Screening analysis of candidate gene mutations in a kindred with polycystic liver disease

AIM:To find potential mutable sites by detecting mutations of the candidate gene in a kindred with polycystic liver disease(PCLD).METHODS:First,we chose a kindred with PCLD and obtained five venous blood samples of this kindred after the family members signed the informed consent form.In the kindred two cases were diagnosed with PCLD,and the left three cases were normal individuals.All the blood samples were preserved at-85?℃.Second,we extracted the genomic DNA from the venous blood samples of the kindred using a QIAamp DNA Mini Kit and then performed long-range polymerase chain reaction(PCR)with different primers.The exons of PKD1 were all sequenced with the forward and reverse primers to ensure the accuracy of the results.Next,we purified the PCR products and directly sequenced them using Big Dye Terminator Chemistry version 3.1.The sequencing reaction was conducted with Biomek FX(Beckman).Finally,we analyzed the results.RESULTS:A total of 42 normal exons were identified in detecting mutations of the PKD1 gene.A synonymous mutation occurred in exon 5.The mutation was a homozygous T in the proband and was C in the reference sequence.This mutation was located in the third codon and did not change the amino acid encoded by the codon.Missense mutations occurred in exons 11 and 35.These mutations were located in the second codon;they changed the amino acid sequence and existed in the db SNP library.A nonsense mutation occurred in exon 15.The mutation was a heterozygous CT in the proband and was C in the referencesequence.This mutation was located in the first codon and resulted in a termination codon.This mutation had an obvious influence on the encoded protein and changed the length of the protein from 4303 to 2246amino acids.This was a new mutation that was not present in the db SNP library.CONCLUSION:The nonsense mutation of exon 15existed in the proband and in the third individual.Additionally,the proband was heterozygous for this mutation,so the mutable site was a pathogenic mutation.

Rapamycin Treatment Attenuates Angiotensin Ⅱ-induced Abdominal Aortic Aneurysm Formation via VSMC Phenotypic Modulation and Down-regulation of ERK1/2 Activity

The aim of the present study is to address the effect of rapamycin on abdominal aortic aneurysm（AAA） and the potential mechanisms. A clinically relevant AAA model was induced in apolipoprotein E-deficient（ApoE-/-） mice, in which miniosmotic pump was implanted subcutaneously to deliver angiotensin Ⅱ（Ang Ⅱ） for 14 days. Male ApoE-/-mice were randomly divided into 3 groups： saline infusion, Ang Ⅱ infusion, and Ang Ⅱ infusion plus intraperitoneal injection of rapamycin. The diameter of the supra-renal abdominal aorta was measured by ultrasonography at the end of the infusion. Then aortic tissue was excised and examined by Western blotting and histoimmunochemistry. Ang Ⅱ with or without rapamycin treatment was applied to the cultured vascular smooth muscle cells（VSMCs） in vitro. The results revealed that rapamycin treatment significantly attenuated the incidence of Ang Ⅱinduced-AAA in ApoE-/-mice. Histologic analysis showed that rapamycin treatment decreased disarray of elastin fibers and VSMCs hyperplasia in the medial layer. Immunochemistry staining and Western blotting documented the increased phospho-ERKl/2 and ERK1/2 expression in aortic walls in Ang Ⅱ induced-AAA,as well as in human lesions. Whereas in the rapamycintreated group, decreased phospho-ERK1/2 expression level was detected. Moreover, rapamycin reversed Ang Ⅱ-induced VSMCs phenotypic change both in vivo and in vitro. Based on those results, we confirmed that rapamycin therapy suppressed Ang Ⅱ-induced AAA formation in mice, partially via VSMCs phenotypic modulation and down-regulation of ERK1/2 activity.

The Change of Quantitative of HBeAg Can Predict the Efficacy of Peg-IFN-α 2a in HBeAgpositive CHB Patients

Objective To investigate the quantitation of hepatitis B e antigen (HBeAg) at week 24 in predicting the efifcacy of pegylated-interferon alfa-2a (Peg-IFN-α2a) in HBeAg-positive chronic hepatitis B (CHB) patients at week 48 and to find a useful predictor for treatment efficacy and investigate individualized treatment of antiviral therapy. Methods Ninety-six HBeAg-positive CHB patients with detectable HBeAg who were treated with Peg-IFN-α2a were enrolled in this trial. They were categorized into 3 groups according to the changes of HBeAg in week 24:HBeAg decline>2 log10 group (group A), HBeAg decline between 1 1og10-2 log10 (group B), HBeAg decline<1 log10 group (group C), and group C was randomly distributed into C1 and C2. The patients in group A, group B, and group C1 continued the original therapy and the patients in group C2 were given lamivudine plus Peg-IFN-α2a for 24 weeks. At week 48, the treatment efifcacy and hepatitis B virus covalently closed circular DNA (HBV cccDNA) in liver biopsies were analyzed. Results At week 48, mean reduction of serum HBV DNA:group A:5.8 log10 copies/ml, group B:3.8 log10 copies/ml, group C1:2.8 log10 copies/ml, group C2:5.7 log10 copies/ml, the reduction of HBV DNA in group A was greater than groups B and C1 (P<0.01), that in group C1 was greater than group C2 (P<0.01), the difference between groups B and C1 had no statistical signiifcance (P=0.19). Mean reduction of HBeAg:group A:2.7 log10S/CO, group B:1.9 log10S/CO, group C1:0.9 log10S/CO, group C2:1.5 log10S/CO, the difference among groups A, B and C1 and between groups C1 and C2 were statistically signiifcant (P<0.01). At week 48, HBV DNA undetectable rate in group A, group B, group C1 and group C2 were 87.5%, 34.5%, 17.4%and 81.9%, respectively, the rate in group A was greater than groups B and C1 (P<0.01),that in group C1 was greater than group C2 (P<0.01). HBeAg seroconversion rate were 75.0%, 24.1%, 13.0%and 22.7%, respectively, that in group A was greater than groups B and C1 (P<0.01). Group A had lower cccDNA in liver tissue than group B and group C1 (P<0.01). The difference of HBV cccDNA between groups B and C1 and that between groups C1 and C2 had no statistical signiifcance. Conclusions HBeAg decline > 2 log10 at week 24 in Peg-IFN-α 2a-treated hepatitis B patients suggested a better efficacy at week 48; HBeAg decline < 2 log10 at week 24 suggests a worse efficacy at week 48, the combined therapy of Peg-IFN-α and lamivudine could improve the clinical responses. The change of quantitative of HBeAg at week 24 may be used as a predictor of treatment effects at week 48.

MDA-JITL model for on-line mechanical property prediction

Mechanical performance prediction is the key to the transformation and upgrading of steel enterprises to intelligent manufacturing.Due to time-varying manufacturing data,the traditional prediction model of mechanical properties of hotrolled strip may cause performance degradation or even failure in its use.An MDA-JITL model was thus proposed to handle the modeling problem of complex time-varying data.Relevant parameters were first chosen and normalized.Then,a distance measurement method combining the importance of data attributes and time characteristics was designed to select the most suitable samples for on-line local modeling.After that,using the chosen dataset,a linear local model was created to predict target sample.Finally,an uncertainty evaluation method was designed to evaluate the uncertainty of prediction results.Furthermore,the appropriate dataset partition and off-line simulation experiment scheme were created based on the peculiarities of hot-rolling production.The suggested model performs much better than the classic global model when applied to actual production data from a steel plant.The stability of its prediction accuracy is demonstrated in a simulation prediction for up to five months.Moreover,there is a high link between the uncertainty evaluation metrics and the prediction error of the model,reducing the field sampling rate by 30%in industrial applications in the latest year.

Active constituents and mechanisms of Respiratory Detox Shot, a traditional Chinese medicine prescription, for COVID-19 control and prevention: Network-molecular docking-LC–MS^E analysis

Objective: Lung-toxin Dispelling Formula No. 1, referred to as Respiratory Detox Shot(RDS), was developed based on a classical prescription of traditional Chinese medicine(TCM) and the theoretical understanding of herbal properties within TCM. Therapeutic benefits of using RDS for both disease control and prevention, in the effort to contain the coronavirus disease 2019(COVID-19), have been shown. However,the biochemically active constituents of RDS and their mechanisms of action are still unclear. The goal of the present study is to clarify the material foundation and action mechanism of RDS.Methods: To conduct an analysis of RDS, an integrative analytical platform was constructed, including target prediction, protein–protein interaction(PPI) network, and cluster analysis;further, the hub genes involved in the disease-related pathways were identified, and the their corresponding compounds were used for in vitro validation of molecular docking predictions. The presence of these validated compounds was also measured in samples of the RDS formula to quantify the abundance of the biochemically active constituents. In our network pharmacological study, a total of 26 bioinformatic programs and databases were used, and six networks, covering the entire Zang-fu viscera, were constructed to comprehensively analyze the intricate connections among the compounds-targets-disease pathways-meridians of RDS.Results: For all 1071 known chemical constituents of the nine ingredients in RDS, identified from established TCM databases, 157 passed drug-likeness screening and led to 339 predicted targets in the constituent–target network. Forty-two hub genes with core regulatory effects were extracted from the PPI network, and 134 compounds and 29 crucial disease pathways were implicated in the target–constitu ent–disease network. Twelve disease pathways attributed to the Lung–Large Intestine meridians, with six and five attributed to the Kidney–Urinary Bladder and Stomach–Spleen meridians, respectively. Onehundred and eighteen candidate constituents showed a high binding affinity with SARS-coronavirus-23-chymotrypsin-like protease(3 CLpro), as indicated by molecular docking using computational pattern recognition. The in vitro activity of 22 chemical constituents of RDS was validated using the 3 CLproinhibition assay. Finally, using liquid chromatography mass spectrometry in data-independent analysis mode,the presence of seven out of these 22 constituents was confirmed and validated in an aqueous decoction of RDS, using reference standards in both non-targeted and targeted approaches.Conclusion: RDS acts primarily in the Lung–Large Intestine, Kidney–Urinary Bladder and Stomach–Spleen meridians, with other Zang-fu viscera strategically covered by all nine ingredients. In the context of TCM meridian theory, the multiple components and targets of RDS contribute to RDS’s dual effects of healthstrengthening and pathogen-eliminating. This results in general therapeutic effects for early COVID-19 control and prevention.

Single molecule electro-catalysis of non-fluorescent molecule

Single molecule catalysis is very powerful in revealing catalytic mechanism at the single molecule level.But fluorescent molecule is always necessary to take part into the catalysis directly in previous research.In order to study the single molecule electro-catalysis of non-fluorescent molecule（SMECNFM） on nanocatalyst, we couple the SMECNFM with a single molecule fluorescence reaction. A certain number of fluorescent molecules will be generated and detected when the SMECNFM happens. Through this method, we can detect the electro-oxidation reaction of one HCOONa molecule. The stability of Pt nanocatalyst supported on active carbon is studied at the single molecule level by this method. This paper also provides a general way to make ultra-sensitive sensor, and to study the SMECNFM for the molecules,such as formic acid, hydrogen, oxygen, etc., on single nanoparticle.

Prediction of mechanical properties of cold rolled strip based on improved extreme random tree

Taking the 2130 cold rolling production line of a steel mill as the research object,feature dimensionality reduction and decoupling processing were realized by fusing random forest and factor analysis,which reduced the generation of weak decision trees while ensured its diversity.The base learner used a weighted voting mechanism to replace the traditional average method,which improved the prediction accuracy.Finally,the analysis method of the correlation between steel grades was proposed to solve the problem of unstable prediction accuracy of multiple steel grades.The experimental results show that the improved prediction model of mechanical properties has high accuracy:the prediction accuracy of yield strength and tensile strength within the error of±20 MPa reaches 93.20%and 97.62%,respectively,and that of the elongation rate under the error of±5%has reached 96.60%.