Pathologically successful conversion hepatectomy for advanced giant hepatocellular carcinoma after multidisciplinary therapy: A case report and review of literature.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the leading causes of death due to its complexity, heterogeneity, rapid metastasis and easy recurrence after surgical resection. We demonstrated that combination therapy with transcatheter arterial chemoembolization (TACE), hepatic arterial infusion chemotherapy (HAIC), Epclusa, Lenvatinib and Sintilimab is useful for patients with advanced HCC. CASE SUMMARY: A 69-year-old man who was infected with hepatitis C virus (HCV) 30 years previously was admitted to the hospital with abdominal pain. Enhanced computed tomography (CT) revealed a low-density mass in the right lobe of the liver, with a volume of 12.9 cm × 9.4 cm × 15 cm, and the mass exhibited a "fast-in/fast-out" pattern, with extensive filling defect areas in the right branch of the portal vein and an alpha-fetoprotein level as high as 657 ng/mL. Therefore, he was judged to have advanced HCC. During treatment, the patient received three months of Epclusa, three TACE treatments, two HAIC treatments, three courses of sintilimab, and twenty-one months of lenvatinib. In the third month of treatment, the patient developed severe side effects and had to stop immunotherapy, and the Lenvatinib dose had to be halved. Postoperative pathological diagnosis indicated a complete response. The patient recovered well after the operation, and no tumor recurrence was found. CONCLUSION: Multidisciplinary conversion therapy for advanced enormous HCC caused by HCV infection has a significant effect. Individualized drug adjustments should be made during any treatment according to the patient's tolerance to treatment.

Protein kinases: The key contributors in pathogenesis and treatment of nonalcoholic fatty liver disease-derived hepatocellular carcinoma.

Protein kinases (PKs), one of the largest protein families, can be further divided into different groups based on their substrate or structure and function. PKs are important signaling messengers in numerous life activities, including cell metabolism, proliferation, division, differentiation, senescence, death, and disease. Among PK-related diseases, nonalcoholic fatty liver disease (NAFLD) has been recognized as a major contributor to hepatocellular carcinoma (HCC) and liver transplantation. Unfortunately, NAFLD-derived HCC (NAFLD-HCC) has poor prognosis because it is typically accompanied by older age, multiple metabolic syndromes, obstacles in early-stage diagnosis, and limited licensed drugs for treatment. Accumulating evidence suggests that PKs are implicated in the pathogenic process of NAFLD-HCC, via aberrant metabolism, hypoxia, autophagy, hypoxia, gut microbiota dysbiosis, and/or immune cell rearrangement. The present review aims to summarize the latest research advances and emphasize the feasibility and effectiveness of therapeutic strategies that regulate the expression and activities of PKs. This might yield clinically significant effects and lead to the design of novel PK-targeting therapies. Furthermore, we discuss emerging PK-based strategies for the treatment of other malignant diseases similar to NAFLD-HCC.

Research progress of proanthocyanidins and anthocyanidins.

Proanthocyanidins (PA) are polyphenol compounds that are widely distributed in the bark, fruit core, skin, or seeds of various plants. Anthocyanidins are water-soluble natural pigments widely found in plants. They are all flavonoids, a major coloring substance in plants and fruits. In recent years, research into PA and anthocyanins has become increasingly popular because of their excellent anti-oxidation, scavenging of reactive oxygen free radicals and other physical and chemical activities, and their anti-cancer, vision protection, aging prevention, skin beauty pharmacological, and nutraceutical effects. Especially, recent systematic reviews and meta-analyses indicate their value, safety, and efficacy in the prevention, adjuvant therapy, and management of cardiometabolic disease. Here, we summarize their research progress from the aspects of chemical structure, biosynthetic pathways, distribution, extraction and separation, coloration, efficacy, and potential. The comparison between them might provide a reference for their development and efficient utilization. However, more large-sample-size randomized controlled trials and high-quality studies are needed to firmly establish their clinical efficacy.

Sequential extraction of RNA, DNA and protein from cultured cells of the same group.

BACKGROUND: Efficient extraction of nucleic acids and proteins (ENAP) from cells is a prerequisite for precise annotation of gene function, and has become laboratory routine for revealing the mysteries of life. However, cell samples are often from different culture dishes, resulting in inevitable experimental errors and sometimes poor repeatability. AIM: To explore a method to improve the efficiency of ENAP, minimizing errors in ENAP processes, enhancing the reliability and repeatability of subsequent experimental results. METHODS: A protocol for the sequential isolation of RNA, DNA, and proteins from the same cultured HepG2 cells using RNAzol reagent is presented here. The first step involves culturing HepG2 cells to the exponential phase, followed by the sequential isolation of RNA, DNA, and proteins from the same cultured cells in the second step. The yield of nucleic acids and proteins is detected in the third step, and their purity and integrity are verified in the last step. RESULTS: The procedure takes as few as 3-4 d from the start to quality verification and is highly efficient. In contrast to the existing kits and reagents, which are primarily based on independent isolation, this RNAzol reagent-based method is characterized by the sequential isolation of RNA, DNA, and proteins from the same cells, and therefore saves time, and has low cost and high efficiency. CONCLUSION: The RNA, DNA, and proteins isolated using this method can be used for reverse transcription-polymerase chain reaction, polymerase chain reaction, and western blotting, respectively.

Proanthocyanidins: Components, Pharmacokinetics and Biomedical Properties.

Proanthocyanidins (PAs) are a group of polyphenols enriched in plant and human food. In recent decades, epidemiological studies have upheld the direct relationship between PA consumption and health benefits; therefore, studies on PAs have become a research hotspot. Although the oral bioavailability of PAs is quite low, pharmacokinetics data revealed that some small molecules and colonic microbial metabolites of PAs could be absorbed and exert their health beneficial effects. The pharmacological effects of PAs mainly include anti-oxidant, anticancer, anti-inflammation, antimicrobial, cardiovascular protection, neuroprotection, and metabolism-regulation behaviors. Moreover, current toxicological studies show that PAs have no observable toxicity to humans. This review summarizes the resources, extraction, structures, pharmacokinetics, pharmacology, and toxicology of PAs and discusses the limitations of current studies. Areas for further research are also proposed.

SNX17 (Sorting Nexin 17) Mediates Atrial Fibrillation Onset Through Endocytic Trafficking of the Kv1.5 (Potassium Voltage-Gated Channel Subfamily A Member 5) Channel.

BACKGROUND: Kv1.5 (Potassium voltage-gated channel subfamily A member 5) has been regarded as a promising target of interventions for atrial fibrillation (AF). SNX17 (sorting nexin 17), a member of the SNXs (sorting nexin family), regulates the intracellular trafficking of membrane proteins through its FERM (four-point-one, ezrin, radixin, moesin) domain. However, whether SNX17 regulates the trafficking process of Kv1.5 remains unknown. METHODS: A SNX17 knockout rat line was generated to test the role of SNX17 in atrial electrophysiology. The protein expression of SNX17 and membrane ion channels was detected by Western blotting. Electrophysiology changes in the atrial tissue and myocytes were analyzed by optical mapping and patch clamp, respectively. Acetylcholine and electrical stimulation were used to induce AF, and ECG recording was adopted to assess the influence of SNX17 deficiency on AF susceptibility. The spatial relationship between Kv1.5 and SNX17 was evaluated by immunostaining and confocal scanning, and the functional region of SNX17 regulating Kv1.5 trafficking was identified using plasmids with truncated SNX17 domains. RESULTS: Embryonic death occurred in homozygous SNX17 knockout rats. SNX17 heterozygous rats survived, and the level of the SNX17 protein in the atrium was decreased by ≈50%. SNX17 deficiency increased the membrane expression of Kv1.5 and atria-specific ultrarapid delayed rectifier outward potassium current ( IKur) density, resulting in a shortened action potential duration, and eventually contributing to AF susceptibility. Mechanistically, SNX17 facilitated the endocytic sorting of Kv1.5 from the plasma membrane to early endosomes via the FERM domain. CONCLUSIONS: SNX17 mediates susceptibility to AF by regulating endocytic sorting of the Kv1.5 channel through the FERM domain. SNX17 could be a potential target for the development of new drugs for AF.

SRPIN340 protects heart muscle from oxidative damage via SRPK1/2 inhibition-mediated AKT activation.

SRPIN340, a selective serine-arginine protein kinase 1/2 (SRPK1/2) inhibitor, has been shown to have antiviral and anti-angiogenesis effects. However, its role in the heart is unknown. The present study explored the role of SRPIN340 in myocardial protection and the related mechanisms. During challenge with H2O2, cardiomyocytes (CMs) pretreated with SRPIN340 showed strikingly more injury tolerance, which was manifested as reduced lactate dehydrogenase (LDH) release and lower apoptotic index. Further research showed that SRPIN340 activated AKT under basal conditions, and AKT inhibition abolished the protective effects of SRPIN340 treatment during H2O2 stress. The protective effect of SRPIN340 was also demonstrated in perfused rat hearts subjected to ischemia/reperfusion (I/R). Collectively, our results reveal the beneficial effects of SRPIN340 against H2O2-induced oxidative damage in CMs and I/R-induced injury in a Langendorff heart model, supporting a potential application of SRPIN340 in the clinically relevant context of reperfusion. The effectiveness of SRPIN340 may be attributed to AKT signal activation.

Nucleoporin 107 facilitates the nuclear export of Scn5a mRNA to regulate cardiac bioelectricity.

Nucleoporins (Nups) are known to be functional in nucleo-cytoplasmic transport, but the roles of nucleoporins in nonproliferating cells, such as cardiac myocytes, are still poorly understood. In this study, we report that Nup107 regulates cardiac bioelectricity by controlling the nucleo-cytoplasmic trafficking of Scn5a mRNA. Overexpression of Nup107 induced the protein expression of Scn5a rather than that of other ion channels, with no effects of their mRNA levels. The analysis for the protein production demonstrated Nup107-facilitated transport of Scn5a mRNA. Using RIP-PCR and luciferase assay, we found that the 5'-UTR of Scn5a mRNA was not involved in the interaction, whereas the spatial interaction between Nup107 protein and Scn5a mRNA was formed when Scn5a mRNA passing through the nuclear pore. Functionally, Nup107 overexpression in neonatal rat ventricle myocytes significantly increased the currents of Scn5a-encoded INa channel. Moreover, the close correlation between Nup107 and Nav1.5 protein expression was observed in cardiomycytes and heart tissues subjected to hypoxia and ischaemic insults, suggesting a fast regulation of Nup107 on Nav1.5 channel in cardiac myocytes in a posttranscriptional manner. These findings may provide insights into the emergent control of cardiac electrophysiology through Nup-mediated modulation of ion channels.

Effects of anthocyanins on the prevention and treatment of cancer.

Anthocyanins are a class of water-soluble flavonoids, which show a range of pharmacological effects, such as prevention of cardiovascular disease, obesity control and antitumour activity. Their potential antitumour effects are reported to be based on a wide variety of biological activities including antioxidant; anti-inflammation; anti-mutagenesis; induction of differentiation; inhibiting proliferation by modulating signal transduction pathways, inducing cell cycle arrest and stimulating apoptosis or autophagy of cancer cells; anti-invasion; anti-metastasis; reversing drug resistance of cancer cells and increasing their sensitivity to chemotherapy. In this review, the latest progress on the anticancer activities of anthocyanins and the underlying molecular mechanisms is summarized using data from basic research in vitro and in vivo, from clinical trials and taking into account theory and practice. LINKED ARTICLES: This article is part of a themed section on Principles of Pharmacological Research of Nutraceuticals. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.11/issuetoc.

Pinus massoniana Bark Extract: Structure-Activity Relationship and Biomedical Potentials.

Proanthocyanidins (PAs) belong to the condensed tannin subfamily of natural flavonoids. Recent studies have shown that the main bioactive compounds of Pinus massoniana bark extract (PMBE) are PAs, especially the proanthocyanidins B series, which play important roles in cell cycle arrest, apoptosis induction and migration inhibition of cancer cells in vivo and in vitro. PA-Bs are mixtures of oligomers and polymers composed of flavan-3-ol, and the relationship between their structure and corresponding biomedical potentials is summarized in this paper. The hydroxyl at certain positions or the linkage between different carbon atoms of different rings determines or affects their anti-oxidant and free radical scavenging bioactivities. The degree of polymerization and the water solubility of the reaction system also influence their biomedical potential. Taken together, PMBE has a promising future in clinical drug development as a candidate anticancer drug and as a food additive to prevent tumorigenesis. We hope this review will encourage interested researchers to conduct further preclinical and clinical studies to evaluate the anticancer activities of PMBE, its active constituents and their derivatives.

Nucleoporin 35 regulates cardiomyocyte pH homeostasis by controlling Na+-H+ exchanger-1 expression.

The mammalian nuclear pore complex is comprised of ∼ 30 different nucleoporins (Nups). It governs the nuclear import of gene expression modulators and the export of mRNAs. In cardiomyocytes, Na(+)-H(+) exchanger-1 (NHE1) is an integral membrane protein that exclusively regulates intracellular pH (pHi) by exchanging one intracellular H(+) for one extracellular Na(+). However, the role of Nups in cardiac NHE1 expression remains unknown. We herein report that Nup35 regulates cardiomyocyte NHE1 expression by controlling the nucleo-cytoplasmic trafficking of nhe1 mRNA. The N-terminal domain of Nup35 determines nhe1 mRNA nuclear export by targeting the 5'-UTR (-412 to -213 nt) of nhe1 mRNA. Nup35 ablation weakens the resistance of cardiomyocytes to an acid challenge by depressing NHE1 expression. Moreover, we identify that Nup35 and NHE1 are simultaneously downregulated in ischemic cardiomyocytes both in vivo and in vitro. Enforced expression of Nup35 effectively counteracts the anoxia-induced intracellular acidification. We conclude that Nup35 selectively regulates cardiomyocyte pHi homeostasis by posttranscriptionally controlling NHE1 expression. This finding reveals a novel regulatory mechanism of cardiomyocyte pHi, and may provide insight into the therapeutic strategy for ischemic cardiac diseases.

Cardiac Nav 1.5 is modulated by ubiquitin protein ligase E3 component n-recognin UBR3 and 6.

The voltage-gated Na(+) channel Nav 1.5 is essential for action potential (AP) formation and electrophysiological homoeostasis in the heart. The ubiquitin-proteasome system (UPS) is a major degradative system for intracellular proteins including ion channels. The ubiquitin protein ligase E3 component N-recognin (UBR) family is a part of the UPS; however, their roles in regulating cardiac Nav 1.5 channels remain elusive. Here, we found that all of the UBR members were expressed in cardiomyocytes. Individual knockdown of UBR3 or UBR6, but not of other UBR members, significantly increased Nav 1.5 protein levels in neonatal rat ventricular myocytes, and this effect was verified in HEK293T cells expressing Nav 1.5 channels. The UBR3/6-dependent regulation of Nav 1.5 channels was not transcriptionally mediated, and pharmacological inhibition of protein biosynthesis failed to counteract the increase in Nav 1.5 protein caused by UBR3/6 reduction, suggesting a degradative modulation of UBR3/6 on Nav 1.5. Furthermore, the effects of UBR3/6 knockdown on Nav 1.5 proteins were abolished under the inhibition of proteasome activity, and UBR3/6 knockdown reduced Nav 1.5 ubiquitylation. The double UBR3-UBR6 knockdown resulted in comparable increases in Nav 1.5 proteins to that observed for single knockdown of either UBR3 or UBR6. Electrophysiological recordings showed that UBR3/6 reduction-mediated increase in Nav 1.5 protein enhanced the opening of Nav 1.5 channels and thereby the amplitude of the AP. Thus, our findings indicate that UBR3/6 regulate cardiomyocyte Nav 1.5 channel protein levels via the ubiquitin-proteasome pathway. It is likely that UBR3/6 have the potential to be a therapeutic target for cardiac arrhythmias.

Low expression of lncRNA-GAS5 is implicated in human primary varicose great saphenous veins.

The cellular mechanisms of primary varicose great saphenous veins (GSVs) involve inflammation, apoptosis, and proliferation of local cells and extracellular matrix degradation. Long non-coding RNAs (lncRNAs) play important roles in these cellular processes; however, which and how lncRNAs related to these mechanisms take effect on GSVs remain unclear. By screening lncRNAs that might experience changes in GSV varicosities, we selected the lower expressed lncRNA-GAS5 (growth arrest specific transcript 5) for functional assessments. Silencing of lncRNA-GAS5 promoted cell proliferation and migration, and cell cycle of the human saphenous vein smooth muscle cells (HSVSMCs), whereas overexpressing it inhibited these cellular behaviors and reduced apoptosis of HSVSMCs. RNA pull-down experiment revealed a direct bind of lncRNA-GAS5 to a Ca2+-dependent RNA-binding protein, Annexin A2. Further experiments showed that silencing of Annexin A2 reduced the HSVSMCs proliferation and vice versa. In the context of lncRNA-GAS5 knockdown, silencing of Annexin A2 reduced the proliferation of HSVSMCs while overexpression of Annexin A2 increased the proliferation. Thus, the low expression of lncRNA-GAS5 may facilitate HSVSMCs proliferation and migration through Annexin A2 and thereby the pathogenesis of GSV varicosities.

Pine bark extracts: nutraceutical, pharmacological, and toxicological evaluation.

Proanthocyanidins are among the most abundant constituents in pine bark extracts (PBEs). This review summarizes medical research on PBEs from Pinus pinaster, Pinus radiata, Pinus massoniana, and other less well characterized species. The precise mechanisms of the important physiologic functions of PBE components remain to be elucidated, but there is evidently great potential for the identification and development of novel antioxidant, anti-inflammatory, cardiovascular, neuroprotective, and anticancer medicines. Although toxicological data for PBEs are limited, no serious adverse effects have been reported. PBEs, therefore, may have potential as nutraceuticals and pharmaceuticals and should be safe for use as food ingredients.

SNX13 reduction mediates heart failure through degradative sorting of apoptosis repressor with caspase recruitment domain.

Heart failure (HF) is associated with complicated molecular remodelling within cardiomyocytes; however, the mechanisms underlying this process remain unclear. Here we show that sorting nexin-13 (SNX13), a member of both the sorting nexin and the regulator of G protein signalling (RGS) protein families, is a potent mediator of HF. Decreased levels of SNX13 are observed in failing hearts of humans and of experimental animals. SNX13-deficient zebrafish recapitulate HF with striking cardiomyocyte apoptosis. Mechanistically, a reduction in SNX13 expression facilitates the degradative sorting of apoptosis repressor with caspase recruitment domain (ARC), which is a multifunctional inhibitor of apoptosis. Consequently, the apoptotic pathway is activated, resulting in the loss of cardiac cells and the dampening of cardiac function. The N-terminal PXA structure of SNX13 is responsible for mediating the endosomal trafficking of ARC. Thus, this study reveals that SNX13 profoundly affects cardiac performance through the SNX13-PXA-ARC-caspase signalling pathway.

Effects of Pinus massoniana bark extract on the adhesion and migration capabilities of HeLa cells.

Pinus massoniana Lamb is a Chinese red pine species used in traditional medicine for the treatment of a variety of human health disorders. Recent studies have shown that P. massoniana bark extract (PMBE) has an anti-proliferation effect on cancer cells. However, it is not clear if PMBE affects cancer cell migration and/or invasion. We tested the effect of PMBE, which has B-type procyanidin as its main constituent, on the adhesion and migration capabilities of HeLa cells, a human cervical cancer cell line, cultured in vitro. Our results showed that PMBE has no significant effect on the adhesion capability of HeLa cells, but strongly inhibits their migration. This finding suggests that PMBE could be a potential therapeutic agent for metastatic cancer.

Expression of NOD2 in a rat model of acute pancreatitis.

OBJECTIVES: To observe the expression of nucleotide-binding oligomerization domain 2 (NOD2) in rats with acute pancreatitis (AP). METHODS: Sprague-Dawley rats were randomly divided into sham operation (SO) groups and AP groups. Acute pancreatitis was induced with retrograde infusion of sodium taurocholate into the biliopancreatic duct. They were then killed at 3, 6, 12, 24, and 48 hours after induction of AP. Blood biochemical indicators were detected with automatic biochemistry analyzer. Nuclear factor-κB (NF-κB) was measured by immunohistochemistry. The NOD2 was detected by real-time quantitative polymerase chain reaction and Western blot. Tumor necrosis factor-α (TNF-α) was determined by enzyme-linked immunosorbent assay. RESULTS: Compared with the SO group, the level of messenger RNA and protein of NOD2 in pancreatic tissue and peritoneal white blood cells (PWBCs) in the AP groups significantly declined (P < 0.05). The messenger RNA level of NOD2 in the AP groups was correlated positively with amylase (P < 0.05) and negatively with TNF-α (P < 0.05); TNF-α significantly decreased in the AP groups, whereas NF-κB significantly increased (P < 0.05). CONCLUSIONS: The NOD2 may play an important role in the up-regulation and activation of NF-κB during inflammation reactions in AP.

Strong and selective binding of amiloride to thymine base opposite AP sites in DNA duplexes: simultaneous binding to DNA phosphate backbone.

Amiloride (N-amidino-3,5-diamino-6-chloro-pyrazinecarboxamide hydrochloride) has two sets of hydrogen-bond forming sites suitable for target nucleotides and the phosphodiester DNA backbone by which a thymine base opposite an abasic site in DNA duplexes can be recognized with high selectivity and affinity, and it is applicable to the fluorescence detection of thymidine-related SNPs (single-nucleotide polymorphisms) of PCR amplification products.

Use of vitamin B2 for fluorescence detection of thymidine-related single-nucleotide polymorphisms.

In combination with abasic site (AP site)-containing DNAs, potential use of a biotic fluorescence compound, Vitamin B2 (riboflavin), is demonstrated for the fluorescence detection of the thymine (T)-related single-nucleotide polymorphisms. Our method is based on construction of the AP site in DNA duplexes, which allows small ligands to bind to target nucleotides accompanied by fluorescence signaling: an AP site-containing probe DNA is hybridized with a target DNA so as to place the AP site toward a target nucleobase, by which hydrophobic microenvironments are provided for ligands to recognize target nucleotides through stacking and hydrogen-bonding interactions. In 10 mM sodium cacodylate buffer solutions (pH 7.0) containing 100 mM NaCl and 1.0 mM EDTA, Vitamin B2 is found to selectively bind to T (K11=1.8x10(6) M(-1) at 5 degrees C) over other nucleobases, and this is accompanied by significant quenching of its fluorescence. While the sensing functions depend on the flanking sequences to the AP site, Vitamin B2 is applicable to the detection of T/C (cytosine), T/G (guanine) and T/A (adenine) mutation sequences of the CYP2A6 gene, where the flanking nucleobases are guanines in both positions (-GXG-, X=AP site).

Effects of Pinus massoniana bark extract on cell proliferation and apoptosis of human hepatoma BEL-7402 cells.

AIM: To study the effects of Pinus massoniana bark extract (PMBE) on cell proliferation and apoptosis of human hepatoma BEL-7402 cells and to elucidate its molecular mechanism. METHODS: BEL-7402 cells were incubated with various concentrations (20-200 microg/mL) of PMBE for different periods of time. After 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was evaluated by morphological observation, agarose gel electrophoresis, and flow cytometry analysis. Possible molecular mechanisms were primarily explored through immunohistochemical staining. RESULTS: PMBE (20-200 microg/mL) significantly suppressed BEL-7402 cell proliferation in a time- and dose-dependent manner. After treatment of BEL-7402 cells with 160 microg/mL PMBE for 24, 48, or 72 h, a typical apoptotic "DNA ladder" was observed using agarose gel electrophoresis. Nuclear condensation and boundary aggregation or split, apoptotic bodies were seen by fluorescence and electron microscopy. Sub-G1 curves were displayed by flow cytometry analysis. PMBE decreased the expression levels of Bcl-2 protein in a time-dependent manner after treatment of cells with 160 microg/mL PMBE. CONCLUSION: PMBE suppresses proliferation of BEL-7402 cells in a time- and dose-dependent manner and induces cell apoptosis by possibly downregulating the expression of the bcl-2 gene.