Roxarsone biotransformation by a nitroreductase and an acetyltransferase in Pseudomonas chlororaphis, a bacterium isolated from soil.

Roxarsone (3-nitro-4-hydroxyphenylarsonic acid, Rox), a widely used organoarsenical feed additive, can enter soils and be further biotransformed into various arsenic species that pose human health and ecological risks. However, the pathway and molecular mechanism of Rox biotransformation by soil microbes are not well studied. Therefore, in this study, we isolated a Rox-transforming bacterium from manure-fertilized soil and identified it as Pseudomonas chlororaphis through morphological analysis and 16S rRNA gene sequencing. Pseudomonas chlororaphis was able to biotransform Rox to 3-amino-4-hydroxyphenylarsonic acid (3-AHPAA), N-acetyl-4-hydroxy-m-arsanilic acid (N-AHPAA), arsenate [As(V)], arsenite [As(III)], and dimethylarsenate [DMAs(V)]. The complete genome of Pseudomonas chlororaphis was sequenced. PcmdaB, encoding a nitroreductase, and PcnhoA, encoding an acetyltransferase, were identified in the genome of Pseudomonas chlororaphis. Expression of PcmdaB and PcnhoA in E. coli Rosetta was shown to confer Rox(III) and 3-AHPAA(III) resistance through Rox nitroreduction and 3-AHPAA acetylation, respectively. The PcMdaB and PcNhoA enzymes were further purified and functionally characterized in vitro. The kinetic data of both PcMdaB and PcNhoA were well fit to the Michaelis-Menten equation, and nitroreduction catalyzed by PcMdaB is the rate-limiting step for Rox transformation. Our results provide new insights into the environmental risk assessment and bioremediation of Rox(V)-contaminated soils.

Tissue-type plasminogen activator (tPA) homozygous Tyr471His mutation associates with thromboembolic disease.

Tissue-type plasminogen activator (tPA) encoded by PLAT is a major mediator that promotes fibrinolysis and prevents thrombosis. Pathogenetic mutations in PLAT associated with venous thromboembolism have rarely been reported. Here, we report the first case of a homozygous point mutation c.1411T>C (p.Y471H) in PLAT leading to thromboembolic events and conduct related functional studies. The corresponding tPA mutant protein (tPA-Y471H) and wild-type tPA (tPA-WT) were synthesized in vitro, and mutant mice (PLATH/H mice) were constructed. The molecular docking and surface plasmon resonance results indicated that the mutation impeded the hydrogen-bonding interactions between the protease domain of tPA and the kringle 4 domain of plasminogen, and the binding affinity of tPA and plasminogen was significantly reduced with a difference of one order of magnitude. mRNA half-life assay showed that the half-life of tPA-Y471H was shortened. The inferior vena cava thrombosis model showed that the rate of venous thrombosis in PLATH/H mice was 80% compared with 53% in wild-type mice. Our data suggested a novel role for the protease domain of tPA in efficient plasminogen activation, and demonstrated that this tPA mutation could reduce the fibrinolysis function of the body and lead to an increased propensity for thrombosis.

Light-Modulated Morphological Transformation of Spiropyran Derivative from Nanosphere to Nanorod.

The construction of tunable morphological systems has important implications for understanding the mechanism of molecular self-assembly. In this study, a spiropyran derivative M1 is reported with light-responsive assembly morphology, which can be tuned from nanosphere to nanorod by ultraviolet light irradiation. The absorption spectra show that M1 molecules are transformed from closed-ring (SP) isomers into open-ring (MC) isomers and start to form H-aggregates with increasing irradiation time. Density functional theory calculations indicate that MC-MC isomers possess stronger binding energy than SP-SP isomers. The MC isomers may thus facilitate the dissociation of the SP-SP aggregates and promote the change of self-assembled morphology with the aid of stronger π-π stackings and dipole-dipole interactions. The research gives an effective method for modulating the morphology of assemblies, with great potential for applications in smart materials.

Inhibitory effect of polyethylene microplastics on roxarsone degradation in soils.

Roxarsone (3-nitro-4-hydroxyphenylarsonic acid, Rox(V)), an extensively used organoarsenical feed additive, enters soils through the application of Rox(V)-containing manure and further degrades to highly toxic arsenicals. Microplastics, as emerging contaminants, are also frequently detected in soils. However, the effects of microplastics on soil Rox(V) degradation are unknown. A microcosm experiment was conducted to investigate soil Rox(V) degradation responses to polyethylene (PE) microplastics and the underlying mechanisms. PE microplastics inhibited soil Rox(V) degradation, with the main products being 3-amino-4-hydroxyphenylarsonic acid [3-AHPAA(V)], N-acetyl-4-hydroxy-m-arsanilic acid [N-AHPAA(V)], arsenate [As(V)], and arsenite [As(III)]. This inhibition was likely driven by the decline in soil pH by PE microplastic addition, which may directly enhance Rox(V) sorption in soils. The decreased soil pH further suppressed the nfnB gene related to nitroreduction of Rox(V) to 3-AHPAA(V) and nhoA gene associated with acetylation of 3-AHPAA(V) to N-AHPAA(V), accompanied by a decrease in the relative abundance of possible Rox(V)-degrading bacteria (e.g., Pseudomonadales), although the diversity, composition, network complexity, and assembly of soil bacterial communities were largely influenced by Rox(V) rather than PE microplastics. Our study emphasizes microplastic-induced inhibition of Rox(V) degradation in soils and the need to consider the role of microplastics in better risk assessment and remediation of Rox(V)-contaminated soils.

Gene editing of human iPSCs rescues thrombophilia in hereditary antithrombin deficiency in mice.

Hereditary antithrombin deficiency is caused by SERPINC1 gene mutations and predisposes to recurrent venous thromboembolism that can be life-threatening. Therefore, lifelong anticoagulation is required, which has side effects and may not be effective. In this study, peripheral blood mononuclear cells from a patient with severe antithrombin deficiency were reprogrammed into induced pluripotent stem cells (iPSCs). The mutation was corrected using CRISPR-Cas9 and Cre/LoxP genome editing. iPSCs were differentiated into hepatocytes, which were injected into the spleen of antithrombin knockout mice to restore the activity of antithrombin and reduce the thrombophilic state. Human iPSC-differentiated hepatocytes colonized mice and secreted antithrombin stably, normalizing antithrombin in plasma (activity: from 46.8 ± 5.7% to 88.6 ± 7.6%, P < 0.0001; antigen: from 146.9 ± 19.5 nanograms per milliliter to 390.7 ± 16.1 nanograms per milliliter, P < 0.0001). In venous thrombosis model, the rate of thrombosis in mice treated with edited hepatocytes, parental hepatocytes, and wild-type mice were 60, 90, and 70%, respectively. The thrombus weight was much lighter in mice treated with edited hepatocytes compared with parental hepatocytes (7.25 ± 2.00 milligrams versus 15.32 ± 2.87 milligrams, P = 0.0025) and showed no notable difference compared with that in wild-type mice (10.41 ± 2.91 milligrams). The activity and concentration of antithrombin remained high for 3 weeks after injection. The liver and kidney function markers showed no obvious abnormality during the observation period. This study provides a proof of principle for correction of mutations in patient-derived iPSCs and potential therapeutic applications for hereditary thrombophilia.

Case Report and Literature Review: Behçet's Disease With a Novel TFPI Gene Mutation.

We report a case of Behçet's disease (BD) with a newly identified tissue factor pathway inhibitor (TFPI) gene mutation. The patient suffered from recurrent deep vein thrombosis and dural sinus thrombosis which could not be relieved by constant anticoagulation therapy. Slight relapsing oral lesion was the initial manifestation of BD but was neglected. Genital ulcers and ocular symptoms were manifest 8-month later than vascular involvement. The patient was diagnosed with BD at last and a novel mutation in TFPI was identified simultaneously. After administration with azathioprine and dexamethasone, the clinical symptoms were quickly gone and no relapse was found during 7-month follow-up.

Epidemiological features for primary lymphoma of the female genital tract patients and development of a nomogram to predict survival.

BACKGROUND: Primary lymphoma of the female genital tract (PLFGT) is a sporadic extranodal lymphoma. Its epidemiology and prognosis are not fully recognized. Our study aimed to construct and validate prognostic nomograms for predicting survival for patients with PLFGT. METHODS: Incidence rate from 1975 to 2017 and patients with PLFGT from 1975 to 2011 in the Surveillance, Epidemiology and End Results (SEER) database were retrospectively reviewed. The nomograms of overall survival (OS) and disease-specific survival (DSS) were established according to the multivariate Cox regression analyses. The concordance index (C-index) and calibration plots were used to demonstrate its robustness and accuracy. RESULTS: A total of 617 PLFGT patients were identified. The overall incidence of PLFGT is 0.437/1,000,000 (adjusted to the US standard population in 2000) from 1975 to 2017. Age, histological subtype, Ann Arbor Stage, and therapeutic strategy were identified as independent prognostic factors for OS and DSS by multivariate Cox regression (p < .05). Nomograms to predict 1-, 5-, and 10-year OS and DSS were established. The C-index and calibration plots showed a good discriminative ability and an optimal accuracy of the nomograms. Patients were divided into three risk groups according to the model of OS. CONCLUSIONS: The incidence of PLFGT has increased in the past 40 years, and the nomograms were developed and validated as an individualized tool to predict OS and DSS for all PLFGT patients and DLBCL patients. All patients are divided into three risk groups to assist clinicians to identify patients at high-risk and choose the optimal individualized treatments for patients.HighlightsThe incident of PLFGT and its subtypes were calculated and compared.Nomograms were constructed to predict the 1-, 5-, and 10-year OS and DSS.Patients are divided into the low-risk, medium-risk, and high-risk according total score of the nomogram.

External fixation in the treatment of displaced supracondylar femoral fracture in children: A retrospective case series.

OBJECTIVE: The aim of this study was to evaluate the clinical and radiological outcomes and complications of external fixation use in the treatment of children with displaced supracondylar femoral fractures.. METHODS: In this retrospective study, 14 children (6 boys, 8 girls; mean age = 7.3 years; age range = 3.9 - 10.3 years) who underwent external fixation for the treatment of a displaced supracondylar femoral fracture from 2010 to 2017 were included. Their medical records were reviewed for general information and surgery details. Postoperative information, such as time to radiographic union, time to regain walking ability, Knee Society Scores (KSS) postoperative score, and KSS functional score were collected. Radiographic images were examined for the measurement of leg length discrepancy and valgus deformity. RESULTS: The mean follow up was 34 (range = 24-50) months. The mean time to radiographic union was 12.3 (range = 10-16) weeks, and the mean time to regain walking ability was 11.8 (range = 11-13) weeks. Leg length discrepancies were all less than 0.8 cm, and valgus deformity was all limited in 10°. The mean KSS postoperative score was 97.5 (range = 93-100), and the mean KSS functional score was 97.1 (range = 90-100). None of the patients exhibited functional deficiency. Neither deep infection nor refracture was detected postoperatively. CONCLUSION: External fixation seems to be an acceptable alternative modality for treatingdisplaced supracondylar femoral fractures in children, with favorable clinical and radiological outcomes as well as a low complication rate. LEVEL OF EVIDENCE: Level IV, Therapeutic Study.

Characterization of a Missense Mutation in the Catalytic Domain and a Splicing Mutation of Coagulation Factor X Compound Heterozygous in a Chinese Pedigree.

BACKGROUND: Congenital coagulation factor X (FX) deficiency is a rare bleeding disorder with an incidence of one in one million caused by mutations in the FX-coding gene(F10), leading to abnormal coagulation activity and a tendency for severe hemorrhage. Therefore, identifying mutations in FX is important for diagnosing congenital FX deficiency. RESULTS: Genetic analysis of the proband identified two single-base substitutions: c.794T > C: p.Ile265Thr and c.865 + 5G > A: IVS7 + 5G > A. His FX activity and antigen levels were < 1% and 49.7%, respectively; aPTT and PT were prolonged to 65.3 and 80.5 s, respectively. Bioinformatics analysis predicted the two novel variants to be pathogenic. In-vitro expression study of the missense mutation c.794T > C: p.Ile265Thr showed normal synthesis and secretion. Activation of FXs by RVV, FVII/TF, and FVIII/FIX all showed no obvious difference between the variant and the reference. However, clotting activity by PT and aPTT assays and activity of thrombin generation in a TGA assay all indicated reduced activity of the mutant FX-Ile265Thr compared to FX-WT. Minigene assay showed a normal splicing mode c.865 + 5G > A: IVS7 + 5G > A, which is inconsistent with clinical phenotype. CONCLUSIONS: The heterozygous variants c.794T > C: p.Ile265Thr or c.865 + 5G > A: IVS7 + 5G > A indicate mild FX deficiency, but the compound heterozygous mutation of the two causes severe congenital FX deficiency. Genetic analysis of these two mutations may help characterize the bleeding tendency and confirm congenital FX deficiency. In-vitro expression and functional study showed that the low activity of the mutant FX-Ile265Thr is caused by decrease in its enzyme activity rather than self-activation. The minigene assay help us explore possible mechanisms of the splicing mutation. However, more in-depth mechanism research is needed in the future.

External fixation and Kirschner wires in the treatment of paediatric displaced supracondylar femur fractures.

PURPOSE: Kirschner-wire fixation (KF) and external fixation (EF) for the treatment of displaced supracondylar femur fractures (SFFs) were demonstrated respectively in previous reports. However, there is no paucity of convincing information on better treatment options for children. The aim of this study was to show results of KF and EF in the treatment of paediatric SFFs according to clinical and radiological outcome. METHODS: A retrospective analysis including 22 displaced closed SFFs was performed. A total of 12 patients were treated with KF, other ten patients were treated with EF. All patients were followed up for at least 24 months. Demographic data, surgical outcomes and postoperative knee function using the Knee Society Score (KSS) scale were evaluated in this research. RESULTS: The patients in the KF group were significantly younger than in the EF group (p < 0.001). The KF group had superiority in operative time (p = 0.001), blood loss (p = 0.027) and length of hospital stay (p = 0.001). Clinical healing outcome did not differ between the two groups. The KF group achieved radiological union in a shorter period (p < 0.001), with a better range of movement (ROM) and KSS postoperative score. CONCLUSION: Both KF and EF can achieve excellent outcomes for paediatric SFFs. KF has many advantages in younger children. LEVEL OF EVIDENCE: IV.

Association between liver cirrhosis and estimated glomerular filtration rates in patients with chronic HBV infection.

To investigate the estimated glomerular filtration rates of chronic hepatitis B (CHB) patients with or without liver cirrhosis, and to explore the related risk factors.A total of 559 CHB patients were enrolled. Liver cirrhosis was diagnosed with ultrasound. The Child-Pugh scoring system was used to stage patients with liver cirrhosis. The Modification of Diet in Renal Disease (MDRD) formula was used to calculate the estimated glomerular filtration rate (eGFR).A total of 296 patients were involved. The results showed that the incidence of renal impairment in patients with liver cirrhosis was 8.45% (25/296). The incidence of renal impairment in Child-Pugh C patients was significantly higher than that in Child-Pugh B and Child-Pugh Grade A patients (i.e., 17.2% [17/99] vs 6.67% [7/105] vs 1.09% [1/92], respectively, P < .001); age, hyperuricemia, and Child-Pugh score are all risk factors for impaired renal function.With the deterioration of liver function in patients with cirrhosis, the incidence of impaired renal function has increased significantly, and renal function should be closely monitored to guide patients in clinical medication.

Locking plate versus external fixation in the treatment of displaced femoral supracondylar fracture in children.

BACKGROUND: Displaced supracondylar femoral fractures (SFF) are difficult injuries to treat in children. Several techniques have been widely used but few studies have compared the merits and drawbacks of each surgical intervention in order to analyze clinical values. The aim of this study was to (1) evaluate postoperative and functional conditions after treatments with locking plate (LP) or external fixation (EF), (2) observe adverse events associated with these two techniques, and (3) evaluate the clinical value of these two techniques. METHODS: Twenty-eight patients less than 14 years of age were included in this study with supracondylar femoral fractures. They underwent locking plate or external fixation in authors' hospital. The postoperative healing and functional outcome were elevated according to radiographic and clinical measures, including American Knee Society Score (KSS). Fisher's exact test and independent samples t test were used for statistical analysis. RESULTS: All fractures healed without delayed union. The KSS scoring results of locking plate and external fixation groups were both excellent. The alignment of lower limbs was acceptable with knee valgus less than 2° for all involved patients. In addition, leg length discrepancy was less than 1 cm. No acute or severe complications were noted. There was significant difference in union time (p = 0.03), operating time (p< 0.001), intraoperative blood loss (p< 0.001), and limb length discrepancy (p = 0.04) between LP group and EF group. CONCLUSIONS: External fixation is superior than locking plate in terms of union, operation time phrases, and intraoperative blood loss. EF techniques are better options for treating displaced supracondylar femoral fracture in children. LEVEL OF EVIDENCE: Retrospective comparative study; level III.

Closed reduction with crossed Kirschner wire fixation for displaced supracondylar femoral fractures in young children.

Supracondylar femoral fractures are uncommon in children, but may result in various deformities. Though many approaches have been used to manage them, there is no literally approved standard yet.From 2015 to 2017, 12 young children at the average age of 2.5 years old (range 3.6-1.6) with displaced supracondylar fractures were admitted to our department and received closed reduction with crossed Kirschner wire (K-wire) fixation as treatment. After the surgeries, we performed follow-up to every patient at the average length of 26 months (range 24-30) and used the Knee Society Score scale to evaluate the outcome.Fracture healing was observed within 10 weeks for all patients and walking was resumed between 10 to 13 weeks. No patient reported a valgus deformity more than 10°, neurovascular injury or knee infection. The average limb length discrepancy was 0.4 cm at the end of our follow-up. Every patient had perfect score on the Knee Society Score scale.Our study suggests that closed reduction with percutaneous crossed Kirschner wire is a favorable method for displaced supracondylar femoral fractures in young children.