Evolutionary Pattern of the FAE1 Gene in Brassicaceae and Its Correlation with the Erucic Acid Trait

The fatty acid elongase 1 (FAE1) gene catalyzes the initial condensation step in the elongation pathway of VLCFA (very long chain fatty acid) biosynthesis and is thus a key gene in erucic acid biosynthesis. Based on a worldwide collection of 62 accessions representing 14 tribes, 31 genera, 51 species, 4 subspecies and 7 varieties, we conducted a phylogenetic reconstruction and correlation analysis between genetic variations in the FAE1 gene and the erucic acid trait, attempting to gain insight into the evolutionary patterns and the correlations between genetic variations in FAE1 and trait variations. The five clear, deeply diverged clades detected in the phylogenetic reconstruction are largely congruent with a previous multiple gene-derived phylogeny. The Ka/Ks ratio (<1) and overall low level of nucleotide diversity in the FAE1 gene suggest that purifying selection is the major evolutionary force acting on this gene. Sequence variations in FAE1 show a strong correlation with the content of erucic acid in seeds, suggesting a causal link between the two. Furthermore, we detected 16 mutations that were fixed between the low and high phenotypes of the FAE1 gene, which constitute candidate active sites in this gene for altering the content of erucic acid in seeds. Our findings begin to shed light on the evolutionary pattern of this important gene and represent the first step in elucidating how the sequence variations impact the production of erucic acid in plants.     

Patrilineal Background of Esophageal Cancer and Gastric Cardia Cancer Patients in a Chaoshan High-Risk Area in China

The Taihang Mountain range of north-central China, the Southern region area of Fujian province, and the Chaoshan plain of Guangdong province are 3 major regions in China well known for their high incidence of esophageal cancer (EC). These areas also exhibit high incidences of gastric cardia cancer (GCC). The ancestors of the Chaoshanese, now the major inhabitants in the Chaoshan plain, were from north-central China. We hypothesized that EC and GCC patients in Chaoshan areas share a common ancestry with Taihang Mountain patients. We analyzed 16 East Asian-specific Y-chromosome biallelic markers (single nucleotide polymorphisms; Y-SNPs) and 6 Y-chromosome short tandem repeat (Y-STR) loci in 72 EC and 48 GCC patients from Chaoshan and 49 EC and 63 GCC patients from the Taihang Mountain range. We also compared data for 32 Chaoshan Hakka people and 24 members of the aboriginal She minority who live near the Chaoshan area. Analysis was by frequency distribution and principal component, correlation and hierarchical cluster analysis of Y-SNP. Chaoshan patients were closely related to Taihang Mountain patients, even though they are geographically distant. Y-STR analysis revealed that the 4 patient groups were more closely related with each other than with other groups. Network analysis of the haplogroup O3a3c1-M117 showed a high degree of patient-specific substructure. We suggest that EC and GCC patients from these 2 areas share a similar patrilineal genetic background, which may play an important role in the genetic factor of EC and GCC in these populations.     

Temperature Sensitivity and Basal Rate of Soil Respiration and Their Determinants in Temperate Forests of North China

The basal respiration rate at 10°C (R₁₀) and the temperature sensitivity of soil respiration (Q₁₀) are two premier parameters in predicting the instantaneous rate of soil respiration at a given temperature. However, the mechanisms underlying the spatial variations in R₁₀ and Q₁₀ are not quite clear. R₁₀ and Q₁₀ were calculated using an exponential function with measured soil respiration and soil temperature for 11 mixed conifer-broadleaved forest stands and nine broadleaved forest stands at a catchment scale. The mean values of R₁₀ were 1.83 µmol CO₂ m⁻² s⁻¹ and 2.01 µmol CO₂ m⁻² s⁻¹, the mean values of Q₁₀ were 3.40 and 3.79, respectively, for mixed and broadleaved forest types. Forest type did not influence the two model parameters, but determinants of R₁₀ and Q₁₀ varied between the two forest types. In mixed forest stands, R₁₀ decreased greatly with the ratio of coniferous to broadleaved tree species; whereas it sharply increased with the soil temperature range and the variations in soil organic carbon (SOC), and soil total nitrogen (TN). Q₁₀ was positively correlated with the spatial variances of herb-layer carbon stock and soil bulk density, and negatively with soil C/N ratio. In broadleaved forest stands, R₁₀ was markedly affected by basal area and the variations in shrub carbon stock and soil phosphorus (P) content; the value of Q₁₀ largely depended on soil pH and the variations of SOC and TN. 51% of variations in both R₁₀ and Q₁₀ can be accounted for jointly by five biophysical variables, of which the variation in soil bulk density played an overwhelming role in determining the amplitude of variations in soil basal respiration rates in temperate forests. Overall, it was concluded that soil respiration of temperate forests was largely dependent on soil physical properties when temperature kept quite low.     

Methylene Blue Modulates Transendothelial Migration of Peripheral Blood Cells

Vasoplegia is a severe complication after cardiac surgery. Within the last years the administration of nitric oxide synthase inhibitor methylene blue (MB) became a new therapeutic strategy. Our aim was to investigate the role of MB on transendothelial migration of circulating blood cells, the potential role of cyclic cGMP, eNOS and iNOS in this process, and the influence of MB on endothelial cell apoptosis. Human vascular endothelial cells (HuMEC-1) were treated for 30 minutes or 2 hours with different concentrations of MB. Inflammation was mimicked by LPS stimulation prior and after MB. Transmigration of PBMCs and T-Lymphocytes through the treated endothelial cells was investigated. The influence of MB upon the different subsets of PBMCs (Granulocytes, T- and B-Lymphocytes, and Monocytes) was assessed after transmigration by means of flow-cytometry. The effect of MB on cell apoptosis was evaluated using Annexin-V and Propidium Iodide stainings. Analyses of the expression of cyclic cGMP, eNOS and iNOS were performed by means of RT-PCR and Western Blot. Results were analyzed using unpaired Students T-test. Analysis of endothelial cell apoptosis by MB indicated a dose-dependent increase of apoptotic cells. We observed time- and dose-dependent effects of MB on transendothelial migration of PBMCs. The prophylactic administration of MB led to an increase of transendothelial migration of PBMCs but not Jurkat cells. Furthermore, HuMEC-1 secretion of cGMP correlated with iNOS expression after MB administration but not with eNOS expression. Expression of these molecules was reduced after MB administration at protein level. This study clearly reveals that endothelial response to MB is dose- and especially time-dependent. MB shows different effects on circulating blood cell-subtypes, and modifies the release patterns of eNOS, iNOS, and cGMP. The transendothelial migration is modulated after treatment with MB. Furthermore, MB provokes apoptosis of endothelial cells in a dose/time-dependent manner.     

Residues 240–250 in the C-Terminus of the Pirh2 Protein Complement the Function of the RING Domain in Self-Ubiquitination of the Pirh2 Protein

Pirh2 is a p53 inducible gene that encodes a RING-H2 domain and is proposed to be a main regulator of p53 protein, thus fine tuning the DNA damage response. Pirh2 interacts physically with p53 and promotes its MDM2-independent ubiquitination and subsequent degradation as well as participates in an auto-regulatory feedback loop that controls p53 function. Pirh2 also self-ubiquitinates. Interestingly, Pirh2 is overexpressed in a wide range of human tumors. In this study, we investigated the domains and residues essential for Pirh2 self-ubiquitination. Deletions were made in each of the three major domains of Pirh2: the N-terminal domain (NTD), Ring domain (RING), and C-terminal domain (CTD). The effects of these deletions on Pirh2 self-ubiquitination were then assessed using in vitro ubiquitination assays. Our results demonstrate that the RING domain is essential, but not sufficient, for Pirh2 self-ubiquitination and that residues 240–250 of the C-terminal domain are also essential. Our results demonstrate that Pirh2 mediated p53 polyubiquitination occurs mainly through the K48 residue of ubiquitin in vitro. Our data further our understanding of the mechanism of Pirh2 self-ubiquitination and may help identify valuable therapeutic targets that play roles in reducing the effects of the overexpression of Pirh2, thus maximizing p53''s response to DNA damage.     

Functional Recovery of Denervated Skeletal Muscle with Sensory or Mixed Nerve Protection: A Pilot Study

Functional recovery is usually poor following peripheral nerve injury when reinnervation is delayed. Early innervation by sensory nerve has been indicated to prevent atrophy of the denervated muscle. It is hypothesized that early protection with sensory axons is adequate to improve functional recovery of skeletal muscle following prolonged denervation of mixed nerve injury. In this study, four groups of rats received surgical denervation of the tibial nerve. The proximal and distal stumps of the tibial nerve were ligated in all animals except for those in the immediate repair group. The experimental groups underwent denervation with nerve protection of peroneal nerve (mixed protection) or sural nerve (sensory protection). The experimental and unprotected groups had a stage II surgery in which the trimmed proximal and distal tibial nerve stumps were sutured together. After 3 months of recovery, electrophysiological, histological and morphometric parameters were assessed. It was detected that the significant muscle atrophy and a good preserved structure of the muscle were observed in the unprotected and protective experimental groups, respectively. Significantly fewer numbers of regenerated myelinated axons were observed in the sensory-protected group. Enhanced recovery in the mixed protection group was indicated by the results of the muscle contraction force tests, regenerated myelinated fiber, and the results of the histological analysis. Our results suggest that early axons protection by mixed nerve may complement sensory axons which are required for promoting functional recovery of the denervated muscle natively innervated by mixed nerve.     

MicroRNA-217 Promotes Angiogenesis of Human Cytomegalovirus-Infected Endothelial Cells through Downregulation of SIRT1 and FOXO3A

Human cytomegalovirus(HCMV) infection has been shown to contribute to vascular disease through the induction of angiogenesis. However, the role of microRNA in angiogenesis induced by HCMV infection remains unclear. The present study was thus designed to explore the potential effect of miR-1217 on angiogenesis and to disclose the underlying mechanism in endothelial cells. We found that HCMV infection of endothelial cells(ECs) enhanced expression of miR-217 and reduced SIRT1 and FOXO3A protein level in 24 hours post infection(hpi). Transfection of miR-217 inhibitor not only depressed cellular migration and tube formation induced by HCMV infection, but also enhanced SIRT1 and FOXO3A protein expression. Additionally, luciferase assay confirmed that miR-217 directly targeted FOXO3A mRNA 3`UTR. Furthermore, pretreatment with resveratrol depressed motility and tube formation of HCMV-infected ECs, which could be reversed by SIRT1 siRNA. Similarly, delivery of FOXO3A overexpression lentivirus suppressed proliferative rate, migration and tube formation of HCMV-infected ECs, which reversed by transfection of FOXO3A siRNA. In summary, HCMV infection of endothelial cells induces angiogenesis by both of miR-217/SIRT1 and miR-217/FOXO3A axis.     

PI3K/Akt Signal Pathway Involved in the Cognitive Impairment Caused by Chronic Cerebral Hypoperfusion in Rats

Chronic cerebral hypoperfusion (CCH) is a common pathophysiological state that usually occurs in conditions such as vascular dementia and Alzheimer''s disease, both of which are characterized by cognitive impairment. In previous studies we found that learning capacity and memory were gradually impaired with CCH, which altered the expression of synaptophysin, microtubule associated protein-2, growth associated protein-43, brain-derived neurotrophic factor, nerve growth factor, N-methyl-D-aspartate receptor subunit 1, cAMP response element-binding protein and tau hyperphosphorylation in the hippocampus. However, the molecular basis of cognitive impairment in CCH remains obscure. Here we explore the hypothesis that the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signal pathway is involved in this type of cognitive impairment. In order to determine if the expression of PI3K, Akt and phosphorylated Akt (p-Akt) proteins are altered at different stages of CCH with differing levels of cognitive impairment. we performed permanent, bilateral occlusion of the common carotid arteries (2-VO) to induce CCH. Adult male SD rats were randomly divided into sham-operated group, 2-VO 1 week group, 2-VO 4 weeks group and 2-VO 8 weeks group. Behavior tests were utilized to assess cognitive abilities, while western blots were utilized to evaluate protein expression. Rats in the 2-VO groups spent less time exploring novel objects than those in the sham-operated group, and the discrimination ratio of the 2-VO 8 weeks group and the sham-operated group were higher than chance (0.50). Escape latencies in the Morris water maze task in the 2-VO 1 week group were longer than those in the sham-operated group on day 4 and day 5, while escape latencies in the 2-VO 4 weeks group were longer than those in the sham-operated group from day 3 to day 5. Escape latencies in 2-VO 8 weeks group were longer than those in the sham-operated group from day 2 to day 5. NE (northeast) square swimming times in the 2-VO 1 week group, 2-VO 4 weeks group and 2-VO 8 weeks group were shorter than that in the sham-operated group. Western blotting showed that the PI3K expression in the 2-VO 1 week group was lower than that in sham-operated group, while p-Akt expression in the 2-VO 8 weeks group was higher than that in the sham-operated group. There was a linear relationship between the PI3K expression and the discrimination ratio, as well as a linear relationship between the PI3K and NE square swimming time. Thus, we propose that the PI3K/Akt signal pathway is an important cell pathway that is associated with the cognitive impairment following CCH.     

Suppression of E. multilocularis Hydatid Cysts after Ionizing Radiation Exposure

Background

Heavy-ion therapy has an advantage over conventional radiotherapy due to its superb biological effectiveness and dose conformity in cancer therapy. It could be a potential alternate approach for hydatid cyst treatment. However, there is no information currently available on the cellular and molecular basis for heavy-ion irradiation induced cell death in cystic echinococcosis.

Methododology/Principal Findings

LD50 was scored by protoscolex death. Cellular and ultrastructural changes within the parasite were studied by light and electron microscopy, mitochondrial DNA (mtDNA) damage and copy number were measured by QPCR, and apoptosis was determined by caspase 3 expression and caspase 3 activity. Ionizing radiation induced sparse cytoplasm, disorganized and clumped organelles, large vacuoles and devoid of villi. The initial mtDNA damage caused by ionizing radiation increased in a dose-dependent manner. The kinetic of DNA repair was slower after carbon-ion radiation than that after X-rays radiation. High dose carbon-ion radiation caused irreversible mtDNA degradation. Cysts apoptosis was pronounced after radiation. Carbon-ion radiation was more effective to suppress hydatid cysts than X-rays.

Conclusions

These studies provide a framework to the evaluation of attenuation effect of heavy-ion radiation on cystic echinococcosis in vitro. Carbon-ion radiation is more effective to suppress E. multilocularis than X-rays.