ECT2 promotes proliferation and metastasis of esophageal squamous cell carcinoma via the RhoA-ERK signaling pathway.

OBJECTIVE: In this study, the effect of epithelial cell transformation sequence 2 (ECT2) on the proliferation, invasion and migration of esophageal squamous cell carcinoma (ESCC) was investigated by interfering the expression of ECT2. PATIENTS AND METHODS: Interfering with the expression level of ECT2 in human squamous cell carcinomas KYSE140 and EC9706 cell lines, the changes of KYSE140 and EC9706 cell proliferation, invasion, and migration were measured using the CCK-8 method, transwell test, and scratch test, respectively. The effects of ECT2 on the Ras homolog gene family, member A-extracellular regulated protein kinases (RhoA-ERK) signaling pathway were also observed. RESULTS: Compared with the control group, the proliferation, migration, and invasion ability of EC9706 and KYSE140 cells after ECT2 knockout were significantly reduced (p <0.05). The knockdown of ECT2 expression in ESCC cell lines suppressed the activation of RhoA-ERK signaling pathway and protein expression of VEGF and MMP9. CONCLUSIONS: ECT2 could regulated the expression of VEGF and MMP9 to inhibit cells proliferation, invasion, migration and tumor development through RhoA-ERK signaling pathway. Therefore, ECT2 could be an available marker, and provide a new theoretical basis for the treatment of ESCC.

[Predictors of fatigue among individuals with primary immune thrombocytopenia in China].

Objective: To study the fatigue symptoms of adult patients with primary immune thrombocytopenia (ITP) and to analyze the possible factors that affect the severity of fatigue. Methods: Eligible adult patients with ITP who admitted to Institute of Hematology & Blood Diseases Hospital were enrolled in this study and the questionnaires including a Chinese version of the Functional Assessment of Chronic Illness Therapy-Fatigue (FACIT-Fatigue) , the Pittsburgh sleep quality index (PSQI) , the Hospital Anxiety and Depression scale (HADS) and demographic information were completed. The predictors of fatigue were determined with multiple linear regression analyses. Results: A total of 207 patients with ITP were enrolled, including 70 males (33.8%) and 137 females (66.2%) , the median age was 42 (18-72) years old. The FACIT-F score in ITP patients was (37.50±9.05) . The FACIT-F severity of ITP patients was positively correlated with the platelet count (r=0.307, P<0.001) . The FACIT-F severity was negatively correlated with bleeding severity (r=-0.276, P<0.001) , sleep quality (r=-0.654, P<0.001) , depression (r=-0.598, P<0.001) and anxiety (r=-0.616, P<0.001) . A multiple linear regression analysis revealed that the severity of ITP-related fatigue was significantly correlated with platelet count (P<0.001) , bleeding severity (P=0.004) , sleep quality (P<0.001) and depression (P<0.001) . Conclusion: Fatigue was determined by complicated factors in adult ITP patients. Interventions addressing depressive symptoms, sleep quality, bleeding symptoms and platelet count could be potential avenues for treatment of fatigue in patients with ITP.

[Retrospectively analysis of the difference of bleeding frequency and hemophilic arthropathy between hemophilia A and hemophilia B patients].

Objective: To analyze the difference of bleeding frequency, plain radiographic (X-ray) , risk factors in hemophilic arthropathy progression and the Arnold-Hilgartner classification. Methods: A retrospective study was conducted in 211 hemophilia patients hospitalized in our medical center between January 2007 and December 2010, some patients with hemarthrosis were followed up for 5 years. Results: All patients were male, including 150 hemophilia A (HA) and 61 hemophilia B (HB) . The HA patients bled more frequently than HB patients with annualized total bleeding rate 20.5 (0-48) vs 13 (1-40) ; annualized joint bleeding rate 13.5 (0-38) vs 8 (0-33) , especially in moderate hemophilia [26 (1-48) vs 12 (1-36) , P<0.001; 18 (0-36) vs 7.5 (0-26) , P=0.001], but severe hemophilia had no difference in bleeding frequency [33 (1-41) vs 26 (1-40) , P=0.702; 22 (0-36) vs 18 (0-33) , P=0.429]. The condition of the affected joints of 108 HA and 54 HB was evaluated on roentgenography. In HA patients, the Arnold-Hilgartner classification increased with the severity ratings (r=0.063, P=0.004) . However, similar associations were not found in HB patients (r=0.045, P=0.082) . Five years later, 36 HA and 19 HB patients received the same joint X-ray, there were no significant differences in joints radiographic progression between the total HA and HB groups (z=1.941, P=0.052) . However, significant difference between moderate HA and HB was observed (z=0.076, P=0.002) . Multivariate unconditioned Logistic analysis showed that annualized joint bleeding rate [P<0.001, OR=1.166 (95%CI 1.097-1.239) ] and articular structural injuries [P=0.018, OR=2.842 (95% CI 1.196-6.755) ] were independent risk factors for the joints radiographic progression. Conclusion: The study suggests that there was a difference in bleeding phenotype between HA and HB, especially in moderate hemophilia. HB patients showed mild but progressive development over time, compared with HA patients. Annualized joint bleeding rate and articular structural injuries were independent risk factors for the joints radiographic progression.

Expression and localization of Yap and Taz during development of the mandibular first molar in rats.

Yes-associated protein (Yap) and transcriptional coactivator with PDZ-binding motif (Taz) are two downstream factors in the Hippo signaling pathway. Yap and Taz participate in regulating organ size, stem cell self-renewal, proliferation and differentiation. We investigated the spatial-temporal expression and relative expression levels of Yap and Taz using immunohistochemistry and real-time polymerase chain reaction. We found Yap and Taz in the oral epithelium and mesenchyme at embryonic (E) day 14.5 (E14.5) and E16.5. By E18.5, Yap and Taz were detected in the dental papilla and the entire enamel organ. At postnatal (P) day 0 (PN0), PN3 and PN7, Yap and Taz expression was localized in ameloblasts, odontoblasts and stratum intermedium. Yap and Taz were expressed in Hertwig's epithelial root sheath (HERS) at PN7. At PN3, PN7 and PN14, Yap was detected in the enamel matrix. From PN21 to PN28, Yap and Taz were absent from differentiated ameloblasts, but they were expressed in odontoblasts. From PN0 to PN10, the Yap and Taz mRNA expression increased, then decreased. We found that Yap and Taz may influence the differentiation of ameloblasts and odontoblasts; they also may contribute to enamel mineralization, crown morphogenesis and root formation.

Online monitoring of oil film using electrical capacitance tomography and level set method.

In the application of oil-air lubrication system, electrical capacitance tomography (ECT) provides a promising way for monitoring oil film in the pipelines by reconstructing cross sectional oil distributions in real time. While in the case of small diameter pipe and thin oil film, the thickness of the oil film is hard to be observed visually since the interface of oil and air is not obvious in the reconstructed images. And the existence of artifacts in the reconstructions has seriously influenced the effectiveness of image segmentation techniques such as level set method. Besides, level set method is also unavailable for online monitoring due to its low computation speed. To address these problems, a modified level set method is developed: a distance regularized level set evolution formulation is extended to image two-phase flow online using an ECT system, a narrowband image filter is defined to eliminate the influence of artifacts, and considering the continuity of the oil distribution variation, the detected oil-air interface of a former image can be used as the initial contour for the detection of the subsequent frame; thus, the propagation from the initial contour to the boundary can be greatly accelerated, making it possible for real time tracking. To testify the feasibility of the proposed method, an oil-air lubrication facility with 4 mm inner diameter pipe is measured in normal operation using an 8-electrode ECT system. Both simulation and experiment results indicate that the modified level set method is capable of visualizing the oil-air interface accurately online.

[The regulation of NO on sleep-wakefulness in rats].

AIM AND METHODS: The effects of an inhibitor of nitric oxide synthase L-NAME and a precursor of NO(L-arginine) on sleep-wakefulness in rats were examined. RESULTS: 1. The time of waking was increased(P < 0.01) and the time of SWS was decreased(P < 0.01) during the first two hours after injection intracerebroventricularly(i.c.v.) of L-NAME 1 mg, this effect was showed across the 4 h recording period(P < 0.05). The i.c.v. injection of L-NAME 0.2 mg had no significant effect. 2. The time of waking was su-ppressed during the third and fourth hours after i.c.v. injection of L-arginine 300 micrograms(P < 0.05). At the same time the increase of SWS was observed (P < 0.01). 3. There was no change on W, SWS, PS when L-NAME 1 mg was given after administration of L-arginine. CONCLUSION: The excitatory effect of L-NAME on wakefulness is mediated by inhibition of NOS. This effect could be antagonized by L-arginine. It suggests that NO is involved in regulation of sleep-wakefulness.

Structure, interaction and electron transfer between cytochrome b5, its E44A and/or E56A mutants and cytochrome c.

Site-directed mutagenesis has been used to produce variants of a tryptic fragment of bovine liver cytochrome b5 in which Glu44 and Glu56 are mutated to alanine. The reduction potentials measured by spectroelectrochemical titration (in the presence of 1 mM (Ru(NH3)6)3+, pH 7.0 and I=0.1 M) are 4.5, 6.0, 6.0 and 7.5 mV versus the standard hydrogen electrode (SHE) for the wild-type and E44A, E56A and E44/56A mutants of cytochrome b5, respectively. A comparative two-dimensional NMR study of cytochrome b5 and its E44/56A mutant in water solution has been achieved. Resonance assignments of side-chains have been completed successfully. The NMR results suggest that the secondary structures and global folding of the E44/56A mutant remain unchanged, but the mutation of both Glu44 and Glu56 to hydrophobic alanine may lead to the two helices containing mutated residues contracting towards the heme center. The inner mobility of the Gly42 approximately Glu44 segment in cytochrome b5 may be responsible for the difference of the binding mode between Glu44 and Glu56 with cytochrome c. The binding between cytochrome c and cytochrome b5 was studied by optical difference spectra of cytochrome c and variants of cytochrome b5. The association constants (KA) for the wild-type, E44A, E56A, and E44/56A mutants of cytochrome b5 with cytochrome c, are 4.70(+/-0. 10)x10(6) M-1, 1.88(+/-0.03)x10(6) M-1, 2.70(+/-0.13)x10(6) M-1, and 1.14(+/-0.05)x10(6) M-1, respectively. This is indicative that both Glu44 and Glu56 are involved in the complex formation between cytochrome b5 and cytochrome c. The reduction of horse heart ferricytochrome c by recombinant ferrocytochrome b5 and its mutants has been studied. The rate constant of the electron transfer reaction between ferricytochrome c and wild-type ferrocytochrome b5 (1.074(+/-0.49)x10(7) M-1 s-1) is higher than those of the mutant protein E44A (8.98(+/-0.20)x10(6) M-1 s-1), E56A (8.76(+/-0. 39)x10(6) M-1 s-1), and E44/56A (8.02(+/-0.38)x10(6) M-1 s-1) at 15 degreesC, pH 7.0, I=0.35 M. The rate constants are strongly dependent on ionic strength and temperature. These studies, by means of a series of techniques, provide conclusive results that the interaction between cytochrome b5 and cytochrome c is electrostatically guided, and, more importantly, that both Glu44 and Glu56 participate in the electron transfer reaction.

[Changes of angiotensin II autocrine by cultured pulmonary artery smooth muscle cells exposed to anoxia].

Alterations of autocrine function of pulmonary arterial smooth muscle cells (PASMCs) might play an important role in development of hypoxic pulmonary hypertension (HPH). To test this hypothesis, the effects of hypoxia on angiotensin II (AT II) secretion by cultured new born bovine PASMCs were investigated. AT II secretion decreased significantly when PASMCs were incubated under 2.5% O2 hypoxic condition for 3 to 48 h (P < 0.01 vs control), but decreased further under anoxic condition (P < 0.01 vs control and 2.5% O2 group). Nitric oxide (NO) donor SIN-1 inhibited AT II secretion significantly under normoxic condition, but NO synthase inhibitor L-nitro-arginine (LNA) eliminated the inhibitory effect of anoxia on AT II autocrine and promoted AT II release. It was also found that the concentration of cGMP in PASMCs increased significantly (P < 0.01) at 24 h incubation in 0% O2, an effect that can be attenuated by LNA. Hypoxia stimulated 3H-TdR incorporation of PASMCs significantly (P < 0.001), while captopril exerts an inhibition in normoxic condition (P < 0.001), but without effect under hypoxic condition. The above results suggest that hypoxia can inhibit AT II secretion by inducing endogenous NO production in PASMCs. AT II autocrine is not involved in hypoxic proliferation of PASMCs, but a decrease of AT II may contribute to prevention of the development of HPH.

[Modulations of proliferation between in vitro pulmonary artery endothelial cells and smooth muscle cells under hypoxia].

Vascular endothelial cells (VECs) and vascular smooth muscle cells (VSMCs) are structurally and functionally closely connected. Their interactions may play important roles in the development of hypoxic pulmonary artery hypertension. In the present study, mitogenic regulations between cultured new bovine pulmonary arterial endothelial cells (PAECs) and pulmonary arterial smooth muscle cells (PASMCs) under both normoxic and hypoxic conditions were investigated. Cell proliferation was assessed by 3H-thymidine incorporation and flow cytometry. Normoxic conditioned medium (CM) from cultured PASMCs inhibited 3H-thymidine incorporation of PAECs by 58% (P < 0.01), but normoxic or hypoxic CM from PAECs promoted or inhibited 3H-thymidine incorporation of PASM by 60% and 27% respectively (P < 0.01). When mixed PAECs and PASMCs were cultured for 24 h, relative 3H-thymidine incorporation of the mixed cells decreased by 22% (P < 0.01 vs monoculture) under normoxic condition, and increased by 75% or 44% under 0% O2 or 2.5% O2 (P < 0.01 vs normoxic mixculture). When PAECs and PASMCs were cocultured in either normoxic or hypoxic conditions, proliferation of PAECs was inhibited while that of PASMs was stimulated significantly (P < 0.01), as compared with that of homotypic cultured cells. These findings suggest that PAECs and PASMCs may mutually regulate their proliferation each other, and this mutual modulation may be changed under hypoxic condition.

[Effects of serotonin on the proliferation of pulmonary arterial smooth muscle cell under hypoxia].

The effects of hypoxia on the bovine pulmonary arterial smooth muscle cell (PASM) proliferation and serotonin transporter gene expression were investigated using 3H-Thymidine incorporation, immunocytochemical staining and mRNA dot blot hybridyzation, the results were as follows, PASM 3H-Thymidine incorporation increased in response to anoxia for 24 h (P < 0.05). The anoxia-induced increased in DNA synthesis of PASM was significantly stimulated by 5-HT (100, 10 and 1 mumol/L) (P < 0.001), but 5-HT did not increase 3H-TdR incorporation under normoxic condition. Immunocytochemical reaction of serotonin in PASM under normoxic condition was markedly stronger than that of anoxic PASM (P < 0.001). Dot blot analysis showed hypoxia increased serotonin transporter mRNA expression of PASM (P < 0.01). These results indicate that stimulation of PASM proliferation with enhanced serotonin transporter gene expression and high consumption of 5-HT in hypoxia or/and anoxia may take part in the development of hypoxic pulmonany hypertention.

Recurrence rate and risk factors for urinary calculi after extracorporeal shock wave lithotripsy.

PURPOSE: We determined the recurrence rate after extracorporeal shock wave lithotripsy (ESWL*) and evaluated its associated risk factors, such as patient sex and age, volume of daily water intake, related diseases, family history of urinary calculi, number, burden and location of stones, residual stones after ESWL, serum uric acid level before ESWL and so forth. MATERIALS AND METHODS: We conducted a retrospective study of 841 patients who underwent ESWL for primary treatment of urinary calculi between 1985 and 1987. Charts were reviewed for stone characteristics, outcomes and associated laboratory data. Followup 6 to 9 years later was done by mail and telephone interviews. RESULTS: Followup data were available for 436 patients (52%). Overall, stones recurred in 51% of the 436 patients at a mean followup of 7.1 years. Late recurrences were common, and were noted in as many as 70% of the patients after 9 years. This rate is greater than what has been previously reported after other treatments. Cox regression analysis indicated that the major risk factors associated with recurrence were male gender (relative risk 1.406, p < 0.05) and multiple stones (relative risk 1.777, p < 0.05). CONCLUSIONS: The frequent late recurrences support the speculation that ESWL might lead to greater recurrence rates. Therefore, the cost-effectiveness of ESWL versus other modalities for stone removal warrants more long-term evaluation.

The study of serum apoprotein levels as indicators for the severity of angiographically assessed coronary artery disease.

Serum apoprotein A-I (Apo A-I) and B (Apo-B) concentrations were determined in 40 subjects undergoing coronary angiography for past myocardial infarction and angina pectoris, and the authors studied the relationship between the apoprotein concentrations and the severity of coronary artery disease (CAD). During this study, serum total cholesterol, triglyceride, and high-density lipoprotein, low-density lipoprotein, and very low density lipoprotein cholesterol concentrations were determined to control analysis. The results showed that the decrease in serum Apo A-I levels was the best indicator distinguishing CAD from non-coronary artery disease; the Apo B/Apo A-I ratio had the most consistent association with the severity of CAD as assessed by angiography; Apo B/Apo A-I values ranging from 0.98 to 1.00 might be considered critical values for early CAD.

Effects of hypoxia and complement activation on lung hemodynamics and fluid exchanges in unanesthetized goats.

The effects of hypoxia and zymosan-activated plasma (ZAP) on lung hemodynamics and fluid exchange were studied in unanesthetized goats. Hypoxia produced a sustained increase in pulmonary arterial pressure (PPA), but had no effect on lung lymph flow and protein content. Following ZAP infusion there was an early phase of leukopenia and marked pulmonary hypertension followed by a phase characterized by a modest increase in the flow of protein-rich lung lymph. When ZAP was infused at a simulated altitude of 4,000 m, there was a further increase in PPA followed by pulmonary vasodilatation, and about 1 h later it was almost back to the control level. No obvious difference in lung lymph flow and protein content was observed between the groups treated with ZAP alone and ZAP plus hypoxia. We conclude that infusion of ZAP into hypoxic animals reduces pulmonary vasoconstriction and that the increase in pulmonary microvascular permeability associated with ZAP is not enhanced by hypoxic conditions occurring at a simulated altitude of 4,000 m.