Biomedical subjects

Xiao-Yu Li

Publications and source records attributed to Xiao-Yu Li.

At least 19 recordsLinked to original sources

PubMed2006

Toxicity of microcystins in the isolated hepatocytes of common carp (Cyprinus carpio L.).

The toxicity of hepatotoxic microcystins produced mainly by Microcystis aeruginosa in mammals and fishes was well studied in recent years. However, there were scarcely reports in toxic effects of microcystins on isolated hepatocytes of fishes, especially investigation of microcystin-induced apoptosis and/or necrosis in carp hepatocytes. In the present study, the isolated hepatocytes of common carp were exposed to various concentrations of microcystins (0.01, 0.1, 1, 10, 100, 1000 microg L(-1)) for 2, 4, 8, 16 and 24h, respectively, and cytotoxicity of microcystins in the toxin-treated cells was determined. Results of this study showed that cytotoxicity of microcystins on carp hepatocytes was time and dose-dependent, and the approximate LC(50) of microcystins in carp hepatocytes was 169.2 microg L(-1). The morphological changes typical of apoptosis, such as blebbing of cell membrane, condensation and fragmentation of cell nucleus were observed in the hepatocytes exposed to microcystins (1, 10 and 100 microg L(-1)) using fluorescence and differential interference contrast microscopy. Agarose gel electrophoresis of DNA demonstrated a typical apoptotic "ladder pattern" in microcystin-treated hepatocytes after 16 h of exposure. Results of the present study indicated that the form of cell death in microcystin-treated hepatocytes depend on the exposure dose of toxin. When lower concentration of microcystins (10 and 100 microg L(-1)) was used for exposure, carp hepatocytes died in apoptosis while, when higher one used (1000 microg L(-1)), they died in the form of necrosis.

Animals↗

PubMed2006

Toxic effects of Microcystis cell extracts on the reproductive system of male mice.

In this paper, the reproductive toxicity of male mice treated with Microcystis aeruginosa cell extracts containing microcystins was examined. In contrast to the control group, male mice exposed intraperitoneally to 3.33 or 6.67 microg microcystins/kg body weight for 14 days had decreased mean body weight, and the mean absolute weight of the testes and epididymides was decreased. However, the mean relative weight of the testes increased compared to the controls. In addition, histological examination of microcystin-treated mice indicated that the testes were damaged and the space between the seminiferous tubules was more pronounced compared to control mice. The quality of mature sperm in the seminiferous tubules was also decreased in treated mice compared with the control group. Further studies showed that motility and viability of the sperm from microcystin-treated mice were reduced, but no significant difference was found in the concentration and abnormality of the sperm from treated mice compared to the control. This study indicated that microcystins had numerous toxic effects on the reproductive system of male mice.

Animals↗

PubMed2006

Microcystins cause embryonic toxicity in mice.

Microcystins produced by freshwater cyanobacteria are potent hepatotoxins and can cause animal intoxications and human illnesses. In the present study, the effects of microcystins on the embryonic development of Kunming mice were determined using cell extracts of Microcystis aeruginosa from the Nanwan reservoir, China. Forty-eight pregnant mice were divided into four groups of 12 mice. Pregnant mice in three experimental groups were injected intraperitoneally with cell extracts at doses equivalent to 3, 6, or 12 microg microcystins/kg body weight daily from gestational days 6-15, while the mice in the control group were injected on the same schedule with sterilized saline. Mice were killed on the 18th day of gestation and embryonic and fetal developmental indexes checked. The fetal mice were also examined for anomalies of external, skeletal, and internal organs. The results demonstrated a significant decrease in body weight gain of pregnant mice in the 12 microg/kg dose group when compared to the control group (p<0.05). Differences in mean body weight, body length, and tail length of the fetuses were also found in these two groups (p<0.05). However, no significant difference in these characteristics was detected in the 6 or 3 microg/kg dose groups when compared to the control group (p>0.05). Four fetuses in the 6 microg/kg body dose group were found to have a curving tail. Additionally, petechial hemorrhage and hydropic degeneration were observed in the livers of fetuses in the 6 and 12 microg/kg experimental groups. These results suggested that microcystins had both maternal and embryonic toxicity in mice.

Animals↗

PubMed2006

Ultrasound-targeted microbubble destruction improves the low density lipoprotein receptor gene expression in HepG2 cells.

Ultrasound-targeted microbubble destruction had been employed in gene delivery and promised great potential. Liver has unique features that make it attractive for gene therapy. However, it poses formidable obstacles to hepatocyte-specific gene delivery. This study was designed to test the efficiency of therapeutic gene transfer and expression mediated by ultrasound/microbubble strategy in HepG2 cell line. Air-filled albumin microbubbles were prepared and mixed with plasmid DNA encoding low density lipoprotein receptor (LDLR) and green fluorescent protein. The mixture of the DNA and microbubbles was administer to cultured HepG2 cells under variable ultrasound conditions. Transfection rate of the transferred gene and cell viability were assessed by FACS analysis, confocal laser scanning microscopy, Western blot analysis and Trypan blue staining. The result demonstrated that microbubbles with ultrasound irradiation can significantly elevate exogenous LDLR gene expression and the expressed LDLRs were functional and active to uptake their ligands. We conclude that ultrasound-targeted microbubble destruction has the potential to promote safe and efficient LDLR gene transfer into hepatocytes. With further refinement, it may represent an effective nonviral avenue of gene therapy for liver-involved genetic diseases.

Cell Line, Tumor↗

PubMed2006

[Long-term effects of early hyperbaric oxygen therapy on neonatal rats with hypoxic-ischemic brain damage].

OBJECTIVE: The application and therapeutic effect of hyperbaric oxygen (HBO) in hypoxic-ischemic brain damage (HIBD) remains controversial. Previous studies have focused on the early pathological and biochemical outcomes and there is a lack of long-term functional evaluation. This study was designed to evaluate the long-term pathological and behavioral changes of early HBO therapy on neonatal rats with HIBD. METHODS: Postnatal 7 days (PD7) rat pups were randomly assigned into Control (n=18), HIBD (n=17) and HBO treatment groups (n=17). HIBD was induced by ligating the left common carotid, followed by 2 hrs hypoxia exposure in the HIBD and HBO treatment groups. The Control group was sham-operated and was not subjected to hypoxia exposure. The HBO therapy with 2 atmosphere absolutes began 0.5-1 hr after HIBD in the HIBD treatment group, once daily for 2 days. The spatial learning and memory ability were evaluated by the Morris water maze test at PD37 to PD41. The morphological and histological changes of the brain, including brain weight, survival neurons, AchE positive unit and NOS positive neurons in hippocampal CA1 region, were detected at PD42. RESULTS: The rats in the HIBD group displayed significant morphological and histological deficits, as well as severe spatial learning and memory disability. In the Morris water maze test, the mean escape latency were longer (56.35 +/- 22.37 s vs 23.07 +/- 16.28 s; P < 0.05) and the probe time and probe length were shorter in the HIBD group (29.29 +/- 6.06 s vs 51.21 +/- 4.59 s and 548 +/- 92 cm vs 989 +/- 101 cm; both P < 0.05) compared with the Control group. The left brain weight in the HIBD group was lighter than that in the Control group (0.601 +/- 0.59 g vs 0.984 +/- 0.18 g; P < 0.05). The survival neurons in the hippocampal CA1 region were less (100 +/- 27/mm vs 183 +/- 8/mm; P < 0.05), as well as the AchE-positive unit and NOS-positive neurons (18.50 +/- 2.24% vs 27.50 +/- 2.18% and 19.25 +/- 4.33 vs 33.75 +/- 5.57 respectively; P < 0.05) after HIBD. Early HBO treatment improved the abilities of spatial learning and alleviated the morphological and histological damage. The mean escape latency (39.17 +/- 21.20 s) was shortened, the probe time (36.84 +/- 4.36 s) and the probe length (686 +/- 76 cm) were longer, and the brain weight (0.768 +/- 0.85 g), the survival neurons (133 +/- 25/mm) and the AchE-positive unit (21.94 +/- 2.73%) increased significantly compared with those of the HIBD group (P < 0.05). CONCLUSIONS: Early HBO treatment resulted in a protective effect against HIBD-induced long-term brain morphological and histological deficits and spatial learning and memory disability.

Acetylcholinesterase↗

PubMed2005

S-adenosyl-L-homocysteine hydrolase inactivation curtails ovalbumin-induced immune responses.

The reversible S-adenosyl-L-homocysteine (AdoHcy) hydrolase inhibitor methyl 4-(adenin-9-yl)-2-hydroxybutanoate (DZ2002) suppresses macrophage activation and function. The effects of DZ2002 on T cell function, however, are still unclear. Here, we examined whether DZ2002 alters type 1 helper T cell (Th1) and/or type 2 helper T cell (Th2) immune responses, and whether these effects are associated with both the inhibition of AdoHcy hydrolase and intracellular elevation of endogenous AdoHcy. Male C57BL/6 mice immunized with ovalbumin (OVA) were treated with DZ2002 (1, 5, and 25 mg/kg/day) after which lymphocyte proliferation, cytokine production, and IgG responses to OVA were monitored. Administration of DZ2002 dose dependently suppressed OVA-specific lymphocyte proliferation and anti-OVA IgG production compared with controls. Interleukin (IL)-2 and interferon (IFN)-gamma as well as anti-OVA IgG2a and IgG3, indicators of Th1 immune responses, were markedly decreased in mice treated with DZ2002, whereas IL-4 and anti-OVA IgG1, indicators of Th2 immune responses, were only mildly suppressed. AdoHcy hydrolase activity in spleens of DZ2002-treated mice was substantially blocked, and not surprisingly, AdoHcy levels were significantly elevated compared with controls. Finally, similar immunosuppressive effects were also observed in mice treated with AdoHcy. These data strongly indicate that DZ2002 suppresses antigen-induced specific immune responses, particularly Th1 responses, through inhibition of AdoHcy hydrolase and elevation of endogenous AdoHcy.

Adenine↗

PubMed2005

Periplocoside E, an effective compound from Periploca sepium Bge, inhibited T cell activation in vitro and in vivo.

Periploca sepium Bge, a traditional Chinese herb medicine, is used for treating rheumatoid arthritis in China. Followed the bioactivity-guided isolation, the most potent immunosuppressive compound, periplocoside E (PSE), a pregnane glycoside, had been identified from P. sepium Bge. We investigated the immunosuppressive effects of PSE in vitro and in vivo. The results showed that PSE in a dose-dependent manner significantly inhibited the proliferation of splenocytes induced by concanavalin A and mixed lymphocyte culture reaction at no cytotoxic concentrations (<5 microM). Administration of PSE suppressed a delayed-type hypersensitivity reaction, and ovalbumin (OVA) induced antigen-specific immune responses in mice. In vivo treatment with PSE dose dependently suppressed OVA-induced proliferation and cytokine [interleukin (IL)-2 and interferon (IFN)-gamma] production from splenocytes in vitro. Purified T cells from OVA-immunized mice with PSE treatment showed its low ability for activation by OVA plus normal antigen presenting cell stimulation again in vitro. Further studies showed PSE dose dependently inhibited anti-CD3-induced primary T cell proliferation, activation for IL-2Ralpha (CD25) expression, and cytokine (IFN-gamma and IL-2) production also at the transcriptional level. PSE was highly specific and significantly inhibited the activation of extracellular signal-regulated kinase and Jun N-terminal kinase, whereas activation of p38 was not affected in T cells stimulated with anti-CD3. These results demonstrated that PSE is an immunosuppressive compound in P. sepium Bge, which directly inhibits T cell activation in vitro and in vivo. This study provided evidence to understand the therapeutic effects of P. sepium Bge and indicated that this herb is appropriate for treatment of T cell-mediated disorders, such as autoimmune diseases.

Animals↗

PubMed2005

Inhibition of inducible nitric-oxide synthase expression by (5R)-5-hydroxytriptolide in interferon-gamma- and bacterial lipopolysaccharide-stimulated macrophages.

(5R)-5-Hydroxytriptolide (LLDT-8) is a novel analog of triptolide that has antiarthritic, hepatoprotective, and antiallogenic transplantation-rejective effects. In the present study, we report that LLDT-8 inhibited nitric oxide (NO) production and inducible nitric-oxide synthase (iNOS) expression in macrophages. LLDT-8 significantly attenuated NO production, in a dose-dependent manner, in primary peritoneal macrophages and a macrophage cell line of Raw 264.7 cells following stimulation with interferon (IFN)-gamma, lipopolysaccharide (LPS), and IFN-gamma plus LPS. It also reduced the production of tumor necrosis factor-alpha from LPS-stimulated Raw 264.7 cells. To further elucidate the mechanism responsible for the inhibition of NO, we examined the effect of LLDT-8 on IFN-gamma and LPS-induced iNOS expression. Indeed, LLDT-8 prevented NO generation by inhibiting iNOS expression at mRNA level and protein level, rather than by interfering its enzymatic activity. In IFN-gamma-stimulated Raw 264.7 cells, LLDT-8 suppressed the gene transcription of signal transducer and activator of transcription 1alpha and interferon regulatory factor (IRF)-1, but it displayed no apparent effect on IFN-gamma receptor level on cell surface. After LPS challenge, LLDT-8 further abrogated the expression of LPS receptor complex, including CD14, Toll-like receptor 4, and myeloid differentiation protein-2; decreased the LPS-induced phosphorylation of stress-activated protein kinase/c-Jun NH(2)-terminal kinase, extracellular signal-regulated kinase 1/2, and p38 mitogen-activated protein kinase (MAPK); retarded the degradation of IkappaBalpha; and ameliorated the DNA binding activity of nuclear factor-kappaB (NF-kappaB) to nuclear proteins that accounts for transcriptional regulation of iNOS. Taken together, these results suggest that LLDT-8 reduces NO production and iNOS expression by inhibiting IFN-gamma-triggered IRF-1 expression and LPS-triggered MAPK phosphorylation and NF-kappaB activation.

Animals↗

PubMed2005

(5R)-5-hydroxytriptolide (LLDT-8), a novel triptolide analog mediates immunosuppressive effects in vitro and in vivo.

A series of triptolide analogs have been successfully synthesized. In the present study we demonstrated one of them, (5R)-5-hydroxytriptolide (LLDT-8), showed low cytotoxicity and relative high immunosuppressive activities as compared with its parent compound triptolide in vitro. The CC50 values of triptolide and LLDT-8 were 2.1+/-0.3 and 256.6+/-73.8 nM, respectively. LLDT-8 significantly inhibited the proliferation of splenocytes induced by concanavalin A (ConA), lipopolysaccharide (LPS), or mixed lymphocyte reaction (MLR), and the IC50 values were 131.7+/-32.4, 171.5+/-17.3, and 38.8+/-5.1 nM, respectively. LLDT-8 (25, 50, 100 nM) dose-dependently reduced the production of Th1 type cytokines (IFN-gamma, IL-2) and inflammatory cytokines (TNF-alpha, IL-6) in vitro. Administration of LLDT-8 (at the low dose of 0.4 microg/kg, i.p.; 40 microg/kg, p.o.) intensively suppressed 2,4-dinitrofluorobenzene (DNFB)-induced delayed type hypersensitivity (DTH) reactions. Treatment with LLDT-8 (40 microg/kg, i.p. and p.o.) also markedly inhibited the sheep red blood cell (SRBC)-induced antibody production in BLAB/c mice. Most importantly, comparing with triptolide, LLDT-8 significantly reduced toxicity, with a 122-fold lower cytotoxicity in vitro and 10-fold lower acute toxicity in vivo. The results suggested that LLDT-8 had immunosuppressive activities in both cellular and humoral immune responses. LLDT-8 might be a potential therapeutic agent for immune-related diseases.

Animals↗

PubMed2005

Inhibition of S-adenosyl-L-homocysteine hydrolase induces immunosuppression.

Lymphocytes depend on transmethylation reactions for efficient activation and function. These reactions are primarily catalyzed by S-adenosylmethionine-dependent methyltransferases, which convert S-adenosylmethionine to S-adenosyl-L-homocysteine. S-adenosyl-L-homocysteine is then hydrolyzed by S-adenosyl-L-homocysteine hydrolase to prevent feedback inhibition of transmethylation reactions. By impeding S-adenosyl-L-homocysteine hydrolase, a build-up of S-adenosyl-L-homocysteine occurs, and most intracellular transmethylation reactions cease. Thus, a nontoxic inhibitor of this enzyme might be a useful immunosuppressive therapeutic agent. We identified a potent reversible type III inhibitor of S-adenosyl-L-homocysteine hydrolase, DZ2002 [methyl 4-(adenin-9-yl)-2-hydroxybutanoate], and determined its cytotoxic and immunologic effects. We demonstrated that DZ2002 blocked S-adenosyl-L-homocysteine hydrolase more effectively than a type I inhibitor, but cytotoxicity from DZ2002 was greatly reduced. Although DZ2002 did not prevent concanavalin A-induced T cell proliferation or interleukin (IL)-2 production, it significantly reduced both a mixed lymphocyte reaction and IL-12 production from in vitro-stimulated splenocytes. In addition, levels of CD80 and CD86 on human monocytic THP-1 cells were decreased in a dose-dependent manner in the presence of 0.1 to 10 microM DZ2002, and decreases were also seen in IL-12 and tumor necrosis factor-alpha production from both mouse thioglycollate-stimulated peritoneal macrophages and THP-1 cells. In vivo, DZ2002 significantly suppressed a delayed-type hypersensitivity reaction as well as antibody secretion. We conclude that DZ2002's immunosuppressive effects are likely not solely attributed to T cell inhibition but also to the obstruction of macrophage activation and function through reductions in cytokine output and/or T cell costimulation. These data suggest an important dual role for the S-adenosyl-l-homocysteine hydrolase in both macrophage and T cell function.

Adenosylhomocysteinase↗

PubMed2005

Oral exposure to Microcystis increases activity-augmented antioxidant enzymes in the liver of loach (Misgurnus mizolepis) and has no effect on lipid peroxidation.

Recently, eutrophication has induced severe cyanobacterial blooms in the Naktong River, the second largest river of Korea. In the present study, lipid peroxidation and the antioxidant enzymes superoxide dismutase, catalase, and glutathione peroxidase, were evaluated in the liver of loach (Misgurnus mizolepis) that were orally exposed to a low dose of Microcystis through dietary supplementation with bloom scum. Loach received 75 mg of dry cells/kg body weight mass (equal to 10 microg microcystin-RR/kg body mass), for 28 days under controlled conditions. Antioxidant enzymatic activity and lipid peroxidation were measured after termination of exposure. The activities of antioxidant enzyme were significantly increased in the livers of toxin-exposed loach after 28 days of exposure, as compared to control fish. However, lipid peroxidation remained stable in both groups. These results suggest that antioxidant enzymes were able to eliminate oxidative stress induced by low concentrations of microcystins and to prevent increased lipid peroxidation in the liver of loach.

Animals↗

PubMed2005

Triptolide suppresses CD80 and CD86 expressions and IL-12 production in THP-1 cells.

AIM: To investigate the effects of triptolide, a diterpenoid triepoxide from Tripterygium wilfordii Hook F (TWHF), on the co-stimulatory molecule expression and interleukin-12 (IL-12) production from THP-1 cells. METHODS: THP-1 cells were differentiated into macrophage-like cells by Me2SO, and then cultured with IFN-gamma (500 kU/L) and lipopolysaccharide (LPS) (1 mg/L) with or without triptolide. The surface molecule expressions were analyzed on a FACScan flow cytometer. IL-12p40, IL-12p70 were assayed by ELISA. RESULTS: Triptolide suppressed CD80 and CD86 expressions on IFN-gamma (500 kU/L) and LPS (1 mg/L) activated THP-1 cells at nontoxic dosages of 2.5-0.625 microg/L. Furthermore, the production of IL-12p40 and IL-12p70 were also significantly reduced in THP-1 cells exposed to triptolide. CONCLUSION: Triptolide impairs the antigen-presenting function by inhibiting CD80 and CD86 expressions and decreased IL-12p40 and IL-12p70 (bioactive form) productions from the activated THP-1 cells.

Antigens, CD↗

PubMed2005

PGE2 up-regulates vascular endothelial growth factor expression in MKN28 gastric cancer cells via epidermal growth factor receptor signaling system.

AIM: 1) To evaluate the effect of prostaglandin E2 (PGE2) on the regulation of vascular endothelial growth factor (VEGF) expression in gastric MKN28 cells, and 2) to investigate the role of the epidermal growth factor receptor (EGFR) signal transduction pathway in any effect exerted by PGE2 on VEGF expression. METHODS: MKN28 cells were incubated with the vehicle (control) or with PGE2 in the presence or absence of AG1478, a selective inhibitor of EGFR tyrosine kinase, or PD098059, a selective inhibitor of the kinase responsible for ERK2 phosphorylation (mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK)). Real-time quantitative polymerase chain reaction and Western blot analysis were used to evaluate VEGF mRNA and protein expression. The activity of EGFR and ERK2 was measured by Western blot analysis. RESULTS: PGE2 significantly up-regulated VEGF mRNA and protein expression and increased the activation of EGFR and ERK2. Incubation of MKN28 cells with AG1478 significantly reduced PGE2-induced EGFR activity, ERK2 activity, and VEGF mRNA and protein expression. Meanwhile, incubation of MKN28 with PD098059 reduced PGE2-induced ERK2 activity and VEGF mRNA and protein expression, but had no effect on EGFR activity. CONCLUSION: Our data suggested that PGE2 up-regulates VEGF expression in gastric cancer cells via transactivation of EGFR-MAPK signaling pathways, which may be mechanisms underlying the contribution of COX-2 to tumor angiogenesis in gastric cancer.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed2004

Subchronic oral toxicity of microcystin in common carp (Cyprinus carpio L.) exposed to Microcystis under laboratory conditions.

The subchronic oral toxicity of microcystin in common carp (Cyprinus carpio L.) was investigated in this study. The fish (mean body weight of 322+/-36 g, n=10) were orally exposed to Microcystis by feeding with bloom scum at a dose of 50 microg microcystins/kg body weight under laboratory conditions for 28 days. Growth assay results showed that microcystin could completely inhibit the growth of carp, but failed to change the fish hepatosomatic index. Ultrastructural examination by electron microscope revealed severe damage in hepatocytes derived from the treated fish. Serum biochemical assays with commercial kits indicated that alanine aminotransferase and aspartate aminotransferase activities were significantly increased as compared to control levels, but gamma-glutamyl transferase, alkaline phosphatase and lactate dehydrogenase activities remained unchanged. Protein phosphatase inhibition assay revealed that the microcystin concentrations were 261.0+/-108.3 ng microcystin-LR equivalent/g fresh weight in hepatopancreas and 38.3+/-12.3 ng microcystin-LR equivalent/g fresh weight in muscle. The latter is above the limit recommended by the World Health Organization for human consumption. Therefore, we recommend that a warning system be instituted for announcing the occurrence of microcystin-producing water bloom and the possible risk of human intoxication.

Alanine Transaminase↗

PubMed2004

Recent pharmacological studies on natural products in China.

Natural products have been used as medicinal agents for many years. In addition, these compounds have also served as the starting points for semisynthetic analogs with improved properties. This review focuses on recent advances in the pharmacological studies on natural products mainly performed and published in China. Emphasis will be placed on those compounds that show the greatest promise clinically such as huperzine A (9-amino-13-ethylidene-11-methyl-4-azatricyclo[7.3.1.0(3.8)]trideca-3(8),6,11-trien-5-one), s-(-)-3-n-butylphthalide (s-(-)-3-butyl-1(3H)-isobenzofuranone), (-)-clausenamide (3-hydroxy-4-phenyl-5a-hydroxybenzyl-N-methyl-gamma-lactam) and Ginkgo biloba extract and its active components.

Alkaloids↗

PubMed2004

Differential regulation of resveratrol on lipopolysacchride-stimulated human macrophages with or without IFN-gamma pre-priming.

Resveratrol, a polyphenol compound found in grapes and red wines, is a prominent anti-cancer agent. In this study, we demonstrate that resveratrol enhanced TNF-alpha, IL-12 and IL-1beta production from LPS activated phorbol myristate acetate (PMA) differentiated THP-1 human macrophages. Expression of CD86 on macrophages was enhanced by resveratrol alone and with LPS. When macrophages were primed with IFN-gamma, resveratrol suppressed the expression of HLA-ABC, HLA-DR, CD80, CD86 and inhibited production of TNF-alpha, IL-12, IL-6 and IL-1beta induced by LPS. The differential impact of resveratrol on expression of CD14 might be correlated with differential response of macrophages to LPS with or without IFN-gamma priming.

Antigen-Presenting Cells↗

PubMed2004

Ultrasound/microbubble enhances foreign gene expression in ECV304 cells and murine myocardium.

Although viral vectors are efficient systems to transfer foreign genes into cells or target tissues, safety issues remain in relation to human gene therapy. Microbubbles currently used as ultrasound contrast agents have been applied in transfection of genes. This study was designed to test the transfection efficiency and the expression of exogenous gene mediated by ultrasound irradiation enhanced air filled albumin microbubbles in ECV304 cell line in vitro and the heart of the mouse in vivo. Air filled microbubbles (2.0-4.0 microm in diameter) were created by sonicating the mixture of human albumin, glucose, mannitol and special additive that was designed for stabilization. Plasmid DNA loading the reporter genes was gently mixed with microbubbles. The mixture of plasmid DNA and microbubbles was administrated to cultured ECV304 cells and BALB/c mice (tail vein injection) under different ultrasound/microbubble conditions, and then the transfection and expression efficiency were examined. The results both in vivo and in vitro demonstrated that microbubble with ultrasound irradiation could significantly elevate the exogenous gene expression as compared with microbubble or ultrasound only. Overall, the present study showed that the ultrasound-target microbubble destruction method enhanced the exogenous gene expression in vivo and in vitro, and provided a gene therapy way not only efficient but also easy to be manipulated and carried out in clinical.

Animals↗

PubMed2003

MARKK, a Ste20-like kinase, activates the polarity-inducing kinase MARK/PAR-1.

MARK, a kinase family related to PAR-1 involved in establishing cell polarity, phosphorylates microtubule-associated proteins (tau/MAP2/MAP4) at KXGS motifs, causes detachment from microtubules, and their disassembly. The sites are prominent in tau from Alzheimer's disease brains. We studied the activation of MARK and identified the upstream kinase, MARKK, a member of the Ste20 kinase family. It phosphorylates MARK within the activation loop (T208 in MARK2). A fraction of MARK in brain tissue is doubly phosphorylated (at T208/S212), reminiscent of the activation of MAP kinase; however, the phosphorylation of the second site in MARK (S212) is inhibitory. In cells the activity of MARKK enhances microtubule dynamics through the activation of MARK and leads to phosphorylation and detachment of tau or equivalent MAPs from microtubules. Overexpression of MARK eventually leads to microtubule breakdown and cell death, but in neuronal cells the primary effect is to allow the development of neurites during differentiation.

Amino Acid Sequence↗