Success rates of in vitro fertilization versus intracytoplasmic sperm injection in men with serum anti-sperm antibodies: a consecutive cohort study.

Antisperm antibodies (ASAs) are assumed to be a possible causative factor for male infertility, with ASAs detected in 5%-15% of infertile men but in only 1%-2% of fertile ones. It remains unclear whether ASAs have an adverse effect on the outcome of in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). This study investigated differences in the rates of fertilization, pregnancy, and live births associated with serum ASA-positive and ASA-negative men following IVF or ICSI. Five hundred and fifty-four consecutive infertile couples undergoing IVF (n = 399) or ICSI (n = 155) were included. The two-sample two-sided t-test and Chi-square or Fisher's exact test was used for statistical analysis. Lower rates of fertilization (41.7% vs 54.8%, P = 0.03), good embryos (18.9% vs 35.2%, P = 0.00), pregnancy (38.5% vs 59.4%, P = 0.00), and live births (25.8% vs 42.5%, P = 0.00) were observed in men of the IVF group with a positive serum ASA than in those with a negative ASA. ASA positivity/negativity correlated with pregnancy rates (P = 0.021, odds ratio [OR]: 0.630, 95% confidence interval [CI]: 0.425-0.932) and live birth rates (P = 0.010, OR: 1.409, 95% CI: 1.084-1.831) after controlling for the female serum follicle-stimulating hormone level and the couple's ages at IVF. Women coupled with ASA-positive men had lower live birth rates with IVF than with ICSI (25.8% and 47.4%, respectively; P = 0.07). Women coupled with ASA-positive men had lower rates of pregnancy and live births following IVF than those coupled with ASA-negative men but had a similar outcome with ICSI.

Hematopoietic stem and progenitor cells can be enriched by implanting biomaterial into spatium intermusculare.

Hematopoietic stem and progenitor cells (HSPCs) have been used successfully to treat patients with cancer and disorders of the blood and immune systems. In this study, we tried to enrich HSPCs by implanting biomaterials into the spatium intermusculare of mice hind limbs. Gelatine sponges were implanted into the spatium intermusculare of mice and then retrieved after 12 days. The presence of HSPCs in the migrating cells (MCs) was detected by phenotypically probing with CD34(+)Sca-1(+) and functionally confirming the presence of using colony-forming cell assay and assessing the long-term reconstitution ability. The frequency of CD34(+), Sca-1(+), and CD34(+)Sca-1(+) cells and colony formation unit in the MCs was much higher than that in the bone marrow (BM). Moreover, transplanted MCs were able to home to BM, muscle, and spleen, which induced an efficient long-term hematopoietic reconstitution in vivo. In addition, HSPCs within the MCs originated from the BM. Furthermore, the administration of G-CSF greatly reduced the time of implantation, and increased the number of MCs and frequency of HSPCs in the MCs. These data provide compelling evidence that HSPCs can be enriched by implanting biomaterial into spatium intermusculare. Implantation of biomaterial may be seen as the first step to a proof of their applicability to clinical practice in enriching HSPCs.

[Effects of rhBMP-7 on osteoblast differentiation of murine bone marrow mesenchymal stem cells in vitro].

AIM: To study the effects of recombinant human bone morphogenetic protein-7 (rhBMP-7) on osteoblast differentiation of murine bone marrow mesenchymal stem cells (BMSCs). METHODS: The BMSCs were collected from murine bone marrow by density gradient centrifugation. Cells were adherent cultured and expanded in vitro. RhBMP-7 was added into the culture medium of the 3rd passage BMSCs. Five days later, we performed alkaline phosphatase (ALP) staining and detected ALP activity and osteocalcin (OC) content. Osteoblast differentiation marker gene including OC and collage I (Col-I) were assayed by RT-PCR. Total protein was isolated and the secretion of Col-I in the cells was measured by Western blotting. RESULTS: The staining intensity of ALP in the group with rhBMP-7 was stronger than that in the negative control group. ALP activity and OC content of rhBMP-7 group were obviously higher than that of the negative control group. The mRNA of OC and Col-I and the protein of Col-I were highly expressed in the group induced by rhBMP-7. CONCLUSION: RhBMP-7 can induce murine BMSCs to differentiate into osteoblast in vitro.

[Association of susceptibility to chronic rhinosinusitis with genetic polymorphisms of IL-4 and IL-10].

OBJECTIVE: To investigate the relationship between the promoter polymorphism of IL-4 and IL-6 and chronic rhinosinusitis (CRS). METHODS: One hundred and twenty-three patients with CRS and 239 healthy controls in Shanghai region were chosen in this study. The genotype of IL-4 gene -33T>C and -590C>T were determined using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method and the genotype of IL-10 gene -1082A>G was determined using amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) method. Statistical calculations were performed using SAS 8.2 software. RESULTS: Significant differences were found in genotype distribution of -33T>C and -590C>T between the CRS group and the control group (χ2=6.6013, P=0.0102, χ2=6.6013, P=0.0304), and -33T>C remained significant following application of the Bonferroni correction (P<0.025). The relative risks of CRS with -33T>C and -590C>T were 1.818(P=0.0236, 95%CI 1.084-3.050) and 1.838 (P=0.0147, 95%CI 1.127-2.997). There was linkage disequilibrium (LD) between the -33T>C and -590C>T. The coefficient of linkage disequilibrium (D') was 0.77 and the related coefficient (r2) was 0.54. The -33T/-590T haplotype was associated with CRS and the relative risk was 1.653 (P=0.0130, 95%CI 1.107-2.469). There were only two genotypes of IL-10 gene-1082A>G and the frequencies of the AA and AG genotypes were not different between the CRS and control groups. CONCLUSION: The promoter polymorphism of IL-4 -33T>C and -590C>T were associated with the susceptibility of CRS and the -33T/-590T haplotype was a risk factor for CRS, but there were no association between the -1082A>G and CRS.

[Suppression of Aurora-A by RNA interference inhibits laryngeal cancer Hep-2 cell growth].

OBJECTIVE: To investigate the effects of knockdown of Aurora-A by RNA interference on laryngeal cancer Hep-2 cell growth in vitro and in vivo. METHODS: A plasmid containing siRNA against Aurora-A was constructed and transfected into human laryngeal cancer cell line Hep-2. Measurements included the CCK-8 assay for viability and proliferation, Transwell assay for invasion, colony formation assay for cell anchorage-independent growth. Western blot and immunohistochemistry assay for protein expression. Tumorigenicity was observed in vivo. RESULTS: In Hep-2 cells transfected by Aurora-A siRNA (designated as siRNA-3), protein expression of Aurora-A was suppressed by 52%. In CCK-8 assay, absorbance value of siRNA-3 cells (3.268 ± 0.106, (x(-) ± s)) was lower than that of Hep-2 cells (3.722 ± 0.152, F = 17.634, P < 0.001). In Transwell assay, the average invasive cells per field in siRNA-3 cells (110.0 ± 18.0) was less than that in Hep-2 cells (236.0 ± 26.0, F = 26.462, P < 0.01). In colony formation assay, the average colony number of siRNA-3 cells (31.0 ± 6.6) was lower than that of Hep-2 cells (104.0 ± 14.0). The average tumor size in siRNA-3 group was (127.77 ± 174.83) mm(3), which was less than Hep-2 cell group (837.26 ± 101.80) mm(3), (F = 28.187, P < 0.001). Silencing of Aurora-A decreased the expression of focal adhesion kinase (FAK) and matrix metalloproteinase-2 (MMP-2), key regulators in cell adhesion and invasion. CONCLUSIONS: The knockdown of Aurora-A inhibits the growth and invasiveness of Hep-2 cells in vitro and in vivo, which may be a promising therapeutic strategy for LSCC.

High-throughput analysis of DNA fragments using a miniaturized CE system combined with a slotted-vial array sample introduction system.

An automated nanoliter sample introduction system was combined to a liquid-core waveguide (LCW)-based microfluidic CE system for high-throughput analysis of DNA fragments. The main component of the sample introduction system was a motor-driven plate, on which a circular array of bottom-slotted vials containing sample/buffer solutions was placed. A 7 cm-long LCW capillary served as both the sample probe and separation channel. The inlet terminal of the capillary could pass through the slots of the vials for electrokinetic sample introduction, and the capillary outlet was immersed in the solution of a reservoir, behind which a PMT facing directly to the outlet was positioned. A diode laser was used as excitation source for LCW LIF detection. Performance of the system was demonstrated through the separation of DNA fragments. Baseline separation was achieved for all 11 fragments of PhiX174-HaeIII digest DNA with a throughput of 33/h. Theoretical plate number for 603 bp fragment was 7.3x10(6)/m, corresponding to a plate height 0.14 microm. The detection limitation for 603 bp fragment was 0.4 ng/microL with a precision of 2.2% RSD for the peak height. Automated sample changing and introduction were achieved with only 0.3 nL gross sample consumption for each cycle.

[Regional evapotranspiration of different land covers based on remote sensing].

In this paper, surface energy balance system (SEBS) was extended into a regional daily evapotranspiration (ET) estimation model based on remote sensing data, and the extended SEBS was applied to estimate the regional daily ET of Huanghe-Huaihe-Haihe rivers region in Northern China Plain by using MODIS/TERRA data. An analysis was made on the estimated daily ET characteristics of different land covers in the study area by using the spatial analysis module of ArcGIS. Since there were no field observations of ET on each land cover, the estimated daily ET of different land covers was compared with each other, taking the data on April 17, 2001 as an example. The results showed that the regional daily ET estimated by SEBS was reasonable. Wetland and cultivated land had the highest daily ET value, followed by forest-, bush- and grassland, and waste land. The characteristics of the daily ET over these land covers were accorded with the existing knowledge of ET over this region, and coincident to the results of previous work in this area. It was interesting that the residential area also had a higher ET value, which was explained as the higher ET of the land use types, e. g. , water body, street trees, and grass parcels in the resident areas within the pixel scale. The spatial inhomogeneity of ET among the forest-, bush-, grass- and cultivated land covers were caused by the spatial inhomogeneous soil water content under these land covers, and the spatial inhomogeneity of ET over cultivated land could be a potential indicator of making reasonable and effective irrigation schedule for the farmland. The limitations of using SEBS model in daily ET estimation were discussed, especially the possibility of underestimating the ET over water body and wetland covers due to the unsuitable surface parameterization scheme for these land types in the model.

[Experimental study of baculovirus-mediated transfection of spiral ganglion cells in rats].

OBJECTIVE: To study the feasibility and the characteristics of recombinant baculovirus as spiral ganglion cells (SGC) gene transfer vector. METHODS: After the generation of baculovirus- green fluorescent protein( Bac-GFP) according to Bac-to-Bac baculovirus expression system, SGC were infected by Bac-GFP with different multiplicities of infection (MOI) and different concentrations of sodium butyrate. The transfection cell rate and mean fluorescence strength (MFS) were detected by fluorescence microscopy and flow cytometry. Toxicity effects of recombinant baculovirus vectors and sodium butyrate on SGC were determined by spectroscopic measurement of 3-(4, 5)-dimethylthiahiazo (-z-y1)-3, 5-diphenytetrazoliumromide (MTF). RESULTS: Baculovirus was able to infect primary SGC cultures. The dose-response characteristics of Bac-GFP were determined on SGC, and the expression level could be up-regulated by sodium butyrate. Infection with Bac-GFP in the absence or presence of sodium butyrate (< or =10 mmol/L) was considered to be non-cytotoxic to primary SGC. GFP had been expressed in SGC at 6 h post-infection and the highest numbers of cells expressing GFP were observed at approximately 48 h post-infection. CONCLUSIONS: Baculovirus is a novel and promising tool for gene transferring into the cochlear nervous system both for studies of the function of foreign genes and the development of gene therapy strategies.

Study of recombinant human osteogenic protein-1 expressed in prokaryocyte on the repair of extracted socket in rabbits.

The study investigated the effect of recombinant human osteogenic protein-1 (rhOP-1) expressed in prokaryocyte, to promote the healing of alveolar socket. A model of rabbit extracted socket into which the composites of rhOP-1 and gelatin sponge was immediately implanted was created and the osteoinduction of rhOP-1 was assessed by histological method, quantitative measurement of calcium content and alkaline phosphatase (ALP) activity. The result of histology showed that bone healing in rhOP-1 side is 4-6 weeks earlier than that of the control side. ALP activity and calcium content in rhOP-1 side were significantly high compared with that of the control side. rhOP-1 has a satisfactory osteoinduction ability to promote the healing of extracted socket.

[Determination of sulfur dioxide in air by sequential injection spectrophotometry].

A sequential injection spectophotometric method for the determination of sulfur dioxide by decolorizing malachite green in an alkali medium is studied; the decreasing in color is proportional to the amount of sulfur dioxide. The loading sequence, the concentration of reagents, the amount of sample and the flow of reagent rate are optimized by univariate method. With sequential injection sample introducing, the volumes of sample and reagent are reduced greatly and the speed of determination is increased apparently, a throughput of 60 samples/h can be achieved. A liner calibration curve was obtained in range of 0.5-4.0 microg x mL(-1). The determination limit is 0.15 microg x mL(-1), the RSD and the recovery are 1.5% and 94%-108% respectively.

A simple microfluidic system for efficient capillary electrophoretic separation and sensitive fluorimetric detection of DNA fragments using light-emitting diode and liquid-core waveguide techniques.

A miniaturized CE system has been developed for fast DNA separations with sensitive fluorimetric detection using a rectangle type light-emitting diode (LED). High sensitivity was achieved by combining liquid-core waveguide (LCW) and lock-in amplification techniques. A Teflon AF-coated silica capillary on a compact 6x3 cm baseplate served as both the separation channel for CE separation and as an LCW for light transmission of fluorescence emission to the detector. An electronically modulated LED illuminated transversely through a 0.2 mm aperture, the detection point on the LCW capillary without focusing, and fluorescence light was transmitted to the capillary outlet. To simplify the optics and enhance collection of light from the capillary outlet, an outlet reservoir was designed, with a light transmission window, positioned directly in front of a photomultiplier tube (PMT), separated only by a high pass filter. Automated sample introduction was achieved using a sequential injection system through a split-flow interface that allowed effective release of gas bubbles. In the separation of a phiX174 HaeIII DNA digest sample, using ethidium bromide as labeling dye, all 11 fragments of the sample were effectively resolved in 400 s, with an S/N ratio comparable to that of a CE system with more sophisticated LIF.

Feature characterization of microfabricated microfluidic chips by PDMS replication and CCD imaging.

This paper presents a new approach for the metrological characterization of microfabricated features on microfluidic chips, based on a combination of poly(dimethylsiloxane) (PDMS) replication and charge-coupled device (CCD) imaging. A PDMS replicate was cast from the original chip sample, and a 2-mm thick sample slice was cut from the replica at the cross-section to be studied. The digital image of the revealed structural profile was captured by a CCD camera under a microscope, and the image was processed using specially-developed algorithms for CCD image calibration and edge detection. Depth and width measurements obtained using the method agreed well with those gained using a stylus profiler and universal measuring microscope, with a deviation of below 0.9 mum, while profile distortions of deeper structures using stylus profilers were avoided. The method is reliable, non-destructive, and cheap and simple to implement in any analytical laboratory.

Enhancement of signal-to-noise level by synchronized dual wavelength modulation for light emitting diode fluorimetry in a liquid-core-waveguide microfluidic capillary electrophoresis system.

The signal-to-noise level of light emitting diode (LED) fluorimetry using a liquid-core-waveguide (LCW)-based microfluidic capillary electrophoresis system was significantly enhanced using a synchronized dual wavelength modulation (SDWM) approach. A blue LED was used as excitation source and a red LED as reference source for background-noise compensation in a microfluidic capillary electrophoresis (CE) system. A Teflon AF-coated silica capillary served as both the separation channel and LCW for light transfer, and blue and red LEDs were used as excitation and reference sources, respectively, both radially illuminating the detection point of the separation channel. The two LEDs were synchronously modulated at the same frequency, but with 180 degrees -phase shift, alternatingly driven by a same constant current source. The LCW transferred the fluorescence emission, as well as the excitation and reference lights that strayed through the optical system to a photomultiplier tube; a lock-in amplifier demodulated the combined signal, significantly reducing its noise level. To test the system, fluorescein isothiocyanate (FITC)-labeled amino acids were separated by capillary electrophoresis and detected by SDWM and single wavelength modulation, respectively. Five-fold improvement in S/N ratio was achieved by dual wavelength modulation, compared with single wavelength modulation; and over 100-fold improvement in S/N ratio was achieved compared with a similar LCW-CE system reported previously using non-modulated LED excitation. A detection limit (S/N=3) of 10nM FITC-labeled arginine was obtained in this work. The effects of modulation frequency on S/N level and on the rejection of noise caused by LED-driver current and detector were also studied.

[Studies on sequential injection spectrophotometric system using a reflective flow cell for process analysis].

A reflective flow cell was developed and coupled to a sequential injection system and optical fiber photometric detection system based on emitting diode light source. A multistrand bifurcated optical fiber was coupled with incident light, detector and flow cell. Optical fibers were used to carry light from the electronics unit to a reflective flow-through cell and back. The liquid flow path through the cell is linear with a large exit aperture such that bubbles are not trapped in the optical path. The optical arrangement is such that the incident light crosses the liquid flow orthogonally and is reflected back to the receiver fiber. This arrangement reduces the reflective index sensitivity by an order of magnitude relative to a conventional flow cell. The cell showed good immunity to refractive index and air bubble effects. The chromogenic reaction of chloride ion with mercury thiocyanate-iron(III) was used as a model reaction to optimize the experiment system and check the optical system. The reflectance of the reaction was monitored with blue emitting diode. The linear range was 0-100 mg x L(-1) Cl-. A detection limit (3sigma) of 1.2 mg x L(-1), precision of 1.5% (n = 11), and a throughput of 30 samples per hour were achieved.