We showed decreased medullary R2* values in patients with diabetic nephropathy compared to healthy volunteers. Our results differ from those in several other BOLD MR imaging studies of diabetes in animal models and in patients. Both Santos et al (15
) and Ries et al (14
) found increased R2* values corresponding to decreased oxygenation in the rat kidneys in the first several days following the induction of diabetes. Epstein et al (16
) and Economides et al (17
) did not find any differences in baseline renal medullary R2* values between patients with diabetes but without overt nephropathy and healthy control. In contrast, we found lower medullary R2* values suggesting higher oxygenation in our patients with diabetic nephropathy. We believe the discrepancies may be explained in part by the differences in the stages of diabetic renal injury that were investigated. In the two rat BOLD MR studies, the kidneys were studied within a few days of induction of diabetes, and likely exhibited glomerular hyperfiltration which is known to occur early on in the kidneys due to diabetes (20
). While the exact causes of renal hypoxia at this stage of diabetes are uncertain, it has been suggested that different factors may contribute toward the observed hypoxia in animal model of diabetes. One such factor is the increased oxygen consumption to support sodium reabsorption during the hyperfiltration stage, and the other is oxidative stress with augmented oxygen consumption (21
). It is possible that the observed renal hypoxia in the two rat BOLD MR studies may be in part due to the glomerular hyperfiltration during early stage of diabetes. In the human studies by Epstein et al (16
) and Economides et al (17
), the patients had mean duration of diabetes greater than 5 years but had not developed overt nephropathy. The glomerular filtration rate in those cases was normal or close to normal. In contrast, the patients in our study had already developed presumed diabetic nephropathy with reduced glomerular filtration rate and protein in the urine. The associated decrease in tubular sodium reabsorption may therefore have contributed to the reduced oxygen consumption and the apparent increase in oxygenation in the renal medulla in our study. The relationship between tubular sodium reabsorption and oxygen consumption has been evaluated in previous study which showed improvement in renal medullary oxygenation following administration of furosemide which reduces tubular sodium reabsorption (19
Complex alteration in renal oxygenation through different stages of renal injury has been documented by invasive means in other form of chronic renal disease. In an experimental model of chronic tubulointerstitial disease induced by ischemia-reflow injury, both the hypoxic marker pimonidazole and the hypoxia inducible factor (HIF) expression level were higher in the kidneys with moderate tubulointerstitial damage when compared to more severely damaged kidneys, indicating less renal hypoxia with more severe renal damage (23
). This paradoxical improvement in hypoxia with severe renal injury may be related to a more pronounced reduction of glomerular filtration rate in the more severely injured kidneys, with subsequent lower oxygen consumption from tubular sodium reabsorption (23
). The results from this experimental model of chronic kidney disease lend support to our findings of the apparent increase in renal medullary oxygenation with long standing diabetic nephropathy, possibly due to the diminished tubular sodium reabsorption and reduced oxygen consumption.
In addition to the alteration of tubular reabsorption being a possible contributing factor to the apparent increase in medullary oxygenation in our patients with diabetic nephropathy, another potential contributing factor may be related to the BOLD MR imaging technique. It is important to note that although changes in tissue oxygenation can be reflected as changes in R2* values, changes in R2* values do not necessarily correlate with changes in tissue oxygenation. BOLD MR imaging indirectly measures tissue oxygenation by measuring changes in the deoxyhemoglobin concentration in the adjacent capillaries. R2* values are a measurement of capillary deoxyhemoglobin concentration, which is thought to be in equilibrium with that of the surrounding tissue under normal circumstances. However, the kidneys in long standing diabetic nephropathy as well as in other forms of chronic kidney disease are frequently affected by tubulointerstitial fibrosis and depletion of peritubular capillaries (24
). The tubulointerstitial fibrosis characterized by the deposition of extracellular matrix will likely interfere with oxygen diffusion from the capillary to the surrounding tissue (24
). The depletion of peritubular capillaries will likely further impair oxygen delivery from blood to tissue. This may lead to reduced oxygen extraction by the tissue, and possibly non-equilibrium between capillary and tissue oxygenation. Therefore, the R2* values, which is a measurement of capillary deoxyhemoglobin concentration, may be decreased (corresponding to increased capillary oxygenation) in the face of renal tissue hypoxia because of the reduced oxygen extraction from the capillaries.
The apparent increase in renal medullary oxygenation observed in our patients has also been reported in other forms of chronic kidney disease studied by BOLD MR imaging. In a study of patients with renal artery stenosis, the investigators reported reduced medullary R2* values suggesting increased oxygenation in the atrophic and non-functioning kidneys distal to the stenosed renal artery compared with normal kidneys without renal artery stenosis or to functioning kidneys downstream from mild renal artery stenosis (11
). In a study of renal transplants, the investigators noted reduced R2* values corresponding to an apparent increase in oxygenation in transplants with chronic allograft nephropathy compared with normal functioning transplants (5
). In yet another study, the investigators reported decreased medullary R2* values corresponding to an apparent increase in oxygenation in patients with chronic tubulointerstitial nephropathy compared to healthy control(13
). The diseased kidneys in these studies likely had features common to most forms of chronic kidney disease including reduced tubular reabsorption function and tubulointerstitial fibrosis. We speculate that these factors may have also contributed towards the apparent increase in renal medullary oxygenation as measured by BOLD MR imaging in these studies.
When we subdivided the patients with diabetic nephropathy into those with mild disease (estimated GFR ≥ 60 ml/min per 1.72m2) and those with moderate to severe disease (estimated GFR < 60 ml/min per 1.72m2), we found lower medullary R2* values with worsening chronic kidney disease. The results from this subgroup analysis should be considered preliminary given the small sample size. Also because of the small sample size, we were not able to correlate the medullary R2* values with individual stages of chronic kidney disease (National Kidney Foundation 5 stages of chronic kidney disease) or with individual estimated GFR.
We performed a multivariate logistic regression model analysis which revealed significant independent association between a decrease in medullary R2* values and the degree of kidney disease categorized as none, mild, or moderate to severe. We did not find the age of the subject to be independently associated with a decrease in medullary R2* values, even though the mean age of the healthy volunteers was significantly younger than that of patients with diabetic nephropathy. This is in agreement with a previous study which showed no significant differences in baseline renal medullary R2* values between healthy young and healthy elderly subjects (26
We did not find any significant differences in R2* values in the renal cortex between patients with diabetic nephropathy and healthy volunteers. A potential explanation is that unlike the renal medulla which functions in relative hypoxia even under physiologic conditions, the renal cortex is better oxygenated and may be less susceptible to injury caused by diabetes (27
). Another potential explanation relates to the property of the hemoglobin dissociation curve. Because the renal cortex functions at an oxygen tension greater than 50 mm Hg under normal physiologic condition, the hemoglobin in the cortex is on a shallow portion of its dissociation curve (28
). Hence a small change in oxygenation in the cortex does not affect the hemoglobin saturation to the same degree as in the medulla, and may not be detectable with BOLD MR imaging.
Our study has several limitations. First, the sample size was relatively small. Despite the small number of subjects, however, there were significant differences in the renal medullary R2* values between patients with diabetic nephropathy and healthy control. Of note, both the medullary and cortical R2* values in the healthy volunteers in our study were in close agreement with what had been reported previously (16
). Second, we did not have kidney biopsy data to prove that the underlying cause of CKD was indeed classic diabetic glomerulosclerosis / nephropathy in all of our patients. But the diagnosis of diabetic nephropathy is often made on clinical grounds without a renal biopsy. It is possible that other co-morbidities such as nephrosclerosis may have also contributed to the underlying renal parenchymal disease and the BOLD MR imaging findings in our study. As discussed previously, many forms of chronic kidney disease, such as those from diabetes, hypertension and renal vascular disease, share similar features including reduced glomerular filtration rate and tubulo-interstitial fibrosis. Therefore, it is not likely that the potential effect of other co-morbidities on the kidneys would significantly alter our results. Third, the potential effect on R2* values by ACE inhibitors or angiotensin receptor blockers use in our patients with diabetic nephropathy was not investigated. These drugs have been shown in animal model of renal injury to reduce oxygen consumption, probably by reducing oxidative stress and improving the efficiency of oxygen utilization for sodium reabsorption (30
). Deng et al demonstrated that angiotensin II blockade normalized renal oxygen consumption in the remnant kidney model of chronic kidney disease in rats which had markedly increased oxygen consumption before treatment (30
). However, in a BOLD MRI study of renal allograft with chronic allograft nephropathy, the use of angiotensin receptor blocker did not alter the renal R2* value (5
). In our study, it is possible that the apparent increase in renal medullary oxygenation in the diabetic patients may be in part due to the use of ACE inhibitors or angiotensin receptor blockers. It is, however, unlikely that the use of these drugs alone will correct the medullary oxygenation to a level that is higher in diabetic nephropathy than in healthy control. Future BOLD MR imaging studies are needed to assess the effect of these drugs on renal oxygenation in patients with diabetic nephropathy and other forms of chronic kidney disease by obtaining scans prior to and after the initiation of these drug therapies. Fourth, in the 3 patients with very advanced kidney disease (GFR < 20 ml/min per 1.72m2
) and poor visualization of corticomedullary differentiation on the T1-weighted reference images, it is possible that volume averaging between the cortex and medulla may have occurred due to less precise ROI placement. This could underestimate the R2* values in the medulla. Diminished corticomedullary differentiation on T1-weighted images in patients with renal insufficiency has been described previously, and was thought to be primarily related to an increased T1 relaxation time of the cortex (31
). In that study, visualization of corticomedullary differentiation was poor when GFR was < 20 ml/min. In general, differentiation of the renal cortex from medulla may be limited in cases of advanced renal disease with very low GFR. In our study, however, we were able to visualize sufficient, albeit reduced, cortiomedullary differentiation in 17 of 20 patients on the GRE T1-weighted anatomical reference images. We believe our results were unlikely to be changed significantly by the few cases where there was poor corticomedullary differentiation. Fifth, another potential factor that may alter BOLD MR signal is the hematocrit level. Because BOLD MR technique depends on the presence of hemoglobin, the R2* values may be affected by abnormally low hematocrit level where there may be reduced sensitivity of the MR signal. However, the mean hematocrit level of 41% in our patients with diabetic nephropathy would fall under the normal range for hematocrit, and therefore is unlikely to have contributed significantly to our observation.
In conclusion, we found decreased medullary R2* values in patients with diabetic nephropathy compared to healthy volunteers. The etiology for the decrease in medullary R2* values corresponding to an apparent increase in medullary oxygenation in diabetic nephropathy is not clear. We speculate that it may be related to an alteration in oxygen consumption or possibly to factors inherent to BOLD MR imaging techniques. Future studies with correlative measurement of tissue oxygenation by invasive means are needed to better understand the relationship between the BOLD MR imaging findings and intra-renal oxygenation in a chronic kidney disease model.