Zhengran Li

Partial splenic embolization for hypersplenism in cirrhosis: A long-term outcome in 62 patients

BACKGROUND Although partial splenic embolization (PSE) has been widely used for treatment of leucocytopaenia and thrombocytopaenia in cirrhosis, only few studies on the correlation between splenic infarction rate and long-term outcome of partial splenic embolization have been reported so far. AIM To evaluate long-term results of partial splenic embolization with different infarction rates in cirrhotic patients with hypersplenism. METHODS Sixty-two consecutive patients with hypersplenism in cirrhosis received partial splenic embolization. According to the splenic infarction rate after partial splenic embolization, the patients were divided into three groups: more than 70% in group A (n=12), 50-70% in group B (n=34), and less than 50% in group C (n=16). The post-partial splenic embolization following-up time was 5 years. RESULTS Before partial splenic embolization, there were no significant differences among the three groups with respect to sex, age, splenic volume, Child-Pugh class, oesophageal varices, and peripheral blood cell counts. After partial splenic embolization, the short- and long-term outcomes of leucocyte and platelet counts showed significant difference among the three groups (P<0.001). In groups A and B, the leucocyte and platelet counts after partial splenic embolization remained significantly higher than those before partial splenic embolization for 2 weeks to 5 years (P<0.05), the post-partial splenic embolization leucocyte and platelet counts was even higher in group A than in group B; while in group C, leucocyte and platelet count improvement only lasted for 6 months after partial splenic embolization. No significant changes were observed concerning blood red cell counts and liver function parameters after partial splenic embolization among the three groups. Severe complications occurred in six patients (50%) in group A and three patients (8.8%) in group B (P<0.05), while in group C, no severe complications developed. CONCLUSIONS In partial splenic embolization, the splenic infarction rate should be limited to 50%-70% in order to ensure the long-term efficacy in alleviating hypersplenism and reduce complications.

Optimization of mesenchymal stem cells (MSCs) delivery dose and route in mice with acute liver injury by bioluminescence imaging.

PurposeBoth experimental and initial clinical studies have shown the therapeutic potential of mesenchymal stem cells (MSCs) in liver disease. Noninvasive tracking of MSCs could facilitate its clinical translation. The purpose of this study was to optimize MSCs delivery dose and route in mice with acute liver injury using bioluminescence imaging (BLI) to track the cells.ProceduresMSCs were labeled with the Luc2-mKate2 dual-fusion reporter gene (MSCs-R). The fate of MSCs-R was tracked through in vivo BLI after administration of different doses or delivery through different routes.ResultsWhen delivered via the superior mesenteric vein (SMV), the high-dose (1.0 × 106 and 5.0 × 105) group mice demonstrated high liver BLI signal but also had lethal portal vein embolization (PVE). By contrast, no PVE and its related death occurred in the low-dose (2.5 × 105) group mice. Thus, 2.5 × 105 is the optimal delivery dose. Three delivery routes, i.e., inferior vena cava (IVC), SMV, and intrahepatic (IH) injection, were also systematically compared. After IVC infusion, MSCs-R were quickly trapped inside the lungs, and no detectable homing to the liver and other organs was observed. By IH injection, lung entrapment was bypassed, but MSCs-R distribution was only localized in the injection region of the liver. By contrast, after SMV infusion, MSCs-R were dispersedly distributed and stayed as long as 7-day posttransplantation in the liver. The in vivo imaging results were further validated by ex vivo imaging, digital subtraction angiography (DSA), and tissue analysis. Therefore, SMV is the optimal MSCs delivery route for liver disease.ConclusionsCollectively, BLI, which could dynamically and quantitatively track cellular location and survival, is useful in determining MSCs transplantation parameters.

Different methods of detaching adherent cells significantly affect the detection of TRAIL receptors.

AIMS AND BACKGROUND As a powerful technique allowing analysis of large numbers of cells, fluorescence-activated cell sorting (FACS) is used more and more widely. For FACS analysis, adherent cells are usually detached by trypsinization, followed by centrifugation and resuspension. However, trypsinization can cut off some receptors from the cell surface like fine scissors, which will affect the accuracy of FACS results. Though non-enzymatic methods such as citric saline buffer have been used to determine cell surface receptors, how much of the receptors is cut off by trypsinization has been rarely studied. This work aimed to investigate whether different methods of detaching adherent cells could affect the detection of cell surface receptors. METHODS Human hepatocellular carcinoma cell lines (HepG2, Huh7 and Hep3B) were detached enzymatically with trypsin-EDTA solution or non-enzymatically with citric saline buffer, and then the receptors of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) were detected by FACS analysis. Cell viability, cell cycle and apoptosis (sub-G1 fraction detected by FACS) of the trypsin-EDTA group and citric saline buffer group were also studied. RESULTS Different methods of detaching adherent cells could significantly affect the detection of TRAIL receptors. Compared to the conventional trypsin-EDTA group, the non-enzymatic group showed a 3.42-fold increase in the mean fluorescence intensity index of DcR HepG2 and a 1.25-fold increase in DR Huh 7 (P <0.05). However, the viability, cell cycle and apoptosis of these cells were not affected. CONCLUSIONS Citric saline buffer might be recommended as the first choice to detach adherent cells for FACS analysis of cell surface receptors.

The inhibitory effect of MSCs expressing TRAIL as a cellular delivery vehicle in combination with cisplatin on hepatocellular carcinoma

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been demonstrated to induce cell apoptosis in many types of tumors, while many hepatocellular carcinoma (HCC) cells display high resistance to TRAIL. Another outstanding limitation of TRAIL is the short half-life in vivo. Stem cell-based therapies provide a promising approach for the treatment of many types of tumors because of the ability of tropism. Therefore, as a new therapeutic strategy, the combination of chemotherapeutic agents and TRAIL gene modified MSCs (TRAIL-MSCs) would improve the therapeutic efficacy of HCC in vivo. This is the first time to show the potential of combination of chemotherapeutic agents and MSCs as a gene vector in the therapy of HCC.

Cisplatin sensitizes human hepatocellular carcinoma cells, but not hepatocytes and mesenchymal stem cells, to TRAIL within a therapeutic window partially depending on the upregulation of DR5.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family of cytokines and has been shown to induce cell apoptosis in many types of tumors, but not in normal cells. This tumor-selective property has made TRAIL a promising approach for the development of cancer therapy. However, hepatocellular carcinoma (HCC) cells display a striking resistance to TRAIL. Although some chemotherapeutic agents can overcome this resistance, safety issues remain a concern because the combination of these agents and TRAIL has been reported to induce toxicity in normal hepatocytes. In this study, we examined whether cisplatin could reverse TRAIL resistance in HCC cells with different p53 status and evaluated the toxicity of combination TRAIL and cisplatin to normal hepatocytes and mesenchymal stem cells (MSCs). We observed that cisplatin could efficiently sensitize HCC cells, but not hepatocytes and MSCs to TRAIL-induced apoptosis within a wide therapeutic window. The apoptosis of HCC cells only partially depended on the upregulation of DR5 and the status of p53. In addition, we provide favorable evidence supporting the feasibility of the combination of chemotherapy and MSCs transduced with TRAIL.

MR tracking of SPIO-labeled mesenchymal stem cells in rats with liver fibrosis could not monitor the cells accurately

Our previous study showed that in vivo magnetic resonance (MR) imaging is effective in tracking superparamagnetic iron oxide (SPIO)-labeled bone marrow mesenchymal stem cells (BMSCs) in rats with liver fibrosis. SPIO-labeling-induced signal reduction on MR images was completely reversed within 15 days after transplantation. It is still unclear whether the signal changes in MR imaging could reflect the number of transplanted cells in the liver. In the present study, BMSCs of male rats were doubly labeled with enhanced green fluorescent protein (EGFP) and SPIO and injected intravascularly into female rats with liver fibrosis. At different time points after injection, MR imaging was performed. The distribution of SPIO particles and EGFP-positive cells was determined by Prussian blue staining and EGFP immunohistochemistry, respectively. The distribution of transplanted BMSCs in various organs was assessed by detection of the SRY gene using real-time quantitative PCR. At 15 days post transplantation, the numbers of transplanted cells were significantly decreased in the lung, kidney, spleen and muscle, but not liver and heart, in comparison with those at 7 days after transplantation. EGFP staining-positive cells were observed in the liver intralobular parenchyma, while Prussian blue staining was negative at 42 days after transplantation. Taken together, SPIO particles and EGFP-labeled BMSCs show a different tissue distribution pattern in rats with liver fibrosis after a long-term period of monitoring. SPIO-based MR imaging may not be suitable for long-term tracking of transplanted BMSCs in vivo.

Transvenous embolization of cavernous sinus dural arteriovenous fistulas using detachable coils and Glubran 2 acrylic glue via the inferior petrosal sinus approach.

Objectives:To describe the technique, efficacy, and safety of transvenous embolisation (TVE) of cavernous sinus arteriovenous fistulas (CSDAVFs) via the inferior petrosal sinus (IPS) with detachable coils and acrylic glue.Methods:Spontaneous unilateral CSDAVFs were confirmed by cerebral angiography in eight patients, with angiographic patency of the ipsilateral IPS in three and angiographic non-visualisation of the ipsilateral IPS in five. There were two patients with complete occlusion of the ipsilateral internal jugular vein (IJV). TVE with detachable coils and acrylic glue were performed through a femoral vein and an IPS approach.Results:TVE viaipsilateral IPS was successfully performed in all eight patients in our group. The number of detachable coils for each patient ranged from 2 to 8 (mean, 5.0). Angiography immediately after TVE showed complete occlusion of the CSCAVFs in seven patients and nearly complete occlusion in one. Complete recovery of clinical symptoms was achieved in all eight patients. No recurrence of clinical symptoms was observed at follow-up.Conclusions:Transvenous embolisation via an IPS approach is a highly efficient and safe treatment for CSDAVFs. Embolisation with a combination of coils and acrylic glue may help to achieve complete occlusion of fistulas with fewer coils.

In Vivo Bioluminescence Imaging of Transplanted Mesenchymal Stromal Cells and Their Rejection Mediated by Intrahepatic NK Cells.

PurposeMesenchymal stromal cells (MSCs) hold promise in the treatment of liver disease. However, short survival time of MSCs after intrahepatic transplantation limits their value; therefore, understanding the basis of MSCs survival and rejection may increase their utility. This study was aimed at determining the role of intrahepatic natural killer (NK) cells on MSCs survival and their retention in the liver shortly after transplant.ProceduresHuman MSCs were labeled with the Luc2-mKate2 dual-fusion reporter gene (MSCs-R), and the residence time and survival of MSCs-R xenografts after intrahepatic transplantation were evaluated by in vivo bioluminescence imaging (BLI). Coculture of MSCs and NK cells was performed to assess cytotoxicity. To evaluate the role of NK cells in rejection of the xenografted cells, the fates of transplanted MSCs-R were then assessed in vivo by BLI after activation of intrahepatic NK cells.ResultsWe observed a linear correlation between luciferase activity from live MSCs-R and cell number in vitro (R2 = 0.9956). In vivo, we observed a gradual decline in bioluminescent signals from transplanted MSCs-R over a region corresponding to the liver in both the control group and the NK-activated group. However, the survival time and retention of intrahepatic MSCs-R decreased more rapidly in the NK-activated group of mice compared to the control group. This indicated that activated NK cells accelerate the elimination of transplanted MSCs. Also, we found that the number of hepatic NK cells and the expression of NK activation markers significantly increased after intrahepatic delivery of MSCs. This suggested that resident NK cells, in a resting state, were activated by intrahepatic transplantation of human MSCs. Taken together, the data suggests that activated hepatic NK cells mediate, in part, rejection of the MSCs xenografts. Cytotoxicity assays showed that activated NK cells may inhibit the proliferation of MSCs and, to a certain extent, induce MSCs death.ConclusionHuman MSCs could be followed dynamically in vivo by BLI, and the role of murine hepatic NK cells, especially activated NK cells, could be inferred from the loss of signals from MSCs. This finding may have practical clinical implications in MSCs transplantation in treating liver disease.

Hepatocellular Carcinoma Cells Carrying a Multimodality Reporter Gene for Fluorescence, Bioluminescence, and Magnetic Resonance Imaging In Vitro and In Vivo.

RATIONALE AND OBJECTIVES The study aimed to evaluate the feasibility of imaging or tracking hepatocellular carcinoma cells by modifying these cells to carry a multimodality reporter gene, enabling fluorescence, bioluminescence, and magnetic resonance imaging (MRI) in vitro and in vivo. MATERIALS AND METHODS HepG2 cells were labeled with the enhanced green fluorescent protein (EGFP)/luciferase2/ferritin-the multimodality reporter gene (labeled HepG2 cells). The labeled and unlabeled HepG2 cells were cultured in vitro and then injected subcutaneously into mice as a hepatoma model in vivo. The expressions of EGFP, luciferase2, and ferritin in HepG2 cell suspensions and hepatoma model were investigated using fluorescence, bioluminescence, and MRI. RESULTS Individual HepG2 cells expressing EGFP were identified under blue laser excitation. The linear coefficient between the optical signal intensity of luciferase2 and the number of labeled cells was 0.993. MRI was used to distinguish the T2* signal of 2 × 107 cells/mL between the labeled (6.67 ± 1.88 ms) and unlabeled cells (10.66 ± 2.22 ms) (P = 0.034). In vivo, individual HepG2 cells expressing EGFP in frozen sections were observed. Labeled cells expressing luciferase2 have been detected since the second day after injection, and the bioluminescence increased with the tumor size. The T2* signal was significantly different between the labeled (6.04 ± 1.60 ms) and unlabeled cells (17.06 ± 2.17 ms) (P <0.001). CONCLUSIONS A multimodality reporter gene consisting of EGFP, luciferase2, and ferritin was successfully integrated into the HepG2 cell genome via a lentiviral vector and was highly expressed in the daughter cells. These cells could be detected by fluorescence, bioluminescence, and MRI in vitro and in vivo.

Percutaneous transhepatic intrahepatic portosystemic shunt for variceal bleeding with chronic portal vein occlusion after splenectomy

ObjectivesThe purpose of this study was to introduce a modified transjugular intrahepatic portosystemic shunt (TIPS), a percutaneous transhepatic intrahepatic portosystemic shunt (PTIPS), and to evaluate its feasibility and efficacy in patients with variceal bleeding with chronic portal vein occlusion (CPVO) after splenectomy.MethodsTwenty-four cirrhotic patients with CPVO after splenectomy who received PTIPS between 2010 and 2015 were included in this retrospective study. The indication was elective control of variceal bleeding. Success rates, effectiveness and complications were evaluated, with comparison of the pre- and post-portosystemic pressure gradient (PPG). Patients’ clinical outcomes and shunt patency were followed periodically.ResultsPTIPS was successfully placed in 22 patients (91.7%) and failed in two. The mean PPG fell from 22.0 ± 4.9 mmHg to 10.6 ± 1.6 mmHg after successful PTIPS (p < 0.05). No fatal procedural complications occurred. During the median follow-up of 29 months, shunt dysfunction occurred in five cases and hepatic encephalopathy in four cases. Three patients died because of rebleeding, hepatic failure and pulmonary disease, respectively. The other patients remained asymptomatic and the shunts patent.ConclusionsWe conclude that PTIPS, as a modified TIPS procedure with a high success rate, is safe and effective for variceal bleeding with CPVO after splenectomy.Key Points• Portal vein occlusion used to be contraindication to transjugular intrahepatic portosystemic shunt.• Portal vein thrombosis is common in patients with previous splenectomy.• We developed a new method, percutaneous transhepatic intrahepatic portosystemic shunt (PTIPS).• PTIPS is feasible in patients with portal vein thrombosis and splenectomy.• PTIPS is effective and safe for these kind of complicated portal hypertension.