Japanese Nursing Staff's Knowledge and Attitude toward Bereavement Care for Couples with Miscarriage/Stillbirth and Its Associated Factors.

Bereavement care is conducted to meet the emotional needs of grieving couples who are devastated by the experience of a miscarriage or stillbirth. From January to April 2022, we distributed a questionnaire that assessed the knowledge and attitudes of Japanese nursing staff (nurses and midwives) in Japan's Chugoku-Shikoku region toward bereavement care for couples with miscarriage/stillbirth. The 370 survey respondents' answers revealed that the nursing staff's knowledge regarding recurrent pregnancy loss and subsequent bereavement care was insufficient. About 41.1% and 64.1% of the respondents had received school and on-the-job education in bereavement care, respectively, and 79.2% expressed willingness to provide such care. Our analyses revealed that the following factors were associated with the nursing staff's knowledge level: parent status, age, reproductive history, midwifery license, work experience and environment, and on-the-job education. The following were correlated with the staff's willingness to provide bereavement care: work environment, midwifery license, bereavement care knowledge, and on-the-job education. Together our findings indicate that education plays a significant role in equipping caregivers to provide effective bereavement care for couples who have experienced a miscarriage or stillbirth.

First-Line Selpercatinib or Chemotherapy and Pembrolizumab in RET Fusion-Positive NSCLC.

BACKGROUND: Selpercatinib, a highly selective potent and brain-penetrant RET inhibitor, was shown to have efficacy in patients with advanced RET fusion-positive non-small-cell lung cancer (NSCLC) in a nonrandomized phase 1-2 study. METHODS: In a randomized phase 3 trial, we evaluated the efficacy and safety of first-line selpercatinib as compared with control treatment that consisted of platinum-based chemotherapy with or without pembrolizumab at the investigator's discretion. The primary end point was progression-free survival assessed by blinded independent central review in both the intention-to-treat-pembrolizumab population (i.e., patients whose physicians had planned to treat them with pembrolizumab in the event that they were assigned to the control group) and the overall intention-to-treat population. Crossover from the control group to the selpercatinib group was allowed if disease progression as assessed by blinded independent central review occurred during receipt of control treatment. RESULTS: In total, 212 patients underwent randomization in the intention-to-treat-pembrolizumab population. At the time of the preplanned interim efficacy analysis, median progression-free survival was 24.8 months (95% confidence interval [CI], 16.9 to not estimable) with selpercatinib and 11.2 months (95% CI, 8.8 to 16.8) with control treatment (hazard ratio for progression or death, 0.46; 95% CI, 0.31 to 0.70; P<0.001). The percentage of patients with an objective response was 84% (95% CI, 76 to 90) with selpercatinib and 65% (95% CI, 54 to 75) with control treatment. The cause-specific hazard ratio for the time to progression affecting the central nervous system was 0.28 (95% CI, 0.12 to 0.68). Efficacy results in the overall intention-to-treat population (261 patients) were similar to those in the intention-to-treat-pembrolizumab population. The adverse events that occurred with selpercatinib and control treatment were consistent with those previously reported. CONCLUSIONS: Treatment with selpercatinib led to significantly longer progression-free survival than platinum-based chemotherapy with or without pembrolizumab among patients with advanced RET fusion-positive NSCLC. (Funded by Eli Lilly and others; ClinicalTrials.gov number, NCT04194944.).

Clinical outcomes of radioactive seed brachytherapy and microwave ablation in inoperable stage I non-small cell lung cancer.

This study assessed the efficacy and safety of radioactive iodine-125 seed ablation brachytherapy (RSABT) in comparison to microwave ablation therapy (MWAT) for treating inoperable stage I non-small cell lung cancer (NSCLC). We conducted a retrospective analysis of data from stage I NSCLC patients who underwent CT-guided RSABT or MWAT. The primary outcomes measured were progression-free survival (PFS), overall survival (OS), and the occurrence of adverse events. Of the patients included in the study, 71 underwent RSABT and 105 received MWAT. The median follow-up time for these groups was 47.4 months and 60 months, respectively. The PFS rates at 1-year, 3-year, and 5-year for the RSABT group were 87.3%, 72.6%, and 65.8%, while for the MWAT group, they were 89.5%, 69.3%, and 43.7%, respectively (P = 0.011). The OS rates at 1-year, 3-year, and 5-year for the RSABT group were 97.2%, 78.1%, and 66.1%, and for the MWAT group, they were 99%, 75.8%, and 55%, respectively (P = 0.112). Upon multivariate analysis, the treatment modality was identified as an independent predictor of PFS (P = 0.008). Additionally, both sex and T stage were found to be independent predictors of both PFS and OS (P < 0.05). Adverse events, such as pneumothorax, occurred in 50% of the MWAT group and 39% of the RSABT group (P = 0.313). The incidence of pleural effusion was 44% in the MWAT group compared to 14% in the RSABT group (P < 0.001). Needle bleeding was observed in 32% of the RSABT group and 5% of the MWAT group (P < 0.001). We conclude RSABT demonstrates promising efficacy and safety in the treatment of stage I NSCLC. However, further studies are essential to validate these preliminary findings.

Differences in the topographical distribution of sleep spindles among adult epilepsy with cognitive impairment.

OBJECTIVE: Cognitive comorbidities are common in epilepsy; however, symptomatic treatment is currently the only available effective therapy. Sleep, cognition, and epilepsy are closely associated. Therefore, many studies on epilepsy and cognition have focused on sleep structures, such as sleep spindles, which are considered windows to understanding the sleeping brain. This study aimed to investigate the relationship between sleep spindles and the severity of cognitive impairment in adult epilepsy. METHODS: Fifty-seven adults with epilepsy underwent overnight sleep electroencephalogram recordings and cognitive testing. Slow (9-12 Hz) and fast (12-15 Hz) spindle characteristics during N2 sleep were calculated using a convolutional neural network-based sleep staging system and automatic spindle detection algorithm. Repeated-measures analysis of variance was used to analyze differences in fast and slow spindle densities among subgroups of patients based on cognitive impairment severity. RESULTS: A significant between-group effect was observed for both slow and fast spindle densities. Multiple comparisons showed that slow and fast spindle densities of the severe cognitive impairment subgroup were lower than those of the noncognitive impairment subgroup (P < 0.05). Simple-effect analysis revealed differences in slow spindle density distributed among the EEG channels Fp1, Fp2, F3, C3, P4, O1, O2, F8, T4, T5, T6, Fz, and Cz (P < 0.05). Differences in fast spindle density were distributed among the channels Fp1, Fp2, F3, C3, O1, O2, F7, F8, T4, T5, T6, and Fz (P < 0.05). SIGNIFICANCE: Significant differences in topographical distribution of fast and slow spindle densities were observed at the scalp level among patients with different cognitive statuses. Compared with patients with no cognitive impairment, those with severe cognitive impairment had lower slow and fast spindle densities over multiple scalp regions during N2 sleep. This study provides a reference for objective assessment of cognitive dysfunction in epilepsy patients.

Does eco-friendly tourism necessary for entrepreneurship? The role of tourism and innovation in sustainable development.

Recent years have seen a surge in international interest in the connections between tourism, technological development, and climate change. This research looks at how increased innovation and tourism might help the economies of the Group of Seven to grow sustainably. Multiple panel unit root tests confirmed the unit root properties of the variables, and the cross-sectional dependency of the panel data from 2000 to 2020 was analyzed. Pedroni and Kao's tests reveal a co-integration link between the variables. Full-modified OLS and dynamic OLS studies show that innovation, as measured by patents and scholarly papers, is associated with economic growth and reduced pollution. In order to estimate the variables, this research makes use of the augmented mean group (AMG) and CCEMG. These findings demonstrate tourism's enormous positive effect on lowering pollution levels and growing economies, two key indicators of sustainable progress. According to the study's findings, asylum seekers did not help boost national economies or participate in national environmental improvement initiatives. Evidence suggests that primary enrollment levels promote sustainable development by reducing environmental degradation and fostering economic growth. The findings suggest that the G7 economies would benefit from increased investment in innovation, tourism infrastructure, and technical higher education. These results provide valuable input for businesses, politicians, and the G7 economies' sustainable development goals.

Assessment of Cognitive Function with Sleep Spindle Characteristics in Adults with Epilepsy.

Objective: Epilepsy may cause chronic cognitive impairment by disturbing sleep plasticity. Sleep spindles play a crucial role in sleep maintenance and brain plasticity. This study explored the relationship between cognition and spindle characteristics in adult epilepsy. Methods: Participants underwent one-night sleep electroencephalogram recording and neuropsychological tests on the same day. Spindle characteristics during N2 sleep were extracted using a learning-based system for sleep staging and an automated spindle detection algorithm. We investigated the difference between cognitive subgroups in spindle characteristics. Multiple linear regressions were applied to analyze associations between cognition and spindle characteristics. Results: Compared with no/mild cognitive impairment, epilepsy patients who developed severe cognitive impairment had lower sleep spindle density, the differences mainly distributed in central, occipital, parietal, middle temporal, and posterior temporal (P < 0.05), and had relatively long spindle duration in occipital and posterior temporal (P < 0.05). Mini-Mental State Examination (MMSE) was associated with spindle density (pars triangularis of the inferior frontal gyrus (IFGtri): ß = 0.253, P = 0.015, and P.adjust = 0.074) and spindle duration (IFGtri: ß = -0.262, P = 0.004, and P.adjust = 0.030). Montreal Cognitive Assessment (MoCA) was associated with spindle duration (IFGtri: ß = -0.246, P = 0.010, and P.adjust = 0.055). Executive Index Score (MoCA-EIS) was associated with spindle density (IFGtri: ß = 0.238, P = 0.019, and P.adjust = 0.087; parietal: ß = 0.227, P = 0.017, and P.adjust = 0.082) and spindle duration (parietal: ß = -0.230, P = 0.013, and P.adjust = 0.065). Attention Index Score (MoCA-AIS) was associated with spindle duration (IFGtri: ß = -0.233, P = 0.017, and P.adjust = 0.081). Conclusions: The findings suggested that the altered spindle activity in epilepsy with severe cognitive impairment, the associations between the global cognitive status of adult epilepsy and spindle characteristics, and specific cognitive domains may relate to spindle characteristics in particular brain regions.

A novel homozygous missense mutation in L-2-HGA gene: A case report.

L-2-hydroxyglutaric aciduria (L-2-HGA) is a rare autosomal recessive disease resulted from the mutated gene L-2- hydroxyglutarate dehydrogenase (L2HGDH). We presented a female case who inherited the disease from her consanguineous relatives and suffered from cognitive impairment, seizure, and ataxia. Using cerebral magnetic resonance imaging (MRI), urine organic acid test, and high-throughput DNA sequencing, a novel homozygous missense mutation was found in the L2HGDH gene, namely c 0.847 G>A/p. G283R in exon 7. Summarizing the clinical information of the patient with L-2-HGA exhibited to be beneficial for the diagnosis of this rare disease. In summary, the pathogenic missense mutation in the case was reliably confirmed using the bioinformatics analysis.

Median arcuate ligament syndrome with retroperitoneal haemorrhage: A case report.

BACKGROUND: Median arcuate ligament syndrome (MALS) is relatively rare and is due to extraluminal compression of the coeliac artery by the median arcuate ligament of the diaphragm. Here, we report a case of MALS found in a patient with abdominal pain and retroperitoneal haemorrhage for education and dissemination. CASE SUMMARY: This article describes a 46-year-old female patient who was admitted to our hospital with abdominal pain as her chief complaint. She had experienced no obvious symptoms but had retroperitoneal bleeding during the course of the disease. Contrast-enhanced computed tomography (CT) and noninvasive CT angiography (CTA) led to an initial misdiagnosis of pancreaticoduodenal artery aneurysm (PDAA) causing retroperitoneal hemorrhage. After intraoperative exploration and detailed analysis of enhanced CT and CTA images, a final diagnosis of MALS was made. The cause of the haemorrhage was bleeding from a branch of the gastroduodenal artery, not rupture of a PDAA. The prognosis of MALS combined with PDAA treated by laparoscopy and interventional therapy is still acceptable. The patient was temporarily treated by gastroduodenal suture haemostasis and was referred for further treatment. CONCLUSION: MALS is very rare and usually has postprandial abdominal pain, upper abdominal murmur, and weight loss. It is diagnosed by imaging or due to complications. When a patient has abdominal bleeding or PDAA, we should consider whether the patient has celiac trunk stenosis (MALS or other etiology). When abdominal bleeding is combined with an aneurysm, we generally think of aneurysm rupture and hemorrhage first, but it may also be collateral artery rupture and hemorrhage.

A Novel circRNA hsa_circRNA_002178 as a Diagnostic Marker in Hepatocellular Carcinoma Enhances Cell Proliferation, Invasion, and Tumor Growth by Stabilizing SRSF1 Expression.

Background: Previous research studies have shown that the elevation of circular RNA (circRNA), hsa_circRNA_002178, was associated with the poor prognosis of breast cancer and colorectal cancer, while its molecular mechanisms underlying the effects on hepatocellular carcinoma (HCC) are still elusive. Methods: The microarray dataset GSE97332 was obtained from the Gene Expression Omnibus (GEO) database and calculated by using the GEO2R tool to identify differentially expressed circRNAs. Differentially expressed hsa_circRNA_002178, in 7 HCC tissue samples and paracancerous tissues, as well as in HCC cell lines and normal hepatocytes, was checked by RT-qPCR. Cell proliferation, invasion, migration, and epithelial-to-mesenchymal transition (EMT)-related proteins were tested in hsa_circRNA_002178-overexpressed or hsa_circRNA_002178-knocked down HCC cells. Subsequently, we identified whether hsa_circRNA_002178 binds to serine- and arginine-rich splicing factor 1 (SRSF1) and then analyzed their function in regulating HCC cell behavior. The effect on HCC cell xenograft tumor growth was observed by the knockdown of hsa_circRNA_002178 in vivo. Results: GEO2R-based analysis displayed that hsa_circRNA_002178 was upregulated in HCC tissues. Overexpression or knockdown of hsa_circRNA_002178 encouraged or impeded HCC cell proliferation, migration, invasion, and EMT program. Mechanically, hsa_circRNA_002178 bound to SRSF1 3'-untranslated region (UTR) and stabilized its expression. SRSF1 weakening eliminated the effects of pcDNA-hsa_circRNA_002178 on cell malignant behavior. Finally, the knockdown of hsa_circRNA_002178 was confirmed to prevent xenograft tumor growth. Conclusions: hsa_circRNA_002178 overexpression encouraged the stability of SRSF1 mRNA expression, and it may serve as an upstream factor of SRSF1 for the diagnosis of HCC.

Clinical Outcome of CT-Guided Iodine-125 Radioactive Seed Implantation for Intrahepatic Recurrent Hepatocellular Carcinoma: A Retrospective, Multicenter Study.

The efficacy and safety of CT-Guided Iodine-125 Radioactive Seed Implantation (RSI) for the treatment of intrahepatic recurrent hepatocellular carcinoma (rHCC) were analyzed in this multicenter retrospective study. We reviewed the medical records of patients with rHCC treated with I-125 seed implantation at four different hospitals in China from December 2011 and January 2021. The local progression-free survival (LPFS),liver PFS, and overall survival (OS) were calculated, and the short-term efficacy and treatment-related toxicities were evaluated. A total of 82 patients were enrolled; the median follow-up time was 46 months (range, 3-80 months). The 1-, 3- and 5-year LPFS rates were 63.8%, 27.1%, and 7.9%, respectively, and the corresponding OS rates were 74.8%, 32.9%, and 12.6%, respectively. Univariate analysis showed that factors influencing LPFS included the maximum lesion diameter, Barcelona Clinic Liver Cancer (BCLC) stage, interval between treatment and recurrence, and D90. Multivariate analyses revealed that the BCLC stage, interval between treatment and recurrence, and D90 were independent factors influencing LPFS, whereas BCLC stage, D90, and short-term efficacy were independent factors influencing OS. In summary, I-125 seed implantation is a safe and effective treatment for rHCC. The BCLC stage, interval, and D90 were found to influence the local control. A larger, prospective study is required to confirm the dose-response curve for Iodine-125 RSI of rHCC.

Circ_0067934 reduces JNK phosphorylation through a microRNA-545-3p/PPA1 axis to enhance tumorigenesis and cisplatin resistance in ovarian cancer.

BACKGROUND: Circular RNA 0067934 (circ_0067934) has been revealed as a cancer driver in multiple human malignancies, whereas its action in the pathogenesis of ovarian cancer (OC) remains unclear. This study focuses on the function of circ_0067934 in tumorigenesis and cisplatin (DDP) resistance in OC and the molecular mechanism. METHODS: Expression of circ_0067934 in OC tissues and cells was examined, and its correlation with the clinical characteristics of patients was analyzed. Candidate targets of circ_0067934 were predicted using bioinformatics systems. Binding relationships between circ_0067934 and microRNA (miR)-545-3p and between miR-545-3p and inorganic pyrophosphatase 1 (PPA1) were validated via luciferase assays. Gain- and loss-of functions of circ_0067934, miR-545-3p and PPA1 were performed to determine their functions in proliferation, invasion, apoptosis and DDP resistance of OC cells in vitro and in vivo. RESULTS: Circ_0067934 was overexpressed in OC samples and associated with advanced tumor staging and lymph node metastasis. Downregulation of circ_0067934 reduced DDP resistance of the DDP-resistant A2780/DDP cell line and reduced cell proliferation and invasion, but the malignant behaviors of OC cells were restored after further miR-545-3p downregulation. Circ_0067934 served as a sponge for miR-545-3p and diminished its suppressive effect on PPA1 translation. Artificial upregulation of PPA1 enhanced proliferation, invasion and DDP resistance of A2780/DDP cells, and it reduced phosphorylation of the pro-apoptotic JNK signaling. Similar results were found in vivo. CONCLUSION: This study suggests that circ_0067934 sequesters miR-545-3p and enhances PPA1 expression to promote tumorigenesis and DDP resistance in OC. This study may provide novel approaches in the management of OC.

Clinical Outcome of CT-Guided Stereotactic Ablative Brachytherapy for Unresectable Early Non-Small Cell Lung Cancer: A Retrospective, Multicenter Study.

OBJECTIVE: To analyze the efficacy and safety of low dose rate stereotactic ablative brachytherapy (L-SABT) for treatment of unresectable early-stage non-small cell lung cancer (NSCLC). METHODS: Data of patients with early-stage NSCLC who received CT-guided L-SABT (radioactive I-125 seeds implantation) at eight different centers from December 2010 to August 2020 were retrospectively analyzed. Treatment efficacy and complications were evaluated. RESULTS: A total of 99 patients were included in this study. Median follow-up duration was 46.3 months (6.1-119.3 months). The 1-year, 3-year, and 5-year local control rates were 89.1%, 77.5%, and 75.7%, respectively. The 1-year, 3-year, and 5-year overall survival rates were 96.7%, 70.1%, and 54.4%, respectively. Treatment failure occurred in 38.4% of patients. Local/regional recurrence, distant metastasis, and recurrence combined with metastasis accounted for 15.1%, 12.1%, and 11.1%, respectively. Pneumothorax occurred in 47 patients (47.5%) with 19 cases (19.2%) needing closed drainage. The only radiation-related adverse reaction was two cases of grade 2 radiation pneumonia. KPS 80-100, T1, the lesion was located in the left lobe, GTV D90 ≥150 Gy and the distance between the lesion and chest wall was < 1 cm, were associated with better local control (all P < 0.05); on multivariate analysis KPS, GTV D90, and the distance between the lesion and chest wall were independent prognostic factors for local control (all P < 0.05). KPS 80-100, T1, GTV D90 ≥150 Gy, and the distance between the lesion and chest wall was < 1 cm were also associated with better survival (all P < 0.05); on multivariate analysis KPS, T stage, and GTV D90 were independent prognostic factors for survival (all P < 0.05). The incidence of pneumothorax in patients with lesions <1 cm and ≥1cm from the chest wall was 33.3% and 56.7%, respectively, and the differences were statistically significant (P = 0.026). CONCLUSION: L-SABT showed acceptable efficacy in the treatment of unresectable early-stage NSCLC. But the incidence of pneumothorax is high. For patients with T1 stage and lesions <1 cm from the chest wall, it may have better efficacy. Prescription dose greater than 150 Gy may bring better results.

Genome collinearity analysis illuminates the evolution of donkey chromosome 1 and horse chromosome 5 in perissodactyls: A comparative study.

BACKGROUND: It is important to resolve the evolutionary history of species genomes as it has affected both genome organization and chromosomal architecture. The rapid innovation in sequencing technologies and the improvement in assembly algorithms have enabled the creation of highly contiguous genomes. DNA Zoo, a global organization dedicated to animal conservation, offers more than 150 chromosome-length genome assemblies. This database has great potential in the comparative genomics field. RESULTS: Using the donkey (Equus asinus asinus, EAS) genome provided by DNA Zoo as an example, the scaffold N50 length and Benchmarking Universal Single-Copy Ortholog score reached 95.5 Mb and 91.6%, respectively. We identified the cytogenetic nomenclature, corrected the direction of the chromosome-length sequence of the donkey genome, analyzed the genome-wide chromosomal rearrangements between the donkey and horse, and illustrated the evolution of the donkey chromosome 1 and horse chromosome 5 in perissodactyls. CONCLUSIONS: The donkey genome provided by DNA Zoo has relatively good continuity and integrity. Sequence-based comparative genomic analyses are useful for chromosome evolution research. Several previously published chromosome painting results can be used to identify the cytogenetic nomenclature and correct the direction of the chromosome-length sequence of new assemblies. Compared with the horse genome, the donkey chromosomes 1, 4, 20, and X have several obvious inversions, consistent with the results of previous studies. A 4.8 Mb inverted structure was first discovered in the donkey chromosome 25 and plains zebra chromosome 11. We speculate that the inverted structure and the tandem fusion of horse chromosome 31 and 4 are common features of non-caballine equids, which supports the correctness of the existing Equus phylogeny to an extent.

Molecular Reorganization during the Formation of the Human Skin Barrier Studied In Situ.

In vertebrates, skin upholds homeostasis by preventing body water loss. The skin's permeability barrier is located intercellularly in the stratum corneum and consists of stacked lipid lamellae composed of ceramides, cholesterol, and free fatty acids. We have combined cryo-electron microscopy with molecular dynamics modeling and electron microscopy simulation in our analysis of the lamellae's formation, a maturation process beginning in stratum granulosum and ending in stratum corneum. Previously, we have revealed the lipid lamellae's initial- and end-stage molecular organizations. In this study, we reveal two cryo-electron microscopy patterns representing intermediate stages in the lamellae's maturation process: a single-band pattern with 2.0‒2.5 nm periodicity and a two-band pattern with 5.5‒6.0 nm periodicity, which may be derived from lamellar lipid structures with 4.0‒5.0 nm and 5.5‒6.0 nm periodicity, respectively. On the basis of the analysis of the data now available on the four maturation stages identified, we can present a tentative molecular model for the complete skin barrier formation process.

RAB33B recruits the ATG16L1 complex to the phagophore via a noncanonical RAB binding protein.

Autophagosome formation is a fundamental process in macroautophagy/autophagy, a conserved self-eating mechanism in all eukaryotes, which requires the conjugating ATG (autophagy related) protein complex, ATG12-ATG5-ATG16L1 and lipidated MAP1LC3/LC3 (microtubule associated protein 1 light chain 3). How the ATG12-ATG5-ATG16L1 complex is recruited to membranes is not fully understood. Here, we demonstrated that RAB33B plays a key role in recruiting the ATG16L1 complex to phagophores during starvation-induced autophagy. Crystal structures of RAB33B bound to the coiled-coil domain (CCD) of ATG16L1 revealed the recognition mechanism between RAB33B and ATG16L1. ATG16L1 is a novel RAB-binding protein (RBP) that can induce RAB proteins to adopt active conformation without nucleotide exchange. RAB33B and ATG16L1 mutually determined the localization of each other on phagophores. RAB33B-ATG16L1 interaction was required for LC3 lipidation and autophagosome formation. Upon starvation, a fraction of RAB33B translocated from the Golgi to phagophores and recruited the ATG16L1 complex. In this work, we reported a new mechanism for the recruitment of the ATG12-ATG5-ATG16L1 complex to phagophores by RAB33B, which is required for autophagosome formation.Abbreviations: ATG: autophagy-related; Cα: alpha carbon; CCD: coiled-coil domain; CLEM: correlative light and electron microscopy; DTE: dithioerythritol; EBSS: Earle's balanced salt solution; EDTA: ethylenediaminetetraacetic acid; EGFP: enhanced green fluorescent protein; FBS: fetal bovine serum; FLIM: fluorescence lifetime imaging microscopy; FRET: Förster resonance energy transfer; GDP: guanosine diphosphate; GOLGA2/GM130: golgin A2; GppNHp: guanosine 5'-[ß,γ-imido]triphosphate; GST: glutathione S-transferase; GTP: guanosine triphosphate; GTPγS: guanosine 5'-O-[gamma-thio]triphosphate; HA (tag): hemagglutinin (tag); HEK: human embryonic kidney; HeLa: Henrietta Lacks; HEPES: (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid); IgG: immunoglobulin G; Kd: dissociation constant; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCF7: Michigan cancer foundation-7; MEF: mouse embryonic fibroblast; MEM: minimum essential medium Eagle; MST: microscale thermophoresis; NEAA: non-essential amino acids; PBS: phosphate-buffered saline; PE: phosphatidylethanolamine; PtdIns3P: phosphatidylinositol-3-phosphate; RAB: RAS-associated binding; RB1CC1/FIP200: RB1 inducible coiled-coil protein 1; RBP: RAB-binding protein; SD: standard deviation; SDS: sodium dodecyl sulfate; SQSTM1/p62: sequestosome 1; TBS-T: tris-buffered saline-tween 20; WD (repeat): tryptophan-aspartic acid (repeat); WIPI2B: WD repeat domain phosphoinositide interacting 2B; WT: wild type.

Efficacy and safety of montelukast sodium combined with fluticasone in the treatment of adult bronchial asthma: A protocol for systematic review and meta-analysis.

BACKGROUND: Bronchial asthma (BA) is a chronic airway inflammatory disease with reversible airflow limitation as the main clinical manifestations, such as wheezing, cough, shortness of breath, chest tightness, etc, mediated by a variety of inflammatory cells, which can be recurrent. Clinical can improve symptoms, but cannot be cured; glucocorticoid is the most important first-line medication. Clinical practice has shown that montelukast sodium combined with fluticasone in the treatment of adult BA can improve clinical efficacy and reduce adverse reactions. The purpose of this study is to systematically study the efficacy and safety of montelukast sodium combined with fluticasone in the treatment of adult BA. METHODS: The Chinese databases (CNKI, VIP, Wanfang, Chinese Biomedical Database) and English databases (PubMed, the Cochrane Library, Embase, Web of Science) were searched by computer, for the randomized controlled clinical studies of montelukast sodium combined with fluticasone in the treatment of adult BA from establishment of database to October 2020. Two researchers independently extracted the relevant data and evaluated the quality of the literatures, and used RevMan5.3 software to conduct meta-analyze of the included literatures. RESULTS: This study assessed the efficacy and safety of montelukast sodium combined with fluticasone in the treatment of adult BA through total effective rate, pulmonary function (FEV1, FVC, PEF, FEV1/FVC), and adverse reactions. CONCLUSION: This study will provide reliable evidence-based evidence for the clinical application of montelukast sodium combined with fluticasone in the treatment of adult BA. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/CKQFM.

High expression of WSB1 is associated with poor prognosis in hepatocellular carcinoma and affects epithelial-mesenchymal transition.

PURPOSE: To explore the expression of WD-40 repeat and SOCS box containing 1 (WSB1) and its clinical significance in hepatocellular carcinoma (HCC). METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression level of WSB1 in clinical tissues. Survival curves were platted using the Kaplan-Meier method and log rank test was used to compare survival between groups. The influencing factors for the long-term survival were analysed using Cox univariate and multivariate analysis. In in vitro study, Western blot was used to evaluate the effects of WSB1 on epithelial-mesenchymal transition (EMT) of tumor cells. RESULTS: The expression of WSB1 was much higher in cancer tissues than that in para-normal tissues, and the WSB1 expression was correlated with tumor differentiation, lymph node metastasis and clinical stage. In addition, HCC patients with higher WSB1 expression had significantly poorer progression-free survival (PFS) and overall survival (OS). Both univariate analysis and multivariate analyses indicated that high expression of WSB1 was an independent predictor of poor prognosis in HCC. Further in in vitro study, downregulation of WSB1 in HCC cell line could reduce the expression of epithelial phenotype protein (E-cadherin) while increase the expression of mesenchymal phenotype protein (N-cadherin and Vimentin). CONCLUSIONS: WSB1 might contribute into the development of HCC and serve as a clinical biomarker and a therapeutic target for HCC patients.

Active tyrosine phenol-lyase aggregates induced by terminally attached functional peptides in Escherichia coli.

The formation of inclusion bodies (IBs) without enzyme activity in bacterial research is generally undesirable. Researchers have attempted to recovery the enzyme activities of IBs, which are commonly known as active IBs. Tyrosine phenol-lyase (TPL) is an important enzyme that can convert pyruvate and phenol into 3,4-dihydroxyphenyl-L-alanine (L-DOPA) and IBs of TPL can commonly occur. To induce the correct folding and recover the enzyme activity of the IBs, peptides, such as ELK16, DKL6, L6KD, ELP10, ELP20, L6K2, EAK16, 18A, and GFIL16, were fused to the carboxyl terminus of TPL. The results showed that aggregate particles of TPL-DKL6, TPL-ELP10, TPL-EAK16, TPL-18A, and TPL-GFIL16 improved the enzyme activity by 40.9%, 50.7%, 48.9%, 86.6%, and 97.9%, respectively. The peptides TPL-DKL6, TPL-EAK16, TPL-18A, and TPL-GFIL16 displayed significantly improved thermostability compared with TPL. L-DOPA titer of TPL-ELP10, TPL-EAK16, TPL-18A, and TPL-GFIL16, with cells reaching 37.8 g/L, 53.8 g/L, 37.5 g/L, and 29.1 g/L, had an improvement of 111%, 201%, 109%, and 63%, respectively. A higher activity and L-DOPA titer of the TPL-EAK16 could be valuable for its industrial application to biosynthesize L-DOPA.

Regulating the biosynthesis of pyridoxal 5'-phosphate with riboswitch to enhance L-DOPA production by Escherichia coli whole-cell biotransformation.

Pyridoxal 5'-phosphate (PLP) is an essential cofactor that participates in ∼4% enzymatic activities cataloged by the Enzyme Commission. The intracellular level of PLP is usually lower than that demanded in industrial catalysis. To realize the self-supply of PLP cofactor in whole-cell biotransformation, the de novo ribose 5-phosphate (R5P)-dependent PLP synthesis pathway was constructed. The pdxST genes from Bacillus subtilis 168 were introduced into the tyrosine phenol-lyase (TPL)-overexpressing Escherichia coli BL21(DE3) strain. TPL and PdxST were co-expressed with a double-promoter or a compatible double-plasmid system. The 3,4-dihydroxyphenylacetate-L-alanine (L-DOPA) titer did not increase with the increase in the intracellular PLP concentration in these strains with TPL and PdxST co-expression. Therefore, it is necessary to optimize the intracellular PLP metabolism level so as to achieve a higher L-DOPA titer and avoid the formation of L-DOPA-PLP cyclic adducts. The thi riboswitch binds to PLP and forms a complex such that the ribosome cannot have access to the Shine-Dalgarno (SD) sequence. Therefore, this metabolite-sensing regulation system was applied to regulate the translation of pdxST mRNA. Riboswitch was introduced into pET-TPL-pdxST-2 to downregulate the expression of PdxST and biosynthesis of PLP at the translation level by sequestering the ribosome-binding site. As a result, the titer and productivity of L-DOPA using the strain BL21-TPLST-Ribo1 improved to 69.8 g/L and 13.96 g/L/h, respectively, with a catechol conversion of 95.9% and intracellular PLP accumulation of 24.8 µM.

Site-directed mutagenesis to improve the thermostability of tyrosine phenol-lyase.

3,4-Dihydroxyphenyl-L-alanine (L-DOPA) is the most important antiparkinsonian drug, and tyrosine phenol-lyase (TPL)-based enzyme catalysis process is one of the most adopted methods on industrial scale production. TPL activity and stability represent the rate-limiting step in L-DOPA synthesis. Here, 25 TPL mutants were predicted, and two were confirmed as exhibiting the highest L-DOPA production and named E313W and E313M. The L-DOPA production from E313W and E313M was 47.5 g/L and 62.1 g/L, which was 110.2 % and 174.8 % higher, respectively, than that observed from wild-type (WT) TPL. The Km of E313W and E313M showed no apparent decrease, whereas the kcat of E313W and E313M improved by 45.5 % and 36.4 %, respectively, relative to WT TPL. Additionally, E313W and E313M displayed improved thermostability, a higher melting temperature, and enhanced affinity between for pyridoxal-5'-phosphate. Structural analysis of the mutants suggested increased stability of the N-terminal region via enhanced interactions between the mutated residues and H317. Application of these mutants in a substrate fed-batch strategy as whole-cell biocatalysts allows realization of a cost-efficient short fermentation period resulting in high L-DOPA yield.