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Objective:To investigate the difference in left ventricular dysfunction between type 2 diabetes mellitus (T2DM) patients with hyperlipidemia and those without hyperlipidemia, and analyze the application value of three-dimensional speckle tracking technology.Methods:The clinical data of 70 patients with T2DM admitted to Heji Hospital Affiliated to Changzhi Medical College from January 2020 to June 2021 were retrospectively analyzed. Among these patients, 35 patients had hyperlipidemia and 35 patients had no hyperlipidemia. At the same time, 40 healthy subjects who concurrently underwent health checkups in the same hospital were included as healthy controls. All subjects underwent routine cardiac ultrasound and 3D-STE examinations. General clinical information and left ventricular function-related parameters, including global longitudinal strain, global circumferential strain, global area strain, and global radial strain were compared between healthy controls and T2DM patients with hyperlipidemia and those without hyperlipidemia.Results:The proportion of left ventricular remodeling increased in T2DM patients without hyperlipidemia, and the proportion of left ventricular hypertrophy was the highest in T2DM patients with hyperlipidemia. Global longitudinal strain and global circumferential strain in T2DM patients without hyperlipidemia were (-16.97 ± 2.59)% and (-17.41 ± 2.50)%, respectively, which were significantly higher than (-18.86 ± 2.46)% and (-18.71 ± 2.92)% in healthy controls ( t = 0.95, 0.57, both P < 0.05). Global longitudinal strain, global circumferential strain, and global area strain in T2DM patients with hyperlipidemia were (-14.98 ± 3.15)%, (-15.80 ± 3.16)%, (-27.17 ± 4.54)%, respectively, which were significantly higher than (-18.86 ± 2.46)%, (-18.71 ± 2.92)%, (-30.62 ± 4.02)% in healthy controls ( t = 0.46, 1.37, 0.98, all P < 0.05) and (-16.97 ± 2.59)%, (-17.41 ± 2.50)%, (-30.06 ± 3.59)% in T2DM patients without hyperlipidemia ( t = 0.37, 1.02, 0.77, all P < 0.05). Global radial strain in T2DM patients with hyperlipidemia was significantly higher than [(51.49 ± 8.94)%, t = 1.35, P < 0.05] in healthy controls and [(47.71± 8.46)%, t = 0.98, P < 0.05] in T2DM patients without hyperlipidemia. In patients with T2DM, fasting blood glucose and hyperlipidemia were independently correlated with all strain-related parameters. Conclusion:Hyperlipidemia can aggravate left ventricular remodeling and dysfunction in patients with T2DM and 3D-STE is one of the examination methods for subclinical left ventricular remodeling and dysfunction in T2DM patients with or without hyperlipidemia.
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Objective:To construct a recombinant herpes simplex virus 2 (HSV-2) expressing enhanced green fluorescent protein (EGFP) using clustered, regularly interspaced, short palindromic repeat/CRISPR-associated nuclease 9 (CRISPR/Cas9) technology.Methods:Four strategies for inserting exogenous EGFP gene into HSV-2 genome using CRISPR/Cas9 technology were designed: (1) conventional homology-directed repair: circular two homology arm donor-mediated gene knock-in; (2) linearized single homology arm donor-mediated gene knock-in; (3) homology-independent targeted integration; (4) conventional homology-directed repair-mediated by cell lines stably expressing Cas9 and sgRNA.Results:The recombinant virus HSV-2-EGFP was successfully constructed based on the second, the third and the fourth strategies. The second strategy was the most efficient, followed by the third and the fourth strategies. The purified recombinant virus could stably express green fluorescent protein in seven passages and shared similar growth characteristics in Vero cells to the parental virus.Conclusions:Linearized single homology arm donor could increase the efficiency of gene knock-in, and cell lines stably expressing Cas9 and sgRNA could increase the efficiency of gene knock-in mediated by homology-directed repair.
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Objective:To investigate the effects of genomic location of a foreign gene in Shanghai-191 strain measles virus (MV) vector on gene expression and virus replication.Methods:The nucleotide sequence encoding S1 protein of SARS-CoV-2 was inserted at different positions in MV antigenome (the upstream of the N gene, between P and M genes, between H and L genes), and co-transfected into 293T cells with helper plasmids coding T7 RNA polymerase and N, P, and L proteins, respectively. The transfected cells were lysed and the supernatants were used to infected Vero cells to harvest recombinant viruses. S1 proteins expressed by the recombinant viruses were identified by RT-PCR, indirect immunofluorescence assay, Western blot and ELISA. Growth kinetics of the recombinant viruses were analyzed.Results:Recombinant viruses were failed to be rescued when the S1 protein-coding sequence was cloned into the upstream of N gene. Two recombinant viruses, MV-M-S1 and MV-L-S1, were successfully rescued when cloning the S1 protein-coding sequence into the intergenic region between P and M genes, or H and L genes, and could express S1 protein. MV-M-S1 expressed more S1 protein than MV-L-S1, but the titer of MV-M-S1 was lower.Conclusions:Inserting a foreign gene at different positions in the MV genome might have different effects on gene expression and virus replication. This study provided reference for the subsequent construction of MV vector.
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Objective@#To develop a real-time quantitative PCR (qPCR) assay for detecting Sendai virus gene copy number.@*Methods@#The recombinant plasmid was constructed and a reference used to establish the real-time qPCR method with the TaqMan probe and verified for reproducibility and sensitivity.@*Results@#The linear range of the developed real-time qPCR assay was 1.024 × 103 ~5 × 1010 copies/μl, with an R2 value of more than 0. 99. The standard recovery of the tested samples was 84.56%~119.48%.@*Conclusions@#Compared with traditional hernagglutination assay for particle number detection, this method has higher sensitivity, better repeatability and high quantitative accuracy. It can be used for the detection of particle number of vaccine with sendai virus as the carrier, so as to detect the specific activity of vaccine products.
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In this paper, we designed and evaluated a duplex detection strategy for microRNAs (miRNAs) using universal probe-based target-triggered double hybridization and fluorescent microsphere-based assay system (xMAP array). In the absence of target miRNA, reporter DNA cannot hybridize stably with the immobilized capture DNA due to its low melting temperature. Only after adding target miRNA, can reporter probe hybridize with capture probe to form a stable three-component complex. This targettriggered stable hybridization makes this method possible for highly selective and sensitive detection of multiple miRNAs. We exemplified a quantitative detection of duplex miRNAs with a limit of detection of 40 pM. The xMAP array platform holds the potential of extending this approach to simultaneous detection of up to 100 miRNA targets. Considering the simplicity, rapidity and multiplexing, this work promised a potential detection of multiple miRNA biomarkers for early disease diagnosis and prognosis.
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Objective:To explore the expression of R-spondin family in colorectal cancer tissues and adjacent tissues,and to evaluate its relationship with clinic-pathological stage.Methods:A total of 64 samples of colorectal cancer tissues and adjacent tissues were collected from the patients,who received radical surgery in Xiangya Hospital,Central South University between January 2014 and August 2014.The mRNA and protein expression levels of R-spondin 1-4 and β-catenin in the colorectal cancer tissues and adjacent tissues were detected by qRT-PCR and immunohistochemistry.The relationship between the expression level of R-spondin 1-4 and the clinic-pathological factors were analyzed to explore the correlation between the expression level of R-spondin 1-4 and β-catenin in colorectal cancer.Results:Compared with the adjacent tissues,the mRNA and protein expression levels of R-spondin 1 were elevated in the colorectal cancer tissues (P＜0.05).The mRNA and protein expression levels of R-spondin 2-4 were increased in the colorectal cancer tissues than those in the normal tissues (P＜0.05),but there was no significant difference between the colorectal cancer tissues and adjacent tissues (P＞0.05).The expression level of R-spondin i was positively correlated with the nuclear expression of β-catenin in the colorectal cancer tissues (r=0.6307,P＜0.05).Conclusion:Compared with the adjacent tissues,the mRNA and protein expression levels of R-spondin 1 are significantly elevated in the colorectal cancer tissues.R-spondin 1 may play a role in promoting carcinogenesis by regulating the activity of β-catenin in the downstream of Wnt signaling pathway.
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Objective To optimize the extraction methods of mitochondrial genome DNA(mtDNA)of Oncomelania hupen-sis. Methods The pyrolysis,protein K variable-temperature digestion and high-concentration potassium acetate purification were applied to optimize the high-concentration-salt precipitation method,and then the optimized method was compared with two common extraction methods,the sucrose density gradient centrifugation method and traditional high-concentration-salt pre-cipitation method. The mtDNA samples were identified by using spectrophotometry,agarose gel electrophoresis and the amplifi-cation products of COX1. The nuclear DNA contamination was tested by the amplification products of ITS. Results The concen-tration and yield of the improved method was significantly higher than those of the traditional method(F=3032.65,10185.00, both P<0.01). The mtDNA samples extracted were essentially free of nuclear DNA and protein,meeting PCR,sequence analy-sis and other molecular biology research requirements. Conclusions The improved high-concentration-salt precipitation meth-od for isolating mtDNA is simple,and it has high yield and low cost. The extracted mtDNA can meet relevant analysis require-ments.
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Objective To investigate effects of diabetes mellitus (DM) on pharmacokinetics (PK) of cephradine (CED).Methods DM model was induced by iv.alloxan 60 mg·kg-1.A reversed phase HPLC internal standard method was developed for measurement of CED plasma concentration.After blood collection,rats were sacrificed to collect kidneys for calculating kidney index(KW/BW).DM and normal control (CTL) rats were randomly assigned to receive iv.or ig.CED at a dose of 180 or 90 mg·kg-1.The 3p97 program was used to calculate PK parameters.Results The developed HPLC method was validated to have high specificity,precision,recovery and good storage stability,and met requirements for PK study of CED.The CED in rats of both DM and CTL groups showed the iv.two-compartment PK and ig.one-compartment PK and followed the first-order kinetics.Following iv.dosing,a remarkably decreased t1/2β and MRT,increased CLt were evident in DM group as compared with CTL group (P ＜ 0.05).After ig dosing,a significant decrease in t1/2k and t a remarkable increase in CLt and Cm=were observed for DM group as compared with CTL group (P ＜ 0.05).The DM rats showed a trend of decreased t1/2ka vs CTL rats.There was no significant difference in the oral bioavailability between the two groups (P ＞ 0.05).KW and KW/BW in DM group were increased remarkably compared with CTL group (P ＜ 0.05).Conclusion The DM vs CTL results in faster absorption and elimination of CED in rats,but does not have significantly affect in oral bioavailability.The compensatory hypertrophy and hyperfunction of early-stage diabetic kidneys may constitute one of causes of quick elimination of CED in rats with DM.
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Objective To explore the role and mechanisms of ω-3 polyunsaturated fatty acids (ω-3PUFA) alone or in combination with dexamethasone ( DEX) in inducing cell apoptosis and anti -proliferation in multiple myeloma(MM).Methods DEX resistance MM cell line MM1R were treated with different concentra-tions of Eicosapentaenoic acid(EPA)or Docosahexaenoic acid(DHA)alone or in combination with DEX for 24hrs or 48hrs.Cell proliferation was detected by MTT .Cell cycle and apoptosis were measured by flow cytometry .The levels of apoptosis related proteins were analyzed by Western blot .Results The proliferation of MM1R was in-hibited by different concentrations (10,20,50,100μM)of EPA or DHA alone or in combination with 10μM DEX in a dose-and time-dependent manner .Inhibition effect was significantly higher in combinative groups than in single agent groups(P<0.05).The percentage of G0/G1 phase and cell apoptosis rate in MM1R treated with dif-ferent concentrations of EPA or DHA alone was increased in a dose -dependent manner ,and being significantly higher in combinative groups than in single agent groups (P<0.05).The expressive levels of cleaved caspase -3 and Bax were up-regulated ,while pro-caspase-3 and BCL-2 were down-regulated in a dose-dependent manner.Conclusion ω-3PUFA can inhibit DEX resistant MM cell proliferation ,arrest cell cycle and induce cell apoptosis ,and has a synergistic anti -resistant effect in combination with DEX ,may serve as a new ,effective MM drugs.
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Dengue virus(DENV)is an enveloped single positive-stranded RNA virus that belongs to the Flaviviridae family. It is the cause of dengue fever,life-threatening dengue hemorrhagic fever and den-gue shock syndrome. Aedes aegypti and Aedes albopictus are responsible for the wide transmission of DENV in tropical and subtropical regions throughout the world. Currently,there have been several dengue vaccines entering clinical trials,including the Sanofi Pasteur chimeric yellow fever dengue tetravalent vaccine (CYD),which has been licensed for use in some countries. However,CYD does not provide adequate pro-tection against all four serotypes of DENV and induces severe dengue diseases in young and seronegative vac-cine recipients. Therefore,a more efficacious dengue vaccine is still needed. Here,we reviewed the oppor-tunities and challenges for the development of dengue vaccines.
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Objective A major component of flow cytometry data analysis involves gating , which is the process of identifying homogeneous groups of cells .As manual gating is error-prone, non-reproducible, nonstandardized, and time-consuming , we propose a time-efficient and accurate approach to automated analysis of flow cytometry data .Methods Unlike manual analysis that successively gates the data projected onto a two-dimensional filed, this approach, using the K-means clustering results , directly analyzed multidimensional flow cytometry data via a similar subpopulations-merged algorithm.In order to apply the K-means to analysis of flow cytometric data , kernel density estimation for selecting the initial number of clustering and k-d tree for optimizing efficiency were proposed .After K-means clustering , results closest to the true populations could be achieved via a two-segment line regression algorithm .Results The misclassification rate (MR) was 0.0736 and time was 2 s in Experiment One, but was 0.0805 and 1 s respectively in Experiment Two. Conclusion The approach we proposed is capable of a rapid and direct analysis of the multidimensional flow cytometry data with a lower misclassification rate compared to both nonprobabilistic and probabilistic clustering methods .
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Objective:To investigate the effect of activin A and nerve growth factor ( NGF) NGF on stimulating neurite outgrowth of dorsal root ganglia(DRG)of the embryonic chicken.Methods:In this study,we observed that activin A and NGF together induced neurite outgrowth of DRG and kept survival of DRG neurons by the primary cultured DRGs from embryonic day 8 ( E8 ) chicken.calcitonin gene-related peptide(CGRP)CGRP mRNA expressions were analyzed by RT-PCR.Results: The DRG treated with activin A +NGF had obvious neurite outgrowth ,compared with only NGF group on day 3,and the number of living DRG neurons also increased.Activin A +NGF up-regulated the mRNA expressions of CGRP in DRG.Conclusion:The Data demonstrated that activin A with NGF can synergistically stimulate DRG neurite outgrowth and maintain the DRG neurons survival , suggesting that it is more effective that NGF and activin A together treat the associated disease of nerve system.
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Objective To introduce the outcomes of tracheal resection combined with primary end-to-end anastomosis for benign cervical tracheal stenosis,and to discuss the strategy for prevention of surgical complications.Methods The clinical data of 22 patients due to different causes benign cervical tracheal stenosis surgery were analyzed retrospectively.Results The length of cervical tracheal stenosis ranged was 2.2-4.2 cm.Grade Ⅱ stenosis was present in 6 patients.Grade Ⅲ stenosis was present in 11 patients and grade Ⅳ stenosis in 5 patients.Successful extubation was achieved in all 22 cases.After surgery,temporary hoarseness occurred to 1 case;unilateral pulmonary atelectasis with pleural effusion occurred to 1 case; subcutaneous emphysema with infection occurred to 1 case; mild dysphagia occurred to 3 cases;a slight deepening of the tone of voice in 10 patients with women occurred to 5 cases,granulation tissue growth near the suture occurred to 3 cases,and suture dehiscence did not occur in any patient.The follow-up period ranged from 6-45 months,no patient developed restenosis.Conclusions It presents a high success rate and good functional result of tracheal resection combined with primary end-to-end anastomosis.Therefore,it is an effective and reliable approach for the management of benign cervical tracheal stenosis.To avoid complications,the preoperative assessment,patients selection and postoperative management should be emphasized.
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ObjectiveTo explore the changes of astrocytes in vitro on the expression of Cyclin D1 under normal and hypoxia/reoxygenation conditions.MethodsThe primary astrocytes were isolated from the cortex of SD fetal rats(less than 24 hours) and identified by immunosytochemical method with anti-GFAP antibody after 3 passages.Astrocytes of passage 3 were divided into normal group ( control group),hypoxia/reoxygenation group ( H/R 37℃ ),and mild hypothermiaintervention group( H/R 32℃ )separately.Astrocytes from the later tow groups were reoxygenated with 4,12,and 20 hours separately after exposed to hypoxia conditions for 8 hours.Trypan blue staining was employed to detect the survival rates and immunofluorescence,western-blot were used to analyse the expressin of Cyclin D1 of of astrocytes of different groups and time points.Results 1.The GFAP positive astrocytes from passage 3 exceeded 95 ％.2.With regard to morphology and survival rates,there is no difference between astrocytes of normal and hypothermia groups after 8 hours exposure to hypoxia conditions.Reoxygenation could obviously rise astrocytes mortality with time went by ( H/R 37 ℃ group:12.87 ± 2.76 ( R4 ),31.55 ± 3.00 ( R12 ),46.40 ±8.50(R20) ;H/R 32℃ group:6.77 ± 1.53( R4),15.97 ±4.00(R12),28.33 ±5.69(R20) ;all P＜0.05).3.Immunofluorescence and western-blot revealed that reoxygenation increased Cyclin D1expression markedly,which was proportional to the duration of reoxygenation.Mild hypothermia could reduce Cyclin D1 expression of astrocytes severely under reoxygenation condition.ConclusionCyclin D1 expression can be regarded as a sensitive index of damage to astrocytes caused by hypoxia/reoxygenation conditions.
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BACKGROUND: Acellular dermalmatrix (ADM), as a new material, is generally applied to tissue repair after head and neck tumorectomy. However, hypopharyngeal carcinoma surgery always involves in repair and functional reconstruction of defected tissue; therefore, the application effect of ADM needs to be further studied for repairing laryngopharynx defect. OBJECTIVE: To evaluate the clinical effect of ADM in repairing laryngopharynx defect following tumorectomy. DESIGN, TIME AND SETTING: A retrospective case analysis was performed at the Department of Otolaryngology-Head and Neck Surgery, Institute of Battle Surgery, Daping Hospital, the Third Military Medical University of Chinese PLA from May 2005 to June 2006.PATICIPANTS: A total of 25 male cases aged 45-68 years including 22 patients with tumor in the lateral wall of the unilateral piriform fossa and 3 patients with in the posterior wall of laryngopharynxat were finally diagnosed as squamous cell carcinoma. METHODS: According to clinical stages, patients underwent partial or complete laryngopharyngectomy. ADM was used to repair oropharynx mucosa following the laryngopharyngectomy. However, ADM alone was used to repair piece-shaped oropharynx mucosa, limiting by being unable to complete reconstruction of spatial structure. Therefore, myocutaneous flap or other materials combined with ADM was beneficial for reconstruction of laryngopharyngeal cavity following complete laryngopharyngectomy. MAIN OUTCOME MEASURES: The effect of ADM on functional reconstruction of swallowing, breathing, and pronouncing. RESULTS: A total of 25 patients were included in the final analysis. No patients suffered with the pharyngeal fistula. Only one case encountered the subcutaneous infection at day 20 after surgery, but recovering after regular change of dressing and treatment of antibiotics. About 15 patients who accepted the surgery of partial laryngopharyngectomy suffered with bucking, but after 30-60 days, the symptom relieved, even disappeared, in spite of 3 cases still bucking when they swallowed liquid diet. Breathing function was recovered, and tracheal cannula was removed 6-12 months after surgery. Ten of them who accepted total laryngopharyngectomy had resumption of normal oral diet after 20 days. The follow-up periods after the surgery lasted 12-30 months. The allograft became normal mucosa finally, without rejection or scar formation. The follow-up of 10 patients lasted more than 24 months, one case had relapse of tumor, still alive after the second surgery. CONCLUSION: ADM with perfect histocompatibility and easy operability, can be applied to repair deletion of laryngeal or pharyngeal tissue and recover functions of pronouncing, breathing, and swallowing after tumorectomy. The combined use of ADM and musculo-cutaneous flap was effective and satisfactory.
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Objective To evaluate the international prognostic index (IPI) in peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS). Methods From May 2005 to May 2008, 75 patients of PTCL--NOS were reviewed. All the patients were diagnosed again by immunohistochemical staining. According to IPI, they were divided into four groups:low risk (0-1), intermediate-low(2), intermediate-high(3), high risk (4-5), then the difference of treatment effectiveness and prognosis among them were analysed. Results IPI scoring of 75 patients were classified as low risk , 10 (13.3%); as intermediate-low, 14 (18.7%); as intermediate-high, 28 (37.3 %); as high risk, 23 (30.7%). There was a significant difference in complete remission rates with first line treatment(X2=16.677,P=0.001), and overall survival rates (P=0.0000) among four groups. Median survival time among 4 groups were 36+, 29.00, 17.00, 10.00 months. 1-year OS were 100.00 %, 89.05 %, 64.24 %, 15.73 %; 2-year OS were: 75.00 %, 53.01%, 34.42 %, 2.00 % respectively. Multivariate analysis showed that both complete remission rates of first line treatment(P=0.002) and IPI(P = 0.049) were independent prognostic factor for PTCL-NOS, while single index of IPI was not. Conclusion At a certain extent, IPI model was able to predict response of treatment effective and prognosis in PTCL-NOS.
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Objective To study the expression of NF-κB p50 in nodal peripheral T-cell lymphomasunspecified (PTCL-U),and investigated the relationship between NF-κB and PTCL-U's complex biological behavior. Methods 51 patients with nodal PTCL-U were analysed by detecting the expression of NF-κB p50, p170 by immunohistochemistry and correlation between them and PTCL-U' s clinical feature, treatment effectiveness and prognosis were also studied. Results 11 patients(21.6 %, 11/51) and 31patients (60.8 %,31/51) were respectively positive for N F-κB p50 and p 170 expression. Expression of NF-κB were significantly correlated with p170 expression, poor performance status (PS＞2) and non-complete remission in first line treatment(Spearman correlation= 0.459, 0.313, 0.284; P = 0.001, 0.025, 0.044). Overall survival rate of NFκB p50-positive PTCL-U was significantly lower than that of NF-κB p50-negative patients by Log-Rank test (P =0.0451). Multivariate analysis showed poor performance and higher Ki-67 were independent prognostic factor for PTCL-U, while NF-κB p50 was not. Conclusion The expression of NF-κB pS0 was correlated with muhidrug resistance and poor prognosis in nodal PTCL-U.
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Objective To evaluate the efficacy and safety of desloratadine for the treatment of allergic rhinitis. Methods A total of 50 cases of chronic or acute allergic rhinitis confirmed clinically were randomized into control and experimental groups for double blind study. Scoring of clinical symptoms, examinations of the nasal cavity, blood and urine routine examination, functions of liver and kidney and electrocardiogram (ECG) were conducted on day 14 before and after the experiment. The collected data were analyzed statistically. Results The clinical data of the two groups were comparable. The effective rate of desloratadine was significant in allergic rhinitis, being 95.45%. No remarkable adverse effect was noted in this experiment. Conclusion Desloratadine is more effective and safe than loratadine in the treatment of allergic rhinitis and it may be feasible for clinical application.
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Objective To observe the relationship between hemoglobin (Hb) and vascular endothelial growth factor (VEGF) in serum of the patients with cancer. Methods Enzyme linked immunosorbent assay (ELISA) was used to detect VEGF levels in serum of the patients with cancers. Results The serum level of VEGF in the cancer group are significantly higher than that in the control group. The serum level of VEGF in patients with low level of hemoglobin ( 123.5 g/L) ( P