endemic nephropathy and hypertension · 2012. 3. 28. · jelakovi ćet al kidney int 2012 the...

ENDEMIC NEPHROPATHY AND

HYPERTENSION

Bojan Jelakovi ć

University of ZagrebSchool of Medicine

Department of Nephrology, Hypertension and Dialysi sUniversity Hospital Center Zagreb

ESH European Hypertension Center of Excellence

1. ENDEMIC NEPHROPATHY FOR BEGINNERS

ENDEMIC (BALKAN) NEPHROPATHY IS ARISTOLOCHIC ACID NEPHROPATHY AND IS A WORLDWIDE PROBLEM

2. ENDEMIC NEPHROPATHY AND HYPERTENSION

ENDEMIC NEPHROPATHYENDEMIC NEPHROPATHY

- Chronic tubulointerstitial nephritis

- Danube river tributaries(South East Europe- Balkan)

-Known since 1950s

- WHO 1964 and 1974 ENDEMIC NEPHROPATHY

-- Chronic tubulointerstitial Chronic tubulointerstitial

nephritisnephritis

-- Danube river tributariesDanube river tributaries

(South East Europe(South East Europe

-- Balkan)Balkan)

--Known since Known since 1951950s0s

-- WHO 1964 and 1974 WHO 1964 and 1974


- certain villages- mosaic distribution- family aggregation - households

ENDEMIC NEPHROPATHYENDEMIC NEPHROPATHY-- Prevalence diseased ~ 2 Prevalence diseased ~ 2 -- 3 % (!)3 % (!)

suspect of having EN ~ 1suspect of having EN ~ 100 --1515 % (!)% (!)

- approx 25.000 diseased and approx. 100.000 at risk

-- m : f = 1 : 1.2m : f = 1 : 1.2

-still present in all endemic villages (?)

-never in children (> 18 y.o.)

-the age distribution was shifted to older ages

(is etiological agent still present/active ?)


1. DISEASED of EN:- proteinuria < 1 g/day and / or LMW (beta2m or alfa1m) - anemia Hb < 120 g/l (m) < 113 g/l (f)- renal impairment Serum creatinine > 132.6 µmol/l- positive family history

2. “SUSPECT” of having EN- proteinuria and/or LMW - anemia or positive family history

3. “AT RISK” of EN- positive family history

4. OTHERS

1.1. DISEASED DISEASED of EN:of EN:

-- proteinuria < 1 g/day and / or LMW (beta2m or alfa1m) proteinuria < 1 g/day and / or LMW (beta2m or alfa1m)

-- anemia Hb < 120 g/l (m)anemia Hb < 120 g/l (m) < 113 g/l (f)< 113 g/l (f)

-- renal impairment Serum creatinine > 132.renal impairment Serum creatinine > 132.6 6 µµmol/lmol/l

-- positive family historypositive family history

22. . ““ SUSPECTSUSPECT”” of having ENof having EN

-- proteinuria and/or LMW proteinuria and/or LMW

-- anemia or positive family historyanemia or positive family history

33. . ““ AT RISKAT RISK ”” of ENof EN

-- positive family historypositive family history

4. OTHERS4. OTHERS

per exclusionemper exclusionemper exclusionem

residence in endemic area for more than 15 years

Proximal renal tubule cells �leading to interstitial fibrosis and chronic renal failure

urothelium � cancer

Two target tissues

- good preservation of glomeruli - a gradient of tubular atrophy - interstitial fibrosis that decreases in severity from outer-to-inner cortex

Endemic Nephropathy

• hypocellular interstitial fibrosis decreasing from outer to inner cortexdecreasing from outer to inner cortex

cortical tubular atrophytubular atrophy involving the proximal proximal tubule tubule

juxtaglomerular juxtaglomerular cells arecells are atrophicatrophic in early in early phase of diseasephase of disease

•the medulla appears relatively well preserved

•glomeruli relatively spared compared to the severity of tubulointerstitial fibrosis

Endemic Nephropathy

Insidious onset, progression to ESRD

Polyuria, nocturia

Normal BP, hypertension rareNormal BP, hypertension rare

Anemia (more severe than the CKD stage)

Upper urothelial cancers Tubular proteinuria (beta2, alfa1)

Tubular enzymuria (NAG, AAP)

Renal salt wasting

Impaired concentrating capacity

Decreased GFR

Final answerFew questions

Cosyns first called attentionto SIMILARITIES in the pathologic features of

ARISTOLOCHIC ACID NEPHROPATHYand

ENDEMIC NEPHROPATHY

Misidentification of the plant/substitution by anot her more toxi c compound.

In 1990 Aristolochia fangchi GUANG FANG JIFANG JI

was inadverently put into the slimming tea

INSTEAD OF

Stephania tetrandra HAN FANG JIFANG JI

In 1992, ~100 young women following a “slimming” regime at a Brussels spa clinic presented with end stage renal failure and, later, many of them developed urothelial cell dysplasia or cancer.

HOW ???

ETIOLOGY OF ENDEMIC NEPHROPATHY

1969

M. Ivić

Liječnički vjesnik 1969; 91:1273

investigation in Niš, Serbia

published for the first time in Liječnički vjesnik, Zagreb, Croatia

M. Ivić, Liječnički vjesnik 1969

M. Ivić, Liječnički vjesnik 1969

FIELD INVESTIGATION

... some fields of wheat in nephropathy area in the watershed of the river Južna Morava were heavily contaminated by a weed that I had never seen in the corn-fields of Vojvodina

...there were many fields without of stem of A.clematitis, but there were also fields highly contaminated by A.c., sometimes so much that the whole field became green with weed...

The wheat is used only by the peasants, the owners of the fields.(familial occurrence of EN)

... during the harvest on no occasion did I see that reapers remove the cut A.c...

... furthermore the peasants do not even know that this herb is poisonous...

... the peasants brought grain contaminated by A.c. for grinding into flour.

“Obviously, the bread of those peasants was also poisonous.”

Remarkably, over next 35 years NO ATTEMPT was madeto confirm or follow-up Ivić s prescient observation !

What a pity !What a pity !

You are a fool until your idea becomes a success.

Mark Twain

OR = 7.2

95% CI: (1.8, 29.0)

p-value < 0.01

Aristolochia in Farm Fields

Always or SometimesNever

Fre

quen

cy

20

18

16

14

12

10

8

6

4

2

GROUP

case

outside_control

Q: A. clematitis in farmfields 20 yrs ago?

Hranjec, 2005

1. Seeds of Aristolochia clematitis co-mingle with the wheat grainused by villagers in endemic regions to prepare bread.

OR = 12.1

95% CI: (2.7, 55.5)

p-value = 0.002

Aristolochia in Wheat

Always or SometimesNever

Fre

quen

cy

20

10

0

GROUP

case

outside_control

Q: A. clematitis in harvestedwheat 20 years ago?

A. clematitis Wheat

Hranjec, 2005

1. Seeds of Aristolochia clematitis co-mingle with the wheat grainused by villagers in endemic regions to prepare bread.

>>> 1 seed (1 gr 6.5 mg AA) / 1 kg bread10-15 years

Aristolactam - DNA adducts in renal cortex of Croatian patients with EN

PNAS, 20070.8 – 5.9 dA-AA adducts/10 7 Nt.0.2 – 6.2 dG-AA adducts/10 7 Nt.

PNAS, 2007

p53 mutational spectra in transitional cell carcinomasEN vs. the IARC p53 database

The Incredulity of Saint Thomas

Caravaggio, 1601 - Sanssouci, Potsdam

No PT SEX BORN VILLAGE COUNTRY DG

1MB F 1927 KANIŽA CROATIA

CA URETERIS LAT DEX

2AG M 1940 SL.KOBAŠ CROATIA

CA PYELONIS LAT SIN

3DM F 1930 SL.KOBAŠ CROATIA

CA PYELONIS ET URETERIS LAT DEX

4 EB F 1923 SL.KOBAŠ CROATIA


5PM M 1929 TOLISA BOSNIA

CA URETERIS LAT DEX

6 JČ F 1921 PRUD BOSNIACA PYELONIS LAT DEX

7 AF F 1934 SL.KOBAŠ CROATIACA PYELONIS LAT DEX

8 IG M 1945 SL.KOBAŠ CROATIACA PYELONIS LAT SIN

9 TF M 1932DOMALJEVAC BOSNIA

CA PYELONIS LAT SIN

10 KI F 1930 KANIŽA CROATIA

CA PYELONIS LAT SIN

11

RĐ F 1935

DOMALJEVAC BOSNIA


ARISTOLACTAM -DNA ADDUCTS IN RENAL CORTEX OF

SERBIAN PATIENTS WITH UPPER UROTHELIAL TRACT CANCERS

dA adducts/10 9 nts nd9 nd nd9 5 26971000 10 nd

std AZ ZMMR RP DB RB SK SM ZT AM SMSample ID

dG-AL

dA-AL

Nikolić, 2010

Croatian patient with EN (paraffin embedded tissue)

20 25 30 35Time (min)

0

50

100

0

50

100

Rel

ativ

e A

bund

ance

0

50

100

m/z200 300 400 500

412.2

292.4

395.3280.3

259.1

412.2

292.2

395.2

384.2

369.2

tR: 27.08A: 1,005

tR: 27.22A: 16,673

dA-AL-IMS3 427 →

dA-AL-IMS3 427 →

Exposed kidney DNA

Exposed kidney DNANegative control (unexposed kidney DNA)

Positive Control(dA-AL-I0ligonucleotide)

Positive Control(dA-AL-I 0ligonucleotide)

dA-AL-IMS3 427 →

Jelaković et al Kidney Int 2012

The chemical Identification and Quantitation of ALThe chemical Identification and Quantitation of AL--DNA Adducts was DNA Adducts was performed by LCperformed by LC--ESI/MS/MSESI/MS/MS33

Mass spectroscopic characterization of DNA-aristolactam (AL) adducts in the renal cortex

MUTATIONAL SPECTRA OF TP53 GENE

unpublished data

Endemic nephropathy Aristolochic acid nephropathy

Pathology identical

Clinical course difference - dose dependent

Prevalence 2-4% 3-5%of exposed population

Incidence of 30 - 50%urothelial cancer

Familal aggregation yes noyes

chronic type - China

initially normal blood pressureearly, severe anemiaaseptic leukocyturiamild LMW proteinuraprogression to ESRD

AAN and EN

1. ENDEMIC NEPHROPATHY FOR BEGINNERS

ENDEMIC (BALKAN) NEPHROPATHY IS ARISTOLOCHIC ACID NEPHROPATHY AND IS A WORLDWIDE PROBLEM

2. ENDEMIC NEPHROPATHY AND HYPERTENSION

HYPERTENSION IN ENDEMIC NEPHROPATHY

... Blood pressure is normal , and in some subjects hypertension develops in the uraemic phase

Cvoriscec et al. Clin Chem Clin Lab 1998;36:271

... patients do not suffer from edema, and their blood pressure is usually normal .Djukanovic and Radovanovic, Clinical Nephrotoxins 2003

... progressive renal failure unaccompanied by salt retention or hypertension .Stefanovic et al, Am J Nephrol 2006;26:1

... the disease is characterized by tubular proteinuria,... profound anemia and abscence of hypertension and edema. Batuman Kidney Int 2006;69:644

Mild and labile hypertension is occasionally observe d in the early stage of the disease, but becomes more frequent 30-40% in advanced renal failure

Stefanovic and Cosyns, Oxford Textbook of Nephrology 2003

Endemic Nephropathy

Why is hypertension rare in early phase ?

1. chronic tubulointerstitial nephritis that starts in cortex and gradually spread to inner parts (proximal tubule)

2. salt wasting nephropathy

3. destruction of juxtaglomerular cells(decreased renin synthesis)

4. renal medulla is not affected(preserved synthesis of PGE2 and PGI2)

Could aristolochic acid be related to later developement of hypertension in EN ?

1. epidemiologic evidence-both in “Chinese herbs nephropathy” and in EN hypertension is not characteristic of early phase and occurs in advanced stages of CKD !

2. pathologic evidence - it starts from outer cortex(PGE2 and PGI2 synthesis preserved)

3. aristolochic acid damages proximal tubule (especially S3 segment - place where ACE is mostlysynthesized in the kidney)

4. aristolochic acid destroys macula densa

Is prevalence of hypertension lower in endemic vill ages compared to other rural areas in Croatia ?

6 endemic, 2 non-endemic villages

Door-to-door (consecutive sample)Participation rate 91%> 18 y.o

Cross-sectional epidemiologic survey

1728 inhabitants (693 men 1035 women)

Dika et al, Kidney Blood Press Res, submitted

Exclusion criteria

Pregnancy

Terminal illnes (bed-ridden)

Severe disability

Limb amputation

Mental illnes

Dementia

DiabetesNephrotic proteinuira


Epidemiologic questionnaire

- past medical history,- family history- past exposure to aristolochic acid- life style habits- socioeconomic status

Clinical examBP – mercury sphygmomanometer (and Omron device)

(home and outpatient measurements)- ESH/ESC guidelines

LaboratoryeGFR – MDRD formulaAlbuminuria - Morning spot urine sample Alpha1 MG - Morning spot urine sample


Criteria for classification of inhabitants of endemic villages:a) positive family (household) history of EN; b) tubular proteinuria (α1-microglobulin >10 mg/L or >14 mg/g creatinine); c) serum creatinine >132.6 µmol/L; d) anemia (hemoglobin <120 g/L in males; <113 g/l in females); e) exclusion of other chronic kidney diseases

Diseased of EN a+b+c+d+e, or b+c+d+e, or a+b+d+e. Suspect of having EN a+b+e or b+d+e At risk for EN healthy subjects who have been living in the EN a)“Others“ inhabitants of EN villages who did not fulfillcriteria

Hypertension = BP≥140/90 mmHg, and/or current use of antihypertensive drugsTreatment of hypertension = usage of antihypertensive medication Control of hypertension =BP< 140/90 mmHg (under pharmacological treatment

Low income = monthly income < 400 EurosLow educational level = < 8 years of schooling


Control villages(X±SD)

Endemic villages(X±SD)

t p

Sex M/F (n) 145/211 496/ 750 0.098* 0.754

Age (years) 53.71± 17.24 50.87 ± 16.94 2.781 0.005

SBP (mmHg) 148.26 ± 28.74 145.43 ± 26.08 1.673 0.095

DBP (mmHg) 85.04 ± 12.89 85.24 ± 12.84 -0.259 0.795

BMI (kg/m2) 27.65 ± 5.35 27.01 ± 5.04 1.993 0.046

Hbg (g/L) 136.93 ± 14.30 136.13 ± 15.33 0.880 0.379

Cr (s) (µmol/L) 84.66 ± 53.63 93.07 ± 73.65 -2.386 0.017

eGFR (ml/min/1.73 m2) 71.74 ± 16.70 69.49 ± 19.17 2.002 0.045

Demographic and laboratory characteristics of the study population1602 inhabitants were considered eligible


EN villages Control villages χ2 P

Prevalence (%)

Total 50.8 46.5 1.879 0.005

Male 52.6 53.5* 0.058 0.811

Female 49.7 42.1 6.107 0.014

Treatment (%)

Total 36.2 37.9 0.276 0.6

Male 28.4* 27.3* 0.116 0.734

Female 41.6 46.4 2.415 0.12

Control of all hypertensives (%)

Total 7.1 7.6 0.042 0.837

Male 7 8.1 0.348 0.555

Female 7.2 7.2 0.014 0.908

Control of treated hypertensives (%)

Total 19.7 20 0.002 0.96

Male 24.7* 29.6* 3.222 0.073

Female 17.4 15.5 0.581 0.446

Prevalence, treatment and control of hypertension in endemic and control villages

*P<0.05 for differences between men and women Dika et al, Kidney Blood Press Res, submitted

DiseasedN=40

SuspectedN=76

At riskN=385

OthersN=745

Controlvillages N=356

F P

SexF/M

25/15 35/41 244/141 446/299 211/145 8.177* 0.085

Age

(years)

72.78(6,44)

61.54(15,69)

48.26(15.29)

49.95(17.05)

53.71(17.24)

30.840 <0.001

SBP

(mmHg)

163.2(27.2)

160.2(28.3)

144.5(23.6)

143.4(26.1)

148.2(28.7)

12.537 <0.001

DBP

(mmHg)

85.3(13.9)

90.1(15.2)

85.9(12.1)

84.3(12.7)

85.0(12.8)

3.899 0.004

BMI (kg/m2)

26.80(5.25)

26.79(4.85)

27.51(5.10)

26.78(5.01)

27.65(5.35)

0.060

Demographic characteristics of included subgroups


Diseased(N=40)

Suspected(N=76)

At risk(N=385)

Others(N=745)

Controlvillages (N=356)

F P

Cr (s)µmol/L

379.45(271.29)

103.84(49.88)

82.27(14.28)

82.18(16.21)

84.66(53.63)

315.813 <0.001

eGFR ml/min/1.73 m2

19.69(15.6)

56.38(15.7)

71.84(15.1)

72.29(17.23)

71.74(16.7)

111.247 <0.001

α1MGNmg/L

Med. 79.3 16.15 5.18 5.18 5.37

324.015 <0.001Min -Max

10.2-398 10.1-91.8 2.2-23 2.25-133 5-119

Albumin mg/L

Med. 56.2 16.35 5.47 6.12 5.78

135.561 <0.001Min -Max

3.7-700 2.59-208 2-342 2-482 2-300

Laboratory characteristics of included subgroups


Diseased Suspected At risk Others P

Total 86.0 72.7 54.3 45.8 <0.001

Male 87.5 70.8 58.4 46.6 <0.001

Female 85.2 75 52 45.3 <0.001

Total 56.8 32.8 36.2 35.0 0.061

Male 50.0 20.6* 29.8 27.4* 0.218

Female 60.9 46.7 40.1 40.3 0.243

Total 0 4.7 6.6 8.3 0.21

Male 0 2.9 7.7 7.9 0.528

Female 0 6.7 6 8.6 0.391

Total 0 14.3 18.4 23.8 0.061

Male 0 14.3 25.8* 28.8* 0.362

Female 0 14.3 14.9 21.4 0.204

Prevalence, treatment and control of hypertension in EN village subgroups

Prevalence (%)

Treatment (%)

Control of all hypertensives (%)

Control of treated hypertensives (%)


Hypertension prevalence in endemic population according to WHO criteria (only subjects with GFR > 60 ml/min)

Hypertension prevalence is higher in suspects of havi ng EN than in other farmers.

Prevalence of obesity (BMI > 30) in endemic populat ionand control non EN village

Obesity was the most prevalent in suspects.

Prevalence of hypertension regarding to BMIin EN population

Obesity influences HT prevalence in EN population

Women vs. Men

Women Men pPrevalenceEN villages 49.7 52.6 NS

TreatmentEN villages 41.6 28.4 <0.001control villages 46.4 27.3 <0.001

Control of treated HTEN villages 17.4 24.7 0.002 control villages 15.5 29.6 0.002

Education level < 8y 67.6 48.9 <0.001Income < 400Eu 73.4 61.8 <0.001

RR for hypertensionLow level of education 1.48 (CI 95% 1.35 – 1.62) P<0.001Low income 1.23 (CI 95% 1.12 – 1.62) P<0.001

No differences in age and BMI


1. The prevalence of hypertension is high in the Croatian endemic rural area.

2. Hypertension is not only a clinical feature of the advanced phase of EN, but also of all EN subgroups.

3. Low income and level of education, besides renal impairment and BMI, are important risk factors for hypertension in endemic villages.

4. Low socioeconomic status is a strong risk factor for hypertension and contributes not only to higher prevalence, but also to poorer control of hypertension, especially in women from EN villages

CONCLUSION

The evolution of changes in hypertension in EN is a nice example and a good lessonof how important environmental factors for development of hypertension are.

PAST

1.More exposure to aristolochic acid

2.->Proximal tubule damageDestroyed macula densa

3. ->Salt wasting nephropathyRAS

4. Late development of hypertension

---> HT is not an early feature of EN

PRESENT

1.Advances in agriculture-> exposure to nephrotoxin diminished

disappeared2. -> No tubular damage3. -> No salt wasting

4. Changes in lifestyle- obesity- stress (poor socioeconomic status)

--> Higher (earlier) prevalence of HT in EN

Acknowledgement sZvonimir Medverec urology General Hospital Sl.Brod, CroatiaKrunoslav Jakovina pathology General Hospital Sl.Brod, CroatiaTratinčica Jakovina pathology General Hospital Sl.Brod , CroatiaMato Vukelić pathology General Hospital Sl.Brod, CroatiaNinoslav Leko nephrology General Hospital Sl.Brod, CroatiaAnte Cvitković epidemiology Marica Miletić-Medved epidemiology Insitute for Public Health, County of Sl.Brod, Croatia

Croatian Center for Endemic NephropathyDamir Dittrich urology General Hospital S. Brod, CroatiaFran Borovečki functional genomics School of Medicine, University of Zagreb, CroatiaSandra Karanović nephrology School of Medicine, University of Zagreb, CroatiaIvana Vuković Lela nephrology School of Medicine, University of Zagreb, CroatiaŽivka Dika nephrology School of Medicine, University of Zagreb, CroatiaDubravka Čvorišćec clinical laboratory University Hospital Center Zagreb, CroatiaIvan Pećin internal medicine University Hospital Center Zagreb, CroatiaAnamarija Kovač Peić nephrology General Hospital Sl.Brod , Croatia Jelena Kos nephrology University Hospital Center Zagreb, CroatiaMaja Mišić cytology General Hospital Sl.Brod, CroatiaKarla Tomić pathology General Hospital Sl.Brod , Croatia Branko Brdar molecular biology Institute Ruđer Bošković, Zagreb, CroatiaNeda Slade molecular biology Institute Ruđer Bošković, Zagreb, CroatiaIvan Habuš organic chemistry Institute Ruđer Bošković, Zagreb, Croatia

Francis Johnson organic chemistry SUNY, USARob Turesky mass spectrometry U of Albany, USAKate Dickman epithelial cell biology SUNY, USATom Rosenquist genetics SUNY, USAShinya Shibutani rodent model of AAN SUNY, USAHeidi Einholf drug metabolism Novartis, USAFred Miller pathology SUNY , USA

Jiri Zavadil microarray analysis NYU, USABruce Weir biostatistics U of Washington, USAKaren Edwards genetic epidieiology CDC U of Washington, USA

Želimir Stipančić nephrology General Hospital Odžak, Bosnia and HerzegoviniaJovan Nikolić urology Urology Clinicl Belgrade, Serbia

Supported by NIEHS, the Fogarty Center and the Ministry of Science (Croatia)

Co PI Arthur P GrollmanBojan Jelaković

Croatian endemic area, harvest 2007

Aristolochia clematitis

endemic nephropathy and hypertension · 2012. 3. 28. · jelakovi ćet al kidney int 2012 the...

Documents