BACKGROUND:Telomerase reverse transcriptase (TERT) plays an important role in telomerase activation.
OBJECTIVE:To construct the targeting short hairpin RNA plasmid vector expressing TERT gene from astrocytes by using pLentilox3.7.U6.
METHODS:By using two sequences from TERT gene, we synthetized sense and antisense strand template sequences of RNA interference molecular in vitro, and then obtained the complementary strands through annealing procedure. We connected the strands with pLentilox3.7.U6 that was sequenced and transfected into the Escherichia coli. In the end, we tested its effect of reducing the TERT gene expressing by using cultured astrocytes from rat spinal cord in vitro through western blot and immunofluorescence technique.
RESULTS AND CONCLUSION:Western blot and immunofluorescence assay showed that, compared with the control group, the interference groups had a lower TERT expression in astrocytes. The targeting short hairpin RNA plasmid vector expressing TERT gene is useful to reduce the TERT gene expression. The targeting short hairpin RNA plasmid vector expressing TERT gene is valid for us to do the further test learning the mechanism of astrocytes in spinal cord injury.

Objective To investigate the correlation of left ventricular ejection fraction (LVEF) as well as serum levels of NT-proBNP, Hcy and D-Dimer (D-D) with different traditional Chinese medicine (TCM) syndrome types of chronic heart failure (CHF). Methods A total of 178 CHF patients were divided into heart function normal ejection fracture group (HF?NEF, n=86) and heart function reduction (HFREF, n=92) according to their LVEF performance. Another 35 cases with nor?mal cardiac function were included in control group. All CHF patients was also divided into 3 TCM syndrome types:both de?ficiency of Qi and Yin syndrome group(n=64),Qi asthenia causing blood stasis syndrome group(n=59) andYang defi?ciency water stop group (n=55). All patients were examined with cardiac color doppler and LVEF values were recorded. And serum NT-proBNP、Hcyand D-D levels were all quantified. Results As to serum levels of NT-proBNP, Hcy and D-D, they were higher in HFREF group than those in HFNEF group than those in control group. On the other hand, LVEF was lowest in HFREF group but highest in control group. All differences were statistically significant (P<0.05). Among patient in HFNEF group, LVEF in theYang deficiency water stop groupwas lower than that inboth deficiency of Qi and Yin syn?drome group(P<0.05). Serum levels of NT-proBNP, Hcy, and D-D were not significantly different between different TCM syndrome groups. By contrast, among patients in HFREF group, LVEF values did not differ significantly between different TCM syndrome groups. Serum level of NT-proBNP was lower inboth deficiency of Qi and Yin syndrome groupthan that inQi asthenia causing blood stasis syndrome group than that in Yang deficiency water stop group. As to serum levels of Hcy and D-D, they are higher inYang deficiency water stop groupthat those inboth deficiency of Qi and Yin syndrome group and Qi asthenia causing blood stasis syndrome group(P<0.05). Conclusion Patients with different TCM syn?drome types of CHF present different levels of serum NT-proBNP, Hcy, D-D level and LVEF. Changes of indicators in HFREF groups are more obvious than they did in HFNEF group.

BACKGROUND:Telomerase reverse transcriptase (TERT) plays an important role in telomerase activation, however there is rare report addressing the construction of the lentivirus targeted its genes to inhibit its expression in the spinal cord astrocytes.
OBJECTIVE:To construct recombinant lentivirus vector expressing smal interfering RNA against TERT gene and to evaluate its potential for inhibiting the TERT expression.
METHODS:After shRNA-TERT sequence was designed and synthesized, the sequence was amplified by PCR and then connected to plasmid pLentilox3.7U6-hTERT to construct recombinant plasmid. The recombinant plasmid was then transfected to DH5αcel s to screen positive colony, and the sequence was identified. The recombinant plasmid pLentilox3.7U6-TERT was transfected in 293T cel s, generating recombinant lentivirus Le-TERT. The titer of recombinant lentivirus was determined and Le-TERT was transfected into the rat spinal cord astrocytes. The expression of TERT in astrocytes was detected by RT-PCR, western blot and immunofluorescence assay.
RESULTS AND CONCLUSION:The gene sequencing analysis confirmed that, recombinant plasmid pLentilox3.7U6-TERT was successful y constructed. The real-time quantitative PCR, western blot analysis and immunofluorescence assay indicated that, after Le-TERT was transfected in the astrocytes for 4 days, the inhibition rate of TERT mRNA was (63.98±2.6)%, and Le-TERT was lowly expressed in the transfected astrocytes. Recombinant expression vector pLentilox3.7U6-TERT can produce the lentivirus at high titer and effectively inhibit TERT expression in the transfected astrocytes.

BACKGROUND:The main treatment of lumbar isthmic spondylolisthesis is the surgery, in a broader attempt to decompression, reduction, fixation and fusion of the lesioned segments. The golden standard of the treatment is biological fusion, while internal fixation is a reliable assistance for fusion therapy. ＜br> OBJECTIVE:To discuss the clinical value and curative effect of intervertebral fusion cage combined with pedicle screw systems for the treatment of lumbar isthmic spondylolisthesis. ＜br> METHODS:From March 2010 to March 2013, 21 cases of isthmic spondylolisthesis were treated with intervertebral fusion cage combined with pedicle screw systems, including 18 cases of spondylolisthesis of degree II and 3 cases of spondylolisthesis of degree III. Al patients were fol owed up regularly, taking JOA lumbago score and visual analog scale score as the objective evaluation criteria of pain in postoperative fol ow-ups. The curative effect was assessed by Macrab standard, and the functional recovery was evaluated based on indicators such as Prolo, and the spinal fusion rate was assessed according to Lenke criteria. Changes of slippage rate, slippage angle, sacral inclination angle and intervertebral space post height in preoperative and postoperative periods were evaluated by iconography data. ＜br> RESULTS AND CONCLUSION:Al the 21 patients with isthmic spondylolisthesis were fol owed up for 12-16 months. JOA lumbago score and vasual analog scale score of al patients were improved after treatment, and the difference was statistical y significant compared with before treatment (P=0.000). According to Macrab evaluation criteria, there were 17 excellent cases and 4 good cases. Each indicator evaluated by preoperative Prolo activities and symptom grading showed significant differences in preoperative and postoperative periods (P=0.003). Postoperative lumbar spondylolisthesis was basical y reset, the slippage angle was significantly reduced, the sacral inclination angle was increased, and the height of the intervertebral space was recovered basical y. Intervertebral fusion cage combined with pedicle screw systems was one of the effective strategies to treat lumbar isthmic spondylolisthesis.

Objective To investigate the relationship between telomerase reverse transcriptase (TERT) gene expression and astrocyte activation.Methods Twenty neonatal 3-day-old male SD rats were used for culture of the astrocytes.The astrocytes were divided into Group A (activated,non-transfected astrocytes),Group B (activated,transfected astrocytes),Group C (unactivated astrocytes) and Group D (activated,empty plasmid transfected astrocytes) according to the random number table,with 5 rats per group.The cell proliferation rate in each group was detected by cell counting kit-8 (CCK-8) ;TERT expression by immunocytochemical method; expressions of TERT and glial fibrillary acidic protein (GFAP) genes by RT-PCR assay.Results Astrocytic proliferation ability in Group B lowered significantly as compared with that in Groups A,D and C (F =43.418,P ＜ 0.01).Expressions of TERT and GFAP mRNAs in Groups A and D were significantly higher than those in Group B and C,and no significant difference was found between Groups A and D.Besides,there was a linear correlation between mRNAs expressions of both genes in Groups A and D (r =0.701,0.704,P ＜ 0.01),while no significant linear correlation was observed in Groups B and C (r =0.260,P ＞ 0.05).Expressions of TERT and GFAP proteins in Groups A and D were markedly higher than those in Groups B and C and no significant difference was found between Groups A and D.Conclusion TERT genes are involved in the activation of astrocytes and exert effect on promoting the activation of astrocytes.

Objective To evaluate the risk factors of fluil-term infants with low birth weight.Methods All related Chinese and English literatures published from 1980 to February 25,2016 were collected from CBM,CNKI,Wang Fang Data,Medline and Embase databases,and screened with inclusion and exclusion criteria and Stata13.0 software was used in this Meta-analysis.Results Twenty three studies were included and there were 278 020 subjects.Female infants (pooled OR=1.60,95％CI:1.49-1.72),less antenatal care visits (pooled OR=1.81,95％CI:1.54-2.11),maternal passive smoking (pooled OR=1.49,95％CI:1.08-2.06),pregnancy-induced hypertension (pooled OR=2.96,95％CI:1.85-4.74) and hypamnion (pooled OR=2.71,95％CI:1.87-3.93) were the risk factors for fluil-term infants with low birth weight.Conclusion Departments of maternal and health care should encourage pregnant women to have antenatal care visits to find and treat their pregnancy complications,and avoid passive smoking actively through health education for the purpose to prompt the birth quality of infants.

Acute respiratory distress syndrome(ARDS)is a highly fatal syndrome in the intensive care unit(ICU),with a mortality rate of up to 40%.Early identification and treatment of ARDS are essential to improve the prognosis.Machine learning,the core of artificial intelligence and data science,is a set of computer tools designed to acquire new knowledge from existing data,which can assist medical staff in making clinical decisions.In recent years,machine learning has been increasingly used in the clinical diagnosis,precision treatment,and prognosis assessment of ARDS,which is expected to generate new ideas for diagnosing and treating ARDS.This article summarizes the application of machine learning in the clinical diagnosis of ARDS,classification of ARDS,treatment of ARDS,prognosis evaluation of ARDS,and the shortcomings of machine learning in the application of ARDS to explore the research progress of machine learning in diagnosing and treating ARDS and provide directions for further research.

OBJEVTIVETo investigate the expression of cysteine-rich secretory protein 2 (CRISP2) in spermatozoa of patients with asthenozoospermia and explore its clinical significance.

METHOSSemen samples were collected from 24 normal volunteers and 24 patients with asthenozoospermia for detecting CRISP2 mRNA and protein expressions using qRT-PCR and Western blotting, respectively. The correlation of CRISP2 expressions with sperm morphology, progressive motility and fertility prognosis were analyzed in patients with asthenozoospermia.

RESULTSCRISP2 protein expression was obviously lowered in the ejaculated spermatozoa of patients with asthenozoospermia as compared to the normal volunteers, but no significant difference in CRISP2 mRNA expression was found between the two groups. Correlation analysis showed that CRISP2 protein expression was positively correlated with normal sperm morphology (r=0.6182, P=0.0037) and progressive motility (r=0.6309, P=0.0029). Follow-up study of the patients revealed a higher fertility rate in patients with a relatively high CRISP2 protein expression than in those with low CRISP2 protein expression (80.0% vs 20.0%, P=0.0230).

CONCLUSIONThe expression level of CRISP2 protein is positively correlated with normal sperm morphology and progressive motility. A reduced CRISP2 protein expression indicates poor fertility prognosis of patients with asthenozoospermia, suggesting the potential value of CRISP2 as a novel therapeutic target for treating asthenozoospermia.

Asthenozoospermia ; metabolism ; Case-Control Studies ; Fertility ; Follow-Up Studies ; Glycoproteins ; metabolism ; Humans ; Male ; RNA, Messenger ; Sperm Motility ; Spermatozoa ; metabolism

OBJECTIVE: To investigate the role of MTBP in regulating the migration and invasion of human prostate cancer cells. METHODS: The baseline expressions of MTBP in 3 different human prostate cancer cells lines (22RV1, DU145 and Lncap) were detected using Western blotting. The cells were transfected with a small interfering RNA (siRNA) for MTBP knockdown or MTBP plasmid for MTBP overexpression, and 48 h later, the cells were examined for MTBP expression with Western blotting; the changes in the migration abilities of the cells were evaluated using wound healing assay and Transwell assay, and the cell invasiveness was assessed using Matrigel Transwell assay. The expression of E-cadherin protein, a marker of epithelial mesenchymal transition (EMT), was detected using Western blotting. RESULTS: MTBP expression was the highest in DU145 cells followed by Lncap cells, and was the lowest in 22RV1 cells, indicating a positive correlation of MTBP expression with the level of malignancy of human prostate cancer cells. Transfection of the cells with siRNA or MTBP plasmids efficiently lowered or enhanced the expressions of MTBP in human prostate cancer cells. Wound healing assay showed that inhibition of MTBP expression decreased the migration ability of the prostate cancer cells, and MTBP overexpression significantly promoted the migration of the cells ( < 0.01). Transwell assay showed that MTBP knockdown significantly lowered the migration and invasion ability of the cells, while MTBP overexpression markedly increased the number of migrating and invading cells ( < 0.01); Western blotting results showed that MTBP knockdown increased the expression of E-cadherin protein, and MTBP overexpression decreased E-cadherin expression in the prostate cancer cells. CONCLUSIONS MTBP overexpression promotes the migration and invasion of human prostate cancer cells possibly relation to the induction of EMT.
Antigens, CD ; metabolism ; Cadherins ; metabolism ; Carrier Proteins ; genetics ; metabolism ; Cell Line, Tumor ; Cell Movement ; Epithelial-Mesenchymal Transition ; Gene Expression Regulation, Neoplastic ; Gene Knockdown Techniques ; Humans ; Male ; Neoplasm Invasiveness ; Prostatic Neoplasms ; metabolism ; pathology ; RNA, Small Interfering ; Transfection

Protein lysine methylation is a prevalent post-translational modification (PTM) and plays critical roles in all domains of life. However, its extent and function in photosynthetic organisms are still largely unknown. Cyanobacteria are a large group of prokaryotes that carry out oxygenic photosynthesis and are applied extensively in studies of photosynthetic mechanisms and environmental adaptation. Here we integrated propionylation of monomethylated proteins, enrichment of the modified peptides, and mass spectrometry (MS) analysis to identify monomethylated proteins in Synechocystis sp. PCC 6803 (Synechocystis). Overall, we identified 376 monomethylation sites in 270 proteins, with numerous monomethylated proteins participating in photosynthesis and carbon metabolism. We subsequently demonstrated that CpcM, a previously identified asparagine methyltransferase in Synechocystis, could catalyze lysine monomethylation of the potential aspartate aminotransferase Sll0480 both in vivo and in vitro and regulate the enzyme activity of Sll0480. The loss of CpcM led to decreases in the maximum quantum yield in primary photosystem II (PSII) and the efficiency of energy transfer during the photosynthetic reaction in Synechocystis. We report the first lysine monomethylome in a photosynthetic organism and present a critical database for functional analyses of monomethylation in cyanobacteria. The large number of monomethylated proteins and the identification of CpcM as the lysine methyltransferase in cyanobacteria suggest that reversible methylation may influence the metabolic process and photosynthesis in both cyanobacteria and plants.
Bacterial Proteins/metabolism* ; Lysine/metabolism* ; Methyltransferases/metabolism* ; Photosynthesis ; Protein Processing, Post-Translational ; Synechocystis/growth & development*