Reversed-Phase Medium-Pressure Liquid Chromatography Purification of Omega-3 Fatty Acid Ethyl Esters Using AQ-C18.

Omega-3 fatty acids are in high demand due to their efficacy in treating hypertriglyceridemia and preventing cardiovascular diseases. However, the growth of the industry is hampered by low purity and insufficient productivity. This study aims to develop an efficient RP-MPLC purification method for omega-3 fatty acid ethyl esters with high purity and capacity. The results indicate that the AQ-C18 featuring polar end-capped silanol groups outperformed C18 and others in retention time and impurity separation. By injecting pure fish oil esters with a volume equivalent to a 1.25% bed volume on an AQ-C18 MPLC column using a binary isocratic methanol-water (90:10, v:v) mobile phase at 30 mL/min, optimal omega-3 fatty acid ethyl esters were obtained, with the notable purity of 90.34% and a recovery rate of 74.30%. The total content of EPA and DHA produced increased from 67.91% to 85.27%, meeting the acceptance criteria of no less than 84% set by the 2020 edition of the Pharmacopoeia of the People's Republic of China. In contrast, RP-MPLC significantly enhanced the production efficiency per unit output compared to RP-HPLC. This study demonstrates a pioneering approach to producing omega-3 fatty acid ethyl esters with high purity and of greater quantity using AQ-C18 RP-MPLC, showing this method's significant potential for use in industrial-scale manufacturing.

Targeted Affinity Purification and Mechanism of Action of Angiotensin-Converting Enzyme (ACE) Inhibitory Peptides from Sea Cucumber Gonads.

Protein hydrolysates from sea cucumber (Apostichopus japonicus) gonads are rich in active materials with remarkable angiotensin-converting enzyme (ACE) inhibitory activity. Alcalase was used to hydrolyze sea cucumber gonads, and the hydrolysate was separated by the ultrafiltration membrane to produce a low-molecular-weight peptide component (less than 3 kDa) with good ACE inhibitory activity. The peptide component (less than 3 kDa) was isolated and purified using a combination method of ACE gel affinity chromatography and reverse high-performance liquid chromatography. The purified fractions were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the resulting products were filtered using structure-based virtual screening (SBVS) to obtain 20 peptides. Of those, three noncompetitive inhibitory peptides (DDQIHIF with an IC50 value of 333.5 µmol·L-1, HDWWKER with an IC50 value of 583.6 µmol·L-1, and THDWWKER with an IC50 value of 1291.8 µmol·L-1) were further investigated based on their favorable pharmacochemical properties and ACE inhibitory activity. Molecular docking studies indicated that the three peptides were entirely enclosed within the ACE protein cavity, improving the overall stability of the complex through interaction forces with the ACE active site. The total free binding energies (ΔGtotal) for DDQIHIF, HDWWKER, and THDWWKER were -21.9 Kcal·mol-1, -71.6 Kcal·mol-1, and -69.1 Kcal·mol-1, respectively. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that HDWWKER could significantly decrease the systolic blood pressure (SBP) of SHRs after intravenous administration. The results showed that based on the better antihypertensive activity of the peptide in SHRs, the feasibility of targeted affinity purification and computer-aided drug discovery (CADD) for the efficient screening and preparation of ACE inhibitory peptide was verified, which provided a new idea of modern drug development method for clinical use.

UCMSCs-derived exosomal circHIPK3 promotes ulcer wound angiogenesis of diabetes mellitus via miR-20b-5p/Nrf2/VEGFA axis.

AIMS: Experiments confirmed that circular RNAs contributed to the pathogenesis of diabetic foot ulcers (DFUs). CircHIPK3 was upregulated in type 2 diabetes mellitus (T2DM), but its role in DFU remained unknown. Our study aimed to investigate the regulatory functions of exosomal circHIPK3 and its potential mechanisms in DFU. METHODS: Exosomal size and distribution, marker proteins, and circHIPK3 levels were evaluated by transmission electron microscope, ExoView R200, western blot, and qRT-PCR. Flow cytometry, MTT, Wound healing assays, and tube formation assays were used to assess the roles of exosomal circHIPK3 in high glucose (HG)-treated human umbilical vein endothelial cells (HUVECs). The relationships between Nrf2/VEGFA/circHIPK3 and miR-20b-5p, and between Nrf2 and VEGFA were determined by luciferase reporter assay and RNA immunoprecipitation. We used cell and mice models to investigate the mechanisms of exosomal circHIPK3 under diabetic conditions. RESULTS: CircHIPK3 was significantly upregulated in exo-circHIPK3 rather than exo-vector. Exo-circHIPK3 remarkably inhibited cell apoptosis but promoted cell proliferation, migration, and tube formation in HG-treated HUVECs. Luciferase reporter and RIP assays showed that miR-20b-5p targeted and inhibited Nrf2 and VEGFA, and circHIPK3 acted as a ceRNA of miR-20b-5p to inhibit the binding to its downstream genes Nrf2 and VEGFA. Mechanistically, circHIPK3 promoted cell proliferation, migration, and angiogenesis via downregulating miR-20b-5p to upregulate Nrf2 and VEGFA. However, the overexpressed miR-20b-5p could abolish the promoting effects of circHIPK3 overexpression on cell proliferation, migration, and tube formation under HG conditions. CONCLUSION: UCMSCs-derived exosomal circHIPK3 protected HG-treated HUVECs via miR-20b-5p/Nrf2/VEGFA axis. The exosomal circHIPK3 might be a therapeutic candidate to treat DFU.

Affinity Purification and Molecular Characterization of Angiotensin-Converting Enzyme (ACE)-Inhibitory Peptides from Takifugu flavidus.

An affinity chromatography filler of CNBr-activated Sepharose 4B-immobilized ACE was used to purify ACE-inhibitory peptides from Takifugu flavidus protein hydrolysate (<1 kDa). Twenty-four peptides with an average local confidence score (ALC) ≥ 80% from bounded components (eluted by 1 M NaCl) were identified by LC-MS/MS. Among them, a novel peptide, TLRFALHGME, with ACE-inhibitory activity (IC50 = 93.5 µmol·L-1) was selected. Molecular docking revealed that TLRFALHGME may interact with the active site of ACE through H-bond, hydrophobic, and electrostatic interactions. The total binding energy (ΔGbinding) of TLRFALHGME was estimated to be -82.7382 kJ·mol-1 by MD simulations, indicating the favorable binding of peptides with ACE. Furthermore, the binding affinity of TLRFALHGME to ACE was determined by surface plasmon resonance (SPR) with a Kd of 80.9 µmol, indicating that there was a direct molecular interaction between them. TLRFALHGME has great potential for the treatment of hypertension.

[Electroencephalogram Features of Anti-N-Methyl-D-Aspartate Receptor Encephalitis and Their Value for Clinical Assessment].

Objective: To analyze the electroencephalogram (EEG) features of anti-N-methyl-D-aspartate receptor encephalitis (anti-NMDARE) and to study the clinical assessment value of the degree of EEG background slowing and the presence of δ brush. Methods: We enrolled 52 patients with anti-NMDARE and collected their clinical data, including age, sex, form of disease onset, status of tumor comorbidity, auxiliary examination findings (cerebrospinal fluid [CSF] anti-methyl-D-aspartate receptor antibody titers, magnetic resonance imaging [MRI] reports, and EEG results), treatment status, and follow-up after discharge. The degree of EEG background abnormality and the presence of δ brush in the EEG of patients with different clinical features were analyzed. Results: Among the 52 patients, 7 (14%) had normal EEG, and 45 (87%), abnormal EEG, including 25 (48%) with mild abnormalities, 11 (21%) with moderate abnormalities, and 9 (17%) with severe abnormalities. δ brush was seen in 6 (12%) patients. At the time of EEG, 32 (62%) patients were in the mild condition group and 20 (38%) patients were in the severe condition group. After 1 year of follow-up, there were 45 (86%) patients in the good prognosis group and 7 (14%) patients in the poor prognosis group. The exacerbation of EEG background abnormalities and the presence of δ brush were indications for an increase in the proportion of patients who were in severe condition, who needed ICU admission, and who had poor prognosis ( P<0.01). The worse the EEG background abnormalities, the higher the proportion of CSF antibody titers>1∶10 ( P=0.035), and the higher the proportion of patients initiating second-line immunotherapy ( P=0.008). The δ brush was seen a higher proportion in patients with comorbid tumors ( P=0.012). The probability of δ brush presence was higher in the first-time diagnosis cases than that in recurrent cases ( P=0.023). Conclusions: The degree of EEG slowing and the presence of δ brush have shown consistent performance in assessing patients' condition and predicting prognosis. The slower the EEG, the more severe the disease, and the worse the prognosis. The presence of δ brush indicates severe disease and poor prognosis. EEG slowing is correlated with the immune status of patients with anti-NMDARE. The slower the EEG, the more severe the immune abnormalities. In clinical practice, patient EEG should be under dynamic monitoring in order to evaluate the effect of immunotherapy. If EEG slowing is not improved, enhanced immunotherapy should be considered as early as possible. The δ brush is seen at a higher proportion in patients with comorbid tumors. Therefore, active efforts should be made to screen for tumors when δ brush is present.

The positive contribution of ultrasound technology in muscle food key processing and its mechanism-a review.

Traditional processing methods can no longer meet the demands of consumers for high-quality muscle food. As a green and non-thermal processing technology, ultrasound has the advantage of improving processing efficiency and reducing processing costs. Of these, the positive effect of power ultrasound in the processing of muscle foods is noticeable. Based on the action mechanism of ultrasound, the factors affecting the action of ultrasound are analyzed. On this basis, the effect of ultrasound technology on muscle food quality and its action mechanism and application status in processing operations (freezing-thawing, tenderization, marination, sterilization, drying, and extraction) is discussed. The transient and steady-state effects, mechanical effects, thermal effects, and chemical effects can have an impact on processing operations through complex correlations, such as improving the efficiency of mass and heat transfer. Ultrasound technology has been proven to be valuable in muscle food processing, but inappropriate ultrasound treatment can also have adverse effects on muscle foods. In the future, kinetic models are expected to be an effective tool for investigating the application effects of ultrasound in food processing. Additionally, the combination with other processing technologies can facilitate their intensive application on an industrial level to overcome the disadvantages of using ultrasound technology alone.

Molecular Characterization of a New Tetrodotoxin-Binding Protein, Peroxiredoxin-1, from Takifugu bimaculatus.

Pufferfish are considered a culinary delicacy but require careful preparation to avoid ingestion of the highly toxic tetrodotoxin (TTX), which accumulates in certain tissues. In this study, the tissue distribution of peroxiredoxin-1 from Takifugu bimaculatus was investigated. The peroxiredoxin-1 protein was obtained by in vitro recombinant expression and purification. The recombinant protein had a strong ability to scavenge hydroxyl radicals, protect superhelical DNA plasmids from oxidative damage, and protect L929 cells from H2O2 toxicity through in vitro antioxidant activity. In addition, we verified its ability to bind to tetrodotoxin using surface plasmon resonance techniques. Further, recombinant proteins were found to facilitate the entry of tetrodotoxin into cells. Through these analyses, we identified, for the first time, peroxiredoxin-1 protein from Takifugu bimaculatus as a potential novel tetrodotoxin-binding protein. Our findings provide a basis for further exploration of the application of peroxiredoxin-1 protein and the molecular mechanisms of tetrodotoxin enrichment in pufferfish.

Screening and Molecular Mechanisms of Novel ACE-Inhibitory Peptides from Gracilariopsis lemaneiformis.

Candidate peptides with novel angiotensin-I-converting enzyme (ACE) inhibitor activity were obtained from hydrolysates of Gracilariopsis lemaneiformis by virtual screening method. Our results showed that G. lemaneiformis peptides (GLP) could significantly lower blood pressure in spontaneously hypertensive rats (SHR). At least 101 peptide sequences of GLP were identified by LC-MS/MS analysis and subjected to virtual screening. A total of 20 peptides with the highest docking score were selected and chemically synthesized in order to verify their ACE-inhibitory activities. Among them, SFYYGK, RLVPVPY, and YIGNNPAKG showed good effects with IC50 values of 6.45 ± 0.22, 9.18 ± 0.42, and 11.23 ± 0.23 µmoL/L, respectively. Molecular docking studies revealed that three peptides interacted with the active center of ACE by hydrogen bonding, hydrophobic interactions, and electrostatic forces. These peptides could form stable complexes with ACE. Furthermore, SFYYGK, RLVPVPY, and YIGNNPAKG significantly reduced systolic blood pressure (SBP) in SHR. YIGNNPAKG exhibited the highest antihypertensive effect, with the largest decrease in SBP (approximately 23 mmHg). In conclusion, SFYYGK, RLVPVPY, and YIGNNPAKG can function as potent therapeutic candidates for hypertension treatment.

Research progress on quality deterioration mechanism and control technology of frozen muscle foods.

Freezing can prolong the shelf life of muscle foods and is widely used in their preservation. However, inevitable quality deterioration can occur during freezing, frozen storage, and thawing. This review explores the eating quality deterioration characteristics (color, water holding capacity, tenderness, and flavor) and mechanisms (irregular ice crystals, oxidation, and hydrolysis of lipids and proteins) of frozen muscle foods. It also summarizes and classifies the novel physical-field-assisted-freezing technologies (high-pressure, ultrasound, and electromagnetic) and bioactive antifreeze (ice nucleation proteins, antifreeze proteins, natural deep eutectic solvents, carbohydrate, polyphenol, phosphate, and protein hydrolysates), regulating the dynamic process from water to ice. Moreover, some novel thermal and nonthermal thawing technologies to resolve the loss of water and nutrients caused by traditional thawing methods were also reviewed. We concluded that the physical damage caused by ice crystals was the primary reason for the deterioration in eating quality, and these novel techniques promoted the eating quality of frozen muscle foods under proper conditions, including appropriate parameters (power, time, and intermittent mode mentioned in ultrasound-assisted techniques; pressure involved in high-pressure-assisted techniques; and field strength involved in electromagnetic-assisted techniques) and the amounts of bioactive antifreeze. To obtain better quality frozen muscle foods, more efficient technologies and substances must be developed. The synergy of novel freezing/thawing technology may be more effective than individual applications. This knowledge may help improve the eating quality of frozen muscle foods.

Visible-Light-Mediated Oxidative Cyclization of 2-Aminobenzyl Alcohols and Secondary Alcohols Enabled by an Organic Photocatalyst.

This paper describes a visible-light-mediated oxidative cyclization of 2-aminobenzyl alcohols and secondary alcohols to produce quinolines at room temperature. This photocatalytic method employed anthraquinone as an organic small-molecule catalyst and DMSO as an oxidant. According to this present procedure, a series of quinolines were prepared in satisfactory yields.

A Novel Angiotensin-I-Converting Enzyme (ACE) Inhibitory Peptide from Takifugu flavidus.

Alcalase, neutral protease, and pepsin were used to hydrolyze the skin of Takifugu flavidus. The T. flavidus hydrolysates (TFHs) with the maximum degree of hydrolysis (DH) and angiotensin-I-converting enzyme (ACE)-inhibitory activity were selected and then ultra-filtered to obtain fractions with components of different molecular weights (MWs) (<1, 1-3, 3-10, 10-50, and >50 kDa). The components with MWs < 1 kDa showed the strongest ACE-inhibitory activity with a half-maximal inhibitory concentration (IC50) of 0.58 mg/mL. Purification and identification using semi-preparative liquid chromatography, Sephadex G-15 gel chromatography, RP-HPLC, and LC-MS/MS yielded one new potential ACE-inhibitory peptide, PPLLFAAL (non-competitive suppression mode; IC50 of 28 µmmol·L-1). Molecular docking and molecular dynamics simulations indicated that the peptides should bind well to ACE and interact with amino acid residues and the zinc ion at the ACE active site. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that PPLLFAAL could significantly decrease the systolic blood pressure (SBP) and diastolic blood pressure (DBP) of SHRs after intravenous administration. These results suggested that PPLLFAAL may have potential applications in functional foods or pharmaceuticals as an antihypertensive agent.

ACE Inhibitory Peptide from Skin Collagen Hydrolysate of Takifugu bimaculatus as Potential for Protecting HUVECs Injury.

Angiotensin-I-converting enzyme (ACE) is a crucial enzyme or receptor that catalyzes the generation of potent vasopressor angiotensin II (Ang II). ACE inhibitory peptides from fish showed effective ACE inhibitory activity. In this study, we reported an ACE inhibitory peptide from Takifugu bimaculatus (T. bimaculatus), which was obtained by molecular docking with acid-soluble collagen (ASC) hydrolysate of T. bimaculatus. The antihypertensive effects and potential mechanism were conducted using Ang-II-induced human umbilical vein endothelial cells (HUVECs) as a model. The results showed that FNLRMQ alleviated the viability and facilitated apoptosis of Ang-II-induced HUVECs. Further research suggested that FNLRMQ may protect Ang-II-induced endothelial injury by regulating Nrf2/HO-1 and PI3K/Akt/eNOS signaling pathways. This study, herein, reveals that collagen peptide FNLRMQ could be used as a potential candidate compound for antihypertensive treatment, and could provide scientific evidence for the high-value utilization of marine resources including T. bimaculatus.

Mitochondria-targeted Re(I) complexes bearing guanidinium as ligands and their anticancer activity.

As the "powerhouse" of a cell, mitochondria maintain energy homeostasis, synthesize ATP via oxidative phosphorylation, generate ROS signaling molecules, and modulate cell apoptosis. Herein, three Re(I) complexes bearing guanidinium derivatives have been synthesized and characterized. All of these complexes exhibit moderate anticancer activity in HepG2, HeLa, MCF-7, and A549 cancer cells. Mechanism studies indicate that complex 3, [Re(CO)3(L)(Im)](PF6)2, can selectively localize in the mitochondria and induce cancer cell death through mitochondria-associated pathways. In addition, complex 3 can effectively depress the ability of cell migration, cell invasion, and colony formation.

The Predictive Value of Red Cell Distribution Width and Red Cell Distribution Width to Erythrocyte Count Ratio for Adverse Cardiovascular Events During the Hospitalization of Patients of ST-segment Elevation Myocardial Infarction.

BACKGROUND: High red cell distribution width (RDW) is correlated with poor prognosis in acute coronary syndromes (ACS), including ST-segment elevation myocardial infarction (STEMI). However, the association of red cell distribution width to erythrocyte count ratio (RER) with STEMI undergoing percutaneous coronary intervention (PCI) during hospitalization has not been investigated. Therefore, we performed a retrospective study to investigate whether RER is associated with STEMI patients after PCI during hospitalization. METHODS: A total of 331 patients, who were hospitalized for STEMI and underwent PCI, were enrolled. Receiver operating characteristic curve (ROC) analyses were used to find the cutoff value of RER and classify the patients into two groups including higher RER group and lower RER group by cutoff value. Differences between measured parameters in higher RER and lower RER groups were analyzed by the Mann-Whitney U test. The evaluation correlation of RDW, red blood cell, and RER with major adverse cardiovascular events was determined by bivariate regression analysis. Univariate logistic regression analysis was used to determine the factors associated with adverse cardiovascular events during the hospitalization of STEMI patients. Multivariate logistic regression analysis was performed to evaluate the potential independent predictors of STEMI. RESULTS: According to ROC analysis, the cutoff value of RER and RDW is 3.10 and 13.9, the sensitivity is 51% and 35%, the specificity is 76% and 80%, respectively. RER showed improved diagnostic capacity compared to RDW in correlation with adverse cardiovascular events during hospitalization in STEMI patients (p < 0.001). Compared with the lower RER group, the incidence of adverse cardiovascular events in STEMI patients is elevated in the higher RER group (75% vs. 64.5%, p < 0.05). Bivariate regression analysis indicated that RER and RDW showed a good correlation with adverse cardiovascular events, and the difference was statistically significant (R = 0.10 p < 0.05 vs. R = 0.05 p < 0.05). Univariate logistic regression analysis showed that age, heart rate, left ventricular ejection fraction, hyperlipidemia, RDW, mean platelet volume, total cholesterol, and RER were correlated with the occurrence of adverse cardiovascular events during the hospitalization of STEMI patients (p < 0.05). Multivariate logistic regression analysis demonstrated that RER could be an independent predictor of adverse cardiovascular events in STEMI patients (B: 0.574, OR: 1.776, 95% CI: 1.043 ~ 3.023, p < 0.05). CONCLUSIONS: RER and RDW demonstrated good correlation with adverse cardiovascular events during hospitalization in STEMI patients. RER is a potential independent predictor of adverse cardiovascular events during hospitalization in STEMI patients.

The Relationship between Erythrocyte Sedimentation Rate and Erythrocyte Sedimentation/Red Blood Cell Ratio and Disease Activity in Systemic Lupus Erythematosus.

BACKGROUND: The treatment options of systemic lupus erythematosus (SLE) patients in active and inactive phases are very different clinically, and the prognosis of patients with active SLE is much worse than inactive patients. However, the present indicators for diagnosis of SLE in activity are limited and inefficient. METHODS: Three hundred thirty patients with SLE were included. All patients are classified as SLEDAI (systemic lupus erythematosus disease activity index) > 4 as active and SLEDAI ≤ 4 as inactive. The linear correlation between variables was assessed by Pearson's correlation analysis. The difference between parameters in active and inactive patients was evaluated by the Mann-Whitney U test. The evaluation capacity of erythrocyte sedimenta-tion/red blood cell (ERR) and red blood cell/albumin ratio (RAR) on SLE activity was determined by bivariate regression analysis. Sensitivity and specificity are assessed by receiver operating characteristic curve (ROC). RESULTS: Compared with the inactive SLE, ESR (52.97 ± 35.66 vs. 32.38 ± 29.16 p < 0.001), ERR (15.40 ± 12.41 vs. 8.19 ± 8.10 p < 0.001) and RAR (0.13 ± 0.10 vs. 0.11 ± 0.20 p = 0.038) are all elevated in active SLE (52.97 ± 35.66 vs. 32.38 ±2 9.16 p < 0.001). ERR shows better correlation than RAR with ESR (p < 0.001 vs. p = 0.911). Patients with active SLE exhibited higher SLEDAI than those with inactive SLE (8.67 ± 2.67 vs. 3.27 ± 1.36, p < 0.001). According to ROC analysis, when ESR levels > 58.5 and ERR levels > 13.18, the sensitivity is 37.6% and 45.2%, the specificity is 83.0% and 83.2%. CONCLUSIONS: ESR and ERR are potential indicators for diagnosis of active and inactive SLE.

The Encystment-Related MicroRNAs and Its Regulation Molecular Mechanism in Pseudourostyla cristata Revealed by High Throughput Small RNA Sequencing.

MicroRNAs (miRNAs) regulate the expression of target genes in diverse cellular processes and play important roles in different physiological processes. However, little is known about the microRNAome (miRNAome) during encystment of ciliated protozoa. In the current study, we first investigated the differentially expressed miRNAs and relative signaling pathways participating in the transformation of vegetative cells into dormant cysts of Pseudourostyla cristata (P. cristata). A total of 1608 known miRNAs were found in the two libraries. There were 165 miRNAs with 1217 target miRNAs. The total number of differential miRNAs screened between vegetative cells and dormant cysts databases were 449 with p < 0.05 and |log2 fold changes| > 1. Among them, the upregulated and downregulated miRNAs were 243 and 206, respectively. Furthermore, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that some of the differentially expressed miRNAs were mainly associated with oxidative phosphorylation, two-component system, and biosynthesis of amino acids. Combining with our bioinformatics analyzes, some differentially expressed miRNAs including miR-143, miR-23b-3p, miR-28, and miR-744-5p participates in the encystment of P. cristata. Based on these findings, we propose a hypothetical signaling network of miRNAs regulating or promoting P. cristata encystment. This study shed new lights on the regulatory mechanisms of miRNAs in encystment of ciliated protozoa.

Molecular characterization, purification, and antioxidant activity of recombinant superoxide dismutase from the Pacific abalone Haliotis discus hannai Ino.

Superoxide dismutase (SOD) is an acidic metalloenzyme that scavenges free radicals produced by endogenous and exogenous substances. In the present study, the tissue distribution of the superoxide dismutase HdhCu/Zn-SOD was investigated in Haliotis discus hannai Ino. The expression profile after lipopolysaccharide (LPS) challenge was determined using quantitative real-time polymerase chain reaction (qPCR). To study the antioxidant activity of a recombinant HdhCu/Zn-SOD protein, the HdhCu/Zn-SOD gene was cloned into the pPIC9K vector and transformed into the Pichia pastoris GS115 strain by electroporation. After induction by methanol, the recombinant product was purified using immobilized metal affinity chromatography and confirmed using mass spectrometry. The optimal expression conditions were determined to be incubation with 0.5% methanol at pH 6.0, resulting in a stable expressed product with the molecular weight of approximately 17 kDa and 21 kDa. The enzymatic activity of HdhCu/Zn-SOD consistently increased with increasing Cu2+ concentrations and showed good thermal stability. Recombinant HdhCu/Zn-SOD showed a strong ability to scavenge superoxide anions and hydroxyl radicals and protected L929 cells against the toxicity caused by H2O2 through its in vitro antioxidant activity. The heterologous expression of HdhCu/Zn-SOD in P. pastoris and the antioxidant activity of this enzyme are reported for the first time.

[Molecular diagnosis and functional study of a pedigree affected with Lubs X-linked mental retardation syndrome].

OBJECTIVE: To explore the genetic basis for a pedigree affected with X-linked mental retardation. METHODS: The proband was subjected to chromosomal karyotyping, FMR1 mutation testing and copy number variation analysis with a single nucleotide polymorphism microarray (SNP array). His family members were subjected to multiplex ligation-dependent probe amplification (MLPA) assaying. Expression of genes within the repeated region were analyzed. RESULTS: The proband had a normal chromosomal karyotype and normal number of CGG repeats within the FMR1 gene. SNP array identified a 370 kb duplication in Xq28 (ChrX: 153 027 633-153 398 515), which encompassed 14 genes including MECP2. The patient was diagnosed as Lubs X-linked mental retardation syndrome (MRXSL). MLPA confirmed the presence of copy number variation, its co-segregation with the disease, in addition with the carrier status of females. Genes from the duplicated region showed higher levels of expression (1.79 to 5.38 folds) within peripheral blood nucleated cells of the proband. CONCLUSION: The patients were diagnosed with MRXSL. The expression of affected genes was up-regulated due to the duplication. Genetic counseling and prenatal diagnosis may be provided based on the results.

[BRAF V600E Mutation and TERT Promoter Mutation in Papillary Thyroid Carcinomas and Their Association with Clinicopathological Characteristics].

OBJECTIVE: To explore the relationships of BRAF V600E and TERT promoter mutations with the clinicopathological features in papillary thyroid carcinoma (PTC). METHODS: The mutations of BRAF V600E and TERT promoters were examined by PCR-direct sequencing in tumor tissues from 326 PTC patients, while the relationships between the gene mutations and clinicopathological features were analyzed. RESULTS: BRAF V600E mutation was found in 269/326 (82.52%), and TERT promoter mutation in 11/326 (3.37%) of PTC patients. In site mutations of TERT promoter, 9 cases were C228T and 2 cases were C250T. Single factor analysis showed that BRAF V600E mutations were significantly associated with age and recurrence/distant metastasis of tumor (P < 0.05), while TERT promoter mutations were significantly associated with age, tumor size, extrathyroidal extension, T stage, AJCC stage and recurrence/distant metastasis of tumor (P < 0.05). Coexistence of BRAF V600E and TERT promoter mutations (BRAF+/TERT+) were particularly associated with age, tumor size, extrathyroidal extension, T stage and AJCC stage (P < 0.05). CONCLUSION: Coexistence of BRAF V600E and TERT promoter mutations in PTC shows more aggressive tumor behavior.

Edge optical scattering of two-dimensional materials.

Rayleigh scattering has shown powerful abilities to study electron resonances of nanomaterials regardless of the specific shapes. In analogy to Rayleigh scattering, here we demonstrate that edge optical scattering from two-dimensional(2D) materials also has the similar advantage. Our result shows that, in visible spectral range, as long as the lateral size of a 2D sample is larger than 2 µm, the edge scattering intensity distribution of the high-angle scattering in k space is nearly independent of the lateral size and the shape of the 2D samples. The high-angle edge scattering spectra are purely determined by the intrinsic dielectric properties of the 2D materials. As an example, we experimentally verify this feature in single-layer MoS2, in which A and B excitons are clearly detected in the edge scattering spectra, and the scattering images in k space and real space are consistent with our theoretical model. This study shows that the edge scattering is a highly practical and efficient method for optical studies of various 2D materials as well as thin films with clear edges.