The key finding of the present study is that urinary transferrin was significantly related with subclinical atherogenesis, particularly in patients with diagnosis of t2DM, who had not developed renal disease yet. This was primarily evidenced by a positive correlation between urinary transferrin and CIMT, being particularly significant for subjects with t2DM. Such correlation results in a potentially useful tool for clinical assessment of ED/vascular damage in patients with very early diabetic nephropathy.
Our primary endpoint was to explore whether transferrinuria was related with early ED, known to occur during renal hyperfiltration phase and just before clinically significant events develop, like microalbuminuria and the decline of glomerular filtration rate.
Microalbuminuria has been traditionally used for the assessment of early renal damage in patients with t2DM. However, approximately 30% to 45% of t2DM patients may show a diminished GFR (glomerular filtration rate), even in the absence of albuminuria. Therefore, clinical characterization of new biomarkers is required . To date, the role of transferrinuria as a marker of ED/vascular damage in patients with t2DM without known renal disease has not been described.
Urinary transferrin, as well as several urine biomarkers such as ceruloplasmin, immunoglobulin G, podocalyxin, neutrophil gelatinase-associated lipocalin (NGAL), N-acetyl-beta-glycosaminidase, α-1-microglobulin, 8-hydroxy-deoxyguanosine, tumor necrosis factor-alpha (TNF-α), interleukin-18 and cystatin C, have shown to reflect early renal damage progression when assessed altogether with microalbuminuria . However, urinary transferrin alone may also correlate with early vascular damage reflected by ED and subclinical atherogenesis, leading to arterioles inability of adaptive vasodilation and to progressive increase of vascular stiffness . This indicates that urinary transferrin is an early marker of vascular disease, occurring even before overt microalbuminuria in patients with t2DM.
In the present study, patients showed an apparently “normal” GFR without microalbuminuria. The relation between GFR and microalbuminuria, during very early phases of diabetic renal damage is not yet clear. Previous reports describe that early decline of renal filtration rate may occur in the absence of microalbuminuria in approximately 30% to 45% of cased with t2DM, which may be associated to an early renal hyperfiltration rate, denoting renal ED.
Consistently, a concept of a “renal reserve” has been proposed, which describes that a reserved renal function can be utilized when the physiological demand for single nephron GFR increases. This concept also supports that in subclinical renal disease like t2DM, at stage before basal GFR begins to reduce, renal functional reserve can be recruited in a manner that preserves renal function. The extension of this concept is that once a decline in basal GFR can be detected, renal disease is already well progressed . We acknowledge that we did not perform a direct measure of renal ED, such as determination of vasoactive mediators, either in plasma or urine. However, studies of patients on hemodialysis, showing that regression of CIMT occurs after renal transplant , as well as the role of carotid plaques as predictors of renal outcomes in individuals with t2DM without other microvascular complications  strongly suggest the relation between ED and accelerated subclinical atherogenesis, as assessed by CIMT.
While the relation of urinary transferrin with early renal damage and subclinical atherogenesis is not completely clear, possible explanation is acting through different pathological mechanisms. Although the underlying problem often cannot be treated, extensive experimental models and human studies suggest that progressive CKD may be largely due to secondary factors that are sometimes unrelated to the activity of the initial disease . Some of these secondary factors include: (a) compensatory response to nephron loss by maintaining total GFR; (b) direct endothelial cell damage, like that induced by systemic hypertension; and c marked tubulointerstitial injury (tubular dilatation, interstitial fibrosis), even if the primary process is a glomerulopathy .
Zylka et al. found the highest relationship between biomarkers associated with glomerular damage (in comparison to albuminuria) and diminished GFR, evaluated in a population similar to our study. Such biomarkers included transferrin, immunoglobulin G, ceruloplasmin, type IV collagen, glycosaminoglycans, prostaglandin D synthase lipocalin type, fibronectin, vascular endothelial growth factor, cystatin C, and nefinina . Besides, Kim et al. described a correlation between tubulointerstitial damage in t2DM with nephropathy, including different biomarkers such as urinary transferrin, that could lead to evaluate histopathological damage . Therefore, the correlation of urinary transferrin with ED suggests that glomerular and tubulointerstitial damages are related with vascular dysfunction, and urinary transferrin may be a useful marker of such dysfunction.
A specific study is required to determine whether urine leakage of transferrin and albumin may occur in the absence of endothelial damage. Unfortunately, the present study design did not allow exploring such specific mechanism of damage.
In addition, our results suggest that either the urinary transferrin or the urinary/index transferrin ratio are useful to identify patients with diabetic nephropathy at early risk for ED/vascular damage. These findings suggess the dynamic participation of the whole transferrin metabolism as related with early renal and vascular damages. The clinical usefulness of urine transferrin and plasma/urine transferrin ratio in the evaluation of early renal damage and subclinical atherogenesis, as a part of a routine test to identify high risk population with t2DM, are to be developed, and deserves further evaluation.
Interestingly, ED was more associated with several t2DM-related variables than urine transferrin during exploratory risk factor analyses. Since this analysis requires that variables are dichotomic and considering the significant correlation between urine transferrin and CIMT, it is suggested that optimal cutoff is still to be established, which at the present study design was not able to address.
The main limitation of our study was the heterogeneous characteristics between cases and controls within the study population, particularly in terms of age. However, no significant association was found between age and FMD nor CIMT values that could affect the study results. Another limitation was the determination of albuminuria with a bedside urine dipstick test in a urine single sample. Since this method is more sensitive for albumin (even with the false positive, false negative results that can occur), it resulted adequate for our analysis. Finally, we used urine transferrin measured in a single morning urine sample, which did not show significant difference with 24 h urine transferrin.
In conclusion, urine transferrin may be related with early ED and subclinical atherogenesis in patients with t2DM at a very early renal damage. This relation may be useful to stratify patients at higher vascular risk as well as early nephropathy, even before microalbuminuria becomes evident.