Biomedical subjects

Li Liu

Publications and source records attributed to Li Liu.

At least 235 records · Page 13Linked to original sources

PubMed2004

Liposome-mediated extracellular superoxide dismutase gene delivery protects against acute liver injury in mice.

Our previous study demonstrated that polycationic liposomes are highly stable in the bloodstream and represent an effective agent for liver gene delivery. We report here that liposome-mediated extracellular superoxide dismutase (EC-SOD) gene delivery successfully prevented acute liver injury in mice. The therapeutic efficacy of EC-SOD gene delivery by polycationic liposomes was determined against the toxicity of superoxide anions and hydroxyethyl radicals in HepG2 cells and in a mouse model of acute liver injury caused by D-galactosamine and lipopolysaccharide intoxication. Transfection of HepG2 cells with an EC-SOD plasmid led to a striking increase in superoxide dismutase activity in the medium. The transfected cells had much less cell death after reactive oxygen species exposure compared with untransfected or control plasmid-transfected cells. In a model of acute liver injury, serum alanine aminotransferase levels in mice receiving portal vein injections of EC-SOD lipoplexes were much lower than in those receiving normal saline, liposomes alone, or control lipoplexes. Liver histology confirmed that there was less cell death in the EC-SOD lipoplex-treated group. Quantitative reverse transcriptase polymerase chain reaction showed a 55-fold increase in human EC-SOD gene expression in the liver of mice injected with EC-SOD lipoplexes. Serum superoxide dismutase activity in EC-SOD lipoplex-treated mice was higher than in the control groups; this was associated with higher liver glutathione levels and reduced lipid peroxidation. In conclusion, polycationic liposome-mediated EC-SOD gene delivery protects against reactive oxygen species toxicity in vitro and against lipopolysaccharide-induced acute liver injury in D-galactosamine-sensitized mice.

Acute Disease↗

PubMed2004

Irradiation induces a biphasic expression of pro-inflammatory cytokines in the lung.

BACKGROUND AND PURPOSE: The precise pathophysiological mechanisms of radiation-induced lung injury are poorly understood, but have been shown to correlate with dysregulation of different cytokines. The purpose of this study was to evaluate the time course of the pro-inflammatory cytokines tumor necrosis factor-(TNF-)alpha, interleukin-(IL-)1alpha and IL-6 after whole-lung irradiation. MATERIAL AND METHODS: The thoraces of C57BL/6J mice were irradiated with 12 Gy. Treated and control mice were sacrificed at 0.5, 1, 3, 6, 12, 24, 48, 72 h, 1, 2, 4, 8, 16, and 24 weeks post irradiation (p. i.). Real-time multiplex RT-PCR (reverse transcriptase polmyerase chain reaction) was established to evaluate the expression of TNF-alpha, IL-1alpha and IL-6 in the lung tissue of the mice. For histological analysis, lung tissue sections were stained by hematoxylin and eosin. RESULTS: Multiplex RT-PCR analysis revealed a biphasic expression of these pro-inflammatory cytokines in the lung tissue after irradiation. After an initial increase at 1 h p. i. for TNF-alpha and at 6 h p. i. for IL-1alpha and IL-6, the mRNA expression of these pro-inflammatory cytokines returned to basal levels (48 h, 72 h, 1 week, 2 weeks p. i.). During the pneumonic phase, TNF-alpha, IL-1alpha and IL-6 were significantly elevated and revealed their maximum at 8 weeks p. i. Histopathologic evaluation of the lung sections obtained within 4 weeks p. i. revealed only minor lung damage in 5-30% of the lung tissue. By contrast, at 8, 16, and 24 weeks p. i., 70-90% of the lung tissue revealed histopathologically detectable organizing alveolitis. CONCLUSION: Irradiation induces a biphasic expression of pro-inflammatory cytokines in the lung. The initial transitory cytokine response occurred within the first hours after lung irradiation with no detectable histopathologic alterations. The second, more persistent cytokine elevation coincided with the onset of histologically discernible organizing acute pneumonitis.

Animals↗

PubMed2004

Percutaneous transluminal septal myocardial ablation for hypertrophic obstructive cardiomyopathy; The Chinese experience in 119 patients from a single center.

BACKGROUND: Percutaneous transluminal septal myocardial ablation (PTSMA) has been introduced as an alternative treatment for hypertrophic obstructive cardiomyopathy (HOCM). We report on the acute, short- and long-term results of our experiences in 119 patients from a single center in China. METHODS AND RESULTS: PTSMA was performed in 119 patients with symptomatic HOCM (mean age 35.4+/-14.8 years, male 80, female 39). All patients had echocardiography performed prior to the procedure, 2-week post-PTSMA, and 6-month post-PTSMA, and 65 patients had echocardiography repeated at 2-year follow-up. The average left ventricular outflow tract (LVOT) gradient was 67.3+/-7.8 mm Hg before the procedure, and 15.9+/-6.8 mm Hg after the procedure (p<0.05). The thickness of interventricular septum (IVS) was 23.3+/-5.6 mm before the procedure, 18.6+/-4.8 mm 2-week post-PTSMA (p<0.05), and 16.8+/-3.4 mm 6-month post-PTSMA in all of the patients, and 15.6+/-3.1 mm 2-year post-PTSMA in 65 patients. The mean width of LVOT was 6.7+/-2.0 mm before the procedure, 8.2+/-3.4 mm 2-week post-PTSMA (p<0.05), 13.7+/-6.3 mm 6-month post-PTSMA and 15.1+/-2.4 mm 2-year post-PTSMA. The incidence of right bundle branch block development post-PTSMA was 52.9%, and three patients (2.5%) had complete heart block. There was no death. CONCLUSIONS: PTSMA is a promising non-surgical procedure for symptomatic patients with HOCM because of its low risk and its significant hemodynamic, echocardiographic and clinical improvement. The significant therapeutic remodeling period was up to 6 months rather than 2 years following the procedure.

Adult↗

PubMed2004

Longitudinal observation on CSF Abeta42 levels in young to middle-aged amyloid precursor protein/presenilin-1 doubly transgenic mice.

To relate levels of beta-amyloid42 (Abeta42) in the cerebrospinal fluid (CSF) and brain in early Alzheimer's disease, we repeatedly measured CSF Abeta42 levels in transgenic mice carrying Swedish amyloid precursor protein and presenilin-1 mutations, at ages before and after amyloid deposition. Hippocampal Abeta42 levels were measured at the endpoints. In APPswe/PS1(A246E) mice, CSF Abeta42 levels significantly increased between 5 and 7 months of age but did not change between 8 and 13 months despite a rapid increase in brain Abeta42. Furthermore, a decline in CSF Abeta42 levels was observed between 6 and 9 months in APPswe/PS1dE9 mice with faster pathology. Interestingly, the initial CSF Abeta42 concentrations correlated more strongly with brain Abeta42 levels than the endpoint CSF Abeta42. Our results suggest that the levels of CSF Abeta42 initially reflect the rate of Abeta42 production, but after reaching a critical concentration stay in equilibrium, until plaque formation leads to decreased CSF Abeta42 levels.

Aging↗

PubMed2004

Increased expression of pro-inflammatory cytokines as a cause of lung toxicity after combined treatment with gemcitabine and thoracic irradiation.

BACKGROUND AND PURPOSE: Preclinical evidence suggesting gemcitabine potentiates the anti-tumor effects of irradiation has resulted in clinical trials to evaluate the treatment efficacy of gemcitabine and concurrent thoracic irradiation in non-small-cell lung cancer (NSCLC). Although these studies demonstrated favorable tumor response, this combined treatment modality was accompanied by severe treatment-related toxicities predominantly of the lung. In an attempt to elucidate the determinants of lung toxicity for gemcitabine, we analyzed the expression of the pro-inflammatory cytokines TNF-alpha, IL-1alpha and IL-6 in the lung tissue of mice treated with gemcitabine and concurrent thoracic irradiation. MATERIALS AND METHODS: Four study groups were defined: C57BL/6J mice that received neither irradiation nor gemcitabine (NT-group), those that received gemcitabine (120 mg/kg intraperitoneal, i.p.) but no irradiation (GEM-group), those that underwent thoracic irradiation (12 Gy) without gemcitabine (XRT-group), and those that received both gemcitabine (120 mg/kg i.p., 2 h before irradiation) and thoracic irradiation (GEM/XRT-group). The mice were sacrificed at 1 h, 1 and 3 days, 1, 2 and 4 weeks post-treatment (p.t.). The mRNA expression of TNF-alpha, IL-1alpha and IL-6 in the lung tissue was quantified by competitive RT-PCR. The cellular origin of the cytokine expression was identified by immunohistochemistry. The cytokine expression was correlated with histopathological alterations. RESULTS: The TNF-alpha, IL-1alpha and IL-6 expression in the lung tissue of the GEM/XRT mice was clearly higher at all assessment time points compared to the NT mice (statistically significant at 1 h, 1 and 3 days, 1, 2 and 4 weeks p.t.), XRT mice (statistically significant at 1 week p.t.) or GEM mice (statistically significant at 1 h, 1 and 2 weeks p.t.). Maximal treatment-induced cytokine expression in the lung tissue of the GEM/XRT mice occurred already at 1 week p.t. (TNF-alpha: 30.9 +/- 5.3/IL-1alpha: 28.3 +/- 5.0/IL-6: 4.9 +/- 0.1 times basal level), and coincides with pathohistologically discernable interstitial pneumonitis. The elevated levels of TNF-alpha and IL-1alpha have been found to correlate with immunohistochemical staining of the bronchiolar epithelium and predominantly of inflammatory cells. CONCLUSIONS: Our data provide evidence that the increased expression of pro-inflammatory cytokines and the induction of a cytokine-triggered inflammatory response may be a determinant of the observed elevated lung toxicity after concurrent treatment with gemcitabine and thoracic irradiation.

Animals↗

PubMed2004

Endothelial targeting of the Sleeping Beauty transposon within lung.

Endothelial cells have complex roles in the pathophysiology of vascular and heart disease and are increasingly being recognized as targets for gene therapy. The intravenous administration of plasmid DNA complexed to lipid tends to target transfection of endothelial cells within the lung; however, expression from the transgene remains transient. Here we utilize the integrating capability of the Sleeping Beauty (SB) transposon for durable gene transfer within lung endothelia. To restrict expression of the transgene, an endothelial cell-specific promoter, endothelin-1, was placed within the transposon. Further refinements to the transposon increased in vitro transposition efficiency by 3.6-fold. Utilizing this optimized transposon we evaluated the expression of two reporter molecules, secreted alkaline phosphatase (SEAP) and intracellular GFP, following administration of DNA-polyethylenimine complexes to mice. Long-term expression (>2 months) of SEAP occurred only with cotransfection of adequate amounts of transposase. Localization studies using the GFP reporter, at 3 days and 6 weeks postinjection, demonstrated that the majority of transgene-expressing cells were of endothelial origin, while the second most abundant cell type was type II pneumocyte. These results suggest that the SB transposon can be adapted to target particular cell types, in this case, endothelial cells. Such an approach may be useful for gene therapy paradigms involving the long-term modulation of vascular and endothelial cell biology.

Alkaline Phosphatase↗

PubMed2004

Expression and purification of recombinant proteins from Escherichia coli: Comparison of an elastin-like polypeptide fusion with an oligohistidine fusion.

Thermally responsive elastin like polypeptides (ELPs) can be used to purify proteins from Escherichia coli culture when proteins are expressed as a fusion with an ELP. Nonchromatographic purification of ELP fusion proteins, termed inverse transition cycling (ITC), exploits the reversible soluble-insoluble phase transition behavior imparted by the ELP tag. Here, we quantitatively compare the expression and purification of ELP and oligohistidine fusions of chloramphenicol acetyltransferase (CAT), blue fluorescent protein (BFP), thioredoxin (Trx), and calmodulin (CalM) from both a 4-h culture with chemical induction of the plasmid-borne fusion protein gene and a 24-h culture without chemical induction. The total protein content and functional activity were quantified at each ITC purification step. For CAT, BFP, and Trx, the 24-h noninduction culture of ELP fusion proteins results in a sevenfold increase in the yield of each fusion protein compared to that obtained by the 4-h-induced culture, and the calculated target protein yield is similar to that of their equivalent oligohistidine fusion. For these proteins, ITC purification of fusion proteins also results in approximately 75% recovery of active fusion protein, similar to affinity chromatography. Compared to chromatographic purification, however, ITC is inexpensive, requires no specialized equipment or reagents, and because ITC is a batch purification process, it is easily scaled up to accommodate larger culture volumes or scaled down and multiplexed for high-throughput, microscale purification; thus, potentially impacting both high-throughput protein expression and purification for proteomics and large scale, cost-effective industrial bioprocessing of pharmaceutically relevant proteins.

Elastin↗

PubMed2004

The c-Raf inhibitor GW5074 provides neuroprotection in vitro and in an animal model of neurodegeneration through a MEK-ERK and Akt-independent mechanism.

Cerebellar granule neurons undergo apoptosis when switched from a medium containing high potassium (HK) to one that has low potassium (LK). LK-induced cell death is blocked by GW5074 [5-Iodo-3-[(3,5-dibromo-4-hydroxyphenyl) methylene]-2-indolinone], a synthetic drug that inhibits c-Raf activity in vitro. GW5074 has no direct effect on the activities of several apoptosis-associated kinases when assayed in vitro. In contrast to its effect in vitro, treatment of neurons with GW5074 causes c-Raf activation (when measured in vitro in the absence of the drug) and stimulates the Raf-MEK-ERK pathway. Treatment of neurons with GW5074 also leads to an increase in the activity of B-Raf, which is not inhibited by GW5074 in vitro at concentrations at which the drug exerts its neuroprotective effect. PD98059 and U0126, two distinct inhibitors of MEK, block the activation of ERK by GW5074 but have no effect on its ability to prevent cell death. Overexpression of a dominant-negative form of Akt does not reduce the efficacy of GW5074, demonstrating an Akt-independent mechanism of action. Neuroprotection is inhibited by SN-50, a specific inhibitor of nuclear factor-kappa B (NF-kappaB) and by the Ras inhibitor S-trans, trans-farnesylthiosalicylic acid (FTS) implicating NF-kappaB and Ras in the neuroprotective signaling pathway activated by GW5074. In addition to preventing LK-induced apoptosis, treatment with GW5074 protects against the neurotoxic effects of MPP+ and methylmercury in cerebellar granule neurons, and glutathione depletion-induced oxidative stress in cortical neurons. Furthermore, GW5074 prevents neurodegeneration and improves behavioral outcome in an animal model of Huntington's disease. Given its neuroprotective effect on distinct types of cultured neurons, in response to different neurotoxic stimuli, and in an animal model of neurodegeneration, GW5074 could have therapeutic value against neurodegenerative pathologies in humans.

Animals↗

PubMed2004

Disease-causing missense mutations in NPHS2 gene alter normal nephrin trafficking to the plasma membrane.

BACKGROUND: Podocin is a membrane-integrated protein that is located at the glomerular slit diaphragm and directly interacts with nephrin. The gene encoding podocin, NPHS2, is mutated in patients with autosomal-recessive steroid-resistant nephrotic syndrome (SRN). In order to study a potential pathomechanism of massive proteinuria in patients with SRN, we have investigated the trafficking and subcellular localization of five common disease-causing missense mutants of human podocin. METHODS: Site-directed mutagenesis was applied to generate cDNA constructs encoding five different missense mutations of human podocin (P20L, G92C, R138Q, V180M, and R291W). To identify the subcellular localization of each mutant in transfected human embryonic kidney (HEK)293 cells, we have generated and characterized a rabbit polyclonal antibody against the human podocin. Specificity of the antibody was determined by light and immunoelectron microscopy, as well as immunoblot analysis using human glomeruli. Confocal microscopy was applied to determine subcellular localization of the wild-type and the mutated podocin molecules, as well as wild-type nephrin in transfected cells. Immunoprecipitation and pull-down studies were carried out to investigate the molecular interaction of podocin mutants and wild-type nephrin. RESULTS: Immunofluorescence and confocal microscopy showed that wild-type podocin located to the plasma membrane when expressed in HEK293 cells. Two missense mutations, P20L and G92C, located at the N-terminus part of the molecule, were also present at the plasma membrane, indicating that these mutations did not affect the subcellular localization of the mutated podocin molecules. In contrast, subcellular localization of three other missense mutants located in the proximal C-terminus part of the protein was drastically altered, in which R138Q was retained in the endoplasmic reticulum (ER), V180M formed inclusion bodies in the cytoplasm, and the R291W mutant was trapped both in the ER and in small intracellular vesicles. Interestingly, this abnormal subcellular localization of podocin missense mutants also resulted in alteration in protein trafficking of wild-type nephrin in cotransfected cells through the strong protein binding between both molecules. CONCLUSION: In patients with SRN, some missense mutations in the NPHS2 gene not only lead to misfolding and mislocalization of the mutated podocin, but they can also interfere with slit diaphragm structure and function by altering the proper trafficking of nephrin to the plasma membrane.

Cell Line↗

PubMed2004

Genetic variability of the G glycoprotein gene of human metapneumovirus.

Human metapneumovirus (hMPV) has been associated with respiratory illnesses like those caused by human respiratory syncytial virus (HRSV) infection. Similar to other pneumoviruses, genetic diversity has been reported for hMPV. Little information is currently available on the genetic variability of the G glycoprotein (G), which is the most variable gene in RSV and avian pneumovirus. The complete nucleotide sequences of the G open reading frame (ORF) of 24 Canadian hMPV isolates were determined. Phylogenetic analysis showed the existence of two major groups or clusters (1 and 2). All but one of the hMPV isolates that we examined belonged to cluster 1. Additional genetic variability was observed in cluster 1, which separated into two genetic subclusters. Within cluster 1 the nucleotide sequence identity for the G ORF was 74.2 to 100%, and the identity for the predicted amino acid sequence was 61.4 to 100%. The G genes of cluster 1 isolates were more divergent from the cluster 2 isolates, with 45.6 to 50.5% and 34.2 to 37.2% identity levels for the nucleotide and amino acid sequences, respectively. Sequence analysis also revealed changes in stop codon usage, resulting in G proteins of different lengths (217, 219, 228, and 236 residues). Western blot analysis with the use of hMPV-specific polyclonal antisera to each hMPV cluster showed significant antigenic divergence between the G proteins of clusters 1 and 2. These results suggest that the G protein of hMPV is continuously evolving and that the genetic diversity observed for the hMPV genes is reflected in the antigenic variability, similar to HRSV.

Amino Acid Sequence↗

PubMed2004

Microarray analysis of the failure of filtering blebs in a rat model of glaucoma filtering surgery.

PURPOSE: To generate data concerning changes in levels of protein expression associated with wound healing and bleb failure in a rat model of glaucoma filtration surgery (GFS), and to identify factors that may play a role in this process. METHODS: Of 36 Sprague-Dawley rats, GFS was performed on 27 by introducing a silicone cannula through a scleral tunnel under a conjunctival flap, resulting in aqueous-filtering blebs that failed over 8 to 13 days. The additional nine rats were used as the nonsurgical control. Nine blebs were harvested at each of days 0, 2, 5, and 12 and pooled, yielding three replicates of three blebs per time point. RNA was extracted, labeled, and hybridized to 230A rat GeneChip arrays (Affymetrix, Santa Clara, CA). RESULTS: Of the 15,924 probe sets/genes present on the array, 923 genes were indicated to have a significant treatment effect at P < 0.005. Eight gene expression clusters were identified that could be broadly classified into three basic patterns. These were an increase on day 2, a decrease on day 2, or an increase on either day 5 or 12. The greatest change occurred between days 0 and 2. The most heavily populated functional categories included growth factors, structural proteins, and matrix metalloproteinases. CONCLUSIONS: This study represents the first large-scale gene expression analysis after GFS. Expression patterns for known mediators of the bleb scarring process, including transforming growth factor-beta, connective tissue growth factor, and matrix metalloproteinases, were confirmed, and a number of mediators not previously associated with this process were identified.

Animals↗

PubMed2004

Phytochemical analysis of anti-atherogenic constituents of Xue-Fu-Zhu-Yu-Tang using HPLC-DAD-ESI-MS.

Xue-Fu-Zhu-Yu-Tang is a famous traditional Chinese medicine (TCM) formula for treating cardiovascular disease and related ailments in China for centuries. To profile the phytochemical constituents of the formula, an HPLC-DAD-ESI-MS analytical method has been developed to separate and determinate the medium- or non-polar fraction of the decoction, which has been demonstrated potency to lower the serum total triglyceride concentration, strongly decrease the TXA(2)/PGI(2) ratio and attenuate production of proinflammatory cytokines in high cholesterol-fed rats. By comparing their retention time, UV and MS data with those obtained from the authentic compounds, ferulic acid (1), naringin (2), neohesperindin (3), naringenin (8), marmin (13), senkyunolide A (14), dehydrosafynol (16), safynol (17) and Z-ligustlide (18) are unequivocally determined. Moreover, additional thirteen compounds are tentatively identified as senkyunolide I (4), senkyunolide H (5), poncirin (7), benzoylpaeoniflorin (10), (Z)-6,7-epoxyligustilide (11), senkyunolide G (12), 2-methoxy-safynol (15), cnidilide (19), tangeritin (20), saikosaponin b(2) (21), 29-O-acetylsaikosaponin b(2) (22), saikosaponin b(1) (23) and auraptene (24), according to the comparison of their UV and MS data with the published data. The present study provides an approach to rapidly characterize bioactive constituents in TCM formulae.

Animals↗

PubMed2004

The synergetic effects of two CCAAT boxes in Aspergillus niger glaA gene promoter on activation of PglaA transcription.

EMSA and footprinting analyses have revealed that the -489-- -414 bp and the -390-- -345 bp (designated DC and PC respectively) upstream of the Aspergillus nigerT21 glaA gene were bound by one protein factor in the A. nigerT21 whole cell extract. Both DC and PC contained CCAAT pentanucleotides. The functions of DC and PC in regulation of expression of glucoamylase (GLA) were studied. CCAAT pentanucleotides were replaced with CGTAA and the mutated DNA fragments DCm and PCm lost the binding activities of protein factors in vitro. In vivo when either DC or PC was mutated or the relative orientations between them were changed on the PglaA, the transcriptional activity of PglaA decreased to a basal level. Introduction of multi-copies of DC into the original site at the PglaA in A. nigerT21 decreased the expression of endogenous GLA expression and the exogenous reporter E. coli uidA gene introduced under the PglaA promoter, while having no effect on the uidA gene under the control of PgpdA. EMSA revealed that the levels of the specific DNA-binding protein factors in the transformants maintained the same meaning that introduction of multi-copies of DC caused the titration effect. AnghapC gene was cloned from A. nigerT21 cDNA and introduced into the DC multi-copied strains. The expression of AnghapC improved the expression of the endogenous GLA and the exogenous gene controlled by PglaA. These results showed that both the CCAAT pentanucleotides were necessary for DC and PC binding to the protein factors, and the simultaneous binding of DC and PC to the protein was necessary for promoting the transcriptional activity of PglaA. AngHapC was the specific positive trans-acting protein factor binding to DC.

Amino Acid Sequence↗

PubMed2004

[Expression of cyclooxygenase-2 in esophageal squamous cell carcinogenesis].

OBJECTIVES: To investigate the expression of cyclooxygenase-2 (cox-2) protein in normal squamous epithelium, squamous dysplasia and squamous cell carcinoma of the esophagus, and to elucidate the role of cox-2 in esophageal carcinogenesis. METHODS: Biopsy specimens of atypical esophageal dysplasia (n = 47) and surgical resection of squamous cell carcinoma (n = 86) were compared with normal esophageal specimens (n = 42) and the expression of cox-2 in those specimens was analyzed by immunohistochemistry and western blotting. RESULTS: A significant elevated cox-2 expression was shown in atypical esophageal squamous dysplasia and squamous cell carcinoma, as compared to that in normal esophageal squamous epithelium, with immunohistochemical stain scores of 2.67 +/- 1.77, 2.19 +/- 1.79 and 0.71 +/- 0.46, respectively. Results of western blotting analysis confirmed those obtained by immunohistochemistry. Cox-2 expression significantly correlated with proliferation activity assessed by proliferating cell nuclear antigen index in dysplastic and carcinomous lesions, respectively, and no such correlation could be found in normal esophageal mucosa. Elevated cox-2 expression was not associated with clinical-pathological features of esophageal squamous carcinoma, including age, gender, tumor size, histological grade, lymph node metastasis and clinical stage. CONCLUSION: Elevated expression of cox-2 in atypical squamous dysplasia and squamous cell carcinoma of the esophagus, which correlated with cell proliferation activity, indicated that cox-2 may be involved in the early stage of squamous carcinogenesis of the esophagus, and may be a target of prevention and treatment for esophageal squamous cell carcinoma.

Adult↗

PubMed2004

[Changes of biological characteristics of colon carcinoma cell line - LoVo induced by Kai1/CD82 transfection].

OBJECTIVE: To study the influence of Kai1/CD82 transfection on the growth, adherence, separation and invasion potential of LoVo colon carcinoma cell line. METHODS: Kai1/CD82 cDNA was transfected into LoVo cells, and a stable expressing clone was established. In vitro methodology was used to obtain the growth curve and also to detect the adherence, separation and invasion potential of the transfected LoVo cells, in comparison with those of control cells without transfection. RESULTS: Compared with the control, no change was observed in the growth pattern of transfected LoVo cells. The numbers of adherent cells in the two groups were 0.08, 0.63, 0.83, 0.91 (x 10(5)) for the transfected cells and 0.04, 0.48, 0.71, 0.82 (x 10(5)) for the control cells respectively after 10, 20, 30, 40 minutes culture with shaking. The difference at 20, 30 and 40 minutes was statistically significant (P < 0.05). The separation rates of each group were 13%, 20%, 53% for the transfected cells and 11%, 28%, 60% for the control cells, respectively after 5, 10, 15 minutes culture with shaking. The difference at 10 and 15 minutes was statistically significant (P < 0.05). The aggregation rate of the transfected cells was higher than that of the control cells after culture with mild shaking for 5 hours (64.8% vs. 58.6%, P < 0.05). After co-incubation with endothelium cells ECV304, the number of invading cells decreased more in the transfected cells than that in the control cells (6.33/field and 17.67/field, P < 0.05). CONCLUSION: Transfection expression of Kail/CD82 into LoVo cell line results in an increase of cell adherence and aggregation, but a diminished capability of separation and invasion, suggesting that the expression of Kai1/CD82 gene may inhibit the metastatic potential of colon carcinoma.

Antigens, CD↗

PubMed2004

[Studies on synthesis and properties of N-benzoyl-THA targeting for brain].

It is the intent of these studies to synthesize acylated prodrugs of tacrine hydrochloride for improving the penetrating ability across the blood-brain barrier (BBB). Prodrugs of tacrine were prepared by benzoylation of the 9 amino group of THA. The structures of the synthesized prodrugs were determined by IR, 1H NMR, MS and UV. Their physical-chemical properties and stability under different conditions were investigated. The function of prodrugs targeting for brain were evaluated in vitro by biopartition micellar chromatography (BMC). The results showed that prodrugs were stable in different environments. Ethyl acetate/water partition coefficient indicated that the lipophilicity of prodrugs was increased, compared with THA. BMC predicted that prodrugs could improve the infiltration ability across BBB of parent-drug. It was suggested that this kind of prodrugs be a promising brain-targeting delivery system.

Animals↗

PubMed2004

[Preparation and transfection of GFP plasmid DNA/cationic liposome complex].

OBJECTIVE: This work was directed at obtaining a better gene carrier to improve the effects of gene delivery. METHODS: Cationic liposomes made from cholesterol, lecithin and Eighteenth Amic by reverse evaporation technique were used for encapsulating plasmid DNA containing gfp gene. The DNA/liposome complexes differed in quantity of plasmid DNA. The sizes of complexes and the efficiency of encapsulation were detected. MTT assay was used to measure the cytotoxicities of complexes. The efficiency of transfection into COS1 cells was shown by observation of green fluorescent and measurement of fluorescent intensity. RESULTS: The average size of complexes was 562 nm, the encapsulating efficiency of DNA in microspheres reached 55%-65%. These DNA/Cationic liposome complexes could be transfected into mammalian cells, but they were different from each other in efficiency of transfection. The cytotoxicities of these complexes varied with the concentration of DNA in complexes, the quantities of complexes and the time of treatment by complexes. CONCLUSION: DNA/Cationic liposome complexes prepared by reverse evaporation technique could be applied in DNA delivery system.

Cations↗

PubMed2004

[Radiosensitization by pentoxifylline in human hepatoma cell line HepG2 and its mechanism].

OBJECTIVE: To investigate the radiosensitization by pentoxifylline (PTX) in human hepatoma cell line HepG2 and its possible mechanism. METHODS: MTT assay was used to evaluate the cytotoxicity of PTX and clonogenic assay employed to observe its effects on the radiosensitivity of the cells quantified by calculating the sensitive enhancement ratio (SER). Flow cytometry was performed to observe the changes in the cell cycle of HepG2 cells in response to X-ray irradiation and the effect of PTX intervention. RESULTS: The cytotoxicity of PTX increased in a dose-dependent manner following a 24-hour treatment, with the optimal dose range of 1-5 mmol/L. A sub-toxic dose of PTX at 2 mmol/L was used in the subsequent experiments. PTX significantly reduced the clonogenic activity and enhanced the radiosensitivity of HepG2 cells with a SER of 1.3+/-0.16 at the dose of 2 mmol/L. Irradiation resulted in cell cycle arrest at G2 phase in HepG2 cells, and the percentages of HepG2 cells in G2-M phase were 32.15% and 19.52% respectively after exposure to 6 Gy radiation alone and to 6 Gy plus 2 mmol/L PTX at 20 h, demonstrating the effectiveness of PTX in resolving radiation-induced G2 arrest. CONCLUSION: Radiosensitization by PTX is possibly associated with the abrogation of G2 arrest in HepG2 cells following radiation exposure.

Carcinoma, Hepatocellular↗