Early effects of spinal endoscopic fusion technology in the treatment of degenerative lumbar disease / 西安交通大学学报(医学版)

【Objective】 To report the early clinical effects and surgical complications of endoscopic spinal fusion technology （Endo-LIF） in the treatment of degenerative lumbar disease. 【Methods】 The clinical data of 31 patients with degenerative lumbar spine disease treated with Endo-LIF from June 2019 to May 2021 were retrospectively analyzed. All the 31 patients underwent endoscopic spinal fusion therapy. We recorded the operation time， hospital stay duration， postoperative complications， visual analogue scale for pain （VAS）， oswestry dysfunction Index （ODI） and low back pain in the Chinese Orthopaedic Association Spine Group Surgery scoring standards before operation， immediately after operation， and the last follow-up to evaluate clinical efficacy. 【Results】 The operation time of the 31 patients was （134.80±34.98） min， the intraoperative blood loss was （100.13±18.49） mL， the hospital stay was （6.65±0.17） days， and the follow-up time was 6 to 18 （14±2.3） months. One patient had hematoma compression after surgery； he had incision made immediately to clear the hematoma and healed after bed rest. Two patients developed spinal hypertension and healed after bed rest. All the patients had no symptoms of nerve injury after operation， and the clinical symptoms were significantly relieved. We compared the perioperative VAS score and ODI index of all the patients， which were lower immediately after operation and at the last follow-up than those before the operation （P<0.05）， and the difference was statistically significant. 【Conclusion】 Endo-LIF technology has good short-term clinical effects and the advantages of milder trauma， less blood loss， and quick recovery after surgery. It is a safe and minimally invasive lumbar fusion surgery.

Eukaryotic translation initiation factors as promising targets in cancer therapy.

The regulation of the translation of messenger RNA (mRNA) in eukaryotic cells is critical for gene expression, and occurs principally at the initiation phase which is mainly regulated by eukaryotic initiation factors (eIFs). eIFs are fundamental for the translation of mRNA and as such act as the primary targets of several signaling pathways to regulate gene expression. Mis-regulated mRNA expression is a common feature of tumorigenesis and the abnormal activity of eIF complexes triggered by upstream signaling pathways is detected in many tumors, leading to the selective translation of mRNA encoding proteins involved in tumorigenesis, metastasis, or resistance to anti-cancer drugs, and making eIFs a promising therapeutic target for various types of cancers. Here, we briefly outline our current understanding of the biology of eIFs, mainly focusing on the effects of several signaling pathways upon their functions and discuss their contributions to the initiation and progression of tumor growth. An overview of the progress in developing agents targeting the components of translation machinery for cancer treatment is also provided. Video abstract.

Body Movement Synchrony Predicts Degrees of Information Exchange in a Natural Conversation.

Human interaction has two principle functions: building and maintaining relationships with others and exchanging information. The function of building and maintaining relationships with others relates to interpersonal coordination; this behavior pattern is expected to predict the outcome of social relationships, such as between therapists and patients. It is unclear, however, whether the exchange of information is associated with interpersonal coordination. In the present study, we tested a hypothesis of whether body movement synchrony occurs in a natural conversation and whether this synchrony has a positive correlation with the degree of information exchange. Fifty participants were engaged in a conversation task; each had different roles in the conversation. We measured their body movements during this conversation using an optical motion capture system. Similar to methods that can be found in previous research, we calculated body movements and quantified their synchrony applying the methods previously reported that automatically quantified their body movements. Moreover, we determined the participants' degree of information exchange concerning the conversation using a questionnaire. We observed that the body movement synchrony of pairs who talked with each other was significantly higher than that of pairs who did not talk with each other, and that this synchrony was positively associated with the degree of information exchange. These results suggest that body movement synchrony predicted information exchange.

Effects of Human Synchronous Hand Movements in Eliciting a Sense of Agency and Ownership.

The self is built as an entity independent from the external world using the human ability to experience the senses of agency and ownership. Humans usually experience these senses during movement. Nevertheless, researchers recently reported that another person's synchronous mirror-symmetrical movements elicited both agency and ownership in research participants. However, it is unclear whether this elicitation was caused by the synchronicity or the mirror symmetry of the movements. To address this question, we investigated the effect of interpersonal synchronization on the self-reported sense of agency and ownership in two conditions, using movements with and without mirror symmetry. Participants performed rhythmic hand movements while viewing the experimenter's synchronous or random hand movements, and then reported their perceptions of agency and ownership in a questionnaire. We observed that agency and ownership were significantly elicited by the experimenter's synchronous hand movements in both conditions. The results suggested that the synchronous movements of another person-rather than mirror- or non-mirror-symmetrical movements-appear to elicit the experience of a sense of agency and ownership. The results also suggested that people could experience these senses not only from their own movements but also from another person's synchronous movements.

Voluntary movement affects simultaneous perception of auditory and tactile stimuli presented to a non-moving body part.

The simultaneous perception of multimodal sensory information has a crucial role for effective reactions to the external environment. Voluntary movements are known to occasionally affect simultaneous perception of auditory and tactile stimuli presented to the moving body part. However, little is known about spatial limits on the effect of voluntary movements on simultaneous perception, especially when tactile stimuli are presented to a non-moving body part. We examined the effect of voluntary movement on the simultaneous perception of auditory and tactile stimuli presented to the non-moving body part. We considered the possible mechanism using a temporal order judgement task under three experimental conditions: voluntary movement, where participants voluntarily moved their right index finger and judged the temporal order of auditory and tactile stimuli presented to their non-moving left index finger; passive movement; and no movement. During voluntary movement, the auditory stimulus needed to be presented before the tactile stimulus so that they were perceived as occurring simultaneously. This subjective simultaneity differed significantly from the passive movement and no movement conditions. This finding indicates that the effect of voluntary movement on simultaneous perception of auditory and tactile stimuli extends to the non-moving body part.

The simultaneous perception of auditory-tactile stimuli in voluntary movement.

The simultaneous perception of multimodal information in the environment during voluntary movement is very important for effective reactions to the environment. Previous studies have found that voluntary movement affects the simultaneous perception of auditory and tactile stimuli. However, the results of these experiments are not completely consistent, and the differences may be attributable to methodological differences in the previous studies. In this study, we investigated the effect of voluntary movement on the simultaneous perception of auditory and tactile stimuli using a temporal order judgment task with voluntary movement, involuntary movement, and no movement. To eliminate the potential effect of stimulus predictability and the effect of spatial information associated with large-scale movement in the previous studies, we randomized the interval between the start of movement and the first stimulus, and used small-scale movement. As a result, the point of subjective simultaneity (PSS) during voluntary movement shifted from the tactile stimulus being first during involuntary movement or no movement to the auditory stimulus being first. The just noticeable difference (JND), an indicator of temporal resolution, did not differ across the three conditions. These results indicate that voluntary movement itself affects the PSS in auditory-tactile simultaneous perception, but it does not influence the JND. In the discussion of these results, we suggest that simultaneous perception may be affected by the efference copy.

Structure of MST2 SARAH domain provides insights into its interaction with RAPL.

The STE20 kinases MST1 and MST2 are key players in mammalian Hippo pathway. The SARAH domains of MST1/2 act as a platform to mediate homodimerization and hetero-interaction with a range of adaptors including RASSFs and Salvador, which also possess SARAH domains. Here, we determined the crystal structure of human MST2 SARAH domain, which forms an antiparallel homodimeric coiled coil. Structural comparison indicates that SARAH domains of different proteins may utilize a shared dimerization module to form homodimer or heterodimer. Structure-guided mutational study identified specific interface residues critical for MST2 homodimerization. MST2 mutations disrupting its homodimerization also impaired its hetero-interaction with RAPL (also named RASSF5 and NORE1), which is mediated by their SARAH domains. Further biochemical and cellular assays indicated that SARAH domain-mediated homodimerization and hetero-interaction with RAPL are required for full activation of MST2 and therefore apoptotic functions in T cells.

[Protective effects of polysaccharides from Dendrobium huoshanense on CCl4-induced acute liver injury in mice].

OBJECTIVE: To study the protective effects of polysaccharides from Dendrobium huoshanense (DHP) against CCl4-induced liver injury in mice. METHOD: Eighty male Kunming mice were randomly divided into normal control group, model control group, dextran control group, starch control group, hydrolyzate control group, three different dose of DPH groups consisting of high-dosage group, middle-dosage group and low-dosage group (200, 100, 50 mg x kg(-1)). Each group contained ten mice. The mice were treated with DHP via intragastric administration for 15 days before treatment of 50% CCl4 in olive oil for consecutive two days. Both alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities in serum and superoxide dismutase (SOD) activities and malondialdehyde (MDA) contents in liver tissues were determined in all groups. Immunohistochemistry was used to detect the expression of TNF-alpha in hepatic tissue. Hepatic histopathological examination was observed. RESULT: DHP effectively decreased the activities of ALT and AST in serum and the contents of hepatic MDA, and restored hepatic SOD activities in acute liver injury mice. Liver tissue damage induced by CCl4 was ameliorated in mice with DHP administration through histopathology examination. Furthermore, the expression of TNF-alpha was greatly decreased in groups treated with polysaccharides. CONCLUSION: DHP has a significantly hepatoprotective effect on CCl4-induced acute liver injury in mice. Protective effect of DHP on the liver may be related to its function of scavenging free radicals and inhibiting lipid peroxidation and TNF-alpha expression.

Protective effects of polysaccharides from Dendrobium huoshanense on CCl4-induced acute liver injury in mice / 中国中药杂志

<p><b>OBJECTIVE</b>To study the protective effects of polysaccharides from Dendrobium huoshanense (DHP) against CCl4-induced liver injury in mice.</p><p><b>METHOD</b>Eighty male Kunming mice were randomly divided into normal control group, model control group, dextran control group, starch control group, hydrolyzate control group, three different dose of DPH groups consisting of high-dosage group, middle-dosage group and low-dosage group (200, 100, 50 mg x kg(-1)). Each group contained ten mice. The mice were treated with DHP via intragastric administration for 15 days before treatment of 50% CCl4 in olive oil for consecutive two days. Both alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities in serum and superoxide dismutase (SOD) activities and malondialdehyde (MDA) contents in liver tissues were determined in all groups. Immunohistochemistry was used to detect the expression of TNF-alpha in hepatic tissue. Hepatic histopathological examination was observed.</p><p><b>RESULT</b>DHP effectively decreased the activities of ALT and AST in serum and the contents of hepatic MDA, and restored hepatic SOD activities in acute liver injury mice. Liver tissue damage induced by CCl4 was ameliorated in mice with DHP administration through histopathology examination. Furthermore, the expression of TNF-alpha was greatly decreased in groups treated with polysaccharides.</p><p><b>CONCLUSION</b>DHP has a significantly hepatoprotective effect on CCl4-induced acute liver injury in mice. Protective effect of DHP on the liver may be related to its function of scavenging free radicals and inhibiting lipid peroxidation and TNF-alpha expression.</p>

delta-Aminolevulinic acid dehydratase activity, urinary delta-aminolevulinic acid concentration and zinc protoporphyrin level among people with low level of lead exposure.

To evaluate the relationship of delta-aminolevulinic acid dehydratase (ALAD) activity, urinary delta-aminolevulinic acid (ALAU) level and blood zinc protoporphyrin (ZPP) concentration to low blood lead (PbB) levels, these biomarkers were determined for all subjects enrolled from a rural area of southeast China where people had low levels of exposure to lead. The mean values of PbB, ALAD, ALAU and ZPP were 67.11 microg/L (SD: 1.654, range: 10.90-514.04), 339.66 nmol ml(-1)h(-1) (1.419, 78.33-793.13), 20.64 microg/L (1.603, 2.00-326.00), and 0.14 micromol/L (3.437, 0.01-2.26), respectively. ALAD was inversely associated with low levels of PbB. ZPP was inversely related to low levels of PbB but positively related to relatively higher levels of PbB. Alcohol drinking contributed to low ALAD in men. Women had higher ZPP than men. ALAU had no significant association with PbB. In conclusion, ALAD possibly has a non-linear relation with low to moderate levels of PbB. At moderate levels of PbB, ZPP increases with increasing levels of PbB. ALAU is not suitable as an indicator for low levels of lead exposure.

[The effect of focused ultrasound on the physicochemical properties of Sarcoma 180 cell membrane].

This study was amied to detect the changes in the cell membrane of Sarcoma 180 (S180) cells induced by focused ultrasound and to probe the underlying mechanism. The viability of tumor cells was examined at various intensities and different treatment times by ultrasound at the frequency of 2.2MHz. Flow cytometry and fluorescence microscopy were used to detect the loading of fluorescein isothiocyanate dextran (FD500) which signifies the change of membrane permeability. The results showed that after the cells were treated by ultrasound, especially when irradiated for 60s, the number of fluorescent cell, which represented the transient change of membrane permeabilization with cell survival, increased significantly. Then the damage of cell membrane was evaluated by the measurement of lactate dehydrogenase (LDH) release which became more severe as the radiation time was increasing. The generation of lipid peroxidation was estimated using the Thibabituric Acid (TBA) method after irradiation. The results reveal that the instant cell damage effects induced by ultrasound may be related to the improved membrane lipid peroxidation levels post-treatment. The physicochemical properties of S180 cell membrane were changed by focused ultrasound. The findings also imply an exposure time-dependent pattern and suggest that the lipid peroxidation produced by acoustic cavitation may play important roles in these actions.

Bioeffects of low-energy continuous ultrasound on isolated sarcoma 180 cells.

BACKGROUND: The aim of this study was to investigate the mechanism underlying bioeffects of low-intensity continuous ultrasound on isolated sarcoma 180 (S180) cells and cellular responses to these effects. METHODS: After sonication, several structural and functional parameters were examined to elucidate ultrasound-induced cell damage. RESULTS: Instant disruption of the cell membrane might be caused by acoustic cavitation, producing mechanical and chemical effects that acted simultaneously on S180 cells; this could be reflected by immediate (morphological) changes such as membrane permeability, membrane fluidity, lipid peroxidation and the generation of hydroxyl radicals in culture medium. Our results of the delayed effects also indicated S180 cells were sensitive to ultrasound-induced apoptosis, and the rate of apoptosis rose gradually with a prolonged incubation time. The presence of apoptotic cells was identified by a distinct morphological form characterized by membrane blebbing, cell shrinkage, chromatin condensation and DNA fragmentation. Moreover, delayed cytotoxicity was accompanied by an increase in intracellular reactive oxygen species (ROS) and a decrease in the mitochondrial membrane potential, and the two events presented obviously a negative correlation. CONCLUSION: ROS secondarily generated from damaged mitochondria may play a role in the induction of apoptosis.

Determination of trace tetracycline antibiotics in foodstuffs by liquid chromatography-tandem mass spectrometry coupled with selective molecular-imprinted solid-phase extraction.

A rapid, specific, and sensitive method has been developed using molecularly imprinted polymers (MIPs) as solid-phase extraction sorbents for extraction of trace tetracycline antibiotics (TCs) in foodstuffs. MIPs were prepared by precipitation polymerization using tetracycline as the template. Under the optimal condition, the imprinting factors for MIPs were 4.1 (oxytetracycline), 7.0 (tetracycline), 7.4 (chlortetracycline), 7.7 (doxycycline), respectively. Furthermore, the performance of MIPs as solid-phase extraction sorbents was evaluated and high extraction efficiency of molecularly imprinted solid-phase extraction (MISPE) procedure was demonstrated. Compared with commercial sorbents, MISPE gave a better cleanup efficiency than C18 cartridge and a higher recovery than Oasis HLB cartridge. Finally, the method of liquid chromatography-tandem mass spectrometry coupled with molecular-imprinted solid-phase extraction was validated in real samples including lobster, duck, honey, and egg. The spiked recoveries of TCs ranged from 94.51% to 103.0%. The limits of detection were in the range of 0.1-0.3 microg kg(-1).

Membrane damage effect of therapeutic ultrasound on Ehrlich ascitic tumor cells.

PURPOSE: The biologic effects and the underlying mechanisms of Ehrlich ascitic tumor (EAT) cells induced by ultrasound were investigated in this study. METHODS: Cells were subjected to ultrasonic irradiation with a frequency of 2.17 MHz and an intensity of 3 W/cm(2) for variable periods of time. Trypan blue exclusion was used to detect the integrity of cellular membrane; the membrane permeability was investigated by the incorporation of fluorescein isothiocyanate dextran during ultrasound exposure; and the cell membrane ultrastructure changes were observed under a scanning electron microscope. The potential mechanism was estimated from the generation of hydroxyl radicals, the lipid peroxidation levels, and intracellular reactive oxygen radicals production. RESULTS: The cell membrane damage effects induced by ultrasound increased with a prolonged exposure time; the fluorescent rates of the cells irradiated with ultrasound for 30 and 60 seconds were 11.46% and 18.50%, respectively; the amount of hydroxyl radicals in 30 (26.10 U/mL) and 60 seconds (28.47 U/mL) were significantly enhanced, compared with the control group (24.44 U/mL); then, the level of lipid peroxidation was also changed from 0.27 to 0.54 (30 seconds) and 1.21 nmol/mL (60 seconds). CONCLUSIONS: Shear forces and free radicals produced by acoustic cavitation may play important roles in these actions.

Study of cell killing effect on S180 by ultrasound activating protoporphyrin IX.

The present study was initiated to investigate the potential biological mechanism of cell killing effect on isolate sarcoma 180 (S180) cells induced by ultrasound activating protoporphyrin IX (PPIX). S180 cells were exposed to ultrasound for 30s duration, at a frequency of 2.2 MHz and an acoustic power of 3 W/cm(2) in the presence of 120 microM PPIX. The viability of cells was evaluated using trypan blue staining. The generation of oxygen free radicals in cell suspensions was detected immediately after treatment using a reactive oxygen detection kit. A copper reagent colorimetry method was used to measure the level of FFAs released into cell suspensions by the process of cell damage induced by ultrasound and PPIX treatment. Oxidative stress was assessed by measuring the activities of key antioxidant enzymes (i.e., SOD, CAT, GSH-PX) in S180 tumor cells. Treatment with ultrasound and PPIX together increased the cell damage rate to 50.91%, while treatment with ultrasound alone gave a cell damage rate to 24.24%, and PPIX alone kept this rate unchanged. Colorimetry and enzymatic chemical methods showed that the level of FFAs in cell suspension increased significantly after the treatment, while the activity of all the above enzymes decreased in tumor cells at different levels, and were associated with the generation of oxygen free radicals in cell suspension after treatment. The results indicate that oxygen free radicals may play an important role in improving the membrane lipid peroxidation, degrading membrane phospholipids to release FFAs, and decreasing the activities of the key antioxidant enzymes in cells. This biological mechanism might be involved in mediating the effects on S180 cells and resulting in the cell damage seen with SDT.

[The killing effect of focused ultrsound activating protoporphyrin IX on S180 cells].

The killing effect on S180 cells was studied using the combination of protoporphyrin IX and focused ultrasound at the frequency of 2.2 MHz and different intensities. Cell viability was determined by trypan blue exclusion test, morphology changes were evaluated by means of scanning electron microscopy and transmission electron microscopy after ultrasonic exposure. The results indicated that protoporphyrin IX(PPIX) alone showed no significant effect on S180 cells when compared with that of control group. Ultrasound alone and ultrasound combined with PPIX groups showed some anti-tumor effect, which became more noticeable as the ultrasound intensity and PPIX concentration increased, and when the concentration of PPIX increased to 120 microM, the ultrasound combined with PPIX exerted a more significant anti-tumor effect than did the ultrasound alone in the same experiment.

Comparison between sonodynamic effect with protoporphyrin IX and hematoporphyrin on sarcoma 180.

PURPOSE: The comparison between sonodynamic antitumor effect with protoporphyrin IX (PPIX) and hematoporphyrin (Hp) at a concentration of 5 mg/kg on Sarcoma 180 (S180) cells was studied in vivo, and the potential cell damage mechanism was also investigated. METHODS: The sonodynamically induced anti-tumor effect of PPIX was studied in mice bearing S180 solid tumors. In order to determine the optimum timing of ultrasound exposure after administration of PPIX, the PPIX concentrations in plasma, skin, muscle and tumor were determined by the fluorescence intensity of tissue extractions with a fluorescence spectrophotometer based on the standard curve. Anti-tumor effects were estimated by measuring the tumor size and the tumor weight. Additionally, the morphological changes of S180 cells were evaluated by transmission electron microscope (TEM) observation immediately after sonodynamic therapy (SDT) treatment. RESULTS: A time of 24 h after the intravenous administration of PPIX was chosen as the best time for ultrasound exposure. The antitumor effect induced by PPIX mediated sonodynamic therapy (PPIX-SDT) was in a dose dependent manner when ultrasound intensity was at or above the inertial cavitation threshold (5 W/cm(2)). A significant tumor growth delay was observed both in PPIX mediated sonodynamic therapy and in Hp mediated sonodynamic therapy treatments (Hp-SDT), and the tumor weight inhibition ratios after the synergistic treatments were 42.82 +/- 0.03 and 35.22 +/- 0.03%, respectively, this difference was significant at P < 0.05. While ultrasound alone (5 W/cm(2)) showed a slight tumor growth inhibitory effect compared with the control group, and PPIX or Hp alone showed almost no significant effect. Furthermore, TEM observation indicated cell damage was more serious in PPIX-SDT treatment group than in Hp-SDT treatment group. After sonication, the cell ultra-structure such as cell membrane destruction, mitochondria swelling, chromatin condensation might be important factors that inhibited the tumor growth and even induced cell death. CONCLUSIONS: The comparative results suggested that PPIX as a sonosensitizer might have more potential cytotoxicity than Hp when irradiated with ultrasound, and the ultra-structural changes may account for cell destruction induced by sonodynamic therapy in our experiment mode.

A quantitative DNA methylation assay using mismatch hybridization and chemiluminescence.

OBJECTIVE: To develop a quantitative method for methylation analysis of the p16 gene based on mismatch hybridization and chemiluminescence. METHODS: Genomic DNA was modified by sodium bisulfite to convert all unmethylated but not methylated cytosines to uracil, and subsequently a pair of primer having no CpG sites was designed for amplification target DNA containing methylated or unmethylated CpG sites. The PCR product spanning CpG sites were hybridized with two oligonucleotide probes which perfectly matched the methylated and unmethylated CpG sequences respectively, and the hybrids were detected by chemiluminescent method. The percentage of methylated target sequences could be estimated by calculating the ratio of signals obtained with two probes. RESULTS: The percentage of methylation of artificial mixtures DNA showed a linear relation. There was a negative correlation between the methyaltion index with p16 transcriptional mRNA of p16 gene in tumor cell lines. CONCLUSION: Compared with existing methods, this assay is nonisotopic, rapid, simple, and can be widely applied to the study of DNA methylation.

[Expression of DNA excision repair enzymes and lung cancer prognosis].

OBJECTIVE: To study the expression levels of three DNA damage excision repair enzymes: excision repair cross-complementing rodent repair deficiency gene 2 (ERCC2), uracil DNA glycosylase (UDG), and proliferating cell nuclear antigen (PCNA), in different lung tissues and their relationship with lung cancer prognosis. METHODS: The expression levels of ERCC2, UDG, and PCNA protein were detected using immunohistochemistry method. Cancer tissues and normal tissues adjacent to the cancer from 61 cases of lung carcinoma, and tissue samples from 18 cases of benign lung disease were studied. The relationship among ERCC2, UDG, and PCNA expression levels with lung cancer development and prognosis were studied using Ridit analysis method. RESULTS: The expression levels of all three proteins were not significantly different between cancer tissues and a normal tissues adjacent to the cancer (P > 0.05); but significantly different between cancer tissues and benign tissues, and between normal tissues adjacent to the cancer and benign tissues (P < 0.05). ERCC2 and UDG levels were higher in benign tissues than those in cancer and cancer adjacent tissues, while the PCNA level was the opposite. Only the UDG level in cancer adjacent normal tissues showed no significant difference compared to that of benign tissues (P > 0.05). There were no relationship among age, smoke, cancer metastasis, cancer type and ERCC2, UDG, PCNA expression levels. ERCC2 also showed no relationship with degree of malignancy and cancer size, while UDG and PCNA expression levels were correlated with cancer malignancy and cancer size (P < 0.05). Low UDG and high PCNA expression levels were correlated with higher malignancy and larger tumors. CONCLUSION: The expression levels of ERCC2, UDG and PCNA were significantly different in benign lung tissues, lung cancer tissues and lung cancer adjacent normal tissues. ERCC2 showed no significant relationship with lung cancer prognosis, while patients with low UDG and high PCNA expressions were more likely to have higher malignancy and larger tumors. UDG and PCNA were possible markers for evaluating lung cancer prognosis.

[Construction of nucleotide excision repair gene XPB antisense RNA expression plasmid and its functions].

BACKGROUND & OBJECTIVE: Nucleotide excision repair is an important pathway for cellular DNA damage repair. The drug resistance of tumor cell is often companied with the enhanced expression of DNA repair genes. Down-regulation of DNA repair capacity by antisense strategy can increase the drug sensitivity of tumor cells. The aim of this study was to construct the eukaryotic expression plasmid pcDNA-XPB/AS (XPB: xeroderma pigmentosum B) and to investigate the function of XPB gene and its roles in chemotherapeutic drug sensitivity in lung cancer A549 cell. METHODS: The XPB cDNA (69-520 bp) fragment amplified by reverse transcription polymerase chain reaction (RT-PCR) was inserted into pcDNA3.1/His plasmid with an inverted orientation. The recombinant plasmid was transiently transfected into A549 cells. The Adriamycin-induced DNA damage was compared between the transfected and the untransfected cells by single cell gel electrophoresis assay (SCGE). The cellular sensitivity to Adriamycin of the transfected and the untransfected cells was determined by MTT assay. RESULTS: The successful construction of antisense plasmid was proved by restriction map and sequence analysis. RT-PCR results showed that the XPB mRNA expression was inhibited in transfected A549 cells. SCGE showed that the cellular damage repair ability induced by 4.0 microg/ml Adriamycin was suppressed in transfected cells. MTT assay showed the sensitivity of the transfected cells to Adriamycin was different from the untransfected cells but without statistical meaning. CONCLUSION: The antisense plasmid constructed by the authors can down-regulate the expression of XPB mRNA in the transfected cells and inhibit the cellular DNA damage repair ability, providing a basis to further study the gene function of XPB.