Effects of melanocortin 1 receptor agonists in experimental nephropathies.

Nephrotic syndrome, characterized by massive proteinuria, is caused by a large group of diseases including membranous nephropathy (MN) and focal segmental glomerulosclerosis (FSGS). Although the underlying mechanisms are beginning to unravel, therapy is unspecific and far from efficient. It has been suggested that adrenocorticotropic hormone (ACTH) has beneficial effects in patients with MN and possibly in other nephrotic diseases. We have previously reported that ACTH may act directly on podocytes through the melanocortin 1 receptor (MC1R). In the present study, we evaluate the effect of highly specific MC1R agonists in two different nephrotic disease models. Experimental MN: Passive Heymann nephritis (PHN) was induced in rats that were treated for four weeks with MS05, a selective MC1R agonist, or saline. The degree of albuminuria was significantly reduced over time and the effect was sustained one week after treatment withdrawal (p<0.05). Experimental FSGS: Based on a dose-response study, two doses of adriamycin were used for induction of nephropathy in Balb/c mice. Mice were treated with either a synthetic MC1R agonist (BMS-470539), with α-melanocyte stimulating hormone (α-MSH) or with saline. There was no beneficial effect of treatment. In summary, MC1R agonists reduce albuminuria and improve morphology in experimentally induced MN whereas they have no effect in experimental FSGS. The results illustrate the differences in these podocytopathies in terms of signaling mechanisms underlying proteinuria, and progression of disease

MC1R protein expression.

<p>Western blotting was performed in order to detect MC1R protein expression in mouse and human tissue. Left panel: incubation with a rabbit polyclonal MC1R antibody (Alomone Labs). Right panel: incubation with MC1R antibody and control blocking peptide antigen (BP) in order to show antibody specificity. A) mouse whole glomerular lysate, B) cultured wild type mouse podocytes and C) a human malignant melanoma cell line A375. MC1R protein expression was detected at the expected size of 37 kDa (left panel).</p

Glomerular morphology was disrupted in adriamycin treated mice.

<p>Kidneys were collected for morphological analysis at day 12 after adriamycin induced nephropathy. Slides were blindly examined by a pathologist (n = 15 for all groups). Shown are representative images of (A) controls, which present a normal glomerular structure and foot processes, (B) untreated adriamycin mice, displaying a high degree of foot process effacement, (C) treatment with BMS-470539 or (D) α-MSH, showing disrupted glomerular structures similar to untreated adriamycin mice. (E) The number of podocyte foot processes per 10 µm of glomerular basement membrane was quantified. Untreated adriamycin (AD) and mice treated with BMS-470539 (BMS) or α-MSH had a lower number of foot processes compared to controls (Ctrl). *Normal foot process, → disrupted glomerular barrier structure and loss of foot processes. Scale bar = 2 µm. FP = foot processes, GBM = glomerular basement membrane.</p

Albuminuria was reduced in MC1R agonist treated PHN rats.

<p>After four weeks of MS05 treatment (n = 17), the urinary albumin- to-creatinine ratio (UACR) was significantly reduced compared to untreated PHN (n = 14; p<0.01). One week after treatment withdrawal (week 5), albuminuria was further reduced in MS05-treated (n = 17) compared to untreated PHN rats (n = 13; p<0.05). Results are presented as geometrical mean ± SEM.</p

MC1R agonists did not reduce albuminuria in adriamycin-treated mice.

<p>(A) Five different doses of adriamycin, in the range of 5–10 mg/kg, were intravenously injected and the level of albuminuria was measured after 7 days. The urinary albumin-to-creatinine ratio (UACR) increased with increasing levels of adriamycin. (B) Albuminuria at day 7. Treatment with the MC1R agonist BMS-470539 did not reduce the level of albuminuria for the 8 mg/kg adriamycin dose, on the contrary it slightly increased albuminuria for the 10 mg/kg dose (n = 17–19, p<0.05). The unspecific melanocortin receptor agonist α-MSH did not have any significant effect (n = 9, n.s.) compared with untreated adriamycin mice. Results are presented as mean ± SEM.</p

Blood urea nitrogen (BUN) does not differ in untreated adriamycin mice compared with MC1R agonist treated mice.

<p>BUN was measured in plasma samples taken from mice 8–10 days after injection of adriamycin. There was no difference between the groups (n = 9–10, n.s.). Results are presented as mean ± SEM.</p