De novo thrombotic microangiopathy following treatment with sirolimus: report of two cases

R. John Crew1, Jai Radhakrishnan1, David J. Cohen1, Leonard Stern1, Michael Goldstein2, Mark Hardy2, Vivette D. D'Agati3 and Glen S. Markowitz3

1 Division of Nephrology, Department of Medicine, 2 Department of Surgery and 3 Department of Pathology, Columbia University, College of Physicians and Surgeons, New York, NY, USA

Correspondence and offprint requests to: Glen S. Markowitz, MD, Department of Pathology, Columbia University Medical Center, 630 West 168th Street, Room 14-224, New York, NY 10032, USA. Email: gsm17{at}columbia.edu

Keywords: calcineurin inhibitor; de novo thrombotic microangiopathy; renal transplantation; sirolimus



   Introduction
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 Introduction
 Methods
 Case 1
 Case 2
 References
 
The term thrombotic microangiopathy (TMA) has been applied to a diverse group of conditions that share the common pathomechanism of endothelial damage. TMA is a recognized complication of solid organ transplantation. The majority of cases represent de novo TMA, which occurs in ~2.8–3.5% of renal transplant recipients and is associated with a 22% rate of graft loss [1]. The rate of graft loss is strongly influenced by whether the TMA is systemic or renal limited (38 vs 0% graft loss, respectively) [1].

Treatment with calcineurin inhibitors (CNIs) is a well-established risk factor for the development of de novo TMA. A recent, large analysis of the United States Renal Data System (USRDS) and Medicare claims identified multiple additional risk factors including younger recipient age, older donor age, female gender of the recipient, longer duration of dialysis before transplantation, previous renal transplant, delayed graft function (DGF), allograft rejection, increased peak panel-reactive antibody, and treatment with sirolimus (SRL) [2]. Anti-cardiolipin antibody seropositivity, often in the setting of chronic hepatitis C virus infection, is an additional risk factor for de novo TMA [3].

SRL (rapamycin) is an immunosuppressive agent commonly administered to renal transplant recipients. SRL may be used in combination with a CNI or as an alternative agent. A recent, large trial has shown that SRL facilitates early CNI withdrawal and that this regimen is associated with less long-term nephrotoxicity [4]. The most frequently reported side effects of SRL are thrombocytopenia, leukopenia, hypertriglyceridaemia and hypercholesterolaemia [5].

SRL has a similar mechanism of action to the CNIs. Cyclosporin (CSA), tacrolimus (TAC) and SRL all produce their immunosuppressive effect by binding to cytoplasmic proteins called immunophilins that modify immune function. CSA binds cyclophilin, and TAC binds FK-binding protein 12 (FKBP12); these complexes in turn inhibit calcineurin, a calcium-dependent phosphatase required for interleukin-2 production and progression of T cells from the G0 to G1 phase of the cell cycle. SRL also binds FKBP12, but the complex does not inhibit calcineurin activity. Instead, the SRL–FKBP12 complex inhibits a cell cycle regulatory protein referred to as mammalian target of rapamycin (mTOR) [6]. Inhibition of mTOR blocks the cellular response to cytokines and growth factors and progression of T cells from the G1 to S phase of the cell cycle. Inhibition of mTOR by the FKBP–SRL complex also has been shown to block proliferation in non-lymphoid tissues, including hepatocytes, vascular smooth muscle cells, endothelial cells and renal tubular epithelial cells [7].

Although initial studies suggested that SRL lacked significant nephrotoxicity, recent reports have challenged this concept. Early SRL use is associated with prolongation of DGF and a distinct histological pattern of renal injury described as a ‘cast nephropathy’ in which distal tubular casts are associated with sloughed tubular epithelia [8]. Based on an animal model of ischaemic acute tubular necrosis, the mechanism of SRL-induced DGF probably involves increased tubular apoptosis and reduced cellular proliferation, mediated in part by inhibition of protein kinase p70S6K [7].

Recent evidence suggests that treatment with SRL may be followed by the development of TMA. First, TMA has been noted in patients on protocols containing SRL in conjunction with CNIs [5,9–10]. Secondly, a recent analysis of the USRDS identified SRL use post-transplantation as a risk factor for TMA [2]. Thirdly, a recent report described a patient who developed TMA 16 days after renal transplantation on an immunosuppression regimen containing SRL, mycophenolate mofetil (MMF) and prednisone, after thymoglobulin induction [11]. This is the only report of SRL-associated TMA in the absence of a CNI. We present two additional cases of biopsy-proven TMA in patients treated with SRL who were not receiving CNIs at the time of renal biopsy.



   Methods
 Top
 Introduction
 Methods
 Case 1
 Case 2
 References
 
We identified two cases of TMA in renal transplant recipients who were being treated with SRL in the absence of a CNI. Patient records were reviewed for clinical histories, laboratory findings, medication use and outcomes. Neither patient had evidence of TMA as their primary disease in the native kidney and neither had evidence of acute antibody-mediated rejection, a potential mimic of TMA, at the time of diagnosis. Both biopsies were processed for light microscopy according to standard techniques and were stained for C4d. In case 1, C4d staining was performed on frozen tissue by indirect immunofluorescence at a concentration of 1:100 (Biogenesis, Kingston, NH). In case 2, frozen tissue was unavailable, and immunohistochemical staining for C4d was performed on formalin-fixed, paraffin-embedded tissue at a dilution of 1:25 (ALPCS Diagnostics, Windham, NH).

Of note, SRL was introduced at Columbia University Medical Center on a compassionate use basis in 1999 and for routine use in 2000. Both patients provided consent for the preparation of this manuscript.



   Case 1
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 Introduction
 Methods
 Case 1
 Case 2
 References
 
Clinical history
A 63-year-old female with a history of hypertension and hepatitis B virus (HBV) infection developed end-stage renal disease (ESRD) due to presumed chronic glomerulonephritis (the patient had had long-standing proteinuria and haematuria, without evidence of microangiopathic haemolytic anaemia). The patient was maintained on haemodialysis for 6 years prior to receiving a cadaveric renal transplant (CRT). Prior to transplantation, the patient had a panel of reactive antibodies (PRA) of <10% and there were no anti-donor antibodies against major histocompatibility complex (MHC) antigens. Induction therapy consisted of thymoglobulin for 5 days and intravenous methylprednisolone for 3 days, followed by tapering doses of prednisone. Chronic immunosuppression therapy consisted of MMF 1 g twice per day, SRL 5 mg per day (started on day 4 post-transplantation) and prednisone 20 mg per day. Following transplantation, the patient had DGF, requiring haemodialysis twice during the first week. She had an average trough level of SRL of 11.4 ng/ml (desired range 10–20 ng/ml). She also received valganciclovir 450 mg once per day for prophylaxis against cytomegalovirus (CMV), and lamivudine 150 mg once per day for HBV. At 35 days post-transplantation, her creatinine remained elevated at 2.8 mg/dl, without significant decline over the previous 10 days. Renal allograft biopsy was performed. At the time of biopsy, the patient had a platelet count of 71 000/mm3 (down from a pre-transplant level of 181 000/mm3; normal range 165 000–415 000 platelets/mm3), a haematocrit of 31.4% (unchanged during the post-transplant period; normal range for women 35–44%), a lactate dehydrogenase (LDH) of 291 IU/l (increased from a pre-transplant level of 134 IU/l; normal 115–221 IU/l), a haptoglobin of 148 mg/dl (normal 30–200 mg/dl) and a reticulocyte count of 0.7% (normal 0.8–2.3%).

Renal allograft biopsy findings
Sampling for light microscopy included two cores of renal cortex and 15 glomeruli, two of which were globally sclerotic. Two glomeruli contained intracapillary fibrin thrombi (Figure 1A) and a few additional glomeruli exhibited ischaemic-type wrinkling of the glomerular basement membrane and mesangiolysis. The remaining glomeruli were unremarkable. Proximal tubules displayed mild degenerative changes characterized by luminal ectasia and cytoplasmic simplification. There was mild tubular atrophy and interstitial fibrosis involving 15% of the cortex sampled. Mild interstitial inflammation composed mainly of lymphocytes occupied <20% of the cortex sampled. There was no significant tubulitis or neutrophil margination in peritubular capillaries. A single artery contained a fibrin thrombus that was associated with endothelial necrosis (Figure 1B). Multiple vessels displayed mucointimal oedema (Figure 1C).



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Fig. 1. (A) A glomerulus from patient 1 contains a large intracapillary fibrin thrombus. Focal marginating neutrophils are also seen [haematoxylin and eosin (H&E), x600]. (B) A small artery is occluded by an intraluminal fibrin thrombus, associated with endothelial necrosis (H&E, x630). (C) A small artery displays mucointimal oedema and mild swelling of endothelial cells (trichrome, x630). (D) A glomerulus from patient 2 contains two intracapillary fibrin thrombi. Glomerular capillary lumina are narrowed by swelling of endothelial cells (H&E, x400). (E) Another glomerulus exhibits marked erythrocyte congestion, fibrin thrombi, endothelial necrosis and focal entrapment of schistocytes (H&E, x250).

 
Immunofluorescence staining of glomeruli was negative for IgG, IgM, IgA, C3, C1q, fibrinogen, albumin, and {kappa} and {lambda} light chains. There was low intensity positivity in vessel walls for C3 and C1q. Staining for C4d showed low intensity positivity in glomeruli and vessels, as is typically seen in normal controls. Importantly, staining for C4d was negative in peritubular capillaries, the site at which positivity has specificity for antibody-mediated rejection. Electron microscopy was not performed.

Based on these findings, the patient was diagnosed with acute TMA.

Clinical follow-up
Following renal biopsy, treatment with SRL was discontinued and replaced with TAC 4 mg twice per day. Prednisone and MMF were continued. CMV polymerase chain reaction (PCR) viral loads and anti-cardiolipin antibody were negative. Within 2 weeks, the patient's creatinine declined to 2.1 mg/dl and her platelet count increased to 191 000/mm3. Six months later, the patient has a serum creatinine of 1.8 mg/dl, a haematocrit of 36% and a platelet count of 237 000/mm3.



   Case 2
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 Introduction
 Methods
 Case 1
 Case 2
 References
 
Clinical history
Renal allograft biopsy findings
Sampling for light microscopy included two cores of renal cortex containing 16 glomeruli, none of which were globally sclerotic. Glomeruli appeared normal in size and exhibited a mild increase in mesangial cells. There were segmental to global narrow double contours of the glomerular basement membrane with focal mesangial interposition and focal marginating mononuclear leukocytes. Three glomeruli contained intracapillary fibrin thrombi associated with endothelial swelling and necrosis, and erythrocyte congestion and fragmentation (Figure 1D and E). Four glomeruli displayed prominent ischaemic changes, with global wrinkling and retraction of the glomerular basement membrane. There was moderate tubular atrophy and interstitial fibrosis involving ~40% of the cortex sampled. There was moderate interstitial inflammation composed mainly of lymphocytes and associated with multifocal tubulitis with a maximum of six mononuclear leukocytes per tubular cross-section. Proximal tubules exhibited mild simplification and coarse vacuolization. There was no evidence of neutrophil margination in peritubular capillaries. A single arteriole contained a fibrin thrombus associated with fibrinoid necrosis of the vessel wall. A small artery exhibited early endovasculitis with a few lymphocytes undermining the endothelium. There was also evidence of moderate arteriosclerosis and arteriolosclerosis with hyalinosis.

Immunohistochemical staining for C4d was negative in peritubular capillaries, providing evidence against antibody-mediated rejection. Immunofluorescence and electron microscopy were not performed.

Based on these findings, three diagnoses were rendered: (i) acute TMA; (ii) moderate acute rejection (Banff grade 2A); and (iii) moderate chronic allograft nephropathy.

Clinical follow-up
The patient was treated for acute rejection with intravenous methylprednisolone 500 mg daily for 3 days followed by prednisone taper. The SRL dose was decreased to 5 mg once per day, and his SRL trough level decreased to 11.1 ng/ml. His creatinine decreased to 2.2 mg/dl, but 1 month later climbed again to 3.6 mg/dl. His platelet count remained depressed at 88 000/mm3. Examination of a peripheral blood smear revealed rare schistocytes. A fourth renal biopsy showed (i) findings of TMA, which now appeared more subacute to chronic; (ii) moderate to severe chronic allograft nephropathy; (iii) no evidence of acute rejection; and (iv) negative staining of peritubular capillaries for C4d.

SRL was discontinued and the patient was started on prednisone 80 mg per day and plasma exchange. Despite 18 plasma exchange treatments, his platelet count remained low at 70 000/mm3. Six weeks later, the patient had a creatinine of 2.7 mg/dl. The patient was started on vincristine and, following a single dose, was admitted with catheter-related infection. Plasma exchange was discontinued and the patient was treated with intravenous immunoglobulin (IVIG). Three days later, he developed acute renal failure with a creatinine of 8.3 mg/dl. A fifth renal allograft biopsy revealed chronic sequelae of the previous findings and superimposed acute tubular injury due to osmotic nephrosis, consistent with IVIG nephrotoxicity.

Treatment with IVIG was discontinued. The patient's creatinine declined over 1 month to a new baseline level of 3.0 mg/dl. The patient subsequently progressed to ESRD over the following 8 months. During this time, his immunosuppressive regimen consisted of azathioprine 25 mg once per day and prednisone 10 mg once per day. Two years later, at the time of his fourth renal transplant, allograft nephrectomy revealed prominent changes of chronic thrombotic microangiopathy. At that time, enzyme-linked immunosorbent assay (ELISA) testing did not reveal antibodies against the spouse's (donor's) HLA antigens.

Results
The clinical parameters of the two patients with de novo TMA following treatment with SRL are summarized in Table 1. Both patients had risk factors for development of de novo TMA including, in case 1, female gender, prolonged duration of dialysis pre-transplantation, and DGF, and, in case 2, previous transplantation and previous allograft rejection. Both patients underwent renal allograft biopsy due to an elevation of serum creatinine (rather than systemic symptoms of TMA) and both were diagnosed with TMA based on biopsy findings. The duration of SRL therapy prior to biopsy was relatively short at 4.5 and 2 weeks, respectively. The histological findings of TMA were accompanied by thrombocytopenia, a known side effect of treatment with SRL. Although neither patient had an acute decline in haematocrit, patient 1 had a mild increase in LDH and patient 2 had rare schistocytes on peripheral smear.


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Table 1. Clinical features

 
Following renal allograft biopsy in patient 1, SRL was discontinued and replaced with TAC. This led to a rapid and sustained improvement in serum creatinine. In patient 2, renal allograft biopsy revealed de novo TMA as well as acute rejection and chronic allograft nephropathy. The patient's serum creatinine initially improved following treatment of the acute rejection and a marked reduction in SRL dose. However, the patient's course subsequently was complicated by more chronic findings of TMA, worsening chronic allograft nephropathy, IVIG toxicity and, eventually, irreversible renal failure.

Discussion
We report the clinical and pathological findings in two patients who developed de novo TMA following treatment with SRL. In case 1, the rapid and sustained improvement in renal function following discontinuation of SRL strongly implicates this agent in the development of acute TMA. Case 2 is more complex in that the patient had previous and concurrent episodes of acute cellular rejection and a history of CNI toxicity. For this same reason, this case is also of particular interest. The patient had two recent previous renal allogaft biopsies documenting CNI toxicity (following treatment with TAC and CSA), but neither biopsy exhibited findings of TMA. The close temporal relationship between the onset of treatment with SRL and the development of TMA strongly implicates SRL as the aetiological agent. Unfortunately, due to multiple co-existent processes, SRL withdrawal did not lead to improvement of renal function.

Multiple previous reports have suggested a relationship between SRL and the development of TMA. Barone et al. described a patient who developed TMA 16 days after renal transplantation on an immunosuppression regimen containing SRL, MMF and prednisone, after thymoglobulin induction [11]. Robson et al. reported a patient with stable renal function on CSA for 8 years. Due to mild chronic allograft nephropathy and leukopenia, azathioprine was discontinued and replaced with SRL. Shortly thereafter, the patient developed acute TMA [9]. Similarly, Saikali et al. described a patient with CNI toxicity related to treatment with TAC [10]. Following introduction of SRL and a marked reduction in TAC dose, the patient developed TMA. Treatment with TAC was discontinued but the patient continued to have worsening TMA. Only following discontinuation of SRL and plasma exchange did the patient's TMA resolve and renal function improve [10]. In a large, phase III study comparing SRL with placebo in renal transplant recipients simultaneously treated with CSA and steroids, TMA was among the most frequent diagnoses leading to SRL discontinuation [5]. SRL has also been linked to acute TMA in patients with steroid-refractory graft-versus-host disease following allogeneic haematopoietic stem cell transplantation [12].

A recent study looked at risk factors for TMA in 15 870 renal transplants recipients in the USRDS [2]. The diagnosis of TMA was based on diagnostic coding at the time of hospital discharge. Based on the design of the study, the data must be interpreted with caution. For instance, a patient might have received SRL because of the fear of using a CNI in a patient with a previous history of TMA. Furthermore, a patient may have been discharged while receiving SRL due to previous CSA-induced TMA, followed by SRL rescue. Despite these potential issues, the results in this large cohort were quite striking. The incidence of TMA among patients on initial SRL maintenance therapy was 18.1 in 1000 patient-years (PY), compared with only 5.0 in 1000 PY among patients on initial maintenance CNI therapy. On univariate analysis, SRL therapy, with or without concurrent CNI, was associated with de novo TMA.

Calcineurin inhibitors have multiple effects on the renal vasculature and are established precipitants of TMA in the renal transplant recipient. CNIs are associated with direct endothelial toxicity, as well as vasoconstriction and hyaline arteriolopathy [13]. In cell culture, CSA has an anti-proliferative and pro-apoptotic effect on endothelial cells [14] and blocks angiogenesis induced by vascular endothelial growth factor [15]. In vivo, higher CSA trough levels correlate with elevated levels of circulating endothelial cells, a marker of endothelial damage [16]. CSA also blocks repopulation of the allograft vasculature by recipient-derived endothelial cells [17].

SRL may promote thrombosis and TMA by a mechanism similar to that of the CNIs. The SRL–FKBP12 complex inhibits mTOR, which in turn prevents p70S6K activation. Endothelial cell proliferation in response to growth factors and oscillatory blood flow is highly dependent on p70S6K signalling [18,19]. Thus, similar to the CNIs, SRL acts to inhibit endothelial cell proliferation [7], a property that is the basis for the use of SRL-eluting coronary artery stents to prevent restenosis [20]. In the transplant setting, endothelial cells may be injured by a variety of mechanisms including direct drug toxicity, infection, immune processes and OKT3 use. In response to injury, blood vessels in the allograft may be repopulated by recipient-derived endothelial cells [21]. The anti-proliferative effect of SRL and the CNIs may prevent repopulation of the allograft vasculature by reparative endothelial proliferation, thereby promoting local activation of the clotting cascade, consumption of platelets and red blood cell destruction. This hypothesis is supported by the fact that the majority of de novo TMA represents ‘renal-limited’ rather than systemic disease. SRL has also been shown in cell culture to promote platelet aggregation [22], thus providing another potential mechanism by which SRL could promote TMA.

How can SRL produce TMA when it has been used with success in patients with de novo TMA associated with CNI use [23,24]? This apparent paradox may be explained by SRL's different mechanism of action and the need for specific CNI withdrawal to control TMA, just as TMA associated with CSA may respond solely to switching to TAC [25]. In a similar manner, it is worth noting that in the case reported by Barone et al. [11] and the first case reported herein, de novo TMA improved following discontinuation of SRL and institution of a CNI.

Thrombocytopenia is a well-recognized side effect of SRL, which is dose dependent and rapidly reversible following cessation of SRL therapy [26]. Because of this side effect, one of the main clinical parameters that may suggest the presence of TMA is of limited utility in patients treated with SRL. As a result, TMA in patients treated with this agent may be under-recognized and requires extra diligence on the part of the clinician. Renal biopsy is likely to remain the most useful modality to identify TMA in patients treated with SRL.

SRL has been a useful advance in the field of transplantation. Initial reports that this agent was not associated with significant nephrotoxicity have proven false, although the incidence of renal side effects is far lower than that of the CNIs. We report two patients with de novo TMA following treatment with SRL. These cases add to a growing body of literature that points to an aetiological relationship between SRL and the development of TMA, with or without co-administration of a CNI. Additional studies are needed to compare the ‘thrombogenic potential’ of SRL and the CNIs and to determine whether these effects are additive when the agents are administered together.

Conflict of interest statement. R. John Crew is supported by a grant from the Kidney and Urology Foundation of America.

[See related article by Marti and Frey (this issue pp. 13–15)]



   References
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 Introduction
 Methods
 Case 1
 Case 2
 References
 

  1. Schwimmer J, Nadasdy TA, Spitalnik PF, Kaplan KL, Zand MS. De novo thrombotic microangiopathy in renal transplant recipients: a comparison of hemolytic uremic syndrome with localized renal thrombotic microangiopathy. Am J Kidney Dis 2003; 41: 471–479[CrossRef][ISI][Medline]
  2. Reynolds JC, Agodoa LY, Yuan CM, Abbott KC. Thrombotic microangiopathy after renal transplantation in the United States. Am J Kidney Dis 2003; 42: 1058–1068[ISI][Medline]
  3. Baid S, Pascual M, Williams WW Jr et al. Renal thrombotic microangiopathy associated with anticardiolipin antibodies in hepatitis C-positive renal allograft recipients. J Am Soc Nephrol 1999; 10: 146–153[Abstract/Free Full Text]
  4. Oberbauer R, Kreis H, Johnson RW et al. Long-term improvement in renal function with sirolimus after early cyclosporine withdrawal in renal transplant recipients: 2-year results of the Rapamune Maintenance Regimen Study. Transplantation 2003; 76: 364–370[CrossRef][ISI][Medline]
  5. MacDonald AS. A worldwide, phase III, randomized, controlled, safety and efficacy study of a sirolimus/cyclosporine regimen for prevention of acute rejection in recipients of primary mismatched renal allografts. Transplantation 2001; 71: 271–280[ISI][Medline]
  6. Schmelzle T, Hall MN. TOR, a central controller of cell growth. Cell 2000; 103: 253–262[ISI][Medline]
  7. Lieberthal W, Fuhro R, Andry CC et al. Rapamycin impairs recovery from acute renal failure: role of cell-cycle arrest and apoptosis of tubular cells. Am J Physiol 2001; 281: F693–F706[ISI]
  8. Smith KD, Wrenshall LE, Nicosia RF et al. Delayed graft function and cast nephropathy associated with tacrolimus plus rapamycin use. J Am Soc Nephrol 2003; 14: 1037–1045[Abstract/Free Full Text]
  9. Robson M, Cote I, Abbs I, Koffman G, Goldsmith D. Thrombotic micro-angiopathy with sirolimus-based immunosuppression: potentiation of calcineurin-inhibitor-induced endothelial damage? Am J Transplant 2003; 3: 324–327[CrossRef][ISI][Medline]
  10. Saikali JA, Truong LD, Suki WN. Sirolimus may promote thrombotic microangiopathy. Am J Transplant 2003; 3: 229–230[CrossRef][ISI][Medline]
  11. Barone GW, Gurley BJ, Abul-Ezz SR, Gokden N. Sirolimus-induced thrombotic microangiopathy in a renal transplant recipient. Am J Kidney Dis 2003; 42: 202–206[CrossRef][ISI][Medline]
  12. Benito AI, Furlong T, Martin PJ et al. Sirolimus for the treatment of steroid-refractory acute graft-versus-host disease. Transplantation 2001; 72: 1924–9[ISI][Medline]
  13. Remuzzi G, Bertani T. Renal vascular and thrombotic effects of cyclosporine. Am J Kidney Dis 1989; 13: 261–272[ISI][Medline]
  14. Esposito C, Fornoni A, Cornacchia F et al. Cyclosporine induces different responses in human epithelial, endothelial and fibroblast cell cultures. Kidney Int 2000; 58: 123–130[CrossRef][ISI][Medline]
  15. Hernandez GL, Volpert OV, Iniguez MA et al. Selective inhibition of vascular endothelial growth factor-mediated angiogenesis by cyclosporin A: roles of the nuclear factor of activated T cells and cyclooxygenase 2. J Exp Med 2001; 193: 607–620[Abstract/Free Full Text]
  16. Woywodt A, Schroeder M, Mengel M et al. Circulating endothelial cells are a novel marker of cyclosporine-induced endothelial damage. Hypertension 2003; 41: 720–723[Abstract/Free Full Text]
  17. Hillebrands JL, Klatter FA, van den Hurk BMH, Popa ER, Nieuwenhuis P, Rozing J. Origin of neointimal endothelium and {alpha}-actin-positive smooth muscle cells in transplant arteriosclerosis. J Clin Invest 2001; 107: 1411–1422[Abstract/Free Full Text]
  18. Kraiss LW, Ennis TM, Alto NM. Flow-induced DNA synthesis requires signaling to a translational control pathway. J Surg Res 2001; 97: 20–26[CrossRef][ISI][Medline]
  19. Vinals F, Chambard JC, Pouyssegur J. p70 S6 kinase-mediated protein synthesis is a critical step for vascular endothelial cell proliferation. J Biol Chem 1999; 274: 26776–26782[Abstract/Free Full Text]
  20. Morice MC, Serruys PW, Sousa JE et al. A randomized comparison of a sirolimus-eluting stent with a standard stent for coronary revascularization. N Engl J Med 2002; 346: 1773–1780[Abstract/Free Full Text]
  21. Lagaaij EL, Cramer-Knijnenburg GF, van Kemenade FJ, van Es LA, Bruijn JA, van Krieken JHJM. Endothelial cell chimerism after renal transplantation and vascular rejection. Lancet 2001; 357: 33–37[CrossRef][ISI][Medline]
  22. Babinska A, Markell MS, Salifu MO, Akoad M, Ehrlich YH, Kornecki E. Enhancement of human platelet aggregation and secretion induced by rapamycin. Nephrol Dial Transplant 1998; 13: 3153–9[Abstract]
  23. Edwards C, House A, Shahinian V, Knoll G. Sirolimus-based immunosuppression for transplant-associated thrombotic microangiopathy. Nephrol Dial Transplant 2002; 17: 1524[Free Full Text]
  24. Franco A, Hernandez D, Capdevilla L et al. De novo hemolytic–uremic syndrome/thrombotic microangiopathy in renal transplant patients receiving calcineurin inhibitors: role of sirolimus. Transplant Proc 2003; 35: 1764–1766[CrossRef][ISI][Medline]
  25. Zarifian A, Meleg-Smith S, O’Donovan R, Tesi RJ, Batuman V. Cyclosporine-associated thrombotic microangiopathy in renal allografts. Kidney Int 1999; 55: 2457–2466[CrossRef][ISI][Medline]
  26. Murgia MG, Jordan S, Kahan BD. The side effect profile of sirolimus: a phase I study in quiescent cyclosporine–prednisone-treated renal transplant patients. Kidney Int 1996; 49: 209–216[ISI][Medline]
Received for publication: 4. 2.04
Accepted in revised form: 3. 5.04