Autologous Bone-Marrow Mesenchymal Stem Cell Implantation and Endothelial Function in a Rabbit Ischemic Limb Model

Background

The purpose of this study was to determine whether autologous mesenchymal stem cells (MSCs) implantation improves endothelial dysfunction in a rabbit ischemic limb model.

Methods

We evaluated the effect of MSC implantation on limb blood flow (LBF) responses to acetylcholine (ACh), an endothelium-dependent vasodilator, and sodium nitroprusside (SNP), an endothelium-independent vasodilator, in rabbits with limb ischemia in which cultured MSCs were implanted (n = 20) or saline was injected as a control group (n = 20). LBF was measured using an electromagnetic flowmeter. A total of 10⁶ MSCs were implanted into each ischemic limb.

Results

Histological sections of ischemic muscle showed that capillary index (capillary/muscle fiber) was greater in the MSC implantation group than in the control group. Laser Doppler blood perfusion index was significantly increased in the MSC implantation group compared with that in the control group. LBF response to ACh was greater in the MSC group than in the control group. After administration of N^G-nitro-L-arginine, a nitric oxide synthase inhibitor, LBF response to ACh was similar in the MSC implantation group and control group. Vasodilatory effects of SNP in the two groups were similar.

Conclusions

These findings suggest that MSC implantation induces angiogenesis and augments endothelium-dependent vasodilation in a rabbit ischemic model through an increase in nitric oxide production.     

Spatial covariation of fish population vital rates in a stream network

Animal populations are spatially structured in heterogeneous landscapes, in which local patches with differing vital rates are connected by dispersal of individuals to varying degrees. Although there is evidence that vital rates differ among local populations, much less is understood about how vital rates covary among local patches in spatially heterogeneous landscapes. In this study, we conducted a nine-year annual mark–recapture survey to characterize spatial covariation of survival and growth for two Japanese native salmonids, white-spotted charr Salvelinus leucomaenis japonicus and red-spotted masu salmon Oncorhynchus masou ishikawae, in a headwater stream network composed of distinctly different tributary and mainstem habitats. Spatial structure of survival and growth differed by species and age class, but results provided support for negative covariation between vital rates, where survival was higher in the tributary habitat but growth was higher in the mainstem habitat. Thus, neither habitat was apparently more important than the other, and local habitats with complementary vital rates may make this spatially structured population less vulnerable to environmental change (i.e. portfolio effect). Despite the spatial structure of vital rates and possibilities that fish can exploit spatially distributed resources, movement of fish was limited due partly to a series of low-head dams that prevented upstream movement of fish in the study area. This study shows that spatial structure of vital rates can be complex and depend on species and age class, and this knowledge is likely paramount to elucidating dynamics of spatially structured populations.     

Initial Contact of Glioblastoma Cells with Existing Normal Brain Endothelial Cells Strengthen the Barrier Function via Fibroblast Growth Factor 2 Secretion: A New In Vitro Blood–Brain Barrier Model

Glioblastoma multiforme (GBM) cells invade along the existing normal capillaries in brain. Normal capillary endothelial cells function as the blood–brain barrier (BBB) that limits permeability of chemicals into the brain. To investigate whether GBM cells modulate the BBB function of normal endothelial cells, we developed a new in vitro BBB model with primary cultures of rat brain endothelial cells (RBECs), pericytes, and astrocytes. Cells were plated on a membrane with 8 μm pores, either as a monolayer or as a BBB model with triple layer culture. The BBB model consisted of RBEC on the luminal side as a bottom, and pericytes and astrocytes on the abluminal side as a top of the chamber. Human GBM cell line, LN-18 cells, or lung cancer cell line, NCI-H1299 cells, placed on either the RBEC monolayer or the BBB model increased the transendothelial electrical resistance (TEER) values against the model, which peaked within 72 h after the tumor cell application. The TEER value gradually returned to baseline with LN-18 cells, whereas the value quickly dropped to the baseline in 24 h with NCI-H1299 cells. NCI-H1299 cells invaded into the RBEC layer through the membrane, but LN-18 cells did not. Fibroblast growth factor 2 (FGF-2) strengthens the endothelial cell BBB function by increased occludin and ZO-1 expression. In our model, LN-18 and NCI-H1299 cells secreted FGF-2, and a neutralization antibody to FGF-2 inhibited LN-18 cells enhanced BBB function. These results suggest that FGF-2 would be a novel therapeutic target for GBM in the perivascular invasive front.     

Possible contribution of Bradyrhizobium on nitrogen fixation in sweet potatoes

Aims

Sweet potato (Ipomoea batatas) is known for its ability to grow under nitrogen-limited conditions. To clarify the possible contribution of biological nitrogen fixation, we tried to isolate and identify diazotrophic bacteria from sweet potatoes.

Methods

By using cultivation technique, we isolated putative endophytes, which possess nifH genes, from surface-sterilized sweet potatoes. Their nitrogen-fixing abilities were demonstrated by the acetylene reduction assay in a semi-solid malate medium and sweet potato extracts. We also examined the colonization of an isolated strain (AT1) in sweet potatoes and their influence on growth and nitrogen fixation in plants as assessed by an acetylene reduction assay and ¹⁵N-isotope dilution technique.

Results

The isolates were identified as strains of Bradyrhizobium sp. AT1, Paenibacillus sp. AS2 and Pseudomonas sp. T16 based on their 16S rRNA gene sequences. They showed acetylene reduction activity (ARA) in the semi-solid malate medium. Among them, B. sp. AT1 showed ARA in sweet potato extracts under micro-aerobic conditions whereas both P. sp. AS2 and P. sp. T16 showed no ARA. The inoculation of B. sp. AT1 to the sweet potatoes resulted in increases in the fresh weights and detection of ARA in the inoculated plants. Moreover, the reduction of ¹⁵N atom % was observed in the inoculated plants compared to uninoculated controls.

Conclusions

B. sp. AT1 actively expresses nitrogenase activity in sweet potatoes and may contribute to the nitrogen nutrition of host plants.     

Characterization of Variant Creutzfeldt-Jakob Disease Prions in Prion Protein-humanized Mice Carrying Distinct Codon 129 Genotypes

To date, all clinical variant Creutzfeldt-Jakob disease (vCJD) patients are homozygous for methionine at polymorphic codon 129 (129M/M) of the prion protein (PrP) gene. However, the appearance of asymptomatic secondary vCJD infection in individuals with a PRNP codon 129 genotype other than M/M and transmission studies using animal models have raised the concern that all humans might be susceptible to vCJD prions, especially via secondary infection. To reevaluate this possibility and to analyze in detail the transmission properties of vCJD prions to transgenic animals carrying distinct codon 129 genotype, we performed intracerebral inoculation of vCJD prions to humanized knock-in mice carrying all possible codon 129 genotypes (129M/M, 129M/V, or 129V/V). All humanized knock-in mouse lines were susceptible to vCJD infection, although the attack rate gradually decreased from 129M/M to 129M/V and to 129V/V. The amount of PrP deposition including florid/amyloid plaques in the brain also gradually decreased from 129M/M to 129M/V and to 129V/V. The biochemical properties of protease-resistant abnormal PrP in the brain and transmissibility of these humanized mouse-passaged vCJD prions upon subpassage into knock-in mice expressing bovine PrP were not affected by the codon 129 genotype. These results indicate that individuals with the 129V/V genotype may be more susceptible to secondary vCJD infection than expected and may lack the neuropathological characteristics observed in vCJD patients with the 129M/M genotype. Besides the molecular typing of protease-resistant PrP in the brain, transmission studies using knock-in mice carrying bovine PrP may aid the differential diagnosis of secondary vCJD infection, especially in individuals with the 129V/V genotype.     

Nitrogen Fixation and Allantoin Formation in Soybean Plants

The activity of nitrogenase and the concentration of ammonia and allantoin (+ allantoic acid) in root nodules were measured throughout the growth period of soybean plants. Nitrogenase activity measured by acetylene reduction increased with plant growth and reached a maximum level at the flowering period. The level of ammonia and allantoin concentration in nodules was parallel with increased nitrogenase activity. At the late reproductive stage (pod-forming period), nitrogenase activity showed a marked decrease, but the ammonia and allantoin in the nodules remained at a constant level. Detached nodules from 56 day-old soybean plants were exposed to ¹⁵N₂ gas, and the distribution of ¹⁵N among nitrogen compounds was investigated. Enrichment of ¹⁵N in allantoin and allantoic acid reached a fairly high level after 90 min of nitrogen fixation; ca. 22% of ¹⁵N in acid-soluble nitrogen compounds was incorporated into allantoin + allantoic acid. In contrast, enrichment of ¹⁵N in amide nitrogen was relatively low. No significant ¹⁵N was detected in the RNA fraction. The data suggested that ureide formation in nitrogen-fixing root nodules did not take place through the breakdown of nucleic acids, but directly associated with the assimilating system of biologically fixed nitrogen.     

Localization of 9- and 13-oxo-octadecadienoic acids in tomato fruit

We previously reported that the two peroxisome proliferator-activated receptor-α agonists, 9- and 13-oxo-octadecadienoic acids (oxo-ODAs), were found in the tomato fruit. However, their localization remains unknown. Herein, we showed that oxo-ODAs localize primarily in the fruit peel and their amount increases after the homogenization of the tomato fruit.     

Progress and prospects for the discovery of biomarkers for gastric cancer: a focus on proteomics

Introduction: Patient outcomes from gastric cancer vary due to the complexity of stomach carcinogenesis. Recent research using proteomic technologies has targeted components of all of these systems in order to develop biomarkers to aid the early diagnosis of gastric cancer and to assist in prognostic stratification.

Areas covered: This review is comprised of evidence obtained from literature searches from PubMed. It covers the evidence of diagnostic, prognostic, and predictive biomarkers for gastric cancer using proteomic technologies, and provides up-to-date references.

Expert commentary: The proteomic technologies have not only enabled the screening of a large number of samples, but also enabled the identification of diagnostic, prognostic and predictive biomarkers for gastric cancer. While major challenges still remain, to date, proteomic studies in gastric cancer have provided a wealth of information in revealing proteome alterations associated with the disease.