Effects of an Early Intensive Blood Pressure-lowering Strategy Using Remifentanil and Dexmedetomidine in Patients with Spontaneous Intracerebral Hemorrhage: A Multicenter, Prospective, Superiority, Randomized Controlled Trial.

BACKGROUND: Although it has been established that elevated blood pressure and its variability worsen outcomes in spontaneous intracerebral hemorrhage, antihypertensives use during the acute phase still lacks robust evidence. A blood pressure-lowering regimen using remifentanil and dexmedetomidine might be a reasonable therapeutic option given their analgesic and antisympathetic effects. The objective of this superiority trial was to validate the efficacy and safety of this blood pressure-lowering strategy that uses remifentanil and dexmedetomidine in patients with acute intracerebral hemorrhage. METHODS: In this multicenter, prospective, single-blinded, superiority randomized controlled trial, patients with intracerebral hemorrhage and systolic blood pressure (SBP) 150 mmHg or greater were randomly allocated to the intervention group (a preset protocol with a standard guideline management using remifentanil and dexmedetomidine) or the control group (standard guideline-based management) to receive blood pressure-lowering treatment. The primary outcome was the SBP control rate (less than 140 mmHg) at 1 h posttreatment initiation. Secondary outcomes included blood pressure variability, neurologic function, and clinical outcomes. RESULTS: A total of 338 patients were allocated to the intervention (n = 167) or control group (n = 171). The SBP control rate at 1 h posttreatment initiation in the intervention group was higher than that in controls (101 of 161, 62.7% vs. 66 of 166, 39.8%; difference, 23.2%; 95% CI, 12.4 to 34.1%; P < 0.001). Analysis of secondary outcomes indicated that patients in the intervention group could effectively reduce agitation while achieving lighter sedation, but no improvement in clinical outcomes was observed. Regarding safety, the incidence of bradycardia and respiratory depression was higher in the intervention group. CONCLUSIONS: Among intracerebral hemorrhage patients with a SBP 150 mmHg or greater, a preset protocol using a remifentanil and dexmedetomidine-based standard guideline management significantly increased the SBP control rate at 1 h posttreatment compared with the standard guideline-based management.

Modulation of Rac1/PAK1/connexin43-mediated ATP release from astrocytes contributes to retinal ganglion cell survival in experimental glaucoma.

Connexin43 (Cx43) is a major gap junction protein in glial cells. Mutations have been found in the gap-junction alpha 1 gene encoding Cx43 in glaucomatous human retinas, suggestive of the involvement of Cx43 in the pathogenesis of glaucoma. However, how Cx43 is involved in glaucoma is still unknown. We showed that increased intraocular pressure in a glaucoma mouse model of chronic ocular hypertension (COH) downregulated Cx43, which was mainly expressed in retinal astrocytes. Astrocytes in the optic nerve head where they gather and wrap the axons (optic nerve) of retinal ganglion cells (RGCs) were activated earlier than neurons in COH retinas and the alterations in astrocytes plasticity in the optic nerve caused a reduction in Cx43 expression. A time course showed that reductions of Cx43 expression were correlated with the activation of Rac1, a member of the Rho family. Co-immunoprecipitation assays showed that active Rac1, or the downstream signaling effector PAK1, negatively regulated Cx43 expression, Cx43 hemichannel opening and astrocyte activation. Pharmacological inhibition of Rac1 stimulated Cx43 hemichannel opening and ATP release, and astrocytes were identified to be one of the main sources of ATP. Furthermore, conditional knockout of Rac1 in astrocytes enhanced Cx43 expression and ATP release, and promoted RGC survival by upregulating the adenosine A3 receptor in RGCs. Our study provides new insight into the relationship between Cx43 and glaucoma, and suggests that regulating the interaction between astrocytes and RGCs via the Rac1/PAK1/Cx43/ATP pathway may be used as part of a therapeutic strategy for managing glaucoma.

Charge transfer in SnS2/Na0.9Mg0.45Ti3.55O8 heterojunction in photocatalytic process.

SnS2/Na0.9Mg0.45Ti3.55O8 (SNMTO) composite photocatalyst was synthesized by a hydrothermal method. The chemical combination in lattice scale between SnS2 and Na0.9Mg0.45Ti3.55O8 (NMTO) was observed by high-resolution transmission electron microscopy, indicating that heterojunctions were obtained between SnS2 and NMTO. The photocatalytic activity of SNMTO heterojunctions was improved in comparison with that of pure NMTO and SnS2 for the photocatalytic degradation of methylene blue and Rhodamine B. Electrons were excited in n-type semiconductors NMTO and SnS2 under light illumination, and a part of them moved to the interface, determined with the surface potential reduction observed directly by Kelvin probe force microscopy. The charge redistribution in the composite illustrates a high density of interface states between SnS2 and NMTO, which attract lots of photoelectrons, as a result enhancing the photocatalytic performance. This finding is very different from the speculation that the photogenerated electrons and holes migrate from one part to another because it is difficult for charge carriers to travel through the interface with high energy.

Ventricular function and dyssynchrony in children with a functional single right ventricle using real time three-dimensional echocardiography after fontan operation.

BACKGROUND: This study aimed to evaluate the feasibility and clinical value of real time three-dimensional echocardiography (RT-3DE) for assessing ventricular systolic dysfunction and dyssynchrony in children with an functional single right ventricle (FSRV) having undergone the Fontan procedure. METHODS: Twenty-five children with an FSRV and 25 healthy children were enrolled in our study. RV volume analysis was performed compared with magnetic resonance imaging (MRI) as the reference standard in FSRV patients. The patients were divided into wide and narrow QRS interval groups. Global and regional functions of the RV in three compartments (inflow, body, and outflow) were compared between FSRV and control subjects, including RV systolic dyssynchrony indices of maximal difference of time to minimal volume (Tmsv-Dif), standard deviation of time to minimal volume (Tmsv-SD), maximal difference of time to minimal volume corrected by R-R interval (Tmsv-Dif%), and standard deviation of time to minimal volume corrected by R-R interval (Tmsv-SD%). RESULTS: RT-3DE measurements were significantly lower than MRI measurements for RV-EDV, RV-ESV, RV-SV, and RVEF (p < 0.01).Compared with controls, patients with an FRSV had significantly higher dyssynchrony indices and significantly lower global EF in both narrow QRS interval and wide QRS interval groups. Tmsv-SD% was shown to be most strongly correlated with MRI-RVEF (r = -.570, p = 0.003). CONCLUSIONS: RT-3DE tended to underestimate RV ventricular volume in children with FSRV. Children with an FSRV and either a wide or narrow QRS interval had reduced ventricular function and higher dyssynchrony than normal subjects. Worsening RV dyssynchrony is associated with overall decline in function after the Fontan operation.

Increased risk of Ventriculostomy-Associated hemorrhage in patients treated with antiplatelet agents for stent-assisted coiling of ruptured intracranial aneurysms.

PURPOSE: The aim of this study is to evaluate the impact of antiplatelet agents for stent-assisted coiling, including intravenous (IV) tirofiban as an antiplatelet premedication, on rates of external ventricular drain (EVD)-related hemorrhage in acutely ruptured intracranial aneurysms. The impact of IV tirofiban in particular was also evaluated. METHODS: Rates of radiographically identified hemorrhage associated with EVD placement were compared between patients who received an antiplatelet agent for stent-assisted coil embolization (SACE), and patients who did not receive an antiplatelet agent between June 2013 and June 2019. RESULTS: 78 patients treated for a ruptured aneurysm which required an EVD were included. A total of 46 patients who underwent stent-assisted coiling and received IV tirofiban and oral asipirin and clopidogrel (DAPT) were included in the antiplatelet group, while 32 who underwent single coiling and received no antiplatelet therapy were included in the control group. Overall, EVD-related hemorrhage occurred in 13 patients (16.67%): 11 (23.91%) in the antiplatelet group and 2 (6.25%) in the control group (p = 0.040). Of 37 patients who underwent computed tomography after SACE, but before the use of DAPT, 8 (21.62%) exhibited EVD-related hemorrhage after IV tirofiban therapy (p = 0.070 vs. control group). EVD-related hemorrhage was not significantly different between patients with EVD placement after coil embolization versus before coil embolization (p = 0.124). In the subgroup analysis for the antiplatelet group, we did not observed increased EVD-related hemorrhage in patients receiving EVD placement after administration of antiplatelet agents (8/27 [29.63%]) versus before administration of antiplatelet agents (3/19 [15.79%]). CONCLUSION: Patients with ruptured aneurysm who receive an antiplatelet agent for stent-assisted coiling are at a higher risk for EVD-related hemorrhage. The order of EVD placement and EVT, as well as the order of EVD placement and antiplatelet initiation do not appear to be significantly different regarding the outcome of EVD-related hemorrhage.HighlightsPatients with ruptured aneurysm who receive an antiplatelet agent for stent-assisted coiling are at a higher risk for EVD-related hemorrhage.There was a trend towards higher EVD related haemorrhage when tirofiban was used but it did not reach statisitical significance.The order of EVD-whether before vs after endovascular treatment, or before vs after antiplatelet therapy did not influence the EVD-related hemorrhage rates.

[Advances in Imaging Genetics of Suicidal Behavior].

Suicide,a major public health problem,is the death caused by injuring oneself with the intent to die.In this paper,we reviewed the genes encoding serotonin system,calcium voltage-gated channel subunit alpha1 C,γ-aminobutyric acid,and spindle and kinetochore associated complex subunit 2,as well as their related brain regions,from the perspective of imaging genetics,aiming to provide new ideas for the research and intervention on suicidal behavior.

TSPO ligands prevent the proliferation of vascular smooth muscle cells and attenuate neointima formation through AMPK activation.

Abnormal growth of the intimal layer of blood vessels (neointima formation) contributes to the progression of atherosclerosis and in-stent restenosis. Recent evidence shows that the 18-kDa translocator protein (TSPO), a mitochondrial membrane protein, is involved in diverse cardiovascular diseases. In this study we investigated the role of endogenous TSPO in neointima formation after angioplasty in vitro and in vivo. We established a vascular injury model in vitro by using platelet-derived growth factor-BB (PDGF-BB) to stimulate rat thoracic aortic smooth muscle cells (A10 cells). We found that treatment with PDGF-BB (1-20 ng/mL) dose-dependently increased TSPO expression in A10 cells, which was blocked in the presence of PKC inhibitor or MAPK inhibitor. Overexpression of TSPO significantly promoted the proliferation and migration in A10 cells, whereas downregulation of TSPO expression by siRNA or treatment with TSPO ligands PK11195 or Ro5-4864 (104 nM) produced the opposite effects. Furthermore, we found that PK11195 (10-104 nM) dose-dependently activated AMPK in A10 cells. PK11195-induced inhibition on the proliferation and migration of PDGF-BB-treated A10 cells were abolished by compound C (an AMPK-specific inhibitor, 103 nM). In rats with balloon-injured carotid arteries, TSPO expression was markedly upregulated in the carotid arteries. Administration of PK11195 (3 mg/kg every 3 days, ip), starting from the initial balloon injury and lasting for 2 weeks, greatly attenuated carotid neointima formation by suppressing balloon injury-induced phenotype switching of VSMCs (increased α-SMA expression). These results suggest that TSPO is a vascular injury-response molecule that promotes VSMC proliferation and migration and is responsible for the neointima formation after vascular injury, which provides a novel therapeutic target for various cardiovascular diseases including atherosclerosis and restenosis.

Trinickia dinghuensis sp. nov. and Trinickia fusca sp. nov., isolated from forest soil.

Two Gram-stain-negative, aerobic, non-spore forming and rod-shaped bacterial strains, designated DHOM06T and 7MK8-2T, were isolated from forest soil sampled at Dinghushan Biosphere Reserve, Guangdong Province, PR China. Strain DHOM06T grew at 12-37 °C (optimum, 28-33 °C), pH 4.5-7.5 (pH 5.5) and in the presence of 0-0.5 % NaCl (w/v); while strain 7MK8-2T grew at 12-42 °C (28-33 °C), pH 4.0-8.5 (pH 4.5-5.5) and in the presence of 0-1.0 % NaCl (w/v). Strains DHOM06T and 7MK8-2T each has a 16S rRNA gene sequence similarity of 97.1-98.9 % as well as 97.4-97.9 % to Trinickia strains, respectively. In the 16S rRNA gene sequence phylogram, both strains and all five currently described Trinickia species formed a clade but they were all distinct from each other. The average nucleotide identity and digital DNA-DNA hybridization values for strains DHOM06T and 7MK8-2T and all Trinickia species were in the range of 77.4-82.6 % and 21.7-26.2 %, respectively. The DNA G+C content of DHOM06T and 7MK8-2T was 63.2 and 63.5 mol%, respectively, based on total genome calculations. These two strains contained ubiquinone 8 as the major respiratory quinone and C16 : 0, C17 : 0cyclo, C19 : 0cyclo ω8c, summed feature 3 (C16 : 1ω7c/C16 : 1ω6c) and summed feature 8 (C18 : 1ω7c/C18 : 1ω6c) as the major cellular fatty acids. The major polar lipids of DHOM06T and 7MK8-2T were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. On the basis of phenotypic, chemotaxonomic, phylogenetic and genomic analysis data, strains DHOM06T and 7MK8-2T represent two novel species of the genus Trinickia, for which the names Trinickia dinghuensis sp. nov. (type strain DHOM06T=GDMCC 1.1280T=LMG 30259T) and Trinickia fusca sp. nov. (type strain 7MK8-2T=GDMCC 1.1449T=KCTC 62469T) are proposed.

Temperature and humidity index (THI)-induced rumen bacterial community changes in goats.

The rumen microbiome is thought to play an important role in maintaining normal gastrointestinal metabolism and nutrient absorption in ruminants. The present study was designed to investigate the effect of heat stress on the rumen microbiome of goats using 16S rRNA sequencing technology. Six female goats were randomly allocated into two control metabolic chambers: A and B (in which the temperature and humidity could be precisely controlled with a precision deviation of ± 0.5 °C and ± 5%, with three goats/chamber). Dynamic changes in the rumen bacterial community were detected under 16 gradually increasing temperature and humidity indexes (THIs). Heat stress had no significant effect on alpha diversity but affected the main phyla and genera of the goat rumen microbiota. With a deeper level of heat stress, the TH groups formed a distinct cluster that differed from that of the control check (CK) group. The dominant phylum transitioned from Firmicutes to Bacteroidetes, and co-exclusion occurred between these two phyla. With the increase in THI, the content of probiotics in the Lachnospiraceae_ND3007_group (P < 0.05) decreased, and the abundance of pathogenic bacteria, such as Erysipelotrichaceae_UCG-004 and Treponema_2, increased; however, the difference between the groups was not significant (P > 0.05). Phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt) was used to predict bacterial function, and we found that the ambient environment significantly affected the balance between carbohydrate and energy metabolism (P < 0.05). In conclusion, heat stress changed the composition of rumen microbes and affected metabolic function. This experiment provides a theoretical basis for exploring the effects of environmental factors on the rumen of goats.

Paraburkholderia caseinilytica sp. nov., isolated from the pine and broad-leaf mixed forest soil.

A novel Gram-stain-negative, aerobic, non-spore-forming, motile and rod-shaped bacterial strain, designated HM451T, was isolated from forest soil sampled at the Dinghushan Biosphere Reserve, Guangdong Province, PR China (112° 31' E 23° 10' N). It grew optimally at 28 °C, pH 5.0-6.0 and in the presence of 0-2.5 % (w/v) NaCl on R2A medium. Strain HM451T was closely related to Paraburkholderia mimosarum NBRC 106338T (98.6 % 16S rRNA gene sequence similarity), Paraburkholderia heleia NBRC 101817T (98.4 %) and Paraburkholderia silvatlantica SRMrh-20T (98.0 %). The 16S rRNA gene sequence analysis showed that strain HM451T and the three closely related strains formed a clade within the genus Paraburkholderia, but was clearly separated from the established species. The DNA-DNA relatedness value between strain HM451T and its phylogenetically closest relative, P. mimosarum NBRC 106338T, was much lower than 70 %. Strain HM451T contained ubiquinone 8 as the major respiratory quinone. Major fatty acids were C16 : 0, C17 : 0cyclo and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). The DNA G+C content of strain HM451T was 65.4 mol%. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, two unidentified aminophospholipids, one unidentified aminolipid and a polar lipid. The phenotypic, chemotaxonomic and phylogenetic data showed that strain HM451T represents a novel species of the genus Paraburkholderia, for which the name Paraburkholderia caseinilytica sp. nov. is proposed. The type strain is HM451T (=GDMCC 1.1190T=LMG 30092T).

An unusual case of ectopic corticotrophin-releasing hormone syndrome caused by an adrenal noncatecholamine-secreting pheochromocytoma: a case report.

BACKGROUND: Pheochromocytoma, especially for noncatecholamine-secreting pheochromocytoma, is an extremely rare cause of ectopic corticotrophin-releasing hormone (CRH) syndrome. CASE PRESENTATION: A 27-year-old Chinese woman was administered dexamethasone for a skin allergy, but her general condition rapidly deteriorated over a month. She was subsequently hospitalized for typical clinical features of Cushing's syndrome. Endocrinological investigation confirmed severe hypercortisolism along with elevated plasma adrenocorticotropin hormone (ACTH). However, magnetic resonance imaging (MRI) revealed no pituitary adenoma. Abdominal contrast-enhanced computed tomography (CT) revealed a 6.5 cm heterogeneous right adrenal mass with mildly contrast enhancement. The tumor was found during a routine physical check-up at a local hospital 16 months ago; however, the patient did not have any symptoms and did not seek further medical attention at that time. Laparoscopic resection of the right adrenal tumor led to a rapid remission of Cushing's syndrome. Based on pathological findings and the presence of normal catecholamine metabolites in her serum and urine, the patient was diagnosed with noncatecholamine-secreting pheochromocytoma. Immunohistochemical staining of the adrenal tumor revealed positive staining for CRH and negative staining for ACTH. CONCLUSIONS: This is an extremely rare case of ectopic CRH syndrome caused by an adrenal noncatecholamine-secreting pheochromocytoma. Both ectopic ACTH syndrome and ectopic CRH syndrome should be considered in patients presenting with ACTH-dependent Cushing's syndrome caused by extrapituitary diseases.

Reduced glutathione alleviates tunicamycin-induced endoplasmic reticulum stress in mouse preimplantation embryos.

Endoplasmic reticulum (ER) stress, a dysfunction in protein-folding capacity, is involved in many pathological and physiological responses, including embryonic development. This study aims to determine the developmental competence, apoptosis, and stress-induced gene expression in mouse preimplantation embryos grown in an in vitro culture medium supplemented with different concentrations of the ER stress inducer tunicamycin (TM) and the antioxidant glutathione (GSH). Treatment of zygotes with 0.5 µg/ml TM significantly decreased (P < 0.05) the rate of blastocyst formation, whereas 1 mM GSH supplementation improved the developmental rate of blastocysts. Furthermore, TM treatment significantly increased (P < 0.05) the apoptotic index and reduced the total number of cells, whereas GSH significantly increased the total number of cells and decreased the apoptotic index. The expression levels of ER chaperones, including immunoglobulin-binding protein, activating transcription factor 6, double-stranded activated protein kinase-like ER kinase, activating transcription factor 4, and C/EBP homologous protein were significantly increased (P < 0.05) by TM, but significantly decreased (P < 0.05) by GSH treatment. A similar pattern was observed in the case of the pro-apoptotic gene, B cell lymphoma-associated X protein. The expression level of the anti-apoptotic gene B cell lymphoma 2, was decreased by TM, but significantly increased after co-treatment with GSH. In conclusion, GSH improves the developmental potential of mouse embryos and significantly alleviates ER stress.

Postmortem Analysis of 4 Mutation Hotspots of KCNQ1, KCNH2, and SCN5A Genes in Sudden Unexplained Death in Southwest of China.

Long QT syndrome (LQTS) is known to be involved in some sudden unexplained death (SUD) cases. To make clear whether the pathogenic genes of LQTS are involved in SUD in Yunnan province, southwest of China, we examined 4 mutation hotspot segments of KCNQ1, KCNH2, and SCN5A genes in 83 SUD cases using polymerase chain reaction and direct DNA sequencing. Genomic DNA was extracted from paraffin-embedded tissues in 83 cases of sudden cardiac death. One novel homozygous missense variant was identified in exon 3 of KCNQ1, c. 575G>T (p.R192L) in one case. One novel heterozygous missense variant was identified in exon 7 of KCNH2, c.1789T>A (p.Y597N) in 1 case. One novel heterozygous missense variant was identified in exon 7 of KCNH2, c.1800C>A (p.S600R) in 9 cases. In addition, 18 individuals were found to have heterozygous missense variant in exon 7 of KCNH2, c.1801G>A (p.G601S). Our study suggests that some SUDs in Yunnan province may be related with the pathogenic genes of LQTS.

Epitaxial Growth of Single Layer Blue Phosphorus: A New Phase of Two-Dimensional Phosphorus.

Blue phosphorus, a previously unknown phase of phosphorus, has been recently predicted by theoretical calculations and shares its layered structure and high stability with black phosphorus, a rapidly rising two-dimensional material. Here, we report a molecular beam epitaxial growth of single layer blue phosphorus on Au(111) by using black phosphorus as precursor, through the combination of in situ low temperature scanning tunneling microscopy and density functional theory calculation. The structure of the as-grown single layer blue phosphorus on Au(111) is explained with a (4 × 4) blue phosphorus unit cell coinciding with a (5 × 5) Au(111) unit cell, and this is verified by the theoretical calculations. The electronic bandgap of single layer blue phosphorus on Au(111) is determined to be 1.10 eV by scanning tunneling spectroscopy measurement. The realization of epitaxial growth of large-scale and high quality atomic-layered blue phosphorus can enable the rapid development of novel electronic and optoelectronic devices based on this emerging two-dimensional material.

Expression and Function of Long Non-coding RNA CASC19 in Colorectal Cancer.

Objective To investigate the expression, function and significance of long non-coding RNA (lncRNA) CASC19 in colorectal cancer (CRC). Methods Real-time quantitative polymerase chain reaction was employed to determine the expression of CASC19 in 40 paired samples from CRC surgical specimens and 5 CRC cell lines. The correlations of CASC19 expression with clinicopathologic parameters were analyzed. Transwell assay was applied to detect the migration ability of CRC cells after the CASC19 expression was knocked down by small interfering RNA. Results The expression of CASC19 in colorectal cancer was significantly higher than those in adjacent normal mucosa tissues (t=5.527, P<0.000 1) and was associated with metastasis (P=0.044). Knockdown of CASC19 expression in CRC inhibited the migration ability of CRC in vitro. Conclusions The expression of CASC19 increases in CRC. CASC19 expression is not associated with age, gender, or tumor site/differentiation but with tumor size, lymph node metastasis, and distant metastasis, suggesting high CASC19 expression may promote CRC metastasis.

Maturation of Fibrinolytic Bacillopeptidase F Involves both Hetero- and Autocatalytic Processes.

Bacillopeptidase F (Bpr) is a fibrinolytic serine protease produced by Bacillus subtilis. Its precursor is composed of a signal peptide, an N-terminal propeptide, a catalytic domain, and a long C-terminal extension (CTE). Several active forms of Bpr have been previously reported, but little is known about the maturation of this enzyme. Here, a gene encoding a Bpr (BprL) was cloned from B. subtilis LZW and expressed in B. subtilis WB700, and three fibrinolytic mature forms with apparent molecular masses of 45, 75, and 85 kDa were identified in the culture supernatant. After treatment with urea, the 75-kDa mature form had the same molecular mass as the 85-kDa mature form, from which we infer that they adopt different conformations. Mutational analysis revealed that while the 85-kDa mature form is generated via heterocatalytic processing of a BprL proform by an unidentified protease of B. subtilis, the production of the 75- and 45-kDa mature forms involves both hetero- and autocatalytic events. From in vitro analysis of BprL and its sequential C-terminal truncation variants, it appears that partial removal of the CTE is required for the initiation of autoprocessing of the N-terminal propeptide, which is composed of a core domain (N*) and a 15-residue linker peptide, thereby yielding the 45-kDa mature form. These data suggest that the differential processing of BprL, either heterocatalytically or autocatalytically, leads to the formation of multiple mature forms with different molecular masses or conformations.

Autocatalytic activation of a thermostable glutamyl endopeptidase capable of hydrolyzing proteins at high temperatures.

Glutamyl endopeptidases (GSEs) specifically hydrolyze peptide bonds formed by α-carboxyl groups of Glu and Asp residues. We cloned the gene for a thermophilic GSE (designated TS-GSE) from Thermoactinomyces sp. CDF. A proform of TS-GSE that contained a 61-amino acid N-terminal propeptide and a 218-amino acid mature domain was produced in Escherichia coli. We found that the proform possessed two processing sites and was capable of autocatalytic activation via multiple pathways. The N-terminal propeptide could be autoprocessed at the Glu-1-Ser1 bond to directly generate the mature enzyme. It could also be autoprocessed at the Glu-12-Lys-11 bond to yield an intermediate, which was then converted into the mature form after removal of the remaining part of the propeptide. The segment surrounding the two processing sites was flexible, which allowed the proform and the intermediate form to be trans-processed into the mature form by either active TS-GSE or heterogeneous proteases. Deletion analysis revealed that the N-terminal propeptide is important for the correct folding and maturation of TS-GSE. The propeptide, even its last 11-amino acid peptide segment, could inhibit the activity of its cognate mature domain. The mature TS-GSE displayed a temperature optimum of 85 °C and retained approximately 90 % of its original activity after incubation at 70 °C for 6 h, representing the most thermostable GSE reported to date. Mutational analysis suggested that the disulfide bonds Cys32-Cys48 and Cys180-Cys183 cumulatively contributed to the thermostability of TS-GSE.

Ventricular Twisting and Dyssynchrony in Children with Single Left Ventricle Using Three-Dimensional Speckle Tracking Imaging after the Fontan Operation.

BACKGROUND: The importance of left ventricular (LV) twisting has been recognized in various types of heart disease, but no studies have investigated twisting of functional single ventricle using echocardiography. This study aimed to evaluate LV twisting and dyssynchrony of children with single left ventricle (SLV) after the Fontan operation and explore the relationship between twisting motion and ventricular contractility using three-dimensional speckle tracking imaging (3DSTI). METHODS: Thirty-five children with SLV and 35 healthy children (controls) were enrolled. The patients were divided into wide and narrow QRS groups according to the QRS interval. Atrioventricular valve inflow velocity and tissue Doppler imaging velocity were obtained, and the Tei index was calculated. Apical rotation, basal rotation, twist, torsion, time to peak apical rotation, time to peak basal rotation, time to peak twist, apical-basal rotation delay, and the systolic dyssynchrony index (SDI) were measured by 3DSTI. RESULTS: Patients with SLV had significantly lower apical rotation (2.50 ± 2.25° vs. 4.85 ± 2.68°, P < 0.001), basal rotation (-3.46 ± 3.11° vs. -7.76 ± 2.11°, P < 0.001), twist (5.15 ± 4.75° vs. 12.19 ± 3.65°, P < 0.001), and torsion (1.04 ± 0.99°/cm vs. 2.37 ± 0.77°/cm, P < 0.001) compared to controls. Time to peak basal rotation, apical-basal rotation delay, and time to peak twist were significantly longer in patients. Apical rotation was significantly lower in the wide QRS group but similar in the narrow QRS group as compared to controls. Time to peak twist and apical-basal delay were significantly longer in the wide QRS group in contrast to the similar time in the narrow QRS group compared with controls. Among these twisting parameters, twist and torsion were most significantly correlated with left ventricular ejection fraction (LVEF), the Tei index, and SDI. Twist and age were significantly correlated. CONCLUSION: Twisting is reduced in children with SLV after the Fontan operation. Torsion is a good indicator of LV global function because of good reproducibility and its lack of association with age.

Degradation of intact chicken feathers by Thermoactinomyces sp. CDF and characterization of its keratinolytic protease.

Thermoactinomyces is known for its resistance to extreme environmental conditions and its ability to digest a wide range of hard-to-degrade compounds. Here, Thermoactinomyces sp. strain CDF isolated from soil was found to completely degrade intact chicken feathers at 55 °C, with the resulting degradation products sufficient to support growth as the primary source of both carbon and nitrogen. Although feathers were not essential for the expression of keratinase, the use of this substrate led to a further 50-300 % increase in enzyme production level under different nutrition conditions, with extracellular keratinolytic activity reaching its highest level (∼400 U/mL) during the late-log phase. Full degradation of feathers required the presence of living cells, which are thought to supply reducing agents necessary for the cleavage of keratin disulfide bonds. Direct contact between the hyphae and substrate may enhance the reducing power and protease concentrations present in the local microenvironment, thereby facilitating keratin degradation. The gene encoding the major keratinolytic protease (protease C2) of strain CDF was cloned, revealing an amino acid sequence identical to that of subtilisin-like E79 protease from Thermoactinomyces sp. E79, albeit with significant differences in the upstream flanking region. Exogenous expression of protease C2 in Escherichia coli resulted in the production of inclusion bodies with proteolytic activity, which could be solubilized to an alkaline solution to produce mature protease C2. Purified protease C2 was able to efficiently hydrolyze α- and ß-keratins at 60-80 °C and pH 11.0, representing a promising candidate for enzymatic processing of hard-to-degrade proteins such as keratinous wastes.