Systematic study of human long intergenic non-coding RNAs and their impact on cancer

The functional impact of several long intergenic non-coding RNAs (lincRNAs) has been characterized in previous studies. However, it is difficult to identify lincRNAs on a large-scale and to ascertain their functions or predict their structures in laboratory experiments because of the diversity, lack of knowledge and specificity of expression of lincRNAs. Furthermore, although there are a few well-characterized examples of lincRNAs associated with cancers, these are just the tip of the iceberg owing to the complexity of cancer. Here, by combining RNA-Seq data from several kinds of human cell lines with chromatin-state maps and human expressed sequence tags, we successfully identified more than 3000 human lincRNAs, most of which were new ones. Subsequently, we predicted the functions of 105 lincRNAs based on a coding-non-coding gene co-expression network. Finally, we propose a genetic mediator and key regulator model to unveil the subtle relationships between lincRNAs and lung cancer. Twelve lincRNAs may be principal players in lung tumorigenesis. The present study combines large-scale identification and functional prediction of human lincRNAs, and is a pioneering work in characterizing cancer-associated lincRNAs by bioinformatics.     

Optimization-driven identification of genetic perturbations accelerates the convergence of model parameters in ensemble modeling of metabolic networks

The ensemble modeling (EM) approach has shown promise in capturing kinetic and regulatory effects in the modeling of metabolic networks. Efficacy of the EM procedure relies on the identification of model parameterizations that adequately describe all observed metabolic phenotypes upon perturbation. In this study, we propose an optimization-based algorithm for the systematic identification of genetic/enzyme perturbations to maximally reduce the number of models retained in the ensemble after each round of model screening. The key premise here is to design perturbations that will maximally scatter the predicted steady-state fluxes over the ensemble parameterizations. We demonstrate the applicability of this procedure for an Escherichia coli metabolic model of central metabolism by successively identifying single, double, and triple enzyme perturbations that cause the maximum degree of flux separation between models in the ensemble. Results revealed that optimal perturbations are not always located close to reaction(s) whose fluxes are measured, especially when multiple perturbations are considered. In addition, there appears to be a maximum number of simultaneous perturbations beyond which no appreciable increase in the divergence of flux predictions is achieved. Overall, this study provides a systematic way of optimally designing genetic perturbations for populating the ensemble of models with relevant model parameterizations.     

Molecular Epidemiology of Porcine Cytomegalovirus (PCMV) in Sichuan Province,China: 2010–2012

Porcine cytomegalovirus (PCMV) is an immunosuppressive virus that mainly inhibits the immune function of the macrophage and T-cell lymphatic systems, and has caused huge economic losses to the porcine breeding industry. Molecular epidemiological investigation of PCMV is important for prevention and treatment, and this study is the first such investigation in Sichuan Province, Southwest China. A PCMV positive infection rate of 84.4% (865/1025) confirmed that PCMV is widely distributed in Sichuan Province. A phylogenetic tree was constructed based on the PCMV glycoprotein B gene (gB) nucleotide and amino acid sequences from 24 novel Sichuan isolates and 18 other PCMV gB sequences from Genbank. PCMV does not appear to have evolved into different serotypes, and two distinct sequence groups were identified (A and B). However, whether PCMV from this region has evolved into different genotypes requires further research. Analysis of the amino acid sequences confirmed the conservation of gB, but amino acid substitutions in the major epitope region have caused antigenic drift, which may have altered the immunogenicity of PCMV.     

Adaptation of Phenylalanine and Tyrosine Catabolic Pathway to Hibernation in Bats

Some mammals hibernate in response to harsh environments. Although hibernating mammals may metabolize proteins, the nitrogen metabolic pathways commonly activated during hibernation are not fully characterized. In contrast to the hypothesis of amino acid preservation, we found evidence of amino acid metabolism as three of five key enzymes, including phenylalanine hydroxylase (PAH), homogentisate 1,2-dioxygenase (HGD), fumarylacetoacetase (FAH), involved in phenylalanine and tyrosine catabolism were co-upregulated during hibernation in two distantly related species of bats, Myotis ricketti and Rhinolophus ferrumequinum. In addition, the levels of phenylalanine in the livers of these bats were significantly decreased during hibernation. Because phenylalanine and tyrosine are both glucogenic and ketogenic, these results indicate the role of this catabolic pathway in energy supply. Since any deficiency in the catabolism of these two amino acids can cause accumulations of toxic metabolites, these results also suggest the detoxification role of these enzymes during hibernation. A higher selective constraint on PAH, HPD, and HGD in hibernators than in non-hibernators was observed, and hibernators had more conserved amino acid residues in each of these enzymes than non-hibernators. These conserved amino acid residues are mostly located in positions critical for the structure and activity of the enzymes. Taken together, results of this work provide novel insights in nitrogen metabolism and removal of harmful metabolites during bat hibernation.     

Clinical Significance in Oral Cavity Squamous Cell Carcinoma of Pathogenic Somatic Mitochondrial Mutations

Somatic mutations affecting the mitochondrial DNA (mtDNA) have been frequently observed in human cancers and proposed as important oncological biomarkers. However, the clinical significance of mtDNA mutations in cancer remains unclear. This study was therefore performed to explore the possible clinical use in assessing oral squamous cell carcinoma (OSCC) of pathogenic mtDNA mutations. The entire mitochondrial genome of 300 OSCC with their matched control DNAs was screened by direct sequencing and criteria were set to define a pathogenic somatic mutation. The patients'' TP53 R72P genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism. The relationships between pathogenic somatic mutations, clinicopathogical features, TP53 R72P genotype and clinical prognosis were analyzed. Overall, 645 somatic mtDNA mutations were identified and 91 of these mutations were defined as pathogenic. About one quarter (74/300) of the OSCC tumor samples contained pathogenic mutations. Individuals with the TP53 R allele had a higher frequency of pathogenic somatic mutation than those with the PP genotype. Kaplan-Meier analysis indicated that TP53 R allele patients with pathogenic somatic mutations demonstrated a significant association with a poorer disease-free survival than other individuals (HR = 1.71; 95% CI, 1.15–2.57; p = 0.009) and this phenomenon still existed after adjusting for mtDNA haplogroup, tumor stage with treatment regimens, differentiation and age at diagnosis (HR = 1.59; 95% CI, 1.06–2.40; p = 0.03). Subgroup analyses showed that this phenomenon was limited to patients who received adjuvant radiotherapy/chemo-radiotherapy after surgery. The results strongly indicated that pathogenic mtDNA mutations are a potential prognostic marker for OSCCs. Furthermore, functional mitochondria may play an active role in cancer development and the patient''s response to radiotherapy/chemo-radiotherapy.     

RhoA/Rho-Kinase and Nitric Oxide in Vascular Reactivity in Rats with Endotoxaemia

RhoA/Rho-kinase (RhoA/ROK) pathway promotes vasoconstriction by calcium sensitivity mechanism. LPS causes nitric oxide (NO) overproduction to induce vascular hyporeactivity. Thus, we tried to examine the role of RhoA/ROK and NO in the regulation of vascular reactivity in different time-point of endotoxaemia. Male Wistar rats were intravenously infused for 10 min with saline or E. coli endotoxin (lipopolysaccharide, LPS, 10 mg/kg) and divided to five groups (n = 8 in each group): (i) Control, sacrificed at 6 h after saline infusion; (ii) LPS1h, sacrificed at 1 h after LPS infusion; (iii) LPS2h, sacrificed at 2 h after LPS infusion; (iv) LPS4h, sacrificed at 4 h after LPS infusion; and (v) LPS6h, sacrificed at 6 h after LPS infusion. LPS1h and LPS2h were regarded as early endotoxaemia, whereas LPS4h and LPS6h were regarded as late endotoxaemia. Indeed, our results showed that LPS reproduced a biphasic hypotension and sustained vascular hyporeactivity to noradrenaline (NA) in vivo. Interestingly, this hyporeactivity did not occur in ex vivo during early endotoxaemia. This could be due to increases of aortic RhoA activity (n = 5, P<0.05) and myosin phosphatase targeting subunit 1 phosphorylation (n = 3, P<0.05). In addition, pressor response to NA and vascular reactivity in early endotoxaemia were inhibited by ROK inhibitor, Y27632. Furthermore, plasma bradykinin was increased at 10 min (24.6±13.7 ng/mL, n = 5, P<0.05) and aortic endothelial NO synthase expression was increased at 1 h (+200%. n = 3, P<0.05) after LPS. In late endotoxaemia, the vascular hyporeactivity was associated with aortic inducible NO synthase expression (n = 3, P<0.05) and an increased serum NO level (n = 8, P<0.05). Thus, an increased RhoA activity could compensate vascular hyporeactivity in early endotoxaemia, and the large NO production inhibiting RhoA activity would lead to vascular hyporeactivity eventually.     

Regional Cardiac Dysfunction and Dyssynchrony in a Murine Model of Afterload Stress

Small animal models of afterload stress have contributed much to our present understanding of the progression from hypertension to heart failure. High-sensitivity methods for phenotyping cardiac function in vivo, particular in the setting of compensated cardiac hypertrophy, may add new information regarding alterations in cardiac performance that can occur even during the earliest stages of exposure to pressure overload. We have developed an echocardiographic analytical method, based on speckle-tracking-based strain analyses, and used this tool to rapidly phenotype cardiac changes resulting from afterload stress in a small animal model. Adult mice were subjected to ascending aortic constriction, with and without subsequent reversal of the pressure gradient. In this model of compensated hypertrophic cardiac remodeling, conventional echocardiographic measurements did not detect changes in left ventricular (LV) function at the early time points examined. Strain analyses, however, revealed a decrement in basal longitudinal myofiber shortening that was induced by aortic constriction and improved following relief of the pressure gradient. Furthermore, we observed that pressure overload resulted in LV segmental dyssynchrony that was attenuated with return of the afterload to baseline levels. Herein, we describe the use of echocardiographic strain analyses for cardiac phenotyping in a mouse model of pressure overload. This method provides evidence of dyssynchrony and regional myocardial dysfunction that occurs early with compensatory hypertrophy, and improves following relief of aortic constriction. Importantly, these findings illustrate the utility of a rapid, non-invasive method for characterizing early cardiac dysfunction, not detectable by conventional echocardiography, following afterload stress.     

Quantification of Blood Flow in Internal Cerebral Artery by Optical Flow Method on Digital Subtraction Angiography in Comparison with Time-Of-Flight Magnetic Resonance Angiography

Objective

This study compared data on the blood flow velocity in the internal carotid artery, which was obtained using the optical flow method (OFM) with digital subtraction angiography (DSA) and the time-of-flight (TOF) technique using magnetic resonance angiography (MRA).

Materials and Methods

Images were obtained from 12 cerebrovascular patients who underwent both brain DSA and MRA imaging. The OFM was applied on the DSA images to determine the average blood flow velocity. The calculated results were compared with the values obtained from the TOF-MRA data. A linear fit was performed on the data and Bland-Altman plots were analyzed.

Results

The blood flow velocity was closely associated with vascular diseases. Color-coding of the OFM measurements were superimposed on to the DSA images, which quantitatively illustrated the relative flow in the vessels. The average blood flow velocity was calculated using OFM and DSA, which demonstrated a high correlation with the MRA measurements in the anterior-posterior (AP) view (R = 0.71). In contrast, the average blood flow velocity was low in the lateral view (R = 0.28). The consistency between the high and low blood velocity in the AP view was better compared to the lateral view. The blood flow velocity distribution in the AP view was statistically closer to the MRA measurement compared to the lateral view.

Conclusions

This study evaluated the correlation of blood flow measured using DSA and TOF-MRA in a small heterogeneous group of patients with cerebrovascular lesions. OFM with DSA imaging reveals hemodynamic information and TOF-MRA.     

Characterization of Tat Antibody Responses in Chinese Individuals Infected with HIV-1

HIV-1 Tat is an important regulatory protein involved in AIDS pathogenesis. However, the immunoprofiles of anti-Tat responses remain unclear. We analysed the immunoprofiles of the anti-Tat antibody responses and the neutralizing activities. Out of 326 HIV-1-seropositive individuals, 12.9% were positive for anti-Tat antibodies. We found six different immunological profiles of anti-Tat antibody responses: full-potential response, combined response, N-specific response, C-specific response, full-length Tat-specific response and Tat-related response. These responses represent two types of anti-Tat responses: the major complete response and the alternative C-prone response. A Tat-neutralizing activity is significantly higher in anti-Tat-seropositive samples than anti-Tat-negative or healthy blood-donor samples, and significantly correlates with the anti-Tat reactivities. The data here could contribute to a better understanding of the significance of anti-Tat responses in preventing HIV pathogenesis and could be useful for designing more effective vaccines in the future.