Biomedical subjects

L M Zhang

Publications and source records attributed to L M Zhang.

At least 19 recordsLinked to original sources

PubMed2006

Improved constraints on D0-D0 mixing in D0 --> K+ pi- decays from the Belle detector.

We report the results of a search for D0-D0 mixing in D0 --> K+ pi- decays based on 400 fb(-1) of data accumulated by the Belle detector at KEKB. Both assuming CP conservation and allowing for CP violation, we fit the decay-time distribution for the mixing parameters x and y, as well as for the parameter R(D), the ratio of doubly Cabibbo-suppressed decays to Cabibbo-favored decays. The 95% confidence level region in the (x'2,y') plane is obtained using a frequentist method. Assuming CP conservation, we find x'2 < 0.72 x 10(-3) and -9.9 x 10(-3) < y' < 6.8 x 10(-3) at the 95% confidence level; these are the most stringent constraints on the mixing parameters to date. The no-mixing point (0,0) has a confidence level of 3.9%. Assuming no mixing, we measure R(D) = (0.377 +/- 0.008 +/- 0.005)%.

Journal Article↗

PubMed2003

Vascular endothelial growth factor inhibits outward delayed-rectifier potassium currents in acutely isolated hippocampal neurons.

In the present study, whole-cell patch-clamp recording was used to study whether vascular endothelial growth factor (VEGF) had a regulatory effect on the potassium-channel currents. The outward delayed-rectifier potassium currents (I(K)) were recorded in acutely isolated hippocampal neurons from 14-day-old rat brains. A local application of VEGF at the concentrations from 50 ng/ml to 200 ng/ml dose-dependently inhibited I(K). Administration of VEGF (100 ng/ml) to the neurons only for seconds could significantly reduce I(K) in 26 of 39 recorded cells. The currents could recover to 82.8+/-3.7% of the control level at 60 s after removing VEGF in the buffer. In the I-V curve analysis, VEGF negatively shifted the I-V curve of I(K); the inhibition was gradually enhanced as the membrane potential increased from -40 mV to 50 mV in 13 cells. Thus, the results reveal that VEGF inhibits I(K) in acute, reversible and voltage-dependent manners. Double staining combined with confocal laser scanning microscopy was used to simultaneously detect the distribution of VEGF receptors (flt-1 and flk-1) in the hippocampal section and isolated neuron. Results showed that flt-1-positive staining, but not flk-1, could be observed on the membrane of the hippocampal neuron in both preparations, suggesting the presence of neuronal membrane VEGF flt-1 receptors in the hippocampus. To investigate if the inhibition by VEGF on I(K) is related to the presence of flt-1 receptors, we further did flt-1-receptor immunostaining for the recorded neurons, which was labeled with Lucifer Yellow during the recording. Among nine recorded cells, five showing the inhibition by VEGF had detectable signals for flt-1 receptors on their membrane, whereas the other four showing no inhibition had no flt-1 receptors either. The results suggest that VEGF can acutely inhibit I(K) in the hippocampal neurons probably related to the presence of membrane flt-1 receptors in the neurons.

Action Potentials↗

PubMed2001

Effects of intraplantar injection of carrageenan on central dopamine release.

The present study examined the changes of extracellular concentration of dopamine and its metabolite homovanillic acid in the striatum, the periaqueductal gray and the dorsal horn of lumbar spinal cord following intraplantar injection of carrageenan. In vivo microdialysis and high-performance liquid chromatography with electrochemical detection were performed. The results showed that intraplantar injection of carrageenan significantly increased dopamine or homovanillic acid levels in the different central regions. The present study suggested that the central dopamine system (at least including ascending nigrostrialtal pathway and descending A11--the dorsal horn fibers) be activated following the carrageenan, while the activation probably mediated antinociception.

Animals↗

PubMed2001

Role of excitatory amino acid transporter 1 in neonatal rat neuronal damage induced by hypoxia-ischemia.

The role of excitatory amino acid transporter 1 in neonatal rat neuronal damage was studied following hypoxia-ischemia. To induce hypoxia-ischemia injury, rats on postnatal day 7 were exposed to 8 % oxygen for 2 h following unilateral common carotid artery ligation. According to brain damage scoring based on Cresyl Violet staining, the neuronal damage time-dependently changed in the ischemic regions following hypoxia-ischemia. Immunohistochemical studies showed that excitatory amino acid transporter 1 expression was mainly observed in the cerebral cortex ipsilateral to common carotid artery ligation and markedly increased at 24 h and 48 h following hypoxia-ischemia. Combined with confocal laser scanning microscopic analysis, double staining showed that excitatory amino acid transporter 1 positive staining appeared in neurons as well as astrocytes after hypoxia-ischemia. Most excitatory amino acid transporter 1 positive staining cells exhibited regular morphological characteristics and only a few were double-stained by terminal deoxynucleotidyl transferase-mediated deoxyuridinetriphosphate nick-end labeling. Down-regulation of excitatory amino acid transporter 1 expression by intraventricular administration of specific antisense oligonucleotide exacerbated neuronal damage in hypoxia-ischemia brain. These results suggest that the increase of excitatory amino acid transporter 1 expression may be involved in a pathophysiological process of hypoxia-ischemia brain damage and may reflect a self-compensative mechanism for protecting neurons from further injury.

ATP-Binding Cassette Transporters↗

PubMed2001

Chinese phlebotomine sandflies of subgenus Adlerius nitzulescu, 1931 (Diptera: Psychodidae) and the identity of Phlebotomus sichuanensis Leng & Yin, 1983. Part I--Taxonomical study and geographical distribution.

Four species of Adlerius phlebotomine sandflies have been recorded in China, namely: P. chinensis Newstead, 1916 (Pc), P. fengi Leng & Zhang, 1994; P. longiductus Parrot, 1928 and P. sichuanensis Leng & Yin, 1983 (Ps). Adlerius phlebotomies are the main vectors of visceral leishmaniasis (VL) in China; three of them are acknowledged as VL vectors and P. fengi is considered a potential VL vector for southwestern mountainous region. Different opinion has been raised to the validity of identity of Ps by some investigators from Shanghai and Shanxi who consider Ps to be a large type of Pc instead of an isolate species. The center of controversy is whether Ps is an isolate taxon or a large type of Pc. The present authors have carried out a series of comparative studies for these two flies on: 1 quantitative and qualitative morphological characters of four Chinese Adlerius phlebotomies; and 2. differences in geographical distribution. All specimens of Pc and Ps used in the present study are collected where their holotypes-paratypes were produced--West Mountain, West Suburb, Beijing and Lixian County, Sichuan Province. The results have forcefully proved that Ps is an isolate species instead of a so-called large type Pc according to the concept of species. The clarification of their taxonomical identities is meaningful because both of them are VL vectors in different epidemic areas in China; especially Ps is an important VL vector in high mountainous regions of southwestern China and some extend to the Loess Plateau of northwestern China, where VL still exists and it is also the first Phlebotomine sandfly discovered in Tibet, the locality being near Assam in India (Leng et al. 1990).

Animals↗

PubMed2001

[Aedes albopictus: cloning and identification of the acetylcholinesterase gene fragment from the mosquito].

OBJECTIVE: To isolate, clone and identify the acetylcholinesterase (AChE) fragment from the mosquito, Aedes albopictus, in relation to exploring mechanism of insectide resistance. METHODS: The genome DNA extracted from the mosquito was used for degenerate polymerase chain reaction (PCR) and the two pairs of oligonucleotides encoding the highly conserved protein sequences were used as primers. The reaction products were cloned to T-vector and transfected into E. coli JM 109. The replicative form DNA of recombinant vector extracted from E. coli JM 109 through alkalilysis was identified by the methods of digestion with EcoR I and Sal I and PCR. RESULTS: The products of degenerate primers polymerase chain reaction were obtained and the identified clone belongs to the AChE fragment of the mosquito. CONCLUSION: The clone was identified as the AChE fragment of Aedes albopictus.

Acetylcholinesterase↗

PubMed2000

The release of serotonin in rat spinal dorsal horn and periaqueductal gray following carrageenan inflammation.

The level of serotonin (5-HT) and its major metabolite 5-hydroxyindoleacetic acid (5-HIAA) in dorsal horn of spinal cord and periaqueductal gray (PAG) were measured using in vivo microdialysis coupled with high performance liquid chromatography and electrochemical detection. Intraplantar injection of carrageenan was used to evoke release. Extracellular concentrations of 5-HT and 5-HIAA in spinal dorsal horn and PAG significantly increased following carrageenan inflammation. The peak occurred at 3h and then gradually returned to baseline at 5-7h. The carrageenan-induced release of 5-HT and 5-HIAA in spinal cord rather than in PAG was decreased by intracerebroventricular (i.c.v.) injection of naloxone, and both in spinal dorsal horn and PAG, the release was increased by i.c.v. injection of bicuculine. These results suggest that activity of 5-HT in spinal cord and PAG increases with carrageenan inflammation; the carrageenan-induced release of 5-HT and 5-HIAA in spinal cord may be tonically modulated by supraspinal opioid and GABA systems, whereas the release in PAG may only be tonically modulated by endogenous GABA system in supraspinal level.

Animals↗

PubMed2000

NEC1, a novel gene, highly expressed in nectary tissue of Petunia hybrida.

To study the molecular regulation of nectary development, we cloned NEC1, a gene predominantly expressed in the nectaries of Petunia hybrida, by using the differential display RT-PCR technique. The secondary structure of the putative NEC1 protein is reminiscent of a transmembrane protein, indicating that the protein is incorporated into the cell membrane or the cytoplast membrane. Immunolocalization revealed that NEC1 protein is present in the nectaries. Northern blot analyses showed that NEC1 is highly expressed in nectary tissue and weakly in the stamen. GUS expression driven by the NEC1 promoter revealed GUS activity in the outer nectary parenchyma cells, the upper part of the filament and the anther stomium. The same expression pattern was observed in Brassica napus. GUS expression was observed as blue spots on the surface of very young nectaries that do not secrete nectar and do accumulate starch. GUS expression was highest in open flowers in which active secretion of nectar and starch hydrolysis had taken place. Ectopic expression of NEC1 resulted in transgenic plants that displayed a phenotype with leaves having 3-4 times more phloem bundles in mid-veins than the wild-type Petunia. The possible role of NEC1 gene in sugar metabolism and nectar secretion is discussed.

Amino Acid Sequence↗

PubMed2000

[The research on the expression of rabbit defensin (NP-1) gene in transgenic tomato].

Rabbit defensin NP-1 is one kind of alpha-defensins. It is composed of 33 amino acids. It was firstly extracted from polymorphonnuclear neutrophile of rabbits. It displayed resistance to bacteria, fungi and virus, especially high resistance to bacteria. In our experiments NP-1 gene was constructed into a plant expression vector. Eight transgenic plants containing the rabbit defensin gene (NP-1) were obtained through agrobacterium-mediated transformation. The transgenic plants were analysized by PCR, Southern hybridization, Northern dot blot hybridization and in vitro microbicidical activity against E. coli and Fusarium oxysporum. The results showed that NP-1 gene was transformed into the tomato, and the transgene displayed physiological-level expression. The transgenic tomato also showed resistance to pathogen Fusarium oxysporum in vivo. Our experiments paved a way for pathogen resistance breeding of tomato.

Animals↗

PubMed1999

Paradoxical increase in retinoblastoma protein in colorectal carcinomas may protect cells from apoptosis.

The retinoblastoma (Rb) gene is inactivated in a variety of human cancers, but in colorectal carcinomas there is frequently increased expression of this gene. This is paradoxical in view of the known role of Rb as a tumor suppressor gene. In the present study, we compared the levels of expression of the Rb protein (pRb) in normal human colorectal mucosa, adenomatous polyps, and carcinomas by immunohistochemistry. In vitro studies were also done to examine the phenotypic effects of an antisense oligodeoxynucleotide (AS-Rb) targeted to Rb mRNA in the HCT116 colon carcinoma cell line that expresses a relatively high level of pRb. The incidence of pRb-positive cells was increased during multistage colorectal carcinogenesis. In vitro treatment of HCT116 cells with AS-Rb decreased the level of pRb by about 70% and also decreased the levels of the cyclin D1 protein and cyclin D1-associated kinase activity. AS-Rb inhibited growth of HCT116 cells and induced apoptosis. Reporter assays indicated about a 17-fold increase in E2F activity. These findings suggest that the increased expression of pRb in colorectal carcinoma cells may provide a homeostatic mechanism that protects them from growth inhibition and apoptosis, perhaps by counterbalancing potentially toxic effects of excessive E2F activity.

Apoptosis↗

PubMed1999

Neuronal ERCC6 mRNA expression in rat brain induced by a transient focal cerebral ischemia.

AIM: To study whether the excision repair cross-complementing group 6 (ERCC6) is involved in the neuronal pathophysiological process following cerebral ischemia-reperfusion injury. METHODS: A transient middle cerebral artery occlusion (MCAO) was used to induce cerebral ischemia-reperfusion injury in rat brain. Northern blot was used to check a specific signal for oligonucleotide probe. The expression of ERCC6 mRNA in the rat brain was observed by in situ hybridization. The specific cellular distribution of ERCC6 mRNA in the neuron or glia of the rat brain was analyzed by double staining combined with confocal laser scanning microscopic analysis. RESULT: The expression of ERCC6 mRNA in the penumbra area increased following ischemia and reperfusion with a time-dependent manner. ERCC6 was expressed on d 2, reached peak values on d 3, and kept high level even on d 14 of reperfusion following ischemia. Number of ERCC6 mRNA expressive cell in the penumbra area on d 1, d 2, d 3, d 7, d 14 of reperfusion following ischemia were (0 +/- 0), (253 +/- 56), (816 +/- 355), (341 +/- 185), (128 +/- 95) x 10(6) cells/m2, respectively. Confocal microscopic analysis showed that ERCC6 mRNA coexpressed with phosphopyruvate hydratase in the neurons and with glial fibrillary acidic protein (GFAP) in a few proliferation astrocyte glia. CONCLUSION: The expression of transcription-repair coupling factor ERCC6 mRNA in the neuron and glia was induced by ischemia-reperfusion injury.

Animals↗

PubMed1999

Hypoxia-ischemia altered expression of glutamate transporter EAAT1 in neonatal rat brain.

AIM: To observe altered expression of glutamate transporter EAAT1 after hypoxia-ischemia (H-I) in newborn rat brain. METHODS: Expression levels of EAAT1 were detected with immunohistochemistry method. RESULTS: EAAT1 was a little expressed in cerebral cortex at sham-operated group [(36 +/- 10) cells/slice]. Its expression in cerebral cortex increased at 24 h and 48 h following H-I [(314 +/- 162) cells/slice and (431 +/- 149) cells/slice, respectively], and recovered to control level at 72 h following H-I [(52 +/- 8) cells/slice]. The expression of EAAT1 in the ipsilateral cortex to common carotid artery (CCA) ligation was higher than that in the contralateral cortex. CONCLUSION: After H-I, the expression of EAAT1 had a temporal change in cerebral cortex of newborn rat, and was mainly located in the ipsilateral cortex to CCA ligation.

ATP-Binding Cassette Transporters↗

PubMed1999

Protective effect of melatonin on injuried cerebral neurons is associated with bcl-2 protein over-expression.

AIM: To study the protective effect of melatonin against neuronal injury and the possible roles of alteration in the expression of bcl-2 and bax following brain ischemia. METHODS: Brain ischemia was induced by left middle cerebral artery occlusion (MCAO) for 60 min in rats. Brain damage was evaluated by the infarct area and the neuronal cell counting. The expression of bcl-2 and bax was analyzed by immunohistochemical method. RESULTS: Melatonin decreased the infarct area and prevented the neuronal death after 24-h reperfusion following 1-h MCAO. Melatonin given before the ischemia enhanced the expression of bcl-2 in the penumbra area and had no significant effect on the expression of bax. CONCLUSION: Melatonin effectively attenuated ischemic brain injury and increased the expression of neuronal bcl-2 in the ischemic brain, indicating that the protective effect of melatonin was associated with up-regulation of bcl-2 in ischemia-induced neuronal death.

Animals↗

PubMed1998

Release of tumor necrosis factor-alpha from coronary smooth muscle: activation of NF-kappaB and inhibition by elevated cyclic AMP.

BACKGROUND: Evidence suggests that tumor necrosis factor-alpha (TNF-alpha) is involved in heart diseases such as atherosclerosis. We used porcine coronary arteries and smooth muscle cells cultured from these vessels to study the regulation of production of TNF-alpha. The aims were to determine if bacterial lipopolysaccharide (LPS) could stimulate production; if activation of the nuclear regulatory factor, NF-kappaB, was associated with production; and if intracellular cAMP regulates TNF-alpha in coronary vasculature through a mechanism involving NF-kappaB. MATERIAL AND METHODS: LPS was used to stimulate TNF-alpha production. Forskolin (FSK) and 8-Br-cAMP were added to tissue and cells in order to elevate intracellular cAMP. TNF-alpha release into the bathing medium was measured by the L929 cell cytotoxicity assay. Intracellular cAMP was determined by radioimmunoassay. NF-kappaB activation was determined in whole cell extracts by electrophoretic mobility shift assay. RESULTS: In segments of coronary arteries, LPS stimulated TNF-alpha release which increased with time to a maximum at 6 h (485 +/- 19 units/g tissue) and remained elevated at this level for 24 h. In contrast, the level of TNF-alpha measured at 24 h in medium from coronary tissue not exposed to LPS was 11.1 +/- 4.1 units/g tissue. In the presence of LPS, both FSK and 8-Br-cAMP significantly reduced TNF-alpha release. For instance at 6 h in the presence of LPS and FSK or 8-Br-cAMP, TNF-alpha was 126 +/- 24 and 71.6 +/- 22 units/g tissue, respectively (P < 0.05 vs LPS alone). Tissue levels of cAMP were significantly elevated in the presence of FSK. Similar results were obtained with smooth muscle cells cultured from the coronary arteries; i.e., LPS stimulated TNF-alpha release which was inhibited in a concentration-dependent manner by a rise in intracellular cAMP induced by FSK. In cultured cells release of TNF-alpha stimulated by LPS was associated with activation of NF-kappaB. Neither FSK nor 8-Br cAMP inhibited activation of NF-kappaB by LPS. CONCLUSIONS: Porcine coronary arteries produce TNF-alpha from a smooth muscle cell source. Production stimulated by LPS was inhibited by elevated intracellular cAMP and was associated with activation of NF-kappaB. However, activation of NF-kappaB was not inhibited by elevated cAMP, suggesting that the regulatory action of this cyclic nucleotide could lie downstream from activation of the TNF-alpha gene. These results support the view that coronary vessels can be a source of TNF-alpha possibly involved in heart disease.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1997

Cell-surface perturbations of the epidermal growth factor and vascular endothelial growth factor receptors by phosphorothioate oligodeoxynucleotides.

Antisense oligodeoxynucleotides offer potential as therapeutic agents to inhibit gene expression. Recent evidence indicates that oligodeoxynucleotides designed to target specific nucleic acid sequences can interact nonspecifically with proteins. This report describes the interactive capabilities of phosphorothioate oligodeoxynucleotides of defined sequence and length with two essential protein tyrosine receptors, flk-1 and epidermal growth factor receptor (EGFR), and their effects on receptor signaling in a transfected and tumor cell line, respectively. Phosphorothioate oligodeoxynucleotides bound to the cell surface, as demonstrated by fluorescence-activated cell-sorter analyses (FACS), and perturbed receptor activation in the presence and absence of cognate ligands, EGF (EGFR) and vascular endothelial growth factor (flk-1), in phosphorylation assays. Certain phosphorothioate oligodeoxynucleotides interacted relatively selectively with flk-1 and partially blocked the binding of specific anti-receptor monoclonal antibodies to target sites. They stimulated EGFR phosphorylation in the absence of EGF but antagonized ligand-mediated activation of EGFR and flk-1. In vivo studies showed that a nonspecific phosphorothioate oligodeoxynucleotide suppressed the growth of glioblastoma in a mouse model of tumorigenesis. These results emphasize the capacity of phosphorothioate oligodeoxynucleotides to interact with cells in a sequence-selective nonantisense manner, while associating with cellular membrane proteins in ways that can inhibit cellular metabolic activities.

3T3 Cells↗

PubMed1997

Eighty-year research of phlebotomine sandflies (Diptera: Psychodidae) in China (1915-1995). II. Phlebotomine vectors of leishmaniasis in China.

The human leishmaniasis vector--phlebotomine sandfly--was proven in China, i.e. P. alexandri, P. chinesis, P. longiductus, P. sichuanesis and P. smirnovi. The infectious rate, biology and distribution of these vectors are shown in this review. Two disputed questions about P. chinensis and P. sichuanesis, P. smirnovi and P. wui are discussed. At last, the vectors of animal leishmania are provided, i.e. the vectors of great gerbil leishmania: P. alexandri, P. andrejievi, P. caucasicus, P. mongolensis and P. smirnovi, and the vector of lizards leishmania: S. sinkiangensis.

Animals↗

PubMed1997

Inhibition of release of tumor necrosis factor-alpha from human vascular tissue and smooth muscle cells by glucocorticoids.

OBJECTIVES: Based on our previous study that bacterial lipopolysaccharide stimulates release of tumor necrosis factor (TNF)-alpha from human vascular tissue and smooth muscle cells, we tested the hypothesis that release of TNF could be inhibited by pretreatment with glucocorticoids. DESIGN: Prospective, repeated-measures analysis of concentration-response relationships. SETTING: Academic anesthesiology research laboratory. SUBJECTS: Segments of internal mammary artery and saphenous vein were obtained during coronary artery bypass surgery. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Confluent human smooth muscle cells, cultured from saphenous vein and internal mammary artery, were exposed to 20 micrograms/mL of bacterial lipopolysaccharide following pretreatment for 18 hrs with either 0.1, 1.0, or 10.0 microM of dexamethasone. At 1, 3, 6, 18, and 24 hrs, the culture medium was removed and analyzed for biologically active TNF-alpha using the L929 cell cytotoxicity assay. Smooth muscle cells exposed to bacterial lipopolysaccharide but not treated with dexamethasone served as controls. In control internal mammary cells, bacterial lipopolysaccharide stimulated TNF-alpha release in a time-dependent manner to a peak of 36 +/- 2.3 U/mg of cell protein at 6 hrs, compared with 0.7 +/- 0.3 U/mg of cell protein in cells not exposed to lipopolysaccharide. Dexamethasone inhibited bacterial lipopolysaccharide-stimulated release at all time points in a concentration-dependent manner. For instance, at 6 hrs, TNF-alpha was 12 +/- 2.2, 6.9 +/- 1.7, and 2.3 +/- 0.9 U/mg of cell protein for cells pretreated with 0.1, 1.0, and 10.0 microM of dexamethasone, respectively (p < .05 vs. control). In separate experiments, segments of internal mammary artery and saphenous vein were obtained from five patients who received 1 g of methylprednisolone intravenously during induction of anesthesia, and from seven patients who did not receive methylprednisolone. Bacterial lipopolysaccharide induced release of TNF-alpha from vascular tissues of untreated patients in a time-dependent manner (e.g., 733 +/- 44 U/g of tissue at 6 hrs in saphenous vein). In contrast, in patients treated with methylprednisolone, bacterial lipopolysaccharide did not stimulate release from vascular tissues incubated for up to 24 hrs. CONCLUSIONS: These results indicate that human vascular tissue, particularly the smooth muscle cell, may be a source of TNF-alpha and that glucocorticoids inhibit release stimulated by bacterial lipopolysaccharide.

Cells, Cultured↗

PubMed1997

Increased intracellular cyclic adenosine 3', 5'-monophosphate inhibits release of tumor necrosis factor-alpha from human vascular tissue and cultured smooth muscle cells.

OBJECTIVES: We recently reported that bacterial lipopolysaccharide stimulates release of tumor necrosis factor (TNF)-alpha from both human vascular tissue and cultured smooth muscle cells. In the current study, we tested the hypothesis that increased intracellular cyclic adenosine 3',5'-monophosphate (cAMP) could inhibit TNF-alpha release. DESIGN: Prospective, repeated-measures analysis. SETTING: Academic research laboratory. SUBJECTS: Segments of internal mammary artery and saphenous vein from patients undergoing coronary artery bypass surgery. MEASUREMENTS AND MAIN RESULTS: Segments of saphenous vein and internal mammary artery and confluent smooth muscle cells cultured from these vessels were incubated in the presence of 20 micrograms/mL bacterial lipopolysaccharide, alone or with the addition of forskolin or 8-Br-cAMP. At 0, 1, 3, 6, 18, and 24 hrs, the incubation medium was removed from vessel segments or cells and was analyzed for biologically active TNF-alpha, using the L929 cell cytotoxicity assay. cAMP was extracted from tissue and cells with 0.1 N HCl and was analyzed by radioimmunoassay. Bacterial lipopolysaccharide stimulated the release of TNF-alpha from internal mammary smooth muscle cells at all time points. For example, at 6 hrs, TNF-alpha concentration in the medium from lipopolysaccharide-stimulated cells was 20 +/- 1.6 U/mg of cell protein, compared with 0.9 +/- 0.5 U/mg of cell protein in control cell medium (p < .05). Forskolin-inhibited bacterial lipopolysaccharide stimulated TNF-alpha release. In the presence of lipopolysaccharide and forskolin, TNF-alpha release at 6 hrs was 8.6 +/- 1.5 U/mg of cell protein (p < .05 vs. in the presence of bacterial lipopolysaccharide alone). Bacterial lipopolysaccharide, alone, had no effect on intracellular cAMP. Forskolin increased intracellular cAMP levels to 74.0 +/- 12 pmol/mg of cell protein at 6 hrs from a control level of 7.7 +/- 0.4 pmol/mg (p < .05). The 8-Br-cAMP, an agent that mimics the action of intracellular cAMP, also inhibited TNF-alpha release stimulated by lipopolysaccharide. Similar inhibition by forskolin and 8-Br-cAMP on TNF-alpha release was obtained with smooth muscle cells from saphenous vein. Finally, in tissue segments from either internal mammary artery or saphenous vein, both forskolin and 8-Br-cAMP inhibited lipopolysaccharide-stimulated TNF-alpha release. CONCLUSIONS: These results are consistent with the conclusion that vascular tissue, particularly the smooth muscle cell, is a source of TNF-alpha. Further, bacterial lipopolysaccharide-stimulated tumor TNF-alpha release can be inhibited by increased intracellular cAMP.

Animals↗