Singlet oxygen is generated by bacteriochlorophylls when light and oxygen are simultaneously present in Rhodobacter sphaeroides. Singlet oxygen triggers a specific response that is partly regulated ...by the alternative sigma factor RpoHI/HII. The sRNA RSs2461 has previously been identified as an RpoHI/HII-dependent sRNA and is derived from the 3′ UTR of the mRNA for an OmpR-type transcriptional regulator. Similar to the RpoHI/HII-dependent CcsR and SorY sRNAs, RSs2461 affects the resistance of R. sphaeroides against singlet oxygen and was therefore renamed here SorX. Furthermore, SorX has a strong impact on resistance against organic hydroperoxides that usually occur as secondary damages downstream of singlet oxygen. The 75-nt SorX 3′ fragment, which is generated by RNase E cleavage and highly conserved among related species, represents the functional entity. A target search identified potA mRNA, which encodes a subunit of a polyamine transporter, as a direct SorX target and stress resistance via SorX could be linked to potA. The PotABCD transporter is an uptake system for spermidine in E. coli. While spermidine is generally described as beneficial during oxidative stress, we observed significantly increased sensitivity of R. sphaeroides to organic hydroperoxides in the presence of spermidine. We therefore propose that the diminished import of spermidine, due to down-regulation of potA by SorX, counteracts oxidative stress. Together with results from other studies this underlines the importance of regulated transport to bacterial stress defense.
As glioblastomas are mostly localized infiltrative lesions, gene therapy based on the retroviral replicating vector (RRV) system is considered an attractive strategy. Combinations of multiple suicide ...genes can circumvent the limitations associated with each gene, achieving direct and synergistic cytotoxic effects, along with bystander cell killing. In this study, we constructed a semi-and pseudotyped-RRV (sp-RRV) system harboring two suicide genes-herpes simplex virus type 1 thymidine kinase (
) and yeast cytosine deaminase (CD)
to verify the dissemination and antitumor efficacy of our sp-RRV system (spRRVe-sEF1α-TK/sRRVgp-sEF1α-CD) in seven patient-derived glioblastoma stem-like cells (GSCs). Flow cytometry and high-content analysis revealed a wide range of transduction efficiency and good correlation between the delivery of therapeutic genes and susceptibility to the prodrugs ganciclovir and 5-fluorocytosine in patient-derived GSCs in vitro. Intra-tumoral delivery of spRRVe-sEF1α-TK/sRRVgp-sEF1α-CD, combined with prodrug treatment, synergistically inhibited cell proliferation and angiogenesis while increasing apoptosis and the depletion of tumor-associated macrophages in orthotopic glioblastoma xenografts. Genomic profiling of patient-derived GSCs revealed that the key genes preventing sp-RRV infection and transmission were associated with cell adhesion, migration, development, differentiation, and proliferation. This is the first report demonstrating that a novel sp-RRV-mediated TK/CD double suicide gene transfer system has high oncolytic power against extremely heterogeneous and treatment-refractory glioblastomas.
To maximize the biological activity of branched-chain amino acids (BCAAs), it is necessary to find a new excipient agent to increase the bioavailability of BCAAs in protein mixtures. The aim of the ...current study was to investigate the effects of soy lecithin (SLC), zinc oxide (ZnO), and methylsulfonylmethane (MSM) on the bioaccessibility and intestinal transport of BCAAs from animal and plant protein mixtures (PMs)
an
digestion model with human intestinal epithelial (Caco-2) cells. The bioaccessibility of total BCAAs in PMs considerably increased by 107.51 ± 1.50% with the addition of SLC, and the combined effects of SLC, ZnO, and MSM on enhancing the bioaccessibility of total BCAAs was observed (107.14 ± 0.18%). Interestingly, SLC showed a major role in binding bile acid, showing 65.78 ± 1.66% of binding capacity. Intestinal transport of BCAAs was measured to be at 100.48, 110.86, and 130.29 μg mL
for leucine, isoleucine, and valine, respectively, in PMs with SLC + ZnO + MSM, and it eventually amplified the amount of the total transported BCAAs (341.63 ± 6.34 μg mL
), which was about 8.72 times higher than that of PM only. The cellular integrity of digesta-treated Caco-2 cells tended to decrease according to the incubation time, but it was recovered in the treatment of PM + SLC + ZnO + MSM, and nearly reached the control levels with 92.82 ± 0.53%. Results from the current study suggest that the co-consumption of proteins equally consisting of plant and animal sources with SLC, ZnO, and MSM could improve the bioavailability of total BCAAs, resulting in the improvement of health benefits.
Colorectal cancer (CRC) screening is the most efficient strategy to reduce disease-related mortality. Frequent aberrant DNA methylation is known to occur in selected genes and early during CRC ...development, which has emerged as a new epigenetic biomarker for early detection of CRC. Previously, we reported that we identified that CpG sites of
were aberrantly methylated in tumor tissues of most CRC patients through comprehensive methylation analysis and demonstrated a high potential of quantification of
methylation in blood for early detection of colorectal cancer. In this study, we aim to investigate the feasibility of quantifying
methylation in stool DNA for the early detection of CRC. The objective of this study was to confirm a high frequency of
methylation in tumor tissues at various stages of CRC and investigate the feasibility of a quantitative test for
methylation in fecal DNA by highly sensitive and accurate real-time PCR for early detection of CRC.
Bisulfite-pyrosequencing assay was performed to measure the
methylation status in tissue samples. For methylation analysis in stool DNA, a highly sensitive and accurate method was applied which implements consecutive two rounds of PCR consisting of unidirectional linear target enrichment (LTE) of
and quantitative methylation-specific real time PCR (qMSP) for
, named as me
LTE-qMSP assay. Its limit of detection was 0.1% methylation (corresponding to ~ 6 copies in total ~ 6200 genome copies).
Positive
methylation was observed in 100% of primary tumors, 90.6% of adenomatous polyps, 94.1% of hyperplastic polyps, and 0% of normal tissues.
methylation level also significantly (
< 0.01) increased according to the severity of lesions. In stool DNA test for
methylation by LTE-qMSP comparing CRC patients with various stages (I to IV) (
= 50) and precancerous lesions (
= 21) with healthy subjects (
= 22), the overall sensitivity was 90.0% for detecting CRC and 33.3% for detecting small polyps, with a specificity of 90.9%.
Taken together, our result indicates that stool DNA-based
methylation test by LTE-qMSP is a potential noninvasive diagnostic tool for early detection of CRC.
ABSTRACT
Dynamic changes in the actin network are crucial for the cortical migration of spindles and establishment of polarity, to ensure asymmetric division during meiotic maturation. In this study, ...filamin A (FLNA) was found to be an essential actin regulator that controlled spindle migration and asymmetric division during oocyte meiosis. FLNA was localized in the cytoplasm and enriched at the cortex and near the chromosomes. Knockdown of FLNA impaired meiotic asymmetric division and spindle migration with a decrease in the amount of cytoplasmic actin mesh and cortical actin levels. Moreover, FLNA knockdown reduced the phosphorylation of cofilin and Rho kinase (ROCK) near the spindle. Similar phenotypes, such as decreased filament actin levels, impaired spindle migration and polar body extrusion, were observed when active cofilin (S3A) was overexpressed or ROCK was inhibited. Notably, we found that FLNA and ROCK interacted directly in mouse oocytes. Taken together, our results show that FLNA plays crucial roles in asymmetric division during meiotic maturation by regulating ROCK‐cofilin–mediated actin reorganization.—Wang, H., Guo J., Lin, Z., Namgoong, S., Oh, J. S., Kim, N.‐H. Filamin A is required for spindle migration and asymmetric division in mouse oocytes. FASEB J. 31, 3677–3688 (2017). www.fasebj.org
Endocrine-cerebro-osteodysplasia (ECO) syndrome is a genetic disorder associated with congenital defects of the endocrine, cerebral, and skeletal systems in humans. ECO syndrome is caused by ...mutations of the intestinal cell kinase (ICK) gene, which encodes a mitogen-activated protein (MAP) kinase-related kinase that plays a critical role in controlling the length of primary cilia. Lack of ICK function disrupts transduction of sonic hedgehog (SHH) signaling, which is important for development and homeostasis in humans and mice. Craniofacial structure abnormalities, such as cleft palate, are one of the most common defects observed in ECO syndrome patients, but the role of ICK in palatal development has not been studied.
Using Ick-mutant mice, we investigated the mechanisms by which ICK function loss causes cleft palate and examined pharmacological rescue of the congenital defects.
SHH signaling was compromised with abnormally elongated primary cilia in the developing palate of Ick-mutant mice. Cell proliferation was significantly decreased, resulting in failure of palatal outgrowth, although palatal adhesion and fusion occurred normally. We thus attempted to rescue the congenital palatal defects of Ick mutants by pharmacological activation of SHH signaling. Treatment of Ick-mutant mice with an agonist for Smoothened (SAG) rescued several congenital defects, including cleft palate.
The recovery of congenital defects by pharmacological intervention in the mouse models for ECO syndrome highlights prenatal SHH signaling modulation as a potential therapeutic measure to overcome congenital defects of ciliopathies.
Oocyte maturation, fertilization, and early embryonic development occur in the absence of gene transcription. Therefore, it is critical to understand at a global level the post-transcriptional events ...that are driving these transitions. Here we used a systems approach by combining polysome mRNA profiling and bioinformatics to identify RNA-binding motifs in mRNAs that either enter or exit the polysome pool during mouse oocyte maturation. Association of mRNA with the polysomes correlates with active translation. Using this strategy, we identified highly specific patterns of mRNA recruitment to the polysomes that are synchronized with the cell cycle. A large number of the mRNAs recovered with translating ribosomes contain motifs for the RNA-binding proteins DAZL (deleted in azoospermia-like) and CPEB (cytoplasmic polyadenylation element-binding protein). Although a Dazl role in early germ cell development is well established, no function has been described during oocyte-to-embryo transition. We demonstrate that CPEB1 regulates Dazl post-transcriptionally, and that DAZL is essential for meiotic maturation and embryonic cleavage. In the absence of DAZL synthesis, the meiotic spindle fails to form due to disorganization of meiotic microtubules. Therefore, Cpeb1 and Dazl function in a progressive, self-reinforcing pathway to promote oocyte maturation and early embryonic development.
Objective
To evaluate the clinical significance of various vital signs in women referred for postpartum hemorrhage (PPH).
Methods
This retrospective study included patients with primary PPH who were ...referred to Korea University Medical Center, Ansan, between January 1, 2004, and December 31, 2016. We analyzed data for systolic and diastolic blood pressure, heart rate, and shock index (heart rate divided by systolic blood pressure) at time of arrival. Significant morbidity, such as massive transfusion, invasive procedures, and admission to the intensive care unit were reviewed. We used the area under the receiver operating characteristic curve (AUROC) for each vital sign to predict adverse maternal outcomes. Sensitivity, specificity, and negative and positive predictive values were assessed.
Results
118 women with PPH were identified. The shock index had the highest AUROC to predict massive transfusion (0.815, 95% confidence interval CI 0.727–0.883). A shock index greater than 0.9 had 93.8% (95% CI 69.8–99.8) sensitivity and 51.2% (35.1–67.1) specificity for prediction of massive transfusion, and 93.6% (78.6–99.2) sensitivity and 31.0% (15.3–50.8) specificity for prediction of invasive procedures.
Conclusion
The shock index has significant ability to predict adverse outcomes of PPH compared with other initial vital signs when patients are referred.
Initial elevated shock index can predict massive transfusion and invasive procedures when women are referred for primary postpartum hemorrhage.
Background
Intracorporeal Billroth I (ICBI) (delta-shaped) anastomosis is being increasingly used for laparoscopic distal gastrectomy. However, few studies have focused on the safety and feasibility ...of adopting this new technique. The present study aimed to review the surgical outcomes after the initial experience of performing ICBI anastomosis and to evaluate whether this technique can be safely adopted without increasing operative risk during the early learning process.
Methods
Forty-two consecutive patients who underwent ICBI anastomosis with laparoscopic distal gastrectomy by a single surgeon were enrolled, and their operative outcomes and hospital course were compared with those of 179 patients who underwent conventional extracorporeal Billroth I (ECBI) anastomosis by the same operator. The learning curve was assessed by evaluating the moving average of anastomosis time.
Results
The operating time in the ICBI group was significantly longer than that in the ECBI group (142 vs. 116 min,
p
< 0.001). However, there were no significant differences in the postoperative hospital course such as gas passage, diet initiation, postoperative fever, and hospital stay between the two groups. Postoperative morbidity did not significantly differ between the ICBI and ECBI groups (7.1 vs. 12.3 %,
p
= 0.428). No anastomosis-related complications occurred in the ICBI group. The mean anastomosis time for ICBI anastomosis was 24 ± 5 min, and the anastomosis average time curve showed that it reached a plateau approximately after the 14th case.
Conclusions
ICBI anastomosis has a steep learning curve without increasing operative risk in the early learning process, when performed by experienced laparoscopic surgeons. The technical feasibility and clinical advantages of intracorporeal anastomosis need to be proven in future clinical trials.
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