Tissue factor related thrombogenesis induced by heterologous human adult liver-derived progenitor cell infusion : modulation by cell dose escalation and anticoagulant drugs : clinical application and relevance

Mesenchymal stem cells (MSCs) are used in numerous clinical trials nowadays, but their blood compatibility is still an important concern. MSCs express a procoagulant activity, linked to the expression of tissue factor (TF), leading to thrombi formation after blood exposure. However, different approaches could be useful to prevent these side effects. In this thesis, we demonstrated that TF, known to be expressed by liver-derived MSCs, induces in a cell dose dependent manner, the activation of coagulation, resulting in fibrin generation and platelet activation. Thrombogenic adverse events induced by MSCs infusions could either be prevented by using low cell doses (5 × 106 cells/kg) or high cell doses (50 × 106 cells/kg) combined with anticoagulant drugs. Next, we also observed that activation of coagulation by liver-derived MSCs is less strong in blood from acute decompensated patients compared to healthy control subjects. Finally, during a clinical trial on eleven patients presenting with metabolic liver disorders, we showed that adding anticoagulant drugs during cell infusions is safe and limits infusion related thrombogenesis to subclinical signs in most of the patients.(MED - Sciences médicales) -- UCL, 202

Hepatocyte Transplantation in Children

The liver has an important function in the human body and plays a crucial role in its metabolism. Orthotopic liver transplantation (OLT) is the gold standard treatment for patients presenting liver failure or end stage liver diseases, and is also applied for liver based intractable metabolic disorders. Due to organ shortage, invasive surgery and persistent mortality/morbidity, other treatments have to be explored. Amongst these, hepatocyte transplantation is an attractive alternative and has shown promising results in the treatment of miscellaneous metabolic disorders

Thrombogenic Risk Induced by Intravascular Mesenchymal Stem Cell Therapy: Current Status and Future Perspectives

Mesenchymal stem cells (MSCs) are currently studied and used in numerous clinical trials. Nevertheless, some concerns have been raised regarding the safety of these infusions and the thrombogenic risk they induce. MSCs express procoagulant activity (PCA) linked to the expression of tissue factor (TF) that, when in contact with blood, initiates coagulation. Some even describe a dual activation of both the coagulation and the complement pathway, called Instant Blood-Mediated Inflammatory Reaction (IBMIR), explaining the disappointing results and low engraftment rates in clinical trials. However, nowadays, different approaches to modulate the PCA of MSCs and thus control the thrombogenic risk after cell infusion are being studied. This review summarizes both in vitro and in vivo studies on the PCA of MSC of various origins. It further emphasizes the crucial role of TF linked to the PCA of MSCs. Furthermore, optimization of MSC therapy protocols using different methods to control the PCA of MSCs are described

A Mixed Quantitative-Qualitative Approach to Disagreement in Online News Comments on Social Networking Sites

This paper investigates disagreement constructions on online social networking sites (SNS). It forms part of a wider project on hate and conflict speech modelling. Combining different research theories from conversational analysis and corpus linguistics, we have devised a six-class disagreement typology that we have manually tested on a 20 000-word corpus of Reddit comments on media posts. We then completed this analysis with a description of linguistic markers that pave the way towards future automated research. Finally, we present politeness strategies and repairs that maintain mutual understanding in media posts’ comments. Our analysis proposes new classifications adapted to SNS. Moreover, it highlights regular forum trends, face-to-face and group threatening acts and Reddit-specific strategies to maintain or repair disagreement

Infusion-related thrombogenesis by liver-derived mesenchymal stem cells controlled by anticoagulant drugs in 11 patients with liver-based metabolic disorders

BACKGROUND: Mesenchymal stem cell (MSC) transplantation is a fast-developing therapy in regenerative medicine. However, some concerns have been raised regarding their safety and the infusion-related pro-coagulant activity. The aim of this study is to analyze the induced thrombogenic risk and the safety of adding anticoagulants during intraportal infusions of liver-derived MSCs (HepaStem), in patients with Crigler-Najjar (CN) and urea cycle disorders (UCD). METHODS: Eleven patients (6 CN and 5 UCD patients) were included in this partially randomized phase 1/2 study. Three cell doses of HepaStem were investigated: low (12.5 × 106 cells/kg), intermediate (50 × 106 cells/kg), and high doses (200 × 106 cells/kg). A combination of anticoagulants, heparin (10 I.U./5 × 106cells), and bivalirudin (1.75 mg/kg/h) were added during cell infusions. The infusion-related thrombogenic risk and anticoagulation were evaluated by clinical monitoring, blood sampling (platelet and D-dimer levels, activated clotting time, etc.) and liver Doppler ultrasound. Mixed effects linear regression models were used to assess statistically significant differences. RESULTS: One patient presented a thrombogenic event such as a partial portal vein thrombus after 6 infusions. Minor adverse effects such as petechiae, epistaxis, and cutaneous hemorrhage at the site of catheter placement were observed in four patients. A significant decrease in platelet and increase in D-dimer levels were observed at the end of the infusion cycle, normalizing spontaneously after 7 days. No significant and clinically relevant increase in portal vein pressure could be observed once the infusion cycle was completed. CONCLUSIONS: The safety- and the infusion-related pro-coagulant activity remains a concern in MSC transplantation. In our study, a combination of heparin and bivalirudin was added to prevent the thrombogenic risk induced by HepaStem infusions in 11 patients. A significant decrease in platelet and increase in D-dimer levels were observed, suggesting the activation of coagulation in these patients; however, this was spontaneously reversible in time. We can conclude that adding this combination of anticoagulants is safe and limits infusion-related thrombogenesis to subclinical signs in most of the patients

Vitamin K May Be A-OK, in Kids and Cholestatic Patients.

Comment on Routine Use of Vitamin K in the Treatment of Cirrhosis-Related Coagulopathy: Is it A-O-K? Maybe Not, We Say

Le risque thrombogène induit par l’infusion portale de cellules souches mésenchymateuse hépatiques humaines chez le rat Wistar peut être contrôlé par un cocktail d’anticoagulants, comprenant l’héparine et la bivalirudine.

Contexte : La thérapie cellulaire par cellules souches mésenchymateuse (MSC) est un traitement en plein essor. Cependant les MSCs présentent une activité procoagulante (APC) induisant un risque thrombogène chez les patients receveurs. Ce risque résulte d’une activation de la cascade de coagulation, initié par le Facteur Tissulaire (FT), conduisant in vitro à une production de complexe thrombine-antithrombine (TAT), de fibrine et une consommation de plaquettes. L’APC des MSCs hépatiques humaines peut être contrôlée in vitro par un cocktail d’anticoagulants, comprenant un activateur de l’antithrombine (Héparine) et un inhibiteur de la thrombine (Bivalirudine). But de l’étude : L’objectif principal est de réduire le risque thrombogène induit par l’infusion cellulaire chez les patients receveurs. Pour cela nous étudions in vivo l’APC des MSCs hépatiques humaines et l’effet du cocktail anticoagulant sur celle-ci. Patients et méthodes de l'étude : Différentes doses de MSCs hépatiques humaines (50, 12.5 et 5 million cellules/kg) ont été infusés par voie intraportale à des rats Wistar avec ou sans cocktail anticoagulant (n=3 par condition). Du PBS a été administré au groupe contrôle. Des prélèvements sanguins ont été réalisés avant et 1 heure après l’infusion, au sacrifice. Par un système d’imagerie digitale, Visiopharm et TissueIA, nous avons analysé la présence de fibrine et des cellules sur des coupes sériées hépatiques. Résultats de l'étude : L’infusion cellulaire de 50 millions cellules/kg tend à induire après 1heure une consommation plaquettaire avec production du complexe TAT, absent dans le groupe contrôle (PBS). L’anticoagulation à tendance à réduire cette chute de plaquettes, de 70% ± 24 à 18% ± 7. L’infusion de 12.5x106cellules/kg tend à diminuer la chute plaquettaire et la production du TAT, et à être absente avec l’administration d’anticoagulation. L’infusion de 5x106cellules/kg n’induit pas de chute plaquettaire ou de production du TAT. L’infusion de 50 et 12.5x106cellules/kg a induit une production de fibrine entourant les ADHLSCs dans les veines portes (VP). L’anticoagulant diminue significativement (p <0.01) cette production de fibrine de 85% ± 8 à 29% ± 10, non influencée par le dosage cellulaire. De nouvelles expériences seront effectuées pour augmenter le nombre de rat par condition et obtenir des valeurs statistiquement significatives. Conclusion: L’infusion intraportale de MSCs hépatiques humaines active la cascade de coagulation in vivo 1 heure après infusion. Cependant en utilisant des dosages cellulaires plus faibles et/ou un cocktail anticoagulant, héparine et bivalirudine, le risque thrombogène peut être contrôlé

The thrombogenic risk induced by intraportal infusion of Adult Derived Human Liver Stem/Mesenchymal Cells in Wistar rats can be controlled with a combination of anticoagulant drugs, heparin and bivalirudin.

Intro : Mesenchymal Stem cell (MSC) transplantation is a fast emerging therapy for regenerative medicine. However MSCs express a procoagulant activity (PCA) inducing a thrombogenic risk in recipient patients. This thrombogenic risk is due to activation of the coagulation cascade, triggered by Tissue Factor (TF) on MSCs, resulting in fibrin production and platelet consumption. In vitro the PCA of Adult Derived Human Liver Stem/Mesenchymal Cells (ADHLSCs) can be controlled by an anticoagulant cocktail, combining an antithrombin activator (Heparin) and a thrombin inhibitor (Bivalirudin). Aim : The aim was to study in vivo, the effect of an anticoagulant cocktail on thrombogenic risk induced by intraportal ADHLSCs. First we wanted to characterize the thrombogenic events induced by ADHLSCs infusion. Then we wanted to study the effect of the anticoagulant cocktail on these events. Finally the effect of cell dosage on the thrombogenic risk was studied. Method : Wistar rats (n=3 per group) were infused with ADHLSCs through an intraportal catheter. Rats were infused with 3 different cell dosages (50, 12.5 and 5 million ADHLSCs/kg), with or without administration of anticoagulant therapy (heparin and bivalirudin). Control group was infused with PBS. Blood samples were collected before and 1 hour after cell infusion. Hemoglobin and platelet levels were analysed with an automated hematology analyzer, Cell-Dynn Emerald (Abbott). Rats were sacrified and liver tissue was collected 1 hour after cell infusion. Serial slides were performed to analyse the localisation of AHDLSCs (human Beta 1 integrin staining) and fibrin (P.T.A.H. coloration). Cell quantification and localisation was analysed with digital imaging system, Visiopharm. Fibrin was analysed in Portal Veins (PV) containing ADHLSCs by digital imaging system, TissueIA. Result : ADHLSCs intraportal infusion induced after 1 hour a significant decrease (p < 0.05) in platelet count with production of fibrin around the infused cells localized in PVs. No platelet consumption or fibrin production was observed in the control (PBS) group. In the anticoagulated group, platelets also decreased significantly (p < 0.05) but fibrin production was significantly (p < 0.001) reduced, from 86.6% ± 6.4 to 26.4% ± 13.8. When different cell dosages were infused, we observed a correlation between the number of infused cells and the number of cells in the liver. The number of cells in liver tissue (cells/mm2) and the number of PVs containing cells (PVs containing cells/total PV) were significantly (p < 0.01) lower when lower cell dosages were used, compared to the higher cell dosage group. Consumption of platelets was also correlated to cell dosage. No decrease in platelets was observed when 5x106cells/kg were infused without anticoagulation, and when 12.5x106cells/kg were infused with anticoagulation. Platelets tended to decrease less in the high cell dose group, 50x106cells/kg when anticoagulant therapy was administered. No significant difference has been observed in the production of fibrin in PVs between the different cell dosages. In all conditions, hemoglobin did not decrease significantly, meaning that blood sampling did not interfere with our results. Conclusion : Intraportal infusion of ADHLSCs activated the coagulation cascade resulting in fibrin formation and platelet consumption 1hour after cell infusion. The use of a combination of anticoagulant drugs, heparin and bivalirudin, could prevent the formation of fibrin, but a significant decrease in platelets was still observed. However this decrease tends to be less and can even be controlled when lower cell dosages are used. The production of fibrin and thus the activation of the coagulation cascade was not dependent of cell dosage

L'infusion intraportale de cellules souches mésenchymateuses hépatiques humaines chez le rat induit une altération transitoire du flux vasculaire: analyse par microscopie intravitale et anapathologique.

Background and Aims: Previous studies showed that liver Mesenchymal Stem Cells (MSC) express a procoagulant activity (PCA) that can be controlled in vitro by an anticoagulant cocktail, combining an antithrombin activator (Heparin) and a thrombin inhibitor (Bivalirudin). First we would like to confirm in vivo the PCA of human liver MSCs in a rat model, then study it’s inhibition by a specific anticoagulant cocktail and finally assess the effect of this cocktail on cell implantation. The main aim of this study is to control the PCA induced by infused liver MSCs without reducing the implantation of cells. Methods: Wistar rat were transplanted with 50x106/kg fluorescent (cell tracker red) human liver MSCs by intraportal infusion with (n=12) or without (n=13) anticoagulant drugs. Using an intra-femoral catheter we injected FITC-dextran and Hoechst to visualise liver vasculature and cell nuclei. By intravital microscopy (IVM) we analysed the left liver lobe at different time points, 1h-24h-48h and 7days post-infusion to study liver MSCs localisation and their effect on liver microvasculature. By pathological examination, cell localisation, cell implantation, vascular and tissue alterations were studied at the same time points. Serial slides were performed for a standard haematoxylin eosin staining and a human cell staining (human B-1-integrin). Results: By IVM we observed that the transplanted cells in the first hour formed aggregates in the larger liver vessels, then cells migrate to the sinusoids after 24h, to be quickly cleared after 7days. After 24h, large defect of perfusion were observed in both groups, but normal hepatic vascularisation was restored after 48h. These observations were confirmed by pathological examination. Large necrotic zones surrounding the infused cells were observed after 24h, with respectively 14.7% and 19.5% of the liver tissue in the non anticoagulated and anticoagulated group. This necrosis decreased rapidly after 48h to 0.5% and 1.9%. The anticoagulant drugs didn’t prevent this necrosis, no difference has been observed between the 2 groups (p>0.05).We confirmed that the infused cells are rapidly cleared after 24h from the liver over time, with respectively after 1h, 24h, 48h and 7days a decrease from 1.3%, 0.3%, 0.07% till 0.03% of infused cells. No difference in cell implantation has been observed between the groups with or without anticoagulation (p>0.05). Conclusion: Intraportal infusion of human liver MSCs to Wistar rats induces a transient alteration in liver vascular flow after 24h. This could be explained by the temporary localisation of liver MSCs in large portal veins and sinusoids up to 1hour after the infusion, in addition to a possible xenotransplantation acute reaction. After 24h more than 70% of cells are cleared and cells are surrounded by transient necrotico-hemorragic regions that regress almost completely after 48h. After 7days no necrosis and very few cells are seen. We observed no difference between the two groups, with or without anticoagulation. From these results we hypothesise that infused liver-MSCs not only express a PCA, but they also induce an Instant Blood Mediated Inflammatory Reaction (IBMIR) that isn’t controlled by our anticoagulant cocktail. With an in vitro technique called Tubing Loop, who mimics the portal vein circulation, we want to confirm this hypothesis by dosing coagulation (fragment 1-2 prothrombin) and IBMIR (platelets, granulocytes and complement C3A) markers after liver-MSCs are exposed to circulating blood, with or without the anticoagulant cocktail