Comparison of injury markers and total protein in perfusate from DBD (n = 17) versus cDCDD (n = 9) human donors.

<p>A: NGAL activity B: LDH activity C: Cytochrome c oxidase activity D: Total protein level Error bars represent Standard Error of the Mean (SEM). * p<0.05 using Student’s T test. DBD = Donation after Brain Death cDCDD = controlled Donation after Circulatory Determination of Death LDH = lactate dehydrogenase NGAL = neutrophil gelatinase associated lipocalin</p

Comparison of MMP-9 and MMP-2 levels in perfusates from kidneys from DBD (n = 15) and cDCDD (n = 9) human donors.

<p>Error bars represent Standard Error of the Mean (SEM). * p<0.05 using Student’s T test. DBD = Donation after Brain Death cDCDD = controlled Donation after Circulatory Determination of Death MMP = matrix metalloproteinase.</p

Level of undamaged mitochondria measured by the levels of cytochrome c oxidase activity in rat kidney <i>tissue</i> after initial flush and following 22 hours of machine cold perfusion with KPS-1, KPS-1 with doxycycline, and KPS-1 with MMP-2 siRNA (n = 4).

<p>Error bars represent Standard Error of the Mean (SEM). * p<0.05 compared to 0 hours using Student’s T test.# p<0.05 compared to 22 hours using Student’s T test.KPS-1 = Kidney Preservation Solution 1MMP = matrix metalloproteinase.</p

Light microscopy (H&E stain) of representative rat kidney at initial flush and after 22 hours of machine cold perfusion (n = 5 per time point).

<p>Light microscopy (H&E stain) of representative rat kidney at initial flush and after 22 hours of machine cold perfusion (n = 5 per time point).</p

Experimental ProtocolKPS-1 = Kidney Perfusion Solution -1 MMP = matrix metalloproteinase.

<p>Experimental ProtocolKPS-1 = Kidney Perfusion Solution -1 MMP = matrix metalloproteinase.</p

NGAL in perfusate of rat kidney with warm ischemic injury at 22 hours with KPS-1 and KPS-1 supplemented with doxycycline and MMP-2 siRNA compared to control (5 hours) (n = 4).

<p>Error bars represent Standard Error of the Mean (SEM). * p<0.05 compared to 5 hours using Student’s T test.# p<0.05 compared to 22 hours using Student’s T test.KPS-1 = Kidney Preservation Solution 1MMP = matrix metalloproteinaseNGAL = neutrophil gelatinase associated lipocalin.</p

LDH activity and Cytochrome c oxidase level in perfusate of rat kidney with warm ischemic injury at 5, 10, and 22 hours perfused with KPS-1 with scrambled siRNA versus KPS-1 with siRNA 50 nM (n = 4–7 each).

<p>A: LDH activity. B: Cytochrome c oxidase level. Error bars represent Standard Error of the Mean (SEM). * p<0.05 compared to 5 hours using Student’s T test. # p<0.05 compared to 22 hours using Student’s T test.</p

Effect of doxycycline on LDH, cytochrome c oxidase, and total protein release.

<p>A: Confirmation of inhibition of MMP-2 and MMP-9 activity by doxycycline using zymography. B: LDH activity in perfusate of rat kidney with warm ischemic injury at 5, 10, and 22 hours perfused with KPS-1 with and without doxycycline (n = 7 each). C: Cytochrome c oxidase level in perfusate of rat kidney with warm ischemic injury at 5, 10, and 22 hours perfused with KPS-1 with and without doxycycline (n = 7 each). D: Total protein release in perfusate of rat kidney with warm ischemic injury at 5, 10, and 22 hours perfused with KPS-1 with and without doxycycline (n = 6 each). Error bars represent Standard Error of the Mean (SEM). * p<0.05 compared to 5 hours using Student’s T test.# p<0.05 compared to 22 hours using Student’s T test.KPS-1 = Kidney Preservation Solution 1MMP = matrix metalloproteinase.</p

Matrix Metalloproteinase levels and injury markers in perfusate from clinically transplanted kidneys with DGF (n = 5) and without DGF (n = 19).

<p>A: MMP-9 level B: MMP-2 level C: NGAL levelD: LDH activity Error bars represent Standard Error of the Mean (SEM). * p<0.05 using Student’s T test. DGF = Delayed Graft Function MMP = matrix metalloproteinase NGAL = neutrophil gelatinase associated lipocalin.</p

Representative electron micrographs of rat kidneys and comparison of mitochondrial damage at the time of retrieval and after 22 hours of machine cold perfusion. (5K magnification).

<p>A: Clear arrows = mitochondrial swelling and loss of detail at 22 hours Dark arrows = swelling and detachment of the tubular endothelium at 22 hours. <b>B (n = 4).</b> Left panel = cytochrome c oxidaseRight panel = percent undamaged outer membrane of mitochondria. Error bars represent Standard Error of the Mean (SEM). * p<0.05 using Student’s T test.</p