A New Approach Using the SYBR Green-Based Real-Time PCR Method for Detection of Soft Rot Pectobacterium odoriferum Associated with Kimchi Cabbage

Pectobacterium odoriferum is the primary causative agent in Kimchi cabbage soft-rot diseases. The pathogenic bacteria Pectobacterium genera are responsible for significant yield losses in crops. However, P. odoriferum shares a vast range of hosts with P. carotovorum, P. versatile, and P. brasiliense, and has similar biochemical, phenotypic, and genetic characteristics to these species. Therefore, it is essential to develop a P. odoriferum-specific diagnostic method for soft-rot disease because of the complicated diagnostic process and management as described above. Therefore, in this study, to select P. odoriferum-specific genes, species-specific genes were selected using the data of the P. odoriferum JK2.1 whole genome and similar bacterial species registered with NCBI. Thereafter, the specificity of the selected gene was tested through blast analysis. We identified novel species-specific genes to detect and quantify targeted P. odoriferum and designed specific primer sets targeting HAD family hydrolases. It was confirmed that the selected primer set formed a specific amplicon of 360 bp only in the DNA of P. odoriferum using 29 Pectobacterium species and related species. Furthermore, the population density of P. odoriferum can be estimated without genomic DNA extraction through SYBR Green-based real-time quantitative PCR using a primer set in plants. As a result, the newly developed diagnostic method enables rapid and accurate diagnosis and continuous monitoring of soft-rot disease in Kimchi cabbage without additional procedures from the plant tissue.     

Changes in the Pulmonary Function Test after Radioactive Iodine Treatment in Patients with Pulmonary Metastases of Differentiated Thyroid Cancer

ObjectivePulmonary function test (PFT) is a useful tool for an objective assessment of respiratory function. Impaired pulmonary function is critical for the survival and quality of life in patients with pulmonary metastases of solid cancers including thyroid cancer. This study aimed to evaluate clinical factors associated with severely impaired pulmonary function by serial assessment with PFT in patients with pulmonary metastasis of differentiated thyroid cancer (DTC) who received radioactive iodine treatment (RAIT).PatientsThis retrospective study enrolled 31 patients who underwent serial PFTs before and after RAIT for pulmonary metastasis of DTC. We evaluated the risk factors for severe impairment of pulmonary function.ResultsThe median age of the patients was 44.1 years and 18 of them were female patients. Severe impairment of pulmonary function was observed in five patients (16%) after a median of three RAITs (cumulative I-131 activity = 20.4 GBq). These patients were older and more frequently had mild impairment of baseline pulmonary function, respiratory symptoms, or progressive disease compared with patients with stable pulmonary function. Neither cumulative dose nor number of RAIT was associated with decreased pulmonary function. Coexisting pulmonary diseases, presence of respiratory symptoms, and metastatic disease progression were significantly associated with severe decrease in forced vital capacity during follow-up (p =.047, p =.011, and p =.021, respectively).ConclusionsPulmonary function was severely impaired during follow-up in some patients with pulmonary metastasis of DTC after a high-dose RAITs. Neither the number of RAIT nor the cumulative I-131 activity was associated with decreased pulmonary function. Serial PFT might be considered for some high-risk patients during follow-up.     

Efficacy of Intrauterine Bakri Balloon Tamponade in Cesarean Section for Placenta Previa Patients

Purpose

The aims of this study were to analyze the predictive factors for the use of intrauterine balloon insertion and to evaluate the efficacy and factors affecting failure of uterine tamponade with a Bakri balloon during cesarean section for abnormal placentation.

Methods

We reviewed the medical records of 137 patients who underwent elective cesarean section for placenta previa between July 2009 and March 2014. Cesarean section and Bakri balloon insertion were performed by a single qualified surgeon. The Bakri balloon was applied when blood loss during cesarean delivery exceeded 1,000 mL.

Results

Sixty-four patients (46.7%) required uterine balloon tamponade during cesarean section due to postpartum bleeding from the lower uterine segment, of whom 50 (78.1%) had placenta previa totalis. The overall success rate was 75% (48/64) for placenta previa patients. Previous cesarean section history, anterior placenta, peripartum platelet count, and disseminated intravascular coagulopathy all significantly differed according to balloon success or failure (all p<0.05). The drainage amount over 1 hour was 500 mL (20–1200 mL) in the balloon failure group and 60 mL (5–500 mL) in the balloon success group (p<0.01).

Conclusion

Intrauterine tamponade with a Bakri balloon is an adequate adjunct management for postpartum hemorrhage following cesarean section for placenta previa to preserve the uterus. This method is simple to apply, non-invasive, and inexpensive. However, possible factors related to failure of Bakri balloon tamponade for placenta previa patients such as prior cesarean section history, anterior placentation, thrombocytopenia, presence of DIC at the time of catheter insertion, and catheter drainage volume more than 500 mL within 1 hour of catheter placement should be recognized, and the next-line management should be prepared in advance.     

Modelling and optimal control of immune response of renal transplant recipients

We consider the increasingly important and highly complex immunological control problem: control of the dynamics of immunosuppression for organ transplant recipients. The goal in this problem is to maintain the delicate balance between over-suppression (where opportunistic latent viruses threaten the patient) and under-suppression (where rejection of the transplanted organ is probable). First, a mathematical model is formulated to describe the immune response to both viral infection and introduction of a donor kidney in a renal transplant recipient. Some numerical results are given to qualitatively validate and demonstrate that this initial model exhibits appropriate characteristics of primary infection and reactivation for immunosuppressed transplant recipients. In addition, we develop a computational framework for designing adaptive optimal treatment regimes with partial observations and low-frequency sampling, where the state estimates are obtained by solving a second deterministic optimal tracking problem. Numerical results are given to illustrate the feasibility of this method in obtaining optimal treatment regimes with a balance between under-suppression and over-suppression of the immune system.     

Genetic and epigenetic alterations of the NF2 gene in sporadic vestibular schwannomas

Background

Mutations in the neurofibromatosis type 2 (NF2) tumor-suppressor gene have been identified in not only NF2-related tumors but also sporadic vestibular schwannomas (VS). This study investigated the genetic and epigenetic alterations in tumors and blood from 30 Korean patients with sporadic VS and correlated these alterations with tumor behavior.

Methodology/Principal Findings

NF2 gene mutations were detected using PCR and direct DNA sequencing and three highly polymorphic microsatellite DNA markers were used to assess the loss of heterozygosity (LOH) from chromosome 22. Aberrant hypermethylation of the CpG island of the NF2 gene was also analyzed. The tumor size, the clinical growth index, and the proliferative activity assessed using the Ki-67 labeling index were evaluated. We found 18 mutations in 16 cases of 30 schwannomas (53%). The mutations included eight frameshift mutations, seven nonsense mutations, one in-frame deletion, one splicing donor site, and one missense mutation. Nine patients (30%) showed allelic loss. No patient had aberrant hypermethylation of the NF2 gene and correlation between NF2 genetic alterations and tumor behavior was not observed in this study.

Conclusions/Significance

The molecular genetic changes in sporadic VS identified here included mutations and allelic loss, but no aberrant hypermethylation of the NF2 gene was detected. In addition, no clear genotype/phenotype correlation was identified. Therefore, it is likely that other factors contribute to tumor formation and growth.     

Allithiamine Exerts Therapeutic Effects on Sepsis by Modulating Metabolic Flux during Dendritic Cell Activation

Recent studies have highlighted that early enhancement of the glycolytic pathway is a mode of maintaining the pro-inflammatory status of immune cells. Thiamine, a well-known co-activator of pyruvate dehydrogenase complex, a gatekeeping enzyme, shifts energy utilization of glucose from glycolysis to oxidative phosphorylation. Thus, we hypothesized that thiamine may modulate inflammation by alleviating metabolic shifts during immune cell activation. First, using allithiamine, which showed the most potent anti-inflammatory capacity among thiamine derivatives, we confirmed the inhibitory effects of allithiamine on the lipopolysaccharide (LPS)-induced pro-inflammatory cytokine production and maturation process in dendritic cells. We applied the LPS-induced sepsis model to examine whether allithiamine has a protective role in hyper-inflammatory status. We observed that allithiamine attenuated tissue damage and organ dysfunction during endotoxemia, even when the treatment was given after the early cytokine release. We assessed the changes in glucose metabolites during LPS-induced dendritic cell activation and found that allithiamine significantly inhibited glucose-driven citrate accumulation. We then examined the clinical implication of regulating metabolites during sepsis by performing a tail bleeding assay upon allithiamine treatment, which expands its capacity to hamper the coagulation process. Finally, we confirmed that the role of allithiamine in metabolic regulation is critical in exerting anti-inflammatory action by demonstrating its inhibitory effect upon mitochondrial citrate transporter activity. In conclusion, thiamine could be used as an alternative approach for controlling the immune response in patients with sepsis.     

Up-regulation of skeletal muscle LIM protein 1 gene by 25-hydroxycholesterol may mediate morphological changes of rat aortic smooth muscle cells

Changes in the expression level of the skeletal muscle LIM protein 1 (SLIM1) in cultured A10 cells were monitored in response to 25-hydroxycholesterol (25-HC), an oxidized form of cholesterol present in the oxidized low-density lipoproteins. The level of SLIM1 mRNA was elevated in a time- and concentration-dependent manner by treatment of 25-HC. Expressions of smooth muscle (SM) alpha-actin and calponin-1 (CNN-1), early markers for SMC differentiation, were also increased by the 25-HC treatments. Expressions of all three genes (SLIM1, SM alpha-actin and CNN-1) were simultaneously elevated in the cells treated with 9-cis retinoic acid (RA). On the other hand, the SLIM1 expression induced by the 25-HC or 9-cis RA (as well as SM alpha-actin and CNN-1) was decreased by the treatment of 15d-PGJ2. Since the 25-HC, 9-cis RA and 15d-PGJ2 were ligands for the LXR, RXRalpha and PPARgamma respectively, there might be a functional positive cross-talk between LXR and RXRalpha pathways and a negative cross-talk between PPARgamma and LXR and/or RXRalpha pathways in the regulation of SLIM1 expression. The cells stably transfected with the expressional vector for SLIM1 also showed an elevation in the levels of SM alpha-actin and CNN-1. In addition, an over-production of SLIM1 in the cells resulted in a change in the cell-shape into a spindle-like form, which is identical to that observed after a prolonged treatment of the cells with cholesterol.     

Isolation of phosphopeptides by pI-difference-based electrophoresis

Effective proteomics studies of protein phosphorylation require an efficient enrichment method for phosphopeptides, which remains a challenge. Here, we describe the discovery of pI differences between methylated phosphopeptides (typically <7.4) and methylated nonphosphorylated peptides (typically >9.0). This pI difference allows isolation of methylated phosphopeptides from the methylated nonphosphopeptides by in-solution isoelectric focusing. We proved the principle of such a novel approach by isolating a phosphorylated peptide from nonphosphorylated tryptic digest of myoglobin. While the principle for pI-based, in-solution electrophoresis is proven, it requires further development for practical application. The method described here provides a stepping stone toward more reliable, convenient method for efficient isolation of phosphopeptides.     

Synthesis and biodegradation of nanogels as delivery carriers for carbohydrate drugs

Biodegradable nanogels loaded with rhodamine B isothiocyanate-dextran (RITC-Dx) as a model for water-soluble biomacromolecular drugs were prepared using atom-transfer radical polymerization (ATRP) in a cyclohexane inverse miniemulsion in the presence of a disulfide-functionalized dimethacrylate cross linker. UV-vis spectroscopy was used to characterize the extent of incorporation of RITC-Dx into the nanogels. The loading efficiency of RITC-Dx into the nanogels exceeded 80%. These nanogels were degraded into polymeric sols in a reducing environment to release the encapsulated carbohydrate drugs. The released carbohydrate biomolecules specifically interacted with concanavalin A in water, suggesting that the biodegradable nanogels could be used as carriers to deliver carbohydrate drugs that can be released upon degradation to bind to pathogens based on lectins.