CHB patients with rtA181T-mutated HBV infection are associated with higher risk hepatocellular carcinoma due to increases in mutation rates of tumour suppressor genes.

The HBV rtA181T mutation is associated with an increased risk of hepatocellular carcinoma (HCC) in patients with chronic hepatitis B (CHB). This study aimed to evaluate the mechanism by which rtA181T mutation increases the risk of HCC. We enrolled 470 CHB patients with rtA181T and rtA181V mutation in this study; 68 (22.15%) of the 307 patients with rtA181T mutation and 22 (13.5%) of the 163 patients with rtA181V mutation developed HCC (p < .05). The median follow-up periods were 8.148 and 8.055 years (p > .05). Serum HBV DNA and HBsAg levels in rtA181T-positive patients were similar to that in rtA181V-positive patients. However, the serum HBeAg levels in the rtA181T-positive patients were significantly higher than that in rtA181V-positive patients. In situ hybridization experiments showed that the HBV cccDNA and HBV RNA levels were significantly higher in the liver cancer tissues of patients with the rtA181T mutation compared to that in the tissues of patients with the rtA181V mutation. The percentage of anti-tumour hot-gene site mutations was significantly higher in the rtA181T-positive HCC liver tissue compared to that in the rtA181T-negative HCC liver tissue (7.65% and 4.3%, p < .05). This is the first study to use a large cohort and a follow-up of more than 5 years (average 8 years) to confirm that the rtA181T mutation increased the risk of HCC, and that it could be related to the increase in the mutation rate of hotspots of tumour suppressor genes (CTNNB1, TP53, NRAS and PIK3CA).

LINC01806 Promotes Breast Cancer Growth and Metastasis via Sponging miR-1286 to Disinhibit ZEB1 Expression.

Breast cancer (BC) is the most abundant and aggressive cancer that impacts millions of women with poorly understood mechanisms. Here, we aimed to investigate the function of LINC01806 in BC development. Human BC tissues and nearby normal specimens were taken from diagnosed BC patients. The expression levels of LINC01806, miR-1286, ZEB1, and EMT-related markers were evaluated by qRT-PCR and western blotting. FISH was used to visualize the subcellular localization of LINC01806. The viability, proliferation, migration and invasion capacities of BC cells were assessed by MTT, colony formation, and transwell assays. Interactions among LINC01806, miR-1286 and ZEB1 were validated by dual luciferase assay. The unpaired Student t-test (for two groups) or one-way ANOVA following with Tukey post-hoc test (for more than three groups) was employed for statistical analysis. LINC01806 level was elevated in BC tissues. Knockdown of LINC01806 suppressed EMT process and BC cell proliferation, migration, and invasion. LINC01806 co-localized and directly bound with miR-1286 in the cytoplasm. MiR-1286 inhibitor blocked the effects of LINC01806 knockdown on BC cell EMT, proliferation and migration. MiR-1286 targeted ZEB1 and overexpression of ZEB1 blocked the regulatory functions of miR-1286 mimics in BC. LINC01806 facilitates EMT and accelerates BC cell proliferation, migration, and invasion via acting as miR-1286 sponge to disinhibit ZEB1 expression.

ASPP2 enhances chemotherapeutic sensitivity through the down-regulation of XIAP expression in a p53 independent manner in hepatocellular carcinoma.

Apoptosis stimulated protein of p53-2 (ASPP2) induces the transcription of p53-targeted genes to stimulates its pro-apoptosis function. The poor chemotherapeutic sensitivity is associated with the decreased ASPP2 expression in many human cancers. Here, multiple genes real-time RT-PCR array and western blotting analysis show that ASPP2 suppress the expression of X-linked inhibitor of apoptosis protein (XIAP), determinant of chemoresistance in cancer, in hepatocellular carcinoma (HCC) in a p53-independent manner. Further experiments with ASPP2-rAd and ASPP2-Lv confirmed that ASPP2 enhanced sensitivity of sorafenib to HCC via suppressing XIAP expression. XIAP mainly found on the cytoplasm and perinuclear areas of ASPP2 over-expressed HepG2 cells, while both cytoplasm and nucleus in ASPP2 shut down HepG2 cells. The association of poor sensitivity of sorafenib and XIAP expression was also found both in ASPP2 shut down and overexpress mice, where liver tissue with decreased or increased ASPP2 displayed less or more apoptosis, respectively. Finally, ASPP2 and XIAP expression analyzed in 43 hepatocellular carcinoma tumors and 44 adjacent normal tissues from 38 hepatocellular carcinoma patients for fully understand their expression within HCC patients. Compared with the tumor tissues, ASPP2 mRNA levels were increased, and XIAP levels decreased in the adjacent normal tissues. Taken together, XIAP suppressed ASPP2 increased tumor sensitivity to chemotherapy in a p53-independent manner, which was associated with chemotherapy resistance, suggesting that p53 activation and XIAP suppression were two independent ways that ASPP2 enhance the sensitivity of chemotherapy.

Translocation of intranasal (i.n.) instillation of different-sized cerium dioxide (CeO2) particles: potential adverse effects in mice.

Cerium oxide (CeO2), one of many engineered nanomaterials (ENMs), is composed primarily of metal oxides, such as cerium oxide (CeO2). CeO2-containing materials are widely used as a polishing agent for glass mirrors, plate glass, television tubes, ophthalmic lenses, and precision optics. The widespread use of this nanomaterial (NM) resulted in increased environmental contamination levels and consequent human exposure. However, the influence of Ce on humans remains to be determined. The aim of this study was to expose female ICR mice to varying nanoparticle sizes of 35 nm, 300 nm as well as a mixture of 1-5 µM CeO2 particles through intranasal (i.n.) instillation at 40 mg/kg dose on day 1, 3 and 5, and the experiment terminated on day 7. Histopathology findings demonstrated that hydropic degeneration was prominently associated with hemorrhage in renal cortex and medulla in all CeO2-administered groups. In liver of CeO2-exposed mice, hydropic degeneration was also prominent. Serum chemistries also indicated signs of renal and hepatic lesion as evidenced by significantly decreased serum levels of total bilirubin (TBIL) and total phosphate (TP) and activity of alkaline phosphatase (ALP). ICP-MS analysis group demonstrated that Ce levels were not significantly higher in liver and kidneys of mice exposed to 35 nm CeO2. An increase in Ce content was observed in hepatic and renal tissues of mice exposed to 300 nm or 1-5 µM CeO2. The levels of Ce were similar in these two groups suggesting a threshold level of Ce was attained regardless of NP size. Data thus demonstrated that i.n. instillation of different-sized CeO2 particles translocated to liver and kidney and that size difference of CeO2 particles did not exert significant in the observed histopathology responses.

Ceria Oxide Nanoparticles an Ideal Carrier Given Little Stress to Cells and Rats.

Nanoparticles were used as ideal carrier for its passive and active targeting property. Unfortunately, many of them were failed for its biotoxicology. Thus, find a safe and targeted drug delivery was the new goal of pharmaceutical industries. Here, A549 and H1299 cells were exposed to ceria oxide nanoparticles, silicon oxide nanoparticles and zinc oxide nanoparticles for 12 h to induce autophagy and late apoptosis. Rats were exposed to ceria oxide nanoparticles (20 mg/kg · bw) for 1, 7, 14 or 28 days to induce lung injury and cytokines change. Luckily, compare with silicon oxide nanoparticles and zinc oxide nanoparticles, autophagy and late apoptosis were failed to fund in ceria oxide nanoparticles groups in 100 µg/ml in cell lines for 12 h. At the same time, the autophagy related genes LC3, atg5, beclin1 and bcl2 were not change in protein level at 0 to 200 µg/ml. What's more, histopathology change of the lung was recovered at the day of 28, only four of twenty-seven cytokines (IL12P70, RANTES, IL-X and MIP-1α) were changed at the day of 28 after exposed to ceria oxide nanoparticles (20 mg/kg · bw). Therefore, we indicated that ceria oxide nanoparticles can't give a stress both in vivo and vitro, and ceria oxide nanoparticles will be an ideal carrier for targeted drug delivery.

Intervention effect of gamma aminobutyric acid on anxiety behavior induced by phthalate (2-ethylhexyl ester) in rats.

BACKGROUND: Di(2-ethylhexyl) phthalate (DEHP) is one of the most widely used phthalate esters. The application of DEHP has caused serious environmental pollution and posed a threat to human health. METHODS: A total of 30 male Sprague-Dawley rats were randomly divided into control group, DEHP group (500 mg/kg DEHP), low GABA (Gama-aminobutyric acid) group (500 mg/kg DEHP and 1 mg/kg GABA), medium GABA group (500 mg/kg DEHP and 2 mg/kg GABA) and high GABA group (500 mg/kg DEHP and 4 mg/kg GABA). The interventions continued for 30 consecutive days. Open-field test and elevated plus-maze test were used to detect behavioral changes of rats before and after interventions. RESULTS: The levels of nitric oxide and nitric oxide synthase in prefrontal cortex of rats were measured using enzyme-linked immunosorbent assay. DEHP and GABA treatment had no significant effects on the body weight of rats. GABA restored food utilization rate of rats impaired by DEHP to the level of healthy rats. According to open-field test and elevated plus-maze test, GABA alleviated the effects of DEHP on rat behaviors. Enzyme-linked immunosorbent assay showed that GABA was effective in reducing the levels of nitric oxide and nitric oxide synthase in rats treated with DEHP. CONCLUSION: DEHP exposure induced anxiety in rats, which may be achieved through elevating nitric oxide and nitric oxide synthase levels in prefrontal cortex of rats. However, the effects caused by DEHP could be alleviated by GABA.

Acute changes in murine hippocampus and olfactory bulb after nasal instillation of varying size cerium dioxide particles.

Cerium (Ce)-containing compounds are now widely applied in medicine, agriculture, and animal breeding. However, the effects of Ce on humans, especially on the central nervous system (CNS), remain to be determined. In order to investigate whether Ce exposure affected the CNS, the aim of this study was to expose female ICR mice to varying nanoparticle sizes of 35 nm and 300 nm, and to a mixture of 1-5 µM cerium dioxide (CeO2) particles through intranasal (i.n.) instillation at daily dose of 40 mg/kg body weight. Immunohistochemical data showed that glial fibrillary acidic protein expression (GFAP) increased significantly in the hippocampus and olfactory bulb in all Ce-administered groups. The ultrastructure of olfactory bulb cells displayed chromatin reduction. In the hippocampus decreased chromatin was associated with ribosome shedding as evidenced from transmission electron microscopy (TEM). No significant differences in immunohistochemistry were noted between varying sizes of CeO2 groups. The results of inductively coupled plasma-mass spectroscopy (ICP-MS) analysis group exposed to 1-5 µM demonstrated that Ce levels were significantly higher in whole brain (0.17 ng/mg) than for the control (0.04 ng/mg). Data thus demonstrated that i.n. instillation of different sized CeO2 particles induced damage in the olfactory bulb and hippocampus and that CeO2 particle size did not appear to play a role in the observed adverse responses.

Deciphering the genomic insights into the coexistence of congenital scoliosis and congenital anomalies of the kidney and urinary tract.

Background: Congenital scoliosis and congenital anomalies of the kidney and urinary tract are distinct genetic disorders with differing clinical manifestations. Clinically, their coexistence is not rare, but the etiologies of these complex diseases remain largely unknown, especially their shared genetic basis. Methods: We sequenced the genomes of 40 individuals diagnosed with both CS and CAKUT, alongside 2,764 controls from a Chinese Han population cohort. Our analyses encompassed gene-based and pathway-based weighted rare variant association tests, complemented by copy number variant association analyses, aiming to unravel the shared genomic etiology underlying these congenital conditions. Results: Gene-based analysis identified PTPN11 as a pivotal gene influencing both skeletal and urinary system development (P = 1.95E-21), participating in metabolic pathways, especially the MAPK/ERK pathway known to regulate skeletal and urinary system development. Pathway-based enrichment showed a significant signal in the MAPK/ERK pathway (P = 3E-04), reinforcing the potential role of PTPN11 and MAPK/ERK pathway in both conditions. Additionally, CNV analysis pinpointed IGFLR1 haploinsufficiency as a potential influential factor in the combined CS-CAKUT phenotypic spectrum. Conclusion: This study enriches our understanding of the intricate genomic interplay underlying congenital scoliosis and kidney and urinary tract anomalies, emphasizing the shared genetic foundations between these two disorders.

LAPTM4B-mediated hepatocellular carcinoma stem cell proliferation and MDSC migration: implications for HCC progression and sensitivity to PD-L1 monoclonal antibody therapy.

In hepatocellular carcinoma (HCC), immunotherapy is vital for advanced-stage patients. However, diverse individual responses and tumor heterogeneity have resulted in heterogenous treatment outcomes. Our mechanistic investigations identified LAPTM4B as a crucial gene regulated by ETV1 (a transcription factor), especially in liver cancer stem cells (LCSCs). The influence of LAPTM4B on LCSCs is mediated via the Wnt1/c-Myc/ß-catenin pathway. CXCL8 secretion by LAPTM4B drove myeloid-derived suppressor cell (MDSC) migration, inducing unfavorable patient prognosis. LAPTM4B affected PD-L1 receptor expression in tumor microenvironment and enhanced tumor suppression induced by PD-L1 monoclonal antibodies in HCC patients. LAPTM4B up-regulation is correlated with adverse outcomes in HCC patients, sensitizing them to PD-L1 monoclonal antibody therapy.

Radiofrequency ablation plays double role in immunosuppression and activation of PBMCs in recurrent hepatocellular carcinoma.

Background: Radiofrequency ablation (RFA) is the primary curative treatment for hepatocellular carcinoma (HCC) patients who are not eligible for surgery. However, the effects of RFA on the global tumor immune response remain unclear. Method: In this study, we examined the phenotypic and functional changes in peripheral blood mononuclear cells (PBMCs) from recurrent HCC patients who had undergone two RFA treatments using mass cytometry and high-throughput mRNA assays. Results: We observed significant increase in monocytes and decrease in T cell subpopulations three days after the first RFA treatment and three days after the second RFA treatment. The down-regulation of GZMB, GZMH, GZMK, and CD8A, which are involved in the cytotoxic function of T cells, was observed following RFA. Furthermore, the population of CD8 effector and memory T cells (CD8 Teff and CD8 Tem) significantly decreased after RFA. The expression of CD5 and CD161 in various T cell subpopulations also showed significant reductions. Additionally, elevated secretion of VEGF was observed in monocytes, B cells, regulatory T cells (Tregs), and CD4 naive T cells. Conclusion: In recurrent HCC patients, serum components derived from radiofrequency therapy can enhance the antigen-presenting capacity of monocytes. However, they also inhibit the anti-cancer immune response by reducing the population of CD8 effector and memory T cells and suppressing the activation of T cells, as well as down-regulating the expression of CD161 and CD5 in various T cell subpopulations. These tumor-derived components also contribute to an immunosuppressive microenvironment by promoting the secretion of VEGF in monocytes, Tregs, B cells, and CD4 naive T cells.

Heterogeneity-induced NGF-NGFR communication inefficiency promotes mitotic spindle disorganization in exhausted T cells through PREX1 suppression to impair the anti-tumor immunotherapy with PD-1 mAb in hepatocellular carcinoma.

BACKGROUND: The mechanism of decreased T cells infiltrating tumor tissues in hepatocellular carcinoma is poorly understood. METHODS: Cells were separated from the single-cell RNA-sequence dataset of hepatocellular carcinoma patients (GSE149614) for cell-cell communication. Flow cytometry, EDU staining, H3-Ser28 staining, confocal immunofluorescence staining, western blotting and naked microsubcutaneous tumors were performed for the mechanism of NGF-NGFR promoting proliferation. RESULTS: The present study has revealed that during the process of T-cell infiltration from adjacent tissues to tumor tissues, an inefficiency in NGF-NGFR communication occurs in the tumor tissues. Importantly, NGF secreted by tumor cells interacts with NGFR present on the membranes of the infiltrated T cells, thereby promoting the proliferation through the activation of mitotic spindle signals. Mechanistically, the mediation of mitotic spindle signal activation promoting proliferation is executed by HDAC1-mediated inhibition of unclear trans-localization of PREX1. Furthermore, PD-1 mAb acts synergistically with the NGF-NGFR communication to suppress tumor progression in both mouse models and HCC patients. Additionally, NGF-NGFR communication was positively correlates with the PD-1/PDL-1 expression. However, expressions of NGF and NGFR are low in tumor tissues, which is responsible for the invasive clinicopathological features and the disappointing prognosis in HCC patients. CONCLUSION: Inefficiency in NGF-NGFR communication impairs PD-1 mAb immunotherapy and could thus be utilized as a novel therapeutic target in the treatment of HCC patients in clinical practice.

Unveiling the Mystery of SUMO-activating enzyme subunit 1: A Groundbreaking Biomarker in the Early Detection and Advancement of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. The discovery and research of effective biomarkers have become prevailing trends. SUMO-activating enzyme subunit 1 (sae1), an E1-activating enzyme, is indispensable for protein SUMOylation. In this study, we conducted a comprehensive analysis of database contents and found that sae1 is highly expressed in HCC and is correlated with poor prognosis. We also identified its regulated transcription factor, rad51, and related signaling pathways. We conclude that sae1 is a promising cancer metabolic biomarker with diagnostic and prognostic value in HCC.

The hepatitis B virus promotes the progression of non-alcoholic fatty liver disease through incomplete autophagy.

Hepatitis B virus (HBV) infection is still a serious public health problem. In recent years, with the increasing incidence of chronic hepatitis B (CHB) combined with nonalcoholic fatty liver disease (NAFLD), a more in-depth exploration of the pathogenesis of CHB combined with NAFLD is required. HBV can induce autophagy and use to increase replication. The removal of fat by autophagy, also known as lipophagy, is also currently considered an alternative pathway for lipid metabolism in liver cells. This degradation of autophagy prevents hepatotoxicity and steatosis. However, it is not known whether there is a correlation between HBV-related autophagy and the progression of NAFLD. We explored how HBV affects disease progression in NAFLD should be " and determined whether it is associated with HBV-associated autophagy. In this study, we constructed HBV-TG mouse high-fat diet (HFD) models and controls, and the results showed that the presence of HBV promoted the occurrence of NAFLD. We also demonstrated that HBV promotes lipid droplet accumulation in hepatocytes using HBV-stable expression cell lines HepG2.2.15 and AML12-HBV. In addition, this study also found that exogenous OA supplementation reduced HBV replication. We further studied the mechanism and found that HBV-related autophagy can promote the absorption of liver cells to lipid droplets. It can reduce the decomposition of lipid droplets by inhibiting the function of autophagolysosome, and eventually lead to the accumulation of lipid droplets in hepatocytes. In a word, HBV promotes the progression of NAFLD by increasing lipid accumulation in hepatocytes through incomplete autophagy.

Recent advances in recurrent hepatocellular carcinoma therapy.

Hepatocellular carcinoma (HCC) is the most prevalent form of primary liver cancer, accounting for 75%-85% of cases. Although treatments are given to cure early-stage HCC, up to 50%-70% of individuals may experience a relapse of the illness in the liver after 5 years. Research on the fundamental treatment modalities for recurrent HCC is moving significantly further. The precise selection of individuals for therapy strategies with established survival advantages is crucial to ensuring better outcomes. These strategies aim to minimize substantial morbidity, support good life quality, and enhance survival for patients with recurrent HCC. For individuals with recurring HCC after curative treatment, no approved therapeutic regimen is currently available. A recent study presented novel approaches, like immunotherapy and antiviral medication, to improve the prognosis of patients with recurring HCC with the apparent lack of data to guide the clinical treatment. The data supporting several neoadjuvant and adjuvant therapies for patients with recurring HCC are outlined in this review. We also discuss the potential for future clinical and translational investigations.

Landscape of Immune Cells Heterogeneity in Liver Transplantation by Single-Cell RNA Sequencing Analysis.

Rejection is still a critical barrier to the long-term survival of graft after liver transplantation, requiring clinicians to unveil the underlying mechanism of liver transplant rejection. The cellular diversity and the interplay between immune cells in the liver graft microenvironment remain unclear. Herein, we performed single-cell RNA sequencing analysis to delineate the landscape of immune cells heterogeneity in liver transplantation. T cells, NK cells, B cells, and myeloid cell subsets in human liver and blood were enriched to characterize their tissue distribution, gene expression, and functional modules. The proportion of CCR6+CD4+ T cells increased within an allograft, suggesting that there are more memory CD4+ T cells after transplantation, in parallel with exhausted CTLA4+CD8+ T and actively proliferating MKI67+CD8+ T cells increased significantly, where they manifested heterogeneity, distinct function, and homeostatic proliferation. Remarkably, the changes of CD1c+ DC, CADM+ DC, MDSC, and FOLR3+ Kupffer cells increase significantly, but the proportion of CD163+ Kupffer, APOE+ Kupffer, and GZMA+ Kupffer decreased. Furthermore, we identified LDLR as a novel marker of activated MDSC to prevent liver transplant rejection. Intriguingly, a subset of CD4+CD8+FOXP3+ T cells included in CTLA4+CD8+ T cells was first detected in human liver transplantation. Furthermore, intercellular communication and gene regulatory analysis implicated the LDLR+ MDSC and CTLA4+CD8+ T cells interact through TIGIT-NECTIN2 signaling pathway. Taken together, these findings have gained novel mechanistic insights for understanding the immune landscape in liver transplantation, and it outlines the characteristics of immune cells and provides potential therapeutic targets in liver transplant rejection.

Heterogeneity induced GZMA-F2R communication inefficient impairs antitumor immunotherapy of PD-1 mAb through JAK2/STAT1 signal suppression in hepatocellular carcinoma.

Tumor heterogeneity has been associated with immunotherapy and targeted drug resistance in hepatocellular carcinoma (HCC). However, communications between tumor and cytotoxic cells are poorly understood to date. In the present study, thirty-one clusters of cells were discovered in the tumor tissues and adjacent tissues through single-cell sequencing. Moreover, the quantity and function exhaustion of cytotoxic cells was observed to be induced in tumors by the TCR and apoptosis signal pathways. Furthermore, granzyme failure of cytotoxic cells was observed in HCC patients. Importantly, the GZMA secreted by cytotoxic cells was demonstrated to interact with the F2R expressed by the tumor cells both in vivo and in vitro. This interaction induced tumor suppression and T cell-mediated killing of tumor cells via the activation of the JAK2/STAT1 signaling pathway. Mechanistically, the activation of JAK2/STAT1 signaling promoted apoptosis under the mediating effect of the LDPRSFLL motif at the N-terminus of F2R, which interacted with GZMA. In addition, GZMA and F2R were positively correlated with PD-1 and PD-L1 in tumor tissues, while the expressions of F2R and GZMA promoted PD-1 mAb-induced tumor suppression in both mouse model and HCC patients. Finally, in HCC patients, a low expression of GZMA and F2R in the tumor tissues was correlated with aggressive clinicopathological characteristics and poor prognosis. Collectively, GZMA-F2R communication inefficient induces deficient PD-1 mAb therapy and provide a completely novel immunotherapy strategy for tumor suppression in HCC patients.

RDIVpSGP motif of ASPP2 binds to 14-3-3 and enhances ASPP2/k18/14-3-3 ternary complex formulation to promote BRAF/MEK/ERK signal inhibited cell proliferation in hepatocellular carcinoma.

The Apoptosis Stimulating Protein of p53 2 (ASPP2) is a heterozygous insufficient tumor suppressor; however, its molecular mechanism(s) in tumor suppression is not completely understood. ASPP2 plays an essential role in cell growth, as shown by liver hepatocellular carcinoma (LIHC) RNA-seq assay using the Cancer Genome Atlas (TCGA) and High-Throughput-PCR assay using ASPP2 knockdown cells. These observations were further confirmed by in vivo and in vitro experiments. Mechanistically, N-terminus ASPP2 interacted with Keratin 18 (k18) in vivo and in vitro. Interestingly, the RDIVpSGP motif of ASPP2 associates with 14-3-3 and promotes ASPP2/k18/14-3-3 ternary-complex formation which promotes MEK/ERK signal activation by impairing 14-3-3 and BRAF association. Additionally, ASPP2-rAd injection promotes paclitaxel-suppressed tumor growth by suppressing cell proliferation in the BALB/c nude mice model. ASPP2 and k18 were preferentially downregulated in Hepatocellular Carcinoma (HCC), which predicted poor prognosis in HCC patients. Overall, these findings suggested that ASPP2 promoted BRAF/MEK/ERK signal activation by promoting the formation of an ASPP2/k18/14-3-3 ternary complex via the RDIVpSGP motif at the N terminus. Moreover, this study provides novel insights into the molecular mechanism of tumor suppression in HCC patients.

Enriched high­throughput reverse transcription­quantitative PCR template preparation without pre­amplification.

A cDNA template with a high concentration is required to generate a high number of copies for accurate downstream high­throughput reverse transcription­quantitative PCR screening. However, with the traditional method, pre­amplification is not widely available. In the present study, a novel strategy to resolve the pre­amplification limitation has been developed. Total RNA was extracted using a commercially available RNeasy Micro kit then, the cDNA was synthesized using SuperScript® III First­Strand Synthesis system. PCR inhibitors (proteins and soluble salt ions) in the enriched cDNA were removed using saturated phenol­chloroform extraction. Finally, genes were evaluated using PCR amplification and the BioMark™ HD system. The positive detection rate of individual target gene expression reached 70.18%; however, it markedly decreased to 35.42% using PCR amplification without prior dilution. Next, the reverse transcription product was purified using saturated phenol­chloroform extraction, and the positive detection rate increased to 97.04%. Notably, the positive detection rate of cDNA prepared using this method of high­throughput and traditional PCR (97.04 vs. 96.6%) was not significantly different. In conclusion, the results demonstrate the novel method was an easy and reproducible method for performing robust and highly accurate targeted amplification.

Association between donor/recipient MTRR gene polymorphisms and the risk of new-onset neurological complications after liver transplantation.

Neurological complications are very common after liver transplantation. This study focuses on clinical risk factors and susceptibility gene polymorphisms of neurological complications after liver transplantation. A better predictive model is obtained. This study proves that MTRR is an independent susceptibility gene for neurological complications. Compared with the independent risk factor of abdominal infection, MTRR has a more advantageous value in predicting neurological complications after liver transplantation.

Progress and prospects of biomarkers in primary liver cancer (Review).

Tumor biomarkers are important in the early screening, diagnosis, therapeutic evaluation, recurrence and prognosis prediction of tumors. Primary liver cancer is one of the most common malignant tumors; it has high incidence and mortality rates and seriously endangers human health. The main pathological types of primary liver cancer include hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (ICC) and combined HCC­cholangiocarcinoma (cHCC­CC). In the present review, a systematic outline of the current biomarkers of primary liver cancer is presented, from conventional blood biomarkers, histochemical biomarkers and potential biomarkers to resistance­associated biomarkers. The important relationships are deeply elucidated between biomarkers and diagnosis, prognosis, clinicopathological features and resistance, as well as their clinical significance, in patients with the three main types of primary liver cancer. Moreover, a summary of several important biomarker signaling pathways is provided, which is helpful for studying the biological mechanism of liver cancer. The purpose of this review is to provide help for clinical or medical researchers in the early diagnosis, differential diagnosis, prognosis and treatment of HCC.