The effect of macromolecular crowding on single-round transcription by Escherichia coli RNA polymerase.

Previous works have reported significant effects of macromolecular crowding on the structure and behavior of biomolecules. The crowded intracellular environment, in contrast to in vitro buffer solutions, likely imparts similar effects on biomolecules. The enzyme serving as the gatekeeper for the genome, RNA polymerase (RNAP), is among the most regulated enzymes. Although it was previously demonstrated that macromolecular crowding affects association of RNAP to DNA, not much is known about how crowding acts on late initiation and promoter clearance steps, which are considered to be the rate-determining steps for many promoters. Here, we demonstrate that macromolecular crowding enhances the rate of late initiation and promoter clearance using in vitro quenching-based single-molecule kinetics assays. Moreover, the enhancement's dependence on crowder size notably deviates from predictions by the scaled-particle theory, commonly used for description of crowding effects. Our findings shed new light on how enzymatic reactions could be affected by crowded conditions in the cellular milieu.

Cecal epidermoid cyst: a neonatal case with clinicopathological consideration.

BACKGROUND: Only 10 cases of cecal epidermoid cyst (CEC) have been reported in the literature. Furthermore, its pathogenesis remains unclear. We report a rare case of congenital CEC in neonate, and discuss its clinicopathological findings. CASE PRESENTATION: A cystic lesion was incidentally identified in the retroperitoneal area of the abdominal right lower quadrant during a routine prenatal ultrasonography (US), prompting an ileocolectomy 3 days after birth. This congenital cyst was composed of mucosal lining cells and submucosal connective tissues, and the inner lining mucosa was composed of stratified squamous epithelium and focally mucin-producing ciliated epithelium. Based on the macroscopic and microscopic findings, the cystic lesion was diagnosed as a congenital cecal epidermoid cyst. CONCLUSIONS: The management of a fetal abdominal mass should be tailored individually, considering that epidermoid cysts can occur in the cecum during the perinatal period. We report the clinicopathological findings in this case, including its possible pathogenesis.

The Capsicum baccatum-Specific Truncated NLR Protein CbCN Enhances the Innate Immunity against Colletotrichum acutatum.

Chili pepper (Capsicumannuum) is an important fruit and spice used globally, but its yield is seriously threatened by anthracnose. Capsicum baccatum is particularly valuable as it carries advantageous disease resistance genes. However, most of the genes remain to be identified. In this study, we identified the C. baccatum-specific gene CbCN, which encodes a truncated nucleotide-binding and leucine-rich repeat protein in the anthracnose resistant chili pepper variety PBC80. The transcription of CbCN was greater in PBC80 than it was in the susceptible variety An-S after Colletotrichum acutatum inoculation. In order to investigate the biological function of CbCN, we generated transgenic tobacco lines constitutively expressing CbCN. Notably, CbCN-overexpressing transgenic plants exhibited enhanced resistance to C. acutatum compared to wild-type plants. Moreover, the expression of pathogenesis-related (PR) genes was remarkably increased in a CbCN-overexpressing tobacco plants. In order to confirm these results in chili pepper, we silenced the CbCN gene using the virus-induced gene silencing system. The anthracnose resistance and expressions of PR1, PR2, and NPR1 were significantly reduced in CbCN-silenced chili peppers after C. acutatum inoculations. These results indicate that CbCN enhances the innate immunity against anthracnose caused by C. acutatum by regulating defense response genes.

Identification of the Capsicum baccatum NLR Protein CbAR9 Conferring Disease Resistance to Anthracnose.

Anthracnose is caused by Colletotrichum species and is one of the most virulent fungal diseases affecting chili pepper (Capsicum) yield globally. However, the noble genes conferring resistance to Colletotrichum species remain largely elusive. In this study, we identified CbAR9 as the causal locus underlying the large effect quantitative trait locus CcR9 from the anthracnose-resistant chili pepper variety PBC80. CbAR9 encodes a nucleotide-binding and leucine-rich repeat (NLR) protein related to defense-associated NLRs in several other plant species. CbAR9 transcript levels were induced dramatically after Colletotrichum capsici infection. To explore the biological function, we generated transgenic Nicotiana benthamiana lines overexpressing CbAR9, which showed enhanced resistance to C. capsici relative to wild-type plants. Transcript levels of pathogenesis-related (PR) genes increased markedly in CbAR9-overexpressing N. benthamiana plants. Moreover, resistance to anthracnose and transcript levels of PR1 and PR2 were markedly reduced in CbAR9-silenced chili pepper fruits after C. capsici infection. Our results revealed that CbAR9 contributes to innate immunity against C. capsici.

Rice glutaredoxin GRXS15 confers broad-spectrum resistance to Xanthomonas oryzae pv. oryzae and Fusarium fujikuroi.

Rice is a particularly widely consumed food crop globally, but its yield is seriously damaged by bacterial blight due to Xanthomonas oryzae pv. oryzae (Xoo) and bakanae disease due to Fusarium fujikuroi (F. fujikuroi). However, broad-spectrum resistance (BSR) to both Xoo and F. fujikuroi remains largely elusive. In this study, we showed that rice monothiol glutaredoxin GRXS15 localizes in mitochondria and the nucleus, and its transcription is induced by Xoo. Transgenic rice lines constitutively expressing OsGRXS15 showed enhanced disease resistance to Xoo and F. fujikuroi, while CRISPR/Cas9-based knockout mutants showed reduced resistance compared with the wild-type plants. The transcription of pathogenesis-related (PR) genes was significantly induced in OsGRXS15-expressing plants. The rice transcription factor OsWRKY65 was identified as a binding partner, and it directly interacted with OsGRXS15 in the nucleus. Moreover, we revealed that the interaction of OsGRXS15 and OsWRKY65 results in the upregulation of OsPR1. These results suggested that OsGRXS15 interacts with transcription factors, and it confers BSR through regulating the expression of genes related to pathogen response. This is the first report on the nuclear function associated with the monothiol glutaredoxin GRXS15.

Predicting Survival of Congenital Diaphragmatic Hernia on the First Day of Life.

BACKGROUND: This study aimed to determine perinatal risk factors for 30-day mortality of congenital diaphragmatic hernia (CDH) patients and develop a prognostic index to predict 30-day mortality of CDH patients. Identifying risk factors that can prognosticate outcome is critical to obtain the best management practices for patients. METHODS: A retrospective study was performed for patients who were diagnosed with CDH from November 2000 to August 2016. A total of 10 prenatal risk factors and 14 postnatal risk factors were analyzed. All postnatal variables were measured within 24 h after birth. RESULTS: A total of 95 CDH patients were enrolled in this study, including 61 males and 34 females with mean gestational age of 38.86 ± 1.51 weeks. The overall 30-day survival rate was 63.2%. Multivariate analysis revealed that five factors (polyhydramnios, gestational age at diagnosis <25 weeks, observed-to-expected lung-to-head ratio ≤45, best oxygenation index in 24 h >11, and severity of tricuspid regurgitation ≥ mild) were independent predictors of 30-day mortality of CDH. Using these five factors, a perinatal prognostic index for 30-day mortality was developed. Four predictive models (poor, bad, good, and excellent) of the perinatal prognostic index were constructed, and external validation was performed. CONCLUSIONS: Awareness of risk factors is very important for predicting prognosis and managing patients. Five independent perinatal risk factors were identified in this study. A perinatal prognostic index was developed for 30-day mortality for patients with CDH. This index may be used to help manage CDH patients.

Endoplasmic reticulum stress-induced accumulation of VAMP721/722 requires CALRETICULIN 1 and CALRETICULIN 2 in Arabidopsis.

Excessive demand for translation and protein folding in the endoplasmic reticulum (ER) can cause ER stress in plants. Here, we show that CALRETICULIN 1 (CRT1) and CRT2 are critical components in the accumulation of VESICLE-ASSOCIATED MEMBRANE PROTEIN 721 (VAMP721) and VAMP722 during ER stress responses. We show that CRT2 interacts with VAMP722 and that CRT1/2 post-translationally maintain elevated VAMP721/722 levels under ER stress. The greater growth inhibition in VAMP721/722-deficient plants, induced by tunicamycin, suggests that plants under ER stress maintain physiological homeostasis, at least in part, by regulating VAMP721/722 levels, as VAMP721/722 are known to participate in various biological processes.

Trends in Antibiotic Susceptibility in Staphylococcus aureus in Boston, Massachusetts, from 2000 to 2014.

The rate of infection by methicillin-resistant Staphylococcus aureus (MRSA) has declined over the past decade, but it is unclear whether this represents a decline in S. aureus infections overall. To evaluate the trends in the annual rates of infection by S. aureus subtypes and mean antibiotic resistance, we conducted a 15-year retrospective observational study at two tertiary care institutions in Boston, MA, of 31,753 adult inpatients with S. aureus isolated from clinical specimens. We inferred the gain and loss of methicillin resistance through genome sequencing of 180 isolates from 2016. The annual rates of infection by S. aureus declined from 2003 to 2014 by 4.2% (2.7% to 5.6%), attributable to an annual decline in MRSA of 10.9% (9.3% to 12.6%). Penicillin-susceptible S. aureus (PSSA) increased by 6.1% (4.2% to 8.1%) annually, and rates of methicillin-susceptible penicillin-resistant S. aureus (MSSA) did not change. Resistance in S. aureus decreased from 2000 to 2014 by 0.8 antibiotics (0.7 to 0.8). Within common MRSA clonal complexes, 3/14 MSSA and 2/21 PSSA isolates arose from the loss of resistance-conferring genes. Overall, in two tertiary care institutions in Boston, MA, a decline in S. aureus infections has been accompanied by a shift toward increased antibiotic susceptibility. The rise in PSSA makes penicillin an increasingly viable treatment option.

Perineal squamous cell carcinoma arising from an epidermal cyst: a case report.

BACKGROUND: Epidermal cysts and squamous cell carcinomas (SCCs) are common skin lesions. However, a malignant change in an epidermal cyst is very rare. The incidence of a malignant change from an epidermal cyst to cutaneous SCC is 0.011-0.045%. In particular, malignant transformation of an epidermal cyst in the perineum is extremely rare. To date, three cases have been reported in the English literature. CASE PRESENTATION: We report a case of 51-year-old male with an approximately 15-cm perineal mass. This mass started to grow suddenly 4 months previously and caused great discomfort in the perineum due to the large size. The patient underwent excision of the mass with a negative margin. Histopathological analysis confirmed a microinvasive SCC arising from a proliferating epidermoid cyst. CONCLUSIONS: Even if benign tumors are suspected, a change in size, pain, ulceration, or discharge should indicate the need for surgical resection due to the possibility of a malignant change.

Synaptotagmin 1 Negatively Controls the Two Distinct Immune Secretory Pathways to Powdery Mildew Fungi in Arabidopsis.

PEN1, one of the plasma membrane (PM) syntaxins, comprises an immune exocytic pathway by forming the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex with SNAP33 and VAMP721/722 in plants. Although this secretory pathway is also involved in plant growth and development, how plants control their exocytic activity is as yet poorly understood. Since constitutive PEN1 cycling between the PM and endocytosed vesicles is critical for its immune activity, we studied here the relationship of PEN1 to synaptotagmin 1 (SYT1) that is known to regulate endocytosis at the PM. Interestingly, syt1 plants showed enhanced disease resistance to the Arabidopsis-adapted Golovinomyces orontii fungus, and elevated protein but not transcript levels of PEN1 Calcium-dependent promotion of PEN1-SYT1 interaction suggests that SYT1 controls defense activities of the PEN1-associated secretory pathway by post-translationally modulating PEN1. Increased PEN1-SYT1 interaction and inhibited PEN1 SNARE complex induction by G. orontii additionally suggest that the adaption of phytopathogens to host plants might partly result from effective suppression of the PEN1-related secretory pathway. Further genetic analyses revealed that SYT1 also regulates the atypical peroxisomal myrosinase PEN2-associated secretory pathway.

Congenital adhesion band causing small bowel obstruction: What's the difference in various age groups, pediatric and adult patients?

BACKGROUND: A congenital adhesion band is a rare condition, but may induce a small bowel obstruction (SBO) at any age. However, only a few sporadic case reports exit. We aimed to identify the clinical characteristics of congenital adhesion band manifesting a SBO stratified by age group between pediatric and adult patients. METHODS: The medical records of all patients with a SBO between Jan 1, 2009 and Dec 31, 2015 were retrospectively reviewed. Cases associated with previous surgical procedure and cases of secondary obstruction due to inflammatory processes or tumor and other systemic diseases were excluded. The patients were divided into two groups according to age below or above 18 years: pediatric and adult. The basic clinical characteristics were analyzed and compared between groups. RESULTS: Of 251 patients with a SBO, 15 (5.9%) met the inclusion criteria; 10 cases in pediatric group (mean age 17.9 ± 38.7 months) and 5 cases in adult group (mean age 60.0 ± 19.7 years). The pediatric group (66.6%) included 3 neonates, 5 infants, and 2 school children. They usually presented with bilious vomiting (50.0%) and abdominal distention (60.0%), and demonstrated a high rate of early operation (80.0%) and bowel resection (70.0%). In contrast, the adult group (33.3%) presented with abdominal pain (100%) in all cases and underwent a relatively simple procedure of band release using a laparoscopic approach (60%). However, group differences did not reach statistical significance. In addition, two groups did not differ in the time interval to the operation or in the range of the operation (p = 0.089 vs. p = 0.329). No significant correlation was found between the time interval to the operation and the necessity of bowel resection (p = 0.136). There was no mortality in either group. CONCLUSIONS: Congenital adhesion band is a very rare condition with diverse clinical presentations across ages. Unlike adult patients, pediatric patients showed a high proportion of early operation and bowel resection. A good result can be expected with an early diagnosis and prompt management regardless of age.

Neonatal Iliopsoas Abscess: The First Korean Case.

Iliopsoas abscess (IPA) is rare in neonates. We present a case of neonatal IPA that was initially believed to bean inguinal hernia. A 20-day-old male infant was referred to our hospital for herniorrhaphy after a 2-day history of swelling and bluish discoloration of the left inguinal area and leg without limitation of motion. Abdominal and pelvic ultrasonography suggested a femoral hernia, but the anatomy was unclear. Abdominal computed tomography revealed a multi-septated cystic mass extending into the psoas muscle from the lower pole of the left kidney to the femur neck. Broad spectrum antibiotics were initiated, and prompt surgical exploration was planned. After opening the retroperitoneal cavity via an inguinal incision, an IPA was diagnosed and surgically drained. Culture of the abscess fluid detected Staphylococcus aureus, sensitive to methicillin. The patient was discharged without complication on the 17th postoperative day.

Cell adhesion and in vivo osseointegration of sandblasted/acid etched/anodized dental implants.

The authors describe a new type of titanium (Ti) implant as a Modi-anodized (ANO) Ti implant, the surface of which was treated by sandblasting, acid etching (SLA), and anodized techniques. The aim of the present study was to evaluate the adhesion of MG-63 cells to Modi-ANO surface treated Ti in vitro and to investigate its osseointegration characteristics in vivo. Four different types of Ti implants were examined, that is, machined Ti (control), SLA, anodized, and Modi-ANO Ti. In the cell adhesion study, Modi-ANO Ti showed higher initial MG-63 cell adhesion and induced greater filopodia growth than other groups. In vivo study in a beagle model revealed the bone-to-implant contact (BIC) of Modi-ANO Ti (74.20%±10.89%) was much greater than those of machined (33.58%±8.63%), SLA (58.47%±12.89), or ANO Ti (59.62%±18.30%). In conclusion, this study demonstrates that Modi-ANO Ti implants produced by sandblasting, acid etching, and anodizing improve cell adhesion and bone ongrowth as compared with machined, SLA, or ANO Ti implants. These findings suggest that the application of Modi-ANO surface treatment could improve the osseointegration of dental implant.

Maximum intensity projection using bidirectional compositing with block skipping.

BACKGROUND: Maximum intensity projection (MIP) is a volume rendering technique that determines the pixel intensity as the maximum of all values sampled along the viewing direction. MIP has been successfully applied to diagnose bone fractures in computed tomography (CT) and the stenosis of vascular structures in magnetic resonance angiography (MRA). However, MIP has a major drawback in that the depth and occlusion information cannot be perceived in the output images. The most universal way to alleviate this problem is to occasionally change the viewpoint for depth perception. To support this function in real time, MIP should be performed at an interactive frame rate. OBJECTIVE: We develop an efficient rendering algorithm for MIP so that MIP is performed at an interactive frame rate without a loss of image quality. METHODS: The proposed method predicts the position of the maximum intensity for each ray using blockwise maximum bounds, after which it performs bidirectional compositing toward both ends of the ray from this predicted position. During the compositing process, block skipping is used as an acceleration method. RESULTS: The proposed method outperformed the block skipping method using the sequential compositing with a speed-up factor of 2.2 ∼ 2.8 depending on the data set without any degradation of the image quality. CONCLUSION: We proposed an efficient rendering technique for MIP. Our method was superior to the conventional block skipping method with respect to the rendering speed and degree of performance consistency.

High-throughput single-molecule optofluidic analysis.

We describe a high-throughput, automated single-molecule measurement system, equipped with microfluidics. The microfluidic mixing device has integrated valves and pumps to accurately accomplish titration of biomolecules with picoliter resolution. We demonstrate that the approach enabled rapid sampling of biomolecule conformational landscape and of enzymatic activity, in the form of transcription by Escherichia coli RNA polymerase, as a function of the chemical environment.

A rare case of infected urachal cyst leading to intestinal obstruction in a 3-month-old boy with febrile urinary tract infection: Case report.

RATIONALE: Urachal anomalies are rare and can present with various clinical manifestations. Urachal remnants, in particular, can be difficult to diagnose because of atypical symptoms at presentation. This study reports a case of intestinal obstruction in an infant secondary to an infected urachal cyst. PATIENTS CONCERNS: A 3-month-old boy with a known febrile urinary tract infection developed acute abdominal distension. DIAGNOSES: Abdominal ultrasound (US) and computed tomography (CT) revealed a nonspecific, ill-defined soft tissue density at the mid-abdomen, associated with intestinal obstruction. INTERVENTIONS: Emergency exploratory laparotomy was performed. The site of the obstruction was found to be at the mid-small bowel; the proximal small bowel was markedly distended, and the small bowel and sigmoid colon were adherent to urachal remnant. The urachal remnant was excised, and the peritoneal adhesions were lysed. OUTCOMES: The day after surgery, the patient was discharged without any complications. LESSONS: Intestinal obstruction is an exceedingly rare presentation of urachal remnants. This case highlights that urachal anomalies should be considered in the differential diagnosis in patients with intestinal obstruction and a concurrent febrile urinary tract infection.

A novel animal model of abdominal aortic aneurysm by mechanical injury.

The present study established a novel and reproducible animal model to study abdominal aortic aneurysms. In total, 22 adult Lewis rats underwent a procedure to produce mechanical injuries at the infrarenal aorta which was opened temporarily. The aortas were injured 6 times and repaired. Those rats were divided into 2 groups and the aortic aneurysm tissue was harvested after 42 (6-week group) or 63 (9-week group) days and evaluated for the progression of aortic aneurysms. In the 6-week group, changes in the aneurysm were observed in 6/10 (60%) rats and the mean maximum diameter of the aorta demonstrated a 119% increase in size from the baseline measurement. In the 9-week group, changes in the aneurysm were observed in 8/11 (88%) rats and the mean maximum diameter of aorta demonstrated a 133% increase in size. Additional findings from the aortic aneurysm tissue were found microscopically, including the destruction of the tunica media and the elastic fiber. The present study demonstrated that this novel animal model for the development of abdominal aortic aneurysms (AAAs) produced by mechanical injury may have high reproducibility and similar gross and microscopic morphology to humans. This model could be helpful to investigate the treatment of AAAs.

Suppressive role of vascular endothelial growth factor on intestinal apoptosis in induced necrotizing enterocolitis in rats.

Purpose: Necrotizing enterocolitis (NEC) is a devastating disease that can cause mortality in preterm babies. NEC may develop through an apoptotic pathway that is known to be inhibited by vascular endothelial growth factor (VEGF). This study determined whether VEGF exerted a protective effect against the development of NEC and apoptosis in rats. Methods: To determine the effect of VEGF in NEC rats, neonatal rats were randomized into 4 groups: the control group, the NEC group, the NEC + intraperitoneal VEGF (50 ng/kg) group (NEC + VEGF IP group), and the NEC + oral VEGF (50 ng/kg) group (NEC + VEGF OR group). NEC was induced by lipopolysaccharide/hypoxia and cold stress. The animals were sacrificed 72 hours later. After laparotomy, we obtained a region of the proximal small bowel from the ileocecal valve about 18 cm in length. Results: The NEC histological grade, apoptosis histological score, and caspase-3 activity were lower in the NEC + VEGF IP and OR groups than in the NEC group. In the NEC + VEGF IP and OR groups, the messenger RNA expression of apoptotic and inflammatory genes, such as Bax, NF-κB, p53, Fas, FasL, and PAF-R, but not that of Bcl-2, was decreased, as was the Bax/Bcl-2 protein ratio. Histological analysis revealed that the apoptosis-blocking effect of VEGF was more effective in the NEC + VEGF IP group than in the NEC + VEGF OR group. Conclusion: We identified apoptotic and inflammatory genes to confirm the preventive effect of VEGF pretreatment on NEC in rats. This study presents a novel approach to prevent apoptosis via VEGF pretreatment in rats with lipopolysaccharide/hypoxia-induced NEC.

Scrotal pyocele secondary to gastrointestinal perforation in infants: a case series.

Pyocele in infants is rarely described in the literature, but it is an emergent condition that requires rapid recognition and treatment to prevent testicular loss. If peritonitis due to gastrointestinal perforation occurs, abdominal contamination may spread through a patent processus vaginalis in an infant, which may lead to pyocele. We report the cases of three infants with scrotal pyocele due to the spread of infection or inflammatory material from the intraperitoneal cavity through a patent processus vaginalis. Two infants were surgically treated, while the other was treated with percutaneous aspiration and intravenous antibiotic administration. Although rare, pyocele should be considered in the differential diagnosis of acute scrotum in infants, especially in infants who previously had peritonitis due to gastrointestinal perforation.

Distinctive clinical features of spontaneous pneumoperitoneum in neonates: A retrospective analysis.

BACKGROUND: Spontaneous pneumoperitoneum (SP) without gastrointestinal perforation rarely occurs in neonates, with most SP cases being idiopathic. Although SP usually follows a benign clinical course with favorable prognosis, it can become life-threatening in certain situations. In these cases, urgent surgical intervention may be required. Therefore, it may be difficult to decide when or how to perform prompt interventions. AIM: To demonstrate the distinct clinical features of SP to guide appropriate management by comparing characteristics between SP and typical pneumoperitoneum secondary to gastrointestinal perforation. METHODS: We retrospectively reviewed electronic medical records and identified 37 neonates with radiological evidence of pneumoperitoneum who were treated at our institution. Clinical variables were compared between neonates with SP without gastrointestinal perforation (Group A) and those with pneumoperitoneum secondary to gastrointestinal perforation (Group B). Clinical variables between groups were compared using Student's t-test and the chi-square test. The risk factors related to mortality were examined using multi-logistic regression analysis. RESULTS: Group A comprised 35.1% (13/37) of the patients. The frequency of persistent pulmonary hypertension (53.8%) and pneumothorax (46.2%) before the development of pneumoperitoneum was significantly higher in group A than in group B (P = 0.004). Platelet count and partial pressure of arterial oxygen (PaO2) were significantly lower in group A (P = 0.015 and 0.025, respectively). Overall mortality was significantly higher in group A than in group B (76.9% vs 16.7%, P = 0.001). Only preterm infants were significantly associated with high mortality (P = 0.041; odds ratio = 18.0). Accompaniment with persistent pulmonary hypertension and pneumothorax were also significantly high (P = 0.004) in group A, but these were not strongly associated with high mortality. CONCLUSION: This study identified a higher mortality rate in patients with SP than that described in previous reports. Neonates with SP were more likely to have thrombocytopenia, pneumothorax, and persistent pulmonary hypertension. Prematurity was the most significant factor affecting mortality.