The loop diuretic furosemide has an action to inhibit Na -K -2Cl co-transporter at the thick ascending limb of Henle's loop resulting in diuresis. Furosemide also has the non-diuretic effects by binding to GABA-A receptor which may involve the gastrointestinal tract. The aim of this study was to investigate the effects of furosemide on smooth muscle contractions in mice ileum and proximal colon. Each intestinal segment suspended in an organ bath was connected to a force transducer. Signal output of mechanical activity was amplified and recorded for analysis using PowerLab System. After equilibration, the intestine was directly exposed to furosemide, GABA, GABA-A receptor agonist (muscimol), or muscarinic receptor antagonist (atropine). Furosemide (50, 100 and 500 μmol L ) acutely reduced the amplitude of ileal and colonic contraction. In the ileum, 1 mmol L GABA and 10-60 μmol L muscimol significantly increased the amplitude, whereas in the colon, 50-100 mmol L GABA and 60 μmol L muscimol decreased the contractions. The contractions were also significantly suppressed by atropine. To investigate the mechanisms underlying the inhibiting effect of furosemide, furosemide was added to the organ bath prior to the addition of muscimol or atropine. A comparison of furosemide combined with muscimol or atropine group and furosemide group showed no significant difference of the ileal contraction, but the amplitude of colonic contraction significantly decreased when compared to adding furosemide alone. These results suggest that furosemide can reduce the ileal and proximal colonic contraction mediated by blocking and supporting of GABA-A receptor, respectively, resulting in decreased acetylcholine release.

Soluble (pro)renin receptor (sPRR), the extracellular domain of (pro)renin receptor (PRR), is primarily generated by site-1 protease (S1P) and furin. It has been reported that sPRR functions as an important regulator of intrarenal renin contributing to angiotensin II (Ang II)-induced hypertension. Relatively, less is known for the function of sPRR in Ang II-independent hypertension such as mineralocorticoid excess. In the present study, we employed a novel mouse model with mutagenesis of the cleavage site in PRR (termed as PRRR279V/L282V or Mutant) to examine the phenotype during aldosterone (Aldo)-salt treatment. The hypertensive response of Mutant mice to Aldo-salt treatment was blunted in parallel with attenuated response of plasma volume expansion, sodium-water retention and renal medullary α-epithelial sodium channel (α-ENaC) expression. Moreover, Aldo-salt-induced hypertrophy in the heart and kidney as well as proteinuria were improved, accompanied with blunted polydipsia and polyuria. Together, these results represent strong evidence favoring endogenous sPRR as a mediator of Aldo-salt-induced hypertension and renal injury.

Sepsis associated acute kidney injury (S-AKI) is a life-threating disease with poorly defined mechanisms and limited therapeutic options. Here we examined the role of site-1 protease (S1P)-derived soluble PRR (sPRR) in the S-AKI model induced by lipopolysaccharide (LPS). Male 10-week-old B6129SF1/J mice were pretreated with a S1P inhibitor PF-429242 (PF) at 20 mg/kg/day via minipump and treated with a single s.c. injection of LPS at 10 mg/kg and monitored for 24 hours. By radiotelemetry, LPS induced fall in mean arterial pressure (MAP) (Δ mmHg: -14.98 ± 4.13 in LPS+PF group vs . -23.15 ± 1.82 in LPS+ Vehicle group; n = 6, p < 0.05) and heart rate (HR) (Beats/min: -78.60 ± 20.73 in LPS+PF group vs . -133.35 ± 30.92 in LPS group, n = 6, p < 0.05), which was significantly attenuated by PF treatment. In parallel, renal dysfunction as reflected by the rises in plasma creatinine and BUN along with albuminuria, urinary excretion of NGAL and Kim-1 were all improved. Similar improvement was observed for infiltration of macrophage as assessed by immunofluorescence detection of F4/80 as well as renal mRNA expression of M1 markers such as iNOS, IL-1β, IL-6, and TNFα. The results from the pharmacological study were validated using a recently generated mouse model of mutagenesis of the cleavage site of PRR (termed Mutant mice). LPS-induced hypotensive response, renal dysfunction, the markers of renal injury and inflammation and M1 activation were all improved in Mutant mice as seen with PF treatment. LPS induced a 3.35-fold increase in plasma sPRR which was attenuated by PF or in Mutant mice. By immunofluorescence, LPS-induced PRR expression was largely restricted to renal medullary intercalated dells. In cultured RAW264.7 macrophage cells, recombinant sPRR treatment at 30 nM directly promoted macrophage migration and stimulated mRNA levels of M1 markers, all of which were attenuated by losartan. Taken together, the present study suggests that (1) S1P-derived sPRR contributes to LPS-induced septic shock and AKI, and (2) within the kidney, LPS-induced sPRR originates from intercalated cells and may act in a paracrine manner to activate AT1R on macrophages to induce pro-inflammatory response.

Aryl hydrocarbon receptor (AhR), a ligand‐activated transcription factor, plays a major role in detoxification of xenobiotics. AhR deficiency has been shown to attenuate renal injury induced by diabetes. It is interesting whether the beneficial effect of AhR deficiency on renal injury in diabetes involve in alteration of intracellular glucose in renal tubular cells. The aim of this study was to investigate the effect and mechanism of AhR on glucose uptake into human renal proximal tubular cells. Exposure HK‐2 cells, human renal proximal tubular cell line, with omeprazole, an AhR agonist, increased glucose transporter (GLUT)‐mediated [3H]‐2‐Deoxy‐D glucose uptake into the cells in concentration‐dependent manner. The stimulatory effect of omeprazole was reduced by co‐incubation with CH223191, an AhR antagonist. These results indicate that the stimulatory effect of omeprazole on GLUT transport function regards activation of AhR. The kinetic study showed that omeprazole significantly decreased half‐maximal transport (Kt) but not maximal rate of transport (Jmax) of GLUT, suggesting that AhR might increase affinity of GLUT‐mediated glucose uptake into the cells. In addition, activation of AhR significantly induced cellular level of ROS. Inhibition of ROS using N‐acetyl‐L‐cysteine (NAC) inhibited the stimulatory effect of AhR on GLUT‐mediated glucose uptake. These results support that activation of AhR increases GLUT‐mediated glucose accumulation via increase in ROS production in human renal proximal tubular cells. Inhibition of AhR might be a target for treatment of high glucose‐induced renal injury from diabetes.