[Co-overexpression of human tissue kallikrein 1 and human metalloproteinase 1 tissue inhibitor inhibits neointima formation in the rat artery after balloon angioplasty].

OBJECTIVE: To investigate the effects of adenovirus-mediated human tissue kallikrein 1(hTK-1) and/or human tissue metalloproteinase inhibitor 1 (hTIMP-1) gene delivery on the neointima formation in balloon-injured rat carotids and related mechanism. METHODS: Forty-six male Sprague-Dawley rats were randomly assigned into 6 groups with the random number table: (1) sham-operated group(n=6), (2) angioplasty group (n=8), (3) vector virus group (n=8), (4) hTK-1 group (n=8), (5) hTIMP-1 group (n=8), (6) hTK-1-hTIMP-1 group (n=8). Except sham rats, all rats underwent carotid artery balloon injury and local delivery of saline or different recombined adenoviruses respectively. Rats were sacrificed 14 days later. Intima/media area ratio was assessed on hematoxylin-eosin stained tissue section. Immunofluorescence images stained for hTK-1, hTIMP-1 were obtained and analyzed by the confocal microscope for co-localization examination of hTK-1 and hTIMP-1. The protein expression levels of hTK-1, hTIMP-1, matrix metalloproteinases(MMP)-2 and MMP-9 were determined by Western blot. Immune histochemical staining for PCNA was also performed. RESULTS: (1)Intima area, intima/media area ratio, PCNA, MMP-2 and MMP-9 levels were all significantly increased in rats underwent angioplasty (did or did not receive vector virus) compared with sham-operated rats (all P<0.01) while above parameters were similar between rats underwent angioplasty or vector virus delivery (all P>0.05). (2) The intima area of rats received vector virus, hTK-1, hTIMP-1 or dual gene transfer were (0.160±0.010), (0.110±0.015), (0.121±0.016) or (0.081±0.008) mm(2) respectively, intima area was similar between rats received hTK-1 or hTIMP-1 (P>0.05), differences were found between other groups (all P<0.01). The intima/media area ratio of rats received vector virus, hTK-1, hTIMP-1 or dual gene transfer were 2.035±0.117, 1.443±0.097, 1.522±0.078 or 0.972±0.072 respectively, no difference was found between rats received hTK-1 or hTIMP-1 in intima/media area ratio (all P>0.05), differences were found between other groups (all P<0.01). The MMP-2 and MMP-9 expression of rats received vector virus, hTK-1, hTIMP-1 or dual gene transfer were 0.817±0.036, 0.606±0.044, 0.571±0.061 or 0.455±0.030 and 0.745±0.057, 0.613±0.038, 0.582±0.050 or 0.473±0.038 respectively, no difference was found between rats received hTK-1 or hTIMP-1 in MMP-2 or MMP-9 expression (all P>0.05), differences were found between other groups (all P<0.01). The PCNA expression of rats received vector virus, hTK-1, hTIMP-1 or dual gene transfer were 0.065±0.007, 0.052±0.004, 0.055±0.007 or 0.031±0.004 respectively, no difference was found between rats received hTK-1or hTIMP-1 in PCNA expression (all P>0.05), differences were found between other groups (all P<0.01). CONCLUSION: hTK-1 and hTIMP-1 co-overexpression may synergistically inhibit neointimal hyperplasia, attenuate vascular remodeling and reduce restenosis possibly via down regulating the expressions of PCNA, MMP-2 and MMP-9 in balloon-injured rat carotids.

Solid-phase microextraction from small volumes of sample in a glass capillary.

A new sampling method is proposed for solid-phase microextraction (SPME), in which the extraction is carried out in a glass capillary containing a few microliters of sample. When an adsorption-type fiber is used for SPME, the equilibrium between aqueous sample and coating can be described by a Langmuir isotherm. Since the total amount of analytes and coexisting substances stays at a low level in a small volume of sample, the linear concentration range of analytes will be extended for SPME to be applied in quantification and the interference caused by sample matrix will be reduced. In addition, sampling in a capillary has a short diffusion distance and extraction equilibrium is established in 5-10 min. It is important in clinical analysis and therapeutic drug monitoring to be able to analyse sample volumes of samples. The feasibility of the new sampling method is demonstrated by the extractions of p-hydroxybenzaldehyde and a synthetic solution containing 1-naphthol, paeonol and 1-naphthylamine.

Computer-aided development of a high-performance liquid chromatographic method for the determination of hydroxyanthraquinone derivatives in Chinese herb medicine rhubarb.

With computer simulation predicting separation in reversed-phase gradient elution, a method to separate and determine five hydroxyanthraquinone derivatives having a wide range of polarity in extract of Chinese herbal medicine rhubarb has been developed. The software DryLab was used to model the retention behavior of the compounds as a function of gradient conditions, based on data from two scouting gradient runs. Under the optimized conditions, i.e. column, Zorbax RX-C18, 5 microm, 15x0.46 cm; solvent A, 36 mM triethylamine phosphate (TEAP), pH=2.5; solvent B, ACN; gradient, 36/36/80/80% B at 0/5.5/20.5/25.5 min; flow-rate, 1.00 ml/min; temperature, ambient, the method was successfully applied to monitor the quality of rhubarb from different sources. The effect of sample preparation procedures on extraction efficiency was also examined.

Combined use of temperature and solvent strength in reversed-phase gradient elution. I. Predicting separation as a function of temperature and gradient conditions.

It has been shown previously that computer simulation based on two initial experiments can predict separation in reversed-phase gradient elution as a function of gradient conditions (gradient steepness, gradient range and gradient shape) and column conditions (column length, flow-rate and particle size). The present study extends this capability for changes in temperature. Four initial experiments (two different gradient times, two different temperatures) provide input data that allow predictions of separation as a function of temperature as well as gradient and column conditions. A semi-empirical relationship, tR = a + bT, is able to relate gradient retention time tR to column temperature T (other conditions constant). The accuracy of this approach has been evaluated for 102 solutes and a variety of experimental conditions, including the use of five different HPLC instruments (four different models).