Shaorui Hao

Immediate intraportal transplantation of human bone marrow mesenchymal stem cells prevents death from fulminant hepatic failure in pigs

The effectiveness of human bone marrow mesenchymal stem cell (hBMSC) transplantation to treat acute and chronic liver injury has been demonstrated in animal models and in a few nonrandomized clinical trials. However, no studies have investigated hBMSC transplantation in the treatment of fulminant hepatic failure (FHF), especially in large animal (pig) models. The aim of this study was to demonstrate the safety, effectiveness, and underlying mechanism of hBMSC transplantation for treating FHF in pigs through the intraportal route. Human BMSCs (3 × 107) were transplanted into pigs with FHF via the intraportal route or peripheral vein immediately after D‐galactosamine injection, and a sham group underwent intraportal transplantation (IPT) without cells (IPT, peripheral vein transplantation [PVT], and control groups, respectively, n = 15 per group). All of the animals in the PVT and control groups died of FHF within 96 hours. In contrast, 13 of 15 animals in the IPT group achieved long‐term survival (>6 months). Immunohistochemistry demonstrated that transplanted hBMSC‐derived hepatocytes in surviving animals were widely distributed in the hepatic lobules and the liver parenchyma from weeks 2 to 10. Thirty percent of the hepatocytes were hBMSC‐derived. However, the number of transplanted cells decreased significantly at week 15. Only a few single cells were scattered in the regenerated liver lobules at week 20, and the liver tissues exhibited a nearly normal structure. Conclusion: Immediate IPT of hBMSCs is a safe and effective treatment for FHF. The transplanted hBMSCs may quickly participate in liver regeneration via proliferation and transdifferentiation into hepatocytes during the initial stage of FHF. This method can possibly be used in future clinical therapy. (HEPATOLOGY 2012;56:1044–1052)

Quantitative evaluation of human bone mesenchymal stem cells rescuing fulminant hepatic failure in pigs

Objective Stem cell transplantation provides a promising alternative for the treatment of fulminant hepatic failure (FHF). However, it lacks fundamental understanding of stem cells’ activities. Our objective was to clarify stem cell-recipient interactions for overcoming barriers to clinical application. Design We used an in-house large-animal (pig) model of FHF rescue by human bone marrow mesenchymal stem cells (hBMSCs) and profiled the cells’ activities. The control and transplantation groups of pigs (n=15 per group) both received a D-galactosamine (D-Gal) injection (1.5 g/kg). The transplantation group received hBMSCs via intraportal vein infusion (3×106 cells/kg) immediately after D-Gal administration. The stem cell-recipient interactions were quantitatively evaluated by biochemical function, cytokine array, metabolite profiling, transcriptome sequencing and immunohistochemistry. Results All pigs in the control group died within an average of 3.22 days, whereas 13/15 pigs in the transplantation group lived >14 days. The cytokine array and metabolite profiling analyses revealed that hBMSC transplantation suppressed D-Gal-induced life-threatening cytokine storms and stabilised FHF within 7 days, while human-derived hepatocytes constituted only ∼4.5% of the pig hepatocytes. The functional synergy analysis of the observed profile changes indicated that the implanted hBMSCs altered the pigs’ cytokine responses to damage through paracrine effects. Delta-like ligand 4 was validated to assist liver restoration in both pig and rat FHF models. Conclusions Our results delineated an integrated model of the multifaceted interactions between stem cells and recipients, which may open a new avenue to the discovery of single molecule-based therapeutics that simulate stem cell actions.

Plasma exchange-centered artificial liver support system in hepatitis B virus-related acute-on-chronic liver failure: a nationwide prospective multicenter study in China

BACKGROUND Plasma exchange (PE)-centered artificial liver support system reduced the high mortality rate of hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF). But the data were diverse in different medical centers. The present prospective nationwide study was to evaluate the effects of PE on patients with HBV-ACLF at different stages. METHODS From December 2009 to December 2011, we evaluated 250 patients at different stages of HBV-ACLF from 10 major medical centers in China. All the laboratory parameters were collected at admission, before and after PE. RESULTS Among the 250 patients who underwent 661 rounds of PE, one-month survival rate was 61.6%; 141 (56.4%) showed improvement after PE. Variables such as age (P=0.000), levels of total bilirubin (TB, P=0.000), direct bilirubin (P=0.000), total triglycerides (P=0.000), low-density lipoprotein (P=0.022), Na+ (P=0.014), Cl- (P=0.038), creatinine (Cr, P=0.007), fibrinogen (P=0.000), prothrombin time (PT, P=0.000), white blood cell (P=0.000), platelet (P=0.003) and MELD (P=0.000) were significantly related to prognosis. Multivariate logistic regression analysis showed that age, disease stage, TB, Cr and PT levels were independent risk factors of mortality among HBV-ACLF patients. CONCLUSIONS PE can improve the clinical outcome of patients with HBV-ACLF. Levels of TB, Cr and PT, age and disease stage help to predict prognosis.

A serum metabolomic analysis for diagnosis and biomarker discovery of primary biliary cirrhosis and autoimmune hepatitis.

BACKGROUND Because of the diversity of the clinical and laboratory manifestations, the diagnosis of autoimmune liver disease (AILD) remains a challenge in clinical practice. The value of metabolomics has been studied in the diagnosis of many diseases. The present study aimed to determine whether the metabolic profiles, based on ultraperformance liquid chromatography-mass spectrometry (UPLC-MS), differed between autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC), to identify specific metabolomic markers, and to establish a model for the diagnosis of AIH and PBC. METHODS Serum samples were collected from 20 patients with PBC, 19 patients with AIH, and 25 healthy individuals. UPLC-MS data of the samples were analyzed using principal component analysis, partial least squares discrimination analysis and orthogonal partial least squares discrimination analysis. RESULTS The partial least squares discrimination analysis model (R2Y=0.991, Q2=0.943) was established between the AIH and PBC groups and exhibited both sensitivity and specificity of 100%. Five groups of biomarkers were identified, including bile acids, free fatty acids, phosphatidylcholines, lysolecithins and sphingomyelin. Bile acids significantly increased in the AIH and PBC groups compared with the healthy control group. The other biomarkers decreased in the AIH and PBC groups compared with those in the healthy control group. In addition, the biomarkers were downregulated in the AIH group compared with the PBC group. CONCLUSIONS The biomarkers identified revealed the pathophysiological changes in AILD and helped to discriminate between AIH and PBC. The predictability of this method suggests its potential application in the diagnosis of AILD.

Development of diagnostic criteria and a prognostic score for hepatitis B virus-related acute-on-chronic liver failure

Objective The definition of acute-on-chronic liver failure (ACLF) based on cirrhosis, irrespective of aetiology, remains controversial. This study aimed to clarify the clinicopathological characteristics of patients with hepatitis B virus-related ACLF (HBV-ACLF) in a prospective study and develop new diagnostic criteria and a prognostic score for such patients. Design The clinical data from 1322 hospitalised patients with acute decompensation of cirrhosis or severe liver injury due to chronic hepatitis B (CHB) at 13 liver centres in China were used to develop new diagnostic and prognostic criteria. Results Of the patients assessed using the Chronic Liver Failure Consortium criteria with the exception of cirrhosis, 391 patients with ACLF were identified: 92 with non-cirrhotic HBV-ACLF, 271 with cirrhotic HBV-ACLF and 28 with ACLF with cirrhosis caused by non-HBV aetiologies (non-HBV-ACLF). The short-term (28/90 days) mortality of the patients with HBV-ACLF were significantly higher than those of the patients with non-HBV-ACLF. Total bilirubin (TB) ≥12 mg/dL and an international normalised ratio (INR) ≥1.5 was proposed as an additional diagnostic indicator of HBV-ACLF, and 19.3% of patients with an HBV aetiology were additionally diagnosed with ACLF. The new prognostic score (0.741×INR+0.523×HBV-SOFA+0.026×age+0.003×TB) for short-term mortality was superior to five other scores based on both discovery and external validation studies. Conclusions Regardless of the presence of cirrhosis, patients with CHB, TB ≥12 mg/dL and INR ≥1.5 should be diagnosed with ACLF. The new criteria diagnosed nearly 20% more patients with an HBV aetiology with ACLF, thus increasing their opportunity to receive timely intensive management.

Serum macrophage inflammatory protein 3α levels predict the severity of HBV-related acute-on-chronic liver failure

Dear Editor, We read with interest the paper by Angeli et al 1 proposing the value of the acute-on-chronic liver failure (ACLF) stratification in the prediction of short-term mortality in patients with acute decompensation of cirrhosis. Other studies2 ,3 indicated that the dynamic changes in serological markers are associated with patients who have cirrhosis. We hypothesised that the pathological processes of HBV-related ACLF (HBV-ACLF) would result in specific changes in the levels of signalling proteins in the blood. Thus, we aimed to search detectable biomarkers to predict the outcome of HBV-ACLF. We collected serum samples from 420 patients with HBV-ACLF, 121 patients with chronic hepatitis B (CHB) and 25 normal controls to identify novel serological biomarkers of HBV-ACLF (see online supplementary table S1). As an initial screening group, 15 subjects were included in the cytokine antibody array analyses (n=5 per group). The remaining 551 subjects were included in the ELISA measurement group. The enrolment criteria for the patients with HBV-ACLF corresponded to the previously published the chronic liver failure-sequential organ failure assessment (CLIF-SOFA) score.4 The bioinformatics analyses of …

Dynamic Patterns of serum metabolites in fulminant hepatic failure pigs

Fulminant hepatic failure (FHF) is still an intractable disease associated with serious metabolic disorder. Investigating the dynamic changes of serum metabolites during the development of FHF would facilitate revealing the pathogenesis and also promote its treatment. Therefore, this study characterized the dynamic metabonome of serum from FHF Pigs using ultra performance liquid chromatography–mass spectrometry. Based on multiple statistical analysis of the resulting dataset, three types of up-regulated and one type of down-regulated patterns were delineated. Each pattern demonstrated distinct trends at different stages during the whole process of FHF, implying the differential clinical significance of them. Specifically, aromatic amino acids (Pattern 1) and lysophosphatidylcholines (LPCs) (Pattern 4) might be good markers for evaluating the severity of FHF, while some conjugated bile acids, long chain acylcarnitines (Pattern 2) and Glycocholic acid (Pattern 3) could indicate liver injury in the early stage. Inspired from the PCA plot that the pathogenetic condition of FHF aggravated with sampling time, a linear discriminant analysis (LDA) model based on phenylalanine and LPC 18:1 were further constructed for evaluating the severity of FHF. The leave-one-out cross-validation accuracy of 91.67% for the training set and the prediction accuracy of 92.31% for the external validation set confirmed its excellent performance. In conclusion, findings obtained from the present study, including four types of Dynamic Patterns of serum metabolites during FHF development and an LDA model for evaluating the severity of FHF, will be of great help to the research and management of FHF in the future.

Return of the metabolic trajectory to the original area after human bone marrow mesenchymal stem cell transplantation for the treatment of fulminant hepatic failure.

Our recent study first demonstrated that human bone marrow mesenchymal stem cell (hBMSC) transplantation could prevent death from fulminant hepatic failure (FHF) in pigs. To further clarify the metabolic mechanism of hBMSC transplantation in FHF, the plasma collected from FHF pigs that received transplantation of hBMSCs was examined using metabolic analysis to identify the key molecular markers that regulate recovery. The results showed that obvious metabolic disturbance occurred during FHF, whereas the hBMSC transplantation group showed less severe liver injury. The metabolic trajectory returns to its original state at week 3 following the hBMSC transplantation. In total, the concentration of 26 metabolites, including conjugated bile acids, phosphatidylcholines, lysophosphatidylcholines, fatty acids, amino acid and sphingomyelin, are significantly different between the FHF group and the hBMSC transplantation group. Moreover, the time course of changes in the metabolites corresponded with that of the biochemical and histological analyses. Real-time PCR further confirmed that the gene expression of phospholipase A1, lecithin-cholesterol acyltransferase and lysophosphatidylcholine acyltransferase 1 decreased significantly, whereas that of phospholipase A2 remained stable, which explains the decrease of the phosphatidylcholines and lysophosphatidylcholines. These novel results have revealed a metabolic mechanism for the hBMSC transplantation in FHF, which could lead to the future development of treatment strategies for stem cell therapies.

Human bone marrow mesenchymal stem cell‐derived hepatocytes express tissue inhibitor of metalloproteinases 4 and follistatin

Human bone marrow mesenchymal stem cell (hBMSC) transplantation is expected to become an alternative regenerative technique for liver diseases. However, the mechanism by which hBMSCs differentiate into hepatocytes is still unclear. The aim of this study was to establish the specific characteristics of hBMSC‐derived hepatocytes (hBMSC‐Heps) for future clinical applications.

Intelligent diagnosis of jaundice with dynamic uncertain causality graph model

Jaundice is a common and complex clinical symptom potentially occurring in hepatology, general surgery, pediatrics, infectious diseases, gynecology, and obstetrics, and it is fairly difficult to distinguish the cause of jaundice in clinical practice, especially for general practitioners in less developed regions. With collaboration between physicians and artificial intelligence engineers, a comprehensive knowledge base relevant to jaundice was created based on demographic information, symptoms, physical signs, laboratory tests, imaging diagnosis, medical histories, and risk factors. Then a diagnostic modeling and reasoning system using the dynamic uncertain causality graph was proposed. A modularized modeling scheme was presented to reduce the complexity of model construction, providing multiple perspectives and arbitrary granularity for disease causality representations. A “chaining” inference algorithm and weighted logic operation mechanism were employed to guarantee the exactness and efficiency of diagnostic reasoning under situations of incomplete and uncertain information. Moreover, the causal interactions among diseases and symptoms intuitively demonstrated the reasoning process in a graphical manner. Verification was performed using 203 randomly pooled clinical cases, and the accuracy was 99.01% and 84.73%, respectively, with or without laboratory tests in the model. The solutions were more explicable and convincing than common methods such as Bayesian Networks, further increasing the objectivity of clinical decision-making. The promising results indicated that our model could be potentially used in intelligent diagnosis and help decrease public health expenditure.摘 要目 的黄疸待查是一个常见而复杂的临床问题, 涉及到 内、外、妇、儿等多个学科。目前我国医学专家 存在数量相对不足, 分布不均匀等情况, 导致了 区域性和部门性医疗服务水平不足。本研究旨在 建立一个客观的黄疸待查智能诊断系统, 以提高 医学诊断的正确性, 提升基层医院及急诊的诊断 水平, 同时减少病人的花费。创新点本研究采用了国际先进的动态不确定性因果图 (DUCG)模型, 建立了黄疸待查相关疾病的知 识库, 通过203 例临床病例的测试, 其准确率达 99.01%。文章以图形化的方式给出了疾病的诊断 过程, 方便医师理解和学习。方法本研究采用了DUCG 模型进行疾病诊断, 首先根 据DUCG 模型的定义和黄疸诊断思路建立了包 含27 种黄疸相关疾病(表4)的知识库(图2), 其中包括了疾病的危险因素、临床症状和体征、客观检查检验结果等。然后与根据DUCG 算法 (公式1–4)编写的推理软件相结合形成诊断系 统, 对203 例临床黄疸患者进行智能诊断, 准确 率达99.01%。最后对一例丙型病毒性肝炎患者 的具体诊断过程进行了拆解阐述, 体现了DUCG 模型适用于复杂逻辑关系、计算效率高、不依赖 推理概率和结果易于理解等优点。结论DUCG 模型成功实现了对黄疸待查相关疾病的智 能诊断, 准确率高, 实用性好。该方法具有在其 他医学领域推广应用的价值。