Angiotensin II type 1-receptor activating antibodies in renal-allograft rejection (authors reply inN Engl J Med. 2005 May 12;352(19):2027-8)

BACKGROUND: Antibodies against HLA antigens cause refractory allograft rejection with vasculopathy in some, but not all, patients. METHODS: We studied 33 kidney-transplant recipients who had refractory vascular rejection. Thirteen had donor-specific anti-HLA antibodies, whereas 20 did not Malignant hypertension was present in 16 of the patients without anti-HLA antibodies, 4 of whom had seizures. The remaining 17 patients had no malignant hypertension. We hypothesized that activating antibodies targeting the angiotensin II type 1 (AT1) receptor might be involved. RESULTS: Activating IgG antibodies targeting the AT1 receptor were detected in serum from all 16 patients with malignant hypertension and without anti-HLA antibodies, but in no other patients. These receptor-activating antibodies are subclass IgG1 and IgG3 antibodies that bind to two different epitopes on the second extracellular loop of the AT1 receptor. Tissue factor expression was increased in renal-biopsy specimens from patients with these antibodies. In vitro stimulation of vascular cells with an AT1-receptor-activating antibody induced phosphorylation of ERK 1/2 kinase and increased the DNA binding activity of the transcription factors activator protein 1 (AP-1) and nuclear factor-κB. The AT1 antagonist losartan blocked agonistic AT1-receptor antibody-mediated effects, and passive antibody transfer induced vasculopathy and hypertension in a rat kidney-transplantation model. CONCLUSIONS: A non-HLA, AT1-receptor-mediated pathway may contribute to refractory vascular rejection, and affected patients might benefit from removal of AT 1-receptor antibodies or from pharmacologic blockade of AT 1 receptors

EphrinB2 activation enhances vascular repair mechanisms and reduces brain swelling after mild cerebral ischemia

Objective- Cerebral edema caused by the disruption of the blood-brain barrier is a major complication after stroke. Therefore, strategies to accelerate and enhance neurovascular recovery after stroke are of prime interest. Our main aim was to study the role of ephrinB2/EphB4 signaling in mediating the vascular repair and in blood-brain barrier restoration after mild cerebral ischemia occlusion/reperfusion. Approach and results- Here, we show that the guidance molecule ephrinB2 plays a key role in neurovascular protection and blood-brain barrier restoration after stroke. In a focal stroke model, we characterize the stroke-induced damage to cerebral blood vessels and their subsequent endogenous repair on a cellular, molecular, and functional level. EphrinB2 and its tyrosine kinase receptor EphB4 are upregulated early after stroke by endothelial cells and perivascular support cells, in parallel to their reassembly during neurovascular recovery. Using both retroviral and pharmacological approaches, we show that the inhibition of ephrinB2/EphB4 signaling suppresses post-middle cerebral artery occlusion neurovascular repair mechanisms resulting in an aggravation of brain swelling. In contrast, the activation of ephrinB2 after brain ischemia leads to an increased pericyte recruitment and increased endothelial-pericyte interaction, resulting in an accelerated neurovascular repair after ischemia. Conclusions- We show that reducing swelling could result in improved outcome because of reduction in damaged brain tissue. We also identify a novel role for ephrinB2/EphB4 signaling in the maintenance of the neurovascular homeostasis and provide a novel therapeutic approach in reducing brain swelling after stroke

Renal ischemia and transplantation predispose to vascular constriction mediated by angiotensin II type 1 receptor-activating antibodies

BACKGROUND: We previously described angiotensin II type 1 receptor-activating antibodies (AT1R-Abs) in renal transplant recipients with vascular rejection and malignant hypertension. In this study, we tested the hypothesis that AT1R-Abs can cause renal artery contraction by AT1R activation with renal ischemia representing a key permissive factor and therefore contribute to renal pathologic condition. METHODS: Isolated renal and mesenteric arteries from Lewis rats were incubated with purified AT1R-Abs from patients with human leukocyte antigen antibody-negative vascular rejection. Vascular contraction was measured using small vessel myography. The measurements were repeated with renal arteries derived from native kidneys subjected to ischemia-reperfusion or after transplantation in a low-responder Fischer 344-to-Lewis rat kidney-transplantation model. RESULTS: AT1R-Abs acted in a vascular bed-specific manner and caused small contractions only in native rat renal arteries but not in mesenteric arteries. AT1R-Abs did not alter the vascular reactivity to phenylephrine, angiotensin II, or acetylcholine in native renal arteries. In contrast, AT1R-Abs caused a pronounced (>10-fold) contraction of renal arteries after ischemia and after allogeneic transplantation. Pretreatment with pharmacologic AT1R blocker only partially inhibited the AT1R-Abs-induced contraction, which was almost completely abolished by neutralizing peptides targeting epitopes of AT1R-Abs on the second loop of AT1R. CONCLUSIONS: These data demonstrate that AT1R-Abs can induce renal vascular contraction under predisposing conditions such as in ischemic or transplanted kidneys. Neutralizing antibodies against specific epitopes in the AT1R can ameliorate this contraction

Lectin-like oxidized low-density lipoprotein (LDL) receptor (LOX-1)-mediated pathway and vascular oxidative injury in older-age rat renal transplants

BACKGROUND: Older-age renal allografts are associated with inferior survival; however, the mechanisms are unclear. Reactive oxygen species participate in aging and in chronic vascular disease. We investigated how mediators of oxidative stress may increase allograft susceptibility to vascular injury. METHODS: We employed the low-responder allogeneic F344-to-Lew rat renal transplantation model. We used nonimmunosuppressed young (donors and recipients aged 12 weeks), old (donors and recipients aged 52 weeks), and old-to-young animal (donors aged 52 weeks and recipients aged 12 weeks) combinations. Grafts were transplanted after 2 hours cold preservation in University of Wisconsin solution and harvested 1, 2, 7 and 10 days later. Additionally, old animals receiving continuous 1.5 mg/kg cyclosporine (CyA) immunosuppression were included. Renal allograft pathology was scored according to Banff criteria. We studied intragraft vascular adhesion molecule-1 (VCAM-1), lectin-like oxidized low-density lipoprotein (LDL) receptor-1 (LOX-1), and hypochlorite-modified LDL expression as well as ED-1+ monocytes/macrophages and CD8+ lymphocyte infiltration. Intragraft in situ superoxide anion radical production was determined with dihydroethidium assay on cryosections. RESULTS: During the first 2 posttransplant days, old transplants demonstrated higher functional impairment and increased oxidative stress, while young transplant had higher ED-1+ monocytes/macrophage infiltration and VCAM-1 expression. The degree of VCAM-1 expression and ED-1+ monocytes/macrophage and CD8+ lymphocyte infiltration correlated at later time points directly with the transplant age. VCAM-1 and LOX-1 staining were localized predominantly on the endothelium of arterial vessels, shifting the distribution to vascular smooth muscle layer strongly dependent on donor age and the grade of vascular injury. LOX-1 staining colocalized with hypochlorite-modified epitopes in the media of injured arteries. We measured increased in situ superoxide anion radical production in corresponding areas. Immunosuppression with CyA had no protective effect on vascular injury and LOX-1 expression. CONCLUSION: Induction of LOX-1-related oxidation pathways and increased susceptibility to oxidative stress could play an important role in promoting vascular injury in old renal transplants independent of the recipient age

Cytoprotective actions of FTY720 modulate severe preservation reperfusion injury in Rat Renal transplants

BACKGROUND: Fingolimod (FTY720) is a potent agonist of sphingosine 1 phosphate receptors and thereby interferes with lymphocyte trafficking. We previously showed that FTY720 protects from mild preservation reperfusion injury induced by 4 hr of cold ischemia. The purpose of this study was to explore the role of FTY720 in ischemic injury and regeneration using a clinically relevant rat renal transplant model with 24 hr of cold ischemia. METHODS: Donor kidneys were cold stored in the University of Wisconsin solution for 24 hr before transplantation into bilaterally nephrectomized syngeneic recipients (n=6 per group), which received 0.5 mg/kg/d FTY720 or vehicle through oral gavage. Grafts were harvested 2 or 7 days posttransplantation. Renal tissue was examined histologically, stained for apoptosis, proliferation, inflammatory cell infiltrates, and studied for transforming growth factor-beta, and tumor necrosis factor-alpha expression. Rat proximal tubular cells were incubated with 0.1 to 30 mumol/L of phosphorylated FTY720 to test for in vitro cytopathic effects. RESULTS: FTY720 induced peripheral lymphopenia and significantly reduced intragraft CD3 and ED1 infiltrates. Acute tubular damage scores and graft function were not influenced by FTY720. Tubular apoptosis was significantly reduced, whereas the number of proliferating cell nuclear antigen-positive tubular cells were markedly increased. FTY720 attenuated renal tumor necrosis factor-alpha and transforming growth factor-beta expression. In vitro, pharmacologic concentrations up to 1 mumol/L of phosphorylated FTY720 did not affect tubular cell viability. CONCLUSION: FTY720 confers tubular epithelial protection in the presence of severe preservation reperfusion injury. Beneficial effects may in part be due to reduction in cell-mediated immune mechanisms. Furthermore, FTY720 could be helpful in patients with delayed graft function

Physiological variables in association with spreading depolarizations in the late phase of ischemic stroke

Physiological effects of spreading depolarizations (SD) are only well studied in the first hours after experimental stroke. In patients with malignant hemispheric stroke (MHS), monitoring of SDs is restricted to the postoperative ICU stay, typically day 2-7 post-ictus. Therefore, we investigated the role of physiological variables (temperature, intracranial pressure, mean arterial pressure and cerebral perfusion pressure) in relationship to SD during the late phase after MHS in humans. Additionally, an experimental stroke model was used to investigate hemodynamic consequences of SD during this time window. In 60 patients with MHS, the occurrence of 1692 SDs was preceded by a decrease in mean arterial pressure (-1.04 mmHg; p = .02) and cerebral perfusion pressure (-1.04 mmHg; p = .03). Twenty-four hours after middle cerebral artery occlusion in 50 C57Bl6/J mice, hypothermia led to prolonged SD-induced hyperperfusion (+2.8 min; p < .05) whereas hypertension mitigated initial hypoperfusion (-1.4 min and +18.5%Delta rCBF; p < .01). MRI revealed that SDs elicited 24 hours after experimental stroke were associated with lesion progression (15.9 vs. 14.8 mm(3); p < .01). These findings of small but significant effects of physiological variables on SDs in the late phase after ischemia support the hypothesis that the impact of SDs may be modified by adjusting physiological variables