ABSTRACT
Microbial instability is a common problem during bio-production based on microbial hosts. Halomonas bluephagenesis has been developed as a chassis for next generation industrial biotechnology (NGIB) under open and unsterile conditions. However, the hidden genomic information and peculiar metabolism have significantly hampered its deep exploitation for cell-factory engineering. Based on the freshly completed genome sequence of H. bluephagenesis TD01, which reveals 1889 biological process-associated genes grouped into 84 GO-slim terms. An enzyme constrained genome-scale metabolic model Halo-ecGEM was constructed, which showed strong ability to simulate fed-batch fermentations. A visible salt-stress responsive landscape was achieved by combining GO-slim term enrichment and CVT-based omics profiling, demonstrating that cells deploy most of the protein resources by force to support the essential activity of translation and protein metabolism when exposed to salt stress. Under the guidance of Halo-ecGEM, eight transposases were deleted, leading to a significantly enhanced stability for its growth and bioproduction of various polyhydroxyalkanoates (PHA) including 3-hydroxybutyrate (3HB) homopolymer PHB, 3HB and 3-hydroxyvalerate (3HV) copolymer PHBV, as well as 3HB and 4-hydroxyvalerate (4HB) copolymer P34HB. This study sheds new light on the metabolic characteristics and stress-response landscape of H. bluephagenesis, achieving for the first time to construct a long-term growth stable chassis for industrial applications. For the first time, it was demonstrated that genome encoded transposons are the reason for microbial instability during growth in flasks and fermentors.
Subject(s)
Halomonas , Halomonas/genetics , Halomonas/metabolism , Halomonas/enzymology , Halomonas/growth & development , Metabolic Engineering , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Metabolic Networks and Pathways/genetics , Gene Deletion , Models, BiologicalABSTRACT
Articular cartilage defects are among the most common orthopedic diseases, which seriously affect patients' health and daily activities, without prompt treatment. The repair biocarrier-based treatment has shown great promise. Total joint injection and open surgery are two main methods to deliver functional repair biocarriers into the knee joint. However, the exhibited drawbacks of these methods hinder their utility. The repair effect of total joint injection is unstable due to the low targeting rate of the repair biocarriers, whereas open surgery causes serious trauma to patients, thereby prolonging the postoperative healing time. In this study, we develop a magnetic soft catheter robot (MSCR) system to perform precise in situ repair of articular cartilage defects with minimal incision. The MSCR processes a size of millimeters, allowing it to enter the joint cavity through a tiny skin incision to reduce postoperative trauma. Meanwhile, a hybrid control strategy combining neural network and visual servo is applied to sequentially complete the coarse and fine positioning of the MSCR on the cartilage defect sites. After reaching the target, the photosensitive hydrogel is injected and anchored into the defect sites through the MSCR, ultimately completing the in situ cartilage repair. The in vitro and ex vivo experiments were conducted on a 3D printed human femur model and an isolated porcine femur, respectively, to demonstrate the potential of our system for the articular cartilage repair.
ABSTRACT
A novel α-amylase Amy03713 was screened and cloned from the starch utilization strain Vibrio alginolyticus LHF01. When heterologously expressed in Escherichia coli, Amy03713 exhibited the highest enzyme activity at 45 °C and pH 7, maintained >50 % of the enzyme activity in the range of 25-75 °C and pH 5-9, and sustained >80 % of the enzyme activity in 25 % (w/v) of NaCl solution, thus showing a wide range of adapted temperatures, pH, and salt concentrations. Halomonas bluephagenesis harboring amy03713 gene was able to directly utilize starch. With optimized amylase expression, H. bluephagenesis could produce poly(3-hydroxybutyrate) (PHB), poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P34HB). When cultured for PHB production, recombinant H. bluephagenesis was able to grow up to a cell dry weight of 11.26 g/L, achieving a PHB titer of 6.32 g/L, which is the highest titer that has been reported for PHB production from starch in shake flasks. This study suggests that Amy03713 is an ideal amylase for PHA production using starch as the carbon source in H. bluephagenesis.
Subject(s)
Halomonas , Pentanoic Acids , Polyhydroxyalkanoates , Halomonas/genetics , Halomonas/metabolism , Carbon/metabolism , Starch/metabolism , Hydroxybutyrates/metabolism , alpha-Amylases/genetics , alpha-Amylases/metabolism , Polyesters/metabolismABSTRACT
Long-term chronic inflammation after Achilles tendon injury is critical for tendinopathy. Platelet-rich plasma (PRP) injection, which is a common method for treating tendinopathy, has positive effects on tendon repair. In addition, tendon-derived stem cells (TDSCs), which are stem cells located in tendons, play a major role in maintaining tissue homeostasis and postinjury repair. In this study, injectable gelatine methacryloyl (GelMA) microparticles containing PRP laden with TDSCs (PRP-TDSC-GM) were prepared by a projection-based 3D bioprinting technique. Our results showed that PRP-TDSC-GM could promote tendon differentiation in TDSCs and reduce the inflammatory response by downregulating the PI3K-AKT pathway, thus promoting the structural and functional repair of tendons in vivo.
Subject(s)
Platelet-Rich Plasma , Tendinopathy , Rats , Animals , Hydrogels/pharmacology , Phosphatidylinositol 3-Kinases/metabolism , Tendons , Tendinopathy/therapy , Tendinopathy/metabolism , Stem Cells , Platelet-Rich Plasma/metabolism , Printing, Three-DimensionalABSTRACT
Glutathione S-transferase P1(GSTP1) is known for its transferase and detoxification activity. Based on disease-phenotype genetic associations, we found that GSTP1 might be associated with bone mineral density through Mendelian randomization analysis. Therefore, this study was performed both in vitro cellular and in vivo mouse model to determine how GSTP1 affects bone homeostasis. In our research, GSTP1 was revealed to upregulate the S-glutathionylation level of Pik3r1 through Cys498 and Cys670, thereby decreasing its phosphorylation, further controlling the alteration of autophagic flux via the Pik3r1-AKT-mTOR axis, and lastly altering osteoclast formation in vitro. In addition, knockdown and overexpression of GSTP1 in vivo also altered bone loss outcomes in the OVX mice model. In general, this study identified a new mechanism by which GSTP1 regulates osteoclastogenesis, and it is evident that the cell fate of osteoclasts is controlled by GSTP1-mediated S-glutathionylation via a redox-autophagy cascade.
Subject(s)
Glutathione Transferase , Osteogenesis , Animals , Mice , Phosphorylation , Transcription Factors , Autophagy , Oxidation-ReductionABSTRACT
Tendon-bone insertion (TBI) injuries, such as anterior cruciate ligament injury and rotator cuff injury, are the most common soft tissue injuries. In most situations, surgical tendon/ligament reconstruction is necessary for treating such injuries. However, a significant number of cases failed because healing of the enthesis occurs through scar tissue formation rather than the regeneration of transitional tissue. In recent years, the therapeutic potential of mesenchymal stem cells (MSCs) has been well documented in animal and clinical studies, such as chronic paraplegia, non-ischemic heart failure, and osteoarthritis of the knee. MSCs are multipotent stem cells, which have self-renewability and the ability to differentiate into a wide variety of cells such as chondrocytes, osteoblasts, and adipocytes. Numerous studies have suggested that MSCs could promote angiogenesis and cell proliferation, reduce inflammation, and produce a large number of bioactive molecules involved in the repair. These effects are likely mediated by the paracrine mechanisms of MSCs, particularly through the release of exosomes. Exosomes, nano-sized extracellular vesicles (EVs) with a lipid bilayer and a membrane structure, are naturally released by various cell types. They play an essential role in intercellular communication by transferring bioactive lipids, proteins, and nucleic acids, such as mRNAs and miRNAs, between cells to influence the physiological and pathological processes of recipient cells. Exosomes have been shown to facilitate tissue repair and regeneration. Herein, we discuss the prospective applications of MSC-derived exosomes in TBI injuries. We also review the roles of MSC-EVs and the underlying mechanisms of their effects on promoting tendon-bone healing. At last, we discuss the present challenges and future research directions.
Subject(s)
Exosomes , Mesenchymal Stem Cells , MicroRNAs , Rotator Cuff Injuries , Animals , Exosomes/metabolism , Tendons/metabolism , MicroRNAs/metabolismABSTRACT
BACKGROUND: Anterior cruciate ligament (ACL) injuries and bone tunnel enlargement (BTE) after ACL reconstruction (ACLR) remain frequent issues. Bone dust (BD) produced by tunnel preparation with osteogenic ability and reverse drilling (RD), an easy compaction technique, make it accessible to enhance tendon-bone healing in the clinic. HYPOTHESIS: RD and BD synergistically promote tendon-bone healing by improving peritunnel bone and preventing BTE in femurs. STUDY DESIGN: Controlled laboratory study. METHODS: In total, 96 New Zealand White rabbits underwent ACLR. The semitendinosus tendon was freed before medial parapatellar arthrotomy. After the native ACL was transected, bone tunnels were prepared through the footprint of the native ACL. All animals were randomly assigned to 1 of 4 groups according to different tunnel preparation methods: group 1 (irrigation after extraction drilling [ED]; control group), group 2 (irrigation after RD), group 3 (no irrigation after ED), and group 4 (no irrigation after RD). BD was harvested by irrigating tunnels and was characterized by morphology and size. The specimens underwent microarchitectural, histological, and biomechanical evaluations at 4, 8, and 12 weeks postoperatively. RESULTS: Micro-computed tomography demonstrated more peritunnel bone and less BTE in the femurs of group 4 compared with the other groups. Histologically, BD possessed osteogenic activity in bone tunnels postoperatively. Meanwhile, group 4 regenerated a higher amount of the tendon-bone interface and more peritunnel bone than group 1. Biomechanically, group 4 showed higher failure loads and stiffness than group 1. However, peritunnel bone loss, active osteoclasts, and significant BTE were found in the femurs of group 1 and group 3 at 12 weeks postoperatively, while no strong correlation was found between BTE and inflammatory cytokines. Scanning electron microscopy and particle size analysis suggested that BD produced by ED and RD had no difference in size. CONCLUSION: Tendon-bone healing was facilitated by the synergistic effect of RD and BD in femurs. CLINICAL RELEVANCE: This study provides a more accessible and effective surgical strategy to promote tendon-bone healing after ACLR by increasing peritunnel bone and preventing BTE in femurs.
Subject(s)
Anterior Cruciate Ligament Reconstruction , Dust , Animals , Rabbits , Research Design , X-Ray MicrotomographyABSTRACT
Objective: To investigate whether preoperative lateral anterior tibial subluxation (LATS) measured from magnetic resonance imaging (MRI) can influence tibial insertion and postoperative sagittal alignment after anterior cruciate ligament reconstructions (ACLRs). Methods: 84 patients who underwent single-bundle ACLRs were retrospectively investigated. Among them, 39 patients (LATS of <6â mm) 23 patients (LATS of ≥6â mm and <10â mm) and 22 patients (excessive LATS of ≥10â mm) were defined as group 1, 2 and 3, respectively. LATS, the position of graft insertion into tibia as ratio of anterior-posterior width (AP ratio) and the sagittal graft angle (SGA) were postoperatively assessed from MRI at 2-year follow-up. Following linear regression analyses were employed. Results: The group 3 exhibited the largest preoperative LATS and remained the most postoperative LATS. Moreover, the group 3 possessed the most posteriorly located tunnel insertion with the largest AP ratio and the most vertical graft orientation. Of all included patients, a moderate correlation was demonstrated between pre- and postoperative LATS (r = 0.635). A low correlation was observed between preoperative LATS and AP ratio (r = 0.300) and a moderate correlation was displayed between AP ratio and SGA (r = 0.656). Conclusion: For ACL injuries with excessive LATS (≥10 mm), most posteriorly located tibial insertion was found out, and worse sagittal alignment containing high residual LATS was associated with more vertical graft orientation following ACLRs.
ABSTRACT
Background: As a valuable blood glucose measurement, HemoglobinA1c (HbA1c) is of great clinical value for diabetes. However, in previous observational studies, studies on its effect on bone mineral density (BMD) have different results. This study aimed to use Mendelian randomization (MR) to assess the effect of HbA1c on bone mineral density and fracture risk, and try to further explore whether this association was achieved through glycemic or non-glycemic factors. Methods: Take HbA1c measurement as exposure, and BMD estimated from quantitative heel ultrasounds (eBMD) and bone fractures as outcomes. Two-Sample MR Analysis was conducted to assess the causal effect of HbA1C on heel BMD and risk fracture. Then, we performed the analysis using two subsets of these variants, one related to glycemic measurement and the other to erythrocyte indices. Results: Genetically increased HbA1C was associated with the lower heel eBMD [odds ratio (OR) 0.91 (95% CI 0.87, 0.96) per %-unit, P = 3 × 10-4(IVW)]. Higher HbA1C was associated with lower heel eBMD when using only erythrocytic variants [OR 0.87 (0.82, 0.93), P=2× 10-5(IVW)]; However, when using only glycemic variants, this casual association does not hold. In further MR analysis, we test the association of erythrocytic traits with heel eBMD. Conclusion: Our study revealed the significant causal effect of HbA1c on eBMD, and this causal link might achieve through non-glycemic pathways (erythrocytic indices).
Subject(s)
Bone Density , Fractures, Bone , Blood Glucose , Bone Density/genetics , Fractures, Bone/genetics , Glycated Hemoglobin , Humans , Mendelian Randomization Analysis , Risk FactorsABSTRACT
OBJECTIVE: To assess the association of insulin-like growth factor 1 (IGF-1) with the risk of incident ischaemic stroke and outcome after ischaemic stroke. DESIGN: A systematic review of primary studies. SETTING: Hospitals in Western Sweden, Italy, China and Denmark. METHODS: A search was carried out in eligible studies in electronic databases (PubMed, Scopus, Embase, China National Knowledge Infrastructure and Web of Science) updated to 29 December 2020. The relevant data were extracted in order to conduct the meta-analysis. Review Manager V.5.2 was used to pool data and calculate the mean difference (MD) and its 95% CI. Heterogeneity, subgroup analysis, sensitivity analysis and publication bias were also performed in this meta-analysis. RESULTS: A total of 2277 patients were included in 17 studies. This meta-analysis indicated that higher serum IGF-1 levels were significantly correlated with less risk of ischaemic stroke (MD=-45.32 95% CI -63.70 to -26.94], p < 0.00001, I2=99%) and better improvement of outcome after ischaemic stroke (MD=27.52, 95% CI 3.89 to 51.14, p=0.02, I2=96%). According to subgroup analysis, heterogeneity comes from country, sample size, male and the time from symptom onset to blood collection. Sensitivity analysis showed that there was no significant influence of any individual study on the pooled MD. The effect of high heterogeneity on result credibility was eliminated when four included studies were merged (MD=-30.32, 95% CI -36.52 to -24.11, p< 0.00001, I2=0%). Moreover, no potential publication bias was discovered in this meta-analysis. CONCLUSION: Higher serum IGF-1 was significantly correlated with a lower risk of ischaemic stroke. In view of the high degree of heterogeneity, it may need more studies to confirm the prognostic value of serum IGF-1 levels in ischaemic stroke and explore the sources of heterogeneity.
Subject(s)
Brain Ischemia , Insulin-Like Growth Factor I , Ischemic Stroke , Stroke , Brain Ischemia/etiology , Brain Ischemia/metabolism , Humans , Insulin-Like Growth Factor I/metabolism , Ischemic Stroke/etiology , Ischemic Stroke/metabolism , Italy , Male , Risk FactorsABSTRACT
PURPOSE: To investigate whether anterior tibial subluxation obtained from magnetic resonance imaging (MRI) could be a predictor of high-grade rotatory instability for anterior cruciate ligament (ACL) injuries, including acute and chronic cases. METHODS: From September 2016 to August 2018, we retrospectively investigated 163 patients with ACL injuries who subsequently underwent primary ACL reconstruction. Among them, 30 patients with high-grade rotatory instability (grade II/III pivot shift) were included in the high-grade group, and their age and sex were matched 1:2 to low-grade cases (
Subject(s)
Anterior Cruciate Ligament Injuries , Joint Dislocations , Joint Instability , Anterior Cruciate Ligament Injuries/complications , Anterior Cruciate Ligament Injuries/diagnostic imaging , Anterior Cruciate Ligament Injuries/surgery , Case-Control Studies , Female , Humans , Joint Dislocations/surgery , Joint Instability/complications , Joint Instability/etiology , Knee Joint/surgery , Magnetic Resonance Imaging , Male , Retrospective StudiesABSTRACT
BACKGROUND: In this paper, a systematic review and meta-analysis of published randomized controlled trials (RCTs) was conducted to compare the efficacies of acupuncture and antidepressant therapy for the treatment of poststroke depression (PSD). METHODS: The research team searched RCTs published on PubMed; Medline; Cochrane library; Chinese National Knowledge Infrastructure (CNKI); Wanfang; Embase; Scopus, and Sinomed from their respective establishments to January 2019. We evaluated the Hamilton Depression Rating Scale (HAMD) scores, Treatment Emergent Symptom Scale (TESS) scores, National Institute of Health Stroke Scale (NIHSS) scores, and total clinical efficacy using fixed effects models. RESULTS: Fourteen RCTs, representing a total of 1124 patients, were studied. Results showed that acupuncture was more effective in improving HAMD scores at 3 weeks after administration (mean difference [MD]â=â-1.17, 95%CIâ=â-2.18 to -0.16), at 4 weeks (MDâ=â-4.44, 95% CIâ=â-5.64 to -3.23), at 6 weeks (MDâ=â-1.02, 95% CIâ=â-1.68 to -0.36), and at 8 weeks (MDâ=â-4.33, 95% CIâ=â-4.96 to -3.70). Similarly, acupuncture more dramatically decreased NIHSS scores (MDâ=â-2.31, 95% CIâ=â-2.53 to -2.09), and TESS scores (MDâ=â-4.70, 95% CIâ=â-4.93 to -4.48) than conventional Western medicinal therapy. Further, the total clinical efficacy in the acupuncture group was significantly higher than in the antidepressants group (risk ratio [RR]â=â1.15, 95% CIâ=â1.08-1.21). CONCLUSIONS: The results of this study suggest that acupuncture not only can reduce the severity of PSD, but also has significant effects on decreasing the appearance of other adverse events.
Subject(s)
Acupuncture Therapy/methods , Antidepressive Agents/therapeutic use , Depression/etiology , Depression/therapy , Stroke/complications , Depression/drug therapy , Humans , Randomized Controlled Trials as TopicABSTRACT
Heterocation insertion and substitution in tunnels and mezzanines of MnO2 present significant influences on the microchemical environment at the surfaces and interfaces. An innovative surface in situ doping modification method via Cu2+-H+/KMnO4 treatment was applied onto the Mn2O3 surface to provide Mn2O3@MnO2 nanospheres. Cu was stabilized into resulting MnO2 cladding substituting original K+ during a mild comproportionation reaction between Mn(VII) and Mn(III). The Cu25 (Cu(II): Mn(VII)atomic = 25%) catalyst shows significant promotion of the catalytic performance compared with bare Mn2O3 and Cu0 (without Cu involving). Isolated Cuδ+ was predominantly inserted into the mezzanine of the [MnO6]δ- layers in MnO2 instead of K+, leading to slight electron transfer from Cuδ+ to outermost Mn(4-ε)+ and a decrease of interlayer spacing as well as crystallinity. Such a configuration facilitates the formation of additional oxygen vacancies, promoting the redox ability and oxygen mobility at relatively low temperatures. The mechanistic study reveals that Cuδ+ in MnO2 cladding boosts the activation of toluene (methyl) to form benzoates and propene (methyl and double bonds) to form carboxylates, enhancing the chemical adsorption of reactants. Moreover, it also inhibits the unfavored accumulation of incomplete oxidized intermediates on the surface at high temperatures.
ABSTRACT
Demecycline (DMTC) and demeclocycline (DMCTC) are C6-demethylated derivatives of tetracycline (TC) and chlortetracycline (CTC), respectively. They are precursors of minocycline and tigecycline, which showed remarkable bioactivity against TC-resistant bacteria and have been used clinically for decades. In order to biosynthesize drug precursors DMTC and DMCTC, the function of a possible C-methyltransferase encoding gene ctcK was studied systematically in the CTC high-yielding industrial strain Streptomyces aureofaciens F3. The ΔctcK mutant accumulated two new products, which were turned out to be DMTC and DMCTC. Meanwhile, time-course analysis of the fermentation products detected the epimers of DMTC and DMCTC transformed spontaneously. Finally, an engineering strain with higher productivity of DMCTC was constructed by deleting ctcK and overexpressing ctcP of three extra copies simultaneously. Construction of these two engineering strains not only served as a successful example of synthesizing required products through metabolic engineering, but also provided original strains for following elaborate engineering to synthesize more effective tetracycline derivatives.
ABSTRACT
Polycyclic xanthones are characterized by highly oxygenated, angular hexacyclic frameworks and exhibit diverse biological activities. Although many of them have been isolated and chemically synthesized, the detailed biosynthetic machinery awaits discovery. Recently, xanthone construction in the xantholipin (1) pathway was shown to involve cryptic demethoxylation. This suggested a rationale for the existence of three O-methyltransferase (OMT) genes in the gene cluster, although there are only two O-methyl groups in the structure of 1. Here, in vivo and in vitro analysis have been used to show that the three paralogous OMTs, XanM1-M3, introduce individual methyl groups at specific points in the biosynthetic pathway. Each OMT can to some extent take over the role of the other OMTs, although they exhibit highly substrate-dependent regiospecificity. In addition, phylogenetic analysis suggests their evolution from a common ancestor. Four putative ancestral proteins were constructed, and one of them performed all the functions of XanM1-M3, while the others possessed more limited catalytic functions. The results suggest that a promiscuous common ancestor may have been able to catalyze all three reactions prior to gene duplication and functional divergence. The characterization of XanM1-M3 expands the enzyme inventory for polycyclic xanthone biosynthesis and suggests novel directed evolution approaches to diversifying natural product pathways.
Subject(s)
Methyltransferases/metabolism , Polyketides/metabolism , Catalysis , Multigene Family , Phylogeny , Stereoisomerism , Substrate SpecificityABSTRACT
BACKGROUND: Acute ischemic stroke due to large-vessel occlusion is a leading cause of death and disability, and therapeutic time window was limited to 4.5âhour when treated with intravenous thrombolysis. It has been acknowledged that endovascular treatment (EVT) is superior to general treatment (only medication, including intravenous recombinant tissue plasminogen activator (rt-PA)) in improving the outcome of AIS since 2015. However, the benefits were limited to improvement of functional outcomes and functional independence. Hence, this meta-analysis was conducted to summarize the benefits of EVT for acute ischemic stroke, explore underlying indications of EVT for AIS patients and suggest implications for clinical practice and future research. METHODS: A search was performed to identify eligible studies in PubMed, Scopus and Web of Science updated to February 5, 2019. Functional outcomes, the modified Rankin Scale (mRS) 0-1, mRS 0-2, all-cause mortality, symptomatic intracerebral hemorrhage and asymptomatic intracerebral hemorrhage (aICH) at 90 days were selected as outcomes. Data was pooled to calculate the odds ratio (OR) and 95% confidence interval (CI). Heterogeneity, subgroup analysis, sensitivity analysis and publication bias were also performed in this meta-analysis. RESULTS: Eighteen studies comprising 3831 patients were included and analyzed in this meta-analysis. In comparison with general treatment, improved functional outcomes (mRS 0-1: ORâ=â1.68, 95% CIâ=â1.43-1.97, inconsistency index [Iâ=â57%, Pâ<â.00001; mRS 0-2: ORâ=â1.78, 95% CIâ=â1.55-2.03, Iâ=â69%, Pâ<â.00001), reduced risk of all-cause mortality (ORâ=â0.82, 95% CIâ=â0.70-0.98, Iâ=â27%, Pâ=â.03) but higher risk of aICH (ORâ=â1.43, 95% CIâ=â1.05-1.95, Iâ=â0%, Pâ=â.02) at 90 days were found in AIS patients treated with EVT. Ageâ<â70, National Institutes of Health Stroke Scale ≥20 and maximum delay for invention>5âhours could improve clinical outcomes following EVT. In sensitivity analysis, it showed that 2 studies had a great influence on the pooled ORs. No potential publication bias was found in this meta-analysis. CONCLUSION: Taken together, EVT, which led to improved functional outcomes and decreased risk of death, is superior to general treatment for AIS patients with ageâ<â70, National Institutes of Health Stroke Scale ≥20 and maximum delay for invention>5âhours. Moreover, it suggests that "with mechanical thrombectomy" is potential favorable factor for improving aICH in comparison with general treatment.
Subject(s)
Brain Ischemia/mortality , Endovascular Procedures/methods , Stroke/drug therapy , Thrombolytic Therapy/methods , Administration, Intravenous , Adult , Aged , Aged, 80 and over , Brain Ischemia/epidemiology , Cerebral Hemorrhage/epidemiology , Cerebral Hemorrhage/etiology , Endovascular Procedures/statistics & numerical data , Female , Fibrinolytic Agents/administration & dosage , Fibrinolytic Agents/therapeutic use , Humans , Intracranial Hemorrhages/epidemiology , Intracranial Hemorrhages/etiology , Male , Middle Aged , Randomized Controlled Trials as Topic , Sensitivity and Specificity , Stroke/surgery , Thrombectomy/methods , Thrombectomy/statistics & numerical data , Time-to-Treatment/statistics & numerical data , Tissue Plasminogen Activator/administration & dosage , Tissue Plasminogen Activator/therapeutic use , Treatment OutcomeABSTRACT
Stroke is the leading cause of death in the elderly. Traditional Chinese medicine provides an exciting strategy for treating stroke. Previous reports indicated that Longshengzhi capsules (LSZ), a modified Chinese formula, reduced formed thrombi and oxidative stress and were promising in the clinical treatment of ischemic stroke. However, the specific therapeutic effect and mechanism of LSZ are still ambiguous. This study aimed to define the effects of LSZ on proinflammatory mediators and neuroprotective effects on middle cerebral artery occlusion and refusion (MCAO/R) rats. Rats were treated with different doses of LSZ (0.54, 1.62, and 4.32 g/(kg·d)) in a week after model building. LSZ could improve the survival rate, ischemic stroke outcome, and infarct volume. In addition, significant decrease was observed in reactive oxygen species (ROS) levels and inflammatory factor levels in LSZ-treated groups, concomitant with increase in activities of superoxide dismutase (SOD), neurosynaptic remodeling, and decrease in brain edema. It is proposed that LSZ has anti-inflammatory and neuroprotective effects resulting in downregulating matrix metalloproteinase 2/9 (MMP-2/9) and vascular endothelial growth factor (VEGF) and nuclear factor kappa-B (NF-κB) and upregulating microtubule-associated protein-2 (Map-2) and growth-associated protein-43 (GAP-43) via p38 MAPK and HIF-1α signaling pathways in MCAO/R rats. This study provides potential evidences that p38 MAPK and HIF-1α/VEGF signaling pathways play significant roles in the anti-inflammatory and neuroprotective effects of LSZ.
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The protein synthesis inhibitor anisomycin features a unique benzylpyrrolidine system and exhibits potent selective activity against pathogenic protozoa and fungi. It is one of the important effective components in Agricultural Antibiotic120, which has been widely used as naturally-originated agents for treatment of crop decay in China. The chemical synthesis of anisomycin has recently been reported, but the complex process with low productivity made the biosynthesis still to be a vital mainstay in efforts. The biosynthetic gene cluster (BGC) of anisomycin in Streptomyces hygrospinosus var. beijingensis has been identified in our previous work, while poor understanding of the regulatory mechanism limited the yield enhancement via regulation engineering of S. hygrospinosus var. beijingensis. In this study here, we characterized AniF as an indispensable LuxR family transcriptional regulator for the activation of anisomycin biosynthesis. The genetic manipulations of aniF and the real-time quantitative PCR (RT-qPCR) revealed that it positively regulated the transcription of the anisomycin BGC. Moreover, the overexpression of aniF contributed to the improvement of the production of anisomycin and its derivatives. Dissection of the mechanism underlying the function of AniF revealed that it directly activated the transcription of the genes aniR-G involved in anisomycin biosynthesis. Especially, one AniF-binding site in the promoter region of aniR was identified by DNase I footprinting assay and an inverted repeat sequence (5'-GGGC-3') composed of two 4-nt half sites in the protected region was found. Taken together, our systematic study confirmed the positive regulatory role of AniF and might facilitate the future construction of engineering strains with high productivity of anisomycin and its derivatives.
ABSTRACT
Piericidin A1, a member of É-pyridone antibiotic, exhibits various biological activities such as antimicrobial, antifungal, and antitumor properties and possesses potent respiration-inhibitory activity against insects due to its competitive binding capacity to mitochondrial complex I. The biosynthetic pathway of piericidin A1 has been reported in Streptomyces piomogeues var. Hangzhouwanensis, while the regulatory mechanism remains poorly understood. In this study, a Streptomyces antibiotic regulatory protein (SARP) family transcriptional regulator PieR was characterized. Genetic disruption and complementation manipulations revealed that PieR positively regulated the production of piericidin A1. Moreover, the overexpression of pieR contributed to the improvement of piericidin A1 productivity. The real-time quantitative PCR (RT-qPCR) was carried out and the data showed that pieR stimulated the transcription of all the biosynthesis-related genes for piericidin A1. In order to explore the regulatory mechanism, electrophoresis mobility shift assays (EMSA) and DNase I footprinting experiments have been conducted. A protected region covering 50 nucleotides within the upstream region of pieR was identified and two 5-nt direct repeat sequences (5'-CCGGA-3') in the protected region were found. These findings, taken together, set stage for transcriptional control engineering in the view of optimizing piericidin A1 production and thus provide a viable potent route for the construction of strains with high productivity.
ABSTRACT
Kelp waste extracts (KWE) contained massive soluble sugars, amino acids and various mineral elements. To probe the effects of KWE on microalgal physiological and biochemical responses, the cultures were carried out under the different dilutions. The results showed that 8.0% KWE increased the biomass productivities and total lipid contents of Chlorella strains dramatically, which were 1.83-31.86 times and 20.78-25.91% higher than that of the control. Phaeodactylum tricornutum and Spirulina maxima presented a better growth performance in 1.0% and 4.0% treatment respectively, while their lipid accumulation were not enhanced. In Chlorella-Arc, Chlorella sorokiniana and P. tricornutum, the contents of saturated and monounsaturated fatty acids could be increased, and polyunsaturated fatty acids could be decreased under the conditions of high concentration of KWE (6.0-8.0%). Briefly, KWE facilitated to enhance the biomass productivity and lipid content of Chlorella strains, also improved the fatty acid compositions for biodiesel production.