.8%