Kidney International, Vol. 54, Suppl. 67 (1998), pp. S-195–S-198
Insulin-like growth factor-I restores microvascular autoregulation in experimental chronic renal failure JEN-JAR LIN, BURKHARD T¨ ONSHOFF, NATHALIE BOURIQUET, DANIEL CASELLAS, FREDERICK J. KASKEL, and LEON C. MOORE Department of Pediatrics, Department of Physiology and Biophysics, State University of New York, SUNY Stony Brook, New York, USA; Children’s Hospital, University of Heidelberg, Heidelberg, Germany; and Groupe Rein et Hypertension, Institut Universitaire de Recherche Clinique, Montpellier, France
Insulin-like growth factor-I restores microvascular autoregulation in experimental chronic renal failure. Impairment of autoregulation (AR) is associated with accelerated progression of chronic renal failure (CRF). As the bioavailability of insulin-like growth factor-I (IGF-I) is low in CRF, we investigated the effects of acute luminal application of 10 nM recombinant human IGF-I on AR in juxtamedullary (JM) afferent arterioles (AA) perfused in vitro with a blood solution [(;30% hematocrit (HCT)]. Studies were conducted in AA from adult male rats three to four weeks after five-sixths nephrectomy (Nx) by either surgical excision (N 5 7) or infarction (N 5 5) of two thirds of the remnant kidney; controls (N 5 6) had sham surgery. AA from both Nx groups exhibited marked hypertrophy and impaired AR responses (60 to 140 mm Hg perfusion pressure), features more pronounced in the infarction group. Responses to abluminal acetylcholine (10 mM) were similar in sham and excision groups but were significantly blunted in the infarction group. All groups vasodilated significantly after Ca-channel blockade (10 mM MnCl2). IGF-I restored AR in AA from both Nx groups (P , 0.05, analysis of variance) while it vasodilated AA from controls. These results suggest that IGF-I may protect the glomerulus from injury by maintaining autoregulatory control of renal blood flow, thereby slowing the progression of CRF.
Injury to the glomerulus by elevated intravascular pressures and flows is thought to play a major role in the pathogenesis of chronic renal failure (CRF) [1]. In rat models of CRF, progressive loss of renal function and glomerular sclerosis are accelerated by hypertension and impairment of renal blood flow autoregulation (AR) [2, 3]. Another complication in CRF is a reduction in the biological activity of insulin-like growth factor-I (IGF-I), as commonly manifested by reduced longitudinal growth [4]. In the rat, this appears to be related to increased hepatic production of IGF-I– binding proteins [4], as well as decreased renal clearance of IGF-I binding proteins and Key words: hemodynamics, afferent arteriole, juxtamedullary, rectomy, autoregulation of blood pressure, progressive CRF, injury.
© 1998 by the International Society of Nephrology
fragments. Excess IGF binding proteins sequester IGF-I, thereby limiting its binding to the IGF-I receptor ([5] has a comprehensive review). In addition to its effects on metabolism and growth, IGF-I has significant vascular activity. In healthy humans and rats, IGF-I vasodilates the renal vasculature and increases glomerular filtration rate [6, 7], and IGF-I has been proposed for therapeutic use in CRF [8]. In this study, we investigated the effects of acute IGF-I treatment on the reactivity of juxtamedullary (JM) afferent arterioles (AA) from rats with experimental CRF. The objectives were to determine if direct investigation of renal microvascular function is feasible in remnant kidneys and to compare the effects of IGF-I on AR in JM AA from normal and CRF rats. METHODS Experimental CRF was induced in adult male SpragueDawley rats (;200 g body weight) by five-sixths nephrectomy (Nx) under sodium pentobarbital anesthesia (50 mg/kg). As the method of renal mass reduction in the remnant kidney has been shown to influence the progression of CRF in rats [9], two thirds of the cortical mass of the left kidney was ablated, following decapsulation to preserve the adrenal glands, via either surgical excision of both poles or infarction of both poles by ligation of renal artery branches. The right kidney was removed. Controls underwent sham surgery. Experiments were conducted three to four weeks after Nx using the blood-perfused JM nephron technique developed by Casellas and Navar [10]. Following anesthesia (110 mg/kg Inactin), the left kidney was cleared via an aortic catheter, removed, and dissected to expose the perihilar cortex. Major arteries supplying the rest of the kidney were ligated. During dissection, the kidney was perfused with a gassed Krebs-bicarbonate-Ringer (KBR) solution containing 4% albumin. During measurements, the kidney was perfused with a blood solution prepared from fresh, washed red blood cells (collected from nonuremic donor rats) and resuspended in KBR-6% albumin
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Fig. 1. A perfused juxtamedullary nephron in a remnant kidney reduced by surgical excision. Visible are an afferent arteriole (A), efferent arteriole (E), glomerulus (G), and macula densa (M). The image is a composite of overlapping video fields. The bar in the upper left is 25 mm.
solution to a hematocrit of ;30%. The preparation was superfused with warmed (37°C) KBR solution with 1% albumin. The reactivity of the mid-AA segments was assessed using videomicroscopy to measure changes in lumen diameter. Recombinant human IGF-I (10 nM; Genentech) was applied luminally. Acetylcholine (10 mM; Sigma) was applied abluminally to verify basic functionality of the endothelial nitric oxide (NO) system. At the end of the studies, 10 mM MnCl2 was applied abluminally to determine relaxed vessel diameter. Statistical analysis was by one- or two-way analysis of variance (ANOVA), based on repeated measures (RM) where appropriate, followed by Student-Newman-Keuls multiple comparison tests; P , 0.05 was considered significant. RESULTS AND DISCUSSION Figure 1 shows a perfused JM nephron from a uremic rat three weeks after five-sixths Nx by surgical excision. Hypertrophy of the vessels in the remnant kidney is evident in the large diameters of the afferent and efferent arterioles. The macula densa region, including individual cells, is also clearly visible. Figure 2 summarizes the results of the AR studies in mid-AA segments. In comparison to the controls, basal AA diameters were significantly larger in both Nx groups, with the largest diameters seen in the infarction group (two-way ANOVA). This pattern was also evident following Ca channel blockade, which resulted in significant pressure-dependent vasodilation in all groups (twoway RM ANOVA). This indicates that vasodilatory reserve is not substantially reduced in JM AA in remnant kidneys when perfused in vitro. AR responsiveness was attenuated in both CRF groups. In the excision group, this was evident between 100 and 140
mm Hg, whereas in the infarction group, AA diameter was unchanged over the entire pressure range studied. In control AA, application of 10 nM IGF-I significantly shifted the autoregulation curve upward (two-way RM ANOVA), and autoregulatory responsiveness was significantly enhanced (comparison before and after IGF-I of diameter change between 60 and 140 mm Hg; paired t-test). In both groups of CRF rats, IGF-I significantly enhanced autoregulatory responsiveness (two-way RM ANOVA). In the excision group, this resulted in a significant vasodilation at 60 mm Hg and vasoconstriction at 140 mm Hg (RM ANOVA). In the infarction group, IGF-I caused significant vasoconstriction at 100 and 140 mm Hg (RM ANOVA). Figure 3 illustrates the responses to abluminal application of 10 mM acetylcholine. The vasodilatory responses were similar in the control and excision groups, suggesting that the endothelial NO system is functional in this model of CRF. In contrast, the responses in AA from rats subjected to renal infarction were blunted significantly (ANOVA). This study demonstrates the feasibility of direct study of the JM microvasculature in rat models of CRF. As abnormalities in intravascular pressures and flows play a major role in the pathogenesis of CRF, our techniques provide unique insight into the pathophysiology of intact AAs in remnant kidneys. The principle results show that AR ability is impaired in JM AA in remnant kidneys, that AR can be at least partly restored by acute IGF-I treatment, and that the method of five-sixths Nx can significantly influence the functional state of JM AA. The impairment of AR responsiveness in JM AA from CRF kidneys is consistent with other studies in intact rats: Bidani et al have demonstrated impaired AR in conscious
Lin et al: IGF-I restores autoregulation in CRF
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Fig. 2. Autoregulation responses in mid-afferent arterioles (AA) segments from rats subjected to five-sixths nephrectomy by surgical excision (N 5 7), cortical infarction (N 5 5), or control sham surgery (N 5 6). Shown are baseline responses (F) and responses during treatment with 10 nM insulin-like growth factor-I (IGF-I; E) or 10 mM MnCl2 to block calcium channels (f). Autoregulation in both chronic renal failure (CRF) groups was significantly impaired versus the sham controls [P , 0.05, two-way analysis of variance (ANOVA)] and was significantly enhanced by IGF-I in both CRF groups (P , 0.05, two-way repeated measures ANOVA).
Fig. 3. Responses to topical application of 10 mM acetylcholine in midafferent arterioles (AA) segments from rats with five-sixths nephrectomy via surgical excision or cortical infarction and in sham controls. The responses in the infarction group were significantly smaller than in the other two groups (P , 0.05, analysis of variance and Student-Newman-Keuls test).
five-sixths Nx rats and have found a strong association between systemic hypertension and the progression of glomerular sclerosis [2, 3, 11]. This agreement suggests that the behavior of JM AA parallels that in more superficial nephrons. Although the mechanism of AR dysfunction in CRF was not addressed in this study, our results do demonstrate that it is not dependent on the presence of uremic blood. Our results also show that the method of five-sixths Nx can have substantial impact on JM AA. Rats subjected to cortical infarction show greater vascular hypertrophy and AR dysfunction and reduced endotheliumdependent vasodilation ability than rats subjected to surgical excision. Differences in the severity and progression of glomerular sclerosis have been reported between these two models of CRF, with slower progression and less severe hypertension in the surgical excision model [9]. The most important finding in our study is that acute exposure to IGF-I at least partially restores AR ability in AA from CRF rats. In addition, whereas IGF-I vasodilates normal kidneys, it vasoconstricts JM AA from rats with CRF at pressures at or above normotensive levels. These studies offer no insight into the mechanism of IGF-I action in CRF. However, studies in normal rats have implicated
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both NO and vasodilatory prostanoids [5], and studies in cultured endothelial cells show that IGF-I stimulates endothelin release [12]. We have recently reported evidence consistent with the participation of all three agents [13]. Nevertheless, our studies suggest that IGF-I treatment may be able slow the progression of CRF by restoring some degree of AR control of glomerular capillary pressure and flow. Further study is required to evaluate this interesting possibility. ACKNOWLEDGMENTS Recombinant human IGF-I was supplied by Genentech, Inc. B. To ¨nshoff and N. Bouriquet were recipients of Fellowships from the New York State Affiliate of the American Heart Association. J.-J. Lin was a recipient of a Fellowship from the National Kidney Foundation of New York/New Jersey.
APPENDIX Abbreviations used in this article are: AA, afferent arterioles; ANOVA, analysis of variance; AR, autoregulation; CRF, chronic renal failure; IGF-I, insulin-like growth factor-I; JM, juxtamedullary; KBR, Krebsbicarbonate-Ringer; NO, nitric oxide; Nx, nephrectomy; RM, repeated measures. Reprint requests to Leon C. Moore, Ph.D., Department of Physiology and Biophysics, SUNY HSC, Stony Brook, New York 11794-8661, USA. E-mail:
[email protected]
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