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The last decade has witnessed a phenomenal increase in our understanding of the biological role of lysophosphatidic acid (LPA) and has led to an appreciation of this critical serum-derived growth factor released from platelets. We herein summarize recent observations that collectively support the hypothesis that LPA may play a key role in the pathogenesis of initiation and progression of proliferative glomerulonephritis. LPA synergistically stimulates mesangial cell proliferation in combination with platelet-derived growth factor in primary culture. The mechanism of co-mitogenesis is likely to be mediated by the prolonged activation of mitogen-activated protein kinase which is stimulated by platelet-derived growth factor and LPA through different mechanisms. LPA contracts cultured mesangial cells and has properties in common with other pressor molecules including mobilization of intracellular Ca2+ and promotion of Ca2+ entry through dihydropyridine-sensitive calcium channels. LPA receptor mRNA has been identified in isolated glomeruli dissected from renal biopsy samples of patients with IgA nephropathy. All of these facts have led us to postulate that LPA is produced within glomeruli and that LPA's mitogenic as well as haemodynamic action contribute to the pathological process of mesangial proliferative glomerulonephritis. The possible production of LPA as an autocrine factor from mesangial cells themselves has also been discussed.

Lysophosphatidic acid (LPA) is a structurally simple, platelet-derived phospholipid, capable of eliciting a variety of physiological responses. We have demonstrated previously that LPA elicited a marked contractile response in rat mesangial cells (Inoue CN, Forster HG, Epstein M. Circ Res 77:888-896, 1995). In the present study, we examined the potential of this vasoactive substance to induce mesangial cell proliferation. Serum-starved quiescent rat mesangial cells were incubated with either LPA or in combination with platelet-derived growth factor (PDGF). DNA synthesis was assessed by [3H]thymidine incorporation after 24 hr, and cell numbers were determined at 0, 4, and 7 days. LPA- (1 nM-30 microM) stimulated mesangial cell DNA synthesis in a dose-dependent manner. The DNA synthesis stimulated by PDGF (1-100 ng/ml) was characterized by a bell-shaped response curve with a maximum at 40 ng/ml PDGF. The ability of LPA (30 microM) to synergize PDGF was observed over the entire range of PDGF concentrations (1-100 ng/ml). Under optimal concentrations of LPA/PDGF (30 microM40 ng/ml, respectively), mesangial cells displayed a 67-fold increase in [3H]thymidine incorporation, and a 1.9-fold (Day 4) and 2.5-fold (Day 7) increase in cell number as compared with that of quiescent mesangial cells. With an in vitro assay with myelin basic protein as the substrate, both LPA and PDGF induced stimulation of mitogen-activated protein (MAP) kinase activity. In addition, LPA augmented PDGF-induced increase in MAP kinase activity. In summary, these results demonstrate that LPA is mitogenic alone and also acts synergistically in combination with PDGF to promote mesangial cell proliferation. We postulate that these actions of LPA have the potential to play a crucial role in the mitogenic response of mesangial cells seen in a wide array of inflammatory and thrombotic glomerular disorders.

In the present study, we assessed the role of tissue angiotensin-converting enzyme as a determinant of intrarenal hemodynamics by using the angiotensin-converting enzyme inhibitor trandolaprilat and the angiotensin II receptor antagonist losartan. Afferent and efferent arteriolar diameters were measured with computer-assisted vessel imaging in isolated perfused hydronephrotic rat kidneys. In response to the addition of 1.0 nM angiotensin I, afferent arterioles constricted by 27.3 +/- 2.4% and efferent arterioles by 20.9 +/- 2.4%. These constrictions were similar to those observed after the administration of 0.3 nM angiotensin 11 (33.7 +/- 2.3% and 20.9 +/- 2.4% in afferent and efferent arterioles, respectively). Pretreatment with the angiotensin-converting enzyme inhibitor trandolaprilat (0.1-10 microM) blunted the angiotensin I-induced constriction of afferent arterioles (12.7 +/- 1.4%) and completely abolished the angiotensin I-induced constriction of efferent arterioles. Subsequent addition of angiotensin II to the perfusate resulted in a marked decrease of afferent (39.9 +/- 1.8%) and efferent (27.8 +/- 3.3%) arteriolar diameters. Pretreatment with the angiotensin II receptor antagonist losartan completely blocked the angiotensin I-induced constriction of both afferent and efferent arterioles. Collectively, these data suggest that angiotensin I affects renal microvessels through its conversion to angiotensin II, mediated by locally available tissue angiotensin-converting enzyme, which subserves the local control of the renal microcirculation.

By permitting the separation of increased aldose reductase activity from hyperglycaemia and insulin deficiency, galactose-fed rats have constituted a useful model for investigating diabetic complications. Such rats manifest an impaired afferent arteriolar responsiveness to pressure similar to that of rats 4 to 6 weeks after induction of diabetes with streptozotocin. In the present study, we investigated whether treatment of galactose-fed rats with the aldose reductase inhibitor tolrestat prevent this autoregulatory defect and whether the blunted afferent arteriolar responsiveness to pressure is associated with impaired responsiveness to angiotensin II. Pressure-induced vasoconstriction of afferent arterioles was assessed in kidneys made hydronephrotic to allow direct visualization of renal microvessels by computer-assisted image processing. Vessel diameters were quantitated following stepwise increments of renal perfusion pressure (RAP; from 80 to 180 mm Hg) in kidneys of control rats and rats fed a diet for 2 weeks with 50% galactose with or without tolrestat. Subsequent to the pressure studies, angiotensin II (0.3 nmol/l) was added to the perfusate, and vessel diameters were reassessed. Control rats exhibited progressive afferent arteriolar vasoconstriction when RAP was increased from 80 to 180 mm Hg (-17.2 +/- 1.0%; p < 0.001). In contrast, myogenic responses to increases in pressure were absent in the arterioles of the galactose-fed rats (-4.1 +/- 1.9%; N.S.). Treatment with tolrestat completely prevented this impairment in afferent arteriolar responsiveness (-16.5 +/- 1.8%; p < 0.001). The angiotensin II-induced vasoconstriction did not differ between control rats and galactose-fed rats. We conclude that increased aldose reductase activity contributes to impaired renal auto-regulation in galactose-fed rats, a model of diabetic nephropathy, but is not involved in the loss of afferent arteriolar responsiveness to angiotensin II.

Lysophosphatidic acid (LPA), the smallest and structurally simplest phospholipid, has the potential to modulate cellular signaling in diverse tissues and cell types, including fibroblasts. In the present study, we assessed the effects of LPA on cultured rat glomerular mesangial cells. Quantitative changes of [Ca2+]i in response to LPA were measured in monolayers of mesangial cells loaded with the fluorescent Ca2+ indicator fura 2. LPA (10 nmol/L to 100 mumol/L) increased [Ca2+]i in a dose-dependent manner and evoked inositol trisphosphate formation. LPA (1 mumol/L to 30 mumol/L) also elicited a marked, albeit transient, contractile response in mesangial cells cultured on collagen gel, as assessed by a decrease in cell surface area (CSA). The contraction persisted for 5 minutes (CSA decreased by 31%), whereupon the mesangial cells gradually relaxed with a consequent increase in CSA. Pretreatment of mesangial cells with isradipine (1 mumol/L), a dihydropyridine-sensitive Ca2+ channel blocker, completely blocked LPA-induced contraction. Isradipine also decreased resting [Ca2+]i levels as well as the peak and the subsequently sustained [Ca2+]i levels induced by LPA, suggesting that the contractile effects of LPA are dependent on Ca2+ entry through voltage-gated Ca2+ channels. Finally, LPA stimulated an increase in both prostaglandin E2 synthesis and cAMP accumulation, indicating that these mediators may modulate the contractile effects of LPA. Our study is the first demonstration that exogenous LPA induces mesangial cell contraction and suggests that the contraction is mediated by mobilization of intracellular Ca2+ by activation of the phosphoinositide cascade as well as by promotion of Ca2+ entry across the plasma membrane.

Previous studies from our laboratory have demonstrated impaired afferent arteriolar responsiveness to pressure in rats 4-6 weeks after the induction of diabetes mellitus. Although the responsible mechanisms mediating this renal autoregulatory defect have not been fully defined, increased polyol metabolism has been implicated as a possible factor involved in the pathogenesis of diabetic complications. We therefore investigated the possible role of this metabolic disturbance in renal autoregulation using the galactose-fed rat, a model characterized by increased polyol pathway activity independent of hyperglycemia or insulin deficiency. Hydronephrosis was induced to permit direct visualization of renal microvessels. Pressure-induced vasoconstriction of afferent arterioles was assessed by quantitating vessel diameter following stepwise increments of renal perfusion pressure (RAP; from 80 to 180 mm Hg) in the hydronephrotic kidneys from control rats and rats fed a 50% galactose diet for 2 or 4 weeks. Vessel diameters were measured from video images by computer-assisted image processing. Control rats exhibited progressive afferent arteriolar vasoconstriction when RAP was increased from 80 to 180 mm Hg (-17.3% +/- 1.0%; P < 0.001). In contrast, myogenic responses to increases in pressure were absent in the afferent arterioles of rats fed a 50% galactose diet for either 2 (-4.1% +/- 1.9%; not significant) or 4 weeks (-2.9 +/- 3.4%; not significant). Our demonstration that the impairment of afferent arteriolar responsiveness to increasing RAP in the normoglycemic galactose-fed rat was identical to that observed in the STZ-diabetic rat suggests that increased polyol accumulation may contribute to the impairment of renal autoregulation in the diabetic rat.

A study of the comparative survival of islet xenografts using a combination of treatment modalities was carried out in the spontaneously diabetic BB/Wor rat. Islets were isolated from hamster donors, and after 4 to 6 days of incubation the cells were injected either into the immunologically privileged abdominal testis or into the nonimmunologically favored renal subcapsular space. Postoperatively the rats were given cyclosporine at two different dose schedules. The results showed that islets injected into the abdominal testis of nonimmunosuppressed rats caused the induction of normoglycemia with a mean duration of 8.3 +/- 1.3 days. A significant prolongation of normoglycemia to a mean of 36.1 +/- 4.5 days occurred in rats that were given abdominal, intratesticular islet xenografts and cyclosporine for 30 days. The longest average survival in excess of a mean of 90.1 +/- 6.5 days was achieved in rats that were given abdominal, intratesticular islet xenografts and cyclosporine continuously, every other day. All of the grafted rats reverted to diabetes upon the cessation of cyclosporine. A similar cyclosporine regimen failed to prolong islet xenograft survival for longer than a mean of 9.0 +/- 2.2 days in rats that were given islet xenografts injected into the renal subcapsular space. Extended survival of abdominal, intratesticular islet xenografts corresponded with trough plasma and testis cyclosporine levels of 457 +/- 46 ng/mL and 643 +/- 45 ng/g, of wet weight, respectively. It is concluded that islet xenografts are protected against immune destruction in the BB/Wor rat with type 1 diabetes only as long as the cells are injected into an immunologically privileged site and the host is continuously immunosuppressed with cyclosporine.