Complete Mitochondrial Genomes of Chimpanzee- and Gibbon-Derived Ascaris Isolated from a Zoological Garden in Southwest China

Roundworms (Ascaridida: Nematoda), one of the most common soil-transmitted helminths (STHs), can cause ascariasis in various hosts worldwide, ranging from wild to domestic animals and humans. Despite the veterinary and health importance of the Ascaridida species, little or no attention has been paid to roundworms infecting wild animals including non-human primates due to the current taxon sampling and survey bias in this order. Importantly, there has been considerable controversy over the years as to whether Ascaris species infecting non-human primates are the same as or distinct from Ascaris lumbricoides infecting humans. Herein, we first characterized the complete mitochondrial genomes of two representative Ascaris isolates derived from two non-human primates, namely, chimpanzees (Pan troglodytes) and gibbons (Hylobates hoolock), in a zoological garden of southwest China and compared them with those of A. lumbricoides and the congeneric Ascaris suum as well as other related species in the same order, and then used comparative mitogenomics, genome-wide nucleotide sequence identity analysis, and phylogeny to determine whether the parasites from chimpanzees and gibbons represent a single species and share genetic similarity with A. lumbricoides. Taken together, our results yielded strong statistical support for the hypothesis that the chimpanzee- and gibbon-derived Ascaris represent a single species that is genetically similar to A. lumbricoides, consistent with the results of previous morphological and molecular studies. Our finding should enhance public alertness to roundworms originating from chimpanzees and gibbons and the mtDNA data presented here also serves to enrich the resource of markers that can be used in molecular diagnostic, systematic, population genetic, and evolutionary biological studies of parasitic nematodes from either wild or domestic hosts.     

Lamina replacement with titanium plate fixation improves spinal stability after total lumbar laminectomy

Biomechanical experiments and strain analyses were performed to investigate the effects of lamina replacement surgery for intraspinal lesions on postoperative spinal stability. Eight specimens of thoracic and lumbar vertebrae (T12–L4) were collected from adult cadavers. Stepwise lumbar total laminectomy, and laminoplasty with lamina reduction and replacement was undertaken in combination with titanium-plate fixation to simulate the surgical setting. The effects of thoracic and lumbar vertebral strain, displacement, and rigidity on spinal stability were measured following both single and multiple segment laminectomy. Significant differences in mechanical indices of stability were seen between stepwise laminectomy of lumbar vertebrae and normal specimens (p < 0.05), between lamina replacement in combination with titanium-plate fixation and laminectomy (p < 0.05), and between single- and multiple-segment laminectomy (p < 0.05). Differences between laminoplasty with lamina replacement in combination with titanium-plate fixation and normal specimens need to be examined for further study. Lumbar laminectomy followed by reduction and replacement, in combination with titanium-plate fixation, was shown to be beneficial in terms of preserving spinal stability and maintaining biomechanical function and spinal loading capability.     

Involvement of reactive oxygen species in Microcystin-LR-induced cytogenotoxicity

Microcystin-LR (MCLR) is a potent hepatotoxin. Oxidative stress is thought to be implicated in the cytotoxicity of MCLR, but the mechanisms by which MCLR produces reactive oxygen species (ROS) are still unclear. This study investigated the role and possible sources of ROS generation in MCLR-induced cytogenotoxicity in HepG2, a human hepatoma cell line. MCLR increased DNA strand breaks, 8-hydroxydeoxiguanosine formation, lipid peroxidation, as well as LDH release, all of which were inhibited by ROS scavengers. ROS scavengers partly suppressed MCLR-induced cytotoxicity determined by the MTT assay. MCLR induced the generation of ROS, as confirmed by confocal microscopy with 2-[6-(4′-hydroxy)phenoxy-3H-xanthen-3-on-9-yl]benzoic acid, and upregulated the expression of CYP2E1 mRNA. In addition, CYP2E1 inhibitors chlormethiazole and diallyl sulphide inhibited both ROS generation and cytotoxicity induced by MCLR. The results suggest that ROS contribute to MCLR-induced cytogenotoxicity. CYP2E1 might be a potential source responsible for ROS generation by MCLR.     

Chionographis shiwandashanensis sp. nov. (Melanthiaceae) from southern Guangxi,China

Chionographis shiwandashanensis a new species from southern Guangxi, China, is described and illustrated. It is similar to C. koidzumiana Ohwi and C. chinensis Krause in habit, but distinguished by oblong‐elliptic to oblong bractlike leaves and 6, filiform, regular tepals.     

Molecular cloning and characterization of SoNCED, a novel gene encoding 9-cis-epoxycarotenoid dioxygenase from sugarcane (Saccharum officinarum L.)

Abscisic acid (ABA) plays important roles in adaptive responses to various environmental stresses. The rate-limiting step in ABA biosynthesis is the oxidative cleavage of cis-epoxycarotenoids, which is catalyzed by 9-cis-epoxycarotenoid dioxygenase (NCED). In this experiment, a full-length cDNA encoding NCED gene was cloned by RT-PCR and RACE from sugarcane (Saccharum officinarum L.). The full-length of SoNCED is 2,521 bp with 1,827 bp open reading frame, encoding a peptide of 608 amino acids. The calculated molecular weight of protein was 65.9 kDa with isoelectric point of 6.04. Conserved domains prediction indicated a chloroplast-targeting peptide located at N-terminus of SoNCED. Phylogenetic tree, constructed by Neighbor-Joining method indicated that SoNCED shared high identity with the NCEDs reported from other plant species. Sequence alignment revealed that the basic secondary structure including α-helices, β-strands, β-propeller and His residues coordinating catalytic sites of SoNCED were highly conserved as in the NCEDs from other plants. Tissue specific expression analysis using quantitative real-time PCR showed a significant increase in SoNCED mRNA level and its correlation with O₂ ^– production rate and ABA accumulation in leaves and roots of sugarcane variety GT21 when exposed to water stress. Further, the stimulation of SoNCED mRNA level, O₂ ^– production rate and ABA content after exogenous application of ABA (100 μMol l^?1) proved its involvement in pathways providing tolerance to drought stress.     

Silencing of DLGAP5 by siRNA Significantly Inhibits the Proliferation and Invasion of Hepatocellular Carcinoma Cells

Background

The dysregulation of oncogenes and tumor suppressor genes plays an important role in many cancers, including hepatocellular carcinoma (HCC), which is one of the most common cancers in the world. In a previous microarray experiment, we found that DLGAP5 is overexpressed in HCCs. However, whether the up-regulation of DLGAP5 contributes to hepatocarcinogenesis remains unclear.

Methodology/Principal Findings

In this study, we showed that DLGAP5 was significantly up-regulated in 76.4% (168 of 220) of the analyzed HCC specimens when compared with adjacent liver tissue. DLGAP5 overexpression was evident in 25% (22 of 88) of the HCC specimens without AFP expression, suggesting that DLGAP5 may be a novel biomarker for HCC pathogenesis. The silencing of DLGAP5 gene expression by RNA interference significantly suppressed cell growth, migration and colony formation in vitro. The expression level of DLGAP5 was also found to be related to the methylation level of its promoter in the HCC specimens.

Conclusions/Significance

Taken together, these data suggest that the expression of DLGAP5 is regulated by methylation and that the up-regulation of DLGAP5 contributes to HCC tumorigenesis by promoting cell proliferation.     

c-kit+ Cardiac Stem Cells Alleviate Post-Myocardial Infarction Left Ventricular Dysfunction Despite Poor Engraftment and Negligible Retention in the Recipient Heart

Although transplantation of c-kit+ cardiac stem cells (CSCs) has been shown to alleviate left ventricular (LV) dysfunction induced by myocardial infarction (MI), the number of exogenous CSCs remaining in the recipient heart following transplantation and their mechanism of action remain unclear. We have previously developed a highly sensitive and accurate method to quantify the absolute number of male murine CSCs in female recipient organs after transplantation. In the present study, we used this method to monitor the number of donor CSCs in the recipient heart after intracoronary infusion. Female mice underwent a 60-min coronary occlusion followed by reperfusion; 2 days later, 100,000 c-kit+/lin- syngeneic male mouse CSCs were infused intracoronarily. Only 12.7% of the male CSCs present in the heart immediately (5 min) after infusion were still present in the heart at 24 h, and their number declined rapidly thereafter. By 35 days after infusion, only ∼1,000 male CSCs were found in the heart. Significant numbers of male CSCs were found in the lungs and kidneys, but only in the first 24 h. The number of CSCs in the lungs increased between 5 min and 24 h after infusion, indicating recirculation of CSCs initially retained in other organs. Despite the low retention and rapid disappearance of CSCs from the recipient heart, intracoronary delivery of CSCs significantly improved LV function at 35 days (Millar catheter). These results suggest that direct differentiation of CSCs alone cannot account for the beneficial effects of CSCs on LV function; therefore, paracrine effects must be the major mechanism. The demonstration that functional improvement is dissociated from survival of transplanted cells has major implications for our understanding of cell therapy. In addition, this new quantitative method of stem cell measurement will be useful in testing approaches of enhancing CSC engraftment and survival after transplantation.     

Mechanisms of GOLPH3 Associated with the Progression of Gastric Cancer: A Preliminary Study

Study Design

To investigate the specific mechanisms by which Golgi phosphoprotein 3 (GOLPH3) affects the progression of gastric cancer and to explore its clinical significance.

Methods

Immunohistochemical analysis was used to evaluate the correlations between GOLPH3, phosphorylated mTOR (p-mTOR), phosphorylated Akt (p-Akt), phosphorylated p70S6 (p-p70S6), phosphorylated 4E-BP1 (p-4E-BP1) and the clinicopathological features of gastric cancer. The mRNA expression levels of GOLPH3, mTOR, Akt, p70S6 and 4E-BP1 in gastric cancer, carcinoma-adjacent and paired normal tissue were analyzed using RT-PCR. Western blotting was used to determine the protein expression of GOLPH3, p-mTOR, p-Akt, p-p70S6 and p-4E-BP1 in tissues.

Results

High expression protein levels of GOLPH3, p-AKT, p-mTOR, p70S6, p-4E-BP1 were positively associated with histological grade (p<0.05), depth of invasion (p<0.05), distant metastasis (p<0.05) and lymph node involvement (p<0.05). Compared with carcinoma-adjacent and paired normal tissues, the mRNA expression levels of GOLPH3, AKT, mTOR, p70S6 and 4EBP1 in gastric cancer tissues were significantly higher. The protein expression levels of GOLPH3, p-AKT, p-mTOR, p-p70S6 and p-4E-BP1 in gastric cancer tissues were also significantly higher than in carcinoma-adjacent and paired normal tissues. A strong positive correlation was observed between GOLPH3, p-mTOR, p-p70S6 and p-4EBP1 expression (r = 0.410, 0.303 and 0.276, respectively, p<0.05), but no significant correlation between the expression of GOLPH3 and p-Akt was observed.

Conclusions

The GOPLH3 expression level is highly correlated with Akt/mTOR signaling in human gastric cancer samples. GOLPH3 combined with Akt/mTOR signaling activation may play an important role in the development, differentiation, invasion and metastasis of gastric cancer.     

Canine bone marrow mesenchymal stromal cells with lentiviral mHCN4 gene transfer create cardiac pacemakers

Background aimsThe study objective was to test the ability of canine mesenchymal stromal cells (cMSC) transfected with the mouse hyperpolarization-activated cyclic nucleotide-gated channel 4 (mHCN4) gene to deliver a biologic pacemaker to the canine heart.Methods and ResultscMSC that were transfected by lentiviral vector with the cardiac pacemaker gene mHCN4 expressed high levels of Cs⁺ -sensitive current (26.4 ± 1.8pA/pF at –140 mV; (n = 17) and were activated in the diastolic potential range with a reversal potential of –29.7 ± 2.5 mV (n = 14), confirming that the expressed current was Funny current (I_f)-like. Next, 3 × 10⁶ cMSC transfected with either control plasmid or the mHCN4 gene construct were injected subepicardially into the canine right ventricular wall in situ. During sinus arrest, all control hearts had spontaneous atrioventricular node rhythms [rate = 21 ± 5beats per minute (b.p.m.)]. In the mHCN4 group, six of eight animals developed spontaneous ventricular rhythms of right-sided origin (rate = 45 ± 9b.p.m.; P < 0.01). Moreover, immunohistochemical analysis of the injected regions demonstrated neither apoptosis nor cellular or humoral rejection at 2 weeks.ConclusionsThese results demonstrate that genetically modified cMSC can express functional HCN4 channels in vitro and in vivo and represent a novel delivery system for pacemaker genes into the heart.