Biological Evaluation of Lysionotin: a Novel Inhibitor of 5-Lipoxygenase for Anti-glioma.

OBJECTIVE: To explore the potential mechanism of lysionotin in treating glioma. METHODS: First, target prediction based on Bernoulli Naïve Bayes profiling and pathway enrichment was used to predict the biological activity of lysionotin. The binding between 5-lipoxygenase (5-LO) and lysionotin was detected by surface plasmon resonance (SPR) and molecular docking, and the inhibitory effects of lysionotin on 5-LO and proliferation of glioma were determined using enzyme inhibition assay in vitro and cell viability analysis, respectively. Furthermore, the pharmaceutical effect of lysionotin was explored by cell survival rate analysis and liquid chromatography with tandem mass spectrometry (LC-MS/MS). The protein expression, intracellular calcium ion concentration and cytoskeleton detection were revealed by Western blot, flow cytometry and fluorescence labeling, respectively. RESULTS: Target prediction and pathway enrichment revealed that lysionotin inhibited 5-LO, a key enzyme involved in the arachidonic acid metabolism pathway, to inhibit the proliferation of glioma. Molecular docking results demonstrated that 5-LO can be binding to lysionotin through hydrogen bonds, forming bonds with His600, Gln557, Asn554, and His372. SPR analysis further confirmed the interaction between 5-LO and lysionotin. Furthermore, enzyme inhibition assay in vitro and cell survival rate analysis revealed that 50% inhibition concentration of lysionotin and the median effective concentration of lysionotin were 90 and 16.58 µmol/L, respectively, and the results of LC-MS/MS showed that lysionotin inhibited the production of 5S-hydroperoxy-eicosatetraenoic acid (P<0.05), and moreover, the LC-MS/MS results indicated that lysionotin can enter glioma cells well (P<0.01) and inhibit their proliferation. Western blot analysis demonstrated that lysionotin can inhibit the expression of 5-LO (P<0.05) and downstream leukotriene B4 receptor (P<0.01). In addition, the results showed that lysionotin affected intracellular calcium ion concentration by inhibiting 5-LO to affect the cytoskeleton, as determined by flow cytometry and fluorescence labeling. CONCLUSION: Lysionotin binds to 5-LO could suppress glioma by inhibiting arachiodonic acid metabolism pathway.

Glyceroglycolipids in marine algae: A review of their pharmacological activity.

Glyceroglycolipids are major metabolites of marine algae and have a wide range of applications in medicine, cosmetics, and chemistry research fields. They are located on the cell surface membranes. Together with glycoproteins and glycosaminoglycans, known as the glycocalyx, they play critical roles in multiple cellular functions and signal transduction and have several biological properties such as anti-oxidant and anti-inflammatory properties, anti-viral activity, and anti-tumor immunity. This article focused on the sources and pharmacological effects of glyceroglycolipids, which are naturally present in various marine algae, including planktonic algae and benthic algae, with the aim to highlight the promising potential of glyceroglycolipids in clinical treatment.

Salvianolic acid B from Salvia miltiorrhiza bunge: A potential antitumor agent.

Salvia miltiorrhiza Bunge (Lamiaceae) is a perennial herb widely found in China since ancient times with a high economic and medicinal value. Salvianolic acid B (Sal-B) is an important natural product derived from Salvia miltiorrhiza and this review summarizes the anticancer activity of Sal-B. Sal-B inhibits tumor growth and metastasis by targeting multiple cell signaling pathways. This review aims to review experimental studies to describe the possible anticancer mechanisms of Sal-B and confirm its potential as a therapeutic drug.

Deep Sea Water Alleviates Tau Phosphorylation and Cognitive Impairment via PI3K/Akt/GSK-3ß Pathway.

Deep sea water (DSW), as a noticeable natural resource, has been demonstrated to contain high levels of beneficial minerals and exert marked anti-diabetes effects. Epidemiological studies show that type 2 diabetes mellitus (T2DM) is closely related to high danger of Alzheimer's disease (AD); moreover, Akt/GSK-3ß signaling is the main underlying pathway that connects these two diseases. Besides, it has been demonstrated that minerals in DSW, such as Mg, Se, and Zn, could effectively treat cognitive deficits associated with AD. Herein, we first observed the protection of DSW against cognitive dysfunction in T2DM rats, then furtherly explored the neuroprotective mechanism in SH-SY5Y cell model. In T2DM rats, DSW obviously elevated the concentrations of elements Mg, V, Cr, Zn, and Se in brain and improved learning and memory dysfunction in behavior assays, including Morris water maze (MWM) and new object recognition (NOR). Western blot (WB) results demonstrated that DSW could stimulate PI3K/Akt/GSK-3ß signaling, arrest Tau hyperphosphorylation at serine (Ser) 396 and threonine (Thr)231, which was confirmed by immunohistochemistry (IHC). In order to further confirm the mechanism, we employed wortmannin to inhibit PI3K in SH-SY5Y cells; results showed that pretreatment with wortmannin almost abolished DSW-induced decreases in phosphorylated Tau. Taken together, these data elucidated that DSW could improve Tau hyperphosphorylation and cognitive impairment, which were closely related with the stimulation of Akt/GSK-3ß signaling, and the neuroprotective effects of DSW should be contributed to the synergistic effects of major and trace elements in it, such as Mg, V, Cr, Zn, and Se. These experimental evidence indicated that DSW may be explored as natural neuroprotective food for the prevention and treatment of AD.

Meta-analysis of open surgical repair versus hybrid arch repair for aortic arch aneurysm.

OBJECTIVE: To conduct a meta-analysis of available comparative studies evaluating hybrid arch repair versus open surgical repair of aortic arch aneurysm. METHODS: A literature search was performed using PubMed, Embase and Web of Science to identify any studies comparing the results of hybrid arch repair with open surgical repair of aortic arch aneurysm. Study quality was assessed with the Newcastle-Ottawa Scale. Statistical heterogeneity was estimated using the chi-square test. A random-effects model was used to illustrate heterogeneity. Publication bias was evaluated by funnel plots. RESULTS: Seven retrospective cohort studies from 2009 to 2016 comprising 727 patients were included. Among these patients, 269 were treated with hybrid arch repair and 458 with open surgical repair. There was no significant difference in operative mortality (OR 0.75; 95% CI 0.41-1.39; P = 0.37), permanent neurological deficit (OR 1.24; 95% CI 0.73-2.13; P = 0.42), late mortality (2 years) (OR 3.41; 95% CI 0.83-14.03; P = 0.09) or renal failure (OR 0.80; 95% CI 0.40-1.61; P = 0.53). Interestingly, the meta-analysis indicated that the hybrid group needed more reinterventions (OR 3.43; 95% CI 1.72-6.84; P = 0.0005). CONCLUSIONS: We found no strong evidence indicating that hybrid arch repair is superior to open surgical repair. Furthermore, the hybrid arch repair resulted in more reinterventions despite the fact that it was a less invasive procedure; it also required fewer days in the hospital. Further studies with large numbers of participants and long-term follow-ups are necessary to confirm the effectiveness of hybrid arch repair.

Toxicity of Graphene Quantum Dots in Zebrafish Embryo.

OBJECTIVE: To evaluate the bio-safety of graphene quantum dots (GQDs), we studied its effects on the embryonic development of zebrafish. METHODS: In vivo, biodistribution and the developmental toxicity of GQDs were investigated in embryonic zebrafish at exposure concentrations ranging from 12.5-200 µg/mL for 4-96 h post-fertilization (hpf). The mortality, hatch rate, malformation, heart rate, GQDs uptake, spontaneous movement, and larval behavior were examined. RESULTS: The fluorescence of GQDs was mainly localized in the intestines and heart. As the exposure concentration increased, the hatch and heart rate decreased, accompanied by an increase in mortality. Exposure to a high level of GQDs (200 µg/mL) resulted in various embryonic malformations including pericardial edema, vitelline cyst, bent spine, and bent tail. The spontaneous movement significantly decreased after exposure to GQDs at concentrations of 50, 100, and 200 µg/mL. The larval behavior testing (visible light test) showed that the total swimming distance and speed decreased dose-dependently. Embryos exposed to 12.5 µg/mL showed hyperactivity while exposure to higher concentrations (25, 50, 100, and 200 µg/mL) caused remarkable hypoactivity in the light-dark test. CONCLUSION: Low concentrations of GQDs were relatively non-toxic. However, GQDs disrupt the progression of embryonic development at concentrations exceeding 50 µg/mL.

Protective effects of shikonin on brain injury induced by carbon ion beam irradiation in mice.

Radiation encephalopathy is the main complication of cranial radiotherapy. It can cause necrosis of brain tissue and cognitive dysfunction. Our previous work had proved that a natural antioxidant shikonin possessed protective effect on cerebral ischemic injury. Here we investigated the effects of shikonin on carbon ion beam induced radiation brain injury in mice. Pretreatment with shikonin significantly increased the SOD and CAT activities and the ratio of GSH/GSSG in mouse brain tissues compared with irradiated group (P<0.01), while obviously reduced the MDA and PCO contents and the ROS levels derived from of the brain mitochondria. The shikonin also noticeably improved the spatial memory deficits caused by carbon ion beam irradiation. All results demonstrated that shikonin could improve the irradiated brain injury which might resulted from its modulation effects on the oxidative stress induced by the 12C6+ ion beam.

[Effect of 5 warm-hot nature Chinese drugs for promoting blood circulation and removing blood stasis on 5-HT, NE, and endocrine hormones of rats of cold coagulation and blood stasis syndrome].

OBJECTIVE: To study the mechanism of warm-hot nature Chinese drugs (WHNCD) for promoting blood circulation and removing blood stasis (PBCRBS) for intervening model rats of cold coagulation and blood stasis syndrome (CCBSS). METHODS: CCBSS rat model was set up in outbred SD rats using ice water immersion method. Totally 300 successfully modeled CCBSS rats were randomly divided into 5 groups according to the principle of balance weight, 60 in each group. Contents of triothyrone (T3), tetraiodothyroine (T4), progesterone (P), 5-hydroxytryptamine (5-HT), and noradrenalin (NE) were paralleledly detected in all groups. Then rats in each group were subdivided into 6 subgroups as the model group, the curcuma group, the Ligsticum Chuanxiong group, the safflower group, the Rhizoma Corydalis group, and the Olibanumg group. Besides, 5 normal control groups were set up for 5 indices, 50 rats in total. We need 70 rats (7 groups) to finish observing 1 index, 350 rats in total for 5 indices. Except those in the model group and the normal control group, rats were administered with corresponding decoction at 20 g crude drugs/kg body weight by gastrogavage, 3 mL each time, once daily for 7 successive days. Equal volume of normal saline was given to rats in the normal control group and the model group. Contents of T3, T4, P, 5-HT, and NE were detected before treatment and 1 week after treatment. RESULTS: Compared with before treatment in the same group, T3 increased in the Ligsticum Chuanxiong group and the Olibanumg group, 5-HT increased in the Ligsticum Chuanxiong group, T4, NE, and P increased in all medicated groups (P < 0.05). Compared with the normal control group, contents of T3, T4, 5-HT, NE, and P in the model group decreased (P < 0.05). Compared with the model group, contents of T3, T4, 5-HT, and NE increased in each medicated group (P < 0.05). There was statistical difference in contents of P between the Ligsticum Chuanxiong group and the Olibanumg group (P < 0.05). CONCLUSIONS: WHNCD for PBCRBS had regulatory roles in serum contents of T3, T4, P, and NE in modeled rats of CCBSS. They could promote the thyroid gland-gonadal axis function, enhance the function of the endocrine system, which might be one of the pharmacodynamic mechanism of WHNCD for PBCRBS in intervening CCBSS.

Caveolin-1 Tyr14 phosphorylation induces interaction with TLR4 in endothelial cells and mediates MyD88-dependent signaling and sepsis-induced lung inflammation.

Activation of TLR4 by the endotoxin LPS is a critical event in the pathogenesis of Gram-negative sepsis. Caveolin-1, the signaling protein associated with caveolae, is implicated in regulating the lung inflammatory response to LPS; however, the mechanism is not understood. In this study, we investigated the role of caveolin-1 in regulating TLR4 signaling in endothelial cells. We observed that LPS interaction with CD14 in endothelial cells induced Src-dependent caveolin-1 phosphorylation at Tyr(14). Using a TLR4-MD2-CD14-transfected HEK-293 cell line and caveolin-1-deficient (cav-1(-/-)) mouse lung microvascular endothelial cells, we demonstrated that caveolin-1 phosphorylation at Tyr(14) following LPS exposure induced caveolin-1 and TLR4 interaction and, thereby, TLR4 activation of MyD88, leading to NF-κB activation and generation of proinflammatory cytokines. Exogenous expression of phosphorylation-deficient Y14F caveolin-1 mutant in cav-1(-/-) mouse pulmonary vasculature rendered the mice resistant to LPS compared with reintroduction of wild-type caveolin-1. Thus, caveolin-1 Y14 phosphorylation was required for the interaction with TLR4 and activation of TLR4-MyD88 signaling and sepsis-induced lung inflammation. Inhibiting caveolin-1 Tyr(14) phosphorylation and resultant inactivation of TLR4 signaling in pulmonary vascular endothelial cells represent a novel strategy for preventing sepsis-induced lung inflammation and injury.

Functional characterization of three mouse formyl peptide receptors.

The evolutionary relationship and functional correlation between human formyl peptide receptors (FPRs) and their mouse counterparts remain incompletely understood. We examined three members of the mouse formyl peptide receptor subfamily (mFprs) and found that they differ in agonist preference and cellular distributions. When stably expressed in transfected rat basophilic leukemia (RBL-2H3) cells, mFpr1 was readily activated by N-formylated peptides derived from Listeria monocytogenes (fMIVTLF), Staphylococcus aureus (fMIFL), and mitochondria (fMMYALF). In contrast, the Escherichia coli-derived fMLF was 1000-fold less potent. The aforementioned peptides were much less efficacious at mFpr2, which responded better to the synthetic hexapeptide WKYMVm, the synthetic agonists Quin-C1 (a substituted quinazolinone), and compound 43 (a nitrosylated pyrazolone derivative). Saturation binding assays showed that mFpr1 and mFpr2 were expressed at similar levels on the cell surface, although their affinity for N-formyl-Met-Leu-Phe-Ile-Ile-Lys-fluorescein isothiocyanate varied by more than 1000-fold [dissociation constant (K(d)) values of 2.8 nM for mFpr1 and 4.8 µM for mFpr2]). Contrary to these receptors, mFpr-rs1 responded poorly to all the previously mentioned peptides that were tested. Fluorescent microscopy revealed an intracellular distribution pattern of mFpr-rs1. On the basis of these results, we conclude that mFpr1 is an ortholog of human FPR1 with certain pharmacologic properties of human FPR2/ALX, whereas mFpr2 has much lower affinity for formyl peptides. The intracellular distribution of mFpr-rs1 suggests an evolutionary correlation with human FPR3.

Characterization of novel VEGF (vascular endothelial growth factor)-C splicing isoforms from mouse.

VEGF (vascular endothelial growth factor)-C is a major growth factor implicated in various physiological processes, such as angiogenesis and lymphangiogenesis. In the present paper, we report the identification of three short VEGF-C splicing isoforms (VEGF-C62, VEGF-C129 and VEGF-C184) from immortalized mouse kidney PTECs (proximal tubular epithelial cells). Semi-quantitative RT (reverse transcription)-PCR analysis showed these isoforms were universally expressed to varying degrees in different tissues with high expression levels in the kidney. In immortalized PTECs and podocytes, VEGF-C62 can activate phosphorylation of FAK (focal adhesion kinase) and promote cell adhesion to substratum. Cell survival was also increased by VEGF-C62 treatment in the absence of serum. VEGF-C62 can also reduce cell proliferation in PTECs and podocytes. Nucleolin was one of the proteins that associated with VEGF-C62 in pull-down assays using GST (glutathione transferase) fusion proteins as bait, indicating different protein binding requirements for VEGF-C62 compared with VEGF-C. In conclusion, these newly identified VEGF-C isoforms represent a new class of proteins, which are potentially involved in epithelial cell adhesion and proliferation through novel receptor pathways.

[Expression and clinical significance of nuclear matrix protein 22 and cytokeratin 18 in transitional cell carcinoma of the bladder].

OBJECTIVE: To evaluate the expression and clinical significance of urinary nuclear matrix protein (NMP22) and cytokeratin 18 (CK18) for transitional cell carcinoma of the bladder. METHODS: Urinary NMP22 and CK18 levels of 293 patients with transitional cell carcinoma of the bladder, 400 patients with non-transitional cell carcinoma of the bladder, and 105 bladder benign disease were analysed by enzyme-linked-immunosorbent assay (ELISA). RESULTS: The levels of urinary NMP22 and CK18 in the patients with transitional cell carcinoma of the bladder (M = 17.3 U/ml, M(CK18) = 484.2 U/L) were significantly higher than those in the non-transitional cell carcinoma of the bladder (M = 6.8 U/ml, M(CK18) = 156.0 U/L) and the benign disease group (M(NMP22) = 2.3 U/ml, M(CK18) = 66.6 U/L) (P < 0.001). The sensitivity and specificity of urinary NMP22 and CK18 were 79.2%, 88.6% and 78.2%, 82.9%, respectively, for transitional cell carcinoma of the bladder before any treatment. The joint sensitivity of the two markers was 91.7%. The NMP22 and CK18 levels were significantly lower in the recovered patients after surgical operation (P < 0.01), while in patients with recurrence or metastasis the levels of the markers were significantly higher (P < 0.01). There was a significant relationship between NMP22 and CK18, (r = 0.689, P < 0.01). The levels of urinary nmp22 and CK18 were significantly different among pathological grade G1, G2, G3, and stage Ta, T1, T2, T3 (P < 0.01). CONCLUSION: NMP22 and CK18 are useful tumor marker for diagnosis of transitional cell carcinoma of the bladder and for monitoring the state of illness. The joint use of the two markers can improve the sensitivity of cancer detection. NMP22 and CK18 may become a new class of tumor markers, and to be the basis for development of a new assay with an increased efficacy for the detection and treatment of bladder cancer.

[Research progress on effect of coumarins compounds in anti-tumor].

Coumarins are a group of important natural compounds, and have been found to have multi-biological activities such as anti-HIV, anti-tumor, anti-hypertension, anti-arrhythmia, anti-osteoporosis, assuaging pain, preventing asthma and antisepsis. One of which is its anti-tumor effect and that is a research focus on. Therefore, we believe that it is necessaryto carry out further studies on the effect of coumarins compounds in anti-tumor. Investigation should emphasize on improving techniques for extraction and separation, searching the effective precursory compound, and synthesizing and screening out courmarin derivatives with high activity and low toxicity. Here the recent research progress in anti-tumor effect of coumarins compounds is reviewed.

Evaluating apoptosis in tumor cells infected with adenovirus expressing p53 tumor suppressor gene.

This chapter describes several methods for recognizing apoptosis in tumor cells following infection with a replication-deficient adenovirus expressing the tumor suppressor gene p53. We include cytotoxicity assays and assays of apoptosis, including DNA-nucleosomal DNA fragmentation (DNA laddering), TUNEL, DAPI staining, analysis of the sub-G1 (subdiploid) population, and degradation of poly(ADP-ribose) polymerase (as assayed by Western blot). Although this is not a comprehensive list of protocols to evaluate apoptosis, we believe that these will cover the majority of conditions of apoptosis that may arise. The chapter also describes the characteristics of each technique, including the advantages and disadvantages of each method.

Development of oncolytic adenovirus armed with a fusion of soluble transforming growth factor-beta receptor II and human immunoglobulin Fc for breast cancer therapy.

We have developed an approach to cancer gene therapy in which the oncolytic effects of an adenoviral vector have been combined with selective expression of a soluble form of transforming growth factor (TGF)-beta receptor II fused with Fc (sTGFbetaRIIFc). We chose to use adenoviral dl01/07 mutant because it can replicate in all cancer cells regardless of their genetic defects. An oncolytic adenovirus expressing sTGFbetaRIIFc (Ad.sT- betaRFc) was constructed by homologous recombination. Infection of MDA-MB-231 and MCF-7 human breast cancer cells with Ad.sTbetaRFc produced sTGFbetaRIIFc, which was released into the media. The conditioned media containing sTGFbetaRIIFc could bind with TGF-beta 1 and inhibited TGF-beta-dependent transcription in target cells. Infection of MDA-MB-231, MCF-7, and 76NE human breast cancer cells with Ad.sTbetaRFc resulted in high levels of viral replication, comparable to that of a wild-type dl309 virus. Although some viral replication was observed in actively dividing normal human lung fibroblasts, there was no replication in nonproliferating normal cells. Direct injection of Ad.sTbetaRFc into MDA-MB-231 human breast xenograft tumors grown in nude mice resulted in a significant inhibition of tumor growth, causing tumor regression in more than 85% of the animals. These results indicate that it is possible to construct an oncolytic virus expressing sTGFbetaRIIFc in which both viral replication and transgene expression remain intact, and the recombinant adenovirus is oncolytic in a human tumor xenograft model. On the basis of these results we believe that it may be feasible to develop a cancer gene therapy approach using Ad.sTbetaRFc as an antitumor agent.

An oncolytic adenovirus expressing soluble transforming growth factor-beta type II receptor for targeting breast cancer: in vitro evaluation.

In recent years, adenoviruses that selectively replicate in tumor cells have been developed. However, there is a tremendous need to improve their anticancer efficacy. We wish to investigate whether a strategy that combines the oncolytic effects of an adenoviral vector with simultaneous expression of soluble form of transforming growth factor-beta type II receptor (sTGFbetaRII) offers a therapeutic advantage. We chose to target TGF-betas because they play a pivotal role in late-stage tumorigenesis by enhancing tumor invasion and metastasis. A sTGFbetaRII cDNA was cloned in conditionally replicating adenoviral vector rAd-sTRII and in a replication-deficient adenovirus Ad-sTRII. Infection of MDA-MB-231 breast cancer cells with rAd-sTRII or Ad-sTRII followed by Western blot analysis indicated the expression of diffused glycosylated forms of sTGFbetaRII that were also secreted into the extracellular medium. The secreted proteins were shown to bind with TGF-beta and antagonize TGF-beta-induced p38 mitogen-activated protein kinase activity. However, marked differences in the replication potential of rAd-sTRII and Ad-sTRII were observed in breast tumor cells. Infection of MDA-MB-231 cells with rAd-sTRII resulted in cytotoxicity and significant increase in the adenoviral titers that were comparable with a wild-type adenovirus dl309. However, Ad-sTRII was much less toxic to the tumor cells, and the viral titers of Ad-sTRII remained relatively unchanged. These results suggest that the infection of breast tumor cells with conditionally replicating adenoviral vector rAd-sTRII produced sTGFbetaRII that can abrogate TGF-beta signaling while maintaining the replication potential of the virus, indicating that rAd-sTRII could be a potential anticancer agent.

Targeting TGFbeta signaling for cancer therapy.

Transforming growth factor (TGF) betas are multifunctional polypeptides that regulate several cellular functions, including cell growth and differentiation, extra cellular matrix production, motility and immunosuppression. The growth-inhibiting properties of TGFbeta have gained much attention into its role as a tumor suppressor. There is, however, now increasing evidence that TGFbeta switches roles, from tumor suppressor to tumor promoter, as the tumor progresses. Given the integral role of TGFbeta in the tumor progression, it follows that TGFbeta signaling offers an attractive target for cancer therapy. Several strategies including the use of antisense oligonucleotides for TGFbeta, TGFbeta antibodies, dominant negative TGFbeta receptor II, and small drug-molecules to inhibit TGFbeta receptor I kinase have shown great promise in the preclinical studies. These new findings, coupled with progressing clinical trials indicate that inhibition of TGFbeta signaling may, indeed, be a viable option to cancer therapy. This review summarizes the TGFbeta signaling, the dual role of TGFbeta--as a tumor suppressor and tumor promoter, and various strategies targeted against TGFbeta signaling for cancer therapy. The next few years promise to better our understanding of approaching cancer therapy with an eye to the inhibition of TGFbeta signaling.