Genetic and physical fine mapping of a multilocular gene Bjln1 in Brassica juncea to a 208-kb region

Most of the germplasm resources in Brassica juncea produce silique with only two locules, whereas a few varieties can produce silique with three or four locules. The increase in locule number in B. juncea has been shown to cause an increase in the number of seeds per silique, resulting in an increase in the yield per plant. Thus, the development of high-locule-number varieties may be an effective way of improving the yield of B. juncea. Duoshi, a B. juncea landrace originating from the Qinghai–Tibetan plateau, produces silique with 3–4 locules. Genetic analysis has shown that the high-locule-number trait in Duoshi is determined by two recessive genes, tentatively designated as Bjln1 and Bjln2. For fine mapping of the Bjln1 gene, a BC3 population was developed from the cross between Duoshi (multilocular parent) and Xinjie (bilocular parent). Using a combination of amplified fragment length polymorphism (AFLP) and bulked segregant analysis, only two AFLP markers linked to Bjln1 were identified. Preliminary linkage analysis showed that the two AFLP markers were located on the same side of Bjln1. Blast analysis revealed that the sequences of the two AFLP markers had homologues on Scaffold000019 at the bottom of B. rapa A7. Using the results of linkage analysis and BlastN searches, simple sequence repeat (SSR) markers were subsequently developed based on the sequence information from B. rapa A7. Seven SSR markers were eventually identified, of which ln 8 was co-segregated with Bjln1. ln 7 and ln 9, the closest flanking markers, were mapped at 2.0 and 0.4 cM distant from the Bjln1 gene, respectively. The SSR markers were cloned, sequenced and mapped on A7 of B. rapa (corresponding to J7 in the A genome of B. juncea). The two closest flanking markers, ln 7 and ln 9, were mapped within a 208-kb genomic region on B. rapa A7, in which the Bjln1 gene might be included. The present study may facilitate cloning of the Bjln1 gene as well as the selection process for developing multilocular varieties in B. juncea by marker-assisted selection and genetic engineering.     

Patients' & Healthcare Professionals' Values Regarding True- & False-Positive Diagnosis when Colorectal Cancer Screening by CT Colonography: Discrete Choice Experiment

Purpose

To establish the relative weighting given by patients and healthcare professionals to gains in diagnostic sensitivity versus loss of specificity when using CT colonography (CTC) for colorectal cancer screening.

Materials and Methods

Following ethical approval and informed consent, 75 patients and 50 healthcare professionals undertook a discrete choice experiment in which they chose between “standard” CTC and “enhanced” CTC that raised diagnostic sensitivity 10% for either cancer or polyps in exchange for varying levels of specificity. We established the relative increase in false-positive diagnoses participants traded for an increase in true-positive diagnoses.

Results

Data from 122 participants were analysed. There were 30 (25%) non-traders for the cancer scenario and 20 (16%) for the polyp scenario. For cancer, the 10% gain in sensitivity was traded up to a median 45% (IQR 25 to >85) drop in specificity, equating to 2250 (IQR 1250 to >4250) additional false-positives per additional true-positive cancer, at 0.2% prevalence. For polyps, the figure was 15% (IQR 7.5 to 55), equating to 6 (IQR 3 to 22) additional false-positives per additional true-positive polyp, at 25% prevalence. Tipping points were significantly higher for patients than professionals for both cancer (85 vs 25, p<0.001) and polyps (55 vs 15, p<0.001). Patients were willing to pay significantly more for increased sensitivity for cancer (p = 0.021).

Conclusion

When screening for colorectal cancer, patients and professionals believe gains in true-positive diagnoses are worth much more than the negative consequences of a corresponding rise in false-positives. Evaluation of screening tests should account for this.     

Bradykinin Preconditioning Improves Therapeutic Potential of Human Endothelial Progenitor Cells in Infarcted Myocardium

Objectives

Stem cell preconditioning (PC) is a powerful approach in reducing cell death after transplantation. We hypothesized that PC human endothelial progenitor cells (hEPCs) with bradykinin (BK) enhance cell survival, inhibit apoptosis and repair the infarcted myocardium.

Methods

The hEPCs were preconditioned with or without BK. The hEPCs apoptosis induced by hypoxia along with serum deprivation was determined by annexin V-fluorescein isothiocyanate/ propidium iodide staining. Cleaved caspase-3, Akt and eNOS expressions were determined by Western blots. Caspase-3 activity and vascular endothelial growth factor (VEGF) levels were assessed in hEPCs. For in vivo studies, the survival and cardiomyocytes apoptosis of transplanted hEPCs were assessed using 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindodi- carbocyanine,4-chlorobenzenesul-fonate salt labeled hEPCs and TUNEL staining. Infarct size and cardiac function were measured at 10 days after transplantation, and the survival of transplanted hEPCs were visualized using near-infrared optical imaging.

Results

In vitro data showed a marked suppression in cell apoptosis following BK PC. The PC reduced caspase-3 activation, increased the Akt, eNOS phosphorylation and VEGF levels. In vivo data in preconditioned group showed a robust cell anti-apoptosis, reduction in infarct size, and significant improvement in cardiac function. The effects of BK PC were abrogated by the B2 receptor antagonist HOE140, the Akt and eNOS antagonists LY294002 and L-NAME, respectively.

Conclusions

The activation of B2 receptor-dependent PI3K/Akt/eNOS pathway by BK PC promotes VEGF secretion, hEPC survival and inhibits apoptosis, thereby improving cardiac function in vivo. The BK PC hEPC transplantation for stem cell-based therapies is a novel approach that has potential for clinical used.     

Lipidomics Reveals Early Metabolic Changes in Subjects with Schizophrenia: Effects of Atypical Antipsychotics

There is a critical need for mapping early metabolic changes in schizophrenia to capture failures in regulation of biochemical pathways and networks. This information could provide valuable insights about disease mechanisms, trajectory of disease progression, and diagnostic biomarkers. We used a lipidomics platform to measure individual lipid species in 20 drug-naïve patients with a first episode of schizophrenia (FE group), 20 patients with chronic schizophrenia that had not adhered to prescribed medications (RE group), and 29 race-matched control subjects without schizophrenia. Lipid metabolic profiles were evaluated and compared between study groups and within groups before and after treatment with atypical antipsychotics, risperidone and aripiprazole. Finally, we mapped lipid profiles to n3 and n6 fatty acid synthesis pathways to elucidate which enzymes might be affected by disease and treatment. Compared to controls, the FE group showed significant down-regulation of several n3 polyunsaturated fatty acids (PUFAs), including 20:5n3, 22:5n3, and 22:6n3 within the phosphatidylcholine and phosphatidylethanolamine lipid classes. Differences between FE and controls were only observed in the n3 class PUFAs; no differences where noted in n6 class PUFAs. The RE group was not significantly different from controls, although some compositional differences within PUFAs were noted. Drug treatment was able to correct the aberrant PUFA levels noted in FE patients, but changes in re patients were not corrective. Treatment caused increases in both n3 and n6 class lipids. These results supported the hypothesis that phospholipid n3 fatty acid deficits are present early in the course of schizophrenia and tend not to persist throughout its course. These changes in lipid metabolism could indicate a metabolic vulnerability in patients with schizophrenia that occurs early in development of the disease.     

A Preliminary Study of the Heating Effect of the Tibetan Plateau

The immense and towering Tibetan Plateau acts as a heating source and, thus, deeply shapes the climate of the Eurasian continent and even the whole world. However, due to the scarcity of meteorological observation stations and very limited climatic data, little is quantitatively known about the heating effect of the plateau and its implications. This paper firstly collects climate data (2001–2007) from 109 observation stations and MODIS-based estimated monthly mean temperature data in the plateau and the neighboring Sichuan Basin, and conducts correlation and simple linear regression to reveal the altitudinal pattern of temperature. Then, according to the linear relationships of temperature and altitude for each month, it compares air temperature differences on the same elevation between the main plateau and surrounding mountains and the Sichuan Basin so as to quantify the heating effect and discuss its implication on timberline of the plateau. The results show that: 1) the heating effect of the plateau is significant. The temperature of the main plateau area was higher than that of free air on the same elevation above the neighboring areas; on the elevation of 4500 m (the main plateau), temperature is 1–6°C higher in the main Plateau than over the Sichuan Basin for different months and 5.9–10.7°C higher than in the Qilian Mountains in the northeastern corner of the plateau. 2) Even at altitudes of 5000–6000 m in the main Plateau, there are 4 months with a mean temperature above 0°C. The mean temperature of the warmest month (July) can reach 10°C at about 4600–4700 m. This may help explain why the highest timberline in the northern hemisphere is on the southeastern Tibetan Plateau.     

Triterpenoids from the Stem Bark of Melia toosendan and Determination of Their Absolute Configurations at C(24)

Six new triterpenoids, meliasenins S–X ( 1 – 6 , resp.), were isolated from the stem bark of Melia toosendan. Their structures were elucidated by mass spectrometry, NMR experiments, and comparison with the known compounds. Particularly, the absolute configuration at C(24) in new compounds was determined through their CD spectra of the [Pr(FOD)₃] complex (fod=1,1,1,2,2,3,3,7,7,7‐decafluoroheptane‐4,6‐dione) in CCl₄, as well as by using Mosher's method.     

Fundamental differences in promoter CpG island DNA hypermethylation between human cancer and genetically engineered mouse models of cancer

Genetic and epigenetic alterations are essential for the initiation and progression of human cancer. We previously reported that primary human medulloblastomas showed extensive cancer-specific CpG island DNA hypermethylation in critical developmental pathways. To determine whether genetically engineered mouse models (GEMMs) of medulloblastoma have comparable epigenetic changes, we assessed genome-wide DNA methylation in three mouse models of medulloblastoma. In contrast to human samples, very few loci with cancer-specific DNA hypermethylation were detected, and in almost all cases the degree of methylation was relatively modest compared with the dense hypermethylation in the human cancers. To determine if this finding was common to other GEMMs, we examined a Burkitt lymphoma and breast cancer model and did not detect promoter CpG island DNA hypermethylation, suggesting that human cancers and at least some GEMMs are fundamentally different with respect to this epigenetic modification. These findings provide an opportunity to both better understand the mechanism of aberrant DNA methylation in human cancer and construct better GEMMs to serve as preclinical platforms for therapy development.     

Rapid apoplastic pH measurement in Arabidopsis leaves using a fluorescent dye

In plants, the extracellular space (apoplast) is one of the main places where exchange of molecules occurs between cells. Not only is this compartment involved in the storage of multiple metabolites and ions, including calcium and protons, but it also plays a role in the transmission of signaling molecules for cell-to-cell communication. It has recently been shown multiple times that these two aspects are linked and can influence each other. In particular, apoplast pH was shown as a primary regulator of auxin (IAA) transport in Arabidopsis thaliana. To prove the role of apoplastic pH, we have developed a protocol for apoplastic fluid extraction from Arabidopsis leaves, followed by pH determination using the 8-hydroxypyrene-1,3,6-trisulfonic acid (HPTS) fluorescent dye. This technique successfully allows one to monitor apoplastic pH variations among different plant lines and to link changes in apoplastic pH to cellular responses in the plant.