To elucidate the intracellular signaling pathways for VLDL-induced VLDLR transcription, Western blot analysis was used to examine phosphorylated ERK1/2 protein. It was found that that VLDL induced an increase in ERK1/2 activity in a protein kinase C (PKC)-dependent manner in murine RAW264. 7 macrophages. By using different protein kinases inhibitors or activators it was observed that the effect of VLDL-induced VLDL receptor transcription, which is monitored by RT-PCR analysis of VLDL receptor mRNA, was not affected by the inhibitor of p38 kinase and cAMP analog, but completely abolished by pretreatment of the cells with PD 98059, an inhibitor of MEK and GF 109203X, an inhibitor of PKC. These results demonstrated that the PKC/ERK1/2 cascade is the essential signaling pathway by which VLDL activates VLDL receptor mRNA expression.

To observe the inhibitory effects of an antisense u-PAR vector on invasion of highly invasive PC-3M cell subclones, the effects of the antisense u-PAR on activity of MMP-9 in those highly invasive cell subclones were detected by a quantitative RT-PCR and zymography. The monolayer invasion assay and colony formation assay in soft agar were used. And tumorigenesis rate and invasions by the cell subclones with or without the antisense u-PAR were observed in nude mice. It was found thatin vitro growth of highly invasive PC-3M cell subclones transfected with the antisense u-PAR was declined, and the ability of anchorage-independent growth of those cell subclones was found decreased sharply, with the inhibiting rate becoming 79% and 60%, respectively. Although the antisense u-PAR didn’t change MMP-9 gene transcription, they could inhibit the activation of MMP-9 of highly invasive PC-3M cell subclones. Moreover, the tumorigenesis rate of the cell subclones with the antisense u-PAR decreased and the growth of a neoplasm also slowed down. Thet tests showed the difference between experimental and control groups was statistically significant (P<0.01). The antisense u-PAR vector could not only inhibit the invasion ability of highly invasive PC-3M cell subclonesin vitro but also restrain the growth of those cell subclonesin vivo.

In order to testify the antitumor effect, especially its effect against liver carcinomain vivo, of VP3 protein, one kind of protein coded by chicken anemia virus, recombinants pcDNA-vp3 containing chicken anemia virus vp3 gene, and control vector pcDNA3 were mixed with murine liver carcinoma cell lines H22 respectively. The mixture was injected subcutaneously into Balb/C mice. Some days later, the mice were killed and the solid tumor weighed. The antitumor efficiency was evaluated. The manners of VP3 proteinin vivo, inducing tumor cell death were identified by using TUNEL assay. All the results suggested that the injection of pcDNA-vp3 and H22 mixture resulted in a significant reduction of tumor growth in mice when compared with the results of control groups. TUNEL assay revealed that VP3 induced apoptosisin vivo. All these indicated that CAV vp3 might be a potential new gene in reducing the growth rate of tumor cells in liver carcinoma or in other kind of solid tumorsin vivo.

To observe the effect of multiple electroacupuncture (EA) on the pain threshold and the regulation of N-methyl-D-aspartate (NMDA) receptor in dorsal root ganglia (DRG) of neuropathic pain rats. Rats were prepared with a unilateral chronic constriction injury (CCI) to the sciatic nerve. EA was done in acupoints “Huan Tiao” and “Yang Ling Quan” for 30 min every day and the thermal thresholds were detected after EA at 3, 5, 7, 10, 14 days after operation. On day 14 after nerve injury, the in situ hybridization method was used to investigate the change of NMDA R1 mRNA in L4–L5 DRG. The thermal threshold reduced significantly from day 3 after operation in CCI rats. After multiple EA treatment, the ipsilateral thermal hyperalgesia relieved gradually and the thermal threshold had no difference with control side after day 5 (P>0.05). From Day 7 after operation, the thermal threshold at each time point were significantly different compared with CCI group respectively (P>0.05). Moreover the EA had accumulative effect. On Day 14 after operation, the NMDAR1 mRNA positive neurons and the mean optic density in ipsilateral L4–5, DRG were less than that of control side (P<0.05), mainly in medium and small neurons. After EA treatment, the NMDAR1 mRNA positive neurons in ipsilateral DRG had no considerable difference comparing with those of control side, significantly increased comparing with CCI group (P<0.05). It’s concluded that the NMDA receptors in DRG relate closely with the generation and development of neuropathic pain. The multiple EA treatment can attenuate the thermal hyperlgesia of neuropathic pain rats and regulate the NMDA receptor.

To explore the anti-apoptotic role of electroacupuncture (EA) and its molecular mechanisms after cerebral ischemia/reperfusion (IR) of rats, by using animal model of middle cerebral artery occlusion (MCA), the changes of the cleavage of PARP were observed by Western blot and the mRNA of heat shock protein (Hsp) 70 and Hsp90ß detected by competitive RT-PCR after cerebral IR and EA treatment. The results were as follows: (1) The cleavage of PARP was increased in ischemic hippocampus, and EA treatment could attenuate the level of the cleavage remarkably: (2) The mRNA expression of Hsp70 was increased in the ischemic cortex and hippocampus and was futher increased after EA treatment: (3) The mRNA expression of Hsp90ß was decreased in ischemic cortex and hippocampus and the decrease was relatively slight after EA treatment. The above results demonstrated EA treatment could protect neurons from apoptosis after cerebral IR. One of the molecular mechanisms was the promotion of the inducible expression of Hsp70 and the improvement of the inhibition of the expression of Hsp90.

The effects of inhibitors of TNFα converting enzyme (TACE) on TNFα secretion were studied to develop an approach to interfere inflammation processes. The HL-60 cell lines were stimulatedin vitro with LPS intravenously for different time to establish the cellular model of inflammation and simultaneously inducein vivo inflammation animal model by LPS The cytotoxic effects of soluble TNFα were checked using MTT colorimetric method to determine the rate of cell proliferation. The level of expression of TACE was detected by using RT-PCR, FCM and immuno-histochemical technique respectively. It was found Chinese medicine Reduqing (RDQ) could inhibit the transcription of TNFα mRNA induced by LPS stimulation (P<0.01, compared with the control). The antioligodeoxyribonucleotide (anti-ODN) of TNFα mRNA could inhibit 78.96% of TNFα secretion. The mimic peptides of TACE substrates with hydroxamine group showed potencyin vivo andin vitro against converting of pro-TNFα. It was concluded that all the three types of TACE inhibitors can regulate the expression of TACE at different levels and inhibit sTNFα secretion, indicating TACE is a novel target for inflammation therapy.

To obeserve the effect of oxidized low density lipoprotein (OxLDL) on arterial endothelial cells apoptosisin vivo, we established a model in which Sprague-Dawley rats were given intraperitoneal and intravenous injection of unmodified LDL (8 mg/kg every day) via the tail vein. Seven days after the injection, the aortic endothelial cells specimens were prepared by anen face preparation of rat aorta. The apoptotic cells were identified and counted byin situ nick and labelling (TUNEL) method and light microscopy. The numbers of the apoptotic cells were 12.52±4.71/field in the intraperitoneal injection control group, 11.41±2.94/field in the intravenous injection control group, 22.98±8.01/field in the intraperitoneal injection LDL group and 103.8±11.5/field in the intravenous injection LDL group, respectively. The difference was significant between injection LDL group and control (P<0.01), and the difference was also significant between two LDL injection groups (P<0.01). These findings suggest that injection of LDL can induce apoptosis in arterial endothelial cells and the effect is especially significant with intravenous injection LDL. After injection, oxidative modification of LDL may occur in local arteries and causes injury to the endothelial cells.

To understand the expression and effect of mismatch repair genes, hMSH2 and hMLH1, and to investigate anti-leukemic cell proliferation mechanism of curcumin, the levels of both genes were detected by multiple comparative RT-PCR. The protein of hMSH2 was determined by flow cytometry (FCM) and the gene mutation of hMSH2 and hMLH1 were detected by PCR-SSCP and microsatellite instability assay. After UV irradiation, the gene expression of hMSH2 and hMLH1 was not increased and showed no response. This phenomenon was not ascribed to gene mutation, because PCP-SSCP and microsatellite instability assay revealed no abnormal gel-shift band in both genes. After irradiation and addition of curcumin, the expression of hMSH2 mRNA increased and the cellular apoptotic rate also increased at the same time. The difference was statistically significant as compared with groups without addition of curcumin and control groups (P<0.05). Our results suggested that when MMR system was inhibited by the same agents, curcumin can remove this suppression and switch to cellular apoptosis.

The effects of carvedilol on cardiomyocyte apoptosis and expression of bcl-2, bax genes following ischemia (0.5 h) and reperfusion (48 h)in vivo and the possible biological mechanism of carvedilol inhibiting cardiomyocyte apoptosis were studied. The left anterior descending artery in Wistar rats were ligated to establish ischemia-reperfusion (I/R) models. The model animals were divided into two groups: I/R group, the model rats not subject to other treatments except ischemia-reperfusion (n=8): carvedilol-treated group (n=8), I/R model rats treated with carvedilol. Eight rats in the sham-operated group were subjected to only experimental open operation. The number of apoptotic cardiomyocyte was determined by TUNEL staining. Immunohistochemistry andin situ hybridization histochemistry (ISHH) were used to detect the expression of bcl-2 and bax genes. Image processing system was used to quantitatively dispose the positive metric substances of both immunohistochemistry and ISHH through the average optical density (OD) value. The results showed that the number of the apoptotic cells were 36.18±9 (I/R group), 0–1 (sham-operated group) and 9.5±3 (carvedilol-treated group) in each visual field respectively with the difference being very significant among the groups (P<0.001). The OD values of bcl-2 protein in sham-operated group, I/R group and carvedilol-treated group were 0.14±0.01, 0.08±0.02 and 0.15±0.03, respectively. The OD values of bcl-2 mRNA in sham-operated group, I/R group and carvedilol-treated group were 0.08±0.01, 0.06±0.01 and 0.09±0.01, respectively. There was no significant difference between carvedilol-treated group and I/R group (P>0.05). The OD values of bax protein in I/R group, sham-operated and carvedilol-treated-treated group were 0.13±0.02, 0.07±0.01, 0.06±0.01, respectively. There was very significant difference between carvedilol-treated group and I/R group (P<0.01). Bcl-2/bax ratio was 1.07±0.14 (I/R group), 1.28±0.16 (sham-operated group), 2.5±0.26 (carvedilol-treated group) respectively with the difference being very significant between carvedilol-treated group and I/R group (P<0.05). It was indicated that carvedilol could inhibit cardiomyocyte apoptosis following ischemia and reperfusion, which was related to the increased bcl-2/bax ratio due to inhibition of bax gene expression.

In order to investigate the association of fibrin monomer polymerization function (FMPF) with traditional cerebrovascular risk factors and ischemic cerebrovascular disease in old people, 1∶1 paired case-control comparative study was performed for FMPF and traditional cerebrovascular risk factors on 110 cases of old ischemic cerebrovascular disease and 110 controls matched on age, sex and living condition. The results showed that cerebrovascular risk factors were more prevalent in case group than in control group. In the case group, FMPF was significantly higher than in control group. There was a significant positive correlation between hypertension and fibrin monomer polymerization velocity (FMPV), hypertension and fibrinogen (Fbg), alcohol consumption and Fbg, but no significant correlation between diabetic mellitus, smoking and FMPF was found. Among the parameters of blood lipids, there were significant positive correlations between total cholesterol (TC) and parameters of FMPF to varying degrees, triglycerides (TG) and FMPV, TG and Fbg. Our results also showed there were significant linear trends between TC and FMPV (P<0.001), TC and Fbg (P=0.0087), TG and FMPV/Amax (maximum absorbance) (P=0.0143) respectively. Multiple logistic regression analysis revealed that FMPF in case group remained significantly higher than control group after adjustment of all risk factors that were significant in univariate analysis. It was concluded that there is a possible pathophysiological link between FMPF and cerebrovascular risk factors. An elevated FMPF is associated with ischemic cerebrovascular disease and an independent risk factor of this disease. In old people, detection of FMPF might be a useful screening to identify individuals at increased cerebrothrombotic risk.

The relationship between 3 polymorphisms sites [interleulin-4 (IL-4), IL-4 receptor (IL-4R) α chain and activation-induced cytidine deaminase (AICDA)] and adult allergic asthma in China was studied. By using case-control method, DNA and clinical data were obtained from allergic asthmatic patients and compared with those in the control subjects. The subjects were genotyped for the IL-4 C-589T promoter polymorphism, the IL-4R α chain Q576R and the AICDA C8408T by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The results showed that the IL-4 C-589T was not associated with adult allergic asthma in China. However, the IL-4R α chain 576R/R and AICDA 8408T/T frequency was significantly increased in allergic asthma group as compared with that in the control group [odd ratio (OR)=3.797 and 9.127, respectively;P<0.01)] and was correlated with the increased plasma total IgE. These data suggested that the IL-4R α chain 576R/R and AICDA 8408T/T genotypes confer genetic susceptibility to adult allergic asthma in China.

To study the expression of peroxisome proliferator-activated receptor-γ(PPAR-γ) in lung cancer cells, and to testify if the PPAR-γ agonists can inhibit human lung cancer cell growth through induction of apoptosis, PPAR-γ was detected in two lung cancer cell lines by RT-PCR and immunohistochemistry, the inhibition of human lung cancer cell growth was investigated by MTT and cell counts, and the apoptosis was assessed by TUNEL. The results showed that: (1) PPAR-γ expressed on two lung cancer cell lines; (2) PPAR-γ activated by ligands could inhibit human lung cancer cell growth remarkably; (3) PPAR-γ agonists could induce apoptosis to inhibit lung cancer cell growth. It was concluded that PPAR-γ expressed in lung cancer cell can be activated by ligands and can inhibit lung cancer cell growth through induction of apoptosis.

In order to investigate the K⁺ channels and their effects on resting membrane potential (Em) and excitability in rat bronchial smooth muscle cells (BSMCs), the components of outward K⁺ channel currents and the effects of K⁺ channels on Em and tension in rat bronchial smooth muscle were observed by using standard whole-cell recording of patch clamp and isometric tension recording techniques. The results showed that under resting conditions, total outward K⁺ channel currents in freshly isolated BSMCs were unaffected by ATP-sensitive K⁺ channel blocker. There were two types of K⁺ currents: voltage-dependent delayed rectifier K⁺ channel (K_v) and large conductance calcium-activated K⁺ channel (BK_Ca) currents. 1 mmol/L 4-aminopyridine (4-AP, an, inhibitor of K_V) caused a significant depolarization (from −8.7±5.9 mV to −25.4±3.1 mV,n=18,P<0.001). In contrast, 1 mmol/L tetraethylammonium (TEA, an inhibitor of BK_Ca) had no significant effect on Em (from −37.6±4.8 mV to −36.8±4.1 mV,n=12,P>0.05). 4-AP caused a concentration-dependent contraction in resting bronchial strips. TEA had no effect on resting tension, but application of 5 mmol/L TEA resulted in a left shift with bigger pD₂ (the negative logarithm of the drug concentration causing 50% of maximal effect) (from 6.27±0.38 to 6.89±0.54,n=10,P<0.05) in the concentration-effect curve of endothine-1, and a right shift with smaller pD₂ (from 8.10±0.23 to 7.69±0.08,n=10,P<0.05) in the concentration-effect curve of isoprenaline. It was suggested that in rat BSMCs there may be two types of K⁺ channels, K_v and BK_Ca, which serve distinct roles. K_v participates in the control of resting Em and tension. BK_Ca is involved in the regulation of relaxation or contraction associated with excitation.

To investigate the distribution of possible novel mutations from parkin gene in variant subset of patients with Parkinson’s disease (PD) in China and explore whether parkin gene plays an important role in the pathogenesis of PD, 70 patients were divided into early-onset group and late-onset group; 70 healthy subjects were included as controls. Genomic DNA from 70 normal controls and from those of PD patients were extracted from peripheral blood leukocytes by using standard procedures. Mutations of parkin gene (exon 1–12) in all the subjects were screened by PCR-single strand conformation polymorphism (SSCP), and further sequencing was performed in the samples with abnormal SSCP results, in order to confirm the mutation and its location. A new missense mutation Gly284Arg in a patient and 3 abnormal bands in SSCP electrophoresis from samples of another 3 patients were found. All the DNA variants were sourced from the samples of the patients with early-onset PD. It was concluded that Parkin point mutation also partially contributes to the development of early-onset Parkinson’s disease in Chinese.

The protective effect of puerarin on the Parkinson disease (PD) mice with decreased estrogen level was investigated in order to develop a new potential medicine as a substitute for estrogen for preventing and treating PD. By using immunohistochemical method of avidinbiotin peroxidase complex (ABC), the distribution of the cells positive for tyrosine hydroxylase (TH) and fibres in the substantia nigra of the mouse were observed. These mice were divided into three groups randomly: group A, normal-female-mouse models; group B containing three subgroups, B1 (normal saline), B2 (estrogen), B3 (puerarin); group C containing three sub groups, C1 (normal saline), C2 (estrogen), C3 (puerarin). By using TUNEL the numbers of apoptosis cells in every visual field was counted and the difference between the experimental group and control group was compared. The results showed the numbers of the cells positive for TH were more and the numbers of apoptosis cells were less in the normal-female-mouse models group than in the group of model made, after ovariosteresis and the group of model made before ovariosteresis (P<0.05), respectively. However, there was no significant difference, between the group given estrogen/puerarin and the controls, and between the group given estrogen and given puerarin. (P>0.05). It was suggested that puerarin may have protective effect on the nigral neurons to PD. Moreover, the protective effect might serve as a surrogate of estrogen and be associated with the apoptosis.

In order to observe neuronal toxical effect of Levodopa and investigate if using Levodopa together with Ginkgo Bilobar Extract (EGb) would be an workable method to treat Parkinson disease, rat models of Parkinson disease (PD) were made by injecting 6-OHDA stereotaxically to right side of the mesencephic ventral tegmental area (VTA) and substantia nigra pars compacta (SNc). Rotational behavioral observation, TUNEL, immunocytochemistry, Nissl’s body staining were performed to measure the difference between group treated by Levodopa (50 mg/kg every day for 3 days, 5 days, 7 days, L-dopa group) and group treated by Levodopa combined with EGb (100 mg/kg every day, E-D group). The results showed that in the L-dopa group, the numbers of apoptosis of substantial nigra, rings of rotational behavior were more than those in the E-D group (P<0.05). The numbers of Nissl’s cells in L-dopa group were fewer than in E-D group (P<0.05). The results suggested that Levodopa had neur toxic effect and EGb may decrease the toxicity of levodopa. The combined use of EGb with Levodopa may be a workable method to treat PD and may be better than using Levodopa alone.

The feasibility of using cord blood mesenchymal stem/progenitor cells (CB-MSPCs) to regenerate cardiomyocytes and the optimal inducing conditions were investigated. The CB mononuclear cells were cultured in low serum DMEM medium to produce an adherent layer. After expansion, the adherent cells were added into cardiomyocyte inducing medium supplemented with 5-azacytidine. Cardiogenic specific contractile protein troponin T staining was performed to identify the cardiomyocyte-like cells. The results showed that the frequency of CB-MSPCs clones in CB mononuclear cells was 0.5×10⁻⁶ and about 1.3×10⁷-fold expansion was achieved within 20 sub-cultivation. After cardiogenic induction, 70% CB-MSPCs was differentiated into cardiomyocyte-like cells. It was indicated that low serum culture could expand CB-MSPCs extensively and the expanded CB-MSPCs could be induced to differentiate into cardiomyocyte-like cells in high efficiency.

To determine the role of NF-kappaB in ischemia reperfusion (I/R) injury of the rat liver, rats underwent partial hepatic ischemia and reperfusion. The left and median lobes of the liver were subjected to ischemia for 90 min followed by reperfusion for defined times. NF-kappaB activity was analyzed by electrophoretic mobility shift assay (EMSA). Semiquantitative reverse-transcriptase polymerase chain reaction was used to analyze TNF-α and ICAM-1 mRNA levels. Results showed during liver I/R injury, NF-kappaB activation was induced in a time dependent manner. NF-kappaB was activated within 1 h and 2 h after the initiation of reperfusion and decreased after 4 h. Messenger RNA expression of TNF-α and ICAM-1 were increased after the reperfusion of 2 h. It was concluded that during hepatic I/R injury, NF-kappaB was activated and could bind to special sequence in the promoters of budget genes, which can up-regulate the expression of TNF-α and ICAM-1 mRNA to result in ischemia reperfusion injury of the rat liver.

The anti-tumor effect and mechanism of the staphylococcal enterotoxin A (SEA) were studied. The mouse gastric tumor model was produced by subcutaneously inoculating gastric tumor cells (MGC80-3). The experimental group was treated with SEA, and the control group was treated with normal saline. The percentage of tumor generation and tumor mass was measured. The results showed that the percentage of the tumor generation in the SEA-treated mice was lower than in the control group, but there was no significant difference (P>0.05). However, the tumor mass in the experimental group was significantly lighter than in the control group, with the difference being very significant (P<0.001). There were more CD₄⁺ T cells and CD_s⁺ T cells in the tumor of the mice treated with SEA than those of the control group. SEA has an obvious anti-tumor effect on mice gastric tumor. The mechanism might be that SEA induces the effect of superantigen-dependent cell mediated cytotoxicity to the tumor cells.

To construct the antisense transforming growth factorβ1 (TGFβ1) gene and investigate the effect of TGFβ1 autocrine loop blockage on the proliferation of osteosarcoma cells. TGFβ1 cDNA was cloned by RT-PCR from human osteosarcoma cells (MG-63) and inserted into pcDNA₃ to construct an antisense expression vector, which was dubbed pcDNA₃-TGFβ1(−). MTT was used to detect the proliferation of osteosarcoma cells transfected by antisense TGFβ1 gene. Our results showed that the proliferation of the transfected osteosarcoma cells was suppressed markedly. It is concluded that TGFβ1 autocrine loop blockage in osteosarcoma cells could inhibit cell proliferation, which might be helpful for gene therapy of osteosarcoma.

In order to study the effect of tanshinone II_A on growth and apoptosis in human hepatoma cell line BEL-7402in vitro, the human hepatoma cell line BEL-7402 was treated with tanshione I_A at various concentrations for 72 h. Growth suppression was evaluated by MTT assay; apoptosis-related alterations in morphology and biochemistry were ascertained under cytochemical staining (Hoechst 33258), transmission electron microscopy (TEM), and DNA agarose gel electrophoresis. Apoptotic rate was quantified by flow cytometry (FCM). The results showed that Tanshinone II_A could inhibit the growth of hepatoma cells in a dose-dependent manner, with IC₅₀ value being 6.28 μg/ml. After treatment with 1–10 μg/ml tanshione II_A for 72 h, BEL-7402 cells apoptosis with nuclear chromatin condensation and fragmentation as well as cell shrinkage and the formation of apoptotic bodies were observed. DNA ladder could be demonstrated on DNA electrophoresis. FCM analysis showed hypodiploid peaks on histogram, and the apoptotic rates at 5 μg/ml concentration for 12 h, 24 h, 36 h, 48 h and 72 h were (2.32±0.16)%, (3.01±0.35)%, (3.87±0.43)%, (6.73±0.58)% and (20.85±1.74)% respectively, which were all significantly higher than those in the control group (1.07±0.13)%. It is concluded that Tanshione II_A could induce human hepatoma cell line BEL-7402 apoptosis, which may be related to the mechanism of growth inhibition.

To evaluate the therapeutic efficiency of combined use of p16-expressing adenovirus and chemotherapeutic agents CDDP or As₂O₃ on human bladder cancer cell line EJ, the human bladder cancer cell line EJ were transfected with adenovirus-mediated p16 gene (Ad-p16), with administration of cisplatin (CDDP) or arsenic trioxide (As₂O₃). The cell growth, morphological changes, cell cycle, apoptosis and molecular changes were measured using cell counting, reverse microscopy, flow cytometry, cloning formation, immunocytochemical assays andin vivo therapy experiments to evaluate the therapeutic efficacy of such combined regimen. Ad-p16 transfer and CDDP or As₂O₃ administration to EJ cells could exert substantially stronger therapeutic effects than the single agent treatment. Especially inin vivo experiments, combined administration of p16 and CDDP or As₂O₃ induced almost tumor diminish compared to the partial tumor diminish induced by single agent. Moreover, delivery of Ad-p16, or administration of minimal-dose CDDP or As₂O₃ or combined regimen could induce massive apoptosis of EJ cell. Cell cycle analysis demonstrated that administration of CDDP or As₂O₃ remarkably arrested EJ cell in G₁ prior to apoptotic cell death. When treated with combined regimen, cells were arrested in G₁ to a greater extent prior to apoptotic cell death. It is concluded that after introduction into EJ cell, Ad-p16 shows enhanced therapeutic efficacy for EJ cell when used in combination with CDDP or As₂O₃.

To study the expression of mTR gene in the testis of SD rats with varied ages and its significance,in situ hybridization (ISH) techniques were applied to detect the expression of telomerase gene mTR mRNA in the testis of SD rats. The expression of mTR was found in testes of differentage male SD rats. There was a positive correlation between the expression of mTR and the location of germ cells (spermatogonia, spermatocyte, spermatid). In Setoli cells, leydig cell and spermatozoa, no telomerase mTR was detectable. Type A spermatogonia expressed the highest level of telomerase mTR mRNA. It was suggested that the expression of mTR gene in the testis of SD rats is of lifetime and coincides with the telomerase activity.

To explore the causes of the postoperative complications of the penile elongation and the measures to prevent them in order to raise the success rate of the penile elongation. 1 000 patients who had received the penile elongation were reviewed and analyzed for the causes of postoperative complications, and the measures of prevention and treatment were discussed. Our results showed that, of the 1 000 cases, 64 had the postoperative complications, including 20 cases of edema of prepuce, 15 cases of flap necrosis, 12 hematoma, 9 infections, and 8 cases of fat and clumsy penis. It is concluded that correct operative manipulation, strict aseptic measures and necessary postoperative care and management could avoid or reduce the postoperative complications. When complications happened, a satisfactory result can be achieved with timely and correct treatment in the majority of the patients.

To compare the cardiotoxicity induced by ropivacaine and bupivacaine and to investigate the mechanism of cardiotoxicity, 24 mature New Zealand rabbits were divided randomly into control group (group C), ropivacaine group (group R) and bupivacaine group (group B). Hearts were drawn out rapidly from the anesthetized animals and cardiac perfusion was performed immediately. Ropivacaine 500 ng/ml (group R) or bupivacaine 500 ng/ml (group B) was added to the perfusion solution. Ventricular myocardial ATP, ADP and AMP were measured with high performance liquid chromatogram. The ability of myocardial mitochondria oxidation to pyruvate or palmitoylcarnitine was detected with Clark electrode. Our results showed that myocardial ATP and ADP decreased significantly (P<0.05) in group R and most significantly (P<0.01) in group B as compared with group C. Myocardial ATP and ADP decreased most significantly (P<0.01) in group B as compared with group R. The changes of myocardial AMP revealed significant difference among three groups. The changes of pyruvate oxidation exibited no significant difference among the three groups. Palmitoylcarnitine oxidation decreased markedly (P<0.05) in group R and most significantly (P<0.01) in group B as compared with group C. The present study indicated that the inhibition of lipid substrate oxidation may be responsible for the cardiotoxicity induced by bupivacaine and ropivacaine. The cardiotoxicity induced by ropivacaine is far more less than bupivacaine.

The utility of three-dimensional spoiled gradient recalled acquisition in steady state (3D-SPGR) imaging in the cerebral diseases was evaluated and 3D-SPGR after enhancement in depicting contrast enhancement of all lesions and 2D-SE T1WI comparatively analyzed. 117 patients were subjected to MRI by a GE 1. 5T MR system. After performance of axial T1WI and T2WI in all patients, MRA (3D-MOTSA) images were acquired in 6 cases (8 lesions) of aneurysms. After enhancement, 3D-SPGR images were obtained in all the remaining patients. Quality parameters (SNR, C and CNR) were calculated on enhanced 2D-SE T1WI and 3D-SPGR images. And a four-point scale was used to measure the signal intensity of the main lesions on both sequences, then statistical analysis of the average score was performed with “t” test. Except for aneurysms, 2D-SE T1WI detected 134 lesions and 3D-SPGR disclosed 147 lesions. It was found that there was no statistically significant difference between the two average scores as determined by the “t” test (t=1.894,P>0.05). The enhancement degree of the main lesion was equivalent on 3D-SPGR and 2D-SE T1WI. Quality parameters (SNR, C and CNR) on 2D-SE T1WI were much larger than that of 3D-SPGR, increasing by an average of 57%, 20% and 97% respectively. 3D-SPGR imaging with MPR could clearly depict vascularity related to neoplasms in 20 cases and demonstrate shifted, deformed and blocked vessels involved by tumors. Six cases of large aneurysms (8 lesions) were visualized more clearly on 3D-SPGR than MRA (3D-MOTSA): 3D-SPGR could display aneurysm necks and differentiate thrombosed portion from the patent lumen, and disclose relationship of aneurysm to surrounding structures. It was concluded that enhanced 3D-SPGR played an important role in the depiction of the cerebral lesions and was superior to 2D-SE T1WI in many aspects.

To study the expression of placental isoferritin (PLF) in placental tissue of pregnancy-induced hypertension (PIH) and the relationship between the level of expression of PLF and the amount of vascular cell adhesion molecule-1 (VCAM-1) in serum, immunohistochemical technique was used to detect the expression of PLF in placenta tissue in 45 PIH patients (PIH group) and 15 normal pregnant women (normal group). High resolution pathological image analysis system (HPIAS-100) was employed to determine the quantity of PLF. The VCAM-1 in serum was examined by enzyme linked immunoabsorbent assay (ELISA). The results showed that the levels of PLF expressions in moderate and severe PIH patients were significantly lower than that of normal group (P<0.01). The serum VCAM-1 was significantly decreased in PIH group (1310±177 ρ/ng/ml) than that of normal group (609±72 ρ/ng/ml,P<0.01). The significant negative correlation existed between the expression of PLF in placental tissue and the serum VACM-1 (r=0.58,P<0.01). It was concluded that the level of PLF expression in PIH decreases and is negatively correlated with the mount of serum VCAM-1, indicating that these may be involved in the pathogenesis of PIH.

In order to evaluate the effects of sex chromosomal mosaicism on the accuracy of single-cell gender diagnosis, sex chromosomes of 21 normal fertilized embryos were detected by dual color fluorescentin-situ hybridization (FISH). The results showed that 4 embryos had sex chromosomal mosaicism (19%) and the remaining 17 showed uniformly XX or XY signals in all blastomeres. In conclusion, identification of sex by dual color FISH analysis of a single cell was accurate and efficient, and sex chromosomal mosaicism would not affect preimplantation gender diagnosis.

In order to study the angiogenesis in endometriosis, the samples of eutopic and ectopic endometria from patients with endometriosis were quantitatively analyzed by color morphometric image system (CMIS) for vascular surface area, and by examining endometrial blood vessel for microvessel density (MVD). The results showed that within each menstrual phase the vascular surface area and MVD were significantly higher in ectopic endometria with endometriosis than those in eutopic endometria with endometriosis or normal endometrium (P<0.05). It is concluded that angiogenesis might be involved in the development of endometriosis.

The value of ELISA, N-PCR and RT-PCR in clinical practice for pregnant women with HCMV infection was investigated. 5581 pregnant women were screened by ELISA. Among them, 100 cases were positive for IgM (group 1), 69 for both IgM and serous DNA (group 2) and 69 for both IgM and mRNA (group 3). The infectious status, maternal-fetal transmission and pregnancy outcome were monitored. It was demonstrated that the accordance rate of group 3 and group 2 with group 1 was 56.25% and 43.75%, respectively. The maternal-fetal transmission are in the group 1, 2 and 3 was 19.00%, 40.58% and 46.15%, respectively, with a significant difference found between group 2, 3 and group 1 (P<0.01). Incidence of spontaneous abortion, fetal death, fetal abnormality and neonatal death in group 1, 2 and 3 was 10.00%, 15.94% and 30.77%, respectively, and that of group 3, 2 was 4 and 2 times as much as that of group 1, respectively (OR=4. 00,P<0.001; OR=2.343,P<005, respectively). It was concluded that HCMV-IgM(+) can only be considered as an screening indicator for pregnant women with HCMV infection, while IgM(+) combined with serous DNA(+) or mRNA(+) indicates active infection and has a high incidence of maternal-fetal transmission and abnormal pregnancy outcome.

To evaluate the changes of 3′, 5′-cyclic adenosine monophosphate (cAMP), thromboxane A₂(TXA₂) and prostacyclin (PGI₂) in cerebrospinal fluid (CSF) in the asphyxiated newborn and explore their roles in hypoxic-ischamic brain damage (HIBD). Thirty-six full term newborns were divided into 3 groups, including 12 with moderate-severe hypoxic-ischaemic encephalopathy (HIE), 13 with mild HIE, 11 without HIE (control group). The levels of cAMP, TXB₂ (TXA₂ metabolite) and 6-keto-PGF_1α (PGI₂ metabolite) in CSF and plasma were measured 36–72 h after birth by RIA, and the concentrations were expressed as nM/L (cAMP), ng/L(TXB₂ and 6-keto-PGF_1α). The infants were followed-up at 6 and 12 month of age and Mental Development Index (MDI) and Psychomotor Development Index (PDI) were measured using Bayley Scales of Infant Development (BSID). The CSF cAMP level in moderate-severe HIE group was 8.60±2.40, significantly lower than that of the mild HIE group (14.83±2.84) and the control group (24.43±2.39) (for bothP<0.01). The levels of TXB₂ and 6-keto-PGF_1α in CFS in the moderate-severe HIE group (206.06±29.74, 168.47±23.02, respectively) were significantly higher than in the mild HIE group (83.37±28.57, 131.42±16.57, respectively,P<0.01) and the control group (41.77±21.58, 86.23±13.05, respectively,P<0.01). The level changes of cAMP, TXB₂ and 6-keto-PGF_1α in plasma in all groups were similar to those in CSF, but no significant difference was found between mild HIE group and the control group (P>0.05). The follow-up results showed that MDI and PDI of the moderate-severe HIE group were the lowest (84.79±13.34, 83.50±13.28, respectively), followed by mild HIE group (102.19±7.02, 99.94±9.08, respectively), with the control group being the highest (116.63±12.08, 116.69±10.87, respectively). Univariate analysis showed some significant difference (the moderate-severe HIE group vs. the mild HIE group or the control group,P<0.01; the mild HIE group vs. the control groupP<0.05). The results suggested that the concentration of cAMP, TXA₂ and T/K ratio in CSF after neonatal asphyxia might be sensitive markers in evaluating the severity of brain damage in early stage and predicting the future outcome.

To investigate the effects of the free radical, 1,2-Diphenyl-2-picylhydrazyl, on cochlear blood flow, 20 guinea pigs were divided into 3 groups at random, 6 for control group, 6 for 1 mmol/L group and 8 for 0.1 mmol/L group. 2 μl vehicle or drugs were dropped into round window membrane (RWM). Cochlear microcirculation was monitored by laser Doppler flowmeter (LDF), and mean arterial blood flow (MABP), which was transferred by pressure conductor sensor and preamplifier, was simultaneously recorded on the computer. Our results showed that MABP was stable throughout the experiment. Cochlear blood flow (CBF) increased by 10.32% (P <0.05) in 1 mmol/L group, and decreased by 4.89% in 0.1 mmol/L group (P<0.05). In control group cochlear microcirculation showed no significant changes. It is concluded that DPPH exerted effects on cochlear microcirculation.

In order to know the effects of caloric stimulation on neuronal firing in medial vestibular nuclei (MVN) by middle ear irrigation, the middle ear was irrigated with ice (4°C), hot (44°C), and warm (37°C) water, and the firing rate of MVN neuron was extracellularly recorded. The results showed that the firing rate of MVN neuron was changed by caloric stimulation, and the majority of MVN neurons showed excitation by irrigation with hot water and inhibition by ice water (type A). The neuronal firing was recovered immediately after the cessation of the stimulation. I It was concluded that the neuronal firing rate in MVN was changed by caloric stimulation in middle ear cavity. The response was different in various neurons.

In order to investigate the susceptibility of mixed infection ofUreaplasma Urealyticum (UU) andMycoplasma Hominis (MH) to 7 kinds of antimicrobial agents and comparison with that of UU infection in NGU patients, thein vitro susceptibility was determined by using microdilution method. The positive results were analyzed. The results showed that the sequence of susceptibility to 7 kinds of antimicrobial agents for both UU infection group and UU-MH mixed infection group was almost the same from the highest susceptibility to the lowest accordingly: Josamycin, Doxycycline, Minocycline, Sparfloxacin, Roxithromycin, Ofloxacin and Azithromycin. The total drug resistance rate for UU-MH mixed infection group (97.67%) was significantly higher than that for UU infection group (44.67%,P<0.01). The highest drug resistance rate in UU group and UU-MH mixed infection group was 31.33% (Ofloxacin) and 90.48% (Azithromycin) respectively. UU-MH mixed infection showed an increased drug resistance and changes of drug resistance spectrum.

The anti-endotoxic effect of syringic acid (SA) isolated fromRadix Isatidis (Banlangen, BLG) was studied. SA was extracted and isolated from BLG and diluted into 1% solution. The content of SA-pretreated endotoxin (ET) was quantitatively determined using Limulus test. The ability of fever induction of ET pretreated with SA was measured using endotoxin-induced fever test in rabbits. The LPS-induced death in mice pretreated with and without SA was compared. Results showed that after pretreatment with SA, 83.16% of ET was destroyed, the ET-induced fever in rabbits relieved markedly and the LPS-induced death rate in mice dropped from 68 % to 20 %. It was concluded that SA isolated from BLG had anti-endotoxic effects.

In order to investigate the influence of radiation therapy after the treatment of maxillary implant-supported prostheses, 27 patients received a total of 131 implants in maxilla after oral cancer treatment and/or reconstructive surgery. Among them, 25 received maxillary implant-supported prostheses. The cumulative survival rates of implants and prostheses were evaluated by the productlimit-estimates method according to Kaplan-Meier. The cumulative survival rate of implants and prostheses in irradiated patients was compared with that in non-irradiated patients by statistical Logrank test. The results showed that 112 implants were observed after implant loading. The implants cumulative survival rate was approximately 65 % for overall patients. The cumulative prosthesis successful rate was approximately 88% for all 25 patients. Log-rank test analysis revealed that there was a significant difference in cumulative implants survival rates between non-irradiated and irradiated maxillary bone (P<0.01). It was concluded that the implants and prostheses in irradiated patients have significantly lower survival rates than in non-irradiated patients.