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Chloroplasts play a crucial role in plant growth and fruit quality. However, the molecular mechanisms of chloroplast development are still poorly understood in fruits. In this study, we investigated the role of the transcription factor SlBEL2 (BEL1-LIKE HOMEODOMAIN 2) in fruit of Solanum lycopersicum (tomato). Phenotypic analysis of SlBEL2 overexpression (OE-SlBEL2) and SlBEL2 knockout (KO-SlBEL2) plants revealed that SlBEL2 has the function of inhibiting green shoulder formation in tomato fruits by affecting the development of fruit chloroplasts. Transcriptome profiling revealed that the expression of chloroplast-related genes such as SlGLK2 and SlLHCB1 changed significantly in the fruit of OE-SlBEL2 and KO-SlBEL2 plants. Further analysis showed that SlBEL2 could not only bind to the promoter of SlGLK2 to inhibit its transcription, but also interacted with the SlGLK2 protein to inhibit the transcriptional activity of SlGLK2 and its downstream target genes. SlGLK2 knockout (KO-SlGLK2) plants exhibited a complete absence of the green shoulder, which was consistent with the fruit phenotype of OE-SlBEL2 plants. SlBEL2 showed an expression gradient in fruits, in contrast with that reported for SlGLK2. In conclusion, our study reveals that SlBEL2 affects the formation of green shoulder in tomato fruits by negatively regulating the gradient expression of SlGLK2, thus providing new insights into the molecular mechanism of fruit green shoulder formation.

Cognitive deficits are a core feature of major depressive disorder (MDD). However, there are no previous studies that directly compare cognitive performance between first-episode drug-naive depressive patients (FDDP) and medicated depressive patients (MDP). Therefore, the aim of this study was to investigate whether there were the differences in cognitive functions between FDDP and MDP. Sixty-two FDDP, 111 MDP and 90 healthy controls were enrolled in a Chinese population. Cognitive functions were assessed using the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS). There were the differences in the RBANS total score (F = 26.55, p

The wet deformation characteristics of coarse aggregates are critical to engineering but have not been well resolved. Therefore, through the large-scale triaxial tests, the sensitivity of the main factors such as initial dry density, initial moisture content, confining pressure and stress level of the filler to the deformation of the wet deformation was analyzed in detail using the orthogonal test method. The results showed that: Among the factors, the primary and secondary factors affecting the axial strain test indexes of each factor are confining pressure> stress level> initial dry density> initial moisture content, and the obtained preliminary law of coarse-grain wet deformation can be coarse-grained roadbed. The design and filling scheme provide important basis.

Objective: Cognitive impairments have been identified as a core feature of major depressive disorder (MDD). To date, no studies on the association between education level and cognitive impairments of MDD from a Chinese population adopting the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) for cognitive measurement emerged. This study is the first to adopt the RBANS to examine whether cognitive impairments of MDD were influenced by education level in a Chinese population. Methods: 90 patients with MDD and 90 healthy controls with matched gender and age were recruited in a case-control study. Cognitive functions were assessed using the RBANS. Moreover, the demographic and clinical data were collected from patients with MDD and healthy controls. Results: There were significant differences in the RBANS total score (F=19.56, p

According to the FLUENT simulation results of the 90°elbow pipe fluid flow condition, the influence of the pressure distribution about the variation of the flow parameters (v、P0 、ρ) and the geometric parameters (R、d、k=R/d、α、β) is analyzed. By using the dimensionless analysis method, on the basis of the influence of the 32 set of parameter variations on the elbow pipe wall pressure distribution regularity, the pressure qualitative function is established including the geometric parameters and the flow parameters. Through the 155 kinds of the FLUENT simulation results, the fitting software 1stOpt is used to verify the internal pressure model above and fit the undetermined coefficients. The verification shows that the model is of a higher calculation precision and the relative error isn’t exceeding 0.003%. It provides a theoretical basis to check the strength of the bend and design the pipeline with high pressure and speed.

On the premise of the simulation data for the inner fuild passing the elbow pipe by means of software named FLUENT and dimensional analysis, the formula modle of inner surface pressure for elbow pipes came up by analyzing the relationships among the inner surface pressure P ,the fluid outlet pressure P0 ,axial angle α, circular angle β and the degree of curvature k; Depending on the inner surface pressure data simulated under 150 kinds of conditions, the pressure formula was fitted by means of 1stOpt software; The verification results show that the calculated value by the formula matched with the value of simulation by FLUENT software very well, the maximum relative error was 0.004%, and the research conclusion layed the theoretical groundwork for the design of the high pressure and high velocity elbow.

According to the principle of harmonious dimensions, the qualitative distribution model of the internal pressure on elbow pipes were determined; Through FLUENT numerical simulation, the internal pressure were given under 96 kinds of gas fluid conditions; By analyzing the variation of the internal pressure with flow and geometrical parameters, the variation of the internal pressure distribution within the elbow pipes has been studied; By means of 1stOpt nonlinear fitting package, the formula of three-dimensional internal pressure distribution was determined; Compared with the numerical results, the maximum relative error is 0.012%. The formula provides theoretical bases for strength check, the transport pipeline and wall thickness design of the high pressure, high velocity elbow.

Using the methods of dimensional analysis and the FLUENT analytic software, a quantitative description method is built; the influents of parameters, including the degree of curvature k=R/d, fluid density ρ, axial angle α, circle angle β, fluid outlet pressure P0 and fluid velocity v on the distribution of pressure on inner surface of elbow pipe are studied from the simulation results; Finally, a three-dimensional distribution model of pressure which is considering the fluid characteristic and the elbow pipe geometric parameters is built by fitting the simulation data. The research conclusion lays the groundwork for strength check and structural design of the elbow pipe.

The processes of the fuild through the elbow pipe under different conditions are simulated by the FEM software named FLUENT. Then, the effects of varied parameters on the distribution of pressure on inner suface of elbow pipe are studied with dimensionless analysis method. Finally, a distribution model of pressure at the the minimum life section where the axial angle α is 45°, degree of curvature k=R/d, circle angle β, fluid density ρ, fluid outlet pressure P0 and fluid velocity v is built and verified by fitting the simulation data. The research conclusion lays the groundwork for strength check and structural design of the elbow pipe

Two thermostable glucoamylases were produced from Aspergillus niger B-30 by submerged fermentation. The two glucoamylases GAM-1 and GAM-2 were purified by ammonium sulfate precipitation, diethylaminoethyl-cellulose fast flow(DEAE FF) and Superdex G-75 gel filtration columns. The molecular weights of GAM-1 and GAM-2 were determined as 9.72×104 and 7.83×104 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), while the molecular weights of GAM-1 and GAM-2 were determined to be 8.05×104 and 7.04×104 by matrix assisted laser desorption ionizationtime-of-flight(MALDI-TOF) mass spectrometry, respectively. Both the enzymes were glycosylated, with 10.4% and 11.4% carbohydrate content, respectively. The optimal pH and temperature were 4.0–4.6 and 70 °C for both. The two glucoamylases were maintained 100% relative activity after incubation at 60 °C for 120 min. After the hydrolysis of starch for 120 min, glucose was the only product, confirming that the two enzymes were of high efficiency towards starch. The GAM-2 exhibited higher catalytic activity towards oligosaccharides such as maltose than GAM-1, and the kinetic analysis shows that the affinity of GAM-2 to starch was lower than that of GAM-1. The high thermostability and effectiveness make the two glucoamylases potentially attractive for biotechnological application.

The aim of this study was to investigate the effect of a B-cell-specific MLV integration site-1 (Bmi-1) RNA interference (RNAi) expression vector on the proliferation and invasiveness of laryngeal carcinoma. We constructed a lentiviral vector expressing Bmi-1-specific short hairpin RNA (shRNA), and transfected it into HEp-2 cells. Bmi-1 gene expression was detected by real-time RT-PCR and western blot analysis. We used flow cytometry and TUNEL assay to analyze the apoptosis of transfected cells, and examined cellular growth in vitro by MTT assay. We established an animal model and evaluated the therapeutic effects of small interfering RNA (siRNA) against Bmi-1. siRNA against Bmi-1 significantly knocked down Bmi-1 expression in HEp-2 cells, induced cell cycle arrest at the G1 phase, inhibited cell proliferation and promoted cell apoptosis. Lentiviral Bmi-1-shRNA vector transfection also significantly reduced cell migration. The formation and growth rate of xenograft tumors in mice transfected with siRNA against Bmi-1 was significantly reduced. The loss of mitochondrial membrane potential, the release of cytochrome c from the mitochondria into the cytosol, and the increased activity of caspase-3, -8 and -9 occurred concomitantly with the inhibition of Bmi-1. Our data indicate that siRNA against Bmi-1 significantly suppresses tumor growth and induces apoptosis in vitro and in vivo.

Background Cognitive deficits have been identified as one of core clinical symptoms of major depressive disorder (MDD). Accumulating evidence indicated that triglycerides (TG) might be associated with MDD and cognitive decline. Objective This study examined whether patients with MDD had poorer cognitive functions than healthy controls, and further investigate whether TG levels were involved in MDD, and its cognitive impairments in a Han Chinese population. Method 115 patients with MDD and 119 healthy controls were enrolled. Cognitive functions were assessed by the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS), and serum TG levels were examined using enzymatic colorimetry. Results TG levels were higher in patients with MDD than healthy controls after controlling for the variables. Cognitive test scores were lower in patients with MDD than healthy controls except for visuospatial/constructional index after controlling for the variables. TG levels were negatively correlated with visuospatial/constructional score, delayed memory score and RBANS total score of MDD. Further multivariate regression analysis showed that TG levels were negatively associated with visuospatial/constructional score, attention score, delayed memory score and RBANS total score of MDD. Conclusions Our findings supported that serum TG levels might be involved in MDD, and play an important role in cognitive impairments of MDD, especially in delayed memory. Moreover, patients with MDD experienced greater cognitive impairments than healthy controls except for visuospatial/constructional index.

We present the experimental results of picosecond pulse laser microstructuring of silicon in the SF6 atmosphere using the galvanometer scanning technique. With appropriate irradiation conditions, a large variety of surface microstructures even nanostructures were produced. By rapidly scanning, a uniform array of micro-crater structure with 10nm in depth is generated on the crystal silicon surface. At low scanning speed, large area uniform self-organizing micro-grating, micro-hole, or micro-cone structure can be produced by cumulative pulses irradiation. These results suggest that picosecond laser scanning technique is a flexible method to produce uniform surface micro-nanostructures, which exhibits great potential for applications in photoelectron devices, special surface marking and surface modification.

To evaluate the electromagnetic radiation pollution in Changchun City and determine the risk of pollution, the probability theory of extreme value distribution is used to build the computational model of electromagnetic pollution risk and discusses their property according to situation that the current urban growth of electromagnetic energy mainly comes from mobile base stations and radio frequency radiation sources. According to the integrated electric field intensity which monitored from Changchun City’s 442 monitoring station by electromagnetic environment monitoring station in Jilin Province, the risk of electromagnetic pollution in Changchun City probability is calculated, ordinary Kriging method is used to draw the map of Changchun City electromagnetic radiation field strength and calculates electromagnetic pollution risk probability. The result indicates that the electromagnetic environment is safe. The value of electric field strength in the office area is higher than other regions. The map drawing by fuzzy Kriging method is more accurately than the map drawing by ordinary Kriging method.

Sulfur-doping and broad band absorptive black silicon materials were fabricated by picosecond laser irradiation. Two kinds of microstructures, laser induced periodic surface structure (LIPSS) and conical spikes were obtained by changing parameters of laser scanning. Black silicon solar cells with back surface field were explored. Influences of different rear side structures to devices were presented and conversion efficiency of 9% is available.

The microstructured samples were prepared by irradiating silicon surface with picosecond laser pulses in SF6. The surface morphology of microstructured samples irradiated at different laser fluence was characterized by SEM. The samples exhibited high optical absorptance over a wide wavelength range between 300 and 2700 nm. The absorptance of samples irradiated with the fluence of 1.0 J/cm2was measured to be up to 95% between 1100 and 2700 nm. The infared absorptance of the surface-structured samples increased with increasing fluence. Whereas, as the annealing temperature was increased, the infared absorptance of the samples irradiated at the same fluence decreased. A tentative explanation for the effects of laser fluence and annealing temperature on the infared absorptance has been proposed based on the formation of mid-band gap impurity bands and the multiple reflections of light between microstructures.

BACKGROUND Elevated phospholipase A2 (PLA2) activity is reported to be involved in the development of schizophrenia. Further study revealed an association between PLA2 groups XIIA (PLA2G12A) polymorphism and patients with schizophrenia in a northeast Chinese Han population. OBJECTIVE This study will further examine whether PLA2G12A rs3087494 polymorphism is associated with patients with schizophrenia in a southern Chinese Han population. METHODS This polymorphism was genotyped in 438 patients with schizophrenia (diagnosed according to Diagnostic and Statistical Manual of Mental Disorders-IV) and 876 healthy controls using a case-control design. Demographic and clinical data were collected in all subjects. RESULTS The allele and genotype frequencies of PLA2G12A rs3087494 polymorphism significantly differed between groups (both, p

Large amounts of sugar are imported into grape berries from source leaves during ripening, and sucrose transporters play a key role during this process. In this study, a putative grape sucrose transporter gene VvSUC27, primarily expressed in sink tissue, was transformed into a yeast strain to characterize its function as a sucrose transporter. Sucrose was taken up by yeast transformed with VvSUC27 at an optimum pH of 4.0–5.0 and a Km of 8.0–10.5 mM, indicating VvSUC27 is a LAHC (low-affinity/high-capacity) sucrose transporter. The ability of sucrose uptake in transformed yeast was activated by monosaccharides and inhibited by maltose and DEPC.

Electrical and optical coupling in an electroabsorption (EA) modulator integrated with a distributed feedback (DFB) laser have been investigated. The integrated device is treated as a three-port optoelectronic device with two electrical ports and one optical output port. The scattering parameters of this three-port device have been measured in the designed experiment. The measured results indicate that there exists the electrical coupling between the DFB laser and EA modulator of the integrated light source whenever the current applied to the laser section is below or above the threshold current, and the optical coupling will have stronger influence on the frequency responses than the electrical coupling when the bias current is above the threshold. A small-signal equivalent circuit model for the integrated device is established considering both the electrical and internal optical coupling. Experiments show that the equivalent circuit model is reasonable and the determined element values are correct. Based on the measurement and modeling, the influences of the electrical and optical coupling on the high-frequency responses are investigated and the effective measure to eliminate the additional modulation in the DFB laser are discussed.

Recently, ETHYLENE OVERPRODUCER 1 (ETO1) had been cloned and identified as a negative post-transcriptional regulator in the ethylene biosynthesis in Arabidopsis. However, little was known about the role of ETO1 in other species, especially in tomato, which was an ideal model for studying the biosynthesis of ethylene during tomato fruit ripening. In this study, a tomato ETHYLENE OVERPRODUCER 1-LIKE 1 (LeEOL1) was cloned. The LeEOL1 cDNA was 3,515 bp long and carried an ORF that putatively encoded a polypeptide of 886 amino acids with a predicted molecular mass of 95 kDa. It shared 74% identity in amino acid sequence with Arabidopsis EOL1 and had one BTB (Broad-complex, Tramtrack, Bric-a-brac) domain and two TPR (tetratricopeptide repeat) domains, which were also conserved domains in AtEOL1. RT-PCR analysis of the temporal expression of LeEOL1 showed that its transcript decreased companied with increase of ethylene production in tomato ripening. The level of LeEOL1 transcripts in wild type tomato fruit at m...

In this study, we examined the specialized features of the outer hair cells (OHCs) and the stereocilium bundles of the bat cochlear fovea. Bat cochlea hair cells were observed by scanning and transmission electron microscopy, and the auditory brainstem response thresholds were assessed. The stereocilia bundles of the OHCs were extremely short. The OHC bodies were flask-shaped and cambiform or ball-shape in the cochlear fovea. Digitations in the Deiters cells had exaggerated lengths, and cup formation of the Deiters cell, housed at the bottom of the OHC in the base of the cell, showed a specialized shape. Our results provide the first evidence that different shapes of the OHCs in the cochlea fovea are related to the high-frequency function of auditory response. Echolocating bats have cochlear morphologies that differ from those of non-echolocating animals. Bat cochlear foveae are specialized for analyzing the Doppler-shifted echoes of the first-harmonics of the CF2 component; these are overrepresented in the frequency range around the dominant harmonic of the echolocation calls of bats. However, the OHCs of the bat cochlear fovea have not been fully characterized.

1.5µm n-type modulation-doping InGaAsP/InGaAsP strained multiple quantum wells grown by low pressure metalorganic chemistry vapor decomposition technology is reported for the first time in the world. N-type modulation-doped lasers exhibit much lower threshold current densities than conventional lasers with undoped barrier layers. The lowest threshold current density we obtained was 1052.5 A/cm2 for 1000 µm long lasers with seven quantum wells. The estimated threshold current density for an infinite cavity length was 94.72A/cm2/well, reduced by 23.3% compared with undoped barrier lasers. The n-type modulation doping effects on the lasing characteristics in 1.5µm devices have been demonstrated.

A directly-modulated distributed-feedback laser module has been achieved with a ridge-waveguide laser chip in our laboratory. It is packaged in butterfly housing with a K connector, and the bandwidth of the laser module is up to 24 GHz with in-band flatness of ±3 dB. It shows a high linearity with input 1-dB compression point of higher than 23 dBm and third-order intercept of 32.2 dBm at 24 GHz. Meanwhile, the packaged DFB laser module gives rather low relative intensity noise of less than -150 dB/Hz when the bias current is 75 mA and it can show a great performance in analog optical communication systems.

Aim: To investigate its effect on the proliferation and invasion of laryngeal carcinoma and understand the potential underlying mechanisms to provide new targets for the diagnosis and treatment of recurrent laryngeal cancer metastasis. Methods: We constructed a lentiviral vector expressing EGFL7 specific shRNA, and introduced it in EGFL7 functions were attenuated by a lentiviral vector harboring shRNA targeting at EGFL7 in laryngeal carcinoma cell line Hep-2. Prolifereation and invasion assays were carried out in vitro. And in vivo tumor burden assay was done in nude mice. Results: The expression of EGFL7 was knocked-down by 80% in hep-2 cells transfected by the lentiviral EGFL7 shRNA vector and EGFL7 gene expression was detected by realtime PCR and Western blotting analysis respectively. The flow cytometric analysis showed that arrested the cell cycle in G1 phase, In tumor burden assay, to parental And vector control cells, the survival rates Of nude mice in EGFL7 shRNA group dropped down from the first day after implantation as indicated by MTT assay (P < 0.05). The formation and growth rate of xenograft tumor in mice transfected with siRNA against Bmi-1 slowed down significantly. Conclusion: Attenuation of EGFL7 function significantly suppresses tumor growth and induces apoptosis, both in vitro and in vivo. EGFL7 may be play a key role in invasion and metastasis of Laryngeal squamous cell carcinoma (LSCC), thus would to be a new target for gene therapy in LSCC.

To investigate the roles of Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) in host defense against Streptococcus pneumoniae infection in the middle ear.Wild-type (WT) C57BL/6J, TLR2-deficient (TLR2(-/-)) and TLR4-deficient (TLR4(-/-)) mice were inoculated with Streptococcus pneumoniae (1 × 10(6)CFU) through the tympanic membrane. All animals were tested the mouse ABR thresholds and tympanometry measurement before, and 1 day, 3 days and 7 days following pneumococcal challenge. Blood bacterial titer were determined by plating 50 µl volumes of 10-fold diluted blood. Histological analysis of middle ear and inner ear were performed by fixation, decalcification, embedded section, and counterstained with hematoxylin/eosin and toluidine blue staining. Semi-quantitative RT-PCR was applied to determine mRNA accumulation of TLR2 and TLR4 related genes.Forty of 68 TLR2(-/-) mice and twenty-one of 59 TLR4(-/-) mice showed bacteremia and died within 3 days after the pneumococcal challenge, however, only 9 of 52 WT mice died. The survive mice were shown have more severe hearing loss in the TLR2(-/-) and TLR4(-/-) mice than in the WT mice, indicated by ABR thresholds, at 3 or 7 days postinoculation. The histological pathology was characterized by effusion and tissue damage in the middle ear, and in the TLR2(-/-) and TLR4(-/-) mice, the outcome of infection became more severe at 7 days. At both 3 and 7 days after challenge, the TLR2(-/-) mice had higher blood bacterial titers than WT mice (P0.05). Temporal bone histopathologic change indicated that 3 days after the pneumococcal challenge, the TLR2(-/-) and TLR4(-/-) mice showed effusion and tissue damage in the middle ear, and the infection became more severe at 7 days postinoculation. TLR2(-/-) mice showed severe inflammatory cell infiltration in the cochlear, the organ of Corti showed the outer hair cells damage, the tectorial membrane swelling, degeneration of the stria vascularis, and severe loss of spiral ganglion cells; However, the WT mice was not found the cell infiltration and tissue damage in the cochlear, the organ of Corti shown normal of outer hair cells. Mast cells were not found in the middle ear mucosa of TLR2(-/-) mice, but in the TLR4(-/-) and WT mice, more mast cells were found in the middle ear mucosa of effusion ear by 3 and 7 days postchallenge. Moreover, by 3 days postchallenge, the mRNA accumulation levels of NF-κB, tumor necrosis factor alpha (TNFα), interleukin1β, MIP-1α, MUC5AC and MUC5B were significantly lower in the ears of TLR2(-/-) mice than that in WT and TLR4(-/-) mice.TLR2(-/-) mice may produce relatively low levels of proinflammatory cytokines following pneumococcal challenge, thus hindering the clearance of bacteria from the middle ear and leading to sepsis and high mortality rate. This study indicated that TLR2 and TLR4 are important in the molecular pathogenesis and host response to otitis media.

Cadherin 23 (CDH23) is an important constituent of the hair cell tip link in the organ of Corti. Mutations in cdh23 are associated with age-related hearing loss (AHL). In this study, we proposed that the Cdh23(nmf308/nmf308) mice with progressive hair cell loss had specific morphological changes and suffered a base to apex gradient and age-related hearing loss, and that mutations in cdh23 were linked to AHL. The Cdh23(nmf308/nmf308) mice produced by the N-nitrosourea (ENU) mutagenesis program were used as an animal model to study AHL and progressive hair cell loss. RT-PCR was performed to confirm the cdh23 mutation in Cdh23(nmf308/nmf308) mice and genetic analysis was used to map the specific mutation site. Distortion product otoacoustic emission (DPOAE) assay and acoustic brainstem evoked response (ABR) threshold analysis were carried out to evaluate the AHL. Cochlear histology was examined with scanning electron microscope (SEM) and transmission electron microscope (TEM), as well as the nuclear labeling by propidium iodide staining; terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) assay and caspase-3 activities were examined to evaluate cell apoptosis. Genetic mapping identified the candidate gene linking AHL in Cdh23(nmf308/nmf308) mice as cdh23. A mutation in exon3 (63 T>C) was screened as compared with the sequence of the same position of the gene from B6 (+/+) mice. The cochleae outer hair cells were reduced from 5-10% at one month to 100% at three months in the basal region. DPOAE and ABR exhibited an increasing threshold at high frequencies (≥16kHz) from one month of age. Morphological and cellular analysis showed that Cdh23(nmf308/nmf308) mice exhibited a time course of histological alterations and cell apoptosis of outer hair cells. Our results suggest that the cdh23 mutation may be harmful to the stereociliary tip link and cause the hair cell apoptosis. Due to the same cdh23 mutations in human subjects with presbycusis (Petit et al., 2001; Zheng et al., 2005), the Cdh23(nmf308/nmf308) mouse is an excellent animal model for investigating the mechanisms involved in human AHL.

A high-speed DFB laser module in butterfly housing has been fabricated in our laboratory, which is operated in direct modulation for Ku-band. The bandwidth of the laser module is measured up to 18 GHz with highly linear characteristics and low relative intensity noise. The input points for 1 dB compression and third order interception of the laser have also been obtained as 23 dBm and 30 dBm respectively, and it can be utilized in analog optical link systems with good performance.

To observe the effect of the lentiviral vectors expressing small interfering RNA (siRNA) for survivin gene knockdown in inhibiting Hep-2 cell growth in vitro and its tumorigenicity in nude mice.The tumorigenicity of Hep-2 cells transfected with the siRNA mediated by the lentiviral vectors was tested in nude mice. The expression of survivin gene of the transfected cells at the mRNA and protein levels were detected by RT-PCR and Western blotting, respectively, and the cell cycle changes were analyzed by flow cytometry.Transfection of the siRNA targeting survivin significantly decreased the expression of survivin mRNA and protein in Hep-2 cells in vitro by 60%-85% and 70%, respectively, resulting also in increased cell apoptosis as shown by flow cytometry (P0.01). The transfection significantly lowered the tumorigenicity of the cells in nude mice.The lentiviral vectors expressing survivin siRNA can significantly inhibit survivin gene expression in Hep-2 cells and induce the cell apoptosis in vitro, and suppress the tumorigenicity of the cells in nude mice.

This paper presents a new technique to generate microwave signal using an electro-absorption modulator (EAM) integrated with a distributed feedback (DFB) laser subject to optical injection. Experiments show that the frequency of the generated microwave can be tuned by changing the wavelength of the external laser or adjusting the bias voltage of the EAM. The frequency response of the EAM is studied and found to be unsmooth due to packaging parasitic effects and four-wave mixing effect occurring in the active layer of the DFB laser. It is also demonstrated that an EA modulator integrated in between two DFB lasers can be used instead of the EML under optical injection. This integrated chip can be used to realize a monolithically integrated tunable microwave source.

To investigate the apoptosis-inducing effect of caspase-3 on human nasopharyngeal carcinoma cells (CNE2).Recombinant caspase-3 was subcloned into the eukaryotic expression vector PEGFP-C1 containing the reporter gene using DNA recombinant technique. CNE2 cells were transfected with the recombinant caspase-3 gene via lipofectamine 2000 and the expression of caspase-3 mRNA was detected by reverse transcription-polymerase chain reaction. The cell morphological changes were observed under fluorescence microscope and electron microscope and the cell survival rate after the transfection was assessed by MTT assay.Transfection with the recombinant caspase-3 gene induced significant apoptosis in CNE2 cells, which exhibited obvious morphological changes typical of apoptotic cells.The recombinant caspase-3 gene can inhibit the growth and effectively induce apoptosis of CNE2 cells.

Variant stereociliary bundles of cochlear outer hair cells (OHCs) in guinea pigs were discovered fortuitously in approximately 50% of the animals examined by several investigators (1). We also found this phenomenon frequently in our previous observations of guinea pigs, even in human newborns (2). Accordingly, the anomaly has been described in a subset of animals obtained from animal suppliers, including golden hamsters from Japan (3) and guinea pigs from the U.K. and China (1,3-5). Planar cell polarity (PCP) refers to the polarization of cells within the plane of a cell sheet. A distinctive epithelial PCP in vertebrates is the uniform orientation of stereociliary bundles of the sensory hair cells in the mammalian cochlea (6,7). Planar cell polarity is a fundamental aspect in the development of the inner ear. Not only is the uniform orientation of stereociliary bundles on hair cells one of the clearest examples of planar polarity in a mammalian system, the cochlea is also an ideal model system to study this (8,9). Yoshida and Liberman (10) histologically examined the cochleae from 100 albino guinea pigs and observed 24 of these to have rotated stereociliary bundles, with males and females almost equally represented in the affected population. This congenital anomaly was strictly limited to first-row OHCs, of which 15 to 50% were found to be rotated up to a possible 180 degrees from the normal orientation. Compound action potentials (CAPs) were measured on mice that displayed abnormal hair bundles to analyze whether this structural alteration had a functional consequence. Indeed, this structural change resulted in CAP thresholds increased by 5 to 10 dB on average when compared with thresholds of control animals. The level of CAP threshold increase was correlated with a quantitation of the degree of stereociliary rotation to demonstrate that first-row OHCs play a greater role in amplifying signals within the cochlea than do the second- and third-row OHCs (10). Another study found that OHCs from av6J mutant (6J/6J) mice appear rotated from normal position in the basal turn. Additionally, in av2J mice, OHCs are severely deformed and show abnormal organization of the stereocilia (11), whereas some of the cells have rotated stereocilia bundles, with the tip of the V pointing at an angle of up to 90 degrees away from the normal orientation. Normal cochleae were apparent in the first row; however, 50% OHCs were rotated bundles, but they were considered normal by this investigator. It was concluded by this author that the irregular arrangement and natural loss of stereociliary bundles of the OHCs are not considered to be pathologic (12). In this current study, assessment of cochlear OHC rotation was not based on criteria from previous literature. Rather, our study involved the assessment of hearing acuity in 100 guinea pigs that were then independently examined for cochlear morphology and histology. We measured hearing acuity by both auditory brainstem response (ABR) and distortion product otoacoustic emission (DPOAE) testing. It is well established that DPOAE analysis can discern alterations of OHC functionality that are not detected by ABR analysis (9). The cochlea of each animal was examined by electron microscopy for stereociliary bundle misorientation that was mapped to specific cochlear areas. The histologic results of cochlear damage were compared with the DPOAE analysis to see if both methods concomitantly pinpointed similar areas of stereociliary bundle misorientation. The goal is to use DPOAE analysis to discern subtle changes in stereociliary bundle misorientation.

To construct a recombinant adenovirus vector carrying antisense heat shock protein 70 (HSP70) cDNA and observe its effect on inhibiting the growth of laryngeal carcinoma Hep-2 cells.The HSP70 gene fragment encoding the 5' region was cloned reversely into the shuttle plasmid PAdTrack-CMV, and the resultant plasmid was recombined with the backbone plasmid PadEasy-1 in the E.coli Bj5183 cells to generate the recombinant adenovirus vector. The adenovirus were then packaged and amplified in 293 cells, and the viral titer was determined using GFP.The recombinant adenovirus vector carrying antisense HSP70 cDNA was constructed successfully with a viral titer of 8 x 10(9). HSP70 expression of Hep-2 cells was obviously blocked by antisense HSP70 RNA, and Western blotting and immuohistochemistry demonstrated that cells transfected with antisense HSP70 did not express or express HSP70 at low levels. Flow cytometry presented apoptotic peak in the antisense HSP70-transfected cells, but not in the control cells.The recombinant adenovirus vector containing antisense HSP70 cDNA can effectively deliver antisense HSP70 gene into Hep-2 cells, suggesting the great potential of this gene therapy strategy in management of human laryngeal carcinoma.

To study the protective effect of local gene therapy with adeno-associated virus (AAV)-mediated neurotrophin-3 (NT-3) on the function and morphology of the cochlea of guinea pigs with gentamicin-induced hearing loss.Hearing loss was induced with gentamicin (80 mg.kg(-1).day(-1) injected intramuscularly) in 18 pigmented guinea pigs 4 days prior to gene transfer. The guinea pigs were then divided into groups A, B, and C for AAV-mediated NT-3 gene transfer (n=7), AAV infection (n=7) or no particular intervention (n=4), respectively. Mini-Osmotic pump were implanted in either side of the ears in groups A and B, and the guinea pigs were injected with gentamicin (80 mg.kg(-1).day(-1)) intramuscularly since the operation day for 10 consecutive days. In group C, only gentamicin was administrated. Before and 14 days after gentamicin administration, auditory brainstem response audiometry (ABR) and distort-product otoacoustic emissions (DPOAE) were recorded, and the animals sacrificed to observe the morphological changes of the cochlear microscopically.Compared with groups B and C, the animals in group A showed better auditory ability (ABR and DPOAE) and significantly higher surviving rate of the outer hair cells (P0.05).AAV-mediated NT-3 gene transfer may protect and repair the cochlear hair cells and auditory function damaged by aminoglycoside ototoxicity in guinea pigs, and aseptic procedure is of vital importance in cochlear local gene therapy.

Recently, ETHYLENE OVERPRODUCER 1 (ETO1) had been cloned and identified as a negative post-transcriptional regulator in the ethylene biosynthesis in Arabidopsis. However, little was known about the role of ETO1 in other species, especially in tomato, which was an ideal model for studying the biosynthesis of ethylene during tomato fruit ripening. In this study, a tomato ETHYLENE OVERPRODUCER 1-LIKE 1 (LeEOL1) was cloned. The LeEOL1 cDNA was 3,515 bp long and carried an ORF that putatively encoded a polypeptide of 886 amino acids with a predicted molecular mass of 95 kDa. It shared 74% identity in amino acid sequence with Arabidopsis EOL1 and had one BTB (Broad-complex, Tramtrack, Bric-à-brac) domain and two TPR (tetratricopeptide repeat) domains, which were also conserved domains in AtEOL1. RT-PCR analysis of the temporal expression of LeEOL1 showed that its transcript decreased companied with increase of ethylene production in tomato ripening. The level of LeEOL1 transcripts in wild type tomato fruit at mature green stage did not distinctively change when treated with exogenous ethylene.

Virus-induced gene silencing (VIGS) is an attractive reverse-genetics tool for studying gene function in plants. We showed that silencing of a phytoene desaturase (PDS) gene is maintained throughout TRV-PDS-inoculated tomato plants as well as in their flowers and fruit and is enhanced by low temperature (15 degrees C) and low humidity (30%). RT-PCR analysis of the PDS gene revealed a dramatic reduction in the level of PDS mRNA in leaves, flowers and fruits. Silencing of PDS results in the accumulation of phytoene, the desaturase substrate. In addition, the content of chlorophyll a, chlorophyll b and total chlorophyll in the leaves of PDS-silenced plants was reduced by more than 90%. We also silenced the LeEIN2 gene by infecting seedlings, and this suppressed fruit ripenning. We conclude that this VIGS approach should facilitate large-scale functional analysis of genes involved in the development and ripening of tomato.

Virus-induced gene silencing (VIGS) is a powerful tool for the study of gene function in plants. Here we report that either by syringe-infiltrating the tobacco rattle virus (TRV)-vector into the surface, stem or carpopodium of a tomato fruit attached to the plant or by vacuum-infiltrating into a tomato fruit detached from the plant, TRV can efficiently spread and replicate in the tomato fruit. Although VIGS can be performed in tomato fruit by all of the means mentioned above, the most effective method is to inject the TRV-vector into the carpopodium of young fruit attached to the plant about 10 days after pollination. Several reporter genes related to ethylene responses and fruit ripening, including LeCTR1 and LeEILs genes, were also successfully silenced by this method during fruit development. In addition, we found that the silencing of the LeEIN2 gene results in the suppression of tomato fruit ripening. The results of our study indicate that the application of VIGS techniques by the described methods can be successfully applied to tomato fruit and is a valuable tool for studying functions of the relevant genes during fruit developing.