Abnormal adenosine metabolism of neutrophils inhibits airway inflammation and remodeling in asthma model induced by Aspergillus fumigatus.

BACKGROUND: Neutrophils consume a large amount of energy when performing their functions. Compared with other white blood cells, neutrophils contain few mitochondria and mainly rely on glycolysis and gluconeogenesis to produce ATP. The inflammatory site is hypoxic and nutrient poor. Our aim is to study the role of abnormal adenosine metabolism of neutrophils in the asthmatic airway inflammation microenvironment. METHOD: In this study, an asthma model was established by intratracheal instillation of Aspergillus fumigatus extract in Ecto-5'-Nucleotidase (CD73) gene-knockout and wild-type mice. Multiple analyses from bronchoalveolar lavage fluid (BALF) were used to determine the levels of cytokines and chemokines. Immunohistochemistry was used to detect subcutaneous fibrosis and inflammatory cell infiltration. Finally, adenosine 5'-(α, ß-methylene) diphosphate (APCP), a CD73 inhibitor, was pumped subcutaneously before Aspergillus attack to observe the infiltration of inflammatory cells and subcutaneous fibrosis to clarify its therapeutic effect. RESULT: PAS staining showed that CD73 knockout inhibited pulmonary epithelial cell proliferation and bronchial fibrosis induced by Aspergillus extract. The genetic knockdownof CD73 significantly reduced the production of Th2 cytokines, interleukin (IL)-4, IL-6, IL-13, chemokine (C-C motif) ligand 5 (CCL5), eosinophil chemokine, neutrophil IL-17, and granulocyte colony-stimulating factor (G-CSF). In addition, exogenous adenosine supplementation increased airway inflammation. Finally, the CD73 inhibitor APCP was administered to reduce inflammation and subcutaneous fibrosis. CONCLUSION: Elevated adenosine metabolism plays an inflammatory role in asthma, and CD73 could be a potential therapeutic target for asthma.

Intermittent Hypoxia Alleviates ß-Aminopropionitrile Monofumarate Induced Thoracic Aortic Dissection in C57BL/6 Mice.

OBJECTIVE: Thoracic aortic dissection (TAD) has a high mortality rate. Intermittent hypoxia (IH) triggers both harmful and beneficial effects in numerous physiological systems. The effects of IH on TAD development were explored in a mouse model. METHODS: ß-Aminopropionitrile monofumarate (BAPN) was used to induce TAD in C57BL/6 mice. Three week old male mice were treated with 1 g/kg/day BAPN in drinking water for four weeks and simultaneously subjected to IH (n = 30) (21%-5% O2, 90 s/cycle, 10 h/day, IH + BAPN group) or normoxia (n = 30) (21% O2, 24 h/day, BAPN group). Human VSMCs (HUASMCs) exposed to IH (30 min, 5% O2)/re-oxygenation (30 min, 21% O2) cycles with a maximum of 60 min/cycle to detect the effect of IH on HIF-1α and LOX via HIF-1α-siRNA. RESULTS: It was found that BAPN administration significantly increased the lumen size and wall thickness of aortas compared with the normal group, but was significantly reversed by IH exposure. Additionally, IH exposure significantly increased the survival rate of BAPN induced TAD (70% vs. 40%). Furthermore, IH exposure reduced BAPN induced elastin breaks and apoptosis of vascular smooth muscle cells. IH exposure also reversed BAPN induced upregulation of inflammation and extracellular matrix (ECM) degradation. Real time polymerase chain reaction (RT-PCR) confirmed that IH inhibited inflammation and ECM degradation related genes interleukin (IL)-1ß, IL-6, cathepsin S (Cat S), and matrix metalloproteinase 9 (MMP-9), but upregulated the ECM synthesis related genes lysyl oxidase (LOX) and collagen type I alpha2 (Col1a2) compared with the BAPN group. In vitro results suggest that IH promotes the expression of LOX via HIF-1α. CONCLUSION: The results suggest that IH alleviates BAPN induced TAD in C57BL/6 mice.

Mechanical stretch-induced endoplasmic reticulum stress, apoptosis and inflammation contribute to thoracic aortic aneurysm and dissection.

Thoracic aortic aneurysm/dissection (TAAD) is characterized by excessive smooth muscle cell (SMC) loss, extracellular matrix (ECM) degradation and inflammation. In response to certain stimuli, endoplasmic reticulum (ER) stress is activated and regulates apoptosis and inflammation. Excessive apoptosis promotes aortic inflammation and degeneration, leading to TAAD. Therefore, we studied the role of ER stress in TAAD formation. A lysyl oxidase inhibitor, 3-aminopropionitrile fumarate (BAPN), was administrated to induce TAAD formation in mice, which showed significant SMC loss (α-SMA level). Excessive apoptosis (TUNEL staining) and ER stress (ATF4 and CHOP), along with inflammation, were present in TAAD samples from both mouse and human. Transcriptional profiling of SMCs after mechanical stress demonstrated the expression of genes for ER stress and inflammation. To explore the causal role of ER stress in initiating degenerative signalling events and TAAD, we treated wild-type (CHOP(+/+)) or CHOP(-/-) mice with BAPN and found that CHOP deficiency protected against TAAD formation and rupture, as well as reduction in α-SMA level. Both SMC apoptosis and inflammation were significantly reduced in CHOP(-/-) mice. Moreover, SMCs isolated from CHOP(-/-) mice were resistant to mechanical stress-induced apoptosis. Taken together, our results demonstrated that mechanical stress-induced ER stress promotes SMCs apoptosis, inflammation and degeneration, providing insight into TAAD formation and progression.

[Report of 3162 cases of holmium laser enucleation of the prostate and review].

OBJECTIVE: To evaluate the safety, effectiveness, and outcomes of holmium laser enucleation of the prostate (HoLEP) for patients with symptomatic enlarged prostate after 11 years of experience. METHODS: The 3162 evaluable patients treated with holmium laser enucleation of the prostate at our institution between August 2001 and August 2011 were retrospectively analyzed. Study variables included International Prostate Symptom Score, quality of life, maximum urinary flow rate, and incidence of complications. RESULTS: HoLEP were performed successfully completed, not patients which occurs as electric cutting syndrome. The operation time was (60.8 ± 18.4) minutes; average resection of prostate quality was (45.4 ± 24.4) g. The hemoglobin reduce though surgery was (1.81 ± 0.93) g/L; percentage of red blood cell change was 1.24% ± 0.43%, and sodium blood drop was (1.14 ± 0.35) mmol/L. Postoperative patients of hospital stay (3.1 ± 1.1) days, average time of indwelling catheter time was (2.3 ± 0.8) days. Patients were followed up for 6-131 months time, an average of 32.4 months. Postoperative patients with international prostate symptom score progressive declined. The quality of life score was 2.2 ± 1.7, and it less than preoperative (5.7 ± 3.3, t = 2.447, P < 0.01). The time of follow-up droped further, and postoperative comparative differences have statistical significance (t = 2.179, 2.228, 2.306 and 2.365, P < 0.05). The maximum urinary flow rate also improved (P < 0.05). Postoperative complications included bladder neck contracture (4 cases), urinary tract infection (107 cases), urethral stricture (11 cases) and urinary incontinence (11 cases). The 11 patients reoperation. CONCLUSIONS: HoLEP treatment of benign prostatic hyperplasia could achieve the advantages of open surgery the same effect. It had fewer damage, faster recovery, fewer complications, and is a good treatment option.

Dynamic changes in cardiac morphology, function, and diffuse myocardial fibrosis duration of diabetes in type 1 and type 2 diabetic mice models using 7.0 T CMR and echocardiography.

Background: Diabetes mellitus (DM) is associated with an increased risk of cardiovascular disease (CVD). Hence, early detection of cardiac changes by imaging is crucial to reducing cardiovascular complications. Purpose: Early detection of cardiac changes is crucial to reducing cardiovascular complications. The study aimed to detect the dynamic change in cardiac morphology, function, and diffuse myocardial fibrosis(DMF) associated with T1DM and T2DM mice models. Materials and methods: 4-week-old C57Bl/6J male mice were randomly divided into control (n=30), T1DM (n=30), and T2DM (n=30) groups. A longitudinal study was conducted every 4 weeks using serial 7.0T CMR and echocardiography imaging. Left ventricular ejection fraction (LV EF), tissue tracking parameters, and DMF were measured by cine CMR and extracellular volume fraction (ECV). Global peak circumferential strain (GCPS), peak systolic strain rate (GCPSSR) values were acquired by CMR feature tracking. LV diastolic function parameter (E/E') was acquired by echocardiography. The correlations between the ECV and cardiac function parameters were assessed by Pearson's test. Results: A total of 6 mice were included every 4 weeks in control, T1DM, and T2DM groups for analysis. Compared to control group, an increase was detected in the LV mass and E/E' ratio, while the values of GCPS, GCPSSR decreased mildly in DM. Compared to T2DM group, GCPS and GCPSSR decreased earlier in T1DM(GCPS 12W,P=0.004; GCPSSR 12W,P=0.04). ECV values showed a significant correlation with GCPS and GCPSSR in DM groups. Moreover, ECV values showed a strong positive correlation with E/E'(T1DM,r=0.757,P<0.001;T2DM, r=0.811,P<0.001). Conclusion: The combination of ECV and cardiac mechanical parameters provide imaging biomakers for pathophysiology, early diagnosis of cardiac morphology, function and early intervention in diabetic cardiomyopathy in the future.

Change and impact of left ventricular global longitudinal strain during transcatheter aortic valve implantation.

BACKGROUND: Although transcatheter aortic valve implantation (TAVI) is a safe and effective treatment for aortic stenosis, it still carries some risks, such as valve leaks, stroke, and even death. The left ventricular global longitudinal strain (LVGLS) measurement may be useful for the prediction of adverse events during this operation. AIM: To explore the change of LVGLS during TAVI procedure and the relationship between LVGLS and perioperative adverse events. METHODS: In this study, 61 patients who had undergone percutaneous transfemoral TAVI were evaluated by transthoracic echocardiography. Before surgery, data on left ventricular ejection fraction (LVEF) and LVGLS were collected separately following balloon expansion and stent implantation. Difference in values of LVGLS and LVEF during preoperative balloon expansion (pre-ex), preoperative stent implantation (pre-im) and balloon expansion-stent implantation (ex-im) were also examined. Adverse events were defined as perioperative death, cardiac rupture, heart arrest, moderate or severe perivalvular leakage, significant mitral regurgitation during TAVI, perioperative moderate or severe mitral regurgitation, perioperative left ventricular outflow tract obstruction, reoperation, and acute heart failure. RESULTS: The occurrence of perioperative adverse events was associated with differences in pre-ex LVGLS, but not with difference in pre-ex LVEF. There were significant differences between pre-LVGLS and ex-LVGLS, and between pre-LVGLS and im-LVGLS (P = 0.037 and P = 0.020, respectively). However, differences in LVEF were not significant (P = 0.358, P = 0.254); however differences in pre-ex LVGLS were associated with pre-LVGLS (P = 0.045). Compared to LVEF, LVGLS is more sensitive as a measure of left heart function during TAVI and the perioperative period. Moreover, the differences in LVGLS were associated with the occurrence of perioperative adverse events, and changes in LVGLS were apparent in patients with undesirable LVGLS before the surgery. Furthermore, LVGLS is useful to predict changes in cardiac function during TAVI. CONCLUSION: Greater attention should be paid to the patients who plan to undergo TAVI with normal LVEF but poor LVGLS.

Short-term waterlogging-induced autophagy in root cells of wheat can inhibit programmed cell death.

Autophagy is a pathway for the degradation of cytoplasmic components in eukaryotes. In wheat, the mechanism by which autophagy regulates programmed cell death (PCD) is unknown. Here, we demonstrated that short-term waterlogging-induced autophagy inhibited PCD in root cells of wheat. The waterlogging-tolerant wheat cultivar Huamai 8 and the waterlogging-sensitive wheat cultivar Huamai 9 were used as experimental materials, and their roots were waterlogged for 0-48 h. Waterlogging stress increased the number of autophagic structures, the expression levels of autophagy-related genes (TaATG), and the occurrence of PCD in root cells. PCD manifested as morphological changes in the cell nucleus, significant enhancement of DNA laddering bands, and increases in caspase-like protease activity and the expression levels of metacaspase genes. The autophagy promoter rapamycin (RAPA) reduced PCD levels, whereas the autophagy inhibitor 3-methyladenine (3-MA) enhanced them. The expression levels of TaATG genes and the number of autophagic structures were lower in cortex cells than in stele cells, but the levels of PCD were higher in cortex cells. The number of autophagic structures was greater in Huamai 8 than in Huamai 9, but the levels of PCD were lower. In summary, our results showed that short-term waterlogging induced autophagy which could inhibit PCD. Mechanisms of response to waterlogging stress differed between cortex and stele cells and between two wheat cultivars of contrasting waterlogging tolerance.

Validating scores predicting atrial fibrillation recurrence post catheter ablation in patients with concurrent atrial fibrillation and pulmonary diseases.

BACKGROUND: Several scores were available for predicting atrial fibrillation (AF) recurrence post radiofrequency ablation. However, the role of different scores predicting AF recurrence after ablation in patients with concurrent AF and pulmonary diseases (PDs) remained obscure. Herein, we aimed to investigate their predicting values and differences in patients with concurrent AF and PDs. METHODS: From January 2008 to April 2015, 304 patients with concurrent AF and PDs treated with catheter ablation were divided into 2 groups according to whether they experienced AF recurrence in our centers. Factors related with AF recurrence were explored using Cox regression and scores predicting recurrent AF were compared in these patients using ROC curves. RESULTS: During a median of 6-month of follow-up, factors correlating with late AF recurrence included heart failure (HF) history [hazard ratio (HR): 2.79; 95% confidence interval (CI): 1.49-5.22, P=0.001], current smoking (1.73; 1.13-2.68, P=0.01) and early AF recurrence (3.85; 95% CI: 2.62-5.66, P<0.001) according to univariate Cox regression analysis. When analyzed using multivariate Cox model, HF history (2.21; 1.12-4.37, P=0.02), hypertension history (1.54; 1.02-2.33, P=0.04) and early AF recurrence (3.90; 2.60-5.85, P<0.001) were related to late AF recurrence. The BASE-AF2 score had higher c-index than the MB-LATER, APPLE, CHADS2, CHA2DS2-VASc, CAAP-AF and HATCH scores when compared using ROC curves analysis (all P<0.05). The optimal point for predicting AF recurrence of the BASE-AF2 score in the ROC analysis was 1 point with sensitivity of 69.03% and specificity of 60.21%. CONCLUSIONS: The predicting AF recurrence value of BASE-AF2 score was superior to MB-LATER, APPLE, CHADS2, CHA2DS2-VASc, CAAP-AF and HATCH scores in patients with concurrent AF and PDs, which can be an effective and helpful score for making AF treatment decisions.

Mutual regulation of ROS accumulation and cell autophagy in wheat roots under hypoxia stress.

Here, we explored the mutual regulation of radical oxygen species (ROS) and autophagy in wheat (Triticum aestivum L.) roots under hypoxia stress. We also analyzed differences between the responses of the stele and the cortex in the two wheat cultivars Huamai 8 (waterlogging-tolerant) and Huamai 9 (waterlogging-sensitive) to hypoxia stress. In situ detection and ultracytochemical localization analysis in wheat roots showed that hypoxia stress caused greater increases in ROS levels and the expression levels of alternative oxidase (AOX) and antioxidant enzymes in the stele than in the cortex. The analysis of exogenous ROS addition and the inhibition of its production revealed the pivotal roles played by ROS in autophagy. Moreover, transmission electron microscopy and qRT-PCR analysis indicated that the stele had a higher level of autophagy than the cortex and that the two wheat cultivars primarily differed in the type and number of autophagosomes. Additional research revealed that autophagy could remove excess ROS, as pre-treatment with the autophagy inhibitor 3-methyladenine increased ROS levels in roots and the addition of the autophagy inducer rapamycin reduced root ROS levels. In conclusion, hypoxia stress induced ROS accumulation in wheat roots where ROS acted as an autophagy signal. Furthermore, higher levels of autophagy and antioxidant enzyme expression in the stele facilitated the elimination of oxidative damage caused by excessive ROS and thereby increased cell survival; in the cortex, a large number of cells died and formed aerenchyma.

The use of cIMT as a predictor of postoperative stroke in patients undergoing surgical repair of acute type a aortic dissection.

BACKGROUND: Acute type A aortic dissection (ATAAD) is a life-threatening condition that requires surgical intervention. Stroke remains an extremely serious adverse outcome that can occur in ATAAD patients undergoing aortic arch repair, leading to higher rates of patient mortality and decreased postoperative quality of life. In the present study, we sought to determine whether carotid intima-media thickness (cIMT) is a reliable predictor of postoperative stroke risk. MATERIALS AND METHODS: This was a prospective study of 76 patients with ATAAD undergoing aortic arch repair. For all patients, cIMT was determined preoperatively through a Doppler-based method. Incidence of different forms of neurological dysfunction, including temporary neurological dysfunction (TND) and stroke, was monitored in these patients, and the relationship between cIMT and stroke incidence was assessed using a receiver-operating characteristic (ROC) curve. Prognostic variables associated with stroke risk were further identified through univariate and multivariate analyses. RESULTS: A total of 26/76 (34.2%) patients in the present study suffered from neurological dysfunction, of whom 16 (21.0%) suffered from TND and 10 (13.2%) suffered a stroke. The remaining 50 patients (65.8%) did not suffer from neurological dysfunction. The cIMT values in the stroke, TND, and neurological dysfunction-free patients in this study were 1.12 ± 0.19 (mm), 0.99 ± 0.13 (mm), and 0.87 ± 0.13 (mm), respectively. A total of 4 patients in this cohort died during the study, including 1 in the TND group and 3 in the stroke group. An ROC curve analysis indicated that cIMT could predict stroke with an area under the curve value of 0.844 (95% CI, 0.719-0.969; p < 0.001). A multivariate analysis revealed that cIMT > 0.9 mm was independently associated with stroke risk (p = 0.018). CONCLUSION: We found that cIMT can be used to predict postoperative stroke risk in ATAAD patients undergoing aortic arch repair, with a cIMT > 0.9 mm coinciding with increased stroke risk in these patients. TRIAL REGISTRATION: ChiCTR1900022289. Date of registration 4 April 2019 retrospectively registered.

Clinical anatomic study of the lower lumbar anterolateral vein: with respect to retroperitoneal endoscopic surgery.

OBJECTIVE: To study the anatomy of veins of the lower lumbar spine and provide the anatomic basement for laparoscopic lumbar surgery. METHODS: A total of 15 formaldehyde-preserved cadavers were studied with special attention to the variety and surrounding structure of ascending lumbar vein (ALV) and iliolumbar veins (ILV), and their relationship with lumbar plexus. RESULTS: ALV and ILV can be found on every sides, which have four variants including separate entry and common entry. The ascending vein and iliolumbar vein separately enter common iliac vein in 18 cases, and as a common stem enter the common iliac vein in 12 cases. Retracting common iliac vein medially both the ascending lumbar and the iliolumbar veins are always at risk of avulsion on exposure of the disc space. The injury of obturator nerve and lumbosacral trunk of lumbar plexus should be avoided. CONCLUSION: Awareness of these anatomic variation can prevent the hemorrhage and be helpful for the surgeon in performing a careful ligation of these veins before medial retraction of the common iliac vein. Our findings emphasize the need for proper dissection of ALV and ILV before ligature during exposure of the lower lumbar spine.

[The clinic anatomy of lumbar plexus in the lumbar anterolaterally approach minimally invasive surgery].

OBJECTIVE: To provide anatomic data for reducing lumbar plexus nerve injury. METHODS: The applied anatomy of lumbar plexus was studied by 15 formaldehyde-preserved cadavers, two groups of sectional images of lumbar segment and three series of virtual chinese human dataset. RESULTS: Arrangement of the lumbar nerve was regular. From anterior view, lumbar plexus nerve arranged from lateral to medial from L2 to L5; from lateral view, lumbar nerve arrange from ventral to dorsal from L2 to L5. The angle degree between the lumbar nerve and lumbar increased from L1 to L5. The lumbar plexus nerve was revealed to be in close contact with transverse process. By sectional anatomy, all parts of the lumbar plexus nerve were located in the dorsal third of the psoas major. The safety zone of the psoas major to prevent nerve injuries was ventrally 2/3. CONCLUSIONS: Psoas major can be considered as surgery landmark when expose the lateral anterior of lumbar by incising the psoas muscle. Incising the psoas muscle ventral 2/3 can prevent lumbar plexus injury. Transverse process can be considered as landmark for the position of lumbar plexus in operation.

Differences of Matrix Metalloproteinase 2 Expression between Left and Right Ventricles in Response to Nandrolone Decanoate and/or Swimming Training in Mice.

BACKGROUND: Matrix metalloproteinase (MMP)-2 plays an important role in the remodeling of left ventricles (LVs) and right ventricles (RVs). We investigated the differences of MMP-2 expression between LV and RV in response to nandrolone decanoate (ND), swimming training (ST), and combined ND and ST (NS) in mice, based on their structural, functional, and biochemical characteristics. METHODS: Totally 28 male C57B1 mice (6 weeks old; 20-23 g) were divided into four groups, including the control (n = 7), ND (n = 6), ST (n = 8), and NS (n = 7) groups. After respective treatments for 8 weeks, echocardiographic examination was used to assess the cardiac structure and function. Van Gieson stain was used to examine the fibrosis of LV and RV in response to different treatments, and Western blotting analysis was performed to explore different MMP-2 expressions between LV and RV in response to ND and/or ST. Analysis of variance was used for comparing the four groups. RESULTS: At 8 weeks, right ventricular dimension/body weight in the ND group was larger than the other three groups (F = 7.12, P < 0.05) according to the echocardiographic examination. Fibrosis induced by ND administration was increased more in RV (2.59%) than that in LV (2.21%). MMP-2 expression of the ND group in RV was significantly greater than the control and NS groups in RV and the corresponding ND group in LV. CONCLUSION: The experimental data support the hypothesis that ND administration induces greater MMP-2 expression increase in RV compared to LV, leading to consequent RV dilation.

[Physiological and biochemical mechanism of spermidine improving drought resistance in maize seedlings under drought stress.] / 干旱胁迫下亚精胺对玉米幼苗抗旱性影响的生理生化机制.

To explore the role of exogenous spermidine (Spd) in enhancing the resistance of maize to drought stress, 15% polyethylene glycol (PEG-6000) was used to simulate drought stress and with 'Xianyu 335' (drought-insensitive) and 'Fenghe 1' (drought sensitive) as the experiment materials, the effects of Spd (0.1 mmol·L-1) on the growth, photosynthetic characteristics, chlorophyll content, osmotic adjustment substance, membrane lipid peroxidation and root activity of maize seedlings were investigated. The results showed that the application of Spd significantly promoted the growth of maize seedlings under drought stress, increased chlorophyll content, net photosynthetic rate (Pn), stomatal conductance (gs), transpiration rate (Tr) and water use efficiency (WUE), and decreased the enhancement of intercellular carbon dioxide concentration (Ci) in 'Fenghe 1'. Moreover, the stomatal and non-stomatal limitations of photosynthetic ability caused by drought stress in 'Fenghe 1' were effectively reduced by exogenous Spd. The application of Spd increased the content of proline and soluble sugar, decreased theO2-· generation rate, contents of H2O2 and MDA and cell membrane permeability, enhanced the root activity. The changes of drought-sensitive 'Fenghe 1' were greater than drought-tolerant 'Xianyu 335'. These results indicated that exogenous Spd had positive effects on the seedlings to capture and converse solar energy, thus promoting photosynthesis and the growth of maize seedlings. It would also enhance the adaptability of seedlings to drought stress by reducing the production of reactive oxygen species (ROS), increasing the accumulation of osmotic adjustment substances to stabilize the cell membrane system and improving the root vigor. The positive effects of Spd was more obvious for drought-sensitive variety 'Fenghe 1'.

Biomechanical effect of vertebroplasty on the adjacent intervertebral levels using a three-dimensional finite element analysis.

OBJECTIVE: To investigate the biomechanical effect of different volume, distribution and leakage to adjacent disc of bone cement on the adjacent vertebral body by three-dimensional osteoporosis finite element model of lumbar. METHODS: L(4)-L(5) motion segment data of the cadaver of an old man who had no abnormal findings on roentgenograms were obtained from computed tomography (CT) scans. Three-dimensional model of L(4)-L(5) was established with Mimics software, and finite element model of L(4)-L(5) functional spinal unit (FSU) was established by Ansys 7.0 software. The effect of different loading conditions and distribution of bone cement after vertebroplasty on the adjacent vertebral body was investigated. RESULTS: This study presented a validated finite element model of L(4)-L(5) FSU with a simulated vertebroplasty augmentation to predict stresses and strains of adjacent untreated vertebral bodies. The findings from this FSU study suggested the endplate and disc stress of the adjacent vertebral body was not influenced by filling volume of bone cement but unipedicle injection and leakage to the disc of bone cement could concentrate the stress of adjacent endplate. CONCLUSIONS: Asymmetric distributions and leakage of cement into intervertebral disc can improve the stress of endplate in adjacent vertebral body. These results suggest that optimal biomechanical configuration should have symmetric placement and avoid leakage of cement in operation.

[Case-control study on the effect of core strength training on the function of anterior cruciate ligament reconstruction].

OBJECTIVE: To observe the effect of core strength training on knee joint function and postural stability after anterior cruciate ligament reconstruction (ACLR). METHODS: A total of 80 ACLR patients were randomly allocated into conventional rehabilitation training group and core strength training group from May 2013 to May 2015 with 40 patients in each group. The patients in conventional rehabilitation training group underwent conventional ACLR rehabilitation training, in which 28 males and 12 females. The mean age was(30.5±5.2) years old(ranged, 22 to 42 years old). The mean BMI was(23.8±2.4) kg/m²(ranged, 18.2 to 25.9 kg/m²). Thirty patients had injuries on the dominant side and 10 patients had injuries on the non-dominant side. The core strength training group received conventional ACLR rehabilitation training and core strength training, in which 31 males and 9 females. The mean age was(31.1±4.8) years old(ranged, 21 to 45 years old). The mean BMI was(24.1±2.7) kg/m²(ranged, 18.5 to 26.1 kg/m²) . Twenty-seven patients had injuries on the dominant side and 13 patients had injuries on the non-dominant side. The Lysholm score, tibial anterior transition measured by KT-1000 before and after treatment, and the Star Excursion Balance Test results after treatment were compared between the two groups. RESULTS: Six months after rehabilitation training, the tibial anterior transition of the conventional rehabilitation training group and the core strength training group were(3.4±1.0) mm and(3.3±1.2) mm respectively, which were less than(12.1±1.8) mm and(12.5±2.0) mm before treatment. But there was no significant difference in anterior tibial translation between two groups(P>0.05). The Lysholm score of the conventional rehabilitation training group and the core strength training group were 91.8±4.3 and 92.1±3.9 individually, which were higher than 69.2±5.8 and 70.2±5.1 before treatment. But there was no significant difference in Lysholm score between two groups(P>0.05). Six months after rehabilitation training, the results of Star Excursion Balance Test showed the reach distance with the support in the injured side and healthy side in the core strength training group were greater than that of the conventional rehabilitation training group in the eight directions(P<0.05). CONCLUSIONS: The core strength training could improve the dynamic balance of ACLR patients.

Construction of the POT1 promoter report gene vector, and the effect and underlying mechanism of the POT1 promoter in regulating telomerase and telomere length.

By using human genomic DNA as a template to clone protection of telomere 1 (POT1) promoter gene segments and construct the POT1 promoter luciferase report gene vector (pGL3-Control-POT1-promoter), the association between POT1, and the regulation of telomerase and telomere length was investigated. In the present study, two recombinant luciferase report gene vectors were constructed, which included different regions of the POT1 promoter. The plasmids were transformed into DH5α and the positive clones were obtained. The two plasmids termed as pGL3-Control-POT1-promoter-1 and pGL3-Control-POT1-promoter-2, were confirmed using restriction enzyme analysis and sequencing. They were separately and transiently transfected into four types of human tumor cells (A549, H460, HepG2 and HeLa). The transcriptional activities of the POT1 promoter were verified using the dual-luciferase assay. The relative expression of POT1 and human telomerase reverse transcriptase (hTERT), and telomere length were analyzed using quantitative polymerase chain reaction in the four types of non-transfected tumor cells. Using SPSS software, correlations between POT1 promoter activity, and POT1 expression, hTERT expression and telomere length were analyzed. Two POT1 promoter fragments (POT1-promoter-1 and -2) were successfully constructed into the pGL3-Control luciferase report gene vector. POT1-promoter-1 exhibited significantly stronger transcription activity compared with POT1-promoter-2. The results of the partial correlation and linear regression analyses were similar: POT1 promoter activity was identified to be significantly and positively correlated with POT1 expression and telomere length (partial correlation coefficients, both P<0.05; linear regression, both P<0.01). However, POT1 promoter activity and hTERT expression were significantly negatively correlated (both P<0.05). The results obtained in the present study suggest that the POT1 promoter influences telomere length. Furthermore, these data indicated that POT1 promoter activity and POT1, as well as telomere length, may be a useful biomarker for tumor detection and future patient prognosis.

MiR-551b-5p Contributes to Pathogenesis of Vein Graft Failure via Upregulating Early Growth Response-1 Expression.

BACKGROUND: Vein graft failure (VGF) is a serious complication of coronary artery bypass graft, although the mechanism remains unclear. The study aimed to investigate the effects of microRNAs (miRNAs) on the endothelial dysfunction involved in VGF. METHODS: Human umbilical vein endothelial cells (HUVECs) were subjected to mechanical stretch stimulation to induce endothelial dysfunction. Genome-wide transcriptome profiling was performed using the Human miRNA OneArray® V4 (PhalanxBio Inc., San Diego, USA). The miRNA-messenger RNA (mRNA) network was investigated using gene ontology and Kyoto Encyclopedia of Genes and Genomes. The miR-551b-5p mimic and inhibitor were applied to regulate miR-551b-5p expression in the HUVECs. The 5-ethynyl-2'-deoxyuridine assay, polymerase chain reaction (PCR), and Western blotting (WB) were used to assess HUVECs proliferation, mRNA expression, and protein expression, respectively. The vein graft model was established in early growth response (Egr)-1 knockout (KO) mice and wide-type (WT) C57BL/6J mice for pathological and immunohistochemical analysis. Endothelial cells isolated from the veins of WT and Egr-1 KO mice were subjected to mechanical stretch stimulation; PCR and WB were conducted to confirm the regulatory effect of Egr-1 on Intercellular adhesion molecule (Icam-1). One-way analysis of variance and independent t-test were performed for data analysis. RESULTS: Thirty-eight miRNAs were differentially expressed in HUVECs after mechanical stretch stimulation. The bioinformatics analysis revealed that Egr-1 might be involved in VGF and was a potential target gene of miR-551b-5p. The mechanical stretch stimulation increased miR-551b-5p expression by 2.93 ± 0.08 fold (t = 3.07, P < 0.05), compared with the normal HUVECs. Transfection with the miR-551b-5p mimic or inhibitor increased expression of miR-551b-5p by 793.1 ± 171.6 fold (t = 13.84, P < 0.001) or decreased by 26.3% ± 2.4% (t = 26.39, P < 0.05) in the HUVECs, respectively. HUVECs proliferation and EGR-1 mRNA expression were significantly suppressed by inhibiting miR-551b-5p expression (P < 0.05). The lumens of the vein grafts in the Egr-1 KO mice were wider than that in the WT mice. Icam-1 expression was suppressed significantly in the Egr-1 KO vein grafts (P < 0.05). CONCLUSIONS: Increased miR-551b-5p expression leads to endothelial dysfunction by upregulating Egr-1 expression. EGR-1 KO can improve the function of a grafted vein through suppressing Icam-1.

Intron-specific shRNA-mediated downregulation of survivin and promotion of apoptosis in HeLa cells.

Overexpression of the survivin gene contributes to tumorigenesis; it has been recognized as an important target for cancer therapy. In the present study, survivin expression was suppressed using recombinant plasmid mediated short hairpin RNAs (shRNAs) that were constructed to target exonic or intronic sequences of the survivin gene. In addition, a negative control shRNA was constructed. HeLa cells were transfected with specific shRNA constructs and the blocking efficiency of each shRNA was assessed at the mRNA and protein levels; and the five shRNA constructs with higher blocking efficiency were selected. Cell apoptosis was assessed by flow cytometry (FCM) following Annexin V-fluorescein isothiocyanate/propidium iodide double staining. Hoechst staining was used to detect the morphological diversity of the nuclei in apoptotic cells. The results demonstrated that survivin expression was effectively reduced by the transfection of shRNAs in HeLa cells. In addition, the apoptotic rates of the shRNA-treated groups were significantly increased compared with the negative control group according to the FCM results. The nuclei of HeLa cells exhibited apoptotic characteristics in the shRNA-treated groups as identified by Hoechst staining. Survivin-targeting shRNAs effectively downregulated the expression of the gene and markedly increased the apoptotic rate of HeLa cells. Data from the present study also indicated that the intron-specific shRNA demonstrate a high efficiency of inhibition of survivin expression and were able to induce cell apoptosis of HeLa cells through RNAi, potentially providing novel target sites for tumor therapy. In conclusion, the present study suggests that intron-specific blocking of survivin by RNAi may provide a tool for anticancer therapy.

Construction and characterization of a transmembrane eukaryotic expression vector based on the membrane domain structure of TNF-α.

The aim of the present study was to construct a fast-acting, eukaryotic expression vector in eukaryotic cells based on transmembrane-tumor necrosis factor­α (TM­TNF­α) structure. Two types of recombinant eukaryotic expression vectors were constructed, pcDNA3.1-TM-enterokinase-TNF­α and pcDNA3.1­TM­Factor Xa­TNF­α, according to the TNF­α transmembrane segments. Following the generation of these vectors, mouse embryonic 3T3 fibroblasts were transfected and reverse transcription­polymerase chain reaction and western blotting analyses were used to analyze mTNF­α mRNA and protein expression levels, respectively, in total cellular protein extracts and extracellular fluid. The biological activity of TNF-α in the extracellular fluid was then measured using an MTT assay. The vectors were successfully constructed, and mRNA and fusion proteins were detected in the 3T3 cells. Among the fusion proteins, the one observed in pcDNA3.1-TM-FactorXa-TNF-α-transfected 3T3 cells remained as a transmembrane protein. In addition, treatment of L929 cells with TNF­α derived extracellular fluid samples from pcDNA3.1­TM­FactorXa­TNF­α­transfected 3T3 cells was associated with a dose­dependent reduction in in cell­specific activity. The results indicate that proteins expressed using pcDNA3.1­TM­FactorXa­TNF­α vectors form transmembrane proteins. In addition, the results indicate that, only when coupled with FactorXa activity, the extracellular region of TM­TNF­α forms s­TNF­α, and the controlled expression of the fusion protein is initiated.