Profiles of differential expression of miRNAs in the late stage of red blood cell preservation and their potential roles.

OBJECTIVE: To detect the differentially expressed regulatory miRNAs in the late stage of red blood cell (RBC) preservation and predict their roles. METHODS: Suspended RBCs with different storage periods of 35 day, 42 day, and 50 day were collected for routine blood tests, RNA extraction, and preparation of small RNA sequencing libraries. The constructed libraries were sequenced and the biological functions of differential miRNAs in RBCs in the late storage were analyzed by bioinformatics. RESULTS: Routine indicators of RBCs in the late stage were not significantly affected by preservation time. The Pearson correlation analysis performing on RBC miRNAs with different storage days revealed that RBC miRNAs changed with the increase of storage days. RBC miRNAs from day 35 (D35), day 42 (D42) and day 50 (D50) showed significant differences (P < 0.05). Compared RBC miRNAs from D42 with these from D35, there were 690 up-regulated miRNAs and 82 down-regulated miRNAs; compared RBC miRNAs from D50 with these from D35, there were 638 up-regulated miRNAs and 123 down-regulated miRNAs; compared RBC miRNAs from D42 with these from D50, there were 271 up-regulated miRNAs and 515 down-regulated miRNAs. GO enrichment analysis of target genes of differential miRNAs were mainly involved in cell metabolism, biosynthesis, protein modification, gene expression and transcriptional regulation of biological processes. KEGG pathway enrichment analysis of miRNA target genes showed that differential miRNA target genes were closely related to pathways in cancer. CONCLUSION: MiRNAs were differentially expressed in the late stage of RBC preservation, and may be involved in various biological processes, especially cancer.

Forecasting demands of blood components based on prediction models.

BACKGROUND: An adequate blood supply is an important guarantee for saving lives and protecting health. In order to manage the blood supply more effectively when the condition of demand and supply are uncertainty, it is very important to forecast the demands of blood resources. MATERIALS AND METHODS: SARIMAX model and LSTM model were integrated into the prediction system of blood station. The collection and supply data of blood components was directly imported into the forecasting models to achieve automatic data update and model update. The forecasting daily demands of apheresis platelets, washing red blood cells (RBCs), suspended RBCs and plasma were recorded from January to June 2023 and compared with real data. RESULTS: The prediction models had good forecasting performances. In the goodness of fit results of apheresis platelet model, the maximum value of coefficient of determination (R2) could reach 87.6%, and the minimum value of the mean absolute percentage error (MAPE) was only 0.0037. The predicted data of washing RBCs could be basically fitted, and the MAPE was 0.0121. For the prediction of suspended RBCs, the R2 was greater than 66%, and the MAPE could be 0.0372. The plasma model generated very high goodness of fit results, with R2 of over 90% and the lowest MAPE of 0.0394. CONCLUSION: The forecasting models, which predicts future demands of different blood components based on historical data, can help managers to overcome the challenges of blood stock control more effectively, thereby reducing blood waste and blood shortages.

Platelet-rich plasma in the treatment of anal fistula: a systematic review and meta-analysis.

OBJECTIVE: To analyse the safety and effectiveness of platelet-rich plasma (PRP) in anal fistula patients. METHODS: Online databases including PubMed, Embase, Cochrane Library, and Web of Science were searched from inception to December 5, 2022, for eligible studies about evaluating the efficacy of platelet-rich plasma (PRP) in treating anal fistula. Literature search, screening, data extraction, and quality assessment were carried out by two independent investigators. The overall cure rate, the complete cure rate, the recurrence rate, and the adverse event rate with their 95% confidence intervals (95% CI) were the primary calculation indexes. Subgroup analyses were conducted primarily according to whether PRP was combined with other treatments. Softwares of MedCalc 18.2 and Review Manager 5.3 were used for meta-analysis. RESULTS: A total of 14 studies with 514 patients were included in the meta-analysis. The overall cure rate of 14 studies was 72.11% (95% CI 0.64-0.79). The cure rate of PRP alone was 62.39% (95% CI 0.55-0.69). The combined cure rate of PRP with other treatments was 83.12% (95% CI 0.77-0.88). The cure rate of interventions involving PRP were superior to the cure rate of surgery methods without using PRP significantly in the 4 randomized controlled studies (RR = 1.30, 95% CI 1.10-1.54, p = 0.002). The complete cure rate of the 8 studies was 66.37% (95% CI 0.52-0.79). The recurrence rate of the 12 studies was 14.84% (95% CI 0.08-0.24). The adverse event rate of the 12 studies was 6.31% (95% CI 0.02-0.12). CONCLUSION: PRP showed favorable safety and effectiveness in the treatment of anal fistula, especially combined with other treatment procedures.

Targeted delivery of triptolide by dendritic cell-derived exosomes for colitis and rheumatoid arthritis therapy in murine models.

BACKGROUND AND PURPOSE: Triptolide (TP) elicits a beneficial effect in the treatment of autoimmune diseases, such as ulcerative colitis (UC) and rheumatoid arthritis (RA). However, its multiorgan toxicity needs to be resolved. Dendritic cells (DCs) are the primary target of TP, which induces immunosuppression, and DC-derived exosomes (DEX) can selectively enter DCs in vivo. Here, we encapsulated TP with DEX (DEXTP) to generate TP-targeted delivery to reduce toxicity. EXPERIMENTAL APPROACH: The effect of DEXTP was evaluated in murine colitis and RA models. Toxicity was examined by haematoxylin and eosin staining and serum biochemical marker detection. Affinity of DEXs for DCs was tracked by fluorescent labelling. The immune environment was evaluated and mimicked in vitro for further analysis of the mechanism. KEY RESULTS: DEXTP effectively carried TP to DCs in vivo, and alleviated local inflammation and damage in colitis and RA mice with no obvious toxicity. Additionally, DEXTP reshaped the immune milieu by decreasing CD4+ T-cell levels and increasing regulatory T-cell levels in vivo. Furthermore, consistent T-cell differentiation was observed in vitro, and DC activation was inhibited by alterations in surface factors and secrete cytokines, and by induction of apoptosis or other form of death. CONCLUSIONS AND IMPLICATIONS: Encapsulating TP with DEX is a new method that both reduces the toxicity of TP and induces immunosuppression in UC and RA mice. The underlying immune mechanism involves DEXTP targeting DCs in vivo, to inhibit DC activation and induce DC apoptosis, which further induces T-cell immunosuppression.

Adverse transfusion reactions and what we can do.

INTRODUCTION: Transfusions of blood and blood components have inherent risks and the ensuing adverse reactions. It is very important to understand the adverse reactions of blood transfusion comprehensively for ensuring the safety of any future transfusions. AREAS COVERED: According to the time of onset, adverse reactions of blood transfusion are divided into immediate and delayed transfusion reactions. In acute transfusion reactions, timely identification and immediate cessation of transfusion is critical. Vigilance is required to distinguish delayed responses or reactions that present nonspecific signs and symptoms. In this review, we present the progress of mechanism, clinical characteristics and management of commonly encountered transfusion reactions. EXPERT OPINION: The incidence of many transfusion-related adverse events is decreasing, but threats to transfusion safety are always emerging. It is particularly important for clinicians and blood transfusion staff to recognize the causes, symptoms, and treatment methods of adverse blood transfusion reactions to improve the safety. In the future, at-risk patients will be better identified and can benefit from more closely matched blood components.

Dendritic cell combination therapy reduces the toxicity of triptolide and ameliorates colitis in murine models.

Triptolide (TP) exerts a promising effect in the treatment of ulcerative colitis (UC). However, its toxicity seriously hinders its application in the clinic. Previous studies indicated that dendritic cells (DCs) are the main target through which TP exerts its immunoregulatory effect. Thus, we designed an approach to target DCs in vitro to avoid the direct exposure of organs to TP. Our results revealed that DCs pretreated with TP (DCTP) exerted satisfactory therapeutic effects in mice with colitis, resulting in improved colonic inflammation and alleviated local lesion damage. In addition, no obvious toxicity was observed. DCTP also reshaped the immune milieu by decreasing CD4+ T cell numbers and increasing regulatory T cell numbers in the spleen, mesenteric lymph nodes, peripheral blood and colon; these effects were further confirmed in vitro. Downregulation of CD80/86, ICAM-1, MHCI, TLR2/4, TNF-α, and IL-6 expression and upregulation of programmed cell death ligand 1 (PDL1) and IL-10 expression were observed, indicating that DCs were converted into tolerogenic DCs. In conclusion, DCTP can effectively reduce toxicity and alleviate colonic inflammation and local lesion damage in mice with colitis. The immune mechanism underlying the effects of DCTP included the conversion of DCs into tolerogenic DCs and the alteration of T cell differentiation to produce immunoinhibitory rather than immunostimulatory T cells.

Clinical benefits of PD-1/PD-L1 inhibitors in advanced hepatocellular carcinoma: a systematic review and meta-analysis.

BACKGROUND: Significant improvement of objective response rate and overall survival period has been achieved in several types of solid tumors by treatment with PD-1/PD-L1 inhibitors, which shed some light on hepatocellular carcinoma (HCC). Currently, a number of clinical trials concerning the application of checkpoint inhibitors in HCC are ongoing, some of which have shown favorable expectations. Hereby, we conducted a meta-analysis of existing studies to reveal the efficacy and safety of checkpoint inhibitors in advanced HCC. METHODS: Medline, Embase, Cochrane Library, and Web of Science were searched from inception to January 31, 2020. The clinical trials reporting the efficacy of PD-1/PD-L1 inhibitors in advanced HCC patients were eligible. Overall results of complete response (CR), partial response (PR), stable disease (SD), progression of disease (PD), objective response rate (ORR), disease control rate (DCR), overall survival (OS), progression-free survival (PFS) and rate of adverse events (AE) with their 95% confidence intervals (95%CI) were calculated as the primary focus of the meta-analysis. Subgroup analyses were conducted primarily according to the categories of PD-1 inhibitor or PD-L1 inhibitor and combination therapy or monotherapy. In addition, pooled results of PD-1/PD-L1 monoclonal antibodies (mAb) combining with anti-VEGF agents were calculated separately. RESULTS: A total of 20 studies with 1232 patients were included. The overall CR, PR and SD rate were 0.01 (95% CI 0.01-0.03), 0.17 (95% CI 0.14-0.22) and 0.39 (95% CI 0.34-0.43), respectively. The overall ORR and DCR were 0.20 (95% CI 0.16-0.24) and 0.60 (95% CI 0.54-0.67), respectively. The overall PFS and OS were 3.58 months (95% CI 2.65-4.50) and 12.24 months (95% CI 10.48-14.00), respectively. For patients treated with PD-1/PD-L1 mAb combing with anti-VEGF agent, ORR was 29% (95% CI 0.15-0.43) and DCR was 77% (95% CI 0.70-0.84). For all included studies, the overall rate of AE was 0.63 (95% CI 0.45-0.78) and serious adverse events (SAE) was 0.11 (95% CI 0.06-0.22). CONCLUSIONS: PD-1/PD-L1 inhibitors showed favorable outcomes concerning response rates and survival periods in advanced HCC. Updated results from high-quality clinical trials are expected to validate these findings.

Prognostic value of leukemia inhibitory factor and its receptor in pancreatic adenocarcinoma.

Currently, the prognostic effects of leukemia inhibitory factor (LIF) and LIF receptor (LIFR) in pancreatic adenocarcinoma (PAAD) are not clear. In the present study, we utilized the large datasets from four public databases to investigate the expression of LIF and LIFR and their clinical significance in PAAD. Eight cohorts containing 1278 cases with PAAD were identified and the analysis results suggested that LIF was highly expressed while LIFR was lowly expressed in PAAD tissues compared with adjacent or normal tissues. Kaplan-Meier plot curves and univariate and multivariate Cox proportional hazards regression analyses indicated high LIF expression was associated with shorter overall survival (adjusted hazard ratio = 1.641, 95% CI: 1.399-1.925, p < 0.001) whereas high LIFR expression was associated with longer overall survival (adjusted hazard ratio = 0.653, 95% CI: 0.517-0.826, p < 0.001).

Prognosis comparison between wait and watch and surgical strategy on rectal cancer patients after treatment with neoadjuvant chemoradiotherapy: a meta-analysis.

BACKGROUND: After achieving a clinical complete response through neoadjuvant chemoradiotherapy, a nonoperative management approach for rectal cancer patients known as Wait and Watch (W&W) has gained increasing attention. However, the W&W strategy has been related to higher local recurrence and ambiguous long-term survival. This meta-analysis compared key prognosis indicators between W&W and surgical treatment in an effort to clarify some long-standing points of confusion. METHODS: Pubmed, Web of Science, EMbase, Cochrane Library were searched for relevant researches comparing W&W with surgery treatment, with a time criteria set from 1 January 2002 to 4 July 2019. Endpoints were 2-year local regrowth/recurrence, 2-year distant metastasis (plus local regrowth/recurrence), 3- and 5-year disease-free survival (DFS), and overall survival (OS). RESULTS: In total, nine studies with 801 patients were enrolled, of which 348 were managed by W&W and 453 by surgery. Surgery patients were further divided into a pathological complete response (pCR) group (all included patients achieved pCR) and a surgery group (consisting of both pCR and non-pCR patients without deliberate screening). Compared with the surgery group, W&W patients have higher 3- and 5-year OS, and are not inferior on 2-year local regrowth (LR), 2-year distant metastasis (DM)/DM+LR, and 3- and 5-year DFS. On the other hand, compared with the pCR group, the W&W group is inferior on 2-year LR, 3- and 5-year DFS, and 5-year OS, and not inferior on 2-year DM/DM+LR and 3-year OS. CONCLUSIONS: In contrast with patients undergoing surgical treatment, the W&W group has higher 3- and 5-year OS, and is not inferior on other major prognostic indicators, which, however, is based on the fact that the tumor stage in the W&W group is generally earlier. Versus surgically treated patients who acquired pCR, W&W group is inferior on all major prognostic indicators except 2-year DM/DM+LR and 3-year OS. Additionally, by comparison of cCR definitions across different studies, we conclude that implementation of the strictest cCR criteria is critical for W&W patients to acquire maximum prognostic benefit.

Effect of Triptolide on Dextran Sodium Sulfate-Induced Ulcerative Colitis and Gut Microbiota in Mice.

Triptolide is beneficial for the treatment of ulcerative colitis (UC), which is closely related to the gut microbiota. However, whether the therapeutic effects of triptolide involve the regulation of the gut microbiota is still unclear. In the present study, animal models of UC mice induced by dextran sodium sulfate (DSS) were established, the changes of gut microbiota in mice were detected by high-throughput sequencing. The effects of triptolide on DSS-induced UC mouse and its gut microbiota were studied. As a result, we found that triptolide exerted anti-inflammatory and therapeutic effects on UC mice. Sequencing results for the gut microbiota showed that the composition of the gut microbiota from DSS group was disordered as compared with that from the control group, consistent with a decrease in the abundance of flora. Triptolide treatment accelerated the recovery of the population of the gut microbiota and significantly improved the microbial diversity. At the phylum level, the population of Bacteroidetes decreased and that of Firmicutes increased. At the genus level, Bacteroides and Lachnospiraceae counts decreased. Thus, triptolide could regulate the composition of the gut microbiota, accelerate the recovery of microbiota, and exert good therapeutic effects in UC mice. Our results also revealed that fecal transplantation from triptolide-treated mice could relieve UC. This study provides a reference for the rational use of triptolide for the treatment of UC.

Correction: Intrahepatic Tissue Implantation Represents a Favorable Approach for Establishing Orthotopic Transplantation Hepatocellular Carcinoma Mouse Models.

[This corrects the article DOI: 10.1371/journal.pone.0148263.].

Identification of biomarkers for Barcelona Clinic Liver Cancer staging and overall survival of patients with hepatocellular carcinoma.

The aim of the current study was to identify biomarkers that correlate with the Barcelona Clinic Liver Cancer (BCLC) staging system and prognosis of patients with hepatocellular carcinoma (HCC). We downloaded 4 gene expression datasets from the Gene Expression Omnibus database (http://www.ncbi.nlm.nih.gov/geo), and screened for genes that were differentially expressed between HCC and normal liver tissues, using significance analysis of the microarray algorithm. We used a weighted gene co-expression network analysis (WGCNA) to identify hub genes that correlate with BCLC staging, functional enrichment analysis to associate hub genes with their functions, protein-protein interaction network analysis to identify interactions among hub genes, UALCAN analysis to assess gene expression levels based on tumour stage, and survival analyses to clarify the effects of hub genes on patients' overall survival (OS). We identified 50 relevant hub genes using WGCNA; among them, 13 genes (including TIGD5, C8ORF33, NUDCD1, INSB8, and STIP1) correlated with OS and BCLC staging. Significantly enriched gene ontology biological process terms included RNA processing, non-coding RNA processing and phosphodiester bond hydrolysis, and 6 genes were found to interact with 10 or more hub genes. We identified several candidate biomarkers that correlate with BCLC staging and OS of HCC. These genes might be used for prognostic assessment and selection of HCC patients for surgery, especially those with intermediate or advanced disease.

Proposal for subclassification to select patients for hepatectomy with intermediate hepatocellular carcinoma and Child-Pugh A liver function: A double-center study from China.

Increasing evidence has shown that hepatectomy provides a longer overall survival (OS) for patients with hepatocellular carcinoma (HCC) in the intermediate stage. Unfortunately, not all patients benefit from liver resection, even if hepatectomy is feasible. This study aimed to propose a subclassification to select patients for surgical resection.OS of patients with intermediate-stage HCC who underwent hepatectomy at Beijing Friendship Hospital or Peking Union Medical College Hospital were reviewed. Patients were divided into 2 groups based on the results of survival analysis. The prognosis of these patients was compared with that in those who were treated by trans-arterial chemoembolization (TACE) in each subgroup.A total of 259 patients with intermediate-stage HCC who were initially treated by hepatectomy were included. Multivariate analysis showed that cumulative tumor size and tumor number independently affected tumor recurrence and survival time of these patients. Patients were then divided into group A (tumor size <11 cm and tumor number < 4; n = 205) and group B (tumor size ≥11 cm and tumor number ≥ 4; n = 54). Multivariate analysis showed that hepatectomy was independently associated with longer OS compared with TACE in patients in group A (hazard ratio = 0.67, 95% confidence interval = 0.49-0.90), but not in group B.Surgical management of intermediate-stage HCC should be performed with more complexity than current practice. Hepatic resection could be considered as the first-line treatment only for patients with HCC who have a cumulative tumor size of less than 11 cm and <4 tumors.

Immortalized common marmoset (Callithrix jacchus) hepatic progenitor cells possess bipotentiality in vitro and in vivo.

Common marmoset (Callithrix jacchus) is emerging as a clinically relevant nonhuman primate model for various diseases, but is hindered by the availability of marmoset cell lines, which are critical for understanding the disease pathogenesis and drug/toxicological screening prior to animal testing. Here we describe the generation of immortalized marmoset hepatic progenitor cells (MHPCs) by lentivirus-mediated transfer of the simian virus 40 large T antigen gene in fetal liver polygonal cells. MHPCs proliferate indefinitely in vitro without chromosomal alteration and telomere shortening. These cells possess hepatic progenitor cell-specific gene expression profiles with potential to differentiate into both hepatocytic and cholangiocytic lineages in vitro and in vivo and also can be genetically modified. Importantly, injected MHPCs repopulated the injured liver of fumarylacetoacetate hydrolase (Fah)-deficient mice with hepatocyte-like cells. MHPCs also engraft as cholangiocytes into bile ducts of 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-induced bile ductular injured mice. MHPCs provide a tool to enable efficient derivation and genetic modification of both hepatocytes and cholangiocytes for use in disease modeling, tissue engineering, and drug screening.

Dendritic Cells Pulsed with Exosomes in Combination with PD-1 Antibody Increase the Efficacy of Sorafenib in Hepatocellular Carcinoma Model.

Advanced hepatocellular carcinoma (HCC) has limited therapeutic options. Immunotherapy is a promising treatment, while sorafenib is a first-line drug-based treatment for advanced HCC. However, the efficacy of sorafenib and immunotherapy in combination, have not been clearly evaluated. Sorafenib treatment has been shown to promote immunosuppression by increasing hypoxia in orthotopic HCC models. Here, we found that sorafenib treatment in mice with orthotopic HCC increased the expression of inhibitor programmed death-ligand 1 (PD-L1) and T-regulatory cells in tumor tissues. We pulsed dendritic cells with exosomes derived from tumor cells (DC-TEX) and found that the number of T-regulatory cells decreased and the number of CD8+T cells increased. However, combining DC-TEX and sorafenib did not prolong survival in these mice. Moreover, we found that the number of PD-1+CD8+T cells significantly increased after DC-TEX treatment. Therefore, we next added PD-1 antibody (PD-1 Ab) to the treatment regimen to block the PD-1/PD-L1 pathway, and found that the exhausted CD8+T cells were restored, without affecting the number of T-regulatory cells. Thus, our data suggest that the combination of DC-TEX and PD-1 Ab enhanced the efficacy of sorafenib, but treatment with either DC-TEX or PD-1 Ab alone, did not.

Fluorescent peptide highlights micronodules in murine hepatocellular carcinoma models and humans in vitro.

Early detection and clear delineation of microscopic lesions during surgery are critical to the prognosis and survival of patients with hepatocellular carcinoma (HCC), a devastating malignancy without effective treatments except for resection. Tools to specifically identify and differentiate micronodules from normal tissue in HCC patients can have a positive impact on survival. Here, we discovered a peptide that preferentially binds to HCC cells through phage display. Significant accumulation of the fluorescence-labeled peptide in tumor from ectopic and orthotopic HCC mice was observed within 2 hours of systemic injection. Contrast between tumor and surrounding liver is up to 6.5-fold, and useful contrast lasts for 30 hours. Micronodules (0.03 cm in diameter) in liver and lung can clearly be distinguished from normal tissue with this fluorescence-labeled peptide in orthotopic HCC mice and HCC patients. Compared to indocyanine green, a Food and Drug Administration-approved imaging contrast agent, an up to 8.7-fold higher differentiation ratio of tumor to fibrosis is achieved with this fluorescence-labeled peptide. Importantly, this peptide enables up to 10-fold differentiation between HCC and peritumoral tissue in human tissues and the complete removal of tumor in HCC mice with surgical navigation. No abnormalities in behavior or activity are observed after systemic treatment, indicating the absence of overt toxicity. The peptide is metabolized with a half-life of approximately 4 hours in serum. CONCLUSION: Our findings demonstrate that micronodules can be specifically differentiated with high sensitivity from surrounding tissue with this molecule, opening clinical possibilities for early detection and precise surgery of HCC. (Hepatology 2018).

Dendritic cell-derived exosomes elicit tumor regression in autochthonous hepatocellular carcinoma mouse models.

BACKGROUND & AIMS: Dendritic cell (DC)-derived exosomes (DEXs) form a new class of vaccines for cancer immunotherapy. However, their potency in hepatocellular carcinoma (HCC), a life-threatening malignancy with limited treatment options in the clinic that responds poorly to immunotherapy, remains to be investigated. METHODS: Exosomes derived from α-fetoprotein (AFP)-expressing DCs (DEXAFP) were investigated in three different HCC mouse models systemically. Tumor growth and microenvironment were monitored. RESULTS: DEXAFP elicited strong antigen-specific immune responses and resulted in significant tumor growth retardation and prolonged survival rates in mice with ectopic, orthotopic and carcinogen-induced HCC tumors that displayed antigenic and pathological heterogeneity. The tumor microenvironment was improved in DEXAFP-treated HCC mice, demonstrated by significantly more γ-interferon (IFN-γ)-expressing CD8+ T lymphocytes, elevated levels of IFN-γ and interleukin-2, and fewer CD25+Foxp3+ regulatory T (Treg) cells and decreased levels of interleukin-10 and transforming growth factor-ß in tumor sites. Lack of efficacy in athymic nude mice and CD8+ T cell-depleted mice showed that T cells contribute to DEXAFP-mediated antitumor function. Dynamic examination of the antitumor efficacy and the immune microenvironment in DEXAFP-treated orthotopic HCC mice at different time-points revealed a positive correlation between tumor suppression and immune microenvironment. CONCLUSIONS: Our findings provide evidence that AFP-enriched DEXs can trigger potent antigen-specific antitumor immune responses and reshape the tumor microenvironment in HCC mice and thus provide a cell-free vaccine option for HCC immunotherapy. Lay summary: Dendritic cell (DC)-derived exosomes (DEXs) form a new class of vaccines for cancer immunotherapy. However, their potency in hepatocellular carcinoma (HCC) remains unknown. Here, we investigated exosomes from HCC antigen-expressing DCs in three different HCC mouse models and proved their feasibility and capability of treating HCC, and thus provide a cell-free vaccine for HCC immunotherapy.

Tumor-derived exosomes elicit tumor suppression in murine hepatocellular carcinoma models and humans in vitro.

UNLABELLED: Hepatocellular carcinoma (HCC) remains a global challenge due to high morbidity and mortality rates and poor response to treatment. Immunotherapy, based on introduction of dendritic cells (DCs) activated by tumor cell lysates as antigens ex vivo, shows limited response rates in HCC patients. Here, we demonstrate that tumor cell-derived exosomes (TEXs), displaying an array of HCC antigens, can elicit a stronger immune response than cell lysates in vitro and in vivo. Significant tumor growth inhibition was achieved in ectopic and orthotopic HCC mice treated with TEX-pulsed DCs. Importantly, the tumor immune microenvironment was significantly improved in orthotopic HCC mice treated by TEX-pulsed DCs, demonstrated by increased numbers of T lymphocytes, elevated levels of interferon-γ, and decreased levels of interleukin-10 and tumor growth factor-ß in tumor sites. As expected, T cells played an essential role in the TEX-pulsed DC-mediated immune response. Notably, exosomes from HCC cells not only promoted HCC-specific cytolysis but also provided cross-protective effects against pancreatic cancer cells. Moreover, HCC-specific cytolysis, elicited by DCs pulsed with human HepG2 cell-derived exosomes, was observed across different human HCC cells irrespective of human leukocyte antigen types. CONCLUSION: HCC TEXs can potently carry HCC antigens, trigger a strong DC-mediated immune response, and improve the HCC tumor microenvironment. (Hepatology 2016;64:456-472).

Intrahepatic Tissue Implantation Represents a Favorable Approach for Establishing Orthotopic Transplantation Hepatocellular Carcinoma Mouse Models.

Mouse models are commonly used for studying hepatocellular carcinoma (HCC) biology and exploring new therapeutic interventions. Currently three main modalities of HCC mouse models have been extensively employed in pre-clinical studies including chemically induced, transgenic and transplantation models. Among them, transplantation models are preferred for evaluating in vivo drug efficacy in pre-clinical settings given the short latency, uniformity in size and close resemblance to tumors in patients. However methods used for establishing orthotopic HCC transplantation mouse models are diverse and fragmentized without a comprehensive comparison. Here, we systemically evaluate four different approaches commonly used to establish HCC mice in preclinical studies, including intravenous, intrasplenic, intrahepatic inoculation of tumor cells and intrahepatic tissue implantation. Four parameters--the latency period, take rates, pathological features and metastatic rates--were evaluated side-by-side. 100% take rates were achieved in liver with intrahepatic, intrasplenic inoculation of tumor cells and intrahepatic tissue implantation. In contrast, no tumor in liver was observed with intravenous injection of tumor cells. Intrahepatic tissue implantation resulted in the shortest latency with 0.5 cm (longitudinal diameter) tumors found in liver two weeks after implantation, compared to 0.1cm for intrahepatic inoculation of tumor cells. Approximately 0.1cm tumors were only visible at 4 weeks after intrasplenic inoculation. Uniform, focal and solitary tumors were formed with intrahepatic tissue implantation whereas multinodular, dispersed and non-uniform tumors produced with intrahepatic and intrasplenic inoculation of tumor cells. Notably, metastasis became visible in liver, peritoneum and mesenterium at 3 weeks post-implantation, and lung metastasis was visible after 7 weeks. T cell infiltration was evident in tumors, resembling the situation in HCC patients. Our study demonstrated that orthotopic HCC mouse models established via intrahepatic tissue implantation authentically reflect clinical manifestations in HCC patients pathologically and immunologically, suggesting intrahepatic tissue implantation is a preferable approach for establishing orthotopic HCC mouse models.

Profiles of differential expression of circulating microRNAs in hepatitis B virus-positive small hepatocellular carcinoma.

BACKGROUND: Abnormally expressed circulating microRNA (miRNA) may serve as a potential biomarker for the diagnosis of cancer patients. OBJECTIVE: We sought to determine the differentially expressed circulating microRNAs in patients with hepatitis B virus (HBV)-positive small hepatocellular carcinoma (HCC) compared to other HBV-positive benign liver diseases. METHODS: The miScript miRNA PCR Array was used to detect the levels of 84 miRNAs in plasma or serum samples of patients with HBV-related small HCC (23 cases), liver cirrhosis (LC) (20 cases), chronic hepatitis B (CHB) (20 cases) and healthy controls (16 cases). MiRNAs with fold-change values ⩾ 2 or ⩽ 0.5 compared to healthy controls were considered to be deregulated miRNAs. RESULTS: The results of duplicate plasma experiments were not reliable. Comprehensive analysis of the two serum experiments showed that the quality controls all met the requirements. We found 18 differentially expressed miRNAs. Relative to healthy controls, nine, three, and 11 miRNAs were up-regulated in the CHB group, LC group and small HCC group, respectively. In contrast, one, three, and three miRNAs were down-regulated in the same patient groups, respectively. Interestingly, miR-195, miR-25 and miR-16 were up-regulated, and miR-205 was down-regulated, in all three experimental groups. Moreover, only in the HCC group, miR-18a, miR-100, miR-145 and miR-223 were up-regulated 3.48-, 2.95-, 2.12- and 3.91-fold, respectively, and miR-200a and miR-222 were down-regulated 2.56- and 2.00-fold, respectively. CONCLUSIONS: Our study demonstrated the presence of six differentially expressed serum microRNAs in HBV-positive small HCC compared to other benign liver diseases associated with HBV.