Sandrine Florquin

Manifestations of Fabry disease in placental tissue

SummaryFabry disease is an X-linked lysosomal storage disorder caused by deficiency of the lysosomal enzyme α-galactosidase A. Manifestations of the disease in placental tissue have been reported only twice. We report for the first time on the biochemical, histological and genetic features of two cases: placenta A derived from a mother heterozygous for Fabry disease who gave birth to a hemizygous son, and placenta B obtained from a healthy mother who carried a heterozygous daughter. Biopsies of placentae A, B and of four healthy controls were taken directly after birth. Assessment of α-galactosidase A (α-Gal) activity was performed both in fetal leukocytes (derived from umbilical cord blood) and in the biopsy specimens. The tissue was further examined by electron microscopy, immunohistochemistry and biochemical analysis for the presence of storage material (ceramide trihexoside (CTH)). In placenta A, characteristic zebra bodies reflecting accumulated storage material were seen in all biopsies evaluated. CTH values were markedly elevated as compared to the controls and α-Gal activity in both fetal leukocytes and placental tissue was very low. Placenta B showed no storage material at all. CTH values were within the control range. α-Gal activity ranged from intermediate to near normal; enzyme activity in fetal leukocytes was significantly decreased. As placental tissue is mainly derived from fetal cells, we may conclude that, in a boy suffering from Fabry disease, extensive storage of CTH is already present at birth. As complications develop only around the age of 10 years, it may be not the CTH itself but secondary processes that cause cellular and organ damage.

The role of plasminogen activator inhibitor type 1 in the inflammatory response to local tissue injury.

Summary.u2002 Background: The plasma levels of the plasminogen activator‐inhibitor type 1 (PAI‐1) are consistently elevated in patients with sterile tissue injury, often accompanied by a systemic acute phase protein response. It remains unknown, however, whether and to what extent PAI‐1 affects the host response to trauma. Methods and results: By using the well‐established murine model of turpentine‐induced tissue injury we compared local and systemic inflammatory responses in PAI‐1 gene‐deficient (PAI‐1–/–) and normal wild‐type (Wt) mice. Subcutaneous turpentine injection elicited strong increases in PAI‐1 protein concentration in plasma and at the site of injury, but not in liver. PAI‐1 mRNA was locally increased and expressed mainly by macrophages and endothelial cells. PAI‐1 deficiency greatly enhanced the early influx of neutrophils to the site of inflammation, which was associated with increased edema and necrosis at 8 h after injection. Furthermore, PAI‐1–/– mice showed a reduced early interleukin (IL)‐6 induction with subsequently lower acute phase protein levels and a much slower recovery of body weight loss. Conclusion: These findings suggest that PAI‐1 is not merely a marker of tissue injury but plays a functional role in the local and systemic host response to trauma.

Atypical relapse of hemolytic uremic syndrome after transplantation

Atypical hemolytic uremic syndrome (HUS) frequently leads to end-stage renal failure and can relapse after transplantation. A 12-year-old girl presenting with familial atypical HUS with a factor H mutation was successfully transplanted 6xa0years after a first transplant that had failed because of immediate recurrent HUS. Prophylactic plasma exchange before and after transplantation was used. Two months after transplantation, concomitant with a reduction in plasma exchange frequency, the plasma creatinine increased from 70xa0µmol/l to 194xa0µmol/l in 2xa0weeks without thrombocytopenia or signs of hemolytic anemia. The patient had minimal clinical symptoms and a presumptive diagnosis of graft rejection was made. Despite treatment with six daily pulses of methylprednisolone, plasma creatinine continued to increase and a graft biopsy was therefore undertaken. This showed the typical appearance of a thrombotic microangiopathy without any evidence of rejection. Despite daily plasmapheresis and replacement of cyclosporine with tacrolimus, there was no improvement and transplant nephrectomy was undertaken. This patient demonstrates that HUS can recur in a kidney transplant without the diagnostic hematological features and emphasizes the need for early transplant biopsy in such patients showing a decline in transplant function.

Stimulation of acetylcholine receptors impairs host defence during pneumococcal pneumonia

The cholinergic nervous system can inhibit the systemic inflammation accompanying sepsis by virtue of a specific action of acetylcholine on α7 cholinergic receptors. The current authors sought to determine the effect of nicotine, an α7 cholinergic receptor agonist, on the host response to pneumonia caused by Streptococcus pneumoniae. Mice were intranasally infected with S. pneumoniae and treated with nicotine or saline intraperitoneally using a treatment schedule shown to improve host defence against abdominal sepsis. Nicotine treatment was associated with a transiently enhanced growth of S. pneumoniae, as indicated by higher bacterial loads in both lungs and blood at 24u2005h after infection. At 48u2005h after infection, bacterial burdens had increased in both treatment groups and differences were no longer present. Remarkably, mice treated with nicotine showed enhanced lung inflammation at 24u2005h after infection. Moreover, both lung and plasma concentrations of the pro-inflammatory cytokines tumour necrosis factor-α and interferon-γ were higher in nicotine-treated animals at this time-point. Additional studies examining the effect of nicotine on the immediate (4-h) inflammatory response to S. pneumoniae did not reveal an anti-inflammatory effect of nicotine either. The present data suggest that nicotine transiently impairs host defence in pneumococcal pneumonia.

Diagnostic dilemmas in Fabry disease: a case series study on GLA mutations of unknown clinical significance

Fabry disease (FD) phenotype is heterogeneous: alpha‐galactosidase A gene mutations (GLA) can lead to classical or non‐classical FD, or no FD. The aim of this study is to describe pitfalls in diagnosing non‐classical FD and assess the diagnostic value of plasma globotriaosylsphingosine. This is a case series study. Family 1 (p.A143T) presented with hypertrophic cardiomyopathy (HCM), absent classical FD signs, high residual alpha‐galactosidase A activity (AGAL‐A) and normal plasma globotriaosylsphingosine. Co‐segregating sarcomeric mutations were found. Cardiac biopsy excluded FD. In family 2 (p.P60L), FD was suspected after kidney biopsy in a female with chloroquine use. Males had residual AGAL‐A, no classical FD signs and minimally increased plasma globotriaosylsphingosine, indicating that p.P60L is most likely non‐pathogenic. Non‐specific complications and histology can be explained by chloroquine and alternative causes. Males of two unrelated families (p.R112H) show AGAL‐A <5%, but slightly elevated plasma globotriaosylsphingosine (1.2–2.0 classical males >50 nmol/l). Histological evidence suggests a variable penetrance of this mutation. Patients with GLA mutations and non‐specific findings such as HCM may have non‐classical FD or no FD. Other (genetic) causes of FD‐like findings should be excluded, including medication inducing FD‐like storage. Plasma globotriaosylsphingosine may serve as a diagnostic tool, but histology of an affected organ is often mandatory.

Treatment with everolimus is associated with a procoagulant state.

INTRODUCTIONnRenal transplant recipients are at increased risk of venous thromboembolic events, which is in part caused by their treatment with maintenance immunosuppressive drugs. Because we observed an increased incidence of venous thromboembolic events in renal transplant recipients treated with the mTOR inhibitor (mTORi) everolimus, we aimed to identify prothrombotic mechanisms of this immunosuppressive drug.nnnMATERIALS AND METHODSnIn a single center study, nested in a multi-center randomized controlled trial, we measured parameters of coagulation, anti-coagulation and fibrinolysis in renal transplant recipients, receiving the mTORi everolimus (n=16, mTOR group) and compared them to a similar patient group, receiving a calcineurin inhibitor and/or mycophenolate sodium (n=20, non-mTOR group). All patients were at least 6 months following transplantation with a stable transplant function.nnnRESULTSnThe use of an mTORi was associated with significantly higher levels of von Willebrand factor, prothrombin fragment 1+2, thrombin-activatable fibrinolysis inhibitor and plasminogen activator inhibitor-1 as compared to a non-mTORi based immunosuppressive regimen.nnnCONCLUSIONSnTreatment with an mTORi leads to increased endothelial activation, thrombin formation and impaired fibrinolysis in renal transplant recipients. This suggests an increased risk of thrombotic events in renal transplant recipients treated with mTOR inhibitors. A prospective study to establish the precise risk of thrombotic events in these patients is urgently needed.

Functional thrombomodulin deficiency causes enhanced thrombus growth in a murine model of carotid artery thrombosis

Abstract.Thrombomodulin (TM) bound thrombin initiates the protein Cnanticoagulant pathway and defects in TM result in enhancedncoagulation. Recent studies suggest a role for TM in arterialnvascular disease. In order to corroborate this association wenstudied arterial thrombus formation in mice with a functional TMndefect. We used mice homozygous for a n404Glu-to-Pro mutation in the TM genen(TMpro/pro) and compared these withnwildtype littermates in a model of FeCl3ninduced carotid artery thrombosis. Time-to-occlusion (TTO) wasnassessed by arterial blood flow measurement, using a Doppler flownprobe. Complete occlusion occurred in 8/10 (80%)nTMpro/pro mice and in 3/11 (27%)nlittermate controls. Mean time to occlusion (TTO) [± SE] was 767n± 196 s in the F2-TMpro/pro mice,nversus 1507 ± 159 s in controls (p = 0.007, Mann Whitney Untest). Histology and immunostaining for tissue factor did notnreveal any differences in thrombus morphology or thrombogenicitynbetween the two groups.These data confirm and extend the fiding that a functionalndeficiency in TM results in enhanced thrombus formation in anmurine model of carotid artery thrombosis and support a role fornTM defects in arterial thrombotic disease.

RAGE does not contribute to renal injury and damage upon ischemia/reperfusion-induced injury.

The receptor for advanced glycation end products (RAGE) mediates a variety of inflammatory responses in renal diseases, but its role in renal ischemia/reperfusion (I/R) injury is unknown. We showed that during renal I/R, RAGE ligands HMGB1 and S100B are expressed. However, RAGE deficiency does not affect renal injury and function upon I/R-induced injury.

Cecal ligation and puncture induced sepsis impairs host defense against Enterococcus faecium peritonitis.

PurposeMultiresistant and vancomycin resistant Enterococcus faecium (VRE) can cause serious infections in hospitalized patients with various co-morbid diseases. We investigated the course of VRE peritonitis after cecal ligation and puncture (CLP)-induced sepsis and compared this to sham operated mice.MethodsMice were subjected to CLP or sham surgery. Forty-eight hours thereafter four groups were created by subjecting mice to peritoneal injection of either VRE or saline.ResultsMice infected with VRE after CLP were severely impaired in eliminating VRE from the peritoneal cavity and distant body sites. These mice failed to mount an early inflammatory response at the primary site of VRE infection. VRE superinfection did not influence CLP-induced organ damage or polymicrobial bacterial loads.ConclusionsSublethal polymicrobial sepsis greatly facilitates infection and dissemination of VRE. VRE does not influence the course of CLP-induced sepsis.

Early Conversion to Prednisolone/Everolimus as an Alternative Weaning Regimen Associates With Beneficial Renal Transplant Histology and Function: The Randomized-Controlled MECANO Trial

In renal transplantation, use of calcineurin inhibitors (CNIs) is associated with nephrotoxicity and immunosuppression with malignancies and infections. This trial aimed to minimize CNI exposure and total immunosuppression while maintaining efficacy. We performed a randomized controlled, open‐label multicenter trial with early cyclosporine A (CsA) elimination. Patients started with basiliximab, prednisolone (P), mycophenolate sodium (MPS), and CsA. At 6 months, immunosuppression was tapered to P/CsA, P/MPS, or P/everolimus (EVL). Primary outcomes were renal fibrosis and inflammation. Secondary outcomes were estimated glomerular filtration rate (eGFR) and incidence of rejection at 24 months. The P/MPS arm was prematurely halted. The trial continued with P/CsA (N = 89) and P/EVL (N = 96). Interstitial fibrosis and inflammation were significantly decreased and the eGFR was significantly higher in the P/EVL arm. Cumulative rejection rates were 13% (P/EVL) and 19% (P/CsA), (p = 0.08). A post hoc analysis of HLA and donor‐specific antibodies at 1 year after transplantation revealed no differences. An individualized immunosuppressive strategy of early CNI elimination to dual therapy with everolimus was associated with decreased allograft fibrosis, preserved allograft function, and good efficacy, but also with more serious adverse events and discontinuation. This can be a valuable alternative regimen in patients suffering from CNI toxicity.