[ANXA8 Regulates Proliferation of Human Non-Small Lung Cancer Cells A549 via EGFR-AKT-mTOR Signaling Pathway].

Annexin A8 (ANXA8) is a member of the annexin family, which had been reported to regulate multiple cancer cellular processes including proliferation, metastasis and inflammation. However, the specific role of ANXA8 in lung cancer cell biology remains unknown. Our previous transcriptome study revealed that ANXA8 mRNA was downregulated in curcumin analog (MHMD) -treated human non-small lung cancer cells (A549 cell line). Here, we continued to study the ANXA8 expression in A549 cells using reverse transcription-quantitative PCR and Western blotting, compared with that in human normal bronchial epithelium cells (BE-AS-2B cell line). Overexpression of ANXA8 via transfection of pEGFP-ANXA8 recombinant vector contributed to the proliferation and migration of A549 cells. Moreover, the cell cycle protein cyclin E1 was upregulated in ANXA8-transfected A549 cells. Knockdown of ANXA8 using an RNA interference technique decreased A549 cell viability and restrained their migration in vitro. The expression levels of multiple cellular factors, including EGFR, PI3K, Akt, mTOR, p70S6K and 4EBP1, in the epidermal growth factor receptor (EGFR) signaling pathway were also altered by ANXA8 knockdown or overexpression in A549 cells, which confirmed the activation of the EGFR/Akt/mTOR signaling pathway by ANXA8. The present results provided evidence to support further investigation of the functional identification of ANXA8 in lung cancer cells in the future.

[Study on the risk factors for hemorrhagic transformation of cerebellar infarction after posterior fossa decompression surgery].

Objective: To investigate the related risk factors for hemorrhagic transformation (HT) of cerebellar infarction after posterior fossa decompression surgery. Methods: A total of 91 patients with cerebellar infarction were treated by posterior fossa decompression surgery in Department of Neurosurgery of Jinhua Municipal Central Hospital from Jan 2010 to Jan 2018, were selected as study subjects. The HT group included 17 cases, while the Non-HT group included 74 cases. The clinical data of the two groups were analyzed retrospectively, the univariate and non-conditional lgistic regression analysis were performed to detect the relevant risk factors for hemorrhagic transformation of cerebellar infarction after posterior fossa decompression surgery. Results: By univariate analysis, the differences of these seven risk factors, the large area cerebellar infarction (the diameter of area was larger than 5 cm), pre-op thrombolysis, pre-op mild HT, oral anticoagulants, atrial fibrillation, hyperglycemia and fluctuation of BP in post-op, between two groups were statistically significant (P<0.05). By multivariate logistic analysis, the large area cerebellar infarction (P<0.05), pre-op thrombolysis(P<0.01), pre-op mild HT (P<0.01), oral anticoagulants (P<0.01) were the independent risk factors for post-op HT. Conclusions: The large area cerebellar infarction (the diameter of area was more than 5 cm), pre-op thrombolysis, pre-op mild HT, oral anticoagulants, atrial fibrillation, hyperglycemia and fluctuation of BP in post-op are important risk factors for post-op HT. The large area cerebellar infarction, pre-op thrombolysis, pre-op mild HT, oral anticoagulants are the independent risk factors for post-op HT. A proper pre-op evaluation of these risk factors and an individualized treatment for post-op HT would help a lot with balancing operational risk and improving prognosis.

CARD9 mutation linked to Corynespora cassiicola infection in a Chinese patient.

Corynespora cassiicola is a plant pathogen associated with leaf-spotting disease. The fungus has been found on diverse substrates: leaves, stems and roots of plants; nematode cysts and human skin. It rarely causes human infections. Here we report one case of subcutaneous phaeohyphomycosis caused by C. cassiicola with prominent tissue necrosis in a woman. All of her clinical features pointed towards a genetic linkage. Hence, whole-exome sequencing and Sanger sequencing were performed on this patient. One mutation of CARD9 was detected.

In silico identification and characterization of the WRKY gene superfamily in pepper (Capsicum annuum L.).

The WRKY family is one of the most important transcription factor families in plants, involved in the regulation of a broad range of biological roles. The recent releases of whole-genome sequences of pepper (Capsicum annuum L.) allow us to perform a genome-wide identification and characterization of the WRKY family. In this study, 61 CaWRKY proteins were identified in the pepper genome. Based on protein structural and phylogenetic analyses, these proteins were classified into four main groups (I, II, III, and NG), and Group II was further divided into five subgroups (IIa to IIe). Chromosome mapping analysis indicated that CaWRKY genes are distributed across all 12 chromosomes, although the location of four CaWRKYs (CaWRKY58-CaWRKY61) could not be identified. Two pairs of CaWRKYs located on chromosome 01 appear to be tandem duplications. Furthermore, the phylogenetic tree showed a close evolutionary relationship of WRKYs in three species from Solanaceae. In conclusion, this comprehensive analysis of CaWRKYs will provide rich resources for further functional studies in pepper.

Specific Cleavage of the Nucleoprotein of Fish Rhabdovirus.

Siniperca chuatsi rhabdovirus (SCRV) is one of myriad rhabdoviruses recorded in fish. Preliminary data show that inhibition of the SCRV nucleoprotein (N) could significantly reduce the progeny virus titers in infected Epithelioma papulosum cyprinid (EPC) cells. Here, the authors propose that cleavage of the viral 47-kDa N protein is caspase-mediated based on caspase inhibition experiments, transient expression in EPC transfection, and analysis of cleavage sites. Cleavage of the SCRV N protein in culture was prevented by a pan-caspase inhibitor, z-VAD-FMK (z-Val-Ala-DL-Asp-fluoromethyl ketone). Subsequently, N was transiently expressed in EPC cells, the results of which indicated that the specific cleavage of N also occurred in the cells transfected with N-GFP plasmid. Several truncated fragments of the N gene were constructed and transiently transfected into EPC cells. Immunoblotting results indicated that D324 and D374 are the cleavage sites of N by caspases. The authors also found that z-VAD-FMK could inhibit the cytopathic effect in SCRV-infected EPC cells but not affect the production of infectious progeny, suggesting that the caspase-mediated cleavage of N protein is not required for in vitro SCRV replication. To the authors' knowledge, this is the first report on the cleavage of rhabdovirus proteins.

Roles of peripheral B1 cells in the individualized treatment of adult idiopathic thrombocytopenic purpura.

We aimed to explore the changes of peripheral B1 cells before and after treatment of adult idiopathic thrombocytopenic purpura (ITP) and to investigate the association of these changes with the disease condition and prognosis. Ninety-seven ITP patients were divided into the effective or ineffective groups, based on their response to hormone therapy. Forty healthy volunteers were enrolled into the control group (HC). The percentages of CD19+ cells, B1 cells, and platelet-associated immunoglobulin (PAIg) in peripheral blood from healthy volunteers and ITP patients before and after treatment were evaluated, and blood platelet (PLT) counts were determined. The percentages of CD19+ cells [(21 ± 10.0) vs (11.2 ± 7.1)%], B1 cells [(8.85 ± 5.23) vs (2.2 ± 1.3)%], and PAIg [(28 ± 19) vs (11.7 ± 8)%] in whole blood from ITP patients before treatment were significantly higher than those in whole blood from healthy controls (P < 0.05). Before treatment, the percentage of B1 cells and PAIg in ITP patients was negatively correlated with the PLT level (r = -0.89, P < 0.05 and r = -0.814, P < 0.05, respectively). Further, the B1 cell percentage was positively associated with the PAIg percentage in ITP patients before treatment. In the effective group, the B1 cell percentage was reduced sharply at 1 month after treatment [(2.45 ± 1.75) vs (8.74 ± 5.04)%, P < 0.05)], so as at 3 and 6 months. However, in the ineffective group, there was no difference in the B1 cell percentage before and after treatment [(7.9 ± 5.6) vs (8.76 ± 5.26)%]. This obvious association of changes in peripheral B1 cells with disease condition and prognosis in ITP patients may be of certain clinical significance for guiding the individualized treatment of ITP.

Assessment of the genetic diversity of tomato yellow leaf curl virus.

The objective of the present study was to analyze the genetic diversity of tomato yellow leaf curl virus (TYLCV). Representative TYLCV sequences were searched in the National Center for Biotechnology Information database. Comprehensive analysis of TYLCV was performed using bioinformatics by examining gene structure, sequence alignments, phylogeny, GC content, and homology. Forty-eight representative TYLCV sequences were selected from 48 regions in 29 countries. The results showed that all TYLCV sequences were 2752-2794 nucleotides in length, which encoded 6 open reading frames (AV1, AV2, AC1, AC2, AC3, and AC4). GC content ranged from 0.41-0.42. Sequence alignment showed a number of insertions and deletions within these TYLCV sequences. Phylogenetic tree results revealed that the sequences were divided into 10 classes; homology of the sequences ranged from 72.8 to 98.6%. All 48 sequences contained the typical structure of TYLCV, including open reading frames and intergenic regions. These results provide a theoretical basis for the identification and evolution of the virus in the future.

In silico analysis of gene content in tomato genomic regions mapped to the Ty-2 resistance gene.

Tomato yellow leaf curl virus is one of the main diseases affecting tomato production worldwide. Previous studies have shown that Ty-2 is an important resistance gene located between molecular markers C2_At2g28250 (82.3 cM) and T0302 (89.0 cM), and exhibits strong resistance to tomato yellow leaf curl virus in Asia. In this study, Ty-2 candidate genes were subjected to bioinformatic analysis for the sequenced tomato genome. We identified 69 genes between molecular markers C2_At2g28250 and T0302, 22 of which were disease-related resistant genes, including nucleotide binding site-leucine-rich repeat disease resistance genes, protease genes (protein kinase, kinase receptor, and protein isomerase), cytochromes, and transcription factors. Expressed sequence tag analysis revealed that 77.3% (17/22) of candidate disease-resistance genes were expressed, involving 143 expressed sequence tags. Based on full-length cDNA sequence analysis, 7 candidate genes were found, 4 of which were involved in tomato responses to pathogens. Microarray expression analysis also showed that most candidate genes were involved in the tomato responses to multiple pathogens, including fungi, viruses, and bacteria. RNA-seq expression analysis revealed that all candidate genes participated in tomato growth and development.

Long non-coding RNA HOTAIR regulates the development of non-small cell lung cancer through miR-217/DACH1 signaling pathway.

OBJECTIVE: Long non-coding RNA HOX transcript antisense RNA (HOTAIR) is reported to make chromatin state, cell growth and cancer metastasis. However, the role of HOTAIR in non-small cell lung cancer (NSCLC) remains unknown. The aim of this study was to explore the regulatory mechanism of HOTAIR in NSCLC in relation to miR-217/Dachshund homolog 1 (DACH1) signaling pathway. MATERIALS AND METHODS: The expression levels of HOTAIR and miR-217 were measured by quantitative Polymerase Chain Reaction (qPCR) in NSCLC cell lines and human bronchial epithelial cell line HBE. The direct target of HOTAIR and miR-217 in NSCLC was confirmed by a Luciferase reporter assay. The expression of DACH1 protein was examined by Western blot (WB) assay. Cell migration and invasion were examined with transwell assays, and cell proliferation was measured by Cell Counting Kit-8 (CCK8) assay. RESULTS: HOTAIR was up-regulated and miR-217 was down-regulated in NSCLC cell lines. Silencing of HOTAIR significantly repressed cell proliferation and inhibited cell migration and invasion in H1299 and A549 cells by facilitating miR-217 expression. Moreover, bioinformatics analysis and Luciferase reporter assay confirmed that DACH1 was a target of miR-217. Furthermore, the overexpression of miR-217 markedly repressed cell proliferation and inhibited cell migration and invasion in H1299 and A549 cells. DACH1 reverses the effects of miR-217 overexpression in NSCLC cells. CONCLUSIONS: HOTAIR was up-regulated in NSCLC cell and regulates the proliferation, migration, invasion through the miR-217/DACH1 signaling pathway. It provides a novel potential treatment strategy for NSCLC.

Quantitative characterization of multiple binding sites for phencyclidine and N-allylnormetazocine in membranes from rat and guinea pig brain.

Quantitative ligand binding studies have been used to characterize binding sites for N-allylnormetazocine ((+)SKF10,047) (SKF), 1-(1-phenylcyclohexyl) piperidine (PCP), N-[1-(2-thienyl)cyclohexyl]piperidine (TCP) and haloperidol in membranes from the brain of rat and guinea pig under conditions which permitted simultaneous analysis of the binding of both PCP and SKF. Using four labelled ligands (SKF, TCP, PCP and haloperidol), each displaced by the corresponding four unlabelled ligands, four classes of binding sites were observed in membranes from the brain of the rat, corresponding to sigma (sigma), two classes of PCP sites (PCP1, PCP2) and dopamine (D2) sites. The sigma site was suppressed by 50 nM haloperidol, while the PCP1 and PCP2 sites were not. These results were confirmed by studies employing a self- and cross-displacement design and dose-response surfaces for SKF and TCP, with and without blockade by haloperidol of the sigma site. Using mathematical modelling, employing the program LIGAND, it was possible to reject simpler models involving a common "PCP/sigma" site or a model involving only two classes of sites (sigma and PCP). Similar methods were used to identify two classes of sigma binding sites and two classes of PCP binding sites, in membranes prepared from the brain of the guinea pig. The relative potencies of 18 ligands for displacement of (+)[3H]SKF10,047 and [3H]TCP were compared: there were significant qualitative and quantitative differences in the "sigma" binding sites in the brain of rat and guinea pig, while the PCP binding sites were very similar in the two species.

L-homocysteate stimulates [3H]MK-801 binding to the phencyclidine recognition site and is thus an agonist for the N-methyl-D-aspartate-operated cation channel.

Rat brain synaptosomal membranes that are depleted of endogenous excitatory amino acids cannot bind [(+)-5-methyl-10, 11-dihydro-5H-dibenzo(a,d]cyclohept-5,10-imine maleate] ([3H]MK-801). However, they do so upon the restoration of excitatory amino acid agonists such as L-glutamate. [3H]MK-801 provides a molecular probe which is specific for a binding site located within the ionophore of the N-methyl-D-aspartate-type excitatory amino acid receptor, [3H]MK-801 does not bind to non-N-methyl-D-aspartate excitatory amino acid receptors. Exploiting [3H]MK-801 binding as a quantitative measure of agonist activity with respect to ability of inducing the open channel conformation, the present study demonstrates that L-homocysteate is an agonist almost equivalent to L-glutamate in terms of efficacy (maximal N-methyl-D-aspartate response) as well as potency (EC50). The effect of L-homocysteate was dose-dependent, stereospecific (L-homocysteate greater than DL-homocysteate greater than D-homocysteate), suppressible by the N-methyl-D-aspartate-selective competitive antagonist (+/-)-3(2-carboxy-piperazine-4-yl)propyl-l-phosphonate, and potentiated by the N-methyl-D-aspartate-selective "allosteric" modulator glycine. The demonstrated inactivity of L-homocysteine (and virtually all naturally occurring, non-acidic amino acids) implies that the omega-sulphonic acid moiety is an acceptable substitute for the omega carboxyl group for activating the N-methyl-D-aspartate receptor. While the potency of L-homocysteate at N-methyl-D-aspartate receptors was by a factor of only 1.6 smaller than that of L-glutamate, the affinity of L-homocysteate for kainate-type excitatory amino acid receptors was approximately four-fold lower than that of L-glutamate.(ABSTRACT TRUNCATED AT 250 WORDS)

Lower CSF level of dynorphin(1-8) immunoreactivity in schizophrenic patients.

Using a highly specific and sensitive radioimmunoassay for dynorphin(1-8) (D 1-8), and a singly blind test design, we measured D(1-8) immunoreactivity (ir D 1-8) in CSF of 35 first break cases of acute schizophrenic patients. All patients were free of psychotropic medication for at least one week before the study. Another 31 neurological patients suffered from cervical arthrosis, tumor, myelopathy etc. were studied as controls. The ir D(1-8) in CSF of schizophrenic patients were significantly lower than that of controls (91.8 +/- 5.6, means +/- S.E.M., and 131.9 +/- 6.8 fmol/ml CSF respectively, p less than 0.001).

Computer-assisted modeling of multiple dextromethorphan and sigma binding sites in guinea pig brain.

Computer-assisted, simultaneous analysis of self- and cross-displacement experiments demonstrated the existence of several binding sites in guinea pig brain for dextromethorphan, (+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine ((+)-3-PPP), and 1,3-di-o-tolyl guanidine (DTG). Dextromethorphan binds with high affinity to two sites (R1 Kd 50-83 and R2 Kd 8-19 nM) and with low affinity to two additional sites (R3 and R4). (+)-3-PPP binds to one high-affinity (R1 Kd 24-36 nM), to one intermediate-affinity (R3 Kd 210-320 nM), and to two (R2 and R4) low-affinity sites. DTG binds with almost identical high affinity to two different sites (R1 Kd 22-24 and R3 Kd 13-16 nM). These results confirm that dextromethorphan, (+)-3-PPP, and DTG bind to the common DM1/sigma 1 site (R1). The binding of DTG to two different sites with identical affinities precludes the use of this compound as a specific marker for sigma receptors. Besides, haloperidol displaces labeled ligands from both high-affinity DTG sites (R1 and R3) with high affinity. Thus, haloperidol sensitivity should not be used as the single criterion to identify a putative receptor. The resolution of these novel sites also may provide new insights into the multiple effects of antipsychotic drugs. In addition, this investigation has important implications regarding the methods that must be applied to characterize multiple binding sites and their relations with putative receptors.

Rigid spine syndrome. Report of 4 cases.

Four cases of rigid spine syndrome are reported for the first time in China. They are 2 males and 2 females. One girl has a positive family history. Limitation of flexion of the cervical and dorsolumbar spine, contractures of joints, especially the elbow joints, extreme flattening of the chest and scoliosis are the prominent symptoms of this syndrome. The clinical criteria for this rigid spine syndrome tally with those described by Dubowitz (1965). Symptomatology, diagnosis, differential diagnosis and genetics are discussed. However, the question whether rigid spine syndrome is a single, well defined disease entity remains to be explored.

Demonstration of the anti-viral activity of garlic extract against human cytomegalovirus in vitro.

The in vitro anti-viral activity of garlic extract (GE) on human cytomegalovirus (HCMV) was evaluated by tissue culture, plaque reduction and early antigen assay. A dose dependent inhibitory effect of GE was evident when GE was applied simultaneously with HCMV. But the effect was stronger when the monolayers were pretreated with GE. In addition, the anti-viral effect of GE persisted long in infected cells after its being removed from the culture medium. The strongest anti-viral effect of GE was demonstrated when it was applied continuously. It is therefore recommended that clinical use of GE against HCMV infection should be persistent and the prophylactic use of GE is preferable in immunocompromised patients.

Comparison between transcutaneous nerve stimulation analgesic effect and electroacupuncture analgesic effect in rabbits.

To compare Transcutaneous Nerve Stimulation Analgesia (TNSA) with Electroacupuncture Analgesia (AA), we observed their effect and mechanism parallelly. The results are as follows: 1. Both TNSA and AA can produce significant current-dependent analgesia. The latency of AA is shorter than that of TNSA. 2. Under the present condition (lower-frequency, lower-intensity) naloxone can antagonize partly AA but not TNSA. 3. TNSA effect could be blocked partly by deep or intradermal injection of procaine into points. However, AA effect could be almost blocked completely by deep injection and not by intradermal injection of procaine.