Building Streptomyces albus as a chassis for synthesis of bacterial terpenoids.

Terpenoids comprise the most chemically and structurally diverse family of natural products. In contrast to the huge numbers of terpenoids discovered from plants and fungi, only a relatively small number of terpenoids were reported from bacteria. Recent genomic data in bacteria suggest that a large number of biosynthetic gene clusters encoding terpenoids remain uncharacterized. In order to enable the functional characterization of terpene synthase and relevant tailoring enzymes, we selected and optimized an expression system based on a Streptomyces chassis. Through genome mining, 16 distinct bacterial terpene biosynthetic gene clusters were selected and 13 of them were successfully expressed in the Streptomyces chassis, leading to characterization of 11 terpene skeletons including three new ones, representing an ∼80% success rate. In addition, after functional expression of tailoring genes, 18 novel distinct terpenoids were isolated and characterized. This work demonstrates the advantages of a Streptomyces chassis which not only enabled the successful production of bacterial terpene synthases, but also enabled functional expression of tailoring genes, especially P450, for terpenoid modification.

[The Relationship between Occurrence of aGVHD in Patients with Acute Myeloid Leukemia after Allogeneic Hematopoietic Stem Cell Transplantation and Immune Cell Components in Graft].

OBJECTIVE: To explore the relationship between occurrence of acute graft-versus-host disease (aGVHD) and various immune cell composition in patients with acute myeloid leukemia (AML) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). METHODS: The clinical data of 104 patients with AML undergoing allo-HSCT in our hospital were retrospectively analyzed, and the hematopoietic reconstitution and occurrence of GVHD were analyzed. Flow cytometry was used to detect the proportion of various types of immune cells in the grafts, the number of graft composition in patients with different degrees of aGVHD was calculated and compared, and to analyze the correlation between the severity of aGVHD in AML patients after allo-HSCT and the immune cell components in the graft. RESULTS: There was no significant difference in the time of hematopoietic reconstitution between the high number group of total number of nucleated cells (TNC) and the low number group, while the time of neutrophil and platelet reconstruction in the high number of CD34 group was significantly faster than that in the low number of CD34 group (P＜0.05), and the total hospital stay also tends to be shorten. Compared with patients in 0-Ι aGVHD group, both HLA-matched and HLA-haploidentical transplantation, the infusion amounts of CD3+ cells, CD3+CD4+ cells, CD3+CD8+ cells, NK cells and CD14+ monocytes were higher in patients of Ⅱ-Ⅳ aGVHD group， but the difference was not statistically significant (P>0.05); In addition, in patients with HLA-haploidentical transplantation, the number of CD4+CD25+ cells in Ⅱ-Ⅳ aGVHD group was significantly lower than that in 0-Ι aGVHD group (P<0.05), and the same trend was also observed in HLA-matched transplanted patients, but the difference was not significant (P=0.078). CONCLUSION: High number of CD34+ cells in the graft is beneficial to hematopoietic reconstitution in AML patients. To a certain degree, high number of CD3+ cells, CD3+CD4+ cells, CD3+CD8+ cells, NK cells and CD14+ cells tend to increase the occurrence of aGVHD, but high number of CD4+CD25+ regulatory T cells is beneficial to reduce the incidence of aGVHD in AML patients.

Preparation of thioglycerol-modified silica through thiol-epoxy click reaction and its application in HILIC for detection of oligosaccharide in beverages.

A novel preparation scheme of thioglycerol-modified silica through thiol-epoxy click reaction was proposed aiming at introducing additional quantities of hydroxyl groups into the structure. When applied as the stationary phase of hydrophilic interaction liquid chromatography (HILIC) for separation of oligosaccharide compounds, the material revealed higher polar separation capability than which synthesized through traditional thiol-ene click reaction. Hydrogen-bond interactions were speculated to be the predominant retention mechanism, while partitioning also participated in the retention of disaccharides and trisaccharides. The column also showed good stability and inter-batch reproducibility. Finally, the column was employed for determination of oligosaccharide compounds in commercial beverages, and good linearities, high accuracy, favorable precision, satisfactory reproducibility and resistance to matrix interference were achieved. In the detection of real samples, the determined content was consistent with the labeled content. This work provided an efficient and practicable method for quantity monitoring of commercial diet drinks in routine laboratory.

Biosynthesis of Sordarin Revealing a Diels-Alderase for the Formation of the Norbornene Skeleton.

Sordarin (1) is a fungal diterpene glycoside that displays potent antifungal bioactivity through inhibition of elongation factor 2. The structures of sordarin and related compounds feature a highly rearranged tetracyclic diterpene core. In this study, we identified a concise pathway in the biosynthesis of sordarin. A diterpene cyclase (SdnA) generates the 5/8/5 cycloaraneosene framework, which is decorated by a set of P450s that catalyze a series of oxidation reactions, including hydroxylation, desaturation, and C-C bond oxidative cleavage, to give a carboxylate intermediate with a terminal alkene and a cyclopentadiene moiety. A novel Diels-Alderase SdnG catalyzes an intramolecular Diels-Alder (IMDA) reaction on this intermediate to forge the sordarin core structure. Subsequent methyl hydroxylation and glycosylation complete the biosynthesis of sordarin. Our work discloses a new strategy used by nature for the formation of the rearranged diterpene skeleton.

Bacteriological and molecular typing of Clostridium perfringens strains isolated in retail beef in Beijing, China.

Clostridium perfringens is an important zoonotic pathogen. This study was designed to explore the prevalence and toxin types of C. perfringens in retail beef collected from Beijing, China. Among 221 beef samples collected, 53 samples were positive for C. perfringens, resulting in the average prevalence as 23.98%. By toxin gene-based typing, the most C. perfringens strains belong to type A (96.23%, 51/53), only 2 strains were identified as type D. By a multi-locus sequence typing (MLST)-based analysis, a total of 36 sequence types (STs) were detected, and the most STs (n=30) represented just a single strain. These finding suggested that the prevalence of C. perfringens in retail beef in Beijing was considerably high and these bacteria displayed extreme diversity in genetics.

The Role of Imaging Techniques in Management of COVID-19 in China: From Diagnosis to Monitoring and Follow-Up.

In December 2019, an outbreak of coronavirus infection emerged in Wuhan, Hubei Province of China, which is now named Coronavirus Disease 2019 (COVID-19). The outbreak spread rapidly within mainland China and globally. This paper reviews the different imaging modalities used in the diagnosis and treatment process of COVID-19, such as chest radiography, computerized tomography (CT) scan, ultrasound examination, and positron emission tomography (PET/CT) scan. A chest radiograph is not recommended as a first-line imaging modality for COVID-19 infection due to its lack of sensitivity, especially in the early stages of infection. Chest CT imaging is reported to be a more reliable, rapid, and practical method for diagnosis of COVID-19, and it can assess the severity of the disease and follow up the disease time course. Ultrasound, on the other hand, is portable and involves no radiation, and thus can be used in critically ill patients to assess cardiorespiratory function, guide mechanical ventilation, and identify the presence of deep venous thrombosis and secondary pulmonary thromboembolism. Supplementary information can be provided by PET/CT. In the absence of vaccines and treatments for COVID-19, prompt diagnosis and appropriate treatment are essential. Therefore, it is important to exploit the advantages of different imaging modalities in the fight against COVID-19.

Identification of the differentially expressed protein biomarkers in rat blood plasma in response to gamma irradiation.

Purpose: Simple, rapid and high-throughput dose assessment is critical for clinical diagnosis, treatment and emergency intervention in a large-scale radiological accident. The goal of this study is to screen and identify new ionizing radiation-responsive protein biomarkers in rat plasma.Materials and methods: Sprague-Dawley rats were exposed to single doses of 0, 1, 3, 5 Gy of Cobalt-60 γ-rays total body irradiation at a dose rate of 1 Gy/min. The tandem mass tag labeling (TMT) combined with liquid chromatography mass spectrometry (LC-MS/MS) approach was used to screen the differentially expressed proteins in rat plasma collected at 1, 3, 5 and 7 days post-irradiation. Bioinformatics analysis was conducted to explore the biological functions of these proteins. The expression levels of candidate radiation-sensitive protein biomarkers were confirmed using enzyme-linked immune-sorbent assay (ELISA).Results: A total of 503 differentially expressed proteins were identified. Most of these proteins were implicated in immune response, phagocytosis and signal transduction following ionizing radiation. Five up-regulated proteins including alpha-2-macroglobulin (A2m), chromogranin-A (CHGA), glutathione pertidase 3 (GPX3), clusterin (Clu) and ceruloplasmin (Cp) were selected for ELISA analysis. It was found that the expression levels of A2m, CHGA and GPX3 protein were increased in a dose-dependent manner at 1, 3 and 5 days after irradiation.Conclusion: Proteomics analysis revealed radiation-induced differentially expressed proteins in rat plasma. Our results suggested that A2m, CHGA, GPX3 protein expressions alterations in rat plasma may have potential as biomarkers to evaluate radiation exposure.

Potential hepatic and renal toxicity induced by the biflavonoids from Ginkgo biloba.

Evidence continues to grow on potential health risks associated with Ginkgo biloba and its constituents. While biflavonoid is a subclass of the flavonoid family in Ginkgo biloba with a plenty of pharmacological properties, the potential toxicological effects of biflavonoids remains largely unknown. Thus, the aim of this study was to investigate the in vitro and in vivo toxicological effects of the biflavonoids from Ginkgo biloba (i.e., amentoflavone, sciadopitysin, ginkgetin, isoginkgetin, and bilobetin). In the in vitro cytotoxicity test, the five biflavonoids all reduced cell viability in a dose-dependent manner in human renal tubular epithelial cells (HK-2) and human normal hepatocytes (L-02), indicating they might have potential liver and kidney toxicity. In the in vivo experiments, after intragastrical administration of these biflavonoids at 20 mg·kg-1·d-1 for 7 days, serum biochemical analysis and histopathological examinations were performed. The activity of alkaline phosphatase was significantly increased after all the biflavonoid administrations and widespread hydropic degeneration of hepatocytes was observed in ginkgetin or bilobetin-treated mice. Moreover, the five biflavonoids all induced acute kidney injury in treated mice and the main pathological lesions were confirmed to the tubule, glomeruli, and interstitium injuries. As the in vitro and in vivo results suggested that these biflavonoids may be more toxic to the kidney than the liver, we further detected the mechanism of biflavonoids-induced nephrotoxicity. The increased TUNEL-positive cells were detected in kidney tissues of biflavonoids-treated mice, accompanied by elevated expression of proapoptotic protein BAX and unchanged levels of antiapoptotic protein BCL-2, indicating apoptosis was involved in biflavonoids-induced nephrotoxicity. Taken together, our results suggested that the five biflavonoids from Ginkgo biloba may have potential hepatic and renal toxicity and more attentions should be paid to ensure Ginkgo biloba preparations safety.

BCR-ABL enhances the prolyl isomerase activity of Pin 1 by interacting with DAPK1 in ph+ ALL.

Philadelphia chromosome (Ph)/BCR-ABL-positive (ph+ ) ALL is the most common genetic abnormality associated with ALL and has been shown to confer the worst prognosis to both children and adults. Increasing evidence has revealed that the dysregulation of prolyl isomerase Pin 1 contributes to multicancer development and progression, including ALL, although the underlying molecular mechanisms remain unclear. Here, we report that the expression of Pin 1 was enhanced in ph+ ALL patient samples and was associated positively with the expression of BCR-ABL. Genetically or pharmacologically inhibiting Pin 1 expression or activity produces potent therapeutic efficacy against ph+ ALL. We further demonstrated that BCR-ABL enhances the prolyl isomerase activity of Pin 1 by decreasing the phosphorylated level of Pin 1 at Ser 71 and interacting with DAPK1. The inhibition of BCR-ABL activity by imatinib in human ph+ ALL cells reduces the prolyl isomerase activity of Pin 1, further suggesting a key role of the newly identified BCR-ABL-Pin 1 axis in ph+ ALL progression. Thus, the combined suppression of Pin 1 and BCR-ABL proteins may be exploited as an additional target therapy for ph+ ALL.

Sirt6 overexpression suppresses senescence and apoptosis of nucleus pulposus cells by inducing autophagy in a model of intervertebral disc degeneration.

Treatment of intervertebral disc degeneration (IDD) seeks to prevent senescence and death of nucleus pulposus (NP) cells. Previous studies have shown that sirt6 exerts potent anti-senescent and anti-apoptotic effects in models of age-related degenerative disease. However, it is not known whether sirt6 protects against IDD. Here, we explored whether sirt6 influenced IDD. The sirt6 level was reduced in senescent human NP cells. Sirt6 overexpression protected against apoptosis and both replicative and stress-induced premature senescence. Sirt6 also activated NP cell autophagy both in vivo and in vitro. 3-methyladenine (3-MA) and chloroquine (CQ)-mediated inhibition of autophagy partially reversed the anti-senescent and anti-apoptotic effects of sirt6, which regulated the expression of degeneration-associated proteins. In vivo, sirt6 overexpression attenuated IDD. Together, the data showed that sirt6 attenuated cell senescence, and reduced apoptosis, by triggering autophagy that ultimately ameliorated IDD. Thus, sirt6 may be a novel therapeutic target for IDD treatment.

[Beneficial effects of reciprocating gait orthosis on bladder and bowel functions in paraplegia patients].

OBJECTIVE: To evaluate the beneficial effects of application of a self-designed reciprocating gait orthosis (RGO) combined with comprehensive rehabilitation training on bladder and bowel function in paraplegic patients sustaining spinal cord injuries (SCI). METHODS: Twelve paraplegic patients with complete spinal cord injury at the levels between T(4) and L(1) received the RGO with a comprehensive rehabilitation exercise program carried out before and after fixing. Cold water and colon excretion tests were performed before and 3 months after application of the RGO, and the bladder volume, residue bladder volume, and bladder and bowel pressures were measured in these patients. RESULTS: The 12 paraplegic patients all showed positive results in cold water test before and after RGO application. Before RGO application, 11 of these patients were positive for urine bacteria and only 1 patient was still positive 3 months after the application. The bladder volume and bowel pressure of the patients were significantly increased (P<0.05), while the residue volume and pressure of the bladder were significantly reduced 3 months after RGO application (P<0.05). CONCLUSION: RGO combined with comprehensive rehabilitation training can effectively improve the bladder and bowel function and the quality of life of patients with complete spinal cord injury, suggesting much clinical value for its extensive application.

[Infrared radiation and magnetic field therapy ameliorates cartilage damage in rabbits with knee osteoarthritis].

OBJECTIVE: To evaluate the effect of infrared radiation and magnetic field therapy on cartilage damage in rabbits with knee osteoarthritis. METHODS: Knee osteoarthritis was induced in 24 adult New Zealand rabbits by prolonged fixation of the knee joint in extension for 6 weeks. The rabbits were subsequently randomized into control group (without treatment), infrared therapy group, magnetic field therapy group and the combined infrared and magnetic field therapy group. At the end of the first, second and third weeks of the therapy, respectively, 2 rabbits from each group were sacrificed to observe the general changes and histopathology of the condylar cartilage of the femur, and the findings were assessed using Mankin scores. RESULTS: Compared with other groups, the rabbits in the combined therapy group showed significantly milder cartilage damage (including injury of the cartilage surface and chondrocyte's proliferation and disarrangement) with significantly lower Mankin scores (P<0.05). No significant differences were found in the findings between the two groups with exclusive infrared or magnetic field therapy (P>0.1). CONCLUSION: Combined infrared and magnetic field therapy can effectively alleviate cartilage destruction, shortens the disease course and enhance the therapeutic effects in rabbits with knee osteoarthritis.