Figure 4. Gain or loss of microRNA(miR)-16 function attenuated or increased, respectively, kidney function in mice.

(A) Quantitative analysis of miR-16 level after the kidney was infused with lenti-miR16 or lenti-pSin control (n = 6 mice per group as shown in the diagram). (B) Kidney functions assessed with or without miR-16 infusion following ischemia-reperfusion (I/R) injury. Bilateral renal arteries were clamped for 45 mins, and serum urea nitrogen and creatinine levels were measured 6 hours after reperfusion or sham surgery. Values are means ± SEM; n = 7 animals/group. *P < 0.05 compared to control groups. (C) Apoptotic kidney cells in mice infused with or without miR-16 using in vivo TUNEL staining. Without (left) or with infused miR-16 (right) mice underwent a sham operation (top) or 45 mins of renal clamping to induce ischemia, followed by 6 hours of reperfusion (bottom). TUNEL staining of representative kidney sections from each experimental group is shown. Colocalization of blue and brown staining in nuclei reflects apoptotic cells which are indicated with arrows. Bar = 50 μm. Proportions of TUNEL-positive renal epithelial nuclei to total nuclei in mice infused with or without miR-16 and subjected to the sham operation or I/R injury are shown. *P < 0.05; n = 7 animals/group. (D) Active caspase-3 protein expression in mouse kidney with or without lenti-miR-16 infection. Kidney lysates of mice subjected to the sham operation or I/R injury were probed with specific antibody against the cleaved, active form of caspase-3. Scanning densitometry was used for semi-quantitative analysis and compared to β-actin levels. Values are means ± SEM from three experiments. n = 3, **P < 0.01. (E) Urinary miR-16 level in mice after the kidney was infused with lenti-miR16 or lenti-pSin control following I/R. (F) Effect of Lenti-mediated gene transfer of antisense-miR-16 on I/R-induced renal function in mice. (G) Reduced urinal miR-16 content in mice infused with antisense-miR16 following I/R.