In Vivo Visualization of the Antialbuminuric Effects of the Angiotensin-Converting Enzyme Inhibitor Enalapril

Abstract

Angiotensin-converting enzyme (ACE) inhibitors are commonly used antiproteinuric drugs. Here we assessed the effect of the ACE inhibitor enalapril on the glomerular sieving coefficient of albumin (GSC A ) using intravital multiphoton microscopy. Munich Wistar Frömter (MWF) rats were used as a model of hypertension-related glomerular lesions. Young (9-week-old) MWF rats were nonproteinuric, similar to what was observed in control Wistar rats. However, urinary albumin excretion in the MWF rats gradually increased during aging, averaging 0.00062 6 0.0001 at age 9 weeks and 0.0054 6 0.0003 (mg/mOsmol per liter) at age 52 weeks (P , 0.0001). Albuminuria in aged MWF rats was accompanied by structural changes, which were indicative of glomerular lesions. The GSC A was low in young MWF rats but increased markedly during aging, averaging 0.00057 6 4.7 Â 10 25 (n 5 25) in young MWF rats and 0.0027 6 0.00036 in 52-week-old MWF rats (n 5 36; P , 0.0001).Treatment of proteinuric 12-month-old MWF rats with enalapril over a 4-week period reduced the GSC A from 0.0027 6 0.00036 to 0.00139 6 0.00013 (P 5 0.0005). Similarly, urinary albumin excretion was reduced, averaging 0.0051 6 0.0003 and 0.0036 6 0.0005 mg/mOsmol per liter before and after enalapril administration, respectively (P 5 0.0089). In parallel, enalapril treatment reduced the mean arterial blood pressure (144.6 6 6.5 mm Hg in untreated versus 110.9 6 0.6 mm Hg in enalapril-treated MWF rats) and increased the glomerular filtration rate from 1.64 6 0.3 ml/min to 3.58 6 0.3 ml/min (P 5 0.0025 versus baseline). In summary, enalapril reduced the GSC A in proteinuric MWF rats, which was paralleled by a similar reduction in urinary albumin excretion. These data suggest that glomerular rather than tubular mechanisms account for the beneficial antiproteinuric effects of the ACE inhibitor.

Intravital multiphoton microscopy as a tool for studying renal physiology and pathophysiology

“…In theory this would facilitate quantitation by removing the need for background subtraction. However in actuality, setting pixels to zero decreases sensitivity and compromises the detector’s ability to detect low intensity values[19, 20]. We conducted an extensive in vitro and in vivo study on this phenomenon and demonstrated a continued reduction in detector sensitivity as more and more pixels are set to zero.…”

“…As the number of pixels at zero increases, the decrease in sensitivity becomes exponential [17]. This results in erroneous data and incorrect conclusions [16, 19, 20]. See figure 9.…”

Live‐Animal Imaging of Renal Function by Multiphoton Microscopy

“…While it might seem appropriate to adjust detector offset such that pixel values of zero are obtained in the absence of fluorescent probes (Schiessl and Castrop, 2013, Schiessl et al, 2015), such aggressive adjustment of detector offset not only eliminates detection of dim fluorescence, but it also artificially depresses estimates of fluorescence background, resulting in erroneously low measures of GSC. Accurate measures of GSC depend upon adjustment of detector offset such that positive values are obtained in regions lacking fluorescent probe, so that background levels are estimated of means of a distribution of pixel values that haven’t been artificially truncated to zero during the collection process.…”

In vivo microscopy

“…1A–C) to label the filtrate and measure the permeability of the glomerular filtration barrier to various macromolecules including dextrans, ficoll, albumin, and angiotensinogen. [2,20,28–30,32–35]. Importantly, this approach also helped to visualize the podocytes in negative (Fig.…”