Reduced renal Na+-K+-Cl- co-transporter activity and inhibited NKCC2 mRNA expression by Leptospira shermani: from bed-side to bench.
BACKGROUND
Renal involvement is common in leptospirosis. Interstitial nephritis with interstitial oedema and mononuclear cellular infiltration are the usual findings. Clinically, non-oligouric acute renal failure, hypokalaemia and sodium wasting appear frequently in leptospirosis. The outer membrane protein from leptospira has been thought to be responsible for the disorder. However, the exact mechanisms of renal involvement are still unclear.


METHODS
To address these questions, we first performed detailed in vivo clearance tests in three patients with leptospirosis (Leptospira shermani) and hypokalaemia to define the tubular lesion. These tests indicated a defective Na(+)-K(+)-Cl(-) co-transporter and a poor response to furosemide infusion. We performed further in vitro studies in a model of medullary thick ascending limb (mTAL) cultured cells derived from normal mouse.


RESULTS
Outer membrane protein extract from L.shermani (0.3 microg/ml) inhibited Na(+)-K(+)-Cl(-) co-transporter activity in mTAL cells (control, 10.15+/-0.52; L.shermani, 6.47+/-0.47 nmol/min/mg protein). The basolateral Na(+)-K(+) ATPase remained intact. Reverse transcription-polymerase chain reaction (RT-PCR) further revealed that the outer membrane protein extract from L.shermani downregulated Na(+)-K(+)-Cl(-) co-transporter (mNKCC2) mRNA expression. These changes were dose dependent and could be reversed by the antibody against outer membrane protein extract from L.shermani. Experiments with a less pathogenic strain of leptospira (L.bratislava) and Escherichia coli did not affect Na(+)-K(+)-Cl(-) co-transporter activity.


CONCLUSIONS
We conclude that L.shermani leptospirosis downregulates mNKCC2 mRNA expression and inhibits Na(+)-K(+)-Cl(-) co-transporter activity in TAL cells. These alterations may explain the observed electrolyte disorders in leptospirosis.
Wu
Mai-Szu
MS
Division of Nephrology, Chang Gung Memorial Hospital, 222, Mai-Chin Road, Keelung, Taiwan. maxwu1@adm.cgmh.org.tw
Yang
Chih-Wei
CW
Pan
Ming-Jen
MJ
Chang
Chiz-Tzung
CT
Chen
Yung-Chih
YC
D000199:Actins / Q000235:genetics; D000328:Adult; D000368:Aged; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000494:pharmacology; D001692:Biological Transport; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005260:Female; D006801:Humans; D007008:Hypokalemia / Q000382:microbiology; D007106:Immune Sera / Q000494:pharmacology; D007919:Leptospira / Q000276:immunology; D007922:Leptospirosis / Q000150:complications / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D008138:Loop of Henle / Q000382:microbiology / Q000473:pathology; D008297:Male; D051379:Mice; D008875:Middle Aged; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012413:Rubidium / Q000493:pharmacokinetics; D049994:Sodium Potassium Chloride Symporter Inhibitors; D028021:Sodium-Potassium-Chloride Symporters / Q000235:genetics; D064466:Solute Carrier Family 12, Member 1
D000199:Actins; D001425:Bacterial Outer Membrane Proteins; D007106:Immune Sera; D012333:RNA, Messenger; C578714:SLC12A1 protein, human; C578715:Slc12a1 protein, mouse; D049994:Sodium Potassium Chloride Symporter Inhibitors; D028021:Sodium-Potassium-Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; D012413:Rubidium